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  1. Controllability analysis of transcriptional regulatory networks reveals circular control patterns among transcription factors

    DEFF Research Database (Denmark)

    Österlund, Tobias; Bordel, Sergio; Nielsen, Jens

    2015-01-01

    % for the human network. The high controllability (low number of drivers needed to control the system) in yeast, mouse and human is due to the presence of internal loops in their regulatory networks where the TFs regulate each other in a circular fashion. We refer to these internal loops as circular control...... motifs (CCM). The E. coli transcriptional regulatory network, which does not have any CCMs, shows a hierarchical structure of the transcriptional regulatory network in contrast to the eukaryal networks. The presence of CCMs also has influence on the stability of these networks, as the presence of cycles...

  2. Metabolic Network Topology Reveals Transcriptional Regulatory Signatures of Type 2 Diabetes

    DEFF Research Database (Denmark)

    Zelezniak, Aleksej; Pers, Tune Hannes; Pinho Soares, Simao Pedro

    2010-01-01

    mechanisms underlying these transcriptional changes and their impact on the cellular metabolic phenotype is a challenging task due to the complexity of transcriptional regulation and the highly interconnected nature of the metabolic network. In this study we integrate skeletal muscle gene expression datasets...... with human metabolic network reconstructions to identify key metabolic regulatory features of T2DM. These features include reporter metabolites—metabolites with significant collective transcriptional response in the associated enzyme-coding genes, and transcription factors with significant enrichment...... factor regulatory network connecting several parts of metabolism. The identified transcription factors include members of the CREB, NRF1 and PPAR family, among others, and represent regulatory targets for further experimental analysis. Overall, our results provide a holistic picture of key metabolic...

  3. Transcriptional decomposition reveals active chromatin architectures and cell specific regulatory interactions

    DEFF Research Database (Denmark)

    Rennie, Sarah; Dalby, Maria; van Duin, Lucas

    2018-01-01

    Transcriptional regulation is tightly coupled with chromosomal positioning and three-dimensional chromatin architecture. However, it is unclear what proportion of transcriptional activity is reflecting such organisation, how much can be informed by RNA expression alone and how this impacts disease...... proportion of total levels and is highly informative of topological associating domain activities and organisation, revealing boundaries and chromatin compartments. Furthermore, expression data alone accurately predict individual enhancer-promoter interactions, drawing features from expression strength...... between transcription and chromatin architecture....

  4. Hierarchical structure and modules in the Escherichia coli transcriptional regulatory network revealed by a new top-down approach

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    Buer Jan

    2004-12-01

    Full Text Available Abstract Background Cellular functions are coordinately carried out by groups of genes forming functional modules. Identifying such modules in the transcriptional regulatory network (TRN of organisms is important for understanding the structure and function of these fundamental cellular networks and essential for the emerging modular biology. So far, the global connectivity structure of TRN has not been well studied and consequently not applied for the identification of functional modules. Moreover, network motifs such as feed forward loop are recently proposed to be basic building blocks of TRN. However, their relationship to functional modules is not clear. Results In this work we proposed a top-down approach to identify modules in the TRN of E. coli. By studying the global connectivity structure of the regulatory network, we first revealed a five-layer hierarchical structure in which all the regulatory relationships are downward. Based on this regulatory hierarchy, we developed a new method to decompose the regulatory network into functional modules and to identify global regulators governing multiple modules. As a result, 10 global regulators and 39 modules were identified and shown to have well defined functions. We then investigated the distribution and composition of the two basic network motifs (feed forward loop and bi-fan motif in the hierarchical structure of TRN. We found that most of these network motifs include global regulators, indicating that these motifs are not basic building blocks of modules since modules should not contain global regulators. Conclusion The transcriptional regulatory network of E. coli possesses a multi-layer hierarchical modular structure without feedback regulation at transcription level. This hierarchical structure builds the basis for a new and simple decomposition method which is suitable for the identification of functional modules and global regulators in the transcriptional regulatory network of E

  5. Patterns of subnet usage reveal distinct scales of regulation in the transcriptional regulatory network of Escherichia coli.

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    Carsten Marr

    Full Text Available The set of regulatory interactions between genes, mediated by transcription factors, forms a species' transcriptional regulatory network (TRN. By comparing this network with measured gene expression data, one can identify functional properties of the TRN and gain general insight into transcriptional control. We define the subnet of a node as the subgraph consisting of all nodes topologically downstream of the node, including itself. Using a large set of microarray expression data of the bacterium Escherichia coli, we find that the gene expression in different subnets exhibits a structured pattern in response to environmental changes and genotypic mutation. Subnets with fewer changes in their expression pattern have a higher fraction of feed-forward loop motifs and a lower fraction of small RNA targets within them. Our study implies that the TRN consists of several scales of regulatory organization: (1 subnets with more varying gene expression controlled by both transcription factors and post-transcriptional RNA regulation and (2 subnets with less varying gene expression having more feed-forward loops and less post-transcriptional RNA regulation.

  6. Boolean modelling reveals new regulatory connections between transcription factors orchestrating the development of the ventral spinal cord.

    KAUST Repository

    Lovrics, Anna

    2014-11-14

    We have assembled a network of cell-fate determining transcription factors that play a key role in the specification of the ventral neuronal subtypes of the spinal cord on the basis of published transcriptional interactions. Asynchronous Boolean modelling of the network was used to compare simulation results with reported experimental observations. Such comparison highlighted the need to include additional regulatory connections in order to obtain the fixed point attractors of the model associated with the five known progenitor cell types located in the ventral spinal cord. The revised gene regulatory network reproduced previously observed cell state switches between progenitor cells observed in knock-out animal models or in experiments where the transcription factors were overexpressed. Furthermore the network predicted the inhibition of Irx3 by Nkx2.2 and this prediction was tested experimentally. Our results provide evidence for the existence of an as yet undescribed inhibitory connection which could potentially have significance beyond the ventral spinal cord. The work presented in this paper demonstrates the strength of Boolean modelling for identifying gene regulatory networks.

  7. Comparative transcriptional profiling of 3 murine models of SLE nephritis reveals both unique and shared regulatory networks.

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    Ramalingam Bethunaickan

    Full Text Available To define shared and unique features of SLE nephritis in mouse models of proliferative and glomerulosclerotic renal disease.Perfused kidneys from NZB/W F1, NZW/BXSB and NZM2410 mice were harvested before and after nephritis onset. Affymetrix based gene expression profiles of kidney RNA were analyzed using Genomatix Pathway Systems and Ingenuity Pathway Analysis software. Gene expression patterns were confirmed using real-time PCR.955, 1168 and 755 genes were regulated in the kidneys of nephritic NZB/W F1, NZM2410 and NZW/BXSB mice respectively. 263 genes were regulated concordantly in all three strains reflecting immune cell infiltration, endothelial cell activation, complement activation, cytokine signaling, tissue remodeling and hypoxia. STAT3 was the top associated transcription factor, having a binding site in the gene promoter of 60/263 regulated genes. The two strains with proliferative nephritis shared a macrophage/DC infiltration and activation signature. NZB/W and NZM2410 mice shared a mitochondrial dysfunction signature. Dominant T cell and plasma cell signatures in NZB/W mice reflected lymphoid aggregates; this was the only strain with regulatory T cell infiltrates. NZW/BXSB mice manifested tubular regeneration and NZM2410 mice had the most metabolic stress and manifested loss of nephrin, indicating podocyte loss.These findings identify shared inflammatory mechanisms of SLE nephritis that can be therapeutically targeted. Nevertheless, the heterogeneity of effector mechanisms suggests that individualized therapy might need to be based on biopsy findings. Some common mechanisms are shared with non-immune-mediated renal diseases, suggesting that strategies to prevent tissue hypoxia and remodeling may be useful in SLE nephritis.

  8. Boolean modelling reveals new regulatory connections between transcription factors orchestrating the development of the ventral spinal cord.

    KAUST Repository

    Lovrics, Anna; Gao, Yu; Juhá sz, Bianka; Bock, Istvá n; Byrne, Helen M; Dinnyé s, Andrá s; Ková cs, Krisztiá n A

    2014-01-01

    with the five known progenitor cell types located in the ventral spinal cord. The revised gene regulatory network reproduced previously observed cell state switches between progenitor cells observed in knock-out animal models or in experiments where

  9. Next-Generation Sequencing of Genomic DNA Fragments Bound to a Transcription Factor in Vitro Reveals Its Regulatory Potential

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    Yukio Kurihara

    2014-12-01

    Full Text Available Several transcription factors (TFs coordinate to regulate expression of specific genes at the transcriptional level. In Arabidopsis thaliana it is estimated that approximately 10% of all genes encode TFs or TF-like proteins. It is important to identify target genes that are directly regulated by TFs in order to understand the complete picture of a plant’s transcriptome profile. Here, we investigate the role of the LONG HYPOCOTYL5 (HY5 transcription factor that acts as a regulator of photomorphogenesis. We used an in vitro genomic DNA binding assay coupled with immunoprecipitation and next-generation sequencing (gDB-seq instead of the in vivo chromatin immunoprecipitation (ChIP-based methods. The results demonstrate that the HY5-binding motif predicted here was similar to the motif reported previously and that in vitro HY5-binding loci largely overlapped with the HY5-targeted candidate genes identified in previous ChIP-chip analysis. By combining these results with microarray analysis, we identified hundreds of HY5-binding genes that were differentially expressed in hy5. We also observed delayed induction of some transcripts of HY5-binding genes in hy5 mutants in response to blue-light exposure after dark treatment. Thus, an in vitro gDNA-binding assay coupled with sequencing is a convenient and powerful method to bridge the gap between identifying TF binding potential and establishing function.

  10. Decoding genome-wide GadEWX-transcriptional regulatory networks reveals multifaceted cellular responses to acid stress in Escherichia coli

    DEFF Research Database (Denmark)

    Seo, Sang Woo; Kim, Donghyuk; O'Brien, Edward J.

    2015-01-01

    The regulators GadE, GadW and GadX (which we refer to as GadEWX) play a critical role in the transcriptional regulation of the glutamate-dependent acid resistance (GDAR) system in Escherichia coli K-12 MG1655. However, the genome-wide regulatory role of GadEWX is still unknown. Here we comprehens...

  11. A unified architecture of transcriptional regulatory elements

    DEFF Research Database (Denmark)

    Andersson, Robin; Sandelin, Albin Gustav; Danko, Charles G.

    2015-01-01

    Gene expression is precisely controlled in time and space through the integration of signals that act at gene promoters and gene-distal enhancers. Classically, promoters and enhancers are considered separate classes of regulatory elements, often distinguished by histone modifications. However...... and enhancers are considered a single class of functional element, with a unified architecture for transcription initiation. The context of interacting regulatory elements and the surrounding sequences determine local transcriptional output as well as the enhancer and promoter activities of individual elements....

  12. Transcription regulatory networks analysis using CAGE

    KAUST Repository

    Tegnér, Jesper N.

    2009-10-01

    Mapping out cellular networks in general and transcriptional networks in particular has proved to be a bottle-neck hampering our understanding of biological processes. Integrative approaches fusing computational and experimental technologies for decoding transcriptional networks at a high level of resolution is therefore of uttermost importance. Yet, this is challenging since the control of gene expression in eukaryotes is a complex multi-level process influenced by several epigenetic factors and the fine interplay between regulatory proteins and the promoter structure governing the combinatorial regulation of gene expression. In this chapter we review how the CAGE data can be integrated with other measurements such as expression, physical interactions and computational prediction of regulatory motifs, which together can provide a genome-wide picture of eukaryotic transcriptional regulatory networks at a new level of resolution. © 2010 by Pan Stanford Publishing Pte. Ltd. All rights reserved.

  13. DNA residence time is a regulatory factor of transcription repression

    Science.gov (United States)

    Clauß, Karen; Popp, Achim P.; Schulze, Lena; Hettich, Johannes; Reisser, Matthias; Escoter Torres, Laura; Uhlenhaut, N. Henriette

    2017-01-01

    Abstract Transcription comprises a highly regulated sequence of intrinsically stochastic processes, resulting in bursts of transcription intermitted by quiescence. In transcription activation or repression, a transcription factor binds dynamically to DNA, with a residence time unique to each factor. Whether the DNA residence time is important in the transcription process is unclear. Here, we designed a series of transcription repressors differing in their DNA residence time by utilizing the modular DNA binding domain of transcription activator-like effectors (TALEs) and varying the number of nucleotide-recognizing repeat domains. We characterized the DNA residence times of our repressors in living cells using single molecule tracking. The residence times depended non-linearly on the number of repeat domains and differed by more than a factor of six. The factors provoked a residence time-dependent decrease in transcript level of the glucocorticoid receptor-activated gene SGK1. Down regulation of transcription was due to a lower burst frequency in the presence of long binding repressors and is in accordance with a model of competitive inhibition of endogenous activator binding. Our single molecule experiments reveal transcription factor DNA residence time as a regulatory factor controlling transcription repression and establish TALE-DNA binding domains as tools for the temporal dissection of transcription regulation. PMID:28977492

  14. Global Analysis of Photosynthesis Transcriptional Regulatory Networks

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    Imam, Saheed; Noguera, Daniel R.; Donohue, Timothy J.

    2014-01-01

    Photosynthesis is a crucial biological process that depends on the interplay of many components. This work analyzed the gene targets for 4 transcription factors: FnrL, PrrA, CrpK and MppG (RSP_2888), which are known or predicted to control photosynthesis in Rhodobacter sphaeroides. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) identified 52 operons under direct control of FnrL, illustrating its regulatory role in photosynthesis, iron homeostasis, nitrogen metabolism and regulation of sRNA synthesis. Using global gene expression analysis combined with ChIP-seq, we mapped the regulons of PrrA, CrpK and MppG. PrrA regulates ∼34 operons encoding mainly photosynthesis and electron transport functions, while CrpK, a previously uncharacterized Crp-family protein, regulates genes involved in photosynthesis and maintenance of iron homeostasis. Furthermore, CrpK and FnrL share similar DNA binding determinants, possibly explaining our observation of the ability of CrpK to partially compensate for the growth defects of a ΔFnrL mutant. We show that the Rrf2 family protein, MppG, plays an important role in photopigment biosynthesis, as part of an incoherent feed-forward loop with PrrA. Our results reveal a previously unrealized, high degree of combinatorial regulation of photosynthetic genes and significant cross-talk between their transcriptional regulators, while illustrating previously unidentified links between photosynthesis and the maintenance of iron homeostasis. PMID:25503406

  15. Global analysis of photosynthesis transcriptional regulatory networks.

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    Saheed Imam

    2014-12-01

    Full Text Available Photosynthesis is a crucial biological process that depends on the interplay of many components. This work analyzed the gene targets for 4 transcription factors: FnrL, PrrA, CrpK and MppG (RSP_2888, which are known or predicted to control photosynthesis in Rhodobacter sphaeroides. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq identified 52 operons under direct control of FnrL, illustrating its regulatory role in photosynthesis, iron homeostasis, nitrogen metabolism and regulation of sRNA synthesis. Using global gene expression analysis combined with ChIP-seq, we mapped the regulons of PrrA, CrpK and MppG. PrrA regulates ∼34 operons encoding mainly photosynthesis and electron transport functions, while CrpK, a previously uncharacterized Crp-family protein, regulates genes involved in photosynthesis and maintenance of iron homeostasis. Furthermore, CrpK and FnrL share similar DNA binding determinants, possibly explaining our observation of the ability of CrpK to partially compensate for the growth defects of a ΔFnrL mutant. We show that the Rrf2 family protein, MppG, plays an important role in photopigment biosynthesis, as part of an incoherent feed-forward loop with PrrA. Our results reveal a previously unrealized, high degree of combinatorial regulation of photosynthetic genes and significant cross-talk between their transcriptional regulators, while illustrating previously unidentified links between photosynthesis and the maintenance of iron homeostasis.

  16. The regulatory mechanism of fruit ripening revealed by analyses of direct targets of the tomato MADS-box transcription factor RIPENING INHIBITOR

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    Fujisawa, Masaki; Ito, Yasuhiro

    2013-01-01

    The developmental process of ripening is unique to fleshy fruits and a key factor in fruit quality. The tomato (Solanum lycopersicum) MADS-box transcription factor RIPENING INHIBITOR (RIN), one of the earliest-acting ripening regulators, is required for broad aspects of ripening, including ethylene-dependent and -independent pathways. However, our knowledge of direct RIN target genes has been limited, considering the broad effects of RIN on ripening. In a recent work published in The Plant Cell, we identified 241 direct RIN target genes by chromatin immunoprecipitation coupled with DNA microarray (ChIP-chip) and transcriptome analysis. Functional classification of the targets revealed that RIN participates in the regulation of many biological processes including well-known ripening processes such as climacteric ethylene production and lycopene accumulation. In addition, we found that ethylene is required for the full expression of RIN and several RIN-targeting transcription factor genes at the ripening stage. Here, based on our recently published findings and additional data, we discuss the ripening processes regulated by RIN and the interplay between RIN and ethylene. PMID:23518588

  17. Reconstructing transcriptional regulatory networks through genomics data

    OpenAIRE

    Sun, Ning; Zhao, Hongyu

    2009-01-01

    One central problem in biology is to understand how gene expression is regulated under different conditions. Microarray gene expression data and other high throughput data have made it possible to dissect transcriptional regulatory networks at the genomics level. Owing to the very large number of genes that need to be studied, the relatively small number of data sets available, the noise in the data and the different natures of the distinct data types, network inference presents great challen...

  18. The transcriptional regulatory network of Mycobacterium tuberculosis.

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    Joaquín Sanz

    Full Text Available Under the perspectives of network science and systems biology, the characterization of transcriptional regulatory (TR networks beyond the context of model organisms offers a versatile tool whose potential remains yet mainly unexplored. In this work, we present an updated version of the TR network of Mycobacterium tuberculosis (M.tb, which incorporates newly characterized transcriptional regulations coming from 31 recent, different experimental works available in the literature. As a result of the incorporation of these data, the new network doubles the size of previous data collections, incorporating more than a third of the entire genome of the bacterium. We also present an exhaustive topological analysis of the new assembled network, focusing on the statistical characterization of motifs significances and the comparison with other model organisms. The expanded M.tb transcriptional regulatory network, considering its volume and completeness, constitutes an important resource for diverse tasks such as dynamic modeling of gene expression and signaling processes, computational reliability determination or protein function prediction, being the latter of particular relevance, given that the function of only a small percent of the proteins of M.tb is known.

  19. In silico discovery of transcription regulatory elements in Plasmodium falciparum

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    Le Roch Karine G

    2008-02-01

    Full Text Available Abstract Background With the sequence of the Plasmodium falciparum genome and several global mRNA and protein life cycle expression profiling projects now completed, elucidating the underlying networks of transcriptional control important for the progression of the parasite life cycle is highly pertinent to the development of new anti-malarials. To date, relatively little is known regarding the specific mechanisms the parasite employs to regulate gene expression at the mRNA level, with studies of the P. falciparum genome sequence having revealed few cis-regulatory elements and associated transcription factors. Although it is possible the parasite may evoke mechanisms of transcriptional control drastically different from those used by other eukaryotic organisms, the extreme AT-rich nature of P. falciparum intergenic regions (~90% AT presents significant challenges to in silico cis-regulatory element discovery. Results We have developed an algorithm called Gene Enrichment Motif Searching (GEMS that uses a hypergeometric-based scoring function and a position-weight matrix optimization routine to identify with high-confidence regulatory elements in the nucleotide-biased and repeat sequence-rich P. falciparum genome. When applied to promoter regions of genes contained within 21 co-expression gene clusters generated from P. falciparum life cycle microarray data using the semi-supervised clustering algorithm Ontology-based Pattern Identification, GEMS identified 34 putative cis-regulatory elements associated with a variety of parasite processes including sexual development, cell invasion, antigenic variation and protein biosynthesis. Among these candidates were novel motifs, as well as many of the elements for which biological experimental evidence already exists in the Plasmodium literature. To provide evidence for the biological relevance of a cell invasion-related element predicted by GEMS, reporter gene and electrophoretic mobility shift assays

  20. Systematic Profiling of Poly(A)+ Transcripts Modulated by Core 3’ End Processing and Splicing Factors Reveals Regulatory Rules of Alternative Cleavage and Polyadenylation

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    Li, Wencheng; You, Bei; Hoque, Mainul; Zheng, Dinghai; Luo, Wenting; Ji, Zhe; Park, Ji Yeon; Gunderson, Samuel I.; Kalsotra, Auinash; Manley, James L.; Tian, Bin

    2015-01-01

    Alternative cleavage and polyadenylation (APA) results in mRNA isoforms containing different 3’ untranslated regions (3’UTRs) and/or coding sequences. How core cleavage/polyadenylation (C/P) factors regulate APA is not well understood. Using siRNA knockdown coupled with deep sequencing, we found that several C/P factors can play significant roles in 3’UTR-APA. Whereas Pcf11 and Fip1 enhance usage of proximal poly(A) sites (pAs), CFI-25/68, PABPN1 and PABPC1 promote usage of distal pAs. Strong cis element biases were found for pAs regulated by CFI-25/68 or Fip1, and the distance between pAs plays an important role in APA regulation. In addition, intronic pAs are substantially regulated by splicing factors, with U1 mostly inhibiting C/P events in introns near the 5’ end of gene and U2 suppressing those in introns with features for efficient splicing. Furthermore, PABPN1 inhibits expression of transcripts with pAs near the transcription start site (TSS), a property possibly related to its role in RNA degradation. Finally, we found that groups of APA events regulated by C/P factors are also modulated in cell differentiation and development with distinct trends. Together, our results support an APA code where an APA event in a given cellular context is regulated by a number of parameters, including relative location to the TSS, splicing context, distance between competing pAs, surrounding cis elements and concentrations of core C/P factors. PMID:25906188

  1. Systematic profiling of poly(A+ transcripts modulated by core 3' end processing and splicing factors reveals regulatory rules of alternative cleavage and polyadenylation.

    Directory of Open Access Journals (Sweden)

    Wencheng Li

    2015-04-01

    Full Text Available Alternative cleavage and polyadenylation (APA results in mRNA isoforms containing different 3' untranslated regions (3'UTRs and/or coding sequences. How core cleavage/polyadenylation (C/P factors regulate APA is not well understood. Using siRNA knockdown coupled with deep sequencing, we found that several C/P factors can play significant roles in 3'UTR-APA. Whereas Pcf11 and Fip1 enhance usage of proximal poly(A sites (pAs, CFI-25/68, PABPN1 and PABPC1 promote usage of distal pAs. Strong cis element biases were found for pAs regulated by CFI-25/68 or Fip1, and the distance between pAs plays an important role in APA regulation. In addition, intronic pAs are substantially regulated by splicing factors, with U1 mostly inhibiting C/P events in introns near the 5' end of gene and U2 suppressing those in introns with features for efficient splicing. Furthermore, PABPN1 inhibits expression of transcripts with pAs near the transcription start site (TSS, a property possibly related to its role in RNA degradation. Finally, we found that groups of APA events regulated by C/P factors are also modulated in cell differentiation and development with distinct trends. Together, our results support an APA code where an APA event in a given cellular context is regulated by a number of parameters, including relative location to the TSS, splicing context, distance between competing pAs, surrounding cis elements and concentrations of core C/P factors.

  2. Transcriptional regulatory programs underlying barley germination and regulatory functions of Gibberellin and abscisic acid

    Science.gov (United States)

    2011-01-01

    Background Seed germination is a complex multi-stage developmental process, and mainly accomplished through concerted activities of many gene products and biological pathways that are often subjected to strict developmental regulation. Gibberellins (GA) and abscisic acid (ABA) are two key phytohormones regulating seed germination and seedling growth. However, transcriptional regulatory networks underlying seed germination and its associated biological pathways are largely unknown. Results The studies examined transcriptomes of barley representing six distinct and well characterized germination stages and revealed that the transcriptional regulatory program underlying barley germination was composed of early, late, and post-germination phases. Each phase was accompanied with transcriptional up-regulation of distinct biological pathways. Cell wall synthesis and regulatory components including transcription factors, signaling and post-translational modification components were specifically and transiently up-regulated in early germination phase while histone families and many metabolic pathways were up-regulated in late germination phase. Photosynthesis and seed reserve mobilization pathways were up-regulated in post-germination phase. However, stress related pathways and seed storage proteins were suppressed through the entire course of germination. A set of genes were transiently up-regulated within three hours of imbibition, and might play roles in initiating biological pathways involved in seed germination. However, highly abundant transcripts in dry barley and Arabidopsis seeds were significantly conserved. Comparison with transcriptomes of barley aleurone in response to GA and ABA identified three sets of germination responsive genes that were regulated coordinately by GA, antagonistically by ABA, and coordinately by GA but antagonistically by ABA. Major CHO metabolism, cell wall degradation and protein degradation pathways were up-regulated by both GA and seed

  3. Transcriptional regulatory programs underlying barley germination and regulatory functions of Gibberellin and abscisic acid

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    Lin Li

    2011-06-01

    Full Text Available Abstract Background Seed germination is a complex multi-stage developmental process, and mainly accomplished through concerted activities of many gene products and biological pathways that are often subjected to strict developmental regulation. Gibberellins (GA and abscisic acid (ABA are two key phytohormones regulating seed germination and seedling growth. However, transcriptional regulatory networks underlying seed germination and its associated biological pathways are largely unknown. Results The studies examined transcriptomes of barley representing six distinct and well characterized germination stages and revealed that the transcriptional regulatory program underlying barley germination was composed of early, late, and post-germination phases. Each phase was accompanied with transcriptional up-regulation of distinct biological pathways. Cell wall synthesis and regulatory components including transcription factors, signaling and post-translational modification components were specifically and transiently up-regulated in early germination phase while histone families and many metabolic pathways were up-regulated in late germination phase. Photosynthesis and seed reserve mobilization pathways were up-regulated in post-germination phase. However, stress related pathways and seed storage proteins were suppressed through the entire course of germination. A set of genes were transiently up-regulated within three hours of imbibition, and might play roles in initiating biological pathways involved in seed germination. However, highly abundant transcripts in dry barley and Arabidopsis seeds were significantly conserved. Comparison with transcriptomes of barley aleurone in response to GA and ABA identified three sets of germination responsive genes that were regulated coordinately by GA, antagonistically by ABA, and coordinately by GA but antagonistically by ABA. Major CHO metabolism, cell wall degradation and protein degradation pathways were up

  4. Transcription factor trapping by RNA in gene regulatory elements.

    Science.gov (United States)

    Sigova, Alla A; Abraham, Brian J; Ji, Xiong; Molinie, Benoit; Hannett, Nancy M; Guo, Yang Eric; Jangi, Mohini; Giallourakis, Cosmas C; Sharp, Phillip A; Young, Richard A

    2015-11-20

    Transcription factors (TFs) bind specific sequences in promoter-proximal and -distal DNA elements to regulate gene transcription. RNA is transcribed from both of these DNA elements, and some DNA binding TFs bind RNA. Hence, RNA transcribed from regulatory elements may contribute to stable TF occupancy at these sites. We show that the ubiquitously expressed TF Yin-Yang 1 (YY1) binds to both gene regulatory elements and their associated RNA species across the entire genome. Reduced transcription of regulatory elements diminishes YY1 occupancy, whereas artificial tethering of RNA enhances YY1 occupancy at these elements. We propose that RNA makes a modest but important contribution to the maintenance of certain TFs at gene regulatory elements and suggest that transcription of regulatory elements produces a positive-feedback loop that contributes to the stability of gene expression programs. Copyright © 2015, American Association for the Advancement of Science.

  5. Transcriptional Regulatory Network Analysis of MYB Transcription Factor Family Genes in Rice

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    Shuchi eSmita

    2015-12-01

    Full Text Available MYB transcription factor (TF is one of the largest TF families and regulates defense responses to various stresses, hormone signaling as well as many metabolic and developmental processes in plants. Understanding these regulatory hierarchies of gene expression networks in response to developmental and environmental cues is a major challenge due to the complex interactions between the genetic elements. Correlation analyses are useful to unravel co-regulated gene pairs governing biological process as well as identification of new candidate hub genes in response to these complex processes. High throughput expression profiling data are highly useful for construction of co-expression networks. In the present study, we utilized transcriptome data for comprehensive regulatory network studies of MYB TFs by top down and guide gene approaches. More than 50% of OsMYBs were strongly correlated under fifty experimental conditions with 51 hub genes via top down approach. Further, clusters were identified using Markov Clustering (MCL. To maximize the clustering performance, parameter evaluation of the MCL inflation score (I was performed in terms of enriched GO categories by measuring F-score. Comparison of co-expressed cluster and clads analyzed from phylogenetic analysis signifies their evolutionarily conserved co-regulatory role. We utilized compendium of known interaction and biological role with Gene Ontology enrichment analysis to hypothesize function of coexpressed OsMYBs. In the other part, the transcriptional regulatory network analysis by guide gene approach revealed 40 putative targets of 26 OsMYB TF hubs with high correlation value utilizing 815 microarray data. The putative targets with MYB-binding cis-elements enrichment in their promoter region, functional co-occurrence as well as nuclear localization supports our finding. Specially, enrichment of MYB binding regions involved in drought-inducibility implying their regulatory role in drought

  6. Transcription regulatory networks analysis using CAGE

    KAUST Repository

    Tegné r, Jesper N.; Bjö rkegren, Johan L M; Ravasi, Timothy; Bajic, Vladimir

    2009-01-01

    and the fine interplay between regulatory proteins and the promoter structure governing the combinatorial regulation of gene expression. In this chapter we review how the CAGE data can be integrated with other measurements such as expression, physical

  7. A deeper look into transcription regulatory code by preferred pair distance templates for transcription factor binding sites

    KAUST Repository

    Kulakovskiy, Ivan V.

    2011-08-18

    Motivation: Modern experimental methods provide substantial information on protein-DNA recognition. Studying arrangements of transcription factor binding sites (TFBSs) of interacting transcription factors (TFs) advances understanding of the transcription regulatory code. Results: We constructed binding motifs for TFs forming a complex with HIF-1α at the erythropoietin 3\\'-enhancer. Corresponding TFBSs were predicted in the segments around transcription start sites (TSSs) of all human genes. Using the genome-wide set of regulatory regions, we observed several strongly preferred distances between hypoxia-responsive element (HRE) and binding sites of a particular cofactor protein. The set of preferred distances was called as a preferred pair distance template (PPDT). PPDT dramatically depended on the TF and orientation of its binding sites relative to HRE. PPDT evaluated from the genome-wide set of regulatory sequences was used to detect significant PPDT-consistent binding site pairs in regulatory regions of hypoxia-responsive genes. We believe PPDT can help to reveal the layout of eukaryotic regulatory segments. © The Author 2011. Published by Oxford University Press. All rights reserved.

  8. Transcription profile of Escherichia coli: genomic SELEX search for regulatory targets of transcription factors.

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    Ishihama, Akira; Shimada, Tomohiro; Yamazaki, Yukiko

    2016-03-18

    Bacterial genomes are transcribed by DNA-dependent RNA polymerase (RNAP), which achieves gene selectivity through interaction with sigma factors that recognize promoters, and transcription factors (TFs) that control the activity and specificity of RNAP holoenzyme. To understand the molecular mechanisms of transcriptional regulation, the identification of regulatory targets is needed for all these factors. We then performed genomic SELEX screenings of targets under the control of each sigma factor and each TF. Here we describe the assembly of 156 SELEX patterns of a total of 116 TFs performed in the presence and absence of effector ligands. The results reveal several novel concepts: (i) each TF regulates more targets than hitherto recognized; (ii) each promoter is regulated by more TFs than hitherto recognized; and (iii) the binding sites of some TFs are located within operons and even inside open reading frames. The binding sites of a set of global regulators, including cAMP receptor protein, LeuO and Lrp, overlap with those of the silencer H-NS, suggesting that certain global regulators play an anti-silencing role. To facilitate sharing of these accumulated SELEX datasets with the research community, we compiled a database, 'Transcription Profile of Escherichia coli' (www.shigen.nig.ac.jp/ecoli/tec/). © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Conserved Transcriptional Regulatory Programs Underlying Rice and Barley Germination

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    Lin, Li; Tian, Shulan; Kaeppler, Shawn; Liu, Zongrang; An, Yong-Qiang (Charles)

    2014-01-01

    Germination is a biological process important to plant development and agricultural production. Barley and rice diverged 50 million years ago, but share a similar germination process. To gain insight into the conservation of their underlying gene regulatory programs, we compared transcriptomes of barley and rice at start, middle and end points of germination, and revealed that germination regulated barley and rice genes (BRs) diverged significantly in expression patterns and/or protein sequences. However, BRs with higher protein sequence similarity tended to have more conserved expression patterns. We identified and characterized 316 sets of conserved barley and rice genes (cBRs) with high similarity in both protein sequences and expression patterns, and provided a comprehensive depiction of the transcriptional regulatory program conserved in barley and rice germination at gene, pathway and systems levels. The cBRs encoded proteins involved in a variety of biological pathways and had a wide range of expression patterns. The cBRs encoding key regulatory components in signaling pathways often had diverse expression patterns. Early germination up-regulation of cell wall metabolic pathway and peroxidases, and late germination up-regulation of chromatin structure and remodeling pathways were conserved in both barley and rice. Protein sequence and expression pattern of a gene change quickly if it is not subjected to a functional constraint. Preserving germination-regulated expression patterns and protein sequences of those cBRs for 50 million years strongly suggests that the cBRs are functionally significant and equivalent in germination, and contribute to the ancient characteristics of germination preserved in barley and rice. The functional significance and equivalence of the cBR genes predicted here can serve as a foundation to further characterize their biological functions and facilitate bridging rice and barley germination research with greater confidence. PMID

  10. Regulatory hotspots in the malaria parasite genome dictate transcriptional variation.

    Directory of Open Access Journals (Sweden)

    Joseph M Gonzales

    2008-09-01

    Full Text Available The determinants of transcriptional regulation in malaria parasites remain elusive. The presence of a well-characterized gene expression cascade shared by different Plasmodium falciparum strains could imply that transcriptional regulation and its natural variation do not contribute significantly to the evolution of parasite drug resistance. To clarify the role of transcriptional variation as a source of stain-specific diversity in the most deadly malaria species and to find genetic loci that dictate variations in gene expression, we examined genome-wide expression level polymorphisms (ELPs in a genetic cross between phenotypically distinct parasite clones. Significant variation in gene expression is observed through direct co-hybridizations of RNA from different P. falciparum clones. Nearly 18% of genes were regulated by a significant expression quantitative trait locus. The genetic determinants of most of these ELPs resided in hotspots that are physically distant from their targets. The most prominent regulatory locus, influencing 269 transcripts, coincided with a Chromosome 5 amplification event carrying the drug resistance gene, pfmdr1, and 13 other genes. Drug selection pressure in the Dd2 parental clone lineage led not only to a copy number change in the pfmdr1 gene but also to an increased copy number of putative neighboring regulatory factors that, in turn, broadly influence the transcriptional network. Previously unrecognized transcriptional variation, controlled by polymorphic regulatory genes and possibly master regulators within large copy number variants, contributes to sweeping phenotypic evolution in drug-resistant malaria parasites.

  11. Iterative reconstruction of transcriptional regulatory networks: an algorithmic approach.

    Directory of Open Access Journals (Sweden)

    Christian L Barrett

    2006-05-01

    Full Text Available The number of complete, publicly available genome sequences is now greater than 200, and this number is expected to rapidly grow in the near future as metagenomic and environmental sequencing efforts escalate and the cost of sequencing drops. In order to make use of this data for understanding particular organisms and for discerning general principles about how organisms function, it will be necessary to reconstruct their various biochemical reaction networks. Principal among these will be transcriptional regulatory networks. Given the physical and logical complexity of these networks, the various sources of (often noisy data that can be utilized for their elucidation, the monetary costs involved, and the huge number of potential experiments approximately 10(12 that can be performed, experiment design algorithms will be necessary for synthesizing the various computational and experimental data to maximize the efficiency of regulatory network reconstruction. This paper presents an algorithm for experimental design to systematically and efficiently reconstruct transcriptional regulatory networks. It is meant to be applied iteratively in conjunction with an experimental laboratory component. The algorithm is presented here in the context of reconstructing transcriptional regulation for metabolism in Escherichia coli, and, through a retrospective analysis with previously performed experiments, we show that the produced experiment designs conform to how a human would design experiments. The algorithm is able to utilize probability estimates based on a wide range of computational and experimental sources to suggest experiments with the highest potential of discovering the greatest amount of new regulatory knowledge.

  12. Memory functions reveal structural properties of gene regulatory networks

    Science.gov (United States)

    Perez-Carrasco, Ruben

    2018-01-01

    Gene regulatory networks (GRNs) control cellular function and decision making during tissue development and homeostasis. Mathematical tools based on dynamical systems theory are often used to model these networks, but the size and complexity of these models mean that their behaviour is not always intuitive and the underlying mechanisms can be difficult to decipher. For this reason, methods that simplify and aid exploration of complex networks are necessary. To this end we develop a broadly applicable form of the Zwanzig-Mori projection. By first converting a thermodynamic state ensemble model of gene regulation into mass action reactions we derive a general method that produces a set of time evolution equations for a subset of components of a network. The influence of the rest of the network, the bulk, is captured by memory functions that describe how the subnetwork reacts to its own past state via components in the bulk. These memory functions provide probes of near-steady state dynamics, revealing information not easily accessible otherwise. We illustrate the method on a simple cross-repressive transcriptional motif to show that memory functions not only simplify the analysis of the subnetwork but also have a natural interpretation. We then apply the approach to a GRN from the vertebrate neural tube, a well characterised developmental transcriptional network composed of four interacting transcription factors. The memory functions reveal the function of specific links within the neural tube network and identify features of the regulatory structure that specifically increase the robustness of the network to initial conditions. Taken together, the study provides evidence that Zwanzig-Mori projections offer powerful and effective tools for simplifying and exploring the behaviour of GRNs. PMID:29470492

  13. A systems level approach reveals new gene regulatory modules in the developing ear

    OpenAIRE

    Chen, Jingchen; Tambalo, Monica; Barembaum, Meyer; Ranganathan, Ramya; Simões-Costa, Marcos; Bronner, Marianne E.; Streit, Andrea

    2017-01-01

    The inner ear is a complex vertebrate sense organ, yet it arises from a simple epithelium, the otic placode. Specification towards otic fate requires diverse signals and transcriptional inputs that act sequentially and/or in parallel. Using the chick embryo, we uncover novel genes in the gene regulatory network underlying otic commitment and reveal dynamic changes in gene expression. Functional analysis of selected transcription factors reveals the genetic hierarchy underlying the transition ...

  14. Global transcriptional regulatory network for Escherichia coli robustly connects gene expression to transcription factor activities

    DEFF Research Database (Denmark)

    Fang, Xin; Sastry, Anand; Mih, Nathan

    2017-01-01

    Transcriptional regulatory networks (TRNs) have been studied intensely for >25 y. Yet, even for the Escherichia coli TRN-probably the best characterized TRN-several questions remain. Here, we address three questions: (i) How complete is our knowledge of the E. coli TRN; (ii) how well can we predi...

  15. Elucidating MicroRNA Regulatory Networks Using Transcriptional, Post-transcriptional, and Histone Modification Measurements

    Directory of Open Access Journals (Sweden)

    Sara J.C. Gosline

    2016-01-01

    Full Text Available MicroRNAs (miRNAs regulate diverse biological processes by repressing mRNAs, but their modest effects on direct targets, together with their participation in larger regulatory networks, make it challenging to delineate miRNA-mediated effects. Here, we describe an approach to characterizing miRNA-regulatory networks by systematically profiling transcriptional, post-transcriptional and epigenetic activity in a pair of isogenic murine fibroblast cell lines with and without Dicer expression. By RNA sequencing (RNA-seq and CLIP (crosslinking followed by immunoprecipitation sequencing (CLIP-seq, we found that most of the changes induced by global miRNA loss occur at the level of transcription. We then introduced a network modeling approach that integrated these data with epigenetic data to identify specific miRNA-regulated transcription factors that explain the impact of miRNA perturbation on gene expression. In total, we demonstrate that combining multiple genome-wide datasets spanning diverse regulatory modes enables accurate delineation of the downstream miRNA-regulated transcriptional network and establishes a model for studying similar networks in other systems.

  16. Uncovering Transcriptional Regulatory Networks by Sparse Bayesian Factor Model

    Directory of Open Access Journals (Sweden)

    Qi Yuan(Alan

    2010-01-01

    Full Text Available Abstract The problem of uncovering transcriptional regulation by transcription factors (TFs based on microarray data is considered. A novel Bayesian sparse correlated rectified factor model (BSCRFM is proposed that models the unknown TF protein level activity, the correlated regulations between TFs, and the sparse nature of TF-regulated genes. The model admits prior knowledge from existing database regarding TF-regulated target genes based on a sparse prior and through a developed Gibbs sampling algorithm, a context-specific transcriptional regulatory network specific to the experimental condition of the microarray data can be obtained. The proposed model and the Gibbs sampling algorithm were evaluated on the simulated systems, and results demonstrated the validity and effectiveness of the proposed approach. The proposed model was then applied to the breast cancer microarray data of patients with Estrogen Receptor positive ( status and Estrogen Receptor negative ( status, respectively.

  17. Analysis of a Plant Transcriptional Regulatory Network Using Transient Expression Systems.

    Science.gov (United States)

    Díaz-Triviño, Sara; Long, Yuchen; Scheres, Ben; Blilou, Ikram

    2017-01-01

    In plant biology, transient expression systems have become valuable approaches used routinely to rapidly study protein expression, subcellular localization, protein-protein interactions, and transcriptional activity prior to in vivo studies. When studying transcriptional regulation, luciferase reporter assays offer a sensitive readout for assaying promoter behavior in response to different regulators or environmental contexts and to confirm and assess the functional relevance of predicted binding sites in target promoters. This chapter aims to provide detailed methods for using luciferase reporter system as a rapid, efficient, and versatile assay to analyze transcriptional regulation of target genes by transcriptional regulators. We describe a series of optimized transient expression systems consisting of Arabidopsis thaliana protoplasts, infiltrated Nicotiana benthamiana leaves, and human HeLa cells to study the transcriptional regulations of two well-characterized transcriptional regulators SCARECROW (SCR) and SHORT-ROOT (SHR) on one of their targets, CYCLIN D6 (CYCD6).Here, we illustrate similarities and differences in outcomes when using different systems. The plant-based systems revealed that the SCR-SHR complex enhances CYCD6 transcription, while analysis in HeLa cells showed that the complex is not sufficient to strongly induce CYCD6 transcription, suggesting that additional, plant-specific regulators are required for full activation. These results highlight the importance of the system and suggest that including heterologous systems, such as HeLa cells, can provide a more comprehensive analysis of a complex gene regulatory network.

  18. Identification of germline transcriptional regulatory elements in Aedes aegypti

    Science.gov (United States)

    Akbari, Omar S.; Papathanos, Philippos A.; Sandler, Jeremy E.; Kennedy, Katie; Hay, Bruce A.

    2014-02-01

    The mosquito Aedes aegypti is the principal vector for the yellow fever and dengue viruses, and is also responsible for recent outbreaks of the alphavirus chikungunya. Vector control strategies utilizing engineered gene drive systems are being developed as a means of replacing wild, pathogen transmitting mosquitoes with individuals refractory to disease transmission, or bringing about population suppression. Several of these systems, including Medea, UDMEL, and site-specific nucleases, which can be used to drive genes into populations or bring about population suppression, utilize transcriptional regulatory elements that drive germline-specific expression. Here we report the identification of multiple regulatory elements able to drive gene expression specifically in the female germline, or in the male and female germline, in the mosquito Aedes aegypti. These elements can also be used as tools with which to probe the roles of specific genes in germline function and in the early embryo, through overexpression or RNA interference.

  19. Oncogenes Activate an Autonomous Transcriptional Regulatory Circuit That Drives Glioblastoma

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    Dinesh K. Singh

    2017-01-01

    Full Text Available Efforts to identify and target glioblastoma (GBM drivers have primarily focused on receptor tyrosine kinases (RTKs. Clinical benefits, however, have been elusive. Here, we identify an SRY-related box 2 (SOX2 transcriptional regulatory network that is independent of upstream RTKs and capable of driving glioma-initiating cells. We identified oligodendrocyte lineage transcription factor 2 (OLIG2 and zinc-finger E-box binding homeobox 1 (ZEB1, which are frequently co-expressed irrespective of driver mutations, as potential SOX2 targets. In murine glioma models, we show that different combinations of tumor suppressor and oncogene mutations can activate Sox2, Olig2, and Zeb1 expression. We demonstrate that ectopic co-expression of the three transcription factors can transform tumor-suppressor-deficient astrocytes into glioma-initiating cells in the absence of an upstream RTK oncogene. Finally, we demonstrate that the transcriptional inhibitor mithramycin downregulates SOX2 and its target genes, resulting in markedly reduced proliferation of GBM cells in vivo.

  20. Assessing quality and completeness of human transcriptional regulatory pathways on a genome-wide scale

    Directory of Open Access Journals (Sweden)

    Aifantis Iannis

    2011-02-01

    Full Text Available Abstract Background Pathway databases are becoming increasingly important and almost omnipresent in most types of biological and translational research. However, little is known about the quality and completeness of pathways stored in these databases. The present study conducts a comprehensive assessment of transcriptional regulatory pathways in humans for seven well-studied transcription factors: MYC, NOTCH1, BCL6, TP53, AR, STAT1, and RELA. The employed benchmarking methodology first involves integrating genome-wide binding with functional gene expression data to derive direct targets of transcription factors. Then the lists of experimentally obtained direct targets are compared with relevant lists of transcriptional targets from 10 commonly used pathway databases. Results The results of this study show that for the majority of pathway databases, the overlap between experimentally obtained target genes and targets reported in transcriptional regulatory pathway databases is surprisingly small and often is not statistically significant. The only exception is MetaCore pathway database which yields statistically significant intersection with experimental results in 84% cases. Additionally, we suggest that the lists of experimentally derived direct targets obtained in this study can be used to reveal new biological insight in transcriptional regulation and suggest novel putative therapeutic targets in cancer. Conclusions Our study opens a debate on validity of using many popular pathway databases to obtain transcriptional regulatory targets. We conclude that the choice of pathway databases should be informed by solid scientific evidence and rigorous empirical evaluation. Reviewers This article was reviewed by Prof. Wing Hung Wong, Dr. Thiago Motta Venancio (nominated by Dr. L Aravind, and Prof. Geoff J McLachlan.

  1. Global transcriptional regulatory network for Escherichia coli robustly connects gene expression to transcription factor activities

    Science.gov (United States)

    Fang, Xin; Sastry, Anand; Mih, Nathan; Kim, Donghyuk; Tan, Justin; Lloyd, Colton J.; Gao, Ye; Yang, Laurence; Palsson, Bernhard O.

    2017-01-01

    Transcriptional regulatory networks (TRNs) have been studied intensely for >25 y. Yet, even for the Escherichia coli TRN—probably the best characterized TRN—several questions remain. Here, we address three questions: (i) How complete is our knowledge of the E. coli TRN; (ii) how well can we predict gene expression using this TRN; and (iii) how robust is our understanding of the TRN? First, we reconstructed a high-confidence TRN (hiTRN) consisting of 147 transcription factors (TFs) regulating 1,538 transcription units (TUs) encoding 1,764 genes. The 3,797 high-confidence regulatory interactions were collected from published, validated chromatin immunoprecipitation (ChIP) data and RegulonDB. For 21 different TF knockouts, up to 63% of the differentially expressed genes in the hiTRN were traced to the knocked-out TF through regulatory cascades. Second, we trained supervised machine learning algorithms to predict the expression of 1,364 TUs given TF activities using 441 samples. The algorithms accurately predicted condition-specific expression for 86% (1,174 of 1,364) of the TUs, while 193 TUs (14%) were predicted better than random TRNs. Third, we identified 10 regulatory modules whose definitions were robust against changes to the TRN or expression compendium. Using surrogate variable analysis, we also identified three unmodeled factors that systematically influenced gene expression. Our computational workflow comprehensively characterizes the predictive capabilities and systems-level functions of an organism’s TRN from disparate data types. PMID:28874552

  2. Effector Regulatory T Cell Differentiation and Immune Homeostasis Depend on the Transcription Factor Myb.

    Science.gov (United States)

    Dias, Sheila; D'Amico, Angela; Cretney, Erika; Liao, Yang; Tellier, Julie; Bruggeman, Christine; Almeida, Francisca F; Leahy, Jamie; Belz, Gabrielle T; Smyth, Gordon K; Shi, Wei; Nutt, Stephen L

    2017-01-17

    FoxP3-expressing regulatory T (Treg) cells are essential for maintaining immune homeostasis. Activated Treg cells undergo further differentiation into an effector state that highly expresses genes critical for Treg cell function, although how this process is coordinated on a transcriptional level is poorly understood. Here, we demonstrate that mice lacking the transcription factor Myb in Treg cells succumbed to a multi-organ inflammatory disease. Myb was specifically expressed in, and required for the differentiation of, thymus-derived effector Treg cells. The combination of transcriptome and genomic footprint analyses revealed that Myb directly regulated a large proportion of the gene expression specific to effector Treg cells, identifying Myb as a critical component of the gene regulatory network controlling effector Treg cell differentiation and function. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Integration and diversity of the regulatory network composed of Maf and CNC families of transcription factors.

    Science.gov (United States)

    Motohashi, Hozumi; O'Connor, Tania; Katsuoka, Fumiki; Engel, James Douglas; Yamamoto, Masayuki

    2002-07-10

    Recent progress in the analysis of transcriptional regulation has revealed the presence of an exquisite functional network comprising the Maf and Cap 'n' collar (CNC) families of regulatory proteins, many of which have been isolated. Among Maf factors, large Maf proteins are important in the regulation of embryonic development and cell differentiation, whereas small Maf proteins serve as obligatory heterodimeric partner molecules for members of the CNC family. Both Maf homodimers and CNC-small Maf heterodimers bind to the Maf recognition element (MARE). Since the MARE contains a consensus TRE sequence recognized by AP-1, Jun and Fos family members may act to compete or interfere with the function of CNC-small Maf heterodimers. Overall then, the quantitative balance of transcription factors interacting with the MARE determines its transcriptional activity. Many putative MARE-dependent target genes such as those induced by antioxidants and oxidative stress are under concerted regulation by the CNC family member Nrf2, as clearly proven by mouse germline mutagenesis. Since these genes represent a vital aspect of the cellular defense mechanism against oxidative stress, Nrf2-null mutant mice are highly sensitive to xenobiotic and oxidative insults. Deciphering the molecular basis of the regulatory network composed of Maf and CNC families of transcription factors will undoubtedly lead to a new paradigm for the cooperative function of transcription factors.

  4. Prediction of transcriptional regulatory elements for plant hormone responses based on microarray data

    Directory of Open Access Journals (Sweden)

    Yamaguchi-Shinozaki Kazuko

    2011-02-01

    Full Text Available Abstract Background Phytohormones organize plant development and environmental adaptation through cell-to-cell signal transduction, and their action involves transcriptional activation. Recent international efforts to establish and maintain public databases of Arabidopsis microarray data have enabled the utilization of this data in the analysis of various phytohormone responses, providing genome-wide identification of promoters targeted by phytohormones. Results We utilized such microarray data for prediction of cis-regulatory elements with an octamer-based approach. Our test prediction of a drought-responsive RD29A promoter with the aid of microarray data for response to drought, ABA and overexpression of DREB1A, a key regulator of cold and drought response, provided reasonable results that fit with the experimentally identified regulatory elements. With this succession, we expanded the prediction to various phytohormone responses, including those for abscisic acid, auxin, cytokinin, ethylene, brassinosteroid, jasmonic acid, and salicylic acid, as well as for hydrogen peroxide, drought and DREB1A overexpression. Totally 622 promoters that are activated by phytohormones were subjected to the prediction. In addition, we have assigned putative functions to 53 octamers of the Regulatory Element Group (REG that have been extracted as position-dependent cis-regulatory elements with the aid of their feature of preferential appearance in the promoter region. Conclusions Our prediction of Arabidopsis cis-regulatory elements for phytohormone responses provides guidance for experimental analysis of promoters to reveal the basis of the transcriptional network of phytohormone responses.

  5. Predictive modelling of gene expression from transcriptional regulatory elements.

    Science.gov (United States)

    Budden, David M; Hurley, Daniel G; Crampin, Edmund J

    2015-07-01

    Predictive modelling of gene expression provides a powerful framework for exploring the regulatory logic underpinning transcriptional regulation. Recent studies have demonstrated the utility of such models in identifying dysregulation of gene and miRNA expression associated with abnormal patterns of transcription factor (TF) binding or nucleosomal histone modifications (HMs). Despite the growing popularity of such approaches, a comparative review of the various modelling algorithms and feature extraction methods is lacking. We define and compare three methods of quantifying pairwise gene-TF/HM interactions and discuss their suitability for integrating the heterogeneous chromatin immunoprecipitation (ChIP)-seq binding patterns exhibited by TFs and HMs. We then construct log-linear and ϵ-support vector regression models from various mouse embryonic stem cell (mESC) and human lymphoblastoid (GM12878) data sets, considering both ChIP-seq- and position weight matrix- (PWM)-derived in silico TF-binding. The two algorithms are evaluated both in terms of their modelling prediction accuracy and ability to identify the established regulatory roles of individual TFs and HMs. Our results demonstrate that TF-binding and HMs are highly predictive of gene expression as measured by mRNA transcript abundance, irrespective of algorithm or cell type selection and considering both ChIP-seq and PWM-derived TF-binding. As we encourage other researchers to explore and develop these results, our framework is implemented using open-source software and made available as a preconfigured bootable virtual environment. © The Author 2014. Published by Oxford University Press. For Permissions, please email: journals.permissions@oup.com.

  6. Modelling reveals kinetic advantages of co-transcriptional splicing.

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    Stuart Aitken

    2011-10-01

    Full Text Available Messenger RNA splicing is an essential and complex process for the removal of intron sequences. Whereas the composition of the splicing machinery is mostly known, the kinetics of splicing, the catalytic activity of splicing factors and the interdependency of transcription, splicing and mRNA 3' end formation are less well understood. We propose a stochastic model of splicing kinetics that explains data obtained from high-resolution kinetic analyses of transcription, splicing and 3' end formation during induction of an intron-containing reporter gene in budding yeast. Modelling reveals co-transcriptional splicing to be the most probable and most efficient splicing pathway for the reporter transcripts, due in part to a positive feedback mechanism for co-transcriptional second step splicing. Model comparison is used to assess the alternative representations of reactions. Modelling also indicates the functional coupling of transcription and splicing, because both the rate of initiation of transcription and the probability that step one of splicing occurs co-transcriptionally are reduced, when the second step of splicing is abolished in a mutant reporter.

  7. Modelling reveals kinetic advantages of co-transcriptional splicing.

    Science.gov (United States)

    Aitken, Stuart; Alexander, Ross D; Beggs, Jean D

    2011-10-01

    Messenger RNA splicing is an essential and complex process for the removal of intron sequences. Whereas the composition of the splicing machinery is mostly known, the kinetics of splicing, the catalytic activity of splicing factors and the interdependency of transcription, splicing and mRNA 3' end formation are less well understood. We propose a stochastic model of splicing kinetics that explains data obtained from high-resolution kinetic analyses of transcription, splicing and 3' end formation during induction of an intron-containing reporter gene in budding yeast. Modelling reveals co-transcriptional splicing to be the most probable and most efficient splicing pathway for the reporter transcripts, due in part to a positive feedback mechanism for co-transcriptional second step splicing. Model comparison is used to assess the alternative representations of reactions. Modelling also indicates the functional coupling of transcription and splicing, because both the rate of initiation of transcription and the probability that step one of splicing occurs co-transcriptionally are reduced, when the second step of splicing is abolished in a mutant reporter.

  8. In silico transcriptional regulatory networks involved in tomato fruit ripening

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    Stilianos Arhondakis

    2016-08-01

    Full Text Available ABSTRACTTomato fruit ripening is a complex developmental programme partly mediated by transcriptional regulatory networks. Several transcription factors (TFs which are members of gene families such as MADS-box and ERF were shown to play a significant role in ripening through interconnections into an intricate network. The accumulation of large datasets of expression profiles corresponding to different stages of tomato fruit ripening and the availability of bioinformatics tools for their analysis provide an opportunity to identify TFs which might regulate gene clusters with similar co-expression patterns. We identified two TFs, a SlWRKY22-like and a SlER24 transcriptional activator which were shown to regulate modules by using the LeMoNe algorithm for the analysis of our microarray datasets representing four stages of fruit ripening, breaker, turning, pink and red ripe. The WRKY22-like module comprised a subgroup of six various calcium sensing transcripts with similar to the TF expression patterns according to real time PCR validation. A promoter motif search identified a cis acting element, the W-box, recognized by WRKY TFs that was present in the promoter region of all six calcium sensing genes. Moreover, publicly available microarray datasets of similar ripening stages were also analyzed with LeMoNe resulting in TFs such as SlERF.E1, SlERF.C1, SlERF.B2, SLERF.A2, SlWRKY24, SLWRKY37 and MADS-box/TM29 which might also play an important role in regulation of ripening. These results suggest that the SlWRKY22-like might be involved in the coordinated regulation of expression of the six calcium sensing genes. Conclusively the LeMoNe tool might lead to the identification of putative TF targets for further physiological analysis as regulators of tomato fruit ripening.

  9. A gene regulatory network controlling hhex transcription in the anterior endoderm of the organizer

    Science.gov (United States)

    Rankin, Scott A.; Kormish, Jay; Kofron, Matt; Jegga, Anil; Zorn, Aaron M.

    2011-01-01

    The homeobox gene hhex is one of the earliest markers of the anterior endoderm, which gives rise to foregut organs such as the liver, ventral pancreas, thyroid, and lungs. The regulatory networks controlling hhex transcription are poorly understood. In an extensive cis-regulatory analysis of the Xenopus hhex promoter we determined how the Nodal, Wnt, and BMP pathways and their downstream transcription factors regulate hhex expression in the gastrula organizer. We show that Nodal signaling, present throughout the endoderm, directly activates hhex transcription via FoxH1/Smad2 binding sites in the proximal −0.44 Kb promoter. This positive action of Nodal is suppressed in the ventral-posterior endoderm by Vent 1 and Vent2, homeodomain repressors that are induced by BMP signaling. Maternal Wnt/β-catenin on the dorsal side of the embryo cooperates with Nodal and indirectly activate hhex expression via the homeodomain activators Siamois and Twin. Siamois/Twin stimulate hhex transcription through two mechanisms: 1) They induce the expression of Otx2 and Lim1 and together Siamois, Twin, Otx2 and Lim1 appear to promote hhex transcription through homeobox sites in a Wnt-responsive element located between −0.65 to −0.55 Kb of the hhex promoter. 2) Siamois/Twin also induce the expression of the BMP-antagonists Chordin and Noggin, which are required to exclude Vents from the organizer allowing hhex transcription. This work reveals a complex network regulating anterior endoderm transcription in the early embryo. PMID:21215263

  10. Information processing in the transcriptional regulatory network of yeast: Functional robustness

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    Dehmer Matthias

    2009-03-01

    Full Text Available Abstract Background Gene networks are considered to represent various aspects of molecular biological systems meaningfully because they naturally provide a systems perspective of molecular interactions. In this respect, the functional understanding of the transcriptional regulatory network is considered as key to elucidate the functional organization of an organism. Results In this paper we study the functional robustness of the transcriptional regulatory network of S. cerevisiae. We model the information processing in the network as a first order Markov chain and study the influence of single gene perturbations on the global, asymptotic communication among genes. Modification in the communication is measured by an information theoretic measure allowing to predict genes that are 'fragile' with respect to single gene knockouts. Our results demonstrate that the predicted set of fragile genes contains a statistically significant enrichment of so called essential genes that are experimentally found to be necessary to ensure vital yeast. Further, a structural analysis of the transcriptional regulatory network reveals that there are significant differences between fragile genes, hub genes and genes with a high betweenness centrality value. Conclusion Our study does not only demonstrate that a combination of graph theoretical, information theoretical and statistical methods leads to meaningful biological results but also that such methods allow to study information processing in gene networks instead of just their structural properties.

  11. Identification of transcript regulatory patterns in cell differentiation.

    Science.gov (United States)

    Gusnanto, Arief; Gosling, John Paul; Pope, Christopher

    2017-10-15

    Studying transcript regulatory patterns in cell differentiation is critical in understanding its complex nature of the formation and function of different cell types. This is done usually by measuring gene expression at different stages of the cell differentiation. However, if the gene expression data available are only from the mature cells, we have some challenges in identifying transcript regulatory patterns that govern the cell differentiation. We propose to exploit the information of the lineage of cell differentiation in terms of correlation structure between cell types. We assume that two different cell types that are close in the lineage will exhibit many common genes that are co-expressed relative to those that are far in the lineage. Current analysis methods tend to ignore this correlation by testing for differential expression assuming some sort of independence between cell types. We employ a Bayesian approach to estimate the posterior distribution of the mean of expression in each cell type, by taking into account the cell formation path in the lineage. This enables us to infer genes that are specific in each cell type, indicating the genes are involved in directing the cell differentiation to that particular cell type. We illustrate the method using gene expression data from a study of haematopoiesis. R codes to perform the analysis are available in http://www1.maths.leeds.ac.uk/∼arief/R/CellDiff/. a.gusnanto@leeds.ac.uk. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  12. Multiple post-transcriptional regulatory mechanisms in ferritin gene expression

    International Nuclear Information System (INIS)

    Mattia, E.; Den Blaauwen, J.; Van Renswoude, J.; Ashwell, G.

    1989-01-01

    The authors have investigated the mechanisms involved in the regulation of ferritin biosynthesis in K562 human erythroleukemia cells during prolonged exposure to iron. They show that, upon addition of hemin (an efficient iron donor) to the cell culture, the rate of ferritin biosynthesis reaches a maximum after a few hours and then decreases. During a 24-hr incubation with the iron donor the concentrations of total ferritin heavy (H) and light (L) subunit mRNAs rise 2- to 5-fold and 2- to 3-fold, respectively, over the control values, while the amount of the protein increases 10- to 30-fold. The hemin-induced increment in ferritin subunit mRNA is not prevented by deferoxamine, suggesting that it is not directly mediated by chelatable iron. In vitro nuclear transcription analyses performed on nuclei isolated from control cells and cells grown in the presence of hemin indicate that the rates of synthesis of H- and L-subunit mRNAs remain constant. They conclude that iron-induced ferritin biosynthesis is governed by multiple post-transcriptional regulatory mechanisms. They propose that exposure of cells to iron leads to stabilization of ferritin mRNAs, in addition to activation and translation of stored H-and L-subunit mRNAs

  13. Inferring the role of transcription factors in regulatory networks

    Directory of Open Access Journals (Sweden)

    Le Borgne Michel

    2008-05-01

    Full Text Available Abstract Background Expression profiles obtained from multiple perturbation experiments are increasingly used to reconstruct transcriptional regulatory networks, from well studied, simple organisms up to higher eukaryotes. Admittedly, a key ingredient in developing a reconstruction method is its ability to integrate heterogeneous sources of information, as well as to comply with practical observability issues: measurements can be scarce or noisy. In this work, we show how to combine a network of genetic regulations with a set of expression profiles, in order to infer the functional effect of the regulations, as inducer or repressor. Our approach is based on a consistency rule between a network and the signs of variation given by expression arrays. Results We evaluate our approach in several settings of increasing complexity. First, we generate artificial expression data on a transcriptional network of E. coli extracted from the literature (1529 nodes and 3802 edges, and we estimate that 30% of the regulations can be annotated with about 30 profiles. We additionally prove that at most 40.8% of the network can be inferred using our approach. Second, we use this network in order to validate the predictions obtained with a compendium of real expression profiles. We describe a filtering algorithm that generates particularly reliable predictions. Finally, we apply our inference approach to S. cerevisiae transcriptional network (2419 nodes and 4344 interactions, by combining ChIP-chip data and 15 expression profiles. We are able to detect and isolate inconsistencies between the expression profiles and a significant portion of the model (15% of all the interactions. In addition, we report predictions for 14.5% of all interactions. Conclusion Our approach does not require accurate expression levels nor times series. Nevertheless, we show on both data, real and artificial, that a relatively small number of perturbation experiments are enough to determine

  14. Predictive regulatory models in Drosophila melanogaster by integrative inference of transcriptional networks

    Science.gov (United States)

    Marbach, Daniel; Roy, Sushmita; Ay, Ferhat; Meyer, Patrick E.; Candeias, Rogerio; Kahveci, Tamer; Bristow, Christopher A.; Kellis, Manolis

    2012-01-01

    Gaining insights on gene regulation from large-scale functional data sets is a grand challenge in systems biology. In this article, we develop and apply methods for transcriptional regulatory network inference from diverse functional genomics data sets and demonstrate their value for gene function and gene expression prediction. We formulate the network inference problem in a machine-learning framework and use both supervised and unsupervised methods to predict regulatory edges by integrating transcription factor (TF) binding, evolutionarily conserved sequence motifs, gene expression, and chromatin modification data sets as input features. Applying these methods to Drosophila melanogaster, we predict ∼300,000 regulatory edges in a network of ∼600 TFs and 12,000 target genes. We validate our predictions using known regulatory interactions, gene functional annotations, tissue-specific expression, protein–protein interactions, and three-dimensional maps of chromosome conformation. We use the inferred network to identify putative functions for hundreds of previously uncharacterized genes, including many in nervous system development, which are independently confirmed based on their tissue-specific expression patterns. Last, we use the regulatory network to predict target gene expression levels as a function of TF expression, and find significantly higher predictive power for integrative networks than for motif or ChIP-based networks. Our work reveals the complementarity between physical evidence of regulatory interactions (TF binding, motif conservation) and functional evidence (coordinated expression or chromatin patterns) and demonstrates the power of data integration for network inference and studies of gene regulation at the systems level. PMID:22456606

  15. Predictive regulatory models in Drosophila melanogaster by integrative inference of transcriptional networks.

    Science.gov (United States)

    Marbach, Daniel; Roy, Sushmita; Ay, Ferhat; Meyer, Patrick E; Candeias, Rogerio; Kahveci, Tamer; Bristow, Christopher A; Kellis, Manolis

    2012-07-01

    Gaining insights on gene regulation from large-scale functional data sets is a grand challenge in systems biology. In this article, we develop and apply methods for transcriptional regulatory network inference from diverse functional genomics data sets and demonstrate their value for gene function and gene expression prediction. We formulate the network inference problem in a machine-learning framework and use both supervised and unsupervised methods to predict regulatory edges by integrating transcription factor (TF) binding, evolutionarily conserved sequence motifs, gene expression, and chromatin modification data sets as input features. Applying these methods to Drosophila melanogaster, we predict ∼300,000 regulatory edges in a network of ∼600 TFs and 12,000 target genes. We validate our predictions using known regulatory interactions, gene functional annotations, tissue-specific expression, protein-protein interactions, and three-dimensional maps of chromosome conformation. We use the inferred network to identify putative functions for hundreds of previously uncharacterized genes, including many in nervous system development, which are independently confirmed based on their tissue-specific expression patterns. Last, we use the regulatory network to predict target gene expression levels as a function of TF expression, and find significantly higher predictive power for integrative networks than for motif or ChIP-based networks. Our work reveals the complementarity between physical evidence of regulatory interactions (TF binding, motif conservation) and functional evidence (coordinated expression or chromatin patterns) and demonstrates the power of data integration for network inference and studies of gene regulation at the systems level.

  16. Deciphering Fur transcriptional regulatory network highlights its complex role beyond iron metabolism in Escherichia coli

    DEFF Research Database (Denmark)

    Seo, Sang Woo; Kim, Donghyuk; Latif, Haythem

    2014-01-01

    The ferric uptake regulator (Fur) plays a critical role in the transcriptional regulation of iron metabolism. However, the full regulatory potential of Fur remains undefined. Here we comprehensively reconstruct the Fur transcriptional regulatory network in Escherichia coli K-12 MG1655 in response...

  17. Redefining the transcriptional regulatory dynamics of classically and alternatively activated macrophages by deepCAGE transcriptomics

    KAUST Repository

    Roy, S.

    2015-06-27

    Classically or alternatively activated macrophages (M1 and M2, respectively) play distinct and important roles for microbiocidal activity, regulation of inflammation and tissue homeostasis. Despite this, their transcriptional regulatory dynamics are poorly understood. Using promoter-level expression profiling by non-biased deepCAGE we have studied the transcriptional dynamics of classically and alternatively activated macrophages. Transcription factor (TF) binding motif activity analysis revealed four motifs, NFKB1_REL_RELA, IRF1,2, IRF7 and TBP that are commonly activated but have distinct activity dynamics in M1 and M2 activation. We observe matching changes in the expression profiles of the corresponding TFs and show that only a restricted set of TFs change expression. There is an overall drastic and transient up-regulation in M1 and a weaker and more sustainable up-regulation in M2. Novel TFs, such as Thap6, Maff, (M1) and Hivep1, Nfil3, Prdm1, (M2) among others, were suggested to be involved in the activation processes. Additionally, 52 (M1) and 67 (M2) novel differentially expressed genes and, for the first time, several differentially expressed long non-coding RNA (lncRNA) transcriptome markers were identified. In conclusion, the finding of novel motifs, TFs and protein-coding and lncRNA genes is an important step forward to fully understand the transcriptional machinery of macrophage activation.

  18. Heart morphogenesis gene regulatory networks revealed by temporal expression analysis.

    Science.gov (United States)

    Hill, Jonathon T; Demarest, Bradley; Gorsi, Bushra; Smith, Megan; Yost, H Joseph

    2017-10-01

    During embryogenesis the heart forms as a linear tube that then undergoes multiple simultaneous morphogenetic events to obtain its mature shape. To understand the gene regulatory networks (GRNs) driving this phase of heart development, during which many congenital heart disease malformations likely arise, we conducted an RNA-seq timecourse in zebrafish from 30 hpf to 72 hpf and identified 5861 genes with altered expression. We clustered the genes by temporal expression pattern, identified transcription factor binding motifs enriched in each cluster, and generated a model GRN for the major gene batteries in heart morphogenesis. This approach predicted hundreds of regulatory interactions and found batteries enriched in specific cell and tissue types, indicating that the approach can be used to narrow the search for novel genetic markers and regulatory interactions. Subsequent analyses confirmed the GRN using two mutants, Tbx5 and nkx2-5 , and identified sets of duplicated zebrafish genes that do not show temporal subfunctionalization. This dataset provides an essential resource for future studies on the genetic/epigenetic pathways implicated in congenital heart defects and the mechanisms of cardiac transcriptional regulation. © 2017. Published by The Company of Biologists Ltd.

  19. Pleiotropy constrains the evolution of protein but not regulatory sequences in a transcription regulatory network influencing complex social behaviours

    Directory of Open Access Journals (Sweden)

    Daria eMolodtsova

    2014-12-01

    Full Text Available It is increasingly apparent that genes and networks that influence complex behaviour are evolutionary conserved, which is paradoxical considering that behaviour is labile over evolutionary timescales. How does adaptive change in behaviour arise if behaviour is controlled by conserved, pleiotropic, and likely evolutionary constrained genes? Pleiotropy and connectedness are known to constrain the general rate of protein evolution, prompting some to suggest that the evolution of complex traits, including behaviour, is fuelled by regulatory sequence evolution. However, we seldom have data on the strength of selection on mutations in coding and regulatory sequences, and this hinders our ability to study how pleiotropy influences coding and regulatory sequence evolution. Here we use population genomics to estimate the strength of selection on coding and regulatory mutations for a transcriptional regulatory network that influences complex behaviour of honey bees. We found that replacement mutations in highly connected transcription factors and target genes experience significantly stronger negative selection relative to weakly connected transcription factors and targets. Adaptively evolving proteins were significantly more likely to reside at the periphery of the regulatory network, while proteins with signs of negative selection were near the core of the network. Interestingly, connectedness and network structure had minimal influence on the strength of selection on putative regulatory sequences for both transcription factors and their targets. Our study indicates that adaptive evolution of complex behaviour can arise because of positive selection on protein-coding mutations in peripheral genes, and on regulatory sequence mutations in both transcription factors and their targets throughout the network.

  20. Transcriptional regulation of rod photoreceptor homeostasis revealed by in vivo NRL targetome analysis.

    Directory of Open Access Journals (Sweden)

    Hong Hao

    Full Text Available A stringent control of homeostasis is critical for functional maintenance and survival of neurons. In the mammalian retina, the basic motif leucine zipper transcription factor NRL determines rod versus cone photoreceptor cell fate and activates the expression of many rod-specific genes. Here, we report an integrated analysis of NRL-centered gene regulatory network by coupling chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq data from Illumina and ABI platforms with global expression profiling and in vivo knockdown studies. We identified approximately 300 direct NRL target genes. Of these, 22 NRL targets are associated with human retinal dystrophies, whereas 95 mapped to regions of as yet uncloned retinal disease loci. In silico analysis of NRL ChIP-Seq peak sequences revealed an enrichment of distinct sets of transcription factor binding sites. Specifically, we discovered that genes involved in photoreceptor function include binding sites for both NRL and homeodomain protein CRX. Evaluation of 26 ChIP-Seq regions validated their enhancer functions in reporter assays. In vivo knockdown of 16 NRL target genes resulted in death or abnormal morphology of rod photoreceptors, suggesting their importance in maintaining retinal function. We also identified histone demethylase Kdm5b as a novel secondary node in NRL transcriptional hierarchy. Exon array analysis of flow-sorted photoreceptors in which Kdm5b was knocked down by shRNA indicated its role in regulating rod-expressed genes. Our studies identify candidate genes for retinal dystrophies, define cis-regulatory module(s for photoreceptor-expressed genes and provide a framework for decoding transcriptional regulatory networks that dictate rod homeostasis.

  1. Redefining the transcriptional regulatory dynamics of classically and alternatively activated macrophages by deepCAGE transcriptomics

    KAUST Repository

    Roy, S.; Schmeier, S.; Arner, E.; Alam, Tanvir; Parihar, S. P.; Ozturk, M.; Tamgue, O.; Kawaji, H.; de Hoon, M. J. L.; Itoh, M.; Lassmann, T.; Carninci, P.; Hayashizaki, Y.; Forrest, A. R. R.; Bajic, Vladimir B.; Guler, R.; Consortium, F.; Brombacher, F.; Suzuki, H.

    2015-01-01

    Classically or alternatively activated macrophages (M1 and M2, respectively) play distinct and important roles for microbiocidal activity, regulation of inflammation and tissue homeostasis. Despite this, their transcriptional regulatory dynamics

  2. Medusa structure of the gene regulatory network: dominance of transcription factors in cancer subtype classification.

    Science.gov (United States)

    Guo, Yuchun; Feng, Ying; Trivedi, Niraj S; Huang, Sui

    2011-05-01

    Gene expression profiles consisting of ten thousands of transcripts are used for clustering of tissue, such as tumors, into subtypes, often without considering the underlying reason that the distinct patterns of expression arise because of constraints in the realization of gene expression profiles imposed by the gene regulatory network. The topology of this network has been suggested to consist of a regulatory core of genes represented most prominently by transcription factors (TFs) and microRNAs, that influence the expression of other genes, and of a periphery of 'enslaved' effector genes that are regulated but not regulating. This 'medusa' architecture implies that the core genes are much stronger determinants of the realized gene expression profiles. To test this hypothesis, we examined the clustering of gene expression profiles into known tumor types to quantitatively demonstrate that TFs, and even more pronounced, microRNAs, are much stronger discriminators of tumor type specific gene expression patterns than a same number of randomly selected or metabolic genes. These findings lend support to the hypothesis of a medusa architecture and of the canalizing nature of regulation by microRNAs. They also reveal the degree of freedom for the expression of peripheral genes that are less stringently associated with a tissue type specific global gene expression profile.

  3. A comprehensive analysis of microProteins reveals their potentially widespread mechanism of transcriptional regulation

    NARCIS (Netherlands)

    Magnani, Enrico; de Klein, Niek; Nam, Hye-In; Kim, Jung-Gun; Pham, Kimberly; Fiume, Elisa; Mudgett, Mary Beth; Rhee, Seung Yon

    2014-01-01

    Truncated transcription factor-like proteins called microProteins (miPs) can modulate transcription factor activities, thereby increasing transcriptional regulatory complexity. To understand their prevalence, evolution, and function, we predicted over 400 genes that encode putative miPs from

  4. Comprehensive transcriptional profiling of NaCl-stressed Arabidopsis roots reveals novel classes of responsive genes

    Directory of Open Access Journals (Sweden)

    Deyholos Michael K

    2006-10-01

    Full Text Available Abstract Background Roots are an attractive system for genomic and post-genomic studies of NaCl responses, due to their primary importance to agriculture, and because of their relative structural and biochemical simplicity. Excellent genomic resources have been established for the study of Arabidopsis roots, however, a comprehensive microarray analysis of the root transcriptome following NaCl exposure is required to further understand plant responses to abiotic stress and facilitate future, systems-based analyses of the underlying regulatory networks. Results We used microarrays of 70-mer oligonucleotide probes representing 23,686 Arabidopsis genes to identify root transcripts that changed in relative abundance following 6 h, 24 h, or 48 h of hydroponic exposure to 150 mM NaCl. Enrichment analysis identified groups of structurally or functionally related genes whose members were statistically over-represented among up- or down-regulated transcripts. Our results are consistent with generally observed stress response themes, and highlight potentially important roles for underappreciated gene families, including: several groups of transporters (e.g. MATE, LeOPT1-like; signalling molecules (e.g. PERK kinases, MLO-like receptors, carbohydrate active enzymes (e.g. XTH18, transcription factors (e.g. members of ZIM, WRKY, NAC, and other proteins (e.g. 4CL-like, COMT-like, LOB-Class 1. We verified the NaCl-inducible expression of selected transcription factors and other genes by qRT-PCR. Conclusion Micorarray profiling of NaCl-treated Arabidopsis roots revealed dynamic changes in transcript abundance for at least 20% of the genome, including hundreds of transcription factors, kinases/phosphatases, hormone-related genes, and effectors of homeostasis, all of which highlight the complexity of this stress response. Our identification of these transcriptional responses, and groups of evolutionarily related genes with either similar or divergent

  5. A trans-acting Variant within the Transcription Factor RIM101 Interacts with Genetic Background to Determine its Regulatory Capacity.

    Directory of Open Access Journals (Sweden)

    Timothy Read

    2016-01-01

    Full Text Available Most genetic variants associated with disease occur within regulatory regions of the genome, underscoring the importance of defining the mechanisms underlying differences in regulation of gene expression between individuals. We discovered a pair of co-regulated, divergently oriented transcripts, AQY2 and ncFRE6, that are expressed in one strain of Saccharomyces cerevisiae, ∑1278b, but not in another, S288c. By combining classical genetics techniques with high-throughput sequencing, we identified a trans-acting single nucleotide polymorphism within the transcription factor RIM101 that causes the background-dependent expression of both transcripts. Subsequent RNA-seq experiments revealed that RIM101 regulates many more targets in S288c than in ∑1278b and that deletion of RIM101 in both backgrounds abrogates the majority of differential expression between the strains. Strikingly, only three transcripts undergo a significant change in expression after swapping RIM101 alleles between backgrounds, implying that the differences in the RIM101 allele lead to a remarkably focused transcriptional response. However, hundreds of RIM101-dependent targets undergo a subtle but consistent shift in expression in the S288c RIM101-swapped strain, but not its ∑1278b counterpart. We conclude that ∑1278b may harbor a variant(s that buffers against widespread transcriptional dysregulation upon introduction of a non-native RIM101 allele, emphasizing the importance of accounting for genetic background when assessing the impact of a regulatory variant.

  6. Structural studies of bacterial transcriptional regulatory proteins by multidimensional heteronuclear NMR

    Energy Technology Data Exchange (ETDEWEB)

    Volkman, Brian Finley [Univ. of California, Berkeley, CA (United States)

    1995-02-01

    Nuclear magnetic resonance spectroscopy was used to elucidate detailed structural information for peptide and protein molecules. A small peptide was designed and synthesized, and its three-dimensional structure was calculated using distance information derived from two-dimensional NMR measurements. The peptide was used to induce antibodies in mice, and the cross-reactivity of the antibodies with a related protein was analyzed with enzyme-linked immunosorbent assays. Two proteins which are involved in regulation of transcription in bacteria were also studied. The ferric uptake regulation (Fur) protein is a metal-dependent repressor which controls iron uptake in bacteria. Two- and three-dimensional NMR techniques, coupled with uniform and selective isotope labeling allowed the nearly complete assignment of the resonances of the metal-binding domain of the Fur protein. NTRC is a transcriptional enhancer binding protein whose N-terminal domain is a "receiver domain" in the family of "two-component" regulatory systems. Phosphorylation of the N-terminal domain of NTRC activates the initiation of transcription of aeries encoding proteins involved in nitrogen regulation. Three- and four-dimensional NMR spectroscopy methods have been used to complete the resonance assignments and determine the solution structure of the N-terminal receiver domain of the NTRC protein. Comparison of the solution structure of the NTRC receiver domain with the crystal structures of the homologous protein CheY reveals a very similar fold, with the only significant difference being the position of helix 4 relative to the rest of the protein. The determination of the structure of the NTRC receiver domain is the first step toward understanding a mechanism of signal transduction which is common to many bacterial regulatory systems.

  7. Selection Shapes Transcriptional Logic and Regulatory Specialization in Genetic Networks.

    Science.gov (United States)

    Fogelmark, Karl; Peterson, Carsten; Troein, Carl

    2016-01-01

    Living organisms need to regulate their gene expression in response to environmental signals and internal cues. This is a computational task where genes act as logic gates that connect to form transcriptional networks, which are shaped at all scales by evolution. Large-scale mutations such as gene duplications and deletions add and remove network components, whereas smaller mutations alter the connections between them. Selection determines what mutations are accepted, but its importance for shaping the resulting networks has been debated. To investigate the effects of selection in the shaping of transcriptional networks, we derive transcriptional logic from a combinatorially powerful yet tractable model of the binding between DNA and transcription factors. By evolving the resulting networks based on their ability to function as either a simple decision system or a circadian clock, we obtain information on the regulation and logic rules encoded in functional transcriptional networks. Comparisons are made between networks evolved for different functions, as well as with structurally equivalent but non-functional (neutrally evolved) networks, and predictions are validated against the transcriptional network of E. coli. We find that the logic rules governing gene expression depend on the function performed by the network. Unlike the decision systems, the circadian clocks show strong cooperative binding and negative regulation, which achieves tight temporal control of gene expression. Furthermore, we find that transcription factors act preferentially as either activators or repressors, both when binding multiple sites for a single target gene and globally in the transcriptional networks. This separation into positive and negative regulators requires gene duplications, which highlights the interplay between mutation and selection in shaping the transcriptional networks.

  8. Bayesian error analysis model for reconstructing transcriptional regulatory networks

    OpenAIRE

    Sun, Ning; Carroll, Raymond J.; Zhao, Hongyu

    2006-01-01

    Transcription regulation is a fundamental biological process, and extensive efforts have been made to dissect its mechanisms through direct biological experiments and regulation modeling based on physical–chemical principles and mathematical formulations. Despite these efforts, transcription regulation is yet not well understood because of its complexity and limitations in biological experiments. Recent advances in high throughput technologies have provided substantial amounts and diverse typ...

  9. Biological data warehousing system for identifying transcriptional regulatory sites from gene expressions of microarray data.

    Science.gov (United States)

    Tsou, Ann-Ping; Sun, Yi-Ming; Liu, Chia-Lin; Huang, Hsien-Da; Horng, Jorng-Tzong; Tsai, Meng-Feng; Liu, Baw-Juine

    2006-07-01

    Identification of transcriptional regulatory sites plays an important role in the investigation of gene regulation. For this propose, we designed and implemented a data warehouse to integrate multiple heterogeneous biological data sources with data types such as text-file, XML, image, MySQL database model, and Oracle database model. The utility of the biological data warehouse in predicting transcriptional regulatory sites of coregulated genes was explored using a synexpression group derived from a microarray study. Both of the binding sites of known transcription factors and predicted over-represented (OR) oligonucleotides were demonstrated for the gene group. The potential biological roles of both known nucleotides and one OR nucleotide were demonstrated using bioassays. Therefore, the results from the wet-lab experiments reinforce the power and utility of the data warehouse as an approach to the genome-wide search for important transcription regulatory elements that are the key to many complex biological systems.

  10. Uncovering transcription factor and microRNA risk regulatory pathways associated with osteoarthritis by network analysis.

    Science.gov (United States)

    Song, Zhenhua; Zhang, Chi; He, Lingxiao; Sui, Yanfang; Lin, Xiafei; Pan, Jingjing

    2018-05-01

    Osteoarthritis (OA) is the most common form of joint disease. The development of inflammation have been considered to play a key role during the progression of OA. Regulatory pathways are known to play crucial roles in many pathogenic processes. Thus, deciphering these risk regulatory pathways is critical for elucidating the mechanisms underlying OA. We constructed an OA-specific regulatory network by integrating comprehensive curated transcription and post-transcriptional resource involving transcription factor (TF) and microRNA (miRNA). To deepen our understanding of underlying molecular mechanisms of OA, we developed an integrated systems approach to identify OA-specific risk regulatory pathways. In this study, we identified 89 significantly differentially expressed genes between normal and inflamed areas of OA patients. We found the OA-specific regulatory network was a standard scale-free network with small-world properties. It significant enriched many immune response-related functions including leukocyte differentiation, myeloid differentiation and T cell activation. Finally, 141 risk regulatory pathways were identified based on OA-specific regulatory network, which contains some known regulator of OA. The risk regulatory pathways may provide clues for the etiology of OA and be a potential resource for the discovery of novel OA-associated disease genes. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Computational methods to dissect cis-regulatory transcriptional ...

    Indian Academy of Sciences (India)

    The formation of diverse cell types from an invariant set of genes is governed by biochemical and molecular processes that regulate gene activity. A complete understanding of the regulatory mechanisms of gene expression is the major function of genomics. Computational genomics is a rapidly emerging area for ...

  12. Multiple oxygen tension environments reveal diverse patterns of transcriptional regulation in primary astrocytes.

    Directory of Open Access Journals (Sweden)

    Wayne Chadwick

    Full Text Available The central nervous system normally functions at O(2 levels which would be regarded as hypoxic by most other tissues. However, most in vitro studies of neurons and astrocytes are conducted under hyperoxic conditions without consideration of O(2-dependent cellular adaptation. We analyzed the reactivity of astrocytes to 1, 4 and 9% O(2 tensions compared to the cell culture standard of 20% O(2, to investigate their ability to sense and translate this O(2 information to transcriptional activity. Variance of ambient O(2 tension for rat astrocytes resulted in profound changes in ribosomal activity, cytoskeletal and energy-regulatory mechanisms and cytokine-related signaling. Clustering of transcriptional regulation patterns revealed four distinct response pattern groups that directionally pivoted around the 4% O(2 tension, or demonstrated coherent ascending/decreasing gene expression patterns in response to diverse oxygen tensions. Immune response and cell cycle/cancer-related signaling pathway transcriptomic subsets were significantly activated with increasing hypoxia, whilst hemostatic and cardiovascular signaling mechanisms were attenuated with increasing hypoxia. Our data indicate that variant O(2 tensions induce specific and physiologically-focused transcript regulation patterns that may underpin important physiological mechanisms that connect higher neurological activity to astrocytic function and ambient oxygen environments. These strongly defined patterns demonstrate a strong bias for physiological transcript programs to pivot around the 4% O(2 tension, while uni-modal programs that do not, appear more related to pathological actions. The functional interaction of these transcriptional 'programs' may serve to regulate the dynamic vascular responsivity of the central nervous system during periods of stress or heightened activity.

  13. The future of genome-scale modeling of yeast through integration of a transcriptional regulatory network

    DEFF Research Database (Denmark)

    Liu, Guodong; Marras, Antonio; Nielsen, Jens

    2014-01-01

    regulatory information is necessary to improve the accuracy and predictive ability of metabolic models. Here we review the strategies for the reconstruction of a transcriptional regulatory network (TRN) for yeast and the integration of such a reconstruction into a flux balance analysis-based metabolic model......Metabolism is regulated at multiple levels in response to the changes of internal or external conditions. Transcriptional regulation plays an important role in regulating many metabolic reactions by altering the concentrations of metabolic enzymes. Thus, integration of the transcriptional....... While many large-scale TRN reconstructions have been reported for yeast, these reconstructions still need to be improved regarding the functionality and dynamic property of the regulatory interactions. In addition, mathematical modeling approaches need to be further developed to efficiently integrate...

  14. Olig2 and Hes regulatory dynamics during motor neuron differentiation revealed by single cell transcriptomics.

    Directory of Open Access Journals (Sweden)

    Andreas Sagner

    2018-02-01

    Full Text Available During tissue development, multipotent progenitors differentiate into specific cell types in characteristic spatial and temporal patterns. We addressed the mechanism linking progenitor identity and differentiation rate in the neural tube, where motor neuron (MN progenitors differentiate more rapidly than other progenitors. Using single cell transcriptomics, we defined the transcriptional changes associated with the transition of neural progenitors into MNs. Reconstruction of gene expression dynamics from these data indicate a pivotal role for the MN determinant Olig2 just prior to MN differentiation. Olig2 represses expression of the Notch signaling pathway effectors Hes1 and Hes5. Olig2 repression of Hes5 appears to be direct, via a conserved regulatory element within the Hes5 locus that restricts expression from MN progenitors. These findings reveal a tight coupling between the regulatory networks that control patterning and neuronal differentiation and demonstrate how Olig2 acts as the developmental pacemaker coordinating the spatial and temporal pattern of MN generation.

  15. Sucrose-induced anthocyanin accumulation in vegetative tissue of Petunia plants requires anthocyanin regulatory transcription factors.

    Science.gov (United States)

    Ai, Trinh Ngoc; Naing, Aung Htay; Arun, Muthukrishnan; Lim, Sun-Hyung; Kim, Chang Kil

    2016-11-01

    The effects of three different sucrose concentrations on plant growth and anthocyanin accumulation were examined in non-transgenic (NT) and transgenic (T 2 ) specimens of the Petunia hybrida cultivar 'Mirage rose' that carried the anthocyanin regulatory transcription factors B-Peru+mPAP1 or RsMYB1. Anthocyanin accumulation was not observed in NT plants in any treatments, whereas a range of anthocyanin accumulation was observed in transgenic plants. The anthocyanin content detected in transgenic plants expressing the anthocyanin regulatory transcription factors (B-Peru+mPAP1 or RsMYB1) was higher than that in NT plants. In addition, increasing sucrose concentration strongly enhanced anthocyanin content as shown by quantitative real-time polymerase chain reaction (qRT-PCR) analysis, wherein increased concentrations of sucrose enhanced transcript levels of the transcription factors that are responsible for the induction of biosynthetic genes involved in anthocyanin synthesis; this pattern was not observed in NT plants. In addition, sucrose affected plant growth, although the effects were different between NT and transgenic plants. Taken together, the application of sucrose could enhance anthocyanin production in vegetative tissue of transgenic Petunia carrying anthocyanin regulatory transcription factors, and this study provides insights about interactive effects of sucrose and transcription factors in anthocyanin biosynthesis in the transgenic plant. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  16. Nonchaoticity of Ordinary Differential Equations Describing Autonomous Transcriptional Regulatory Circuits

    International Nuclear Information System (INIS)

    Li Pengfei; Hu Gang; Chen Runsheng

    2008-01-01

    Gene transcriptional regulation (TR) processes are often described by coupled nonlinear ordinary differential equations (ODEs). When the dimension of TR circuits is high (e.g. n ≥ 3) the motions of the corresponding ODEs may, very probably, show self-sustained oscillations and chaos. On the other hand, chaoticity may be harmful for the normal biological functions of TR processes. In this letter we numerically study the dynamics of 3-gene TR ODEs in great detail, and investigate many 4-, 5-, and 10-gene TR systems by randomly choosing figures and parameters in the conventionally accepted ranges. And we find that oscillations are very seldom and no chaotic motion is observed, even if the dimension of systems is sufficiently high (n ≥ 3). It is argued that the observation of nonchaoticity of these ODEs agrees with normal functions of actual TR processes

  17. Genome-Wide Mapping of Collier In Vivo Binding Sites Highlights Its Hierarchical Position in Different Transcription Regulatory Networks.

    Directory of Open Access Journals (Sweden)

    Mathilde de Taffin

    Full Text Available Collier, the single Drosophila COE (Collier/EBF/Olf-1 transcription factor, is required in several developmental processes, including head patterning and specification of muscle and neuron identity during embryogenesis. To identify direct Collier (Col targets in different cell types, we used ChIP-seq to map Col binding sites throughout the genome, at mid-embryogenesis. In vivo Col binding peaks were associated to 415 potential direct target genes. Gene Ontology analysis revealed a strong enrichment in proteins with DNA binding and/or transcription-regulatory properties. Characterization of a selection of candidates, using transgenic CRM-reporter assays, identified direct Col targets in dorso-lateral somatic muscles and specific neuron types in the central nervous system. These data brought new evidence that Col direct control of the expression of the transcription regulators apterous and eyes-absent (eya is critical to specifying neuronal identities. They also showed that cross-regulation between col and eya in muscle progenitor cells is required for specification of muscle identity, revealing a new parallel between the myogenic regulatory networks operating in Drosophila and vertebrates. Col regulation of eya, both in specific muscle and neuronal lineages, may illustrate one mechanism behind the evolutionary diversification of Col biological roles.

  18. Genome-Wide Mapping of Collier In Vivo Binding Sites Highlights Its Hierarchical Position in Different Transcription Regulatory Networks

    Science.gov (United States)

    Dubois, Laurence; Bataillé, Laetitia; Painset, Anaïs; Le Gras, Stéphanie; Jost, Bernard; Crozatier, Michèle; Vincent, Alain

    2015-01-01

    Collier, the single Drosophila COE (Collier/EBF/Olf-1) transcription factor, is required in several developmental processes, including head patterning and specification of muscle and neuron identity during embryogenesis. To identify direct Collier (Col) targets in different cell types, we used ChIP-seq to map Col binding sites throughout the genome, at mid-embryogenesis. In vivo Col binding peaks were associated to 415 potential direct target genes. Gene Ontology analysis revealed a strong enrichment in proteins with DNA binding and/or transcription-regulatory properties. Characterization of a selection of candidates, using transgenic CRM-reporter assays, identified direct Col targets in dorso-lateral somatic muscles and specific neuron types in the central nervous system. These data brought new evidence that Col direct control of the expression of the transcription regulators apterous and eyes-absent (eya) is critical to specifying neuronal identities. They also showed that cross-regulation between col and eya in muscle progenitor cells is required for specification of muscle identity, revealing a new parallel between the myogenic regulatory networks operating in Drosophila and vertebrates. Col regulation of eya, both in specific muscle and neuronal lineages, may illustrate one mechanism behind the evolutionary diversification of Col biological roles. PMID:26204530

  19. Evolutionary dynamics of DNA-binding sites and direct target genes of a floral master regulatory transcription factor [ChIP-Seq

    NARCIS (Netherlands)

    Muiño, J.M.; Bruijn, de S.A.; Vingron, Martin; Angenent, G.C.; Kaufmann, K.

    2015-01-01

    Plant development is controlled by transcription factors (TFs) which form complex gene-regulatory networks. Genome-wide TF DNA-binding studies revealed that these TFs have several thousands of binding sites in the Arabidopsis genome, and may regulate the expression of many genes directly. Given the

  20. Evolutionary dynamics of DNA-binding sites and direct target genes of a floral master regulatory transcription factor [RNA-Seq

    NARCIS (Netherlands)

    Muiño, J.M.; Bruijn, de S.A.; Vingron, Martin; Angenent, G.C.; Kaufmann, Kerstin

    2015-01-01

    Plant development is controlled by transcription factors (TFs) which form complex gene-regulatory networks. Genome-wide TF DNA-binding studies revealed that these TFs have several thousands of binding sites in the Arabidopsis genome, and may regulate the expression of many genes directly. Given the

  1. TIGER: Toolbox for integrating genome-scale metabolic models, expression data, and transcriptional regulatory networks

    Directory of Open Access Journals (Sweden)

    Jensen Paul A

    2011-09-01

    Full Text Available Abstract Background Several methods have been developed for analyzing genome-scale models of metabolism and transcriptional regulation. Many of these methods, such as Flux Balance Analysis, use constrained optimization to predict relationships between metabolic flux and the genes that encode and regulate enzyme activity. Recently, mixed integer programming has been used to encode these gene-protein-reaction (GPR relationships into a single optimization problem, but these techniques are often of limited generality and lack a tool for automating the conversion of rules to a coupled regulatory/metabolic model. Results We present TIGER, a Toolbox for Integrating Genome-scale Metabolism, Expression, and Regulation. TIGER converts a series of generalized, Boolean or multilevel rules into a set of mixed integer inequalities. The package also includes implementations of existing algorithms to integrate high-throughput expression data with genome-scale models of metabolism and transcriptional regulation. We demonstrate how TIGER automates the coupling of a genome-scale metabolic model with GPR logic and models of transcriptional regulation, thereby serving as a platform for algorithm development and large-scale metabolic analysis. Additionally, we demonstrate how TIGER's algorithms can be used to identify inconsistencies and improve existing models of transcriptional regulation with examples from the reconstructed transcriptional regulatory network of Saccharomyces cerevisiae. Conclusion The TIGER package provides a consistent platform for algorithm development and extending existing genome-scale metabolic models with regulatory networks and high-throughput data.

  2. Transcriptional and Epigenetic Regulatory Mechanisms Affecting HTLV-1 Provirus

    Directory of Open Access Journals (Sweden)

    Paola Miyazato

    2016-06-01

    Full Text Available Human T-cell leukemia virus type 1 (HTLV-1 is a retrovirus associated with human diseases, such as adult T-cell leukemia (ATL and HTLV-1-associated myelopathy/Tropic spastic paraparesis (HAM/TSP. As a retrovirus, its life cycle includes a step where HTLV-1 is integrated into the host genomic DNA and forms proviral DNA. In the chronic phase of the infection, HTLV‑1 is known to proliferate as a provirus via the mitotic division of the infected host cells. There are generally tens of thousands of infected clones within an infected individual. They exist not only in peripheral blood, but also in various lymphoid organs. Viral proteins encoded in HTLV-1 genome play a role in the proliferation and survival of the infected cells. As is the case with other chronic viral infections, HTLV-1 gene expression induces the activation of the host immunity against the virus. Thus, the transcription from HTLV-1 provirus needs to be controlled in order to evade the host immune surveillance. There should be a dynamic and complex regulation in vivo, where an equilibrium between viral antigen expression and host immune surveillance is achieved. The mechanisms regulating viral gene expression from the provirus are a key to understanding the persistent/latent infection with HTLV-1 and its pathogenesis. In this article, we would like to review our current understanding on this topic.

  3. A transcription factor collective defines the HSN serotonergic neuron regulatory landscape.

    Science.gov (United States)

    Lloret-Fernández, Carla; Maicas, Miren; Mora-Martínez, Carlos; Artacho, Alejandro; Jimeno-Martín, Ángela; Chirivella, Laura; Weinberg, Peter; Flames, Nuria

    2018-03-22

    Cell differentiation is controlled by individual transcription factors (TFs) that together activate a selection of enhancers in specific cell types. How these combinations of TFs identify and activate their target sequences remains poorly understood. Here, we identify the cis -regulatory transcriptional code that controls the differentiation of serotonergic HSN neurons in Caenorhabditis elegans . Activation of the HSN transcriptome is directly orchestrated by a collective of six TFs. Binding site clusters for this TF collective form a regulatory signature that is sufficient for de novo identification of HSN neuron functional enhancers. Among C. elegans neurons, the HSN transcriptome most closely resembles that of mouse serotonergic neurons. Mouse orthologs of the HSN TF collective also regulate serotonergic differentiation and can functionally substitute for their worm counterparts which suggests deep homology. Our results identify rules governing the regulatory landscape of a critically important neuronal type in two species separated by over 700 million years. © 2018, Lloret-Fernández et al.

  4. Stochasticity in the enterococcal sex pheromone response revealed by quantitative analysis of transcription in single cells.

    Science.gov (United States)

    Breuer, Rebecca J; Bandyopadhyay, Arpan; O'Brien, Sofie A; Barnes, Aaron M T; Hunter, Ryan C; Hu, Wei-Shou; Dunny, Gary M

    2017-07-01

    In Enterococcus faecalis, sex pheromone-mediated transfer of antibiotic resistance plasmids can occur under unfavorable conditions, for example, when inducing pheromone concentrations are low and inhibiting pheromone concentrations are high. To better understand this paradox, we adapted fluorescence in situ hybridization chain reaction (HCR) methodology for simultaneous quantification of multiple E. faecalis transcripts at the single cell level. We present direct evidence for variability in the minimum period, maximum response level, and duration of response of individual cells to a specific inducing condition. Tracking of induction patterns of single cells temporally using a fluorescent reporter supported HCR findings. It also revealed subpopulations of rapid responders, even under low inducing pheromone concentrations where the overall response of the entire population was slow. The strong, rapid induction of small numbers of cells in cultures exposed to low pheromone concentrations is in agreement with predictions of a stochastic model of the enterococcal pheromone response. The previously documented complex regulatory circuitry controlling the pheromone response likely contributes to stochastic variation in this system. In addition to increasing our basic understanding of the biology of a horizontal gene transfer system regulated by cell-cell signaling, demonstration of the stochastic nature of the pheromone response also impacts any future efforts to develop therapeutic agents targeting the system. Quantitative single cell analysis using HCR also has great potential to elucidate important bacterial regulatory mechanisms not previously amenable to study at the single cell level, and to accelerate the pace of functional genomic studies.

  5. Using network component analysis to dissect regulatory networks mediated by transcription factors in yeast.

    Directory of Open Access Journals (Sweden)

    Chun Ye

    2009-03-01

    Full Text Available Understanding the relationship between genetic variation and gene expression is a central question in genetics. With the availability of data from high-throughput technologies such as ChIP-Chip, expression, and genotyping arrays, we can begin to not only identify associations but to understand how genetic variations perturb the underlying transcription regulatory networks to induce differential gene expression. In this study, we describe a simple model of transcription regulation where the expression of a gene is completely characterized by two properties: the concentrations and promoter affinities of active transcription factors. We devise a method that extends Network Component Analysis (NCA to determine how genetic variations in the form of single nucleotide polymorphisms (SNPs perturb these two properties. Applying our method to a segregating population of Saccharomyces cerevisiae, we found statistically significant examples of trans-acting SNPs located in regulatory hotspots that perturb transcription factor concentrations and affinities for target promoters to cause global differential expression and cis-acting genetic variations that perturb the promoter affinities of transcription factors on a single gene to cause local differential expression. Although many genetic variations linked to gene expressions have been identified, it is not clear how they perturb the underlying regulatory networks that govern gene expression. Our work begins to fill this void by showing that many genetic variations affect the concentrations of active transcription factors in a cell and their affinities for target promoters. Understanding the effects of these perturbations can help us to paint a more complete picture of the complex landscape of transcription regulation. The software package implementing the algorithms discussed in this work is available as a MATLAB package upon request.

  6. Transcriptomic profiling-based mutant screen reveals three new transcription factors mediating menadione resistance in Neurospora crassa.

    Science.gov (United States)

    Zhu, Jufen; Yu, Xinxu; Xie, Baogui; Gu, Xiaokui; Zhang, Zhenying; Li, Shaojie

    2013-06-01

    To gain insight into the regulatory mechanisms of oxidative stress responses in filamentous fungi, the genome-wide transcriptional response of Neurospora crassa to menadione was analysed by digital gene expression (DGE) profiling, which identified 779 upregulated genes and 576 downregulated genes. Knockout mutants affecting 130 highly-upregulated genes were tested for menadione sensitivity, which revealed that loss of the transcription factor siderophore regulation (SRE) (a transcriptional repressor for siderophore biosynthesis), catatase-3, cytochrome c peroxidase or superoxide dismutase 1 copper chaperone causes hypersensitivity to menadione. Deletion of sre dramatically increased transcription of the siderophore biosynthesis gene ono and the siderophore iron transporter gene sit during menadione stress, suggesting that SRE is required for repression of iron uptake under oxidative stress conditions. Contrary to its phenotype, the sre deletion mutant showed higher transcriptional levels of genes encoding reactive oxygen species (ROS) scavengers than wild type during menadione stress, which implies that the mutant suffers a higher level of oxidative stress than wild type. Uncontrolled iron uptake in the sre mutant might exacerbate cellular oxidative stress. This is the first report of a negative regulator of iron assimilation participating in the fungal oxidative stress response. In addition to SRE, eight other transcription factor genes were also menadione-responsive but their single gene knockout mutants showed wild-type menadione sensitivity. Two of them, named as mit-2 (menadione induced transcription factor-2) and mit-4 (menadione induced transcription factor-4), were selected for double mutant analysis. The double mutant was hypersensitive to menadione. Similarly, the double mutation of mit-2 and sre also had additive effects on menadione sensitivity, suggesting multiple transcription factors mediate oxidative stress resistance in an additive manner

  7. Aggregation of topological motifs in the Escherichia coli transcriptional regulatory network

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    Barabási Albert-László

    2004-01-01

    Full Text Available Abstract Background Transcriptional regulation of cellular functions is carried out through a complex network of interactions among transcription factors and the promoter regions of genes and operons regulated by them.To better understand the system-level function of such networks simplification of their architecture was previously achieved by identifying the motifs present in the network, which are small, overrepresented, topologically distinct regulatory interaction patterns (subgraphs. However, the interaction of such motifs with each other, and their form of integration into the full network has not been previously examined. Results By studying the transcriptional regulatory network of the bacterium, Escherichia coli, we demonstrate that the two previously identified motif types in the network (i.e., feed-forward loops and bi-fan motifs do not exist in isolation, but rather aggregate into homologous motif clusters that largely overlap with known biological functions. Moreover, these clusters further coalesce into a supercluster, thus establishing distinct topological hierarchies that show global statistical properties similar to the whole network. Targeted removal of motif links disintegrates the network into small, isolated clusters, while random disruptions of equal number of links do not cause such an effect. Conclusion Individual motifs aggregate into homologous motif clusters and a supercluster forming the backbone of the E. coli transcriptional regulatory network and play a central role in defining its global topological organization.

  8. DMPD: Type I interferon [corrected] gene induction by the interferon regulatory factorfamily of transcription factors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 16979567 Type I interferon [corrected] gene induction by the interferon regulatory factorfamily...ng) (.svg) (.html) (.csml) Show Type I interferon [corrected] gene induction by the interferon regulatory factorfamily...orrected] gene induction by the interferon regulatory factorfamily of transcription factors. Authors Honda K

  9. Prediction of transcriptional regulatory sites in the complete genome sequence of Escherichia coli K-12.

    Science.gov (United States)

    Thieffry, D; Salgado, H; Huerta, A M; Collado-Vides, J

    1998-06-01

    As one of the best-characterized free-living organisms, Escherichia coli and its recently completed genomic sequence offer a special opportunity to exploit systematically the variety of regulatory data available in the literature in order to make a comprehensive set of regulatory predictions in the whole genome. The complete genome sequence of E.coli was analyzed for the binding of transcriptional regulators upstream of coding sequences. The biological information contained in RegulonDB (Huerta, A.M. et al., Nucleic Acids Res.,26,55-60, 1998) for 56 different transcriptional proteins was the support to implement a stringent strategy combining string search and weight matrices. We estimate that our search included representatives of 15-25% of the total number of regulatory binding proteins in E.coli. This search was performed on the set of 4288 putative regulatory regions, each 450 bp long. Within the regions with predicted sites, 89% are regulated by one protein and 81% involve only one site. These numbers are reasonably consistent with the distribution of experimental regulatory sites. Regulatory sites are found in 603 regions corresponding to 16% of operon regions and 10% of intra-operonic regions. Additional evidence gives stronger support to some of these predictions, including the position of the site, biological consistency with the function of the downstream gene, as well as genetic evidence for the regulatory interaction. The predictions described here were incorporated into the map presented in the paper describing the complete E.coli genome (Blattner,F.R. et al., Science, 277, 1453-1461, 1997). The complete set of predictions in GenBank format is available at the url: http://www. cifn.unam.mx/Computational_Biology/E.coli-predictions ecoli-reg@cifn.unam.mx, collado@cifn.unam.mx

  10. Bmp indicator mice reveal dynamic regulation of transcriptional response.

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    Anna L Javier

    Full Text Available Cellular responses to Bmp ligands are regulated at multiple levels, both extracellularly and intracellularly. Therefore, the presence of these growth factors is not an accurate indicator of Bmp signaling activity. While a common approach to detect Bmp signaling activity is to determine the presence of phosphorylated forms of Smad1, 5 and 8 by immunostaining, this approach is time consuming and not quantitative. In order to provide a simpler readout system to examine the presence of Bmp signaling in developing animals, we developed BRE-gal mouse embryonic stem cells and a transgenic mouse line that specifically respond to Bmp ligand stimulation. Our reporter identifies specific transcriptional responses that are mediated by Smad1 and Smad4 with the Schnurri transcription factor complex binding to a conserved Bmp-Responsive Element (BRE, originally identified among Drosophila, Xenopus and human Bmp targets. Our BRE-gal mES cells specifically respond to Bmp ligands at concentrations as low as 5 ng/ml; and BRE-gal reporter mice, derived from the BRE-gal mES cells, show dynamic activity in many cellular sites, including extraembryonic structures and mammary glands, thereby making this a useful scientific tool.

  11. A primer on thermodynamic-based models for deciphering transcriptional regulatory logic.

    Science.gov (United States)

    Dresch, Jacqueline M; Richards, Megan; Ay, Ahmet

    2013-09-01

    A rigorous analysis of transcriptional regulation at the DNA level is crucial to the understanding of many biological systems. Mathematical modeling has offered researchers a new approach to understanding this central process. In particular, thermodynamic-based modeling represents the most biophysically informed approach aimed at connecting DNA level regulatory sequences to the expression of specific genes. The goal of this review is to give biologists a thorough description of the steps involved in building, analyzing, and implementing a thermodynamic-based model of transcriptional regulation. The data requirements for this modeling approach are described, the derivation for a specific regulatory region is shown, and the challenges and future directions for the quantitative modeling of gene regulation are discussed. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Short-lived non-coding transcripts (SLiTs): Clues to regulatory long non-coding RNA.

    Science.gov (United States)

    Tani, Hidenori

    2017-03-22

    Whole transcriptome analyses have revealed a large number of novel long non-coding RNAs (lncRNAs). Although the importance of lncRNAs has been documented in previous reports, the biological and physiological functions of lncRNAs remain largely unknown. The role of lncRNAs seems an elusive problem. Here, I propose a clue to the identification of regulatory lncRNAs. The key point is RNA half-life. RNAs with a long half-life (t 1/2 > 4 h) contain a significant proportion of ncRNAs, as well as mRNAs involved in housekeeping functions, whereas RNAs with a short half-life (t 1/2 regulatory ncRNAs and regulatory mRNAs. This novel class of ncRNAs with a short half-life can be categorized as Short-Lived non-coding Transcripts (SLiTs). I consider that SLiTs are likely to be rich in functionally uncharacterized regulatory RNAs. This review describes recent progress in research into SLiTs.

  13. Nascent-Seq reveals novel features of mouse circadian transcriptional regulation

    Science.gov (United States)

    Menet, Jerome S; Rodriguez, Joseph; Abruzzi, Katharine C; Rosbash, Michael

    2012-01-01

    A substantial fraction of the metazoan transcriptome undergoes circadian oscillations in many cells and tissues. Based on the transcription feedback loops important for circadian timekeeping, it is commonly assumed that this mRNA cycling reflects widespread transcriptional regulation. To address this issue, we directly measured the circadian dynamics of mouse liver transcription using Nascent-Seq (genome-wide sequencing of nascent RNA). Although many genes are rhythmically transcribed, many rhythmic mRNAs manifest poor transcriptional rhythms, indicating a prominent contribution of post-transcriptional regulation to circadian mRNA expression. This analysis of rhythmic transcription also showed that the rhythmic DNA binding profile of the transcription factors CLOCK and BMAL1 does not determine the transcriptional phase of most target genes. This likely reflects gene-specific collaborations of CLK:BMAL1 with other transcription factors. These insights from Nascent-Seq indicate that it should have broad applicability to many other gene expression regulatory issues. DOI: http://dx.doi.org/10.7554/eLife.00011.001 PMID:23150795

  14. Inference of Transcription Regulatory Network in Low Phytic Acid Soybean Seeds

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    Neelam Redekar

    2017-11-01

    Full Text Available A dominant loss of function mutation in myo-inositol phosphate synthase (MIPS gene and recessive loss of function mutations in two multidrug resistant protein type-ABC transporter genes not only reduce the seed phytic acid levels in soybean, but also affect the pathways associated with seed development, ultimately resulting in low emergence. To understand the regulatory mechanisms and identify key genes that intervene in the seed development process in low phytic acid crops, we performed computational inference of gene regulatory networks in low and normal phytic acid soybeans using a time course transcriptomic data and multiple network inference algorithms. We identified a set of putative candidate transcription factors and their regulatory interactions with genes that have functions in myo-inositol biosynthesis, auxin-ABA signaling, and seed dormancy. We evaluated the performance of our unsupervised network inference method by comparing the predicted regulatory network with published regulatory interactions in Arabidopsis. Some contrasting regulatory interactions were observed in low phytic acid mutants compared to non-mutant lines. These findings provide important hypotheses on expression regulation of myo-inositol metabolism and phytohormone signaling in developing low phytic acid soybeans. The computational pipeline used for unsupervised network learning in this study is provided as open source software and is freely available at https://lilabatvt.github.io/LPANetwork/.

  15. Transcriptional Dysregulation of MYC Reveals Common Enhancer-Docking Mechanism.

    Science.gov (United States)

    Schuijers, Jurian; Manteiga, John Colonnese; Weintraub, Abraham Selby; Day, Daniel Sindt; Zamudio, Alicia Viridiana; Hnisz, Denes; Lee, Tong Ihn; Young, Richard Allen

    2018-04-10

    Transcriptional dysregulation of the MYC oncogene is among the most frequent events in aggressive tumor cells, and this is generally accomplished by acquisition of a super-enhancer somewhere within the 2.8 Mb TAD where MYC resides. We find that these diverse cancer-specific super-enhancers, differing in size and location, interact with the MYC gene through a common and conserved CTCF binding site located 2 kb upstream of the MYC promoter. Genetic perturbation of this enhancer-docking site in tumor cells reduces CTCF binding, super-enhancer interaction, MYC gene expression, and cell proliferation. CTCF binding is highly sensitive to DNA methylation, and this enhancer-docking site, which is hypomethylated in diverse cancers, can be inactivated through epigenetic editing with dCas9-DNMT. Similar enhancer-docking sites occur at other genes, including genes with prominent roles in multiple cancers, suggesting a mechanism by which tumor cell oncogenes can generally hijack enhancers. These results provide insights into mechanisms that allow a single target gene to be regulated by diverse enhancer elements in different cell types. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  16. Transcriptional Dysregulation of MYC Reveals Common Enhancer-Docking Mechanism

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    Jurian Schuijers

    2018-04-01

    Full Text Available Summary: Transcriptional dysregulation of the MYC oncogene is among the most frequent events in aggressive tumor cells, and this is generally accomplished by acquisition of a super-enhancer somewhere within the 2.8 Mb TAD where MYC resides. We find that these diverse cancer-specific super-enhancers, differing in size and location, interact with the MYC gene through a common and conserved CTCF binding site located 2 kb upstream of the MYC promoter. Genetic perturbation of this enhancer-docking site in tumor cells reduces CTCF binding, super-enhancer interaction, MYC gene expression, and cell proliferation. CTCF binding is highly sensitive to DNA methylation, and this enhancer-docking site, which is hypomethylated in diverse cancers, can be inactivated through epigenetic editing with dCas9-DNMT. Similar enhancer-docking sites occur at other genes, including genes with prominent roles in multiple cancers, suggesting a mechanism by which tumor cell oncogenes can generally hijack enhancers. These results provide insights into mechanisms that allow a single target gene to be regulated by diverse enhancer elements in different cell types. : Schuijers et al. show that a conserved CTCF site at the promoter of the MYC oncogene plays an important role in enhancer-promoter looping with tumor-specific super-enhancers. Perturbation of this site provides a potential therapeutic vulnerability. Keywords: gene regulation, super-enhancers, chromosome structure, enhancer docking

  17. Comparative analysis of regulatory elements between Escherichia coli and Klebsiella pneumoniae by genome-wide transcription start site profiling.

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    Donghyuk Kim

    Full Text Available Genome-wide transcription start site (TSS profiles of the enterobacteria Escherichia coli and Klebsiella pneumoniae were experimentally determined through modified 5' RACE followed by deep sequencing of intact primary mRNA. This identified 3,746 and 3,143 TSSs for E. coli and K. pneumoniae, respectively. Experimentally determined TSSs were then used to define promoter regions and 5' UTRs upstream of coding genes. Comparative analysis of these regulatory elements revealed the use of multiple TSSs, identical sequence motifs of promoter and Shine-Dalgarno sequence, reflecting conserved gene expression apparatuses between the two species. In both species, over 70% of primary transcripts were expressed from operons having orthologous genes during exponential growth. However, expressed orthologous genes in E. coli and K. pneumoniae showed a strikingly different organization of upstream regulatory regions with only 20% identical promoters with TSSs in both species. Over 40% of promoters had TSSs identified in only one species, despite conserved promoter sequences existing in the other species. 662 conserved promoters having TSSs in both species resulted in the same number of comparable 5' UTR pairs, and that regulatory element was found to be the most variant region in sequence among promoter, 5' UTR, and ORF. In K. pneumoniae, 48 sRNAs were predicted and 36 of them were expressed during exponential growth. Among them, 34 orthologous sRNAs between two species were analyzed in depth, and the analysis showed that many sRNAs of K. pneumoniae, including pleiotropic sRNAs such as rprA, arcZ, and sgrS, may work in the same way as in E. coli. These results reveal a new dimension of comparative genomics such that a comparison of two genomes needs to be comprehensive over all levels of genome organization.

  18. Spatially resolved metabolic analysis reveals a central role for transcriptional control in carbon allocation to wood.

    Science.gov (United States)

    Roach, Melissa; Arrivault, Stéphanie; Mahboubi, Amir; Krohn, Nicole; Sulpice, Ronan; Stitt, Mark; Niittylä, Totte

    2017-06-15

    The contribution of transcriptional and post-transcriptional regulation to modifying carbon allocation to developing wood of trees is not well defined. To clarify the role of transcriptional regulation, the enzyme activity patterns of eight central primary metabolism enzymes across phloem, cambium, and developing wood of aspen (Populus tremula L.) were compared with transcript levels obtained by RNA sequencing of sequential stem sections from the same trees. Enzymes were selected on the basis of their importance in sugar metabolism and in linking primary metabolism to lignin biosynthesis. Existing enzyme assays were adapted to allow measurements from ~1 mm3 sections of dissected stem tissue. These experiments provided high spatial resolution of enzyme activity changes across different stages of wood development, and identified the gene transcripts probably responsible for these changes. In most cases, there was a clear positive relationship between transcripts and enzyme activity. During secondary cell wall formation, the increases in transcript levels and enzyme activities also matched with increased levels of glucose, fructose, hexose phosphates, and UDP-glucose, emphasizing an important role for transcriptional regulation in carbon allocation to developing aspen wood. These observations corroborate the efforts to increase carbon allocation to wood by engineering gene regulatory networks. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  19. The transcriptional regulatory network of Corynebacterium jeikeium K411 and its interaction with metabolic routes contributing to human body odor formation.

    Science.gov (United States)

    Barzantny, Helena; Schröder, Jasmin; Strotmeier, Jasmin; Fredrich, Eugenie; Brune, Iris; Tauch, Andreas

    2012-06-15

    Lipophilic corynebacteria are involved in the generation of volatile odorous products in the process of human body odor formation by degrading skin lipids and specific odor precursors. Therefore, these bacteria represent appropriate model systems for the cosmetic industry to examine axillary malodor formation on the molecular level. To understand the transcriptional control of metabolic pathways involved in this process, the transcriptional regulatory network of the lipophilic axilla isolate Corynebacterium jeikeium K411 was reconstructed from the complete genome sequence. This bioinformatic approach detected a gene-regulatory repertoire of 83 candidate proteins, including 56 DNA-binding transcriptional regulators, nine two-component systems, nine sigma factors, and nine regulators with diverse physiological functions. Furthermore, a cross-genome comparison among selected corynebacterial species of the taxonomic cluster 3 revealed a common gene-regulatory repertoire of 44 transcriptional regulators, including the MarR-like regulator Jk0257, which is exclusively encoded in the genomes of this taxonomical subline. The current network reconstruction comprises 48 transcriptional regulators and 674 gene-regulatory interactions that were assigned to five interconnected functional modules. Most genes involved in lipid degradation are under the combined control of the global cAMP-sensing transcriptional regulator GlxR and the LuxR-family regulator RamA, probably reflecting the essential role of lipid degradation in C. jeikeium. This study provides the first genome-scale in silico analysis of the transcriptional regulation of metabolism in a lipophilic bacterium involved in the formation of human body odor. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. Dynamic zebrafish interactome reveals transcriptional mechanisms of dioxin toxicity.

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    Andrey Alexeyenko

    2010-05-01

    Full Text Available In order to generate hypotheses regarding the mechanisms by which 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin causes toxicity, we analyzed global gene expression changes in developing zebrafish embryos exposed to this potent toxicant in the context of a dynamic gene network. For this purpose, we also computationally inferred a zebrafish (Danio rerio interactome based on orthologs and interaction data from other eukaryotes.Using novel computational tools to analyze this interactome, we distinguished between dioxin-dependent and dioxin-independent interactions between proteins, and tracked the temporal propagation of dioxin-dependent transcriptional changes from a few genes that were altered initially, to large groups of biologically coherent genes at later times. The most notable processes altered at later developmental stages were calcium and iron metabolism, embryonic morphogenesis including neuronal and retinal development, a variety of mitochondria-related functions, and generalized stress response (not including induction of antioxidant genes. Within the interactome, many of these responses were connected to cytochrome P4501A (cyp1a as well as other genes that were dioxin-regulated one day after exposure. This suggests that cyp1a may play a key role initiating the toxic dysregulation of those processes, rather than serving simply as a passive marker of dioxin exposure, as suggested by earlier research.Thus, a powerful microarray experiment coupled with a flexible interactome and multi-pronged interactome tools (which are now made publicly available for microarray analysis and related work suggest the hypothesis that dioxin, best known in fish as a potent cardioteratogen, has many other targets. Many of these types of toxicity have been observed in mammalian species and are potentially caused by alterations to cyp1a.

  1. High-resolution structure of TBP with TAF1 reveals anchoring patterns in transcriptional regulation.

    Science.gov (United States)

    Anandapadamanaban, Madhanagopal; Andresen, Cecilia; Helander, Sara; Ohyama, Yoshifumi; Siponen, Marina I; Lundström, Patrik; Kokubo, Tetsuro; Ikura, Mitsuhiko; Moche, Martin; Sunnerhagen, Maria

    2013-08-01

    The general transcription factor TFIID provides a regulatory platform for transcription initiation. Here we present the crystal structure (1.97 Å) and NMR analysis of yeast TAF1 N-terminal domains TAND1 and TAND2 bound to yeast TBP, together with mutational data. We find that yeast TAF1-TAND1, which in itself acts as a transcriptional activator, binds TBP's concave DNA-binding surface by presenting similar anchor residues to TBP as does Mot1 but from a distinct structural scaffold. Furthermore, we show how TAF1-TAND2 uses an aromatic and acidic anchoring pattern to bind a conserved TBP surface groove traversing the basic helix region, and we find highly similar TBP-binding motifs also presented by the structurally distinct TFIIA, Mot1 and Brf1 proteins. Our identification of these anchoring patterns, which can be easily disrupted or enhanced, provides insight into the competitive multiprotein TBP interplay critical to transcriptional regulation.

  2. Localizing potentially active post-transcriptional regulations in the Ewing's sarcoma gene regulatory network

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    Delyon Bernard

    2010-11-01

    Full Text Available Abstract Background A wide range of techniques is now available for analyzing regulatory networks. Nonetheless, most of these techniques fail to interpret large-scale transcriptional data at the post-translational level. Results We address the question of using large-scale transcriptomic observation of a system perturbation to analyze a regulatory network which contained several types of interactions - transcriptional and post-translational. Our method consisted of post-processing the outputs of an open-source tool named BioQuali - an automatic constraint-based analysis mimicking biologist's local reasoning on a large scale. The post-processing relied on differences in the behavior of the transcriptional and post-translational levels in the network. As a case study, we analyzed a network representation of the genes and proteins controlled by an oncogene in the context of Ewing's sarcoma. The analysis allowed us to pinpoint active interactions specific to this cancer. We also identified the parts of the network which were incomplete and should be submitted for further investigation. Conclusions The proposed approach is effective for the qualitative analysis of cancer networks. It allows the integrative use of experimental data of various types in order to identify the specific information that should be considered a priority in the initial - and possibly very large - experimental dataset. Iteratively, new dataset can be introduced into the analysis to improve the network representation and make it more specific.

  3. Exploring the bZIP transcription factor regulatory network in Neurospora crassa.

    Science.gov (United States)

    Tian, Chaoguang; Li, Jingyi; Glass, N Louise

    2011-03-01

    Transcription factors (TFs) are key nodes of regulatory networks in eukaryotic organisms, including filamentous fungi such as Neurospora crassa. The 178 predicted DNA-binding TFs in N. crassa are distributed primarily among six gene families, which represent an ancient expansion in filamentous ascomycete genomes; 98 TF genes show detectable expression levels during vegetative growth of N. crassa, including 35 that show a significant difference in expression level between hyphae at the periphery versus hyphae in the interior of a colony. Regulatory networks within a species genome include paralogous TFs and their respective target genes (TF regulon). To investigate TF network evolution in N. crassa, we focused on the basic leucine zipper (bZIP) TF family, which contains nine members. We performed baseline transcriptional profiling during vegetative growth of the wild-type and seven isogenic, viable bZIP deletion mutants. We further characterized the regulatory network of one member of the bZIP family, NCU03905. NCU03905 encodes an Ap1-like protein (NcAp-1), which is involved in resistance to multiple stress responses, including oxidative and heavy metal stress. Relocalization of NcAp-1 from the cytoplasm to the nucleus was associated with exposure to stress. A comparison of the NcAp-1 regulon with Ap1-like regulons in Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans and Aspergillus fumigatus showed both conservation and divergence. These data indicate how N. crassa responds to stress and provide information on pathway evolution.

  4. RNA-FISH to Study Regulatory RNA at the Site of Transcription.

    Science.gov (United States)

    Soler, Marta; Boque-Sastre, Raquel; Guil, Sonia

    2017-01-01

    The increasing role of all types of regulatory RNAs in the orchestration of cellular programs has enhanced the development of a variety of techniques that allow its precise detection, quantification, and functional scrutiny. Recent advances in imaging and fluoresecent in situ hybridization (FISH) methods have enabled the utilization of user-friendly protocols that provide highly sensitive and accurate detection of ribonucleic acid molecules at both the single cell and subcellular levels. We herein describe the approach originally developed by Stellaris ® , in which the target RNA molecule is fluoresecently labeled with multiple tiled complementary probes each carrying a fluorophore, thus improving sensitivity and reducing the chance of false positives. We have applied this method to the detection of nascent RNAs that partake of special regulatory structures called R loops. Their growing role in active gene expression regulation (Aguilera and Garcia-Muse, Mol Cell 46:115-124, 2012; Ginno et al., Mol Cell 45:814-825, 2012; Sun et al., Science 340:619-621, 2013; Bhatia et al., Nature 511:362-365, 2014) imposes the use of a combination of in vivo and in vitro techniques for the detailed analysis of the transcripts involved. Therefore, their study is a good example to illustrate how RNA FISH, combined with transcriptional arrest and/or cell synchronization, permits localization and temporal characterization of potentially regulatory RNA sequences.

  5. Transcriptional and Translational Regulatory Responses to Iron Limitation in the Globally Distributed Marine Bacterium Candidatus Pelagibacter ubique

    Science.gov (United States)

    Smith, Daniel P.; Kitner, Joshua B.; Norbeck, Angela D.; Clauss, Therese R.; Lipton, Mary S.; Schwalbach, Michael S.; Steindler, Laura; Nicora, Carrie D.; Smith, Richard D.; Giovannoni, Stephen J.

    2010-01-01

    Iron is recognized as an important micronutrient that limits microbial plankton productivity over vast regions of the oceans. We investigated the gene expression responses of Candidatus Pelagibacter ubique cultures to iron limitation in natural seawater media supplemented with a siderophore to chelate iron. Microarray data indicated transcription of the periplasmic iron binding protein sfuC increased by 16-fold, and iron transporter subunits, iron-sulfur center assembly genes, and the putative ferroxidase rubrerythrin transcripts increased to a lesser extent. Quantitative peptide mass spectrometry revealed that sfuC protein abundance increased 27-fold, despite an average decrease of 59% across the global proteome. Thus, we propose sfuC as a marker gene for indicating iron limitation in marine metatranscriptomic and metaproteomic ecological surveys. The marked proteome reduction was not directly correlated to changes in the transcriptome, implicating post-transcriptional regulatory mechanisms as modulators of protein expression. Two RNA-binding proteins, CspE and CspL, correlated well with iron availability, suggesting that they may contribute to the observed differences between the transcriptome and proteome. We propose a model in which the RNA-binding activity of CspE and CspL selectively enables protein synthesis of the iron acquisition protein SfuC during transient growth-limiting episodes of iron scarcity. PMID:20463970

  6. The post-transcriptional regulatory system CSR controls the balance of metabolic pools in upper glycolysis of Escherichia coli.

    Science.gov (United States)

    Morin, Manon; Ropers, Delphine; Letisse, Fabien; Laguerre, Sandrine; Portais, Jean-Charles; Cocaign-Bousquet, Muriel; Enjalbert, Brice

    2016-05-01

    Metabolic control in Escherichia coli is a complex process involving multilevel regulatory systems but the involvement of post-transcriptional regulation is uncertain. The post-transcriptional factor CsrA is stated as being the only regulator essential for the use of glycolytic substrates. A dozen enzymes in the central carbon metabolism (CCM) have been reported as potentially controlled by CsrA, but its impact on the CCM functioning has not been demonstrated. Here, a multiscale analysis was performed in a wild-type strain and its isogenic mutant attenuated for CsrA (including growth parameters, gene expression levels, metabolite pools, abundance of enzymes and fluxes). Data integration and regulation analysis showed a coordinated control of the expression of glycolytic enzymes. This also revealed the imbalance of metabolite pools in the csrA mutant upper glycolysis, before the phosphofructokinase PfkA step. This imbalance is associated with a glucose-phosphate stress. Restoring PfkA activity in the csrA mutant strain suppressed this stress and increased the mutant growth rate on glucose. Thus, the carbon storage regulator system is essential for the effective functioning of the upper glycolysis mainly through its control of PfkA. This work demonstrates the pivotal role of post-transcriptional regulation to shape the carbon metabolism. © 2016 John Wiley & Sons Ltd.

  7. Fanconi anemia core complex gene promoters harbor conserved transcription regulatory elements.

    Science.gov (United States)

    Meier, Daniel; Schindler, Detlev

    2011-01-01

    The Fanconi anemia (FA) gene family is a recent addition to the complex network of proteins that respond to and repair certain types of DNA damage in the human genome. Since little is known about the regulation of this novel group of genes at the DNA level, we characterized the promoters of the eight genes (FANCA, B, C, E, F, G, L and M) that compose the FA core complex. The promoters of these genes show the characteristic attributes of housekeeping genes, such as a high GC content and CpG islands, a lack of TATA boxes and a low conservation. The promoters functioned in a monodirectional way and were, in their most active regions, comparable in strength to the SV40 promoter in our reporter plasmids. They were also marked by a distinctive transcriptional start site (TSS). In the 5' region of each promoter, we identified a region that was able to negatively regulate the promoter activity in HeLa and HEK 293 cells in isolation. The central and 3' regions of the promoter sequences harbor binding sites for several common and rare transcription factors, including STAT, SMAD, E2F, AP1 and YY1, which indicates that there may be cross-connections to several established regulatory pathways. Electrophoretic mobility shift assays and siRNA experiments confirmed the shared regulatory responses between the prominent members of the TGF-β and JAK/STAT pathways and members of the FA core complex. Although the promoters are not well conserved, they share region and sequence specific regulatory motifs and transcription factor binding sites (TBFs), and we identified a bi-partite nature to these promoters. These results support a hypothesis based on the co-evolution of the FA core complex genes that was expanded to include their promoters.

  8. Fanconi anemia core complex gene promoters harbor conserved transcription regulatory elements.

    Directory of Open Access Journals (Sweden)

    Daniel Meier

    Full Text Available The Fanconi anemia (FA gene family is a recent addition to the complex network of proteins that respond to and repair certain types of DNA damage in the human genome. Since little is known about the regulation of this novel group of genes at the DNA level, we characterized the promoters of the eight genes (FANCA, B, C, E, F, G, L and M that compose the FA core complex. The promoters of these genes show the characteristic attributes of housekeeping genes, such as a high GC content and CpG islands, a lack of TATA boxes and a low conservation. The promoters functioned in a monodirectional way and were, in their most active regions, comparable in strength to the SV40 promoter in our reporter plasmids. They were also marked by a distinctive transcriptional start site (TSS. In the 5' region of each promoter, we identified a region that was able to negatively regulate the promoter activity in HeLa and HEK 293 cells in isolation. The central and 3' regions of the promoter sequences harbor binding sites for several common and rare transcription factors, including STAT, SMAD, E2F, AP1 and YY1, which indicates that there may be cross-connections to several established regulatory pathways. Electrophoretic mobility shift assays and siRNA experiments confirmed the shared regulatory responses between the prominent members of the TGF-β and JAK/STAT pathways and members of the FA core complex. Although the promoters are not well conserved, they share region and sequence specific regulatory motifs and transcription factor binding sites (TBFs, and we identified a bi-partite nature to these promoters. These results support a hypothesis based on the co-evolution of the FA core complex genes that was expanded to include their promoters.

  9. Transcriptome profiling reveals regulatory mechanisms underlying Corolla Senescence in Petunia

    Science.gov (United States)

    Genetic regulatory mechanisms that govern petal natural senescence in petunia is complicated and unclear. To identify key genes and pathways that regulate the process, we initiated a transcriptome analysis in petunia petals at four developmental time points, including petal opening without anthesis ...

  10. A deeper look into transcription regulatory code by preferred pair distance templates for transcription factor binding sites

    KAUST Repository

    Kulakovskiy, Ivan V.; Belostotsky, A. A.; Kasianov, Artem S.; Esipova, Natalia G.; Medvedeva, Yulia; Eliseeva, Irina A.; Makeev, Vsevolod J.

    2011-01-01

    Motivation: Modern experimental methods provide substantial information on protein-DNA recognition. Studying arrangements of transcription factor binding sites (TFBSs) of interacting transcription factors (TFs) advances understanding

  11. Nsite, NsiteH and NsiteM Computer Tools for Studying Tran-scription Regulatory Elements

    KAUST Repository

    Shahmuradov, Ilham

    2015-07-02

    Summary: Gene transcription is mostly conducted through interactions of various transcription factors and their binding sites on DNA (regulatory elements, REs). Today, we are still far from understanding the real regulatory content of promoter regions. Computer methods for identification of REs remain a widely used tool for studying and understanding transcriptional regulation mechanisms. The Nsite, NsiteH and NsiteM programs perform searches for statistically significant (non-random) motifs of known human, animal and plant one-box and composite REs in a single genomic sequence, in a pair of aligned homologous sequences and in a set of functionally related sequences, respectively.

  12. Increments and duplication events of enzymes and transcription factors influence metabolic and regulatory diversity in prokaryotes.

    Directory of Open Access Journals (Sweden)

    Mario Alberto Martínez-Núñez

    Full Text Available In this work, the content of enzymes and DNA-binding transcription factors (TFs in 794 non-redundant prokaryotic genomes was evaluated. The identification of enzymes was based on annotations deposited in the KEGG database as well as in databases of functional domains (COG and PFAM and structural domains (Superfamily. For identifications of the TFs, hidden Markov profiles were constructed based on well-known transcriptional regulatory families. From these analyses, we obtained diverse and interesting results, such as the negative rate of incremental changes in the number of detected enzymes with respect to the genome size. On the contrary, for TFs the rate incremented as the complexity of genome increased. This inverse related performance shapes the diversity of metabolic and regulatory networks and impacts the availability of enzymes and TFs. Furthermore, the intersection of the derivatives between enzymes and TFs was identified at 9,659 genes, after this point, the regulatory complexity grows faster than metabolic complexity. In addition, TFs have a low number of duplications, in contrast to the apparent high number of duplications associated with enzymes. Despite the greater number of duplicated enzymes versus TFs, the increment by which duplicates appear is higher in TFs. A lower proportion of enzymes among archaeal genomes (22% than in the bacterial ones (27% was also found. This low proportion might be compensated by the interconnection between the metabolic pathways in Archaea. A similar proportion was also found for the archaeal TFs, for which the formation of regulatory complexes has been proposed. Finally, an enrichment of multifunctional enzymes in Bacteria, as a mechanism of ecological adaptation, was detected.

  13. Core Transcriptional Regulatory Circuit Controlled by the TAL1 Complex in Human T Cell Acute Lymphoblastic Leukemia

    OpenAIRE

    Sanda, Takaomi; Lawton, Lee N.; Barrasa, M. Inmaculada; Fan, Zi Peng; Kohlhammer, Holger; Gutierrez, Alejandro; Ma, Wenxue; Tatarek, Jessica; Ahn, Yebin; Kelliher, Michelle A.; Jamieson, Catriona H.M.; Staudt, Louis M.; Young, Richard A.; Look, A. Thomas

    2012-01-01

    The oncogenic transcription factor TAL1/SCL is aberrantly expressed in over 40% of cases of human T-cell acute lymphoblastic leukemia (T-ALL), emphasizing its importance in the molecular pathogenesis of T-ALL. Here we identify the core transcriptional regulatory circuit controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3 and RUNX1. We show that TAL1 forms a positive interconnected auto-regulatory loop with GATA3 and RUNX1, and that the TAL1 complex directly activates the MY...

  14. Comparison of stationary and oscillatory dynamics described by differential equations and Boolean maps in transcriptional regulatory circuits

    Science.gov (United States)

    Ye, Weiming; Li, Pengfei; Huang, Xuhui; Xia, Qinzhi; Mi, Yuanyuan; Chen, Runsheng; Hu, Gang

    2010-10-01

    Exploring the principle and relationship of gene transcriptional regulations (TR) has been becoming a generally researched issue. So far, two major mathematical methods, ordinary differential equation (ODE) method and Boolean map (BM) method have been widely used for these purposes. It is commonly believed that simplified BMs are reasonable approximations of more realistic ODEs, and both methods may reveal qualitatively the same essential features though the dynamical details of both systems may show some differences. In this Letter we exhaustively enumerated all the 3-gene networks and many autonomous randomly constructed TR networks with more genes by using both the ODE and BM methods. In comparison we found that both methods provide practically identical results in most of cases of steady solutions. However, to our great surprise, most of network structures showing periodic cycles with the BM method possess only stationary states in ODE descriptions. These observations strongly suggest that many periodic oscillations and other complicated oscillatory states revealed by the BM rule may be related to the computational errors of variable and time discretizations and rarely have correspondence in realistic biology transcriptional regulatory circuits.

  15. Bovine proteins containing poly-glutamine repeats are often polymorphic and enriched for components of transcriptional regulatory complexes

    LENUS (Irish Health Repository)

    Whan, Vicki

    2010-11-23

    Abstract Background About forty human diseases are caused by repeat instability mutations. A distinct subset of these diseases is the result of extreme expansions of polymorphic trinucleotide repeats; typically CAG repeats encoding poly-glutamine (poly-Q) tracts in proteins. Polymorphic repeat length variation is also apparent in human poly-Q encoding genes from normal individuals. As these coding sequence repeats are subject to selection in mammals, it has been suggested that normal variations in some of these typically highly conserved genes are implicated in morphological differences between species and phenotypic variations within species. At present, poly-Q encoding genes in non-human mammalian species are poorly documented, as are their functions and propensities for polymorphic variation. Results The current investigation identified 178 bovine poly-Q encoding genes (Q ≥ 5) and within this group, 26 genes with orthologs in both human and mouse that did not contain poly-Q repeats. The bovine poly-Q encoding genes typically had ubiquitous expression patterns although there was bias towards expression in epithelia, brain and testes. They were also characterised by unusually large sizes. Analysis of gene ontology terms revealed that the encoded proteins were strongly enriched for functions associated with transcriptional regulation and many contributed to physical interaction networks in the nucleus where they presumably act cooperatively in transcriptional regulatory complexes. In addition, the coding sequence CAG repeats in some bovine genes impacted mRNA splicing thereby generating unusual transcriptional diversity, which in at least one instance was tissue-specific. The poly-Q encoding genes were prioritised using multiple criteria for their likelihood of being polymorphic and then the highest ranking group was experimentally tested for polymorphic variation within a cattle diversity panel. Extensive and meiotically stable variation was identified

  16. The 3' untranslated region of human Cyclin-Dependent Kinase 5 Regulatory subunit 1 contains regulatory elements affecting transcript stability

    Directory of Open Access Journals (Sweden)

    Ratti Antonia

    2007-12-01

    Full Text Available Abstract Background CDK5R1 plays a central role in neuronal migration and differentiation during central nervous system development. CDK5R1 has been implicated in neurodegenerative disorders and proposed as a candidate gene for mental retardation. The remarkable size of CDK5R1 3'-untranslated region (3'-UTR suggests a role in post-transcriptional regulation of CDK5R1 expression. Results The bioinformatic study shows a high conservation degree in mammals and predicts several AU-Rich Elements (AREs. The insertion of CDK5R1 3'-UTR into luciferase 3'-UTR causes a decreased luciferase activity in four transfected cell lines. We identified 3'-UTR subregions which tend to reduce the reporter gene expression, sometimes in a cell line-dependent manner. In most cases the quantitative analysis of luciferase mRNA suggests that CDK5R1 3'-UTR affects mRNA stability. A region, leading to a very strong mRNA destabilization, showed a significantly low half-life, indicating an accelerated mRNA degradation. The 3' end of the transcript, containing a class I ARE, specifically displays a stabilizing effect in neuroblastoma cell lines. We also observed the interaction of the stabilizing neuronal RNA-binding proteins ELAV with the CDK5R1 transcript in SH-SY5Y cells and identified three 3'-UTR sub-regions showing affinity for ELAV proteins. Conclusion Our findings evince the presence of both destabilizing and stabilizing regulatory elements in CDK5R1 3'-UTR and support the hypothesis that CDK5R1 gene expression is post-transcriptionally controlled in neurons by ELAV-mediated mechanisms. This is the first evidence of the involvement of 3'-UTR in the modulation of CDK5R1 expression. The fine tuning of CDK5R1 expression by 3'-UTR may have a role in central nervous system development and functioning, with potential implications in neurodegenerative and cognitive disorders.

  17. Transcriptome sequencing from diverse human populations reveals differentiated regulatory architecture.

    Directory of Open Access Journals (Sweden)

    Alicia R Martin

    2014-08-01

    Full Text Available Large-scale sequencing efforts have documented extensive genetic variation within the human genome. However, our understanding of the origins, global distribution, and functional consequences of this variation is far from complete. While regulatory variation influencing gene expression has been studied within a handful of populations, the breadth of transcriptome differences across diverse human populations has not been systematically analyzed. To better understand the spectrum of gene expression variation, alternative splicing, and the population genetics of regulatory variation in humans, we have sequenced the genomes, exomes, and transcriptomes of EBV transformed lymphoblastoid cell lines derived from 45 individuals in the Human Genome Diversity Panel (HGDP. The populations sampled span the geographic breadth of human migration history and include Namibian San, Mbuti Pygmies of the Democratic Republic of Congo, Algerian Mozabites, Pathan of Pakistan, Cambodians of East Asia, Yakut of Siberia, and Mayans of Mexico. We discover that approximately 25.0% of the variation in gene expression found amongst individuals can be attributed to population differences. However, we find few genes that are systematically differentially expressed among populations. Of this population-specific variation, 75.5% is due to expression rather than splicing variability, and we find few genes with strong evidence for differential splicing across populations. Allelic expression analyses indicate that previously mapped common regulatory variants identified in eight populations from the International Haplotype Map Phase 3 project have similar effects in our seven sampled HGDP populations, suggesting that the cellular effects of common variants are shared across diverse populations. Together, these results provide a resource for studies analyzing functional differences across populations by estimating the degree of shared gene expression, alternative splicing, and

  18. Chromosomal clustering of a human transcriptome reveals regulatory background

    Directory of Open Access Journals (Sweden)

    Purmann Antje

    2005-09-01

    Full Text Available Abstract Background There has been much evidence recently for a link between transcriptional regulation and chromosomal gene order, but the relationship between genomic organization, regulation and gene function in higher eukaryotes remains to be precisely defined. Results Here, we present evidence for organization of a large proportion of a human transcriptome into gene clusters throughout the genome, which are partly regulated by the same transcription factors, share biological functions and are characterized by non-housekeeping genes. This analysis was based on the cardiac transcriptome identified by our genome-wide array analysis of 55 human heart samples. We found 37% of these genes to be arranged mainly in adjacent pairs or triplets. A significant number of pairs of adjacent genes are putatively regulated by common transcription factors (p = 0.02. Furthermore, these gene pairs share a significant number of GO functional classification terms. We show that the human cardiac transcriptome is organized into many small clusters across the whole genome, rather than being concentrated in a few larger clusters. Conclusion Our findings suggest that genes expressed in concert are organized in a linear arrangement for coordinated regulation. Determining the relationship between gene arrangement, regulation and nuclear organization as well as gene function will have broad biological implications.

  19. CoryneRegNet: an ontology-based data warehouse of corynebacterial transcription factors and regulatory networks.

    Science.gov (United States)

    Baumbach, Jan; Brinkrolf, Karina; Czaja, Lisa F; Rahmann, Sven; Tauch, Andreas

    2006-02-14

    The application of DNA microarray technology in post-genomic analysis of bacterial genome sequences has allowed the generation of huge amounts of data related to regulatory networks. This data along with literature-derived knowledge on regulation of gene expression has opened the way for genome-wide reconstruction of transcriptional regulatory networks. These large-scale reconstructions can be converted into in silico models of bacterial cells that allow a systematic analysis of network behavior in response to changing environmental conditions. CoryneRegNet was designed to facilitate the genome-wide reconstruction of transcriptional regulatory networks of corynebacteria relevant in biotechnology and human medicine. During the import and integration process of data derived from experimental studies or literature knowledge CoryneRegNet generates links to genome annotations, to identified transcription factors and to the corresponding cis-regulatory elements. CoryneRegNet is based on a multi-layered, hierarchical and modular concept of transcriptional regulation and was implemented by using the relational database management system MySQL and an ontology-based data structure. Reconstructed regulatory networks can be visualized by using the yFiles JAVA graph library. As an application example of CoryneRegNet, we have reconstructed the global transcriptional regulation of a cellular module involved in SOS and stress response of corynebacteria. CoryneRegNet is an ontology-based data warehouse that allows a pertinent data management of regulatory interactions along with the genome-scale reconstruction of transcriptional regulatory networks. These models can further be combined with metabolic networks to build integrated models of cellular function including both metabolism and its transcriptional regulation.

  20. CoryneRegNet: An ontology-based data warehouse of corynebacterial transcription factors and regulatory networks

    Directory of Open Access Journals (Sweden)

    Czaja Lisa F

    2006-02-01

    Full Text Available Abstract Background The application of DNA microarray technology in post-genomic analysis of bacterial genome sequences has allowed the generation of huge amounts of data related to regulatory networks. This data along with literature-derived knowledge on regulation of gene expression has opened the way for genome-wide reconstruction of transcriptional regulatory networks. These large-scale reconstructions can be converted into in silico models of bacterial cells that allow a systematic analysis of network behavior in response to changing environmental conditions. Description CoryneRegNet was designed to facilitate the genome-wide reconstruction of transcriptional regulatory networks of corynebacteria relevant in biotechnology and human medicine. During the import and integration process of data derived from experimental studies or literature knowledge CoryneRegNet generates links to genome annotations, to identified transcription factors and to the corresponding cis-regulatory elements. CoryneRegNet is based on a multi-layered, hierarchical and modular concept of transcriptional regulation and was implemented by using the relational database management system MySQL and an ontology-based data structure. Reconstructed regulatory networks can be visualized by using the yFiles JAVA graph library. As an application example of CoryneRegNet, we have reconstructed the global transcriptional regulation of a cellular module involved in SOS and stress response of corynebacteria. Conclusion CoryneRegNet is an ontology-based data warehouse that allows a pertinent data management of regulatory interactions along with the genome-scale reconstruction of transcriptional regulatory networks. These models can further be combined with metabolic networks to build integrated models of cellular function including both metabolism and its transcriptional regulation.

  1. Transcriptional regulatory elements in the noncoding region of human papillomavirus type 6

    International Nuclear Information System (INIS)

    Wu, Tzyy-Choou.

    1989-01-01

    The structure and function of the transcriptional regulatory region of human papillomavirus type 6 (HPV-6) has been investigated. To investigate tissue specific gene expression, a sensitive method to detect and localize HPV-6 viral DNA, mRNA and protein in plastic-embedded tissue sections of genital and respiratory tract papillomata by using in situ hybridization and immunoperoxidase assays has been developed. This method, using ultrathin sections and strand-specific 3 H labeled riboprobes, offers the advantages of superior morphological preservation and detection of viral genomes at low copy number with good resolution, and the modified immunocytochemistry provides better sensitivity. The results suggest that genital tract epithelium is more permissive for HPV-6 replication than respiratory tract epithelium. To study the tissue tropism of HPV-6 at the level of regulation of viral gene expression, the polymerase chain reaction was used to isolate the noncoding region (NCR) of HPV-6 in independent isolates. Nucleotide sequence analysis of molecularly cloned DNA identified base substitutions, deletions/insertions and tandem duplications. Transcriptional regulatory elements in the NCR were assayed in recombinant plasmids containing the bacterial gene for chloramphenicol acetyl transferase

  2. Genetic networks of liver metabolism revealed by integration of metabolic and transcriptional profiling.

    Directory of Open Access Journals (Sweden)

    Christine T Ferrara

    2008-03-01

    Full Text Available Although numerous quantitative trait loci (QTL influencing disease-related phenotypes have been detected through gene mapping and positional cloning, identification of the individual gene(s and molecular pathways leading to those phenotypes is often elusive. One way to improve understanding of genetic architecture is to classify phenotypes in greater depth by including transcriptional and metabolic profiling. In the current study, we have generated and analyzed mRNA expression and metabolic profiles in liver samples obtained in an F2 intercross between the diabetes-resistant C57BL/6 leptin(ob/ob and the diabetes-susceptible BTBR leptin(ob/ob mouse strains. This cross, which segregates for genotype and physiological traits, was previously used to identify several diabetes-related QTL. Our current investigation includes microarray analysis of over 40,000 probe sets, plus quantitative mass spectrometry-based measurements of sixty-seven intermediary metabolites in three different classes (amino acids, organic acids, and acyl-carnitines. We show that liver metabolites map to distinct genetic regions, thereby indicating that tissue metabolites are heritable. We also demonstrate that genomic analysis can be integrated with liver mRNA expression and metabolite profiling data to construct causal networks for control of specific metabolic processes in liver. As a proof of principle of the practical significance of this integrative approach, we illustrate the construction of a specific causal network that links gene expression and metabolic changes in the context of glutamate metabolism, and demonstrate its validity by showing that genes in the network respond to changes in glutamine and glutamate availability. Thus, the methods described here have the potential to reveal regulatory networks that contribute to chronic, complex, and highly prevalent diseases and conditions such as obesity and diabetes.

  3. Robust identification of transcriptional regulatory networks using a Gibbs sampler on outlier sum statistic.

    Science.gov (United States)

    Gu, Jinghua; Xuan, Jianhua; Riggins, Rebecca B; Chen, Li; Wang, Yue; Clarke, Robert

    2012-08-01

    Identification of transcriptional regulatory networks (TRNs) is of significant importance in computational biology for cancer research, providing a critical building block to unravel disease pathways. However, existing methods for TRN identification suffer from the inclusion of excessive 'noise' in microarray data and false-positives in binding data, especially when applied to human tumor-derived cell line studies. More robust methods that can counteract the imperfection of data sources are therefore needed for reliable identification of TRNs in this context. In this article, we propose to establish a link between the quality of one target gene to represent its regulator and the uncertainty of its expression to represent other target genes. Specifically, an outlier sum statistic was used to measure the aggregated evidence for regulation events between target genes and their corresponding transcription factors. A Gibbs sampling method was then developed to estimate the marginal distribution of the outlier sum statistic, hence, to uncover underlying regulatory relationships. To evaluate the effectiveness of our proposed method, we compared its performance with that of an existing sampling-based method using both simulation data and yeast cell cycle data. The experimental results show that our method consistently outperforms the competing method in different settings of signal-to-noise ratio and network topology, indicating its robustness for biological applications. Finally, we applied our method to breast cancer cell line data and demonstrated its ability to extract biologically meaningful regulatory modules related to estrogen signaling and action in breast cancer. The Gibbs sampler MATLAB package is freely available at http://www.cbil.ece.vt.edu/software.htm. xuan@vt.edu Supplementary data are available at Bioinformatics online.

  4. Coordinate Regulation of Yeast Sterol Regulatory Element-binding Protein (SREBP) and Mga2 Transcription Factors.

    Science.gov (United States)

    Burr, Risa; Stewart, Emerson V; Espenshade, Peter J

    2017-03-31

    The Mga2 and Sre1 transcription factors regulate oxygen-responsive lipid homeostasis in the fission yeast Schizosaccharomyces pombe in a manner analogous to the mammalian sterol regulatory element-binding protein (SREBP)-1 and SREBP-2 transcription factors. Mga2 and SREBP-1 regulate triacylglycerol and glycerophospholipid synthesis, whereas Sre1 and SREBP-2 regulate sterol synthesis. In mammals, a shared activation mechanism allows for coordinate regulation of SREBP-1 and SREBP-2. In contrast, distinct pathways activate fission yeast Mga2 and Sre1. Therefore, it is unclear whether and how these two related pathways are coordinated to maintain lipid balance in fission yeast. Previously, we showed that Sre1 cleavage is defective in the absence of mga2 Here, we report that this defect is due to deficient unsaturated fatty acid synthesis, resulting in aberrant membrane transport. This defect is recapitulated by treatment with the fatty acid synthase inhibitor cerulenin and is rescued by addition of exogenous unsaturated fatty acids. Furthermore, sterol synthesis inhibition blocks Mga2 pathway activation. Together, these data demonstrate that Sre1 and Mga2 are each regulated by the lipid product of the other transcription factor pathway, providing a source of coordination for these two branches of lipid synthesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Transcriptome landscape of Lactococcus lactis reveals many novel RNAs including a small regulatory RNA involved in carbon uptake and metabolism.

    Science.gov (United States)

    van der Meulen, Sjoerd B; de Jong, Anne; Kok, Jan

    2016-01-01

    RNA sequencing has revolutionized genome-wide transcriptome analyses, and the identification of non-coding regulatory RNAs in bacteria has thus increased concurrently. Here we reveal the transcriptome map of the lactic acid bacterial paradigm Lactococcus lactis MG1363 by employing differential RNA sequencing (dRNA-seq) and a combination of manual and automated transcriptome mining. This resulted in a high-resolution genome annotation of L. lactis and the identification of 60 cis-encoded antisense RNAs (asRNAs), 186 trans-encoded putative regulatory RNAs (sRNAs) and 134 novel small ORFs. Based on the putative targets of asRNAs, a novel classification is proposed. Several transcription factor DNA binding motifs were identified in the promoter sequences of (a)sRNAs, providing insight in the interplay between lactococcal regulatory RNAs and transcription factors. The presence and lengths of 14 putative sRNAs were experimentally confirmed by differential Northern hybridization, including the abundant RNA 6S that is differentially expressed depending on the available carbon source. For another sRNA, LLMGnc_147, functional analysis revealed that it is involved in carbon uptake and metabolism. L. lactis contains 13% leaderless mRNAs (lmRNAs) that, from an analysis of overrepresentation in GO classes, seem predominantly involved in nucleotide metabolism and DNA/RNA binding. Moreover, an A-rich sequence motif immediately following the start codon was uncovered, which could provide novel insight in the translation of lmRNAs. Altogether, this first experimental genome-wide assessment of the transcriptome landscape of L. lactis and subsequent sRNA studies provide an extensive basis for the investigation of regulatory RNAs in L. lactis and related lactococcal species.

  6. Feast/famine regulatory proteins (FFRPs): Escherichia coli Lrp, AsnC and related archaeal transcription factors.

    Science.gov (United States)

    Yokoyama, Katsushi; Ishijima, Sanae A; Clowney, Lester; Koike, Hideaki; Aramaki, Hironori; Tanaka, Chikako; Makino, Kozo; Suzuki, Masashi

    2006-01-01

    Feast/famine regulatory proteins comprise a diverse family of transcription factors, which have been referred to in various individual identifications, including Escherichia coli leucine-responsive regulatory protein and asparagine synthase C gene product. A full length feast/famine regulatory protein consists of the N-terminal DNA-binding domain and the C-domain, which is involved in dimerization and further assembly, thereby producing, for example, a disc or a chromatin-like cylinder. Various ligands of the size of amino acids bind at the interface between feast/famine regulatory protein dimers, thereby altering their assembly forms. Also, the combination of feast/famine regulatory protein subunits forming the same assembly is altered. In this way, a small number of feast/famine regulatory proteins are able to regulate a large number of genes in response to various environmental changes. Because feast/famine regulatory proteins are shared by archaea and eubacteria, the genome-wide regulation by feast/famine regulatory proteins is traceable back to their common ancestor, being the prototype of highly differentiated transcription regulatory mechanisms found in organisms nowadays.

  7. Identification of direct regulatory targets of the transcription factor Sox10 based on function and conservation

    Directory of Open Access Journals (Sweden)

    Lee Sanghyuk

    2008-09-01

    Full Text Available Abstract Background Sox10, a member of the Sry-related HMG-Box gene family, is a critical transcription factor for several important cell lineages, most notably the neural crest stem cells and the derivative peripheral glial cells and melanocytes. Thus far, only a handful of direct target genes are known for this transcription factor limiting our understanding of the biological network it governs. Results We describe identification of multiple direct regulatory target genes of Sox10 through a procedure based on function and conservation. By combining RNA interference technique and DNA microarray technology, we have identified a set of genes that show significant down-regulation upon introduction of Sox10 specific siRNA into Schwannoma cells. Subsequent comparative genomics analyses led to potential binding sites for Sox10 protein conserved across several mammalian species within the genomic region proximal to these genes. Multiple sites belonging to 4 different genes (proteolipid protein, Sox10, extracellular superoxide dismutase, and pleiotrophin were shown to directly interact with Sox10 by chromatin immunoprecipitation assay. We further confirmed the direct regulation through the identified cis-element for one of the genes, extracellular superoxide dismutase, using electrophoretic mobility shift assay and reporter assay. Conclusion In sum, the process of combining differential expression profiling and comparative genomics successfully led to further defining the role of Sox10, a critical transcription factor for the development of peripheral glia. Our strategy utilizing relatively accessible techniques and tools should be applicable to studying the function of other transcription factors.

  8. Genome-wide identification of the regulatory targets of a transcription factor using biochemical characterization and computational genomic analysis

    Directory of Open Access Journals (Sweden)

    Jolly Emmitt R

    2005-11-01

    Full Text Available Abstract Background A major challenge in computational genomics is the development of methodologies that allow accurate genome-wide prediction of the regulatory targets of a transcription factor. We present a method for target identification that combines experimental characterization of binding requirements with computational genomic analysis. Results Our method identified potential target genes of the transcription factor Ndt80, a key transcriptional regulator involved in yeast sporulation, using the combined information of binding affinity, positional distribution, and conservation of the binding sites across multiple species. We have also developed a mathematical approach to compute the false positive rate and the total number of targets in the genome based on the multiple selection criteria. Conclusion We have shown that combining biochemical characterization and computational genomic analysis leads to accurate identification of the genome-wide targets of a transcription factor. The method can be extended to other transcription factors and can complement other genomic approaches to transcriptional regulation.

  9. A genomic approach to identify regulatory nodes in the transcriptional network of systemic acquired resistance in plants.

    Directory of Open Access Journals (Sweden)

    Dong Wang

    2006-11-01

    Full Text Available Many biological processes are controlled by intricate networks of transcriptional regulators. With the development of microarray technology, transcriptional changes can be examined at the whole-genome level. However, such analysis often lacks information on the hierarchical relationship between components of a given system. Systemic acquired resistance (SAR is an inducible plant defense response involving a cascade of transcriptional events induced by salicylic acid through the transcription cofactor NPR1. To identify additional regulatory nodes in the SAR network, we performed microarray analysis on Arabidopsis plants expressing the NPR1-GR (glucocorticoid receptor fusion protein. Since nuclear translocation of NPR1-GR requires dexamethasone, we were able to control NPR1-dependent transcription and identify direct transcriptional targets of NPR1. We show that NPR1 directly upregulates the expression of eight WRKY transcription factor genes. This large family of 74 transcription factors has been implicated in various defense responses, but no specific WRKY factor has been placed in the SAR network. Identification of NPR1-regulated WRKY factors allowed us to perform in-depth genetic analysis on a small number of WRKY factors and test well-defined phenotypes of single and double mutants associated with NPR1. Among these WRKY factors we found both positive and negative regulators of SAR. This genomics-directed approach unambiguously positioned five WRKY factors in the complex transcriptional regulatory network of SAR. Our work not only discovered new transcription regulatory components in the signaling network of SAR but also demonstrated that functional studies of large gene families have to take into consideration sequence similarity as well as the expression patterns of the candidates.

  10. Combined effect of regulatory polymorphisms on transcription of UGT1A1 as a cause of Gilbert syndrome

    Directory of Open Access Journals (Sweden)

    Sato Hiroshi

    2010-06-01

    Full Text Available Abstract Background Gilbert syndrome is caused by defects in bilirubin UDP-glucuronosyltransferase (UGT1A1. The most common variation believed to be involved is A(TA7TAA. Although several polymorphisms have been found to link with A(TA7TAA, the combined effect of regulatory polymorphisms in the development of Gilbert syndrome remains unclear. Methods In an analysis of 15 patients and 60 normal subjects, we detected 14 polymorphisms and nine haplotypes in the regulatory region. We classified the 4-kbp regulatory region of the patients into: the TATA box including A(TA7TAA; a phenobarbital responsive enhancer module including c.-3275T>G; and a region including other ten linked polymorphisms. The effect on transcription of these polymorphisms was studied. Results All haplotypes with A(TA7TAA had c.-3275T>G and additional polymorphisms. In an in-vitro expression study of the 4-kbp regulatory region, A(TA7TAA alone did not significantly reduce transcription. In contrast, c.-3275T>G reduced transcription to 69% of that of wild type, and the linked polymorphisms reduced transcription to 88% of wild type. Transcription of the typical regulatory region of the patients was 56% of wild type. Co-expression of constitutive androstane receptor (CAR increased the transcription of wild type by a factor of 4.3. Each polymorphism by itself did not reduce transcription to the level of the patients, however, even in the presence of CAR. Conclusions These results imply that co-operation of A(TA7TAA, c.-3275T>G and the linked polymorphisms is necessary in causing Gilbert syndrome.

  11. A conserved RNA structural element within the hepatitis B virus post-transcriptional regulatory element enhance nuclear export of intronless transcripts and repress the splicing mechanism.

    Science.gov (United States)

    Visootsat, Akasit; Payungporn, Sunchai; T-Thienprasert, Nattanan P

    2015-12-01

    Hepatitis B virus (HBV) infection is a primary cause of hepatocellular carcinoma and liver cirrhosis worldwide. To develop novel antiviral drugs, a better understanding of HBV gene expression regulation is vital. One important aspect is to understand how HBV hijacks the cellular machinery to export unspliced RNA from the nucleus. The HBV post-transcriptional regulatory element (HBV PRE) has been proposed to be the HBV RNA nuclear export element. However, the function remains controversial, and the core element is unclear. This study, therefore, aimed to identify functional regulatory elements within the HBV PRE and investigate their functions. Using bioinformatics programs based on sequence conservation and conserved RNA secondary structures, three regulatory elements were predicted, namely PRE 1151-1410, PRE 1520-1620 and PRE 1650-1684. PRE 1151-1410 significantly increased intronless and unspliced luciferase activity in both HepG2 and COS-7 cells. Likewise, PRE 1151-1410 significantly elevated intronless and unspliced HBV surface transcripts in liver cancer cells. Moreover, motif analysis predicted that PRE 1151-1410 contains several regulatory motifs. This study reported the roles of PRE 1151-1410 in intronless transcript nuclear export and the splicing mechanism. Additionally, these results provide knowledge in the field of HBV RNA regulation. Moreover, PRE 1151-1410 may be used to enhance the expression of other mRNAs in intronless reporter plasmids.

  12. Transcriptional response of P. pastoris in fed-batch cultivations to Rhizopus oryzae lipase production reveals UPR induction

    Directory of Open Access Journals (Sweden)

    Valero Francisco

    2007-07-01

    Full Text Available Abstract Background The analysis of transcriptional levels of the genes involved in protein synthesis and secretion is a key factor to understand the host organism's responses to recombinant protein production, as well as their interaction with the cultivation conditions. Novel techniques such as the sandwich hybridization allow monitoring quantitatively the dynamic changes of specific RNAs. In this study, the transcriptional levels of some genes related to the unfolded protein response (UPR and central metabolism of Pichia pastoris were analysed during batch and fed-batch cultivations using an X-33-derived strain expressing a Rhizopus oryzae lipase under control of the formaldehyde dehydrogenase promoter (FLD1, namely the alcohol oxidase gene AOX1, the formaldehyde dehydrogenase FLD1, the protein disulfide isomerase PDI, the KAR2 gene coding for the BiP chaperone, the 26S rRNA and the R. oryzae lipase gene ROL. Results The transcriptional levels of the selected set of genes were first analysed in P. pastoris cells growing in shake flask cultures containing different carbon and nitrogen sources combinations, glycerol + ammonium, methanol + methylamine and sorbitol + methylamine. The transcriptional levels of the AOX1 and FLD1 genes were coherent with the known regulatory mechanism of C1 substrates in P. pastoris, whereas ROL induction lead to the up-regulation of KAR2 and PDI transcriptional levels, thus suggesting that ROL overexpression triggers the UPR. This was further confirmed in fed-batch cultivations performed at different growth rates. Transcriptional levels of the analysed set of genes were generally higher at higher growth rates. Nevertheless, when ROL was overexpressed in a strain having the UPR constitutively activated, significantly lower relative induction levels of these marker genes were detected. Conclusion The bead-based sandwich hybridization assay has shown its potential as a reliable instrument for quantification of

  13. Exosome proteomics reveals transcriptional regulator proteins with potential to mediate downstream pathways.

    Science.gov (United States)

    Ung, Timothy H; Madsen, Helen J; Hellwinkel, Justin E; Lencioni, Alex M; Graner, Michael W

    2014-11-01

    Exosomes are virus-sized, membrane-enclosed vesicles with origins in the cellular endosomal system, but are released extracellularly. As a population, these tiny vesicles carry relatively enormous amounts of information in their protein, lipid and nucleic acid content, and the vesicles can have profound impacts on recipient cells. This review employs publically-available data combined with gene ontology applications to propose a novel concept, that exosomes transport transcriptional and translational machinery that may have direct impacts on gene expression in recipient cells. Here, we examine the previously published proteomic contents of medulloblastoma-derived exosomes, focusing on transcriptional regulators; we found that there are numerous proteins that may have potential roles in transcriptional and translational regulation with putative influence on downstream, cancer-related pathways. We expanded this search to all of the proteins in the Vesiclepedia database; using gene ontology approaches, we see that these regulatory factors are implicated in many of the processes involved in cancer initiation and progression. This information suggests that some of the effects of exosomes on recipient cells may be due to the delivery of protein factors that can directly and fundamentally change the transcriptional landscape of the cells. Within a tumor environment, this has potential to tilt the advantage towards the cancer. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  14. Sasquatch: predicting the impact of regulatory SNPs on transcription factor binding from cell- and tissue-specific DNase footprints

    OpenAIRE

    Schwessinger, R; Suciu, MC; McGowan, SJ; Telenius, J; Taylor, S; Higgs, DR; Hughes, JR

    2017-01-01

    In the era of genome-wide association studies (GWAS) and personalized medicine, predicting the impact of single nucleotide polymorphisms (SNPs) in regulatory elements is an important goal. Current approaches to determine the potential of regulatory SNPs depend on inadequate knowledge of cell-specific DNA binding motifs. Here, we present Sasquatch, a new computational approach that uses DNase footprint data to estimate and visualize the effects of noncoding variants on transcription factor bin...

  15. The Rts1 regulatory subunit of protein phosphatase 2A is required for control of G1 cyclin transcription and nutrient modulation of cell size.

    Directory of Open Access Journals (Sweden)

    Karen Artiles

    2009-11-01

    Full Text Available The key molecular event that marks entry into the cell cycle is transcription of G1 cyclins, which bind and activate cyclin-dependent kinases. In yeast cells, initiation of G1 cyclin transcription is linked to achievement of a critical cell size, which contributes to cell-size homeostasis. The critical cell size is modulated by nutrients, such that cells growing in poor nutrients are smaller than cells growing in rich nutrients. Nutrient modulation of cell size does not work through known critical regulators of G1 cyclin transcription and is therefore thought to work through a distinct pathway. Here, we report that Rts1, a highly conserved regulatory subunit of protein phosphatase 2A (PP2A, is required for normal control of G1 cyclin transcription. Loss of Rts1 caused delayed initiation of bud growth and delayed and reduced accumulation of G1 cyclins. Expression of the G1 cyclin CLN2 from an inducible promoter rescued the delayed bud growth in rts1Delta cells, indicating that Rts1 acts at the level of transcription. Moreover, loss of Rts1 caused altered regulation of Swi6, a key component of the SBF transcription factor that controls G1 cyclin transcription. Epistasis analysis revealed that Rts1 does not work solely through several known critical upstream regulators of G1 cyclin transcription. Cells lacking Rts1 failed to undergo nutrient modulation of cell size. Together, these observations demonstrate that Rts1 is a key player in pathways that link nutrient availability, cell size, and G1 cyclin transcription. Since Rts1 is highly conserved, it may function in similar pathways in vertebrates.

  16. Transcript specificity in yeast pre-mRNA splicing revealed by mutations in core spliceosomal components.

    Directory of Open Access Journals (Sweden)

    Jeffrey A Pleiss

    2007-04-01

    Full Text Available Appropriate expression of most eukaryotic genes requires the removal of introns from their pre-messenger RNAs (pre-mRNAs, a process catalyzed by the spliceosome. In higher eukaryotes a large family of auxiliary factors known as SR proteins can improve the splicing efficiency of transcripts containing suboptimal splice sites by interacting with distinct sequences present in those pre-mRNAs. The yeast Saccharomyces cerevisiae lacks functional equivalents of most of these factors; thus, it has been unclear whether the spliceosome could effectively distinguish among transcripts. To address this question, we have used a microarray-based approach to examine the effects of mutations in 18 highly conserved core components of the spliceosomal machinery. The kinetic profiles reveal clear differences in the splicing defects of particular pre-mRNA substrates. Most notably, the behaviors of ribosomal protein gene transcripts are generally distinct from other intron-containing transcripts in response to several spliceosomal mutations. However, dramatically different behaviors can be seen for some pairs of transcripts encoding ribosomal protein gene paralogs, suggesting that the spliceosome can readily distinguish between otherwise highly similar pre-mRNAs. The ability of the spliceosome to distinguish among its different substrates may therefore offer an important opportunity for yeast to regulate gene expression in a transcript-dependent fashion. Given the high level of conservation of core spliceosomal components across eukaryotes, we expect that these results will significantly impact our understanding of how regulated splicing is controlled in higher eukaryotes as well.

  17. Inference of RNA decay rate from transcriptional profiling highlights the regulatory programs of Alzheimer's disease.

    Science.gov (United States)

    Alkallas, Rached; Fish, Lisa; Goodarzi, Hani; Najafabadi, Hamed S

    2017-10-13

    The abundance of mRNA is mainly determined by the rates of RNA transcription and decay. Here, we present a method for unbiased estimation of differential mRNA decay rate from RNA-sequencing data by modeling the kinetics of mRNA metabolism. We show that in all primary human tissues tested, and particularly in the central nervous system, many pathways are regulated at the mRNA stability level. We present a parsimonious regulatory model consisting of two RNA-binding proteins and four microRNAs that modulate the mRNA stability landscape of the brain, which suggests a new link between RBFOX proteins and Alzheimer's disease. We show that downregulation of RBFOX1 leads to destabilization of mRNAs encoding for synaptic transmission proteins, which may contribute to the loss of synaptic function in Alzheimer's disease. RBFOX1 downregulation is more likely to occur in older and female individuals, consistent with the association of Alzheimer's disease with age and gender."mRNA abundance is determined by the rates of transcription and decay. Here, the authors propose a method for estimating the rate of differential mRNA decay from RNA-seq data and model mRNA stability in the brain, suggesting a link between mRNA stability and Alzheimer's disease."

  18. Conservation of lipid metabolic gene transcriptional regulatory networks in fish and mammals.

    Science.gov (United States)

    Carmona-Antoñanzas, Greta; Tocher, Douglas R; Martinez-Rubio, Laura; Leaver, Michael J

    2014-01-15

    Lipid content and composition in aquafeeds have changed rapidly as a result of the recent drive to replace ecologically limited marine ingredients, fishmeal and fish oil (FO). Terrestrial plant products are the most economic and sustainable alternative; however, plant meals and oils are devoid of physiologically important cholesterol and long-chain polyunsaturated fatty acids (LC-PUFA), eicosapentaenoic (EPA), docosahexaenoic (DHA) and arachidonic (ARA) acids. Although replacement of dietary FO with vegetable oil (VO) has little effect on growth in Atlantic salmon (Salmo salar), several studies have shown major effects on the activity and expression of genes involved in lipid homeostasis. In vertebrates, sterols and LC-PUFA play crucial roles in lipid metabolism by direct interaction with lipid-sensing transcription factors (TFs) and consequent regulation of target genes. The primary aim of the present study was to elucidate the role of key TFs in the transcriptional regulation of lipid metabolism in fish by transfection and overexpression of TFs. The results show that the expression of genes of LC-PUFA biosynthesis (elovl and fads2) and cholesterol metabolism (abca1) are regulated by Lxr and Srebp TFs in salmon, indicating highly conserved regulatory mechanism across vertebrates. In addition, srebp1 and srebp2 mRNA respond to replacement of dietary FO with VO. Thus, Atlantic salmon adjust lipid metabolism in response to dietary lipid composition through the transcriptional regulation of gene expression. It may be possible to further increase efficient and effective use of sustainable alternatives to marine products in aquaculture by considering these important molecular interactions when formulating diets. © 2013.

  19. Comprehensive reanalysis of transcription factor knockout expression data in Saccharomyces cerevisiae reveals many new targets.

    Science.gov (United States)

    Reimand, Jüri; Vaquerizas, Juan M; Todd, Annabel E; Vilo, Jaak; Luscombe, Nicholas M

    2010-08-01

    Transcription factor (TF) perturbation experiments give valuable insights into gene regulation. Genome-scale evidence from microarray measurements may be used to identify regulatory interactions between TFs and targets. Recently, Hu and colleagues published a comprehensive study covering 269 TF knockout mutants for the yeast Saccharomyces cerevisiae. However, the information that can be extracted from this valuable dataset is limited by the method employed to process the microarray data. Here, we present a reanalysis of the original data using improved statistical techniques freely available from the BioConductor project. We identify over 100,000 differentially expressed genes-nine times the total reported by Hu et al. We validate the biological significance of these genes by assessing their functions, the occurrence of upstream TF-binding sites, and the prevalence of protein-protein interactions. The reanalysed dataset outperforms the original across all measures, indicating that we have uncovered a vastly expanded list of relevant targets. In summary, this work presents a high-quality reanalysis that maximizes the information contained in the Hu et al. compendium. The dataset is available from ArrayExpress (accession: E-MTAB-109) and it will be invaluable to any scientist interested in the yeast transcriptional regulatory system.

  20. COMK ENCODES THE COMPETENCE TRANSCRIPTION FACTOR, THE KEY REGULATORY PROTEIN FOR COMPETENCE DEVELOPMENT IN BACILLUS-SUBTILIS

    NARCIS (Netherlands)

    VANSINDEREN, D; LUTTINGER, A; KONG, LY; DUBNAU, D; VENEMA, G; HAMOEN, L

    comK is a positive autoregulatory gene occupying a central position in the com petence-signal-transduction network. All regulatory routes identified in this network converge at the level of comK expression. The ComK protein is required for the transcriptional induction of comK and the late

  1. Time-resolved transcriptome and proteome landscape of human regulatory T cell (Treg) differentiation reveals novel regulators of FOXP3

    KAUST Repository

    Schmidt, Angelika; Marabita, Francesco; Kiani, Narsis A.; Gross, Catharina C.; Johansson, Henrik J.; É liá s, Szabolcs; Rautio, Sini; Eriksson, Matilda; Fernandes, Sunjay Jude; Silberberg, Gilad; Ullah, Ubaid; Bhatia, Urvashi; Lä hdesmä ki, Harri; Lehtiö , Janne; Gomez-Cabrero, David; Wiendl, Heinz; Lahesmaa, Riitta; Tegner, Jesper

    2018-01-01

    BackgroundRegulatory T cells (Tregs) expressing the transcription factor FOXP3 are crucial mediators of self-tolerance, preventing autoimmune diseases but possibly hampering tumor rejection. Clinical manipulation of Tregs is of great interest

  2. The transcriptional regulatory network mediated by banana (Musa acuminata) dehydration-responsive element binding (MaDREB) transcription factors in fruit ripening.

    Science.gov (United States)

    Kuang, Jian-Fei; Chen, Jian-Ye; Liu, Xun-Cheng; Han, Yan-Chao; Xiao, Yun-Yi; Shan, Wei; Tang, Yang; Wu, Ke-Qiang; He, Jun-Xian; Lu, Wang-Jin

    2017-04-01

    Fruit ripening is a complex, genetically programmed process involving the action of critical transcription factors (TFs). Despite the established significance of dehydration-responsive element binding (DREB) TFs in plant abiotic stress responses, the involvement of DREBs in fruit ripening is yet to be determined. Here, we identified four genes encoding ripening-regulated DREB TFs in banana (Musa acuminata), MaDREB1, MaDREB2, MaDREB3, and MaDREB4, and demonstrated that they play regulatory roles in fruit ripening. We showed that MaDREB1-MaDREB4 are nucleus-localized, induced by ethylene and encompass transcriptional activation activities. We performed a genome-wide chromatin immunoprecipitation and high-throughput sequencing (ChIP-Seq) experiment for MaDREB2 and identified 697 genomic regions as potential targets of MaDREB2. MaDREB2 binds to hundreds of loci with diverse functions and its binding sites are distributed in the promoter regions proximal to the transcriptional start site (TSS). Most of the MaDREB2-binding targets contain the conserved (A/G)CC(G/C)AC motif and MaDREB2 appears to directly regulate the expression of a number of genes involved in fruit ripening. In combination with transcriptome profiling (RNA sequencing) data, our results indicate that MaDREB2 may serve as both transcriptional activator and repressor during banana fruit ripening. In conclusion, our study suggests a hierarchical regulatory model of fruit ripening in banana and that the MaDREB TFs may act as transcriptional regulators in the regulatory network. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  3. Bladder inflammatory transcriptome in response to tachykinins: Neurokinin 1 receptor-dependent genes and transcription regulatory elements

    Directory of Open Access Journals (Sweden)

    Dozmorov Igor

    2007-05-01

    Full Text Available Abstract Background Tachykinins (TK, such as substance P, and their neurokinin receptors which are ubiquitously expressed in the human urinary tract, represent an endogenous system regulating bladder inflammatory, immune responses, and visceral hypersensitivity. Increasing evidence correlates alterations in the TK system with urinary tract diseases such as neurogenic bladders, outflow obstruction, idiopathic detrusor instability, and interstitial cystitis. However, despite promising effects in animal models, there seems to be no published clinical study showing that NK-receptor antagonists are an effective treatment of pain in general or urinary tract disorders, such as detrusor overactivity. In order to search for therapeutic targets that could block the tachykinin system, we set forth to determine the regulatory network downstream of NK1 receptor activation. First, NK1R-dependent transcripts were determined and used to query known databases for their respective transcription regulatory elements (TREs. Methods An expression analysis was performed using urinary bladders isolated from sensitized wild type (WT and NK1R-/- mice that were stimulated with saline, LPS, or antigen to provoke inflammation. Based on cDNA array results, NK1R-dependent genes were selected. PAINT software was used to query TRANSFAC database and to retrieve upstream TREs that were confirmed by electrophoretic mobility shift assays. Results The regulatory network of TREs driving NK1R-dependent genes presented cRel in a central position driving 22% of all genes, followed by AP-1, NF-kappaB, v-Myb, CRE-BP1/c-Jun, USF, Pax-6, Efr-1, Egr-3, and AREB6. A comparison between NK1R-dependent and NK1R-independent genes revealed Nkx-2.5 as a unique discriminator. In the presence of NK1R, Nkx2-5 _01 was significantly correlated with 36 transcripts which included several candidates for mediating bladder development (FGF and inflammation (PAR-3, IL-1R, IL-6, α-NGF, TSP2. In the absence of

  4. Senataxin Mutation Reveals How R-Loops Promote Transcription by Blocking DNA Methylation at Gene Promoters.

    Science.gov (United States)

    Grunseich, Christopher; Wang, Isabel X; Watts, Jason A; Burdick, Joshua T; Guber, Robert D; Zhu, Zhengwei; Bruzel, Alan; Lanman, Tyler; Chen, Kelian; Schindler, Alice B; Edwards, Nancy; Ray-Chaudhury, Abhik; Yao, Jianhua; Lehky, Tanya; Piszczek, Grzegorz; Crain, Barbara; Fischbeck, Kenneth H; Cheung, Vivian G

    2018-02-01

    R-loops are three-stranded nucleic acid structures found abundantly and yet often viewed as by-products of transcription. Studying cells from patients with a motor neuron disease (amyotrophic lateral sclerosis 4 [ALS4]) caused by a mutation in senataxin, we uncovered how R-loops promote transcription. In ALS4 patients, the senataxin mutation depletes R-loops with a consequent effect on gene expression. With fewer R-loops in ALS4 cells, the expression of BAMBI, a negative regulator of transforming growth factor β (TGF-β), is reduced; that then leads to the activation of the TGF-β pathway. We uncovered that genome-wide R-loops influence promoter methylation of over 1,200 human genes. DNA methyl-transferase 1 favors binding to double-stranded DNA over R-loops. Thus, in forming R-loops, nascent RNA blocks DNA methylation and promotes further transcription. Hence, our results show that nucleic acid structures, in addition to sequences, influence the binding and activity of regulatory proteins. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Core transcriptional regulatory circuit controlled by the TAL1 complex in human T cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Sanda, Takaomi; Lawton, Lee N; Barrasa, M Inmaculada; Fan, Zi Peng; Kohlhammer, Holger; Gutierrez, Alejandro; Ma, Wenxue; Tatarek, Jessica; Ahn, Yebin; Kelliher, Michelle A; Jamieson, Catriona H M; Staudt, Louis M; Young, Richard A; Look, A Thomas

    2012-08-14

    The oncogenic transcription factor TAL1/SCL is aberrantly expressed in over 40% of cases of human T cell acute lymphoblastic leukemia (T-ALL), emphasizing its importance in the molecular pathogenesis of T-ALL. Here we identify the core transcriptional regulatory circuit controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3, and RUNX1. We show that TAL1 forms a positive interconnected autoregulatory loop with GATA3 and RUNX1 and that the TAL1 complex directly activates the MYB oncogene, forming a positive feed-forward regulatory loop that reinforces and stabilizes the TAL1-regulated oncogenic program. One of the critical downstream targets in this circuitry is the TRIB2 gene, which is oppositely regulated by TAL1 and E2A/HEB and is essential for the survival of T-ALL cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Proteomic analysis reveals metabolic and regulatory systems involved the syntrophic and axenic lifestyle of Syntrophomonas wolfei.

    Directory of Open Access Journals (Sweden)

    Jessica Rhea Sieber

    2015-02-01

    Full Text Available Microbial syntrophy is a vital metabolic interaction necessary for the complete oxidation of organic biomass to methane in all-anaerobic ecosystems. However, this process is thermodynamically constrained and represents an ecosystem-level metabolic bottleneck. To gain insight into the physiology of this process, a shotgun proteomic approach was used to quantify the protein landscape of the model syntrophic metabolizer, Syntrophomonas wolfei, grown axenically and syntrophically with Methanospirillum hungatei. Remarkably, the abundance of most proteins as represented by normalized spectral abundance factor (NSAF value changed very little between the pure and coculture growth conditions. Among the most abundant proteins detected were GroEL and GroES chaperonins, a small heat shock protein, and proteins involved in electron transfer, beta-oxidation, and ATP synthesis. Several putative energy conservation enzyme systems that utilize NADH and ferredoxin were present. The abundance of an EtfAB2 and the membrane-bound iron-sulfur oxidoreductase (Swol_0698 gene product delineated a potential conduit for electron transfer between acyl-CoA dehydrogenases and membrane redox carriers. Proteins detected only when S. wolfei was grown with M. hungatei included a zinc-dependent dehydrogenase with a GroES domain, whose gene is present in genomes in many organisms capable of syntrophy, and transcriptional regulators responsive to environmental stimuli or the physiological status of the cell. The proteomic analysis revealed an emphasis macromolecular stability and energy metabolism to S. wolfei and presence of regulatory mechanisms responsive to external stimuli and cellular physiological status.

  7. Cis-regulatory signatures of orthologous stress-associated bZIP transcription factors from rice, sorghum and Arabidopsis based on phylogenetic footprints

    Directory of Open Access Journals (Sweden)

    Xu Fuyu

    2012-09-01

    Full Text Available Abstract Background The potential contribution of upstream sequence variation to the unique features of orthologous genes is just beginning to be unraveled. A core subset of stress-associated bZIP transcription factors from rice (Oryza sativa formed ten clusters of orthologous groups (COG with genes from the monocot sorghum (Sorghum bicolor and dicot Arabidopsis (Arabidopsis thaliana. The total cis-regulatory information content of each stress-associated COG was examined by phylogenetic footprinting to reveal ortholog-specific, lineage-specific and species-specific conservation patterns. Results The most apparent pattern observed was the occurrence of spatially conserved ‘core modules’ among the COGs but not among paralogs. These core modules are comprised of various combinations of two to four putative transcription factor binding site (TFBS classes associated with either developmental or stress-related functions. Outside the core modules are specific stress (ABA, oxidative, abiotic, biotic or organ-associated signals, which may be functioning as ‘regulatory fine-tuners’ and further define lineage-specific and species-specific cis-regulatory signatures. Orthologous monocot and dicot promoters have distinct TFBS classes involved in disease and oxidative-regulated expression, while the orthologous rice and sorghum promoters have distinct combinations of root-specific signals, a pattern that is not particularly conserved in Arabidopsis. Conclusions Patterns of cis-regulatory conservation imply that each ortholog has distinct signatures, further suggesting that they are potentially unique in a regulatory context despite the presumed conservation of broad biological function during speciation. Based on the observed patterns of conservation, we postulate that core modules are likely primary determinants of basal developmental programming, which may be integrated with and further elaborated by additional intrinsic or extrinsic signals in

  8. A DNA-binding-site landscape and regulatory network analysis for NAC transcription factors in Arabidopsis thaliana

    DEFF Research Database (Denmark)

    Lindemose, Søren; Jensen, Michael Krogh; de Velde, Jan Van

    2014-01-01

    regulatory networks of 12 NAC transcription factors. Our data offer specific single-base resolution fingerprints for most TFs studied and indicate that NAC DNA-binding specificities might be predicted from their DNA-binding domain's sequence. The developed methodology, including the application......Target gene identification for transcription factors is a prerequisite for the systems wide understanding of organismal behaviour. NAM-ATAF1/2-CUC2 (NAC) transcription factors are amongst the largest transcription factor families in plants, yet limited data exist from unbiased approaches to resolve...... the DNA-binding preferences of individual members. Here, we present a TF-target gene identification workflow based on the integration of novel protein binding microarray data with gene expression and multi-species promoter sequence conservation to identify the DNA-binding specificities and the gene...

  9. The upstream regulatory sequence of the light harvesting complex Lhcf2 gene of the marine diatom Phaeodactylum tricornutum enhances transcription in an orientation- and distance-independent fashion.

    Science.gov (United States)

    Russo, Monia Teresa; Annunziata, Rossella; Sanges, Remo; Ferrante, Maria Immacolata; Falciatore, Angela

    2015-12-01

    Diatoms are a key phytoplankton group in the contemporary ocean, showing extraordinary adaptation capacities to rapidly changing environments. The recent availability of whole genome sequences from representative species has revealed distinct features in their genomes, like novel combinations of genes encoding distinct metabolisms and a significant number of diatom-specific genes. However, the regulatory mechanisms driving diatom gene expression are still largely uncharacterized. Considering the wide variety of fields of study orbiting diatoms, ranging from ecology, evolutionary biology to biotechnology, it is thus essential to increase our understanding of fundamental gene regulatory processes such as transcriptional regulation. To this aim, we explored the functional properties of the 5'-flanking region of the Phaeodatylum tricornutum Lhcf2 gene, encoding a member of the Light Harvesting Complex superfamily and we showed that this region enhances transcription of a GUS reporter gene in an orientation- and distance-independent fashion. This represents the first example of a cis-regulatory sequence with enhancer-like features discovered in diatoms and it is instrumental for the generation of novel genetic tools and diatom exploitation in different areas of study. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Circuit-wide Transcriptional Profiling Reveals Brain Region-Specific Gene Networks Regulating Depression Susceptibility.

    Science.gov (United States)

    Bagot, Rosemary C; Cates, Hannah M; Purushothaman, Immanuel; Lorsch, Zachary S; Walker, Deena M; Wang, Junshi; Huang, Xiaojie; Schlüter, Oliver M; Maze, Ian; Peña, Catherine J; Heller, Elizabeth A; Issler, Orna; Wang, Minghui; Song, Won-Min; Stein, Jason L; Liu, Xiaochuan; Doyle, Marie A; Scobie, Kimberly N; Sun, Hao Sheng; Neve, Rachael L; Geschwind, Daniel; Dong, Yan; Shen, Li; Zhang, Bin; Nestler, Eric J

    2016-06-01

    Depression is a complex, heterogeneous disorder and a leading contributor to the global burden of disease. Most previous research has focused on individual brain regions and genes contributing to depression. However, emerging evidence in humans and animal models suggests that dysregulated circuit function and gene expression across multiple brain regions drive depressive phenotypes. Here, we performed RNA sequencing on four brain regions from control animals and those susceptible or resilient to chronic social defeat stress at multiple time points. We employed an integrative network biology approach to identify transcriptional networks and key driver genes that regulate susceptibility to depressive-like symptoms. Further, we validated in vivo several key drivers and their associated transcriptional networks that regulate depression susceptibility and confirmed their functional significance at the levels of gene transcription, synaptic regulation, and behavior. Our study reveals novel transcriptional networks that control stress susceptibility and offers fundamentally new leads for antidepressant drug discovery. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Transcriptional Repressor HIC1 Contributes to Suppressive Function of Human Induced Regulatory T Cells

    Directory of Open Access Journals (Sweden)

    Ubaid Ullah

    2018-02-01

    Full Text Available Regulatory T (Treg cells are critical in regulating the immune response. In vitro induced Treg (iTreg cells have significant potential in clinical medicine. However, applying iTreg cells as therapeutics is complicated by the poor stability of human iTreg cells and their variable suppressive activity. Therefore, it is important to understand the molecular mechanisms of human iTreg cell specification. We identified hypermethylated in cancer 1 (HIC1 as a transcription factor upregulated early during the differentiation of human iTreg cells. Although FOXP3 expression was unaffected, HIC1 deficiency led to a considerable loss of suppression by iTreg cells with a concomitant increase in the expression of effector T cell associated genes. SNPs linked to several immune-mediated disorders were enriched around HIC1 binding sites, and in vitro binding assays indicated that these SNPs may alter the binding of HIC1. Our results suggest that HIC1 is an important contributor to iTreg cell development and function.

  12. Muscle and neural isoforms of agrin increase utrophin expression in cultured myotubes via a transcriptional regulatory mechanism.

    Science.gov (United States)

    Gramolini, A O; Burton, E A; Tinsley, J M; Ferns, M J; Cartaud, A; Cartaud, J; Davies, K E; Lunde, J A; Jasmin, B J

    1998-01-09

    . Furthermore, this increase in transcriptional activity in response to agrin resulted from a greater number of myonuclei expressing the 1.3-kilobase pair utrophin promoter-nlsLacZ construct. Deletion of 800 base pairs 5' from this fragment decreased the basal levels of nlsLacZ expression and abolished the sensitivity of the utrophin promoter to exogenously applied agrin. In addition, site-directed mutagenesis of an N-box motif contained within this 800-base pair fragment demonstrated its essential contribution in this regulatory mechanism. Finally, direct gene transfer studies performed in vivo further revealed the importance of this DNA element for the synapse-specific expression of the utrophin gene along multinucleated muscle fibers. These data show that both muscle and neural isoforms of agrin can regulate expression of the utrophin gene and further indicate that agrin is not only involved in the mechanisms leading to the formation of clusters containing presynthesized synaptic molecules but that it can also participate in the local regulation of genes encoding synaptic proteins. Together, these observations are therefore relevant for our basic understanding of the events involved in the assembly and maintenance of the postsynaptic membrane domain of the neuromuscular junction and for the potential use of utrophin as a therapeutic strategy to counteract the effects of Duchenne muscular dystrophy.

  13. Massively parallel amplicon sequencing reveals isotype-specific variability of antimicrobial peptide transcripts in Mytilus galloprovincialis.

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    Umberto Rosani

    Full Text Available BACKGROUND: Effective innate responses against potential pathogens are essential in the living world and possibly contributed to the evolutionary success of invertebrates. Taken together, antimicrobial peptide (AMP precursors of defensin, mytilin, myticin and mytimycin can represent about 40% of the hemocyte transcriptome in mussels injected with viral-like and bacterial preparations, and unique profiles of myticin C variants are expressed in single mussels. Based on amplicon pyrosequencing, we have ascertained and compared the natural and Vibrio-induced diversity of AMP transcripts in mussel hemocytes from three European regions. METHODOLOGY/PRINCIPAL FINDINGS: Hemolymph was collected from mussels farmed in the coastal regions of Palavas (France, Vigo (Spain and Venice (Italy. To represent the AMP families known in M. galloprovincialis, nine transcript sequences have been selected, amplified from hemocyte RNA and subjected to pyrosequencing. Hemolymph from farmed (offshore and wild (lagoon Venice mussels, both injected with 10(7 Vibrio cells, were similarly processed. Amplicon pyrosequencing emphasized the AMP transcript diversity, with Single Nucleotide Changes (SNC minimal for mytilin B/C and maximal for arthropod-like defensin and myticin C. Ratio of non-synonymous vs. synonymous changes also greatly differed between AMP isotypes. Overall, each amplicon revealed similar levels of nucleotidic variation across geographical regions, with two main sequence patterns confirmed for mytimycin and no substantial changes after immunostimulation. CONCLUSIONS/SIGNIFICANCE: Barcoding and bidirectional pyrosequencing allowed us to map and compare the transcript diversity of known mussel AMPs. Though most of the genuine cds variation was common to the analyzed samples we could estimate from 9 to 106 peptide variants in hemolymph pools representing 100 mussels, depending on the AMP isoform and sampling site. In this study, no prevailing SNC patterns related

  14. A ChIP-chip approach reveals a novel role for transcription factor IRF1 in the DNA damage response.

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    Frontini, Mattia; Vijayakumar, Meeraa; Garvin, Alexander; Clarke, Nicole

    2009-03-01

    IRF1 is a transcription factor that regulates key processes in the immune system and in tumour suppression. To gain further insight into IRF1's role in these processes, we searched for new target genes by performing chromatin immunoprecipitation coupled to a CpG island microarray (ChIP-chip). Using this approach we identified 202 new IRF1-binding sites with high confidence. Functional categorization of the target genes revealed a surprising cadre of new roles that can be linked to IRF1. One of the major functional categories was the DNA damage response pathway. In order to further validate our findings, we show that IRF1 can regulate the mRNA expression of a number of the DNA damage response genes in our list. In particular, we demonstrate that the mRNA and protein levels of the DNA repair protein BRIP1 [Fanconi anemia gene J (FANC J)] are upregulated after IRF1 over-expression. We also demonstrate that knockdown of IRF1 by siRNA results in loss of BRIP1 expression, abrogation of BRIP1 foci after DNA interstrand crosslink (ICL) damage and hypersensitivity to the DNA crosslinking agent, melphalan; a characteristic phenotype of FANC J cells. Taken together, our data provides a more complete understanding of the regulatory networks controlled by IRF1 and reveals a novel role for IRF1 in regulating the ICL DNA damage response.

  15. A ChIP–chip approach reveals a novel role for transcription factor IRF1 in the DNA damage response

    Science.gov (United States)

    Frontini, Mattia; Vijayakumar, Meeraa; Garvin, Alexander; Clarke, Nicole

    2009-01-01

    IRF1 is a transcription factor that regulates key processes in the immune system and in tumour suppression. To gain further insight into IRF1's role in these processes, we searched for new target genes by performing chromatin immunoprecipitation coupled to a CpG island microarray (ChIP–chip). Using this approach we identified 202 new IRF1-binding sites with high confidence. Functional categorization of the target genes revealed a surprising cadre of new roles that can be linked to IRF1. One of the major functional categories was the DNA damage response pathway. In order to further validate our findings, we show that IRF1 can regulate the mRNA expression of a number of the DNA damage response genes in our list. In particular, we demonstrate that the mRNA and protein levels of the DNA repair protein BRIP1 [Fanconi anemia gene J (FANC J)] are upregulated after IRF1 over-expression. We also demonstrate that knockdown of IRF1 by siRNA results in loss of BRIP1 expression, abrogation of BRIP1 foci after DNA interstrand crosslink (ICL) damage and hypersensitivity to the DNA crosslinking agent, melphalan; a characteristic phenotype of FANC J cells. Taken together, our data provides a more complete understanding of the regulatory networks controlled by IRF1 and reveals a novel role for IRF1 in regulating the ICL DNA damage response. PMID:19129219

  16. The transcriptional and gene regulatory network of Lactococcus lactis MG1363 during growth in milk.

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    Anne de Jong

    Full Text Available In the present study we examine the changes in the expression of genes of Lactococcus lactis subspecies cremoris MG1363 during growth in milk. To reveal which specific classes of genes (pathways, operons, regulons, COGs are important, we performed a transcriptome time series experiment. Global analysis of gene expression over time showed that L. lactis adapted quickly to the environmental changes. Using upstream sequences of genes with correlated gene expression profiles, we uncovered a substantial number of putative DNA binding motifs that may be relevant for L. lactis fermentative growth in milk. All available novel and literature-derived data were integrated into network reconstruction building blocks, which were used to reconstruct and visualize the L. lactis gene regulatory network. This network enables easy mining in the chrono-transcriptomics data. A freely available website at http://milkts.molgenrug.nl gives full access to all transcriptome data, to the reconstructed network and to the individual network building blocks.

  17. Dissecting the expression relationships between RNA-binding proteins and their cognate targets in eukaryotic post-transcriptional regulatory networks

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    Nishtala, Sneha; Neelamraju, Yaseswini; Janga, Sarath Chandra

    2016-05-01

    RNA-binding proteins (RBPs) are pivotal in orchestrating several steps in the metabolism of RNA in eukaryotes thereby controlling an extensive network of RBP-RNA interactions. Here, we employed CLIP (cross-linking immunoprecipitation)-seq datasets for 60 human RBPs and RIP-ChIP (RNP immunoprecipitation-microarray) data for 69 yeast RBPs to construct a network of genome-wide RBP- target RNA interactions for each RBP. We show in humans that majority (~78%) of the RBPs are strongly associated with their target transcripts at transcript level while ~95% of the studied RBPs were also found to be strongly associated with expression levels of target transcripts when protein expression levels of RBPs were employed. At transcript level, RBP - RNA interaction data for the yeast genome, exhibited a strong association for 63% of the RBPs, confirming the association to be conserved across large phylogenetic distances. Analysis to uncover the features contributing to these associations revealed the number of target transcripts and length of the selected protein-coding transcript of an RBP at the transcript level while intensity of the CLIP signal, number of RNA-Binding domains, location of the binding site on the transcript, to be significant at the protein level. Our analysis will contribute to improved modelling and prediction of post-transcriptional networks.

  18. Transcriptional and physiological data reveal the dehydration memory behavior in switchgrass (Panicum virgatum L.).

    Science.gov (United States)

    Zhang, Chao; Peng, Xi; Guo, Xiaofeng; Tang, Gaijuan; Sun, Fengli; Liu, Shudong; Xi, Yajun

    2018-01-01

    Switchgrass ( Panicum virgatum L.) is a model biofuel plant because of its high biomass, cellulose-richness, easy degradation to ethanol, and the availability of extensive genomic information. However, a little is currently known about the molecular responses of switchgrass plants to dehydration stress, especially multiple dehydration stresses. Studies on the transcriptional profiles of 35-day-old tissue culture plants revealed 741 dehydration memory genes. Gene Ontology and pathway analysis showed that these genes were enriched in phenylpropanoid biosynthesis, starch and sucrose metabolism, and plant hormone signal transduction. Further analysis of specific pathways combined with physiological data suggested that switchgrass improved its dehydration resistance by changing various aspects of its responses to secondary dehydration stress (D2), including the regulation of abscisic acid (ABA) and jasmonic acid (JA) biosynthesis and signal transduction, the biosynthesis of osmolytes (l-proline, stachyose and trehalose), energy metabolism (i.e., metabolic process relating to photosynthetic systems, glycolysis, and the TCA cycle), and lignin biosynthesis. The transcriptional data and chemical substance assays showed that ABA was significantly accumulated during both primary (D1) and secondary (D2) dehydration stresses, whereas JA accumulated during D1 but became significantly less abundant during D2. This suggests the existence of a complicated signaling network of plant hormones in response to repeated dehydration stresses. A homology analysis focusing on switchgrass, maize, and Arabidopsis revealed the conservation and species-specific distribution of dehydration memory genes. The molecular responses of switchgrass plants to successive dehydration stresses have been systematically characterized, revealing a previously unknown transcriptional memory behavior. These results provide new insights into the mechanisms of dehydration stress responses in plants. The genes and

  19. Sialotranscriptomics of Rhipicephalus zambeziensis reveals intricate expression profiles of secretory proteins and suggests tight temporal transcriptional regulation during blood-feeding.

    Science.gov (United States)

    de Castro, Minique Hilda; de Klerk, Daniel; Pienaar, Ronel; Rees, D Jasper G; Mans, Ben J

    2017-08-10

    Ticks secrete a diverse mixture of secretory proteins into the host to evade its immune response and facilitate blood-feeding, making secretory proteins attractive targets for the production of recombinant anti-tick vaccines. The largely neglected tick species, Rhipicephalus zambeziensis, is an efficient vector of Theileria parva in southern Africa but its available sequence information is limited. Next generation sequencing has advanced sequence availability for ticks in recent years and has assisted the characterisation of secretory proteins. This study focused on the de novo assembly and annotation of the salivary gland transcriptome of R. zambeziensis and the temporal expression of secretory protein transcripts in female and male ticks, before the onset of feeding and during early and late feeding. The sialotranscriptome of R. zambeziensis yielded 23,631 transcripts from which 13,584 non-redundant proteins were predicted. Eighty-six percent of these contained a predicted start and stop codon and were estimated to be putatively full-length proteins. A fifth (2569) of the predicted proteins were annotated as putative secretory proteins and explained 52% of the expression in the transcriptome. Expression analyses revealed that 2832 transcripts were differentially expressed among feeding time points and 1209 between the tick sexes. The expression analyses further indicated that 57% of the annotated secretory protein transcripts were differentially expressed. Dynamic expression profiles of secretory protein transcripts were observed during feeding of female ticks. Whereby a number of transcripts were upregulated during early feeding, presumably for feeding site establishment and then during late feeding, 52% of these were downregulated, indicating that transcripts were required at specific feeding stages. This suggested that secretory proteins are under stringent transcriptional regulation that fine-tunes their expression in salivary glands during feeding. No open

  20. Global Transcription Profiling Reveals Comprehensive Insights into Hypoxic Response in Arabidopsis1[w

    Science.gov (United States)

    Liu, Fenglong; VanToai, Tara; Moy, Linda P.; Bock, Geoffrey; Linford, Lara D.; Quackenbush, John

    2005-01-01

    Plants have evolved adaptation mechanisms to sense oxygen deficiency in their environments and make coordinated physiological and structural adjustments to enhance their hypoxic tolerance. To gain insight into how plants respond to low-oxygen stress, gene expression profiling using whole-genome DNA amplicon microarrays was carried out at seven time points over 24 h, in wild-type and transgenic PSAG12:ipt Arabidopsis (Arabidopsis thaliana) plants under normoxic and hypoxic conditions. Transcript levels of genes involved in glycolysis and fermentation pathways, ethylene synthesis and perception, calcium signaling, nitrogen utilization, trehalose metabolism, and alkaloid synthesis were significantly altered in response to oxygen limitation. Analysis based on gene ontology assignments suggested a significant down-regulation of genes whose functions are associated with cell walls, nucleosome structures, water channels, and ion transporters and a significant up-regulation of genes involved in transcriptional regulation, protein kinase activity, and auxin responses under conditions of oxygen shortage. Promoter analysis on a cluster of up-regulated genes revealed a significant overrepresentation of the AtMYB2-binding motif (GT motif), a sugar response element-like motif, and a G-box-related sequence, and also identified several putative anaerobic response elements. Finally, quantitative real-time polymerase chain reactions using 29 selected genes independently verified the microarray results. This study represents one of the most comprehensive analyses conducted to date investigating hypoxia-responsive transcriptional networks in plants. PMID:15734912

  1. Global transcription profiling reveals comprehensive insights into hypoxic response in Arabidopsis.

    Science.gov (United States)

    Liu, Fenglong; Vantoai, Tara; Moy, Linda P; Bock, Geoffrey; Linford, Lara D; Quackenbush, John

    2005-03-01

    Plants have evolved adaptation mechanisms to sense oxygen deficiency in their environments and make coordinated physiological and structural adjustments to enhance their hypoxic tolerance. To gain insight into how plants respond to low-oxygen stress, gene expression profiling using whole-genome DNA amplicon microarrays was carried out at seven time points over 24 h, in wild-type and transgenic P(SAG12):ipt Arabidopsis (Arabidopsis thaliana) plants under normoxic and hypoxic conditions. Transcript levels of genes involved in glycolysis and fermentation pathways, ethylene synthesis and perception, calcium signaling, nitrogen utilization, trehalose metabolism, and alkaloid synthesis were significantly altered in response to oxygen limitation. Analysis based on gene ontology assignments suggested a significant down-regulation of genes whose functions are associated with cell walls, nucleosome structures, water channels, and ion transporters and a significant up-regulation of genes involved in transcriptional regulation, protein kinase activity, and auxin responses under conditions of oxygen shortage. Promoter analysis on a cluster of up-regulated genes revealed a significant overrepresentation of the AtMYB2-binding motif (GT motif), a sugar response element-like motif, and a G-box-related sequence, and also identified several putative anaerobic response elements. Finally, quantitative real-time polymerase chain reactions using 29 selected genes independently verified the microarray results. This study represents one of the most comprehensive analyses conducted to date investigating hypoxia-responsive transcriptional networks in plants.

  2. Genome-wide Reconstruction of OxyR and SoxRS Transcriptional Regulatory Networks under Oxidative Stress in Escherichia coli K-12 MG1655

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    Sang Woo Seo

    2015-08-01

    Full Text Available Three transcription factors (TFs, OxyR, SoxR, and SoxS, play a critical role in transcriptional regulation of the defense system for oxidative stress in bacteria. However, their full genome-wide regulatory potential is unknown. Here, we perform a genome-scale reconstruction of the OxyR, SoxR, and SoxS regulons in Escherichia coli K-12 MG1655. Integrative data analysis reveals that a total of 68 genes in 51 transcription units (TUs belong to these regulons. Among them, 48 genes showed more than 2-fold changes in expression level under single-TF-knockout conditions. This reconstruction expands the genome-wide roles of these factors to include direct activation of genes related to amino acid biosynthesis (methionine and aromatic amino acids, cell wall synthesis (lipid A biosynthesis and peptidoglycan growth, and divalent metal ion transport (Mn2+, Zn2+, and Mg2+. Investigating the co-regulation of these genes with other stress-response TFs reveals that they are independently regulated by stress-specific TFs.

  3. Comparative genetic screens in human cells reveal new regulatory mechanisms in WNT signaling

    Science.gov (United States)

    Lebensohn, Andres M; Dubey, Ramin; Neitzel, Leif R; Tacchelly-Benites, Ofelia; Yang, Eungi; Marceau, Caleb D; Davis, Eric M; Patel, Bhaven B; Bahrami-Nejad, Zahra; Travaglini, Kyle J; Ahmed, Yashi; Lee, Ethan; Carette, Jan E; Rohatgi, Rajat

    2016-01-01

    The comprehensive understanding of cellular signaling pathways remains a challenge due to multiple layers of regulation that may become evident only when the pathway is probed at different levels or critical nodes are eliminated. To discover regulatory mechanisms in canonical WNT signaling, we conducted a systematic forward genetic analysis through reporter-based screens in haploid human cells. Comparison of screens for negative, attenuating and positive regulators of WNT signaling, mediators of R-spondin-dependent signaling and suppressors of constitutive signaling induced by loss of the tumor suppressor adenomatous polyposis coli or casein kinase 1α uncovered new regulatory features at most levels of the pathway. These include a requirement for the transcription factor AP-4, a role for the DAX domain of AXIN2 in controlling β-catenin transcriptional activity, a contribution of glycophosphatidylinositol anchor biosynthesis and glypicans to R-spondin-potentiated WNT signaling, and two different mechanisms that regulate signaling when distinct components of the β-catenin destruction complex are lost. The conceptual and methodological framework we describe should enable the comprehensive understanding of other signaling systems. DOI: http://dx.doi.org/10.7554/eLife.21459.001 PMID:27996937

  4. Analysis of Transcriptional Signatures in Response to Listeria monocytogenes Infection Reveals Temporal Changes That Result from Type I Interferon Signaling

    Science.gov (United States)

    Potempa, Krzysztof; Graham, Christine M.; Moreira-Teixeira, Lucia; McNab, Finlay W.; Howes, Ashleigh; Stavropoulos, Evangelos; Pascual, Virginia; Banchereau, Jacques; Chaussabel, Damien; O’Garra, Anne

    2016-01-01

    Analysis of the mouse transcriptional response to Listeria monocytogenes infection reveals that a large set of genes are perturbed in both blood and tissue and that these transcriptional responses are enriched for pathways of the immune response. Further we identified enrichment for both type I and type II interferon (IFN) signaling molecules in the blood and tissues upon infection. Since type I IFN signaling has been reported widely to impair bacterial clearance we examined gene expression from blood and tissues of wild type (WT) and type I IFNαβ receptor-deficient (Ifnar1-/-) mice at the basal level and upon infection with L. monocytogenes. Measurement of the fold change response upon infection in the absence of type I IFN signaling demonstrated an upregulation of specific genes at day 1 post infection. A less marked reduction of the global gene expression signature in blood or tissues from infected Ifnar1-/- as compared to WT mice was observed at days 2 and 3 after infection, with marked reduction in key genes such as Oasg1 and Stat2. Moreover, on in depth analysis, changes in gene expression in uninfected mice of key IFN regulatory genes including Irf9, Irf7, Stat1 and others were identified, and although induced by an equivalent degree upon infection this resulted in significantly lower final gene expression levels upon infection of Ifnar1-/- mice. These data highlight how dysregulation of this network in the steady state and temporally upon infection may determine the outcome of this bacterial infection and how basal levels of type I IFN-inducible genes may perturb an optimal host immune response to control intracellular bacterial infections such as L. monocytogenes. PMID:26918359

  5. Genome-wide analysis of the Dof transcription factor gene family reveals soybean-specific duplicable and functional characteristics.

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    Yong Guo

    Full Text Available The Dof domain protein family is a classic plant-specific zinc-finger transcription factor family involved in a variety of biological processes. There is great diversity in the number of Dof genes in different plants. However, there are only very limited reports on the characterization of Dof transcription factors in soybean (Glycine max. In the present study, 78 putative Dof genes were identified from the whole-genome sequence of soybean. The predicted GmDof genes were non-randomly distributed within and across 19 out of 20 chromosomes and 97.4% (38 pairs were preferentially retained duplicate paralogous genes located in duplicated regions of the genome. Soybean-specific segmental duplications contributed significantly to the expansion of the soybean Dof gene family. These Dof proteins were phylogenetically clustered into nine distinct subgroups among which the gene structure and motif compositions were considerably conserved. Comparative phylogenetic analysis of these Dof proteins revealed four major groups, similar to those reported for Arabidopsis and rice. Most of the GmDofs showed specific expression patterns based on RNA-seq data analyses. The expression patterns of some duplicate genes were partially redundant while others showed functional diversity, suggesting the occurrence of sub-functionalization during subsequent evolution. Comprehensive expression profile analysis also provided insights into the soybean-specific functional divergence among members of the Dof gene family. Cis-regulatory element analysis of these GmDof genes suggested diverse functions associated with different processes. Taken together, our results provide useful information for the functional characterization of soybean Dof genes by combining phylogenetic analysis with global gene-expression profiling.

  6. Transcriptome analysis reveals regulatory networks underlying differential susceptibility to Botrytis cinerea in response to nitrogen availability in Solanum lycopersicum.

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    Andrea eVega

    2015-11-01

    Full Text Available Nitrogen (N is one of the main limiting nutrients for plant growth and crop yield. It is well documented that changes in nitrate availability, the main N source found in agricultural soils, influences a myriad of developmental programs and processes including the plant defense response. Indeed, many agronomical reports indicate that the plant N nutritional status influences their ability to respond effectively when challenged by different pathogens. However, the molecular mechanisms involved in N-modulation of plant susceptibility to pathogens are poorly characterized. In this work, we show that Solanum lycopersicum defense response to the necrotrophic fungus Botrytis cinerea is affected by plant N availability, with higher susceptibility in nitrate-limiting conditions. Global gene expression responses of tomato against B. cinerea under contrasting nitrate conditions reveals that plant primary metabolism is affected by the fungal infection regardless of N regimes. This result suggests that differential susceptibility to pathogen attack under contrasting N conditions is not only explained by a metabolic alteration. We used a systems biology approach to identify the transcriptional regulatory network implicated in plant response to the fungus infection under contrasting nitrate conditions. Interestingly, hub genes in this network are known key transcription factors involved in ethylene and jasmonic acid signaling. This result positions these hormones as key integrators of nitrate and defense against B. cinerea in tomato plants. Our results provide insights into potential crosstalk mechanisms between necrotrophic defense response and N status in plants.

  7. Hydrogen-Deuterium Exchange Mass Spectrometry Reveals Calcium Binding Properties and Allosteric Regulation of Downstream Regulatory Element Antagonist Modulator (DREAM).

    Science.gov (United States)

    Zhang, Jun; Li, Jing; Craig, Theodore A; Kumar, Rajiv; Gross, Michael L

    2017-07-18

    Downstream regulatory element antagonist modulator (DREAM) is an EF-hand Ca 2+ -binding protein that also binds to a specific DNA sequence, downstream regulatory elements (DRE), and thereby regulates transcription in a calcium-dependent fashion. DREAM binds to DRE in the absence of Ca 2+ but detaches from DRE under Ca 2+ stimulation, allowing gene expression. The Ca 2+ binding properties of DREAM and the consequences of the binding on protein structure are key to understanding the function of DREAM. Here we describe the application of hydrogen-deuterium exchange mass spectrometry (HDX-MS) and site-directed mutagenesis to investigate the Ca 2+ binding properties and the subsequent conformational changes of full-length DREAM. We demonstrate that all EF-hands undergo large conformation changes upon calcium binding even though the EF-1 hand is not capable of binding to Ca 2+ . Moreover, EF-2 is a lower-affinity site compared to EF-3 and -4 hands. Comparison of HDX profiles between wild-type DREAM and two EF-1 mutated constructs illustrates that the conformational changes in the EF-1 hand are induced by long-range structural interactions. HDX analyses also reveal a conformational change in an N-terminal leucine-charged residue-rich domain (LCD) remote from Ca 2+ -binding EF-hands. This LCD domain is responsible for the direct interaction between DREAM and cAMP response element-binding protein (CREB) and regulates the recruitment of the co-activator, CREB-binding protein. These long-range interactions strongly suggest how conformational changes transmit the Ca 2+ signal to CREB-mediated gene transcription.

  8. Concurrent growth rate and transcript analyses reveal essential gene stringency in Escherichia coli.

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    Shan Goh

    Full Text Available BACKGROUND: Genes essential for bacterial growth are of particular scientific interest. Many putative essential genes have been identified or predicted in several species, however, little is known about gene expression requirement stringency, which may be an important aspect of bacterial physiology and likely a determining factor in drug target development. METHODOLOGY/PRINCIPAL FINDINGS: Working from the premise that essential genes differ in absolute requirement for growth, we describe silencing of putative essential genes in E. coli to obtain a titration of declining growth rates and transcript levels by using antisense peptide nucleic acids (PNA and expressed antisense RNA. The relationship between mRNA decline and growth rate decline reflects the degree of essentiality, or stringency, of an essential gene, which is here defined by the minimum transcript level for a 50% reduction in growth rate (MTL(50. When applied to four growth essential genes, both RNA silencing methods resulted in MTL(50 values that reveal acpP as the most stringently required of the four genes examined, with ftsZ the next most stringently required. The established antibacterial targets murA and fabI were less stringently required. CONCLUSIONS: RNA silencing can reveal stringent requirements for gene expression with respect to growth. This method may be used to validate existing essential genes and to quantify drug target requirement.

  9. Co-expression networks reveal the tissue-specific regulation of transcription and splicing.

    Science.gov (United States)

    Saha, Ashis; Kim, Yungil; Gewirtz, Ariel D H; Jo, Brian; Gao, Chuan; McDowell, Ian C; Engelhardt, Barbara E; Battle, Alexis

    2017-11-01

    Gene co-expression networks capture biologically important patterns in gene expression data, enabling functional analyses of genes, discovery of biomarkers, and interpretation of genetic variants. Most network analyses to date have been limited to assessing correlation between total gene expression levels in a single tissue or small sets of tissues. Here, we built networks that additionally capture the regulation of relative isoform abundance and splicing, along with tissue-specific connections unique to each of a diverse set of tissues. We used the Genotype-Tissue Expression (GTEx) project v6 RNA sequencing data across 50 tissues and 449 individuals. First, we developed a framework called Transcriptome-Wide Networks (TWNs) for combining total expression and relative isoform levels into a single sparse network, capturing the interplay between the regulation of splicing and transcription. We built TWNs for 16 tissues and found that hubs in these networks were strongly enriched for splicing and RNA binding genes, demonstrating their utility in unraveling regulation of splicing in the human transcriptome. Next, we used a Bayesian biclustering model that identifies network edges unique to a single tissue to reconstruct Tissue-Specific Networks (TSNs) for 26 distinct tissues and 10 groups of related tissues. Finally, we found genetic variants associated with pairs of adjacent nodes in our networks, supporting the estimated network structures and identifying 20 genetic variants with distant regulatory impact on transcription and splicing. Our networks provide an improved understanding of the complex relationships of the human transcriptome across tissues. © 2017 Saha et al.; Published by Cold Spring Harbor Laboratory Press.

  10. Comprehensive prediction in 78 human cell lines reveals rigidity and compactness of transcription factor dimers

    Science.gov (United States)

    Jankowski, Aleksander; Szczurek, Ewa; Jauch, Ralf; Tiuryn, Jerzy; Prabhakar, Shyam

    2013-01-01

    The binding of transcription factors (TFs) to their specific motifs in genomic regulatory regions is commonly studied in isolation. However, in order to elucidate the mechanisms of transcriptional regulation, it is essential to determine which TFs bind DNA cooperatively as dimers and to infer the precise nature of these interactions. So far, only a small number of such dimeric complexes are known. Here, we present an algorithm for predicting cell-type–specific TF–TF dimerization on DNA on a large scale, using DNase I hypersensitivity data from 78 human cell lines. We represented the universe of possible TF complexes by their corresponding motif complexes, and analyzed their occurrence at cell-type–specific DNase I hypersensitive sites. Based on ∼1.4 billion tests for motif complex enrichment, we predicted 603 highly significant cell-type–specific TF dimers, the vast majority of which are novel. Our predictions included 76% (19/25) of the known dimeric complexes and showed significant overlap with an experimental database of protein–protein interactions. They were also independently supported by evolutionary conservation, as well as quantitative variation in DNase I digestion patterns. Notably, the known and predicted TF dimers were almost always highly compact and rigidly spaced, suggesting that TFs dimerize in close proximity to their partners, which results in strict constraints on the structure of the DNA-bound complex. Overall, our results indicate that chromatin openness profiles are highly predictive of cell-type–specific TF–TF interactions. Moreover, cooperative TF dimerization seems to be a widespread phenomenon, with multiple TF complexes predicted in most cell types. PMID:23554463

  11. Network modeling reveals prevalent negative regulatory relationships between signaling sectors in Arabidopsis immune signaling.

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    Masanao Sato

    Full Text Available Biological signaling processes may be mediated by complex networks in which network components and network sectors interact with each other in complex ways. Studies of complex networks benefit from approaches in which the roles of individual components are considered in the context of the network. The plant immune signaling network, which controls inducible responses to pathogen attack, is such a complex network. We studied the Arabidopsis immune signaling network upon challenge with a strain of the bacterial pathogen Pseudomonas syringae expressing the effector protein AvrRpt2 (Pto DC3000 AvrRpt2. This bacterial strain feeds multiple inputs into the signaling network, allowing many parts of the network to be activated at once. mRNA profiles for 571 immune response genes of 22 Arabidopsis immunity mutants and wild type were collected 6 hours after inoculation with Pto DC3000 AvrRpt2. The mRNA profiles were analyzed as detailed descriptions of changes in the network state resulting from the genetic perturbations. Regulatory relationships among the genes corresponding to the mutations were inferred by recursively applying a non-linear dimensionality reduction procedure to the mRNA profile data. The resulting static network model accurately predicted 23 of 25 regulatory relationships reported in the literature, suggesting that predictions of novel regulatory relationships are also accurate. The network model revealed two striking features: (i the components of the network are highly interconnected; and (ii negative regulatory relationships are common between signaling sectors. Complex regulatory relationships, including a novel negative regulatory relationship between the early microbe-associated molecular pattern-triggered signaling sectors and the salicylic acid sector, were further validated. We propose that prevalent negative regulatory relationships among the signaling sectors make the plant immune signaling network a "sector

  12. Genome-wide comparative analysis reveals human-mouse regulatory landscape and evolution.

    Science.gov (United States)

    Denas, Olgert; Sandstrom, Richard; Cheng, Yong; Beal, Kathryn; Herrero, Javier; Hardison, Ross C; Taylor, James

    2015-02-14

    Because species-specific gene expression is driven by species-specific regulation, understanding the relationship between sequence and function of the regulatory regions in different species will help elucidate how differences among species arise. Despite active experimental and computational research, relationships among sequence, conservation, and function are still poorly understood. We compared transcription factor occupied segments (TFos) for 116 human and 35 mouse TFs in 546 human and 125 mouse cell types and tissues from the Human and the Mouse ENCODE projects. We based the map between human and mouse TFos on a one-to-one nucleotide cross-species mapper, bnMapper, that utilizes whole genome alignments (WGA). Our analysis shows that TFos are under evolutionary constraint, but a substantial portion (25.1% of mouse and 25.85% of human on average) of the TFos does not have a homologous sequence on the other species; this portion varies among cell types and TFs. Furthermore, 47.67% and 57.01% of the homologous TFos sequence shows binding activity on the other species for human and mouse respectively. However, 79.87% and 69.22% is repurposed such that it binds the same TF in different cells or different TFs in the same cells. Remarkably, within the set of repurposed TFos, the corresponding genome regions in the other species are preferred locations of novel TFos. These events suggest exaptation of some functional regulatory sequences into new function. Despite TFos repurposing, we did not find substantial changes in their predicted target genes, suggesting that CRMs buffer evolutionary events allowing little or no change in the TFos - target gene associations. Thus, the small portion of TFos with strictly conserved occupancy underestimates the degree of conservation of regulatory interactions. We mapped regulatory sequences from an extensive number of TFs and cell types between human and mouse using WGA. A comparative analysis of this correspondence unveiled the

  13. An Org-1-Tup transcriptional cascade reveals different types of alary muscles connecting internal organs in Drosophila.

    Science.gov (United States)

    Boukhatmi, Hadi; Schaub, Christoph; Bataillé, Laetitia; Reim, Ingolf; Frendo, Jean-Louis; Frasch, Manfred; Vincent, Alain

    2014-10-01

    The T-box transcription factor Tbx1 and the LIM-homeodomain transcription factor Islet1 are key components in regulatory circuits that generate myogenic and cardiogenic lineage diversity in chordates. We show here that Org-1 and Tup, the Drosophila orthologs of Tbx1 and Islet1, are co-expressed and required for formation of the heart-associated alary muscles (AMs) in the abdomen. The same holds true for lineage-related muscles in the thorax that have not been described previously, which we name thoracic alary-related muscles (TARMs). Lineage analyses identified the progenitor cell for each AM and TARM. Three-dimensional high-resolution analyses indicate that AMs and TARMs connect the exoskeleton to the aorta/heart and to different regions of the midgut, respectively, and surround-specific tracheal branches, pointing to an architectural role in the internal anatomy of the larva. Org-1 controls tup expression in the AM/TARM lineage by direct binding to two regulatory sites within an AM/TARM-specific cis-regulatory module, tupAME. The contributions of Org-1 and Tup to the specification of Drosophila AMs and TARMs provide new insights into the transcriptional control of Drosophila larval muscle diversification and highlight new parallels with gene regulatory networks involved in the specification of cardiopharyngeal mesodermal derivatives in chordates. © 2014. Published by The Company of Biologists Ltd.

  14. Characterization of a putative cis-regulatory element that controls transcriptional activity of the pig uroplakin II gene promoter

    International Nuclear Information System (INIS)

    Kwon, Deug-Nam; Park, Mi-Ryung; Park, Jong-Yi; Cho, Ssang-Goo; Park, Chankyu; Oh, Jae-Wook; Song, Hyuk; Kim, Jae-Hwan; Kim, Jin-Hoi

    2011-01-01

    Highlights: → The sequences of -604 to -84 bp of the pUPII promoter contained the region of a putative negative cis-regulatory element. → The core promoter was located in the 5F-1. → Transcription factor HNF4 can directly bind in the pUPII core promoter region, which plays a critical role in controlling promoter activity. → These features of the pUPII promoter are fundamental to development of a target-specific vector. -- Abstract: Uroplakin II (UPII) is a one of the integral membrane proteins synthesized as a major differentiation product of mammalian urothelium. UPII gene expression is bladder specific and differentiation dependent, but little is known about its transcription response elements and molecular mechanism. To identify the cis-regulatory elements in the pig UPII (pUPII) gene promoter region, we constructed pUPII 5' upstream region deletion mutants and demonstrated that each of the deletion mutants participates in controlling the expression of the pUPII gene in human bladder carcinoma RT4 cells. We also identified a new core promoter region and putative negative cis-regulatory element within a minimal promoter region. In addition, we showed that hepatocyte nuclear factor 4 (HNF4) can directly bind in the pUPII core promoter (5F-1) region, which plays a critical role in controlling promoter activity. Transient cotransfection experiments showed that HNF4 positively regulates pUPII gene promoter activity. Thus, the binding element and its binding protein, HNF4 transcription factor, may be involved in the mechanism that specifically regulates pUPII gene transcription.

  15. Transcriptional regulatory networks downstream of TAL1/SCL in T-cell acute lymphoblastic leukemia

    OpenAIRE

    Palomero, Teresa; Odom, Duncan T.; O'Neil, Jennifer; Ferrando, Adolfo A.; Margolin, Adam; Neuberg, Donna S.; Winter, Stuart S.; Larson, Richard S.; Li, Wei; Liu, X. Shirley; Young, Richard A.; Look, A. Thomas

    2006-01-01

    Aberrant expression of 1 or more transcription factor oncogenes is a critical component of the molecular pathogenesis of human T-cell acute lymphoblastic leukemia (T-ALL); however, oncogenic transcriptional programs downstream of T-ALL oncogenes are mostly unknown. TAL1/SCL is a basic helix-loop-helix (bHLH) transcription factor oncogene aberrantly expressed in 60% of human T-ALLs. We used chromatin immunoprecipitation (ChIP) on chip to identify 71 direct transcriptional targets of TAL1/SCL. ...

  16. Transcriptional Analysis and Subcellular Protein Localization Reveal Specific Features of the Essential WalKR System in Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Olivier Poupel

    Full Text Available The WalKR two-component system, controlling cell wall metabolism, is highly conserved among Bacilli and essential for cell viability. In Staphylococcus aureus, walR and walK are followed by three genes of unknown function: walH, walI and walJ. Sequence analysis and transcript mapping revealed a unique genetic structure for this locus in S. aureus: the last gene of the locus, walJ, is transcribed independently, whereas transcription of the tetra-cistronic walRKHI operon occurred from two independent promoters located upstream from walR. Protein topology analysis and protein-protein interactions in E. coli as well as subcellular localization in S. aureus allowed us to show that WalH and WalI are membrane-bound proteins, which associate with WalK to form a complex at the cell division septum. While these interactions suggest that WalH and WalI play a role in activity of the WalKR regulatory pathway, deletion of walH and/or walI did not have a major effect on genes whose expression is strongly dependent on WalKR or on associated phenotypes. No effect of WalH or WalI was seen on tightly controlled WalKR regulon genes such as sle1 or saouhsc_00773, which encodes a CHAP-domain amidase. Of the genes encoding the two major S. aureus autolysins, AtlA and Sle1, only transcription of atlA was increased in the ΔwalH or ΔwalI mutants. Likewise, bacterial autolysis was not increased in the absence of WalH and/or WalI and biofilm formation was lowered rather than increased. Our results suggest that contrary to their major role as WalK inhibitors in B. subtilis, the WalH and WalI proteins have evolved a different function in S. aureus, where they are more accessory. A phylogenomic analysis shows a striking conservation of the 5 gene wal cluster along the evolutionary history of Bacilli, supporting the key importance of this signal transduction system, and indicating that the walH and walI genes were lost in the ancestor of Streptococcaceae, leading to their

  17. Transcriptional Analysis and Subcellular Protein Localization Reveal Specific Features of the Essential WalKR System in Staphylococcus aureus.

    Science.gov (United States)

    Poupel, Olivier; Moyat, Mati; Groizeleau, Julie; Antunes, Luísa C S; Gribaldo, Simonetta; Msadek, Tarek; Dubrac, Sarah

    2016-01-01

    The WalKR two-component system, controlling cell wall metabolism, is highly conserved among Bacilli and essential for cell viability. In Staphylococcus aureus, walR and walK are followed by three genes of unknown function: walH, walI and walJ. Sequence analysis and transcript mapping revealed a unique genetic structure for this locus in S. aureus: the last gene of the locus, walJ, is transcribed independently, whereas transcription of the tetra-cistronic walRKHI operon occurred from two independent promoters located upstream from walR. Protein topology analysis and protein-protein interactions in E. coli as well as subcellular localization in S. aureus allowed us to show that WalH and WalI are membrane-bound proteins, which associate with WalK to form a complex at the cell division septum. While these interactions suggest that WalH and WalI play a role in activity of the WalKR regulatory pathway, deletion of walH and/or walI did not have a major effect on genes whose expression is strongly dependent on WalKR or on associated phenotypes. No effect of WalH or WalI was seen on tightly controlled WalKR regulon genes such as sle1 or saouhsc_00773, which encodes a CHAP-domain amidase. Of the genes encoding the two major S. aureus autolysins, AtlA and Sle1, only transcription of atlA was increased in the ΔwalH or ΔwalI mutants. Likewise, bacterial autolysis was not increased in the absence of WalH and/or WalI and biofilm formation was lowered rather than increased. Our results suggest that contrary to their major role as WalK inhibitors in B. subtilis, the WalH and WalI proteins have evolved a different function in S. aureus, where they are more accessory. A phylogenomic analysis shows a striking conservation of the 5 gene wal cluster along the evolutionary history of Bacilli, supporting the key importance of this signal transduction system, and indicating that the walH and walI genes were lost in the ancestor of Streptococcaceae, leading to their atypical 3 wal gene

  18. YY1 binding association with sex-biased transcription revealed through X-linked transcript levels and allelic binding analyses.

    Science.gov (United States)

    Chen, Chih-Yu; Shi, Wenqiang; Balaton, Bradley P; Matthews, Allison M; Li, Yifeng; Arenillas, David J; Mathelier, Anthony; Itoh, Masayoshi; Kawaji, Hideya; Lassmann, Timo; Hayashizaki, Yoshihide; Carninci, Piero; Forrest, Alistair R R; Brown, Carolyn J; Wasserman, Wyeth W

    2016-11-18

    Sex differences in susceptibility and progression have been reported in numerous diseases. Female cells have two copies of the X chromosome with X-chromosome inactivation imparting mono-allelic gene silencing for dosage compensation. However, a subset of genes, named escapees, escape silencing and are transcribed bi-allelically resulting in sexual dimorphism. Here we conducted in silico analyses of the sexes using human datasets to gain perspectives into such regulation. We identified transcription start sites of escapees (escTSSs) based on higher transcription levels in female cells using FANTOM5 CAGE data. Significant over-representations of YY1 transcription factor binding motif and ChIP-seq peaks around escTSSs highlighted its positive association with escapees. Furthermore, YY1 occupancy is significantly biased towards the inactive X (Xi) at long non-coding RNA loci that are frequent contacts of Xi-specific superloops. Our study suggests a role for YY1 in transcriptional activity on Xi in general through sequence-specific binding, and its involvement at superloop anchors.

  19. Transcription Profiles Reveal Sugar and Hormone Signaling Pathways Mediating Flower Induction in Apple (Malus domestica Borkh.).

    Science.gov (United States)

    Xing, Li-Bo; Zhang, Dong; Li, You-Mei; Shen, Ya-Wen; Zhao, Cai-Ping; Ma, Juan-Juan; An, Na; Han, Ming-Yu

    2015-10-01

    Flower induction in apple (Malus domestica Borkh.) is regulated by complex gene networks that involve multiple signal pathways to ensure flower bud formation in the next year, but the molecular determinants of apple flower induction are still unknown. In this research, transcriptomic profiles from differentiating buds allowed us to identify genes potentially involved in signaling pathways that mediate the regulatory mechanisms of flower induction. A hypothetical model for this regulatory mechanism was obtained by analysis of the available transcriptomic data, suggesting that sugar-, hormone- and flowering-related genes, as well as those involved in cell-cycle induction, participated in the apple flower induction process. Sugar levels and metabolism-related gene expression profiles revealed that sucrose is the initiation signal in flower induction. Complex hormone regulatory networks involved in cytokinin (CK), abscisic acid (ABA) and gibberellic acid pathways also induce apple flower formation. CK plays a key role in the regulation of cell formation and differentiation, and in affecting flowering-related gene expression levels during these processes. Meanwhile, ABA levels and ABA-related gene expression levels gradually increased, as did those of sugar metabolism-related genes, in developing buds, indicating that ABA signals regulate apple flower induction by participating in the sugar-mediated flowering pathway. Furthermore, changes in sugar and starch deposition levels in buds can be affected by ABA content and the expression of the genes involved in the ABA signaling pathway. Thus, multiple pathways, which are mainly mediated by crosstalk between sugar and hormone signals, regulate the molecular network involved in bud growth and flower induction in apple trees. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

  20. Transcriptional profiling at whole population and single cell levels reveals somatosensory neuron molecular diversity

    Science.gov (United States)

    Chiu, Isaac M; Barrett, Lee B; Williams, Erika K; Strochlic, David E; Lee, Seungkyu; Weyer, Andy D; Lou, Shan; Bryman, Gregory S; Roberson, David P; Ghasemlou, Nader; Piccoli, Cara; Ahat, Ezgi; Wang, Victor; Cobos, Enrique J; Stucky, Cheryl L; Ma, Qiufu; Liberles, Stephen D; Woolf, Clifford J

    2014-01-01

    The somatosensory nervous system is critical for the organism's ability to respond to mechanical, thermal, and nociceptive stimuli. Somatosensory neurons are functionally and anatomically diverse but their molecular profiles are not well-defined. Here, we used transcriptional profiling to analyze the detailed molecular signatures of dorsal root ganglion (DRG) sensory neurons. We used two mouse reporter lines and surface IB4 labeling to purify three major non-overlapping classes of neurons: 1) IB4+SNS-Cre/TdTomato+, 2) IB4−SNS-Cre/TdTomato+, and 3) Parv-Cre/TdTomato+ cells, encompassing the majority of nociceptive, pruriceptive, and proprioceptive neurons. These neurons displayed distinct expression patterns of ion channels, transcription factors, and GPCRs. Highly parallel qRT-PCR analysis of 334 single neurons selected by membership of the three populations demonstrated further diversity, with unbiased clustering analysis identifying six distinct subgroups. These data significantly increase our knowledge of the molecular identities of known DRG populations and uncover potentially novel subsets, revealing the complexity and diversity of those neurons underlying somatosensation. DOI: http://dx.doi.org/10.7554/eLife.04660.001 PMID:25525749

  1. Complexity of CNC transcription factors as revealed by gene targeting of the Nrf3 locus.

    Science.gov (United States)

    Derjuga, Anna; Gourley, Tania S; Holm, Teresa M; Heng, Henry H Q; Shivdasani, Ramesh A; Ahmed, Rafi; Andrews, Nancy C; Blank, Volker

    2004-04-01

    Cap'n'collar (CNC) family basic leucine zipper transcription factors play crucial roles in the regulation of mammalian gene expression and development. To determine the in vivo function of the CNC protein Nrf3 (NF-E2-related factor 3), we generated mice deficient in this transcription factor. We performed targeted disruption of two Nrf3 exons coding for CNC homology, basic DNA-binding, and leucine zipper dimerization domains. Nrf3 null mice developed normally and revealed no obvious phenotypic differences compared to wild-type animals. Nrf3(-/-) mice were fertile, and gross anatomy as well as behavior appeared normal. The mice showed normal age progression and did not show any apparent additional phenotype during their life span. We observed no differences in various blood parameters and chemistry values. We infected wild-type and Nrf3(-/-) mice with acute lymphocytic choriomeningitis virus and found no differences in these animals with respect to their number of virus-specific CD8 and CD4 T cells as well as their B-lymphocyte response. To determine whether the mild phenotype of Nrf3 null animals is due to functional redundancy, we generated mice deficient in multiple CNC factors. Contrary to our expectations, an absence of Nrf3 does not seem to cause additional lethality in compound Nrf3(-/-)/Nrf2(-/-) and Nrf3(-/-)/p45(-/-) mice. We hypothesize that the role of Nrf3 in vivo may become apparent only after appropriate challenge to the mice.

  2. Integrative Genomics Reveals Mechanisms of Copy Number Alterations Responsible for Transcriptional Deregulation in Colorectal Cancer

    Science.gov (United States)

    Camps, Jordi; Nguyen, Quang Tri; Padilla-Nash, Hesed M.; Knutsen, Turid; McNeil, Nicole E.; Wangsa, Danny; Hummon, Amanda B.; Grade, Marian; Ried, Thomas; Difilippantonio, Michael J.

    2016-01-01

    To evaluate the mechanisms and consequences of chromosomal aberrations in colorectal cancer (CRC), we used a combination of spectral karyotyping, array comparative genomic hybridization (aCGH), and array-based global gene expression profiling on 31 primary carcinomas and 15 established cell lines. Importantly, aCGH showed that the genomic profiles of primary tumors are recapitulated in the cell lines. We revealed a preponderance of chromosome breakpoints at sites of copy number variants (CNVs) in the CRC cell lines, a novel mechanism of DNA breakage in cancer. The integration of gene expression and aCGH led to the identification of 157 genes localized within high-level copy number changes whose transcriptional deregulation was significantly affected across all of the samples, thereby suggesting that these genes play a functional role in CRC. Genomic amplification at 8q24 was the most recurrent event and led to the overexpression of MYC and FAM84B. Copy number dependent gene expression resulted in deregulation of known cancer genes such as APC, FGFR2, and ERBB2. The identification of only 36 genes whose localization near a breakpoint could account for their observed deregulated expression demonstrates that the major mechanism for transcriptional deregulation in CRC is genomic copy number changes resulting from chromosomal aberrations. PMID:19691111

  3. Kinome-wide transcriptional profiling of uveal melanoma reveals new vulnerabilities to targeted therapeutics.

    Science.gov (United States)

    Bailey, Fiona P; Clarke, Kim; Kalirai, Helen; Kenyani, Jenna; Shahidipour, Haleh; Falciani, Francesco; Coulson, Judy M; Sacco, Joseph J; Coupland, Sarah E; Eyers, Patrick A

    2018-03-01

    Metastatic uveal melanoma (UM) is invariably fatal, usually within a year of diagnosis. There are currently no effective therapies, and clinical studies employing kinase inhibitors have so far demonstrated limited success. This is despite common activating mutations in GNAQ/11 genes, which trigger signalling pathways that might predispose tumours to a variety of targeted drugs. In this study, we have profiled kinome expression network dynamics in various human ocular melanomas. We uncovered a shared transcriptional profile in human primary UM samples and across a variety of experimental cell-based models. The poor overall response of UM cells to FDA-approved kinase inhibitors contrasted with much higher sensitivity to the bromodomain inhibitor JQ1, a broad transcriptional repressor. Mechanistically, we identified a repressed FOXM1-dependent kinase subnetwork in JQ1-exposed cells that contained multiple cell cycle-regulated protein kinases. Consistently, we demonstrated vulnerability of UM cells to inhibitors of mitotic protein kinases within this network, including the investigational PLK1 inhibitor BI6727. We conclude that analysis of kinome-wide signalling network dynamics has the potential to reveal actionable drug targets and inhibitors of potential therapeutic benefit for UM patients. © 2017 The Authors. Pigment Cell & Melanoma Research Published by John Wiley & Sons.

  4. Dynamic in vivo binding of transcription factors to cis-regulatory modules of cer and gsc in the stepwise formation of the Spemann–Mangold organizer

    Science.gov (United States)

    Sudou, Norihiro; Yamamoto, Shinji; Ogino, Hajime; Taira, Masanori

    2012-01-01

    How multiple developmental cues are integrated on cis-regulatory modules (CRMs) for cell fate decisions remains uncertain. The Spemann–Mangold organizer in Xenopus embryos expresses the transcription factors Lim1/Lhx1, Otx2, Mix1, Siamois (Sia) and VegT. Reporter analyses using sperm nuclear transplantation and DNA injection showed that cerberus (cer) and goosecoid (gsc) are activated by the aforementioned transcription factors through CRMs conserved between X. laevis and X. tropicalis. ChIP-qPCR analysis for the five transcription factors revealed that cer and gsc CRMs are initially bound by both Sia and VegT at the late blastula stage, and subsequently bound by all five factors at the gastrula stage. At the neurula stage, only binding of Lim1 and Otx2 to the gsc CRM, among others, persists, which corresponds to their co-expression in the prechordal plate. Based on these data, together with detailed expression pattern analysis, we propose a new model of stepwise formation of the organizer, in which (1) maternal VegT and Wnt-induced Sia first bind to CRMs at the blastula stage; then (2) Nodal-inducible Lim1, Otx2, Mix1 and zygotic VegT are bound to CRMs in the dorsal endodermal and mesodermal regions where all these genes are co-expressed; and (3) these two regions are combined at the gastrula stage to form the organizer. Thus, the in vivo dynamics of multiple transcription factors highlight their roles in the initiation and maintenance of gene expression, and also reveal the stepwise integration of maternal, Nodal and Wnt signaling on CRMs of organizer genes to generate the organizer. PMID:22492356

  5. Dynamic in vivo binding of transcription factors to cis-regulatory modules of cer and gsc in the stepwise formation of the Spemann-Mangold organizer.

    Science.gov (United States)

    Sudou, Norihiro; Yamamoto, Shinji; Ogino, Hajime; Taira, Masanori

    2012-05-01

    How multiple developmental cues are integrated on cis-regulatory modules (CRMs) for cell fate decisions remains uncertain. The Spemann-Mangold organizer in Xenopus embryos expresses the transcription factors Lim1/Lhx1, Otx2, Mix1, Siamois (Sia) and VegT. Reporter analyses using sperm nuclear transplantation and DNA injection showed that cerberus (cer) and goosecoid (gsc) are activated by the aforementioned transcription factors through CRMs conserved between X. laevis and X. tropicalis. ChIP-qPCR analysis for the five transcription factors revealed that cer and gsc CRMs are initially bound by both Sia and VegT at the late blastula stage, and subsequently bound by all five factors at the gastrula stage. At the neurula stage, only binding of Lim1 and Otx2 to the gsc CRM, among others, persists, which corresponds to their co-expression in the prechordal plate. Based on these data, together with detailed expression pattern analysis, we propose a new model of stepwise formation of the organizer, in which (1) maternal VegT and Wnt-induced Sia first bind to CRMs at the blastula stage; then (2) Nodal-inducible Lim1, Otx2, Mix1 and zygotic VegT are bound to CRMs in the dorsal endodermal and mesodermal regions where all these genes are co-expressed; and (3) these two regions are combined at the gastrula stage to form the organizer. Thus, the in vivo dynamics of multiple transcription factors highlight their roles in the initiation and maintenance of gene expression, and also reveal the stepwise integration of maternal, Nodal and Wnt signaling on CRMs of organizer genes to generate the organizer.

  6. Identification of basic/helix-loop-helix transcription factors reveals candidate genes involved in anthocyanin biosynthesis from the strawberry white-flesh mutant

    OpenAIRE

    Zhao, Fengli; Li, Gang; Hu, Panpan; Zhao, Xia; Li, Liangjie; Wei, Wei; Feng, Jiayue; Zhou, Houcheng

    2018-01-01

    As the second largest transcription factor family in plant, the basic helix-loop-helix (bHLH) transcription factor family, characterized by the conserved bHLH domain, plays a central regulatory role in many biological process. However, the bHLH transcription factor family of strawberry has not been systematically identified, especially for the anthocyanin biosynthesis. Here, we identified a total of 113 bHLH transcription factors and described their chromosomal distribution and bioinformatics...

  7. Sasquatch: predicting the impact of regulatory SNPs on transcription factor binding from cell- and tissue-specific DNase footprints.

    Science.gov (United States)

    Schwessinger, Ron; Suciu, Maria C; McGowan, Simon J; Telenius, Jelena; Taylor, Stephen; Higgs, Doug R; Hughes, Jim R

    2017-10-01

    In the era of genome-wide association studies (GWAS) and personalized medicine, predicting the impact of single nucleotide polymorphisms (SNPs) in regulatory elements is an important goal. Current approaches to determine the potential of regulatory SNPs depend on inadequate knowledge of cell-specific DNA binding motifs. Here, we present Sasquatch, a new computational approach that uses DNase footprint data to estimate and visualize the effects of noncoding variants on transcription factor binding. Sasquatch performs a comprehensive k -mer-based analysis of DNase footprints to determine any k -mer's potential for protein binding in a specific cell type and how this may be changed by sequence variants. Therefore, Sasquatch uses an unbiased approach, independent of known transcription factor binding sites and motifs. Sasquatch only requires a single DNase-seq data set per cell type, from any genotype, and produces consistent predictions from data generated by different experimental procedures and at different sequence depths. Here we demonstrate the effectiveness of Sasquatch using previously validated functional SNPs and benchmark its performance against existing approaches. Sasquatch is available as a versatile webtool incorporating publicly available data, including the human ENCODE collection. Thus, Sasquatch provides a powerful tool and repository for prioritizing likely regulatory SNPs in the noncoding genome. © 2017 Schwessinger et al.; Published by Cold Spring Harbor Laboratory Press.

  8. A Regulatory Circuit Composed of a Transcription Factor, IscR, and a Regulatory RNA, RyhB, Controls Fe-S Cluster Delivery.

    Science.gov (United States)

    Mandin, Pierre; Chareyre, Sylvia; Barras, Frédéric

    2016-09-20

    Fe-S clusters are cofactors conserved through all domains of life. Once assembled by dedicated ISC and/or SUF scaffolds, Fe-S clusters are conveyed to their apo-targets via A-type carrier proteins (ATCs). Escherichia coli possesses four such ATCs. ErpA is the only ATC essential under aerobiosis. Recent studies reported a possible regulation of the erpA mRNA by the small RNA (sRNA) RyhB, which controls the expression of many genes under iron starvation. Surprisingly, erpA has not been identified in recent transcriptomic analysis of the iron starvation response, thus bringing into question the actual physiological significance of the putative regulation of erpA by RyhB. Using an sRNA library, we show that among 26 sRNAs, only RyhB represses the expression of an erpA-lacZ translational fusion. We further demonstrate that this repression occurs during iron starvation. Using mutational analysis, we show that RyhB base pairs to the erpA mRNA, inducing its disappearance. In addition, IscR, the master regulator of Fe-S homeostasis, represses expression of erpA at the transcriptional level when iron is abundant, but depleting iron from the medium alleviates this repression. The conjunction of transcriptional derepression by IscR and posttranscriptional repression by RyhB under Fe-limiting conditions is best described as an incoherent regulatory circuit. This double regulation allows full expression of erpA at iron concentrations for which Fe-S biogenesis switches from the ISC to the SUF system. We further provide evidence that this regulatory circuit coordinates ATC usage to iron availability. Regulatory small RNAs (sRNAs) have emerged as major actors in the control of gene expression in the last few decades. Relatively little is known about how these regulators interact with classical transcription factors to coordinate genetic responses. We show here how an sRNA, RyhB, and a transcription factor, IscR, regulate expression of an essential gene, erpA, in the bacterium E

  9. Elements in the transcriptional regulatory region flanking herpes simplex virus type 1 oriS stimulate origin function.

    Science.gov (United States)

    Wong, S W; Schaffer, P A

    1991-05-01

    Like other DNA-containing viruses, the three origins of herpes simplex virus type 1 (HSV-1) DNA replication are flanked by sequences containing transcriptional regulatory elements. In a transient plasmid replication assay, deletion of sequences comprising the transcriptional regulatory elements of ICP4 and ICP22/47, which flank oriS, resulted in a greater than 80-fold decrease in origin function compared with a plasmid, pOS-822, which retains these sequences. In an effort to identify specific cis-acting elements responsible for this effect, we conducted systematic deletion analysis of the flanking region with plasmid pOS-822 and tested the resulting mutant plasmids for origin function. Stimulation by cis-acting elements was shown to be both distance and orientation dependent, as changes in either parameter resulted in a decrease in oriS function. Additional evidence for the stimulatory effect of flanking sequences on origin function was demonstrated by replacement of these sequences with the cytomegalovirus immediate-early promoter, resulting in nearly wild-type levels of oriS function. In competition experiments, cotransfection of cells with the test plasmid, pOS-822, and increasing molar concentrations of a competitor plasmid which contained the ICP4 and ICP22/47 transcriptional regulatory regions but lacked core origin sequences resulted in a significant reduction in the replication efficiency of pOS-822, demonstrating that factors which bind specifically to the oriS-flanking sequences are likely involved as auxiliary proteins in oriS function. Together, these studies demonstrate that trans-acting factors and the sites to which they bind play a critical role in the efficiency of HSV-1 DNA replication from oriS in transient-replication assays.

  10. Integrative analyses reveal a long noncoding RNA-mediated sponge regulatory network in prostate cancer.

    Science.gov (United States)

    Du, Zhou; Sun, Tong; Hacisuleyman, Ezgi; Fei, Teng; Wang, Xiaodong; Brown, Myles; Rinn, John L; Lee, Mary Gwo-Shu; Chen, Yiwen; Kantoff, Philip W; Liu, X Shirley

    2016-03-15

    Mounting evidence suggests that long noncoding RNAs (lncRNAs) can function as microRNA sponges and compete for microRNA binding to protein-coding transcripts. However, the prevalence, functional significance and targets of lncRNA-mediated sponge regulation of cancer are mostly unknown. Here we identify a lncRNA-mediated sponge regulatory network that affects the expression of many protein-coding prostate cancer driver genes, by integrating analysis of sequence features and gene expression profiles of both lncRNAs and protein-coding genes in tumours. We confirm the tumour-suppressive function of two lncRNAs (TUG1 and CTB-89H12.4) and their regulation of PTEN expression in prostate cancer. Surprisingly, one of the two lncRNAs, TUG1, was previously known for its function in polycomb repressive complex 2 (PRC2)-mediated transcriptional regulation, suggesting its sub-cellular localization-dependent function. Our findings not only suggest an important role of lncRNA-mediated sponge regulation in cancer, but also underscore the critical influence of cytoplasmic localization on the efficacy of a sponge lncRNA.

  11. Functional organization of an Mbp enhancer exposes striking transcriptional regulatory diversity within myelinating glia

    DEFF Research Database (Denmark)

    Dionne, Nancy; Dib, Samar; Finsen, Bente

    2016-01-01

    regulatory element combinations were found to drive expression in oligodendrocytes and Schwann cells with a minimal 129 bp sequence conferring expression in oligodendrocytes throughout myelin elaboration, maintenance and repair. Unexpectedly, M3 derivatives conferred markedly different spatial and temporal...

  12. Identifying significant genetic regulatory networks in the prostate cancer from microarray data based on transcription factor analysis and conditional independency

    Directory of Open Access Journals (Sweden)

    Yeh Cheng-Yu

    2009-12-01

    Full Text Available Abstract Background Prostate cancer is a world wide leading cancer and it is characterized by its aggressive metastasis. According to the clinical heterogeneity, prostate cancer displays different stages and grades related to the aggressive metastasis disease. Although numerous studies used microarray analysis and traditional clustering method to identify the individual genes during the disease processes, the important gene regulations remain unclear. We present a computational method for inferring genetic regulatory networks from micorarray data automatically with transcription factor analysis and conditional independence testing to explore the potential significant gene regulatory networks that are correlated with cancer, tumor grade and stage in the prostate cancer. Results To deal with missing values in microarray data, we used a K-nearest-neighbors (KNN algorithm to determine the precise expression values. We applied web services technology to wrap the bioinformatics toolkits and databases to automatically extract the promoter regions of DNA sequences and predicted the transcription factors that regulate the gene expressions. We adopt the microarray datasets consists of 62 primary tumors, 41 normal prostate tissues from Stanford Microarray Database (SMD as a target dataset to evaluate our method. The predicted results showed that the possible biomarker genes related to cancer and denoted the androgen functions and processes may be in the development of the prostate cancer and promote the cell death in cell cycle. Our predicted results showed that sub-networks of genes SREBF1, STAT6 and PBX1 are strongly related to a high extent while ETS transcription factors ELK1, JUN and EGR2 are related to a low extent. Gene SLC22A3 may explain clinically the differentiation associated with the high grade cancer compared with low grade cancer. Enhancer of Zeste Homolg 2 (EZH2 regulated by RUNX1 and STAT3 is correlated to the pathological stage

  13. Identifying significant genetic regulatory networks in the prostate cancer from microarray data based on transcription factor analysis and conditional independency.

    Science.gov (United States)

    Yeh, Hsiang-Yuan; Cheng, Shih-Wu; Lin, Yu-Chun; Yeh, Cheng-Yu; Lin, Shih-Fang; Soo, Von-Wun

    2009-12-21

    Prostate cancer is a world wide leading cancer and it is characterized by its aggressive metastasis. According to the clinical heterogeneity, prostate cancer displays different stages and grades related to the aggressive metastasis disease. Although numerous studies used microarray analysis and traditional clustering method to identify the individual genes during the disease processes, the important gene regulations remain unclear. We present a computational method for inferring genetic regulatory networks from micorarray data automatically with transcription factor analysis and conditional independence testing to explore the potential significant gene regulatory networks that are correlated with cancer, tumor grade and stage in the prostate cancer. To deal with missing values in microarray data, we used a K-nearest-neighbors (KNN) algorithm to determine the precise expression values. We applied web services technology to wrap the bioinformatics toolkits and databases to automatically extract the promoter regions of DNA sequences and predicted the transcription factors that regulate the gene expressions. We adopt the microarray datasets consists of 62 primary tumors, 41 normal prostate tissues from Stanford Microarray Database (SMD) as a target dataset to evaluate our method. The predicted results showed that the possible biomarker genes related to cancer and denoted the androgen functions and processes may be in the development of the prostate cancer and promote the cell death in cell cycle. Our predicted results showed that sub-networks of genes SREBF1, STAT6 and PBX1 are strongly related to a high extent while ETS transcription factors ELK1, JUN and EGR2 are related to a low extent. Gene SLC22A3 may explain clinically the differentiation associated with the high grade cancer compared with low grade cancer. Enhancer of Zeste Homolg 2 (EZH2) regulated by RUNX1 and STAT3 is correlated to the pathological stage. We provide a computational framework to reconstruct

  14. Large-scale transcriptome data reveals transcriptional activity of fission yeast LTR retrotransposons

    DEFF Research Database (Denmark)

    Mourier, Tobias; Willerslev, Eske

    2010-01-01

    of transcriptional activity are observed from both strands of solitary LTR sequences. Transcriptome data collected during meiosis suggests that transcription of solitary LTRs is correlated with the transcription of nearby protein-coding genes. CONCLUSIONS: Presumably, the host organism negatively regulates...

  15. A chemical perspective on transcriptional fidelity dominant contributions of sugar integrity revealed by unlocked nucleic acids

    DEFF Research Database (Denmark)

    Xu, Liang; Plouffe, Steven W; Chong, Jenny

    2013-01-01

    Transcription unlocked: A synthetic chemical biology approach involving unlocked nucleic acids was used to dissect the contribution of sugar backbone integrity to the RNA Polymerase II (Pol II) transcription process. An unexpected dominant role for sugar-ring integrity in Pol II transcriptional...

  16. Identification of cis-regulatory sequences that activate transcription in the suspensor of plant embryos.

    Science.gov (United States)

    Kawashima, Tomokazu; Wang, Xingjun; Henry, Kelli F; Bi, Yuping; Weterings, Koen; Goldberg, Robert B

    2009-03-03

    Little is known about the molecular mechanisms by which the embryo proper and suspensor of plant embryos activate specific gene sets shortly after fertilization. We analyzed the upstream region of the scarlet runner bean (Phaseolus coccineus) G564 gene to understand how genes are activated specifically within the suspensor during early embryo development. Previously, we showed that the G564 upstream region has a block of tandem repeats, which contain a conserved 10-bp motif (GAAAAG(C)/(T)GAA), and that deletion of these repeats results in a loss of suspensor transcription. Here, we use gain-of-function (GOF) experiments with transgenic globular-stage tobacco embryos to show that only 1 of the 5 tandem repeats is required to drive suspensor-specific transcription. Fine-scale deletion and scanning mutagenesis experiments with 1 tandem repeat uncovered a 54-bp region that contains all of the sequences required to activate transcription in the suspensor, including the 10-bp motif (GAAAAGCGAA) and a similar 10-bp-like motif (GAAAAACGAA). Site-directed mutagenesis and GOF experiments indicated that both the 10-bp and 10-bp-like motifs are necessary, but not sufficient to activate transcription in the suspensor, and that a sequence (TTGGT) between the 10-bp and the 10-bp-like motifs is also necessary for suspensor transcription. Together, these data identify sequences that are required to activate transcription in the suspensor of a plant embryo after fertilization.

  17. Mapping Mammalian Cell-type-specific Transcriptional Regulatory Networks Using KD-CAGE and ChIP-seq Data in the TC-YIK Cell Line

    Science.gov (United States)

    Lizio, Marina; Ishizu, Yuri; Itoh, Masayoshi; Lassmann, Timo; Hasegawa, Akira; Kubosaki, Atsutaka; Severin, Jessica; Kawaji, Hideya; Nakamura, Yukio; Suzuki, Harukazu; Hayashizaki, Yoshihide; Carninci, Piero; Forrest, Alistair R. R.

    2015-01-01

    Mammals are composed of hundreds of different cell types with specialized functions. Each of these cellular phenotypes are controlled by different combinations of transcription factors. Using a human non islet cell insulinoma cell line (TC-YIK) which expresses insulin and the majority of known pancreatic beta cell specific genes as an example, we describe a general approach to identify key cell-type-specific transcription factors (TFs) and their direct and indirect targets. By ranking all human TFs by their level of enriched expression in TC-YIK relative to a broad collection of samples (FANTOM5), we confirmed known key regulators of pancreatic function and development. Systematic siRNA mediated perturbation of these TFs followed by qRT-PCR revealed their interconnections with NEUROD1 at the top of the regulation hierarchy and its depletion drastically reducing insulin levels. For 15 of the TF knock-downs (KD), we then used Cap Analysis of Gene Expression (CAGE) to identify thousands of their targets genome-wide (KD-CAGE). The data confirm NEUROD1 as a key positive regulator in the transcriptional regulatory network (TRN), and ISL1, and PROX1 as antagonists. As a complimentary approach we used ChIP-seq on four of these factors to identify NEUROD1, LMX1A, PAX6, and RFX6 binding sites in the human genome. Examining the overlap between genes perturbed in the KD-CAGE experiments and genes with a ChIP-seq peak within 50 kb of their promoter, we identified direct transcriptional targets of these TFs. Integration of KD-CAGE and ChIP-seq data shows that both NEUROD1 and LMX1A work as the main transcriptional activators. In the core TRN (i.e., TF-TF only), NEUROD1 directly transcriptionally activates the pancreatic TFs HSF4, INSM1, MLXIPL, MYT1, NKX6-3, ONECUT2, PAX4, PROX1, RFX6, ST18, DACH1, and SHOX2, while LMX1A directly transcriptionally activates DACH1, SHOX2, PAX6, and PDX1. Analysis of these complementary datasets suggests the need for caution in interpreting Ch

  18. Transcriptional profiling reveals gland-specific differential expression in the three major salivary glands of the adult mouse.

    Science.gov (United States)

    Gao, Xin; Oei, Maria S; Ovitt, Catherine E; Sincan, Murat; Melvin, James E

    2018-04-01

    RNA-Seq was used to better understand the molecular nature of the biological differences among the three major exocrine salivary glands in mammals. Transcriptional profiling found that the adult murine parotid, submandibular, and sublingual salivary glands express greater than 14,300 protein-coding genes, and nearly 2,000 of these genes were differentially expressed. Principle component analysis of the differentially expressed genes revealed three distinct clusters according to gland type. The three salivary gland transcriptomes were dominated by a relatively few number of highly expressed genes (6.3%) that accounted for more than 90% of transcriptional output. Of the 912 transcription factors expressed in the major salivary glands, greater than 90% of them were detected in all three glands, while expression for ~2% of them was enriched in an individual gland. Expression of these unique transcription factors correlated with sublingual and parotid specific subsets of both highly expressed and differentially expressed genes. Gene ontology analyses revealed that the highly expressed genes common to all glands were associated with global functions, while many of the genes expressed in a single gland play a major role in the function of that gland. In summary, transcriptional profiling of the three murine major salivary glands identified a limited number of highly expressed genes, differentially expressed genes, and unique transcription factors that represent the transcriptional signatures underlying gland-specific biological properties.

  19. Fruit specific variability in capsaicinoid accumulation and transcription of structural and regulatory genes in Capsicum fruit.

    Science.gov (United States)

    Keyhaninejad, Neda; Curry, Jeanne; Romero, Joslynn; O'Connell, Mary A

    2014-02-01

    Accumulation of capsaicinoids in the placental tissue of ripening chile (Capsicum spp.) fruit follows the coordinated expression of multiple biosynthetic enzymes producing the substrates for capsaicin synthase. Transcription factors are likely agents to regulate expression of these biosynthetic genes. Placental RNAs from habanero fruit (Capsicum chinense) were screened for expression of candidate transcription factors; with two candidate genes identified, both in the ERF family of transcription factors. Characterization of these transcription factors, Erf and Jerf, in nine chile cultivars with distinct capsaicinoid contents demonstrated a correlation of expression with pungency. Amino acid variants were observed in both ERF and JERF from different chile cultivars; none of these changes involved the DNA binding domains. Little to no transcription of Erf was detected in non-pungent Capsium annuum or C. chinense mutants. This correlation was characterized at an individual fruit level in a set of jalapeño (C. annuum) lines again with distinct and variable capsaicinoid contents. Both Erf and Jerf are expressed early in fruit development, 16-20 days post-anthesis, at times prior to the accumulation of capsaicinoids in the placental tissues. These data support the hypothesis that these two members of the complex ERF family participate in regulation of the pungency phenotype in chile. Copyright © 2013. Published by Elsevier Ireland Ltd.

  20. Interactome maps of mouse gene regulatory domains reveal basic principles of transcriptional regulation

    DEFF Research Database (Denmark)

    Kieffer-Kwon, Kyong-Rim; Tang, Zhonghui; Mathe, Ewy

    2013-01-01

    IA-PET technologies to map the promoter-enhancer interactomes of pluripotent ES cells and differentiated B lymphocytes. We confirm that enhancer usage varies widely across tissues. Unexpectedly, we find that this feature extends to broadly transcribed genes, including Myc and Pim1 cell-cycle regulators, which...... associate with an entirely different set of enhancers in ES and B cells. By means of high-resolution CpG methylomes, genome editing, and digital footprinting, we show that these enhancers recruit lineage-determining factors. Furthermore, we demonstrate that the turning on and off of enhancers during...

  1. Discovery of a Regulatory Motif for Human Satellite DNA Transcription in Response to BATF2 Overexpression.

    Science.gov (United States)

    Bai, Xuejia; Huang, Wenqiu; Zhang, Chenguang; Niu, Jing; Ding, Wei

    2016-03-01

    One of the basic leucine zipper transcription factors, BATF2, has been found to suppress cancer growth and migration. However, little is known about the genes downstream of BATF2. HeLa cells were stably transfected with BATF2, then chromatin immunoprecipitation-sequencing was employed to identify the DNA motifs responsive to BATF2. Comprehensive bioinformatics analyses indicated that the most significant motif discovered as TTCCATT[CT]GATTCCATTC[AG]AT was primarily distributed among the chromosome centromere regions and mostly within human type II satellite DNA. Such motifs were able to prime the transcription of type II satellite DNA in a directional and asymmetrical manner. Consistently, satellite II transcription was up-regulated in BATF2-overexpressing cells. The present study provides insight into understanding the role of BATF2 in tumours and the importance of satellite DNA in the maintenance of genomic stability. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  2. Sheep skeletal muscle transcriptome analysis reveals muscle growth regulatory lncRNAs.

    Science.gov (United States)

    Chao, Tianle; Ji, Zhibin; Hou, Lei; Wang, Jin; Zhang, Chunlan; Wang, Guizhi; Wang, Jianmin

    2018-01-01

    As widely distributed domestic animals, sheep are an important species and the source of mutton. In this study, we aimed to evaluate the regulatory lncRNAs associated with muscle growth and development between high production mutton sheep (Dorper sheep and Qianhua Mutton Merino sheep) and low production mutton sheep (Small-tailed Han sheep). In total, 39 lncRNAs were found to be differentially expressed. Using co-expression analysis and functional annotation, 1,206 co-expression interactions were found between 32 lncRNAs and 369 genes, and 29 of these lncRNAs were found to be associated with muscle development, metabolism, cell proliferation and apoptosis. lncRNA-mRNA interactions revealed 6 lncRNAs as hub lncRNAs. Moreover, three lncRNAs and their associated co-expressed genes were demonstrated by cis-regulatory gene analyses, and we also found a potential regulatory relationship between the pseudogene lncRNA LOC101121401 and its parent gene FTH1. This study provides a genome-wide resolution of lncRNA and mRNA regulation in muscles from mutton sheep.

  3. Comparative study of discretization methods of microarray data for inferring transcriptional regulatory networks

    Directory of Open Access Journals (Sweden)

    Ji Wei

    2010-10-01

    Full Text Available Abstract Background Microarray data discretization is a basic preprocess for many algorithms of gene regulatory network inference. Some common discretization methods in informatics are used to discretize microarray data. Selection of the discretization method is often arbitrary and no systematic comparison of different discretization has been conducted, in the context of gene regulatory network inference from time series gene expression data. Results In this study, we propose a new discretization method "bikmeans", and compare its performance with four other widely-used discretization methods using different datasets, modeling algorithms and number of intervals. Sensitivities, specificities and total accuracies were calculated and statistical analysis was carried out. Bikmeans method always gave high total accuracies. Conclusions Our results indicate that proper discretization methods can consistently improve gene regulatory network inference independent of network modeling algorithms and datasets. Our new method, bikmeans, resulted in significant better total accuracies than other methods.

  4. Likelihood for transcriptions in a genetic regulatory system under asymmetric stable Lévy noise.

    Science.gov (United States)

    Wang, Hui; Cheng, Xiujun; Duan, Jinqiao; Kurths, Jürgen; Li, Xiaofan

    2018-01-01

    This work is devoted to investigating the evolution of concentration in a genetic regulation system, when the synthesis reaction rate is under additive and multiplicative asymmetric stable Lévy fluctuations. By focusing on the impact of skewness (i.e., non-symmetry) in the probability distributions of noise, we find that via examining the mean first exit time (MFET) and the first escape probability (FEP), the asymmetric fluctuations, interacting with nonlinearity in the system, lead to peculiar likelihood for transcription. This includes, in the additive noise case, realizing higher likelihood of transcription for larger positive skewness (i.e., asymmetry) index β, causing a stochastic bifurcation at the non-Gaussianity index value α = 1 (i.e., it is a separating point or line for the likelihood for transcription), and achieving a turning point at the threshold value β≈-0.5 (i.e., beyond which the likelihood for transcription suddenly reversed for α values). The stochastic bifurcation and turning point phenomena do not occur in the symmetric noise case (β = 0). While in the multiplicative noise case, non-Gaussianity index value α = 1 is a separating point or line for both the MFET and the FEP. We also investigate the noise enhanced stability phenomenon. Additionally, we are able to specify the regions in the whole parameter space for the asymmetric noise, in which we attain desired likelihood for transcription. We have conducted a series of numerical experiments in "regulating" the likelihood of gene transcription by tuning asymmetric stable Lévy noise indexes. This work offers insights for possible ways of achieving gene regulation in experimental research.

  5. A Regulatory Circuit Composed of a Transcription Factor, IscR, and a Regulatory RNA, RyhB, Controls Fe-S Cluster Delivery

    Directory of Open Access Journals (Sweden)

    Pierre Mandin

    2016-09-01

    Full Text Available Fe-S clusters are cofactors conserved through all domains of life. Once assembled by dedicated ISC and/or SUF scaffolds, Fe-S clusters are conveyed to their apo-targets via A-type carrier proteins (ATCs. Escherichia coli possesses four such ATCs. ErpA is the only ATC essential under aerobiosis. Recent studies reported a possible regulation of the erpA mRNA by the small RNA (sRNA RyhB, which controls the expression of many genes under iron starvation. Surprisingly, erpA has not been identified in recent transcriptomic analysis of the iron starvation response, thus bringing into question the actual physiological significance of the putative regulation of erpA by RyhB. Using an sRNA library, we show that among 26 sRNAs, only RyhB represses the expression of an erpA-lacZ translational fusion. We further demonstrate that this repression occurs during iron starvation. Using mutational analysis, we show that RyhB base pairs to the erpA mRNA, inducing its disappearance. In addition, IscR, the master regulator of Fe-S homeostasis, represses expression of erpA at the transcriptional level when iron is abundant, but depleting iron from the medium alleviates this repression. The conjunction of transcriptional derepression by IscR and posttranscriptional repression by RyhB under Fe-limiting conditions is best described as an incoherent regulatory circuit. This double regulation allows full expression of erpA at iron concentrations for which Fe-S biogenesis switches from the ISC to the SUF system. We further provide evidence that this regulatory circuit coordinates ATC usage to iron availability.

  6. Phosphoproteomics Reveals Regulatory T Cell-Mediated DEF6 Dephosphorylation That Affects Cytokine Expression in Human Conventional T Cells

    KAUST Repository

    Joshi, Rubin N.

    2017-09-25

    Regulatory T cells (Tregs) control key events of immune tolerance, primarily by suppression of effector T cells. We previously revealed that Tregs rapidly suppress T cell receptor (TCR)-induced calcium store depletion in conventional CD4CD25 T cells (Tcons) independently of IP levels, consequently inhibiting NFAT signaling and effector cytokine expression. Here, we study Treg suppression mechanisms through unbiased phosphoproteomics of primary human Tcons upon TCR stimulation and Treg-mediated suppression, respectively. Tregs induced a state of overall decreased phosphorylation as opposed to TCR stimulation. We discovered novel phosphosites (T595_S597) in the DEF6 (SLAT) protein that were phosphorylated upon TCR stimulation and conversely dephosphorylated upon coculture with Tregs. Mutation of these DEF6 phosphosites abrogated interaction of DEF6 with the IP receptor and affected NFAT activation and cytokine transcription in primary Tcons. This novel mechanism and phosphoproteomics data resource may aid in modifying sensitivity of Tcons to Treg-mediated suppression in autoimmune disease or cancer.

  7. Human iPSC-Derived Cerebellar Neurons from a Patient with Ataxia-Telangiectasia Reveal Disrupted Gene Regulatory Networks

    Directory of Open Access Journals (Sweden)

    Sam P. Nayler

    2017-10-01

    Full Text Available Ataxia-telangiectasia (A-T is a rare genetic disorder caused by loss of function of the ataxia-telangiectasia-mutated kinase and is characterized by a predisposition to cancer, pulmonary disease, immune deficiency and progressive degeneration of the cerebellum. As animal models do not faithfully recapitulate the neurological aspects, it remains unclear whether cerebellar degeneration is a neurodevelopmental or neurodegenerative phenotype. To address the necessity for a human model, we first assessed a previously published protocol for the ability to generate cerebellar neuronal cells, finding it gave rise to a population of precursors highly enriched for markers of the early hindbrain such as EN1 and GBX2, and later more mature cerebellar markers including PTF1α, MATH1, HOXB4, ZIC3, PAX6, and TUJ1. RNA sequencing was used to classify differentiated cerebellar neurons generated from integration-free A-T and control induced pluripotent stem cells. Comparison of RNA sequencing data with datasets from the Allen Brain Atlas reveals in vitro-derived cerebellar neurons are transcriptionally similar to discrete regions of the human cerebellum, and most closely resemble the cerebellum at 22 weeks post-conception. We show that patient-derived cerebellar neurons exhibit disrupted gene regulatory networks associated with synaptic vesicle dynamics and oxidative stress, offering the first molecular insights into early cerebellar pathogenesis of ataxia-telangiectasia.

  8. The Complexity of Posttranscriptional Small RNA Regulatory Networks Revealed by In Silico Analysis of Gossypium arboreum L. Leaf, Flower and Boll Small Regulatory RNAs.

    Directory of Open Access Journals (Sweden)

    Hongtao Hu

    Full Text Available MicroRNAs (miRNAs and secondary small interfering RNAs (principally phased siRNAs or trans-acting siRNAs are two distinct subfamilies of small RNAs (sRNAs that are emerging as key regulators of posttranscriptional gene expression in plants. Both miRNAs and secondary-siRNAs (sec-siRNAs are processed from longer RNA precursors by DICER-LIKE proteins (DCLs. Gossypium arboreum L., also known as tree cotton or Asian cotton, is a diploid, possibly ancestral relative of tetraploid Gossypium hirsutum L., the predominant type of commercially grown cotton worldwide known as upland cotton. To understand the biological significance of these gene regulators in G. arboreum, a bioinformatics analysis was performed on G. arboreum small RNAs produced from G. arboreum leaf, flower, and boll tissues. Consequently, 263 miRNAs derived from 353 precursors, including 155 conserved miRNAs (cs-miRNAs and 108 novel lineage-specific miRNAs (ls-miRNAs. Along with miRNAs, 2,033 miRNA variants (isomiRNAs were identified as well. Those isomiRNAs with variation at the 3'-miRNA end were expressed at the highest levels, compared to other types of variants. In addition, 755 pha-siRNAs derived 319 pha-siRNA gene transcripts (PGTs were identified, and the potential pha-siRNA initiators were predicted. Also, 2,251 non-phased siRNAs were found as well, of which 1,088 appeared to be produced by so-called cis- or trans-cleavage of the PGTs observed at positions differing from pha-siRNAs. Of those sRNAs, 148 miRNAs/isomiRNAs and 274 phased/non-phased siRNAs were differentially expressed in one or more pairs of tissues examined. Target analysis revealed that target genes for both miRNAs and pha-siRNAs are involved a broad range of metabolic and enzymatic activities. We demonstrate that secondary siRNA production could result from initial cleavage of precursors by both miRNAs or isomiRNAs, and that subsequently produced phased and unphased siRNAs could result that also serve as triggers

  9. The Complexity of Posttranscriptional Small RNA Regulatory Networks Revealed by In Silico Analysis of Gossypium arboreum L. Leaf, Flower and Boll Small Regulatory RNAs.

    Science.gov (United States)

    Hu, Hongtao; Rashotte, Aaron M; Singh, Narendra K; Weaver, David B; Goertzen, Leslie R; Singh, Shree R; Locy, Robert D

    2015-01-01

    MicroRNAs (miRNAs) and secondary small interfering RNAs (principally phased siRNAs or trans-acting siRNAs) are two distinct subfamilies of small RNAs (sRNAs) that are emerging as key regulators of posttranscriptional gene expression in plants. Both miRNAs and secondary-siRNAs (sec-siRNAs) are processed from longer RNA precursors by DICER-LIKE proteins (DCLs). Gossypium arboreum L., also known as tree cotton or Asian cotton, is a diploid, possibly ancestral relative of tetraploid Gossypium hirsutum L., the predominant type of commercially grown cotton worldwide known as upland cotton. To understand the biological significance of these gene regulators in G. arboreum, a bioinformatics analysis was performed on G. arboreum small RNAs produced from G. arboreum leaf, flower, and boll tissues. Consequently, 263 miRNAs derived from 353 precursors, including 155 conserved miRNAs (cs-miRNAs) and 108 novel lineage-specific miRNAs (ls-miRNAs). Along with miRNAs, 2,033 miRNA variants (isomiRNAs) were identified as well. Those isomiRNAs with variation at the 3'-miRNA end were expressed at the highest levels, compared to other types of variants. In addition, 755 pha-siRNAs derived 319 pha-siRNA gene transcripts (PGTs) were identified, and the potential pha-siRNA initiators were predicted. Also, 2,251 non-phased siRNAs were found as well, of which 1,088 appeared to be produced by so-called cis- or trans-cleavage of the PGTs observed at positions differing from pha-siRNAs. Of those sRNAs, 148 miRNAs/isomiRNAs and 274 phased/non-phased siRNAs were differentially expressed in one or more pairs of tissues examined. Target analysis revealed that target genes for both miRNAs and pha-siRNAs are involved a broad range of metabolic and enzymatic activities. We demonstrate that secondary siRNA production could result from initial cleavage of precursors by both miRNAs or isomiRNAs, and that subsequently produced phased and unphased siRNAs could result that also serve as triggers of a second

  10. Mitochondrial biogenesis in brown adipose tissue is associated with differential expression of transcription regulatory factors

    Czech Academy of Sciences Publication Activity Database

    Villena, J. A.; Carmona, M. C.; Rodriguez de la Concepción, M.; Rossmeisl, Martin; Vinas, O.; Mampel, T.; Iglesias, R.; Giralt, M.; Villarroya, F.

    2002-01-01

    Roč. 59, č. 11 (2002), s. 1934-1944 ISSN 1420-682X Grant - others:Ministerio de Ciencia y Tecnología (ES) PM98.0188 Institutional research plan: CEZ:AV0Z5011922 Keywords : brown adipose tissue * mitochondria * transcription factors Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.259, year: 2002

  11. Epigenetic landscapes reveal transcription factors that regulate CD8+ T cell differentiation.

    Science.gov (United States)

    Yu, Bingfei; Zhang, Kai; Milner, J Justin; Toma, Clara; Chen, Runqiang; Scott-Browne, James P; Pereira, Renata M; Crotty, Shane; Chang, John T; Pipkin, Matthew E; Wang, Wei; Goldrath, Ananda W

    2017-05-01

    Dynamic changes in the expression of transcription factors (TFs) can influence the specification of distinct CD8 + T cell fates, but the observation of equivalent expression of TFs among differentially fated precursor cells suggests additional underlying mechanisms. Here we profiled the genome-wide histone modifications, open chromatin and gene expression of naive, terminal-effector, memory-precursor and memory CD8 + T cell populations induced during the in vivo response to bacterial infection. Integration of these data suggested that the expression and binding of TFs contributed to the establishment of subset-specific enhancers during differentiation. We developed a new bioinformatics method using the PageRank algorithm to reveal key TFs that influence the generation of effector and memory populations. The TFs YY1 and Nr3c1, both constitutively expressed during CD8 + T cell differentiation, regulated the formation of terminal-effector cell fates and memory-precursor cell fates, respectively. Our data define the epigenetic landscape of differentiation intermediates and facilitate the identification of TFs with previously unappreciated roles in CD8 + T cell differentiation.

  12. Epigenetic landscapes reveal transcription factors regulating CD8+ T cell differentiation

    Science.gov (United States)

    Yu, Bingfei; Zhang, Kai; Milner, J. Justin; Toma, Clara; Chen, Runqiang; Scott-Browne, James P.; Pereira, Renata M.; Crotty, Shane; Chang, John T.; Pipkin, Matthew E.; Wang, Wei; Goldrath, Ananda W.

    2017-01-01

    Dynamic changes in the expression of transcription factors (TFs) can influence specification of distinct CD8+ T cell fates, but the observation of equivalent expression of TF among differentially-fated precursor cells suggests additional underlying mechanisms. Here, we profiled genome-wide histone modifications, open chromatin and gene expression of naive, terminal-effector, memory-precursor and memory CD8+ T cell populations induced during the in vivo response to bacterial infection. Integration of these data suggested that TF expression and binding contributed to establishment of subset-specific enhancers during differentiation. We developed a new bioinformatics method using the PageRank algorithm to reveal novel TFs influencing the generation of effector and memory populations. The TFs YY1 and Nr3c1, both constitutively expressed during CD8+ T cell differentiation, regulated the formation of terminal-effector and memory-precursor cell-fates, respectively. Our data define the epigenetic landscape of differentiation intermediates, facilitating identification of TFs with previously unappreciated roles in CD8+ T cell differentiation. PMID:28288100

  13. Single nucleotide polymorphism in transcriptional regulatory regions and expression of environmentally responsive genes

    International Nuclear Information System (INIS)

    Wang, Xuting; Tomso, Daniel J.; Liu Xuemei; Bell, Douglas A.

    2005-01-01

    Single nucleotide polymorphisms (SNPs) in the human genome are DNA sequence variations that can alter an individual's response to environmental exposure. SNPs in gene coding regions can lead to changes in the biological properties of the encoded protein. In contrast, SNPs in non-coding gene regulatory regions may affect gene expression levels in an allele-specific manner, and these functional polymorphisms represent an important but relatively unexplored class of genetic variation. The main challenge in analyzing these SNPs is a lack of robust computational and experimental methods. Here, we first outline mechanisms by which genetic variation can impact gene regulation, and review recent findings in this area; then, we describe a methodology for bioinformatic discovery and functional analysis of regulatory SNPs in cis-regulatory regions using the assembled human genome sequence and databases on sequence polymorphism and gene expression. Our method integrates SNP and gene databases and uses a set of computer programs that allow us to: (1) select SNPs, from among the >9 million human SNPs in the NCBI dbSNP database, that are similar to cis-regulatory element (RE) consensus sequences; (2) map the selected dbSNP entries to the human genome assembly in order to identify polymorphic REs near gene start sites; (3) prioritize the candidate polymorphic RE containing genes by searching the existing genotype and gene expression data sets. The applicability of this system has been demonstrated through studies on p53 responsive elements and is being extended to additional pathways and environmentally responsive genes

  14. A central regulatory system largely controls transcriptional activation and repression responses to phosphate starvation in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Regla Bustos

    2010-09-01

    Full Text Available Plants respond to different stresses by inducing or repressing transcription of partially overlapping sets of genes. In Arabidopsis, the PHR1 transcription factor (TF has an important role in the control of phosphate (Pi starvation stress responses. Using transcriptomic analysis of Pi starvation in phr1, and phr1 phr1-like (phl1 mutants and in wild type plants, we show that PHR1 in conjunction with PHL1 controls most transcriptional activation and repression responses to phosphate starvation, regardless of the Pi starvation specificity of these responses. Induced genes are enriched in PHR1 binding sequences (P1BS in their promoters, whereas repressed genes do not show such enrichment, suggesting that PHR1(-like control of transcriptional repression responses is indirect. In agreement with this, transcriptomic analysis of a transgenic plant expressing PHR1 fused to the hormone ligand domain of the glucocorticoid receptor showed that PHR1 direct targets (i.e., displaying altered expression after GR:PHR1 activation by dexamethasone in the presence of cycloheximide corresponded largely to Pi starvation-induced genes that are highly enriched in P1BS. A minimal promoter containing a multimerised P1BS recapitulates Pi starvation-specific responsiveness. Likewise, mutation of P1BS in the promoter of two Pi starvation-responsive genes impaired their responsiveness to Pi starvation, but not to other stress types. Phylogenetic footprinting confirmed the importance of P1BS and PHR1 in Pi starvation responsiveness and indicated that P1BS acts in concert with other cis motifs. All together, our data show that PHR1 and PHL1 are partially redundant TF acting as central integrators of Pi starvation responses, both specific and generic. In addition, they indicate that transcriptional repression responses are an integral part of adaptive responses to stress.

  15. Phenotype specific analyses reveal distinct regulatory mechanism for chronically activated p53.

    Directory of Open Access Journals (Sweden)

    Kristina Kirschner

    2015-03-01

    Full Text Available The downstream functions of the DNA binding tumor suppressor p53 vary depending on the cellular context, and persistent p53 activation has recently been implicated in tumor suppression and senescence. However, genome-wide information about p53-target gene regulation has been derived mostly from acute genotoxic conditions. Using ChIP-seq and expression data, we have found distinct p53 binding profiles between acutely activated (through DNA damage and chronically activated (in senescent or pro-apoptotic conditions p53. Compared to the classical 'acute' p53 binding profile, 'chronic' p53 peaks were closely associated with CpG-islands. Furthermore, the chronic CpG-island binding of p53 conferred distinct expression patterns between senescent and pro-apoptotic conditions. Using the p53 targets seen in the chronic conditions together with external high-throughput datasets, we have built p53 networks that revealed extensive self-regulatory 'p53 hubs' where p53 and many p53 targets can physically interact with each other. Integrating these results with public clinical datasets identified the cancer-associated lipogenic enzyme, SCD, which we found to be directly repressed by p53 through the CpG-island promoter, providing a mechanistic link between p53 and the 'lipogenic phenotype', a hallmark of cancer. Our data reveal distinct phenotype associations of chronic p53 targets that underlie specific gene regulatory mechanisms.

  16. Transcriptional Regulation in Ebola Virus: Effects of Gene Border Structure and Regulatory Elements on Gene Expression and Polymerase Scanning Behavior.

    Science.gov (United States)

    Brauburger, Kristina; Boehmann, Yannik; Krähling, Verena; Mühlberger, Elke

    2016-02-15

    The highly pathogenic Ebola virus (EBOV) has a nonsegmented negative-strand (NNS) RNA genome containing seven genes. The viral genes either are separated by intergenic regions (IRs) of variable length or overlap. The structure of the EBOV gene overlaps is conserved throughout all filovirus genomes and is distinct from that of the overlaps found in other NNS RNA viruses. Here, we analyzed how diverse gene borders and noncoding regions surrounding the gene borders influence transcript levels and govern polymerase behavior during viral transcription. Transcription of overlapping genes in EBOV bicistronic minigenomes followed the stop-start mechanism, similar to that followed by IR-containing gene borders. When the gene overlaps were extended, the EBOV polymerase was able to scan the template in an upstream direction. This polymerase feature seems to be generally conserved among NNS RNA virus polymerases. Analysis of IR-containing gene borders showed that the IR sequence plays only a minor role in transcription regulation. Changes in IR length were generally well tolerated, but specific IR lengths led to a strong decrease in downstream gene expression. Correlation analysis revealed that these effects were largely independent of the surrounding gene borders. Each EBOV gene contains exceptionally long untranslated regions (UTRs) flanking the open reading frame. Our data suggest that the UTRs adjacent to the gene borders are the main regulators of transcript levels. A highly complex interplay between the different cis-acting elements to modulate transcription was revealed for specific combinations of IRs and UTRs, emphasizing the importance of the noncoding regions in EBOV gene expression control. Our data extend those from previous analyses investigating the implication of noncoding regions at the EBOV gene borders for gene expression control. We show that EBOV transcription is regulated in a highly complex yet not easily predictable manner by a set of interacting cis

  17. Analyses of Catharanthus roseus and Arabidopsis thaliana WRKY transcription factors reveal involvement in jasmonate signaling.

    Science.gov (United States)

    Schluttenhofer, Craig; Pattanaik, Sitakanta; Patra, Barunava; Yuan, Ling

    2014-06-20

    To combat infection to biotic stress plants elicit the biosynthesis of numerous natural products, many of which are valuable pharmaceutical compounds. Jasmonate is a central regulator of defense response to pathogens and accumulation of specialized metabolites. Catharanthus roseus produces a large number of terpenoid indole alkaloids (TIAs) and is an excellent model for understanding the regulation of this class of valuable compounds. Recent work illustrates a possible role for the Catharanthus WRKY transcription factors (TFs) in regulating TIA biosynthesis. In Arabidopsis and other plants, the WRKY TF family is also shown to play important role in controlling tolerance to biotic and abiotic stresses, as well as secondary metabolism. Here, we describe the WRKY TF families in response to jasmonate in Arabidopsis and Catharanthus. Publically available Arabidopsis microarrays revealed at least 30% (22 of 72) of WRKY TFs respond to jasmonate treatments. Microarray analysis identified at least six jasmonate responsive Arabidopsis WRKY genes (AtWRKY7, AtWRKY20, AtWRKY26, AtWRKY45, AtWRKY48, and AtWRKY72) that have not been previously reported. The Catharanthus WRKY TF family is comprised of at least 48 members. Phylogenetic clustering reveals 11 group I, 32 group II, and 5 group III WRKY TFs. Furthermore, we found that at least 25% (12 of 48) were jasmonate responsive, and 75% (9 of 12) of the jasmonate responsive CrWRKYs are orthologs of AtWRKYs known to be regulated by jasmonate. Overall, the CrWRKY family, ascertained from transcriptome sequences, contains approximately 75% of the number of WRKYs found in other sequenced asterid species (pepper, tomato, potato, and bladderwort). Microarray and transcriptomic data indicate that expression of WRKY TFs in Arabidopsis and Catharanthus are under tight spatio-temporal and developmental control, and potentially have a significant role in jasmonate signaling. Profiling of CrWRKY expression in response to jasmonate treatment

  18. LexA Binds to Transcription Regulatory Site of Cell Division Gene ftsZ in Toxic Cyanobacterium Microcystis aeruginosa.

    Science.gov (United States)

    Honda, Takashi; Morimoto, Daichi; Sako, Yoshihiko; Yoshida, Takashi

    2018-05-17

    Previously, we showed that DNA replication and cell division in toxic cyanobacterium Microcystis aeruginosa are coordinated by transcriptional regulation of cell division gene ftsZ and that an unknown protein specifically bound upstream of ftsZ (BpFz; DNA-binding protein to an upstream site of ftsZ) during successful DNA replication and cell division. Here, we purified BpFz from M. aeruginosa strain NIES-298 using DNA-affinity chromatography and gel-slicing combined with gel electrophoresis mobility shift assay (EMSA). The N-terminal amino acid sequence of BpFz was identified as TNLESLTQ, which was identical to that of transcription repressor LexA from NIES-843. EMSA analysis using mutant probes showed that the sequence GTACTAN 3 GTGTTC was important in LexA binding. Comparison of the upstream regions of lexA in the genomes of closely related cyanobacteria suggested that the sequence TASTRNNNNTGTWC could be a putative LexA recognition sequence (LexA box). Searches for TASTRNNNNTGTWC as a transcriptional regulatory site (TRS) in the genome of M. aeruginosa NIES-843 showed that it was present in genes involved in cell division, photosynthesis, and extracellular polysaccharide biosynthesis. Considering that BpFz binds to the TRS of ftsZ during normal cell division, LexA may function as a transcriptional activator of genes related to cell reproduction in M. aeruginosa, including ftsZ. This may be an example of informality in the control of bacterial cell division.

  19. Architecture of transcriptional regulatory circuits is knitted over the topology of bio-molecular interaction networks

    DEFF Research Database (Denmark)

    Soberano de Oliveira, Ana Paula; Patil, Kiran Raosaheb; Nielsen, Jens

    2008-01-01

    is to use the topology of bio-molecular interaction networks in order to constrain the solution space. Such approaches systematically integrate the existing biological knowledge with the 'omics' data. Results: Here we introduce a hypothesis-driven method that integrates bio-molecular network topology......Background: Uncovering the operating principles underlying cellular processes by using 'omics' data is often a difficult task due to the high-dimensionality of the solution space that spans all interactions among the bio-molecules under consideration. A rational way to overcome this problem...... with transcriptome data, thereby allowing the identification of key biological features (Reporter Features) around which transcriptional changes are significantly concentrated. We have combined transcriptome data with different biological networks in order to identify Reporter Gene Ontologies, Reporter Transcription...

  20. A regulatory transcriptional loop controls proliferation and differentiation in Drosophila neural stem cells.

    Directory of Open Access Journals (Sweden)

    Tetsuo Yasugi

    Full Text Available Neurogenesis is initiated by a set of basic Helix-Loop-Helix (bHLH transcription factors that specify neural progenitors and allow them to generate neurons in multiple rounds of asymmetric cell division. The Drosophila Daughterless (Da protein and its mammalian counterparts (E12/E47 act as heterodimerization factors for proneural genes and are therefore critically required for neurogenesis. Here, we demonstrate that Da can also be an inhibitor of the neural progenitor fate whose absence leads to stem cell overproliferation and tumor formation. We explain this paradox by demonstrating that Da induces the differentiation factor Prospero (Pros whose asymmetric segregation is essential for differentiation in one of the two daughter cells. Da co-operates with the bHLH transcription factor Asense, whereas the other proneural genes are dispensible. After mitosis, Pros terminates Asense expression in one of the two daughter cells. In da mutants, pros is not expressed, leading to the formation of lethal transplantable brain tumors. Our results define a transcriptional feedback loop that regulates the balance between self-renewal and differentiation in Drosophila optic lobe neuroblasts. They indicate that initiation of a neural differentiation program in stem cells is essential to prevent tumorigenesis.

  1. Transcriptional analysis and molecular dynamics simulations reveal the mechanism of toxic metals removal and efflux pumps in Lysinibacillus sphaericus OT4b.31

    KAUST Repository

    Shaw, Dario Rangel

    2017-11-23

    Lysinibacillus sphaericus strain OT4b.31 is a bacterium widely applied in bioremediation processes of hydrocarbon and metal polluted environments. In this study, we identified the molecular mechanism underlying the Pb2+ and Cr6+ resistance. Metal uptake and temporal transcription patterns of metal resistance operons were evaluated using reverse-transcribed quantitative PCR amplification. The function of the resistance determinants was studied applying docking and in silico mutagenesis methods. The results revealed that the adaptation of Lysinibacillus sphaericus OT4b.31 to elevated levels of lead and chromium involves the pbr and chr operons which comprise a transcriptional regulatory component (pbrR and chrB) and efflux ATPases (pbrA and chrA) to expel ions from the cytoplasm. Expression of metal resistance genes was constitutive and specifically inducible to the exposure of Pb2+ and Cr6+. The simultaneous presence of cations didn\\'t affect the bioaccumulation of metals, evidencing the multimetal resistance of L. sphaericus. Docking analysis revealed the key metal-protein interactions and the conformational changes after metal or ATP binding. Results showed that residues with aromatic rings or imidazole in the catalytic domain are crucial for metal binding and achievement of the function. To our knowledge, this is the first report of a specific mechanism for lead and chromium resistance in Lysinibacillus genus. From the findings of this study, it is possible to suggest the bacterium as a suitable candidate for rapid toxic metals bioremediation processes.

  2. Large-scale analysis of Arabidopsis transcription reveals a basal co-regulation network

    Directory of Open Access Journals (Sweden)

    Chamovitz Daniel A

    2009-09-01

    Full Text Available Abstract Background Analyses of gene expression data from microarray experiments has become a central tool for identifying co-regulated, functional gene modules. A crucial aspect of such analysis is the integration of data from different experiments and different laboratories. How to weigh the contribution of different experiments is an important point influencing the final outcomes. We have developed a novel method for this integration, and applied it to genome-wide data from multiple Arabidopsis microarray experiments performed under a variety of experimental conditions. The goal of this study is to identify functional globally co-regulated gene modules in the Arabidopsis genome. Results Following the analysis of 21,000 Arabidopsis genes in 43 datasets and about 2 × 108 gene pairs, we identified a globally co-expressed gene network. We found clusters of globally co-expressed Arabidopsis genes that are enriched for known Gene Ontology annotations. Two types of modules were identified in the regulatory network that differed in their sensitivity to the node-scoring parameter; we further showed these two pertain to general and specialized modules. Some of these modules were further investigated using the Genevestigator compendium of microarray experiments. Analyses of smaller subsets of data lead to the identification of condition-specific modules. Conclusion Our method for identification of gene clusters allows the integration of diverse microarray experiments from many sources. The analysis reveals that part of the Arabidopsis transcriptome is globally co-expressed, and can be further divided into known as well as novel functional gene modules. Our methodology is general enough to apply to any set of microarray experiments, using any scoring function.

  3. Transcriptional profiling in response to terminal drought stress reveals differential responses along the wheat genome

    Directory of Open Access Journals (Sweden)

    Ferrari Francesco

    2009-06-01

    Full Text Available Abstract Background Water stress during grain filling has a marked effect on grain yield, leading to a reduced endosperm cell number and thus sink capacity to accumulate dry matter. The bread wheat cultivar Chinese Spring (CS, a Chinese Spring terminal deletion line (CS_5AL-10 and the durum wheat cultivar Creso were subjected to transcriptional profiling after exposure to mild and severe drought stress at the grain filling stage to find evidences of differential stress responses associated to different wheat genome regions. Results The transcriptome analysis of Creso, CS and its deletion line revealed 8,552 non redundant probe sets with different expression levels, mainly due to the comparisons between the two species. The drought treatments modified the expression of 3,056 probe sets. Besides a set of genes showing a similar drought response in Creso and CS, cluster analysis revealed several drought response features that can be associated to the different genomic structure of Creso, CS and CS_5AL-10. Some drought-related genes were expressed at lower level (or not expressed in Creso (which lacks the D genome or in the CS_5AL-10 deletion line compared to CS. The chromosome location of a set of these genes was confirmed by PCR-based mapping on the D genome (or the 5AL-10 region. Many clusters were characterized by different level of expression in Creso, CS and CS_AL-10, suggesting that the different genome organization of the three genotypes may affect plant adaptation to stress. Clusters with similar expression trend were grouped and functional classified to mine the biological mean of their activation or repression. Genes involved in ABA, proline, glycine-betaine and sorbitol pathways were found up-regulated by drought stress. Furthermore, the enhanced expression of a set of transposons and retrotransposons was detected in CS_5AL-10. Conclusion Bread and durum wheat genotypes were characterized by a different physiological reaction to water

  4. Spatiotemporal network motif reveals the biological traits of developmental gene regulatory networks in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Kim Man-Sun

    2012-05-01

    Full Text Available Abstract Background Network motifs provided a “conceptual tool” for understanding the functional principles of biological networks, but such motifs have primarily been used to consider static network structures. Static networks, however, cannot be used to reveal time- and region-specific traits of biological systems. To overcome this limitation, we proposed the concept of a “spatiotemporal network motif,” a spatiotemporal sequence of network motifs of sub-networks which are active only at specific time points and body parts. Results On the basis of this concept, we analyzed the developmental gene regulatory network of the Drosophila melanogaster embryo. We identified spatiotemporal network motifs and investigated their distribution pattern in time and space. As a result, we found how key developmental processes are temporally and spatially regulated by the gene network. In particular, we found that nested feedback loops appeared frequently throughout the entire developmental process. From mathematical simulations, we found that mutual inhibition in the nested feedback loops contributes to the formation of spatial expression patterns. Conclusions Taken together, the proposed concept and the simulations can be used to unravel the design principle of developmental gene regulatory networks.

  5. Validation of Skeletal Muscle cis-Regulatory Module Predictions Reveals Nucleotide Composition Bias in Functional Enhancers

    Science.gov (United States)

    Kwon, Andrew T.; Chou, Alice Yi; Arenillas, David J.; Wasserman, Wyeth W.

    2011-01-01

    We performed a genome-wide scan for muscle-specific cis-regulatory modules (CRMs) using three computational prediction programs. Based on the predictions, 339 candidate CRMs were tested in cell culture with NIH3T3 fibroblasts and C2C12 myoblasts for capacity to direct selective reporter gene expression to differentiated C2C12 myotubes. A subset of 19 CRMs validated as functional in the assay. The rate of predictive success reveals striking limitations of computational regulatory sequence analysis methods for CRM discovery. Motif-based methods performed no better than predictions based only on sequence conservation. Analysis of the properties of the functional sequences relative to inactive sequences identifies nucleotide sequence composition can be an important characteristic to incorporate in future methods for improved predictive specificity. Muscle-related TFBSs predicted within the functional sequences display greater sequence conservation than non-TFBS flanking regions. Comparison with recent MyoD and histone modification ChIP-Seq data supports the validity of the functional regions. PMID:22144875

  6. Validation of skeletal muscle cis-regulatory module predictions reveals nucleotide composition bias in functional enhancers.

    Directory of Open Access Journals (Sweden)

    Andrew T Kwon

    2011-12-01

    Full Text Available We performed a genome-wide scan for muscle-specific cis-regulatory modules (CRMs using three computational prediction programs. Based on the predictions, 339 candidate CRMs were tested in cell culture with NIH3T3 fibroblasts and C2C12 myoblasts for capacity to direct selective reporter gene expression to differentiated C2C12 myotubes. A subset of 19 CRMs validated as functional in the assay. The rate of predictive success reveals striking limitations of computational regulatory sequence analysis methods for CRM discovery. Motif-based methods performed no better than predictions based only on sequence conservation. Analysis of the properties of the functional sequences relative to inactive sequences identifies nucleotide sequence composition can be an important characteristic to incorporate in future methods for improved predictive specificity. Muscle-related TFBSs predicted within the functional sequences display greater sequence conservation than non-TFBS flanking regions. Comparison with recent MyoD and histone modification ChIP-Seq data supports the validity of the functional regions.

  7. Transcript Profile of Flowering Regulatory Genes in VcFT-Overexpressing Blueberry Plants.

    Directory of Open Access Journals (Sweden)

    Aaron E Walworth

    Full Text Available In order to identify genetic components in flowering pathways of highbush blueberry (Vaccinium corymbosum L., a transcriptome reference composed of 254,396 transcripts and 179,853 gene contigs was developed by assembly of 72.7 million reads using Trinity. Using this transcriptome reference and a query of flowering pathway genes of herbaceous plants, we identified potential flowering pathway genes/transcripts of blueberry. Transcriptome analysis of flowering pathway genes was then conducted on leaf tissue samples of transgenic blueberry cv. Aurora ('VcFT-Aurora', which overexpresses a blueberry FLOWERING LOCUS T-like gene (VcFT. Sixty-one blueberry transcripts of 40 genes showed high similarities to 33 known flowering-related genes of herbaceous plants, of which 17 down-regulated and 16 up-regulated genes were identified in 'VcFT-Aurora'. All down-regulated genes encoded transcription factors/enzymes upstream in the signaling pathway containing VcFT. A blueberry CONSTANS-LIKE 5-like (VcCOL5 gene was down-regulated and associated with five other differentially expressed (DE genes in the photoperiod-mediated flowering pathway. Three down-regulated genes, i.e., a MADS-AFFECTING FLOWERING 2-like gene (VcMAF2, a MADS-AFFECTING FLOWERING 5-like gene (VcMAF5, and a VERNALIZATION1-like gene (VcVRN1, may function as integrators in place of FLOWERING LOCUS C (FLC in the vernalization pathway. Because no CONSTAN1-like or FLOWERING LOCUS C-like genes were found in blueberry, VcCOL5 and VcMAF2/VcMAF5 or VRN1 might be the major integrator(s in the photoperiod- and vernalization-mediated flowering pathway, respectively. The major down-stream genes of VcFT, i.e., SUPPRESSOR of Overexpression of Constans 1-like (VcSOC1, LEAFY-like (VcLFY, APETALA1-like (VcAP1, CAULIFLOWER 1-like (VcCAL1, and FRUITFULL-like (VcFUL genes were present and showed high similarity to their orthologues in herbaceous plants. Moreover, overexpression of VcFT promoted expression of all of

  8. Transcript Profile of Flowering Regulatory Genes in VcFT-Overexpressing Blueberry Plants

    Science.gov (United States)

    Walworth, Aaron E.; Chai, Benli; Song, Guo-qing

    2016-01-01

    In order to identify genetic components in flowering pathways of highbush blueberry (Vaccinium corymbosum L.), a transcriptome reference composed of 254,396 transcripts and 179,853 gene contigs was developed by assembly of 72.7 million reads using Trinity. Using this transcriptome reference and a query of flowering pathway genes of herbaceous plants, we identified potential flowering pathway genes/transcripts of blueberry. Transcriptome analysis of flowering pathway genes was then conducted on leaf tissue samples of transgenic blueberry cv. Aurora (‘VcFT-Aurora’), which overexpresses a blueberry FLOWERING LOCUS T-like gene (VcFT). Sixty-one blueberry transcripts of 40 genes showed high similarities to 33 known flowering-related genes of herbaceous plants, of which 17 down-regulated and 16 up-regulated genes were identified in ‘VcFT-Aurora’. All down-regulated genes encoded transcription factors/enzymes upstream in the signaling pathway containing VcFT. A blueberry CONSTANS-LIKE 5-like (VcCOL5) gene was down-regulated and associated with five other differentially expressed (DE) genes in the photoperiod-mediated flowering pathway. Three down-regulated genes, i.e., a MADS-AFFECTING FLOWERING 2-like gene (VcMAF2), a MADS-AFFECTING FLOWERING 5-like gene (VcMAF5), and a VERNALIZATION1-like gene (VcVRN1), may function as integrators in place of FLOWERING LOCUS C (FLC) in the vernalization pathway. Because no CONSTAN1-like or FLOWERING LOCUS C-like genes were found in blueberry, VcCOL5 and VcMAF2/VcMAF5 or VRN1 might be the major integrator(s) in the photoperiod- and vernalization-mediated flowering pathway, respectively. The major down-stream genes of VcFT, i.e., SUPPRESSOR of Overexpression of Constans 1-like (VcSOC1), LEAFY-like (VcLFY), APETALA1-like (VcAP1), CAULIFLOWER 1-like (VcCAL1), and FRUITFULL-like (VcFUL) genes were present and showed high similarity to their orthologues in herbaceous plants. Moreover, overexpression of VcFT promoted expression of all

  9. Transcript Profile of Flowering Regulatory Genes in VcFT-Overexpressing Blueberry Plants.

    Science.gov (United States)

    Walworth, Aaron E; Chai, Benli; Song, Guo-Qing

    2016-01-01

    In order to identify genetic components in flowering pathways of highbush blueberry (Vaccinium corymbosum L.), a transcriptome reference composed of 254,396 transcripts and 179,853 gene contigs was developed by assembly of 72.7 million reads using Trinity. Using this transcriptome reference and a query of flowering pathway genes of herbaceous plants, we identified potential flowering pathway genes/transcripts of blueberry. Transcriptome analysis of flowering pathway genes was then conducted on leaf tissue samples of transgenic blueberry cv. Aurora ('VcFT-Aurora'), which overexpresses a blueberry FLOWERING LOCUS T-like gene (VcFT). Sixty-one blueberry transcripts of 40 genes showed high similarities to 33 known flowering-related genes of herbaceous plants, of which 17 down-regulated and 16 up-regulated genes were identified in 'VcFT-Aurora'. All down-regulated genes encoded transcription factors/enzymes upstream in the signaling pathway containing VcFT. A blueberry CONSTANS-LIKE 5-like (VcCOL5) gene was down-regulated and associated with five other differentially expressed (DE) genes in the photoperiod-mediated flowering pathway. Three down-regulated genes, i.e., a MADS-AFFECTING FLOWERING 2-like gene (VcMAF2), a MADS-AFFECTING FLOWERING 5-like gene (VcMAF5), and a VERNALIZATION1-like gene (VcVRN1), may function as integrators in place of FLOWERING LOCUS C (FLC) in the vernalization pathway. Because no CONSTAN1-like or FLOWERING LOCUS C-like genes were found in blueberry, VcCOL5 and VcMAF2/VcMAF5 or VRN1 might be the major integrator(s) in the photoperiod- and vernalization-mediated flowering pathway, respectively. The major down-stream genes of VcFT, i.e., SUPPRESSOR of Overexpression of Constans 1-like (VcSOC1), LEAFY-like (VcLFY), APETALA1-like (VcAP1), CAULIFLOWER 1-like (VcCAL1), and FRUITFULL-like (VcFUL) genes were present and showed high similarity to their orthologues in herbaceous plants. Moreover, overexpression of VcFT promoted expression of all of these

  10. Evolution of Transcriptional Regulatory Networks in Pseudomonas aeruginosa During Long Time Growth in Human Hosts

    DEFF Research Database (Denmark)

    Andresen, Eva Kammer

    extent these observations relate to natural microbial populations. The focus of this thesis has been to study how regulatory networks evolve in natural systems. By using a particular infectious disease scenario (human associated persistent airway infections caused by the bacterium Pseudomonas aeruginosa...... in global regulator genes facilitate the generation of novel phenotypes which again facilitate the shift in life-style of the bacterium from an environmental opportunistic pathogen to a human airway specific pathogen. These findings are not only applicable to P. aeruginosa specific studies, but suggest that...

  11. Post-Transcriptional Regulation by the Csr Global Regulatory System in Escherichia coli

    OpenAIRE

    Suzuki, Kazushi; 鈴木, 一史

    2007-01-01

    In many species of bacteria, the Csr (carbon storage regulator) global regulatory system coordinates the expression of various genes. In Escherichia coli, the central component of this system, CsrA, is a RNA-binding protein. The CsrA is a homodimer and binds to leader segments of target mRNAs, affecting their translation and stability. CsrA activity is regulated by two small non-coding RNAs, CsrB and CsrC. These RNAs contain multiple CsrA-binding sequences and act by sequestering CsrA. In thi...

  12. Integrative analysis of RNA, translation, and protein levels reveals distinct regulatory variation across humans.

    Science.gov (United States)

    Cenik, Can; Cenik, Elif Sarinay; Byeon, Gun W; Grubert, Fabian; Candille, Sophie I; Spacek, Damek; Alsallakh, Bilal; Tilgner, Hagen; Araya, Carlos L; Tang, Hua; Ricci, Emiliano; Snyder, Michael P

    2015-11-01

    Elucidating the consequences of genetic differences between humans is essential for understanding phenotypic diversity and personalized medicine. Although variation in RNA levels, transcription factor binding, and chromatin have been explored, little is known about global variation in translation and its genetic determinants. We used ribosome profiling, RNA sequencing, and mass spectrometry to perform an integrated analysis in lymphoblastoid cell lines from a diverse group of individuals. We find significant differences in RNA, translation, and protein levels suggesting diverse mechanisms of personalized gene expression control. Combined analysis of RNA expression and ribosome occupancy improves the identification of individual protein level differences. Finally, we identify genetic differences that specifically modulate ribosome occupancy--many of these differences lie close to start codons and upstream ORFs. Our results reveal a new level of gene expression variation among humans and indicate that genetic variants can cause changes in protein levels through effects on translation. © 2015 Cenik et al.; Published by Cold Spring Harbor Laboratory Press.

  13. A model for genetic and epigenetic regulatory networks identifies rare pathways for transcription factor induced pluripotency

    Science.gov (United States)

    Artyomov, Maxim; Meissner, Alex; Chakraborty, Arup

    2010-03-01

    Most cells in an organism have the same DNA. Yet, different cell types express different proteins and carry out different functions. This is because of epigenetic differences; i.e., DNA in different cell types is packaged distinctly, making it hard to express certain genes while facilitating the expression of others. During development, upon receipt of appropriate cues, pluripotent embryonic stem cells differentiate into diverse cell types that make up the organism (e.g., a human). There has long been an effort to make this process go backward -- i.e., reprogram a differentiated cell (e.g., a skin cell) to pluripotent status. Recently, this has been achieved by transfecting certain transcription factors into differentiated cells. This method does not use embryonic material and promises the development of patient-specific regenerative medicine, but it is inefficient. The mechanisms that make reprogramming rare, or even possible, are poorly understood. We have developed the first computational model of transcription factor-induced reprogramming. Results obtained from the model are consistent with diverse observations, and identify the rare pathways that allow reprogramming to occur. If validated, our model could be further developed to design optimal strategies for reprogramming and shed light on basic questions in biology.

  14. Differential 3’ processing of specific transcripts expands regulatory and protein diversity across neuronal cell types

    Science.gov (United States)

    Jereb, Saša; Hwang, Hun-Way; Van Otterloo, Eric; Govek, Eve-Ellen; Fak, John J; Yuan, Yuan; Hatten, Mary E

    2018-01-01

    Alternative polyadenylation (APA) regulates mRNA translation, stability, and protein localization. However, it is unclear to what extent APA regulates these processes uniquely in specific cell types. Using a new technique, cTag-PAPERCLIP, we discovered significant differences in APA between the principal types of mouse cerebellar neurons, the Purkinje and granule cells, as well as between proliferating and differentiated granule cells. Transcripts that differed in APA in these comparisons were enriched in key neuronal functions and many differed in coding sequence in addition to 3’UTR length. We characterize Memo1, a transcript that shifted from expressing a short 3’UTR isoform to a longer one during granule cell differentiation. We show that Memo1 regulates granule cell precursor proliferation and that its long 3’UTR isoform is targeted by miR-124, contributing to its downregulation during development. Our findings provide insight into roles for APA in specific cell types and establish a platform for further functional studies. PMID:29578408

  15. Different papillomaviruses have different repertoires of transcription factor binding sites: convergence and divergence in the upstream regulatory region

    Directory of Open Access Journals (Sweden)

    Alonso Ángel

    2006-03-01

    Full Text Available Abstract Background Papillomaviruses (PVs infect stratified squamous epithelia in warm-blooded vertebrates and have undergone a complex evolutionary process. The control of the expression of the early ORFs in PVs depends on the binding of cellular and viral transcription factors to the upstream regulatory region (URR of the virus. It is believed that there is a core of transcription factor binding sites (TFBS common to all PVs, with additional individual differences, although most of the available information focuses only on a handful of viruses. Results We have studied the URR of sixty-one PVs, covering twenty different hosts. We have predicted the TFBS present in the URR and analysed these results by principal component analysis and genetic algorithms. The number and nature of TFBS in the URR might be much broader than thus far described, and different PVs have different repertoires of TFBS. Conclusion There are common fingerprints in the URR in PVs that infect primates, although the ancestors of these viruses diverged a long time ago. Additionally, there are obvious differences between the URR of alpha and beta PVs, despite these PVs infect similar histological cell types in the same host, i.e. human. A thorough analysis of the TFBS in the URR might provide crucial information about the differential biology of cancer-associated PVs.

  16. Expression Pattern of Myogenic Regulatory Transcription Factor mRNAs in the Embryo and Adult Labeo rohita (Hamilton, 1822

    Directory of Open Access Journals (Sweden)

    Archya Sengupta

    2014-01-01

    Full Text Available Understanding the regulation of skeletal muscle development is important to meet the increasing demand of Indian major carp Labeo rohita. Myogenic regulatory factors (MRFs along with myocyte specific enhancer factor 2 (MEF2 play the pivotal role in the determination and differentiation of skeletal muscle. The majority of skeletal muscle genes require both MRFs and MEF2 family members to activate their transcription. In this study, the expression pattern of MyoD, myf-5, myogenin, and MEF2A was observed from 6 h after fertilization to 12 months of age using semiquantitative RT-PCR as well as real-time PCR method. MyoD and myf-5 mRNAs were expressed at high level at the early embryonic stages. Myogenin and MEF2A were expressed after MyoD and myf-5 and remained active up to adult stage. Expression of MyoD was lower than that of Myf-5 after the 5th month. Partial sequencing of MyoD, myf-5, and MEF2A was done to draw phylogeny. In phylogenetic study, Labeo MyoD, MEF2A and myf-5 were found to be closely related to those of common carp. The present investigation suggests that the four transcription factors play pivotal role in the regulation of muscle growth of Labeo rohita in an overlapping and interconnected way.

  17. Genome-wide mRNA processing in methanogenic archaea reveals post-transcriptional regulation of ribosomal protein synthesis.

    Science.gov (United States)

    Qi, Lei; Yue, Lei; Feng, Deqin; Qi, Fengxia; Li, Jie; Dong, Xiuzhu

    2017-07-07

    Unlike stable RNAs that require processing for maturation, prokaryotic cellular mRNAs generally follow an 'all-or-none' pattern. Herein, we used a 5΄ monophosphate transcript sequencing (5΄P-seq) that specifically captured the 5΄-end of processed transcripts and mapped the genome-wide RNA processing sites (PSSs) in a methanogenic archaeon. Following statistical analysis and stringent filtration, we identified 1429 PSSs, among which 23.5% and 5.4% were located in 5΄ untranslated region (uPSS) and intergenic region (iPSS), respectively. A predominant uridine downstream PSSs served as a processing signature. Remarkably, 5΄P-seq detected overrepresented uPSS and iPSS in the polycistronic operons encoding ribosomal proteins, and the majority upstream and proximal ribosome binding sites, suggesting a regulatory role of processing on translation initiation. The processed transcripts showed increased stability and translation efficiency. Particularly, processing within the tricistronic transcript of rplA-rplJ-rplL enhanced the translation of rplL, which can provide a driving force for the 1:4 stoichiometry of L10 to L12 in the ribosome. Growth-associated mRNA processing intensities were also correlated with the cellular ribosomal protein levels, thereby suggesting that mRNA processing is involved in tuning growth-dependent ribosome synthesis. In conclusion, our findings suggest that mRNA processing-mediated post-transcriptional regulation is a potential mechanism of ribosomal protein synthesis and stoichiometry. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  18. Transcriptional regulatory network triggered by oxidative signals configures the early response mechanisms of japonica rice to chilling stress

    Directory of Open Access Journals (Sweden)

    Wijaya Edward

    2010-01-01

    Full Text Available Abstract Background The transcriptional regulatory network involved in low temperature response leading to acclimation has been established in Arabidopsis. In japonica rice, which can only withstand transient exposure to milder cold stress (10°C, an oxidative-mediated network has been proposed to play a key role in configuring early responses and short-term defenses. The components, hierarchical organization and physiological consequences of this network were further dissected by a systems-level approach. Results Regulatory clusters responding directly to oxidative signals were prominent during the initial 6 to 12 hours at 10°C. Early events mirrored a typical oxidative response based on striking similarities of the transcriptome to disease, elicitor and wounding induced processes. Targets of oxidative-mediated mechanisms are likely regulated by several classes of bZIP factors acting on as1/ocs/TGA-like element enriched clusters, ERF factors acting on GCC-box/JAre-like element enriched clusters and R2R3-MYB factors acting on MYB2-like element enriched clusters. Temporal induction of several H2O2-induced bZIP, ERF and MYB genes coincided with the transient H2O2 spikes within the initial 6 to 12 hours. Oxidative-independent responses involve DREB/CBF, RAP2 and RAV1 factors acting on DRE/CRT/rav1-like enriched clusters and bZIP factors acting on ABRE-like enriched clusters. Oxidative-mediated clusters were activated earlier than ABA-mediated clusters. Conclusion Genome-wide, physiological and whole-plant level analyses established a holistic view of chilling stress response mechanism of japonica rice. Early response regulatory network triggered by oxidative signals is critical for prolonged survival under sub-optimal temperature. Integration of stress and developmental responses leads to modulated growth and vigor maintenance contributing to a delay of plastic injuries.

  19. Transcriptional regulatory network triggered by oxidative signals configures the early response mechanisms of japonica rice to chilling stress

    KAUST Repository

    Yun, Kil-Young

    2010-01-25

    Background: The transcriptional regulatory network involved in low temperature response leading to acclimation has been established in Arabidopsis. In japonica rice, which can only withstand transient exposure to milder cold stress (10C), an oxidative-mediated network has been proposed to play a key role in configuring early responses and short-term defenses. The components, hierarchical organization and physiological consequences of this network were further dissected by a systems-level approach.Results: Regulatory clusters responding directly to oxidative signals were prominent during the initial 6 to 12 hours at 10C. Early events mirrored a typical oxidative response based on striking similarities of the transcriptome to disease, elicitor and wounding induced processes. Targets of oxidative-mediated mechanisms are likely regulated by several classes of bZIP factors acting on as1/ocs/TGA-like element enriched clusters, ERF factors acting on GCC-box/JAre-like element enriched clusters and R2R3-MYB factors acting on MYB2-like element enriched clusters.Temporal induction of several H2O2-induced bZIP, ERF and MYB genes coincided with the transient H2O2spikes within the initial 6 to 12 hours. Oxidative-independent responses involve DREB/CBF, RAP2 and RAV1 factors acting on DRE/CRT/rav1-like enriched clusters and bZIP factors acting on ABRE-like enriched clusters. Oxidative-mediated clusters were activated earlier than ABA-mediated clusters.Conclusion: Genome-wide, physiological and whole-plant level analyses established a holistic view of chilling stress response mechanism of japonica rice. Early response regulatory network triggered by oxidative signals is critical for prolonged survival under sub-optimal temperature. Integration of stress and developmental responses leads to modulated growth and vigor maintenance contributing to a delay of plastic injuries. 2010 Yun et al; licensee BioMed Central Ltd.

  20. Deep RNA sequencing reveals hidden features and dynamics of early gene transcription in Paramecium bursaria chlorella virus 1.

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    Guillaume Blanc

    Full Text Available Paramecium bursaria chlorella virus 1 (PBCV-1 is the prototype of the genus Chlorovirus (family Phycodnaviridae that infects the unicellular, eukaryotic green alga Chlorella variabilis NC64A. The 331-kb PBCV-1 genome contains 416 major open reading frames. A mRNA-seq approach was used to analyze PBCV-1 transcriptomes at 6 progressive times during the first hour of infection. The alignment of 17 million reads to the PBCV-1 genome allowed the construction of single-base transcriptome maps. Significant transcription was detected for a subset of 50 viral genes as soon as 7 min after infection. By 20 min post infection (p.i., transcripts were detected for most PBCV-1 genes and transcript levels continued to increase globally up to 60 min p.i., at which time 41% or the poly (A+-containing RNAs in the infected cells mapped to the PBCV-1 genome. For some viral genes, the number of transcripts in the latter time points (20 to 60 min p.i. was much higher than that of the most highly expressed host genes. RNA-seq data revealed putative polyadenylation signal sequences in PBCV-1 genes that were identical to the polyadenylation signal AAUAAA of green algae. Several transcripts have an RNA fragment excised. However, the frequency of excision and the resulting putative shortened protein products suggest that most of these excision events have no functional role but are probably the result of the activity of misled splicesomes.

  1. Analysis of the highly diverse gene borders in Ebola virus reveals a distinct mechanism of transcriptional regulation.

    Science.gov (United States)

    Brauburger, Kristina; Boehmann, Yannik; Tsuda, Yoshimi; Hoenen, Thomas; Olejnik, Judith; Schümann, Michael; Ebihara, Hideki; Mühlberger, Elke

    2014-11-01

    the regulatory role of the structurally unique EBOV gene borders during viral transcription. Our data suggest that transcriptional regulation in EBOV is highly complex and differs from that in prototype viruses and further the understanding of this most fundamental process in the filovirus replication cycle. Moreover, our results with recombinant EBOVs suggest a novel role of the long IR found in all filovirus genomes during the viral replication cycle. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  2. Reverse engineering a mouse embryonic stem cell-specific transcriptional network reveals a new modulator of neuronal differentiation.

    Science.gov (United States)

    De Cegli, Rossella; Iacobacci, Simona; Flore, Gemma; Gambardella, Gennaro; Mao, Lei; Cutillo, Luisa; Lauria, Mario; Klose, Joachim; Illingworth, Elizabeth; Banfi, Sandro; di Bernardo, Diego

    2013-01-01

    Gene expression profiles can be used to infer previously unknown transcriptional regulatory interaction among thousands of genes, via systems biology 'reverse engineering' approaches. We 'reverse engineered' an embryonic stem (ES)-specific transcriptional network from 171 gene expression profiles, measured in ES cells, to identify master regulators of gene expression ('hubs'). We discovered that E130012A19Rik (E13), highly expressed in mouse ES cells as compared with differentiated cells, was a central 'hub' of the network. We demonstrated that E13 is a protein-coding gene implicated in regulating the commitment towards the different neuronal subtypes and glia cells. The overexpression and knock-down of E13 in ES cell lines, undergoing differentiation into neurons and glia cells, caused a strong up-regulation of the glutamatergic neurons marker Vglut2 and a strong down-regulation of the GABAergic neurons marker GAD65 and of the radial glia marker Blbp. We confirmed E13 expression in the cerebral cortex of adult mice and during development. By immuno-based affinity purification, we characterized protein partners of E13, involved in the Polycomb complex. Our results suggest a role of E13 in regulating the division between glutamatergic projection neurons and GABAergic interneurons and glia cells possibly by epigenetic-mediated transcriptional regulation.

  3. Single-cell Transcriptional Analysis Reveals Novel Neuronal Phenotypes and Interaction Networks involved In the Central Circadian Clock

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    James Park

    2016-10-01

    Full Text Available Single-cell heterogeneity confounds efforts to understand how a population of cells organizes into cellular networks that underlie tissue-level function. This complexity is prominent in the mammalian suprachiasmatic nucleus (SCN. Here, individual neurons exhibit a remarkable amount of asynchronous behavior and transcriptional heterogeneity. However, SCN neurons are able to generate precisely coordinated synaptic and molecular outputs that synchronize the body to a common circadian cycle by organizing into cellular networks. To understand this emergent cellular network property, it is important to reconcile single-neuron heterogeneity with network organization. In light of recent studies suggesting that transcriptionally heterogeneous cells organize into distinct cellular phenotypes, we characterized the transcriptional, spatial, and functional organization of 352 SCN neurons from mice experiencing phase-shifts in their circadian cycle. Using the community structure detection method and multivariate analytical techniques, we identified previously undescribed neuronal phenotypes that are likely to participate in regulatory networks with known SCN cell types. Based on the newly discovered neuronal phenotypes, we developed a data-driven neuronal network structure in which multiple cell types interact through known synaptic and paracrine signaling mechanisms. These results provide a basis from which to interpret the functional variability of SCN neurons and describe methodologies towards understanding how a population of heterogeneous single cells organizes into cellular networks that underlie tissue-level function.

  4. Transcriptome profiling of a curdlan-producing Agrobacterium reveals conserved regulatory mechanisms of exopolysaccharide biosynthesis

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    Ruffing Anne M

    2012-02-01

    Full Text Available Abstract Background The ability to synthesize exopolysaccharides (EPS is widespread among microorganisms, and microbial EPS play important roles in biofilm formation, pathogen persistence, and applications in the food and medical industries. Although it is well established that EPS synthesis is invariably in response to environmental cues, it remains largely unknown how various environmental signals trigger activation of the biochemical synthesis machinery. Results We report here the transcriptome profiling of Agrobacterium sp. ATCC 31749, a microorganism that produces large amounts of a glucose polymer known as curdlan under nitrogen starvation. Transcriptome analysis revealed a nearly 100-fold upregulation of the curdlan synthesis operon upon transition to nitrogen starvation, thus establishing the prominent role that transcriptional regulation plays in the EPS synthesis. In addition to known mechanisms of EPS regulation such as activation by c-di-GMP, we identify novel mechanisms of regulation in ATCC 31749, including RpoN-independent NtrC regulation and intracellular pH regulation by acidocalcisomes. Furthermore, we show evidence that curdlan synthesis is also regulated by conserved cell stress responses, including polyphosphate accumulation and the stringent response. In fact, the stringent response signal, pppGpp, appears to be indispensible for transcriptional activation of curdlan biosynthesis. Conclusions This study identifies several mechanisms regulating the synthesis of curdlan, an EPS with numerous applications. These mechanisms are potential metabolic engineering targets for improving the industrial production of curdlan from Agrobacterium sp. ATCC 31749. Furthermore, many of the genes identified in this study are highly conserved across microbial genomes, and we propose that the molecular elements identified in this study may serve as universal regulators of microbial EPS synthesis.

  5. Transcriptional regulation of the outer membrane porin gene ompW reveals its physiological role during the transition from the aerobic to the anaerobic lifestyle of Escherichia coli

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    Minfeng eXiao

    2016-05-01

    Full Text Available Understanding bacterial physiology relies on elucidating the regulatory mechanisms and cellular functions of those differentially expressed genes in response to environmental changes. A widespread Gram-negative bacterial outer membrane protein OmpW has been implicated in the adaptation to stresses in various species. It is recently found to be present in the regulon of the global anaerobic transcription factor FNR and ArcA in E. coli. However, little is known about the physiological implications of this regulatory disposition. In this study, we demonstrate that transcription of ompW is indeed mediated by a series of global regulators involved in the anaerobiosis of E. coli. We show that FNR can both activate and repress the expression of ompW through its direct binding to two distinctive sites, -81.5 and -126.5 bp respectively, on ompW promoter. ArcA also participates in repression of ompW under anaerobic condition, but in an FNR dependent manner. Additionally, ompW is also subject to the regulation by CRP and NarL which senses the availability and types of carbon sources and respiration electron acceptors in the environment respectively, implying a role of OmpW in the carbon and energy metabolism of E. coli during its anaerobic adaptation. Molecular docking reveals that OmpW can bind fumarate, an alternative electron acceptor in anaerobic respiration, with sufficient affinity. Moreover, supplement of fumarate or succinate which belongs to the C4-dicarboxylates family of metabolite, to E. coli culture rescues OmpW-mediated colicin S4 killing. Taken together, we propose that OmpW is involved in anaerobic carbon and energy metabolism to mediate the transition from aerobic to anaerobic lifestyle in E. coli.

  6. A minimal murine Msx-1 gene promoter. Organization of its cis-regulatory motifs and their role in transcriptional activation in cells in culture and in transgenic mice.

    Science.gov (United States)

    Takahashi, T; Guron, C; Shetty, S; Matsui, H; Raghow, R

    1997-09-05

    To dissect the cis-regulatory elements of the murine Msx-1 promoter, which lacks a conventional TATA element, a putative Msx-1 promoter DNA fragment (from -1282 to +106 base pairs (bp)) or its congeners containing site-specific alterations were fused to luciferase reporter and introduced into NIH3T3 and C2C12 cells, and the expression of luciferase was assessed in transient expression assays. The functional consequences of the sequential 5' deletions of the promotor revealed that multiple positive and negative regulatory elements participate in regulating transcription of the Msx-1 gene. Surprisingly, however, the optimal expression of Msx-1 promoter in either NIH3T3 or C2C12 cells required only 165 bp of the upstream sequence to warrant detailed examination of its structure. Therefore, the functional consequences of site-specific deletions and point mutations of the cis-acting elements of the minimal Msx-1 promoter were systematically examined. Concomitantly, potential transcriptional factor(s) interacting with the cis-acting elements of the minimal promoter were also studied by gel electrophoretic mobility shift assays and DNase I footprinting. Combined analyses of the minimal promoter by DNase I footprinting, electrophoretic mobility shift assays, and super shift assays with specific antibodies revealed that 5'-flanking regions from -161 to -154 and from -26 to -13 of the Msx-1 promoter contains an authentic E box (proximal E box), capable of binding a protein immunologically related to the upstream stimulating factor 1 (USF-1) and a GC-rich sequence motif which can bind to Sp1 (proximal Sp1), respectively. Additionally, we observed that the promoter activation was seriously hampered if the proximal E box was removed or mutated, and the promoter activity was eliminated completely if the proximal Sp1 site was similarly altered. Absolute dependence of the Msx-1 minimal promoter on Sp1 could be demonstrated by transient expression assays in the Sp1-deficient

  7. The Transcriptional and Gene Regulatory Network of Lactococcus lactis MG1363 during Growth in Milk

    NARCIS (Netherlands)

    de Jong, Anne; Hansen, Morten E.; Kuipers, Oscar P.; Kilstrup, Mogens; Kok, Jan

    2013-01-01

    In the present study we examine the changes in the expression of genes of Lactococcus lactis subspecies cremoris MG1363 during growth in milk. To reveal which specific classes of genes (pathways, operons, regulons, COGs) are important, we performed a transcriptome time series experiment. Global

  8. Structures of RNA Polymerase Closed and Intermediate Complexes Reveal Mechanisms of DNA Opening and Transcription Initiation.

    Science.gov (United States)

    Glyde, Robert; Ye, Fuzhou; Darbari, Vidya Chandran; Zhang, Nan; Buck, Martin; Zhang, Xiaodong

    2017-07-06

    Gene transcription is carried out by RNA polymerases (RNAPs). For transcription to occur, the closed promoter complex (RPc), where DNA is double stranded, must isomerize into an open promoter complex (RPo), where the DNA is melted out into a transcription bubble and the single-stranded template DNA is delivered to the RNAP active site. Using a bacterial RNAP containing the alternative σ 54 factor and cryoelectron microscopy, we determined structures of RPc and the activator-bound intermediate complex en route to RPo at 3.8 and 5.8 Å. Our structures show how RNAP-σ 54 interacts with promoter DNA to initiate the DNA distortions required for transcription bubble formation, and how the activator interacts with RPc, leading to significant conformational changes in RNAP and σ 54 that promote RPo formation. We propose that DNA melting is an active process initiated in RPc and that the RNAP conformations of intermediates are significantly different from that of RPc and RPo. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  9. A tobacco cDNA reveals two different transcription patterns in vegetative and reproductive organs

    Directory of Open Access Journals (Sweden)

    I. da Silva

    2002-08-01

    Full Text Available In order to identify genes expressed in the pistil that may have a role in the reproduction process, we have established an expressed sequence tags project to randomly sequence clones from a Nicotiana tabacum stigma/style cDNA library. A cDNA clone (MTL-8 showing high sequence similarity to genes encoding glycine-rich RNA-binding proteins was chosen for further characterization. Based on the extensive identity of MTL-8 to the RGP-1a sequence of N. sylvestris, a primer was defined to extend the 5' sequence of MTL-8 by RT-PCR from stigma/style RNAs. The amplification product was sequenced and it was confirmed that MTL-8 corresponds to an mRNA encoding a glycine-rich RNA-binding protein. Two transcripts of different sizes and expression patterns were identified when the MTL-8 cDNA insert was used as a probe in RNA blots. The largest is 1,100 nucleotides (nt long and markedly predominant in ovaries. The smaller transcript, with 600 nt, is ubiquitous to the vegetative and reproductive organs analyzed (roots, stems, leaves, sepals, petals, stamens, stigmas/styles and ovaries. Plants submitted to stress (wounding, virus infection and ethylene treatment presented an increased level of the 600-nt transcript in leaves, especially after tobacco necrosis virus infection. In contrast, the level of the 1,100-nt transcript seems to be unaffected by the stress conditions tested. Results of Southern blot experiments have suggested that MTL-8 is present in one or two copies in the tobacco genome. Our results suggest that the shorter transcript is related to stress while the larger one is a flower predominant and nonstress-inducible messenger.

  10. Transcriptional similarity in couples reveals the impact of shared environment and lifestyle on gene regulation through modified cytosines

    Directory of Open Access Journals (Sweden)

    Ke Tang

    2016-06-01

    Full Text Available Gene expression is a complex and quantitative trait that is influenced by both genetic and non-genetic regulators including environmental factors. Evaluating the contribution of environment to gene expression regulation and identifying which genes are more likely to be influenced by environmental factors are important for understanding human complex traits. We hypothesize that by living together as couples, there can be commonly co-regulated genes that may reflect the shared living environment (e.g., diet, indoor air pollutants, behavioral lifestyle. The lymphoblastoid cell lines (LCLs derived from unrelated couples of African ancestry (YRI, Yoruba people from Ibadan, Nigeria from the International HapMap Project provided a unique model for us to characterize gene expression pattern in couples by comparing gene expression levels between husbands and wives. Strikingly, 778 genes were found to show much smaller variances in couples than random pairs of individuals at a false discovery rate (FDR of 5%. Since genetic variation between unrelated family members in a general population is expected to be the same assuming a random-mating society, non-genetic factors (e.g., epigenetic systems are more likely to be the mediators for the observed transcriptional similarity in couples. We thus evaluated the contribution of modified cytosines to those genes showing transcriptional similarity in couples as well as the relationships these CpG sites with other gene regulatory elements, such as transcription factor binding sites (TFBS. Our findings suggested that transcriptional similarity in couples likely reflected shared common environment partially mediated through cytosine modifications.

  11. Untangling the transcription regulatory network of the bacitracin synthase operon in Bacillus licheniformis DW2.

    Science.gov (United States)

    Wang, Dong; Wang, Qin; Qiu, Yimin; Nomura, Christopher T; Li, Junhui; Chen, Shouwen

    The bacitracin synthetase gene cluster in Bacillus licheniformis DW2 is composed of the bacABC operon encoding a non-ribosomal peptide synthetase and bacT encoding a thioesterase. Although the bacitracin gene cluster has been well studied, little is known about how this gene cluster is regulated. This study provides insight into how the transcription factors Spo0A and AbrB regulate bacitracin biosynthesis. Deletion of spo0A resulted in drastically reduced expression of bacA and bacT, and subsequently bacitracin production. On the other hand, the expression of bacA and bacT increased significantly in B. licheniformis DW2ΔabrB and DW2Δ0AΔabrB compared to the wild-type strain DW2. The bacitracin yields on cell numbers (U/CFU) in DW2ΔabrB and DW2Δ0A/pHY300-0A-sad67 were 17.5% and 14.9% higher than that of the wild-type strain. An electrophoretic mobility shift assay (EMSA) further confirmed that AbrB could directly bind to the promoter regions of bacA and bacT. These results indicate that AbrB acts as a repressor of bacitracin biosynthesis by inhibiting bacA and bacT expression, while Spo0A indirectly promotes bacitracin biosynthesis by repressing abrB expression. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  12. RNA-seq of 272 gliomas revealed a novel, recurrent PTPRZ1-MET fusion transcript in secondary glioblastomas.

    Science.gov (United States)

    Bao, Zhao-Shi; Chen, Hui-Min; Yang, Ming-Yu; Zhang, Chuan-Bao; Yu, Kai; Ye, Wan-Lu; Hu, Bo-Qiang; Yan, Wei; Zhang, Wei; Akers, Johnny; Ramakrishnan, Valya; Li, Jie; Carter, Bob; Liu, Yan-Wei; Hu, Hui-Min; Wang, Zheng; Li, Ming-Yang; Yao, Kun; Qiu, Xiao-Guang; Kang, Chun-Sheng; You, Yong-Ping; Fan, Xiao-Long; Song, Wei Sonya; Li, Rui-Qiang; Su, Xiao-Dong; Chen, Clark C; Jiang, Tao

    2014-11-01

    Studies of gene rearrangements and the consequent oncogenic fusion proteins have laid the foundation for targeted cancer therapy. To identify oncogenic fusions associated with glioma progression, we catalogued fusion transcripts by RNA-seq of 272 gliomas. Fusion transcripts were more frequently found in high-grade gliomas, in the classical subtype of gliomas, and in gliomas treated with radiation/temozolomide. Sixty-seven in-frame fusion transcripts were identified, including three recurrent fusion transcripts: FGFR3-TACC3, RNF213-SLC26A11, and PTPRZ1-MET (ZM). Interestingly, the ZM fusion was found only in grade III astrocytomas (1/13; 7.7%) or secondary GBMs (sGBMs, 3/20; 15.0%). In an independent cohort of sGBMs, the ZM fusion was found in three of 20 (15%) specimens. Genomic analysis revealed that the fusion arose from translocation events involving introns 3 or 8 of PTPRZ and intron 1 of MET. ZM fusion transcripts were found in GBMs irrespective of isocitrate dehydrogenase 1 (IDH1) mutation status. sGBMs harboring ZM fusion showed higher expression of genes required for PIK3CA signaling and lowered expression of genes that suppressed RB1 or TP53 function. Expression of the ZM fusion was mutually exclusive with EGFR overexpression in sGBMs. Exogenous expression of the ZM fusion in the U87MG glioblastoma line enhanced cell migration and invasion. Clinically, patients afflicted with ZM fusion harboring glioblastomas survived poorly relative to those afflicted with non-ZM-harboring sGBMs (P < 0.001). Our study profiles the shifting RNA landscape of gliomas during progression and reveled ZM as a novel, recurrent fusion transcript in sGBMs. © 2014 Bao et al.; Published by Cold Spring Harbor Laboratory Press.

  13. Mixture models reveal multiple positional bias types in RNA-Seq data and lead to accurate transcript concentration estimates.

    Directory of Open Access Journals (Sweden)

    Andreas Tuerk

    2017-05-01

    Full Text Available Accuracy of transcript quantification with RNA-Seq is negatively affected by positional fragment bias. This article introduces Mix2 (rd. "mixquare", a transcript quantification method which uses a mixture of probability distributions to model and thereby neutralize the effects of positional fragment bias. The parameters of Mix2 are trained by Expectation Maximization resulting in simultaneous transcript abundance and bias estimates. We compare Mix2 to Cufflinks, RSEM, eXpress and PennSeq; state-of-the-art quantification methods implementing some form of bias correction. On four synthetic biases we show that the accuracy of Mix2 overall exceeds the accuracy of the other methods and that its bias estimates converge to the correct solution. We further evaluate Mix2 on real RNA-Seq data from the Microarray and Sequencing Quality Control (MAQC, SEQC Consortia. On MAQC data, Mix2 achieves improved correlation to qPCR measurements with a relative increase in R2 between 4% and 50%. Mix2 also yields repeatable concentration estimates across technical replicates with a relative increase in R2 between 8% and 47% and reduced standard deviation across the full concentration range. We further observe more accurate detection of differential expression with a relative increase in true positives between 74% and 378% for 5% false positives. In addition, Mix2 reveals 5 dominant biases in MAQC data deviating from the common assumption of a uniform fragment distribution. On SEQC data, Mix2 yields higher consistency between measured and predicted concentration ratios. A relative error of 20% or less is obtained for 51% of transcripts by Mix2, 40% of transcripts by Cufflinks and RSEM and 30% by eXpress. Titration order consistency is correct for 47% of transcripts for Mix2, 41% for Cufflinks and RSEM and 34% for eXpress. We, further, observe improved repeatability across laboratory sites with a relative increase in R2 between 8% and 44% and reduced standard deviation.

  14. Strand-specific RNA-seq reveals widespread occurrence of novel cis-natural antisense transcripts in rice

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    Lu Tingting

    2012-12-01

    Full Text Available Abstract Background Cis-natural antisense transcripts (cis-NATs are RNAs transcribed from the antisense strand of a gene locus, and are complementary to the RNA transcribed from the sense strand. Common techniques including microarray approach and analysis of transcriptome databases are the major ways to globally identify cis-NATs in various eukaryotic organisms. Genome-wide in silico analysis has identified a large number of cis-NATs that may generate endogenous short interfering RNAs (nat-siRNAs, which participate in important biogenesis mechanisms for transcriptional and post-transcriptional regulation in rice. However, the transcriptomes are yet to be deeply sequenced to comprehensively investigate cis-NATs. Results We applied high-throughput strand-specific complementary DNA sequencing technology (ssRNA-seq to deeply sequence mRNA for assessing sense and antisense transcripts that were derived under salt, drought and cold stresses, and normal conditions, in the model plant rice (Oryza sativa. Combined with RAP-DB genome annotation (the Rice Annotation Project Database build-5 data set, 76,013 transcripts corresponding to 45,844 unique gene loci were assembled, in which 4873 gene loci were newly identified. Of 3819 putative rice cis-NATs, 2292 were detected as expressed and giving rise to small RNAs from their overlapping regions through integrated analysis of ssRNA-seq data and small RNA data. Among them, 503 cis-NATs seemed to be associated with specific conditions. The deep sequence data from isolated epidermal cells of rice seedlings further showed that 54.0% of cis-NATs were expressed simultaneously in a population of homogenous cells. Nearly 9.7% of rice transcripts were involved in one-to-one or many-to-many cis-NATs formation. Furthermore, only 17.4-34.7% of 223 many-to-many cis-NAT groups were all expressed and generated nat-siRNAs, indicating that only some cis-NAT groups may be involved in complex regulatory networks. Conclusions

  15. A Transcriptional Regulatory Mechanism Finely Tunes the Firing of Type VI Secretion System in Response to Bacterial Enemies.

    Science.gov (United States)

    Lazzaro, Martina; Feldman, Mario F; García Véscovi, Eleonora

    2017-08-22

    The ability to detect and measure danger from an environmental signal is paramount for bacteria to respond accordingly, deploying strategies that halt or counteract potential cellular injury and maximize survival chances. Type VI secretion systems (T6SSs) are complex bacterial contractile nanomachines able to target toxic effectors into neighboring bacteria competing for the same colonization niche. Previous studies support the concept that either T6SSs are constitutively active or they fire effectors in response to various stimuli, such as high bacterial density, cell-cell contact, nutrient depletion, or components from dead sibling cells. For Serratia marcescens , it has been proposed that its T6SS is stochastically expressed, with no distinction between harmless or aggressive competitors. In contrast, we demonstrate that the Rcs regulatory system is responsible for finely tuning Serratia T6SS expression levels, behaving as a transcriptional rheostat. When confronted with harmless bacteria, basal T6SS expression levels suffice for Serratia to eliminate the competitor. A moderate T6SS upregulation is triggered when, according to the aggressor-prey ratio, an unbalanced interplay between homologous and heterologous effectors and immunity proteins takes place. Higher T6SS expression levels are achieved when Serratia is challenged by a contender like Acinetobacter , which indiscriminately fires heterologous effectors able to exert lethal cellular harm, threatening the survival of the Serratia population. We also demonstrate that Serratia 's RcsB-dependent T6SS regulatory mechanism responds not to general stress signals but to the action of specific effectors from competitors, displaying an exquisite strategy to weigh risks and keep the balance between energy expenditure and fitness costs. IMPORTANCE Serratia marcescens is among the health-threatening pathogens categorized by the WHO as research priorities to develop alternative antimicrobial strategies, and it was

  16. Pyrosequencing data reveals tissue-specific expression of lineage-specific transcripts in chickpea

    OpenAIRE

    Garg, Rohini; Jain, Mukesh

    2011-01-01

    Chickpea is a very important crop legume plant, which provides a protein-rich supplement to cereal-based diets and has the ability to fix atmospheric nitrogen. Despite its economic importance, the functional genomic resources for chickpea are very limited. Recently, we reported the complete transcriptome of chickpea using next generation sequencing technologies. We analyzed the tissue-specific expression of chickpea transcripts based on RNA-seq data. In addition, we identified two sets of lin...

  17. Iron Starvation Conditions Upregulate Ehrlichia ruminantium Type IV Secretion System, tr1 Transcription Factor and map1 Genes Family through the Master Regulatory Protein ErxR

    Directory of Open Access Journals (Sweden)

    Amal Moumène

    2018-01-01

    Full Text Available Ehrlichia ruminantium is an obligatory intracellular bacterium that causes heartwater, a fatal disease in ruminants. Due to its intracellular nature, E. ruminantium requires a set of specific virulence factors, such as the type IV secretion system (T4SS, and outer membrane proteins (Map proteins in order to avoid and subvert the host's immune response. Several studies have been conducted to understand the regulation of the T4SS or outer membrane proteins, in Ehrlichia, but no integrated approach has been used to understand the regulation of Ehrlichia pathogenicity determinants in response to environmental cues. Iron is known to be a key nutrient for bacterial growth both in the environment and within hosts. In this study, we experimentally demonstrated the regulation of virB, map1, and tr1 genes by the newly identified master regulator ErxR (for Ehrlichia ruminantium expression regulator. We also analyzed the effect of iron depletion on the expression of erxR gene, tr1 transcription factor, T4SS and map1 genes clusters in E. ruminantium. We show that exposure of E. ruminantium to iron starvation induces erxR and subsequently tr1, virB, and map1 genes. Our results reveal tight co-regulation of T4SS and map1 genes via the ErxR regulatory protein at the transcriptional level, and, for the first time link map genes to the virulence function sensu stricto, thereby advancing our understanding of Ehrlichia's infection process. These results suggest that Ehrlichia is able to sense changes in iron concentrations in the environment and to regulate the expression of virulence factors accordingly.

  18. Transcript profiling reveals rewiring of iron assimilation gene expression in Candida albicans and C. dubliniensis.

    LENUS (Irish Health Repository)

    Moran, Gary P

    2012-12-01

    Hyphal growth is repressed in Candida albicans and Candida dubliniensis by the transcription factor Nrg1. Transcript profiling of a C. dubliniensis NRG1 mutant identified a common group of 28 NRG1-repressed genes in both species, including the hypha-specific genes HWP1, ECE1 and the regulator of cell elongation UME6. Unexpectedly, C. dubliniensis NRG1 was required for wild-type levels of expression of 10 genes required for iron uptake including seven ferric reductases, SIT1, FTR1 and RBT5. However, at alkaline pH and during filamentous growth in 10% serum, most of these genes were highly induced in C. dubliniensis. Conversely, RBT5, PGA10, FRE10 and FRP1 did not exhibit induction during hyphal growth when NRG1 is downregulated, indicating that in C. dubliniensis NRG1 is also required for optimal expression of these genes in alkaline environments. In iron-depleted medium at pH 4.5, reduced growth of the NRG1 mutant relative to wild type was observed; however, growth was restored to wild-type levels or greater at pH 6.5, indicating that alkaline induction of iron assimilation gene expression could rescue this phenotype. These data indicate that transcriptional control of iron assimilation and pseudohypha formation has been separated in C. albicans, perhaps promoting growth in a wider range of niches.

  19. Ancestral regulatory circuits governing ectoderm patterning downstream of Nodal and BMP2/4 revealed by gene regulatory network analysis in an echinoderm.

    Directory of Open Access Journals (Sweden)

    Alexandra Saudemont

    2010-12-01

    Full Text Available Echinoderms, which are phylogenetically related to vertebrates and produce large numbers of transparent embryos that can be experimentally manipulated, offer many advantages for the analysis of the gene regulatory networks (GRN regulating germ layer formation. During development of the sea urchin embryo, the ectoderm is the source of signals that pattern all three germ layers along the dorsal-ventral axis. How this signaling center controls patterning and morphogenesis of the embryo is not understood. Here, we report a large-scale analysis of the GRN deployed in response to the activity of this signaling center in the embryos of the Mediterranean sea urchin Paracentrotus lividus, in which studies with high spatial resolution are possible. By using a combination of in situ hybridization screening, overexpression of mRNA, recombinant ligand treatments, and morpholino-based loss-of-function studies, we identified a cohort of transcription factors and signaling molecules expressed in the ventral ectoderm, dorsal ectoderm, and interposed neurogenic ("ciliary band" region in response to the known key signaling molecules Nodal and BMP2/4 and defined the epistatic relationships between the most important genes. The resultant GRN showed a number of striking features. First, Nodal was found to be essential for the expression of all ventral and dorsal marker genes, and BMP2/4 for all dorsal genes. Second, goosecoid was identified as a central player in a regulatory sub-circuit controlling mouth formation, while tbx2/3 emerged as a critical factor for differentiation of the dorsal ectoderm. Finally, and unexpectedly, a neurogenic ectoderm regulatory circuit characterized by expression of "ciliary band" genes was triggered in the absence of TGF beta signaling. We propose a novel model for ectoderm regionalization, in which neural ectoderm is the default fate in the absence of TGF beta signaling, and suggest that the stomodeal and neural subcircuits that we

  20. In vivo transcriptional profile analysis reveals RNA splicing and chromatin remodeling as prominent processes for adult neurogenesis.

    Science.gov (United States)

    Lim, Daniel A; Suárez-Fariñas, Mayte; Naef, Felix; Hacker, Coleen R; Menn, Benedicte; Takebayashi, Hirohide; Magnasco, Marcelo; Patil, Nila; Alvarez-Buylla, Arturo

    2006-01-01

    Neural stem cells and neurogenesis persist in the adult mammalian brain subventricular zone (SVZ). Cells born in the rodent SVZ migrate to the olfactory bulb (Ob) where they differentiate into interneurons. To determine the gene expression and functional profile of SVZ neurogenesis, we performed three complementary sets of transcriptional analysis experiments using Affymetrix GeneChips: (1) comparison of adult mouse SVZ and Ob gene expression profiles with those of the striatum, cerebral cortex, and hippocampus; (2) profiling of SVZ stem cells and ependyma isolated by fluorescent-activated cell sorting (FACS); and (3) analysis of gene expression changes during in vivo SVZ regeneration after anti-mitotic treatment. Gene Ontology (GO) analysis of data from these three separate approaches showed that in adult SVZ neurogenesis, RNA splicing and chromatin remodeling are biological processes as statistically significant as cell proliferation, transcription, and neurogenesis. In non-neurogenic brain regions, RNA splicing and chromatin remodeling were not prominent processes. Fourteen mRNA splicing factors including Sf3b1, Sfrs2, Lsm4, and Khdrbs1/Sam68 were detected along with 9 chromatin remodeling genes including Mll, Bmi1, Smarcad1, Baf53a, and Hat1. We validated the transcriptional profile data with Northern blot analysis and in situ hybridization. The data greatly expand the catalogue of cell cycle components, transcription factors, and migration genes for adult SVZ neurogenesis and reveal RNA splicing and chromatin remodeling as prominent biological processes for these germinal cells.

  1. Microarray analysis of a salamander hopeful monster reveals transcriptional signatures of paedomorphic brain development

    Science.gov (United States)

    2010-01-01

    Background The Mexican axolotl (Ambystoma mexicanum) is considered a hopeful monster because it exhibits an adaptive and derived mode of development - paedomorphosis - that has evolved rapidly and independently among tiger salamanders. Unlike related tiger salamanders that undergo metamorphosis, axolotls retain larval morphological traits into adulthood and thus present an adult body plan that differs dramatically from the ancestral (metamorphic) form. The basis of paedomorphic development was investigated by comparing temporal patterns of gene transcription between axolotl and tiger salamander larvae (Ambystoma tigrinum tigrinum) that typically undergo a metamorphosis. Results Transcript abundances from whole brain and pituitary were estimated via microarray analysis on four different days post hatching (42, 56, 70, 84 dph) and regression modeling was used to independently identify genes that were differentially expressed as a function of time in both species. Collectively, more differentially expressed genes (DEGs) were identified as unique to the axolotl (n = 76) and tiger salamander (n = 292) than were identified as shared (n = 108). All but two of the shared DEGs exhibited the same temporal pattern of expression and the unique genes tended to show greater changes later in the larval period when tiger salamander larvae were undergoing anatomical metamorphosis. A second, complementary analysis that directly compared the expression of 1320 genes between the species identified 409 genes that differed as a function of species or the interaction between time and species. Of these 409 DEGs, 84% exhibited higher abundances in tiger salamander larvae at all sampling times. Conclusions Many of the unique tiger salamander transcriptional responses are probably associated with metamorphic biological processes. However, the axolotl also showed unique patterns of transcription early in development. In particular, the axolotl showed a genome-wide reduction in mRNA abundance

  2. Microarray analysis of a salamander hopeful monster reveals transcriptional signatures of paedomorphic brain development

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    Putta Srikrishna

    2010-06-01

    Full Text Available Abstract Background The Mexican axolotl (Ambystoma mexicanum is considered a hopeful monster because it exhibits an adaptive and derived mode of development - paedomorphosis - that has evolved rapidly and independently among tiger salamanders. Unlike related tiger salamanders that undergo metamorphosis, axolotls retain larval morphological traits into adulthood and thus present an adult body plan that differs dramatically from the ancestral (metamorphic form. The basis of paedomorphic development was investigated by comparing temporal patterns of gene transcription between axolotl and tiger salamander larvae (Ambystoma tigrinum tigrinum that typically undergo a metamorphosis. Results Transcript abundances from whole brain and pituitary were estimated via microarray analysis on four different days post hatching (42, 56, 70, 84 dph and regression modeling was used to independently identify genes that were differentially expressed as a function of time in both species. Collectively, more differentially expressed genes (DEGs were identified as unique to the axolotl (n = 76 and tiger salamander (n = 292 than were identified as shared (n = 108. All but two of the shared DEGs exhibited the same temporal pattern of expression and the unique genes tended to show greater changes later in the larval period when tiger salamander larvae were undergoing anatomical metamorphosis. A second, complementary analysis that directly compared the expression of 1320 genes between the species identified 409 genes that differed as a function of species or the interaction between time and species. Of these 409 DEGs, 84% exhibited higher abundances in tiger salamander larvae at all sampling times. Conclusions Many of the unique tiger salamander transcriptional responses are probably associated with metamorphic biological processes. However, the axolotl also showed unique patterns of transcription early in development. In particular, the axolotl showed a genome

  3. Quantitative transcription dynamic analysis reveals candidate genes and key regulators for ethanol tolerance in Saccharomyces cerevisiae

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    Ma Menggen

    2010-06-01

    Full Text Available Abstract Background Derived from our lignocellulosic conversion inhibitor-tolerant yeast, we generated an ethanol-tolerant strain Saccharomyces cerevisiae NRRL Y-50316 by enforced evolutionary adaptation. Using a newly developed robust mRNA reference and a master equation unifying gene expression data analyses, we investigated comparative quantitative transcription dynamics of 175 genes selected from previous studies for an ethanol-tolerant yeast and its closely related parental strain. Results A highly fitted master equation was established and applied for quantitative gene expression analyses using pathway-based qRT-PCR array assays. The ethanol-tolerant Y-50316 displayed significantly enriched background of mRNA abundance for at least 35 genes without ethanol challenge compared with its parental strain Y-50049. Under the ethanol challenge, the tolerant Y-50316 responded in consistent expressions over time for numerous genes belonging to groups of heat shock proteins, trehalose metabolism, glycolysis, pentose phosphate pathway, fatty acid metabolism, amino acid biosynthesis, pleiotropic drug resistance gene family and transcription factors. The parental strain showed repressed expressions for many genes and was unable to withstand the ethanol stress and establish a viable culture and fermentation. The distinct expression dynamics between the two strains and their close association with cell growth, viability and ethanol fermentation profiles distinguished the tolerance-response from the stress-response in yeast under the ethanol challenge. At least 82 genes were identified as candidate and key genes for ethanol-tolerance and subsequent fermentation under the stress. Among which, 36 genes were newly recognized by the present study. Most of the ethanol-tolerance candidate genes were found to share protein binding motifs of transcription factors Msn4p/Msn2p, Yap1p, Hsf1p and Pdr1p/Pdr3p. Conclusion Enriched background of transcription abundance

  4. Transcriptional analysis of the jamaicamide gene cluster from the marine cyanobacterium Lyngbya majuscula and identification of possible regulatory proteins

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    Dorrestein Pieter C

    2009-12-01

    Full Text Available Abstract Background The marine cyanobacterium Lyngbya majuscula is a prolific producer of bioactive secondary metabolites. Although biosynthetic gene clusters encoding several of these compounds have been identified, little is known about how these clusters of genes are transcribed or regulated, and techniques targeting genetic manipulation in Lyngbya strains have not yet been developed. We conducted transcriptional analyses of the jamaicamide gene cluster from a Jamaican strain of Lyngbya majuscula, and isolated proteins that could be involved in jamaicamide regulation. Results An unusually long untranslated leader region of approximately 840 bp is located between the jamaicamide transcription start site (TSS and gene cluster start codon. All of the intergenic regions between the pathway ORFs were transcribed into RNA in RT-PCR experiments; however, a promoter prediction program indicated the possible presence of promoters in multiple intergenic regions. Because the functionality of these promoters could not be verified in vivo, we used a reporter gene assay in E. coli to show that several of these intergenic regions, as well as the primary promoter preceding the TSS, are capable of driving β-galactosidase production. A protein pulldown assay was also used to isolate proteins that may regulate the jamaicamide pathway. Pulldown experiments using the intergenic region upstream of jamA as a DNA probe isolated two proteins that were identified by LC-MS/MS. By BLAST analysis, one of these had close sequence identity to a regulatory protein in another cyanobacterial species. Protein comparisons suggest a possible correlation between secondary metabolism regulation and light dependent complementary chromatic adaptation. Electromobility shift assays were used to evaluate binding of the recombinant proteins to the jamaicamide promoter region. Conclusion Insights into natural product regulation in cyanobacteria are of significant value to drug discovery

  5. Uncovering packaging features of co-regulated modules based on human protein interaction and transcriptional regulatory networks

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    He Weiming

    2010-07-01

    Full Text Available Abstract Background Network co-regulated modules are believed to have the functionality of packaging multiple biological entities, and can thus be assumed to coordinate many biological functions in their network neighbouring regions. Results Here, we weighted edges of a human protein interaction network and a transcriptional regulatory network to construct an integrated network, and introduce a probabilistic model and a bipartite graph framework to exploit human co-regulated modules and uncover their specific features in packaging different biological entities (genes, protein complexes or metabolic pathways. Finally, we identified 96 human co-regulated modules based on this method, and evaluate its effectiveness by comparing it with four other methods. Conclusions Dysfunctions in co-regulated interactions often occur in the development of cancer. Therefore, we focussed on an example co-regulated module and found that it could integrate a number of cancer-related genes. This was extended to causal dysfunctions of some complexes maintained by several physically interacting proteins, thus coordinating several metabolic pathways that directly underlie cancer.

  6. Does positive selection drive transcription factor binding site turnover? A test with Drosophila cis-regulatory modules.

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    Bin Z He

    2011-04-01

    Full Text Available Transcription factor binding site(s (TFBS gain and loss (i.e., turnover is a well-documented feature of cis-regulatory module (CRM evolution, yet little attention has been paid to the evolutionary force(s driving this turnover process. The predominant view, motivated by its widespread occurrence, emphasizes the importance of compensatory mutation and genetic drift. Positive selection, in contrast, although it has been invoked in specific instances of adaptive gene expression evolution, has not been considered as a general alternative to neutral compensatory evolution. In this study we evaluate the two hypotheses by analyzing patterns of single nucleotide polymorphism in the TFBS of well-characterized CRM in two closely related Drosophila species, Drosophila melanogaster and Drosophila simulans. An important feature of the analysis is classification of TFBS mutations according to the direction of their predicted effect on binding affinity, which allows gains and losses to be evaluated independently along the two phylogenetic lineages. The observed patterns of polymorphism and divergence are not compatible with neutral evolution for either class of mutations. Instead, multiple lines of evidence are consistent with contributions of positive selection to TFBS gain and loss as well as purifying selection in its maintenance. In discussion, we propose a model to reconcile the finding of selection driving TFBS turnover with constrained CRM function over long evolutionary time.

  7. Transcriptome landscape of Lactococcus lactis reveals many novel RNAs including a small regulatory RNA involved in carbon uptake and metabolism

    NARCIS (Netherlands)

    van der Meulen, Sjoerd B; de Jong, Anne; Kok, Jan

    2016-01-01

    RNA sequencing has revolutionized genome-wide transcriptome analyses, and the identification of non-coding regulatory RNAs in bacteria has thus increased concurrently. Here we reveal the transcriptome map of the lactic acid bacterial paradigm Lactococcus lactis MG1363 by employing differential RNA

  8. The Streptococcus pyogenes Serotype M49 Nra-Ralp3 Transcriptional Regulatory Network and Its Control of Virulence Factor Expression from the Novel eno ralp3 epf sagA Pathogenicity Region▿ †

    Science.gov (United States)

    Kreikemeyer, Bernd; Nakata, Masanobu; Köller, Thomas; Hildisch, Hendrikje; Kourakos, Vassilios; Standar, Kerstin; Kawabata, Shigetada; Glocker, Michael O.; Podbielski, Andreas

    2007-01-01

    Many Streptococcus pyogenes (group A streptococcus [GAS]) virulence factor- and transcriptional regulator-encoding genes cluster together in discrete genomic regions. Nra is a central regulator of the FCT region. Previous studies exclusively described Nra as a transcriptional repressor of adhesin and toxin genes. Here transcriptome and proteome analysis of a serotype M49 GAS strain and an isogenic Nra mutant of this strain revealed the complete Nra regulon profile. Nra is active in all growth phases tested, with the largest regulon in the transition phase. Almost exclusively, virulence factor-encoding genes are repressed by Nra; these genes include the GAS pilus operon, the capsule synthesis operon, the cytolysin-mediated translocation system genes, all Mga region core virulence genes, and genes encoding other regulators, like the Ihk/Irr system, Rgg, and two additional RofA-like protein family regulators. Surprisingly, our experiments revealed that Nra additionally acts as a positive regulator, mostly for genes encoding proteins and enzymes with metabolic functions. Epidemiological investigations revealed strong genetic linkage of one particular Nra-repressed regulator, Ralp3 (SPy0735), with a gene encoding Epf (extracellular protein factor from Streptococcus suis). In a serotype-specific fashion, this ralp3 epf gene block is integrated, most likely via transposition, into the eno sagA virulence gene block, which is present in all GAS serotypes. In GAS serotypes M1, M4, M12, M28, and M49 this novel discrete genetic region is therefore designated the eno ralp3 epf sagA (ERES) pathogenicity region. Functional experiments showed that Epf is a novel GAS plasminogen-binding protein and revealed that Ralp3 activity counteracts Nra and MsmR regulatory activity. In addition to the Mga and FCT regions, the ERES region is the third discrete chromosomal pathogenicity region. All of these regions are transcriptionally linked, adding another level of complexity to the known

  9. The Streptococcus pyogenes serotype M49 Nra-Ralp3 transcriptional regulatory network and its control of virulence factor expression from the novel eno ralp3 epf sagA pathogenicity region.

    Science.gov (United States)

    Kreikemeyer, Bernd; Nakata, Masanobu; Köller, Thomas; Hildisch, Hendrikje; Kourakos, Vassilios; Standar, Kerstin; Kawabata, Shigetada; Glocker, Michael O; Podbielski, Andreas

    2007-12-01

    Many Streptococcus pyogenes (group A streptococcus [GAS]) virulence factor- and transcriptional regulator-encoding genes cluster together in discrete genomic regions. Nra is a central regulator of the FCT region. Previous studies exclusively described Nra as a transcriptional repressor of adhesin and toxin genes. Here transcriptome and proteome analysis of a serotype M49 GAS strain and an isogenic Nra mutant of this strain revealed the complete Nra regulon profile. Nra is active in all growth phases tested, with the largest regulon in the transition phase. Almost exclusively, virulence factor-encoding genes are repressed by Nra; these genes include the GAS pilus operon, the capsule synthesis operon, the cytolysin-mediated translocation system genes, all Mga region core virulence genes, and genes encoding other regulators, like the Ihk/Irr system, Rgg, and two additional RofA-like protein family regulators. Surprisingly, our experiments revealed that Nra additionally acts as a positive regulator, mostly for genes encoding proteins and enzymes with metabolic functions. Epidemiological investigations revealed strong genetic linkage of one particular Nra-repressed regulator, Ralp3 (SPy0735), with a gene encoding Epf (extracellular protein factor from Streptococcus suis). In a serotype-specific fashion, this ralp3 epf gene block is integrated, most likely via transposition, into the eno sagA virulence gene block, which is present in all GAS serotypes. In GAS serotypes M1, M4, M12, M28, and M49 this novel discrete genetic region is therefore designated the eno ralp3 epf sagA (ERES) pathogenicity region. Functional experiments showed that Epf is a novel GAS plasminogen-binding protein and revealed that Ralp3 activity counteracts Nra and MsmR regulatory activity. In addition to the Mga and FCT regions, the ERES region is the third discrete chromosomal pathogenicity region. All of these regions are transcriptionally linked, adding another level of complexity to the known

  10. Transcript and metabolite analysis in Trincadeira cultivar reveals novel information regarding the dynamics of grape ripening.

    Science.gov (United States)

    Fortes, Ana M; Agudelo-Romero, Patricia; Silva, Marta S; Ali, Kashif; Sousa, Lisete; Maltese, Federica; Choi, Young H; Grimplet, Jerome; Martinez-Zapater, José M; Verpoorte, Robert; Pais, Maria S

    2011-11-02

    Grapes (Vitis vinifera L.) are economically the most important fruit crop worldwide. However, the complexity of molecular and biochemical events that lead to the onset of ripening of nonclimacteric fruits is not fully understood which is further complicated in grapes due to seasonal and cultivar specific variation. The Portuguese wine variety Trincadeira gives rise to high quality wines but presents extremely irregular berry ripening among seasons probably due to high susceptibility to abiotic and biotic stresses. Ripening of Trincadeira grapes was studied taking into account the transcriptional and metabolic profilings complemented with biochemical data. The mRNA expression profiles of four time points spanning developmental stages from pea size green berries, through véraison and mature berries (EL 32, EL 34, EL 35 and EL 36) and in two seasons (2007 and 2008) were compared using the Affymetrix GrapeGen® genome array containing 23096 probesets corresponding to 18726 unique sequences. Over 50% of these probesets were significantly differentially expressed (1.5 fold) between at least two developmental stages. A common set of modulated transcripts corresponding to 5877 unigenes indicates the activation of common pathways between years despite the irregular development of Trincadeira grapes. These unigenes were assigned to the functional categories of "metabolism", "development", "cellular process", "diverse/miscellanenous functions", "regulation overview", "response to stimulus, stress", "signaling", "transport overview", "xenoprotein, transposable element" and "unknown". Quantitative RT-PCR validated microarrays results being carried out for eight selected genes and five developmental stages (EL 32, EL 34, EL 35, EL 36 and EL 38). Metabolic profiling using 1H NMR spectroscopy associated to two-dimensional techniques showed the importance of metabolites related to oxidative stress response, amino acid and sugar metabolism as well as secondary metabolism. These

  11. Transcriptome profiling of Nasonia vitripennis testis reveals novel transcripts expressed from the selfish B chromosome, paternal sex ratio.

    Science.gov (United States)

    Akbari, Omar S; Antoshechkin, Igor; Hay, Bruce A; Ferree, Patrick M

    2013-09-04

    A widespread phenomenon in nature is sex ratio distortion of arthropod populations caused by microbial and genetic parasites. Currently little is known about how these agents alter host developmental processes to favor one sex or the other. The paternal sex ratio (PSR) chromosome is a nonessential, paternally transmitted centric fragment that segregates in natural populations of the jewel wasp, Nasonia vitripennis. To persist, PSR is thought to modify the hereditary material of the developing sperm, with the result that all nuclear DNA other than the PSR chromosome is destroyed shortly after fertilization. This results in the conversion of a fertilized embryo--normally a female--into a male, thereby insuring transmission of the "selfish" PSR chromosome, and simultaneously leading to wasp populations that are male-biased. To begin to understand this system at the mechanistic level, we carried out transcriptional profiling of testis from WT and PSR-carrying males. We identified a number of transcripts that are differentially expressed between these conditions. We also discovered nine transcripts that are uniquely expressed from the PSR chromosome. Four of these PSR-specific transcripts encode putative proteins, whereas the others have very short open reading frames and no homology to known proteins, suggesting that they are long noncoding RNAs. We propose several different models for how these transcripts could facilitate PSR-dependent effects. Our analyses also revealed 15.71 MB of novel transcribed regions in the N. vitripennis genome, thus increasing the current annotation of total transcribed regions by 53.4%. Finally, we detected expression of multiple meiosis-related genes in the wasp testis, despite the lack of conventional meiosis in the male sex.

  12. Structures of BmrR-Drug Complexes Reveal a Rigid Multidrug Binding Pocket And Transcription Activation Through Tyrosine Expulsion

    Energy Technology Data Exchange (ETDEWEB)

    Newberry, K.J.; Huffman, J.L.; Miller, M.C.; Vazquez-Laslop, N.; Neyfakh, A.A.; Brennan, R.G.

    2009-05-22

    BmrR is a member of the MerR family and a multidrug binding transcription factor that up-regulates the expression of the bmr multidrug efflux transporter gene in response to myriad lipophilic cationic compounds. The structural mechanism by which BmrR binds these chemically and structurally different drugs and subsequently activates transcription is poorly understood. Here, we describe the crystal structures of BmrR bound to rhodamine 6G (R6G) or berberine (Ber) and cognate DNA. These structures reveal each drug stacks against multiple aromatic residues with their positive charges most proximal to the carboxylate group of Glu-253 and that, unlike other multidrug binding pockets, that of BmrR is rigid. Substitution of Glu-253 with either alanine (E253A) or glutamine (E253Q) results in unpredictable binding affinities for R6G, Ber, and tetraphenylphosphonium. Moreover, these drug binding studies reveal that the negative charge of Glu-253 is not important for high affinity binding to Ber and tetraphenylphosphonium but plays a more significant, but unpredictable, role in R6G binding. In vitro transcription data show that E253A and E253Q are constitutively active, and structures of the drug-free E253A-DNA and E253Q-DNA complexes support a transcription activation mechanism requiring the expulsion of Tyr-152 from the multidrug binding pocket. In sum, these data delineate the mechanism by which BmrR binds lipophilic, monovalent cationic compounds and suggest the importance of the redundant negative electrostatic nature of this rigid drug binding pocket that can be used to discriminate against molecules that are not substrates of the Bmr multidrug efflux pump.

  13. Single-Cell Analysis of Human Pancreas Reveals Transcriptional Signatures of Aging and Somatic Mutation Patterns.

    Science.gov (United States)

    Enge, Martin; Arda, H Efsun; Mignardi, Marco; Beausang, John; Bottino, Rita; Kim, Seung K; Quake, Stephen R

    2017-10-05

    As organisms age, cells accumulate genetic and epigenetic errors that eventually lead to impaired organ function or catastrophic transformation such as cancer. Because aging reflects a stochastic process of increasing disorder, cells in an organ will be individually affected in different ways, thus rendering bulk analyses of postmitotic adult cells difficult to interpret. Here, we directly measure the effects of aging in human tissue by performing single-cell transcriptome analysis of 2,544 human pancreas cells from eight donors spanning six decades of life. We find that islet endocrine cells from older donors display increased levels of transcriptional noise and potential fate drift. By determining the mutational history of individual cells, we uncover a novel mutational signature in healthy aging endocrine cells. Our results demonstrate the feasibility of using single-cell RNA sequencing (RNA-seq) data from primary cells to derive insights into genetic and transcriptional processes that operate on aging human tissue. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Intervention of pumpkin seed oil on metabolic disease revealed by metabonomics and transcript profile.

    Science.gov (United States)

    Zhao, Xiu-Ju; Chen, Yu-Lian; Fu, Bing; Zhang, Wen; Liu, Zhiguo; Zhuo, Hexian

    2017-03-01

    Understanding the metabolic and transcription basis of pumpkin seed oil (PSO) intervention on metabolic disease (MD) is essential to daily nutrition and health. This study analyzed the liver metabolic variations of Wistar rats fed normal diet (CON), high-fat diet (HFD) and high-fat plus PSO diet (PSO) to establish the relationship between the liver metabolite composition/transcript profile and the effects of PSO on MD. By using proton nuclear magnetic resonance spectroscopy together with multivariate data analysis, it was found that, compared with CON rats, HFD rats showed clear dysfunctions of choline metabolism, glucose metabolism and nucleotide and amino acid metabolism. Using quantitative real-time polymerase chain reaction (qPCR), it was found that, compared with HFD rats, PSO rats showed alleviated endoplasmic reticulum stress accompanied by lowered unfolded protein response. These findings provide useful information to understand the metabolic alterations triggered by MD and to evaluate the effects of PSO intervention. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  15. Transcriptional Network Analysis Reveals Drought Resistance Mechanisms of AP2/ERF Transgenic Rice

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    Hongryul Ahn

    2017-06-01

    Full Text Available This study was designed to investigate at the molecular level how a transgenic version of rice “Nipponbare” obtained a drought-resistant phenotype. Using multi-omics sequencing data, we compared wild-type rice (WT and a transgenic version (erf71 that had obtained a drought-resistant phenotype by overexpressing OsERF71, a member of the AP2/ERF transcription factor (TF family. A comprehensive bioinformatics analysis pipeline, including TF networks and a cascade tree, was developed for the analysis of multi-omics data. The results of the analysis showed that the presence of OsERF71 at the source of the network controlled global gene expression levels in a specific manner to make erf71 survive longer than WT. Our analysis of the time-series transcriptome data suggests that erf71 diverted more energy to survival-critical mechanisms related to translation, oxidative response, and DNA replication, while further suppressing energy-consuming mechanisms, such as photosynthesis. To support this hypothesis further, we measured the net photosynthesis level under physiological conditions, which confirmed the further suppression of photosynthesis in erf71. In summary, our work presents a comprehensive snapshot of transcriptional modification in transgenic rice and shows how this induced the plants to acquire a drought-resistant phenotype.

  16. In vivo genome-wide profiling of RNA secondary structure reveals novel regulatory features.

    Science.gov (United States)

    Ding, Yiliang; Tang, Yin; Kwok, Chun Kit; Zhang, Yu; Bevilacqua, Philip C; Assmann, Sarah M

    2014-01-30

    RNA structure has critical roles in processes ranging from ligand sensing to the regulation of translation, polyadenylation and splicing. However, a lack of genome-wide in vivo RNA structural data has limited our understanding of how RNA structure regulates gene expression in living cells. Here we present a high-throughput, genome-wide in vivo RNA structure probing method, structure-seq, in which dimethyl sulphate methylation of unprotected adenines and cytosines is identified by next-generation sequencing. Application of this method to Arabidopsis thaliana seedlings yielded the first in vivo genome-wide RNA structure map at nucleotide resolution for any organism, with quantitative structural information across more than 10,000 transcripts. Our analysis reveals a three-nucleotide periodic repeat pattern in the structure of coding regions, as well as a less-structured region immediately upstream of the start codon, and shows that these features are strongly correlated with translation efficiency. We also find patterns of strong and weak secondary structure at sites of alternative polyadenylation, as well as strong secondary structure at 5' splice sites that correlates with unspliced events. Notably, in vivo structures of messenger RNAs annotated for stress responses are poorly predicted in silico, whereas mRNA structures of genes related to cell function maintenance are well predicted. Global comparison of several structural features between these two categories shows that the mRNAs associated with stress responses tend to have more single-strandedness, longer maximal loop length and higher free energy per nucleotide, features that may allow these RNAs to undergo conformational changes in response to environmental conditions. Structure-seq allows the RNA structurome and its biological roles to be interrogated on a genome-wide scale and should be applicable to any organism.

  17. Global mapping of cell type-specific open chromatin by FAIRE-seq reveals the regulatory role of the NFI family in adipocyte differentiation.

    Directory of Open Access Journals (Sweden)

    Hironori Waki

    2011-10-01

    Full Text Available Identification of regulatory elements within the genome is crucial for understanding the mechanisms that govern cell type-specific gene expression. We generated genome-wide maps of open chromatin sites in 3T3-L1 adipocytes (on day 0 and day 8 of differentiation and NIH-3T3 fibroblasts using formaldehyde-assisted isolation of regulatory elements coupled with high-throughput sequencing (FAIRE-seq. FAIRE peaks at the promoter were associated with active transcription and histone modifications of H3K4me3 and H3K27ac. Non-promoter FAIRE peaks were characterized by H3K4me1+/me3-, the signature of enhancers, and were largely located in distal regions. The non-promoter FAIRE peaks showed dynamic change during differentiation, while the promoter FAIRE peaks were relatively constant. Functionally, the adipocyte- and preadipocyte-specific non-promoter FAIRE peaks were, respectively, associated with genes up-regulated and down-regulated by differentiation. Genes highly up-regulated during differentiation were associated with multiple clustered adipocyte-specific FAIRE peaks. Among the adipocyte-specific FAIRE peaks, 45.3% and 11.7% overlapped binding sites for, respectively, PPARγ and C/EBPα, the master regulators of adipocyte differentiation. Computational motif analyses of the adipocyte-specific FAIRE peaks revealed enrichment of a binding motif for nuclear family I (NFI transcription factors. Indeed, ChIP assay showed that NFI occupy the adipocyte-specific FAIRE peaks and/or the PPARγ binding sites near PPARγ, C/EBPα, and aP2 genes. Overexpression of NFIA in 3T3-L1 cells resulted in robust induction of these genes and lipid droplet formation without differentiation stimulus. Overexpression of dominant-negative NFIA or siRNA-mediated knockdown of NFIA or NFIB significantly suppressed both induction of genes and lipid accumulation during differentiation, suggesting a physiological function of these factors in the adipogenic program. Together, our

  18. RNA-Seq reveals spliceosome and proteasome genes as most consistent transcripts in human cancer cells.

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    Tara Macrae

    Full Text Available Accurate quantification of gene expression by qRT-PCR relies on normalization against a consistently expressed control gene. However, control genes in common use often vary greatly between samples, especially in cancer. The advent of Next Generation Sequencing technology offers the possibility to better select control genes with the least cell to cell variability in steady state transcript levels. Here we analyze the transcriptomes of 55 leukemia samples to identify the most consistent genes. This list is enriched for components of the proteasome (ex. PSMA1 and spliceosome (ex. SF3B2, and also includes the translation initiation factor EIF4H, and many heterogeneous nuclear ribonucleoprotein genes (ex. HNRNPL. We have validated the consistency of our new control genes in 1933 cancer and normal tissues using publically available RNA-seq data, and their usefulness in qRT-PCR analysis is clearly demonstrated.

  19. Broad genomic and transcriptional analysis reveals a highly derived genome in dinoflagellate mitochondria

    Directory of Open Access Journals (Sweden)

    Keeling Patrick J

    2007-09-01

    Full Text Available Abstract Background Dinoflagellates comprise an ecologically significant and diverse eukaryotic phylum that is sister to the phylum containing apicomplexan endoparasites. The mitochondrial genome of apicomplexans is uniquely reduced in gene content and size, encoding only three proteins and two ribosomal RNAs (rRNAs within a highly compacted 6 kb DNA. Dinoflagellate mitochondrial genomes have been comparatively poorly studied: limited available data suggest some similarities with apicomplexan mitochondrial genomes but an even more radical type of genomic organization. Here, we investigate structure, content and expression of dinoflagellate mitochondrial genomes. Results From two dinoflagellates, Crypthecodinium cohnii and Karlodinium micrum, we generated over 42 kb of mitochondrial genomic data that indicate a reduced gene content paralleling that of mitochondrial genomes in apicomplexans, i.e., only three protein-encoding genes and at least eight conserved components of the highly fragmented large and small subunit rRNAs. Unlike in apicomplexans, dinoflagellate mitochondrial genes occur in multiple copies, often as gene fragments, and in numerous genomic contexts. Analysis of cDNAs suggests several novel aspects of dinoflagellate mitochondrial gene expression. Polycistronic transcripts were found, standard start codons are absent, and oligoadenylation occurs upstream of stop codons, resulting in the absence of termination codons. Transcripts of at least one gene, cox3, are apparently trans-spliced to generate full-length mRNAs. RNA substitutional editing, a process previously identified for mRNAs in dinoflagellate mitochondria, is also implicated in rRNA expression. Conclusion The dinoflagellate mitochondrial genome shares the same gene complement and fragmentation of rRNA genes with its apicomplexan counterpart. However, it also exhibits several unique characteristics. Most notable are the expansion of gene copy numbers and their arrangements

  20. In Situ Tagged nsp15 Reveals Interactions with Coronavirus Replication/Transcription Complex-Associated Proteins

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    Jeremiah Athmer

    2017-01-01

    Full Text Available Coronavirus (CoV replication and transcription are carried out in close proximity to restructured endoplasmic reticulum (ER membranes in replication/transcription complexes (RTC. Many of the CoV nonstructural proteins (nsps are required for RTC function; however, not all of their functions are known. nsp15 contains an endoribonuclease domain that is conserved in the CoV family. While the enzymatic activity and crystal structure of nsp15 are well defined, its role in replication remains elusive. nsp15 localizes to sites of RNA replication, but whether it acts independently or requires additional interactions for its function remains unknown. To begin to address these questions, we created an in situ tagged form of nsp15 using the prototypic CoV, mouse hepatitis virus (MHV. In MHV, nsp15 contains the genomic RNA packaging signal (P/S, a 95-bp RNA stem-loop structure that is not required for viral replication or nsp15 function. Utilizing this knowledge, we constructed an internal hemagglutinin (HA tag that replaced the P/S. We found that nsp15-HA was localized to discrete perinuclear puncta and strongly colocalized with nsp8 and nsp12, both well-defined members of the RTC, but not the membrane (M protein, involved in virus assembly. Finally, we found that nsp15 interacted with RTC-associated proteins nsp8 and nsp12 during infection, and this interaction was RNA independent. From this, we conclude that nsp15 localizes and interacts with CoV proteins in the RTC, suggesting it plays a direct or indirect role in virus replication. Furthermore, the use of in situ epitope tags could be used to determine novel nsp-nsp interactions in coronaviruses.

  1. RNA-SEQ reveals transcriptional level changes of poplar roots in different forms of nitrogen treatments

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    Chunpu eQu

    2016-02-01

    Full Text Available Poplar has emerged as a model plant for understanding molecular mechanisms of tree growth, development and response to environment. Long-term application of different forms of nitrogen (such as NO3--N and NH4+-N may cause morphological changes of poplar roots; however, the molecular level changes are still not well known. In this study, we analyzed the expression profiling of poplar roots treated by three forms of nitrogen: S1 (NH4+, S2 (NH4NO3 and S3 (NO3- by using RNA-SEQ technique. We found 463 genes significantly differentially expressed in roots by different N treatments, of which a total of 116 genes were found to differentially express between S1 and S2, 173 genes between S2 and S3, and 327 genes between S1 and S3. A cluster analysis shows significant difference in many transcription factor families and functional genes family under different N forms. Through an analysis of Mapman metabolic pathway, we found that the significantly differentially expressed genes are associated with fermentation, glycolysis and tricarboxylic acid cycle (TCA, secondary metabolism, hormone metabolism, and transport processing. Interestingly, we did not find significantly differentially expressed genes in N metabolism pathway, mitochondrial electron transport / ATP synthesis and mineral nutrition. We also found abundant candidate genes (20 transcription factors and 30 functional genes regulating morphology changes of poplar roots under the three N forms. The results obtained are beneficial to a better understanding of the potential molecular and cellular mechanisms regulating root morphology changes under different N treatments.

  2. Diurnal Cycling Transcription Factors of Pineapple Revealed by Genome-Wide Annotation and Global Transcriptomic Analysis.

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    Sharma, Anupma; Wai, Ching Man; Ming, Ray; Yu, Qingyi

    2017-09-01

    Circadian clock provides fitness advantage by coordinating internal metabolic and physiological processes to external cyclic environments. Core clock components exhibit daily rhythmic changes in gene expression, and the majority of them are transcription factors (TFs) and transcription coregulators (TCs). We annotated 1,398 TFs from 67 TF families and 80 TCs from 20 TC families in pineapple, and analyzed their tissue-specific and diurnal expression patterns. Approximately 42% of TFs and 45% of TCs displayed diel rhythmic expression, including 177 TF/TCs cycling only in the nonphotosynthetic leaf tissue, 247 cycling only in the photosynthetic leaf tissue, and 201 cycling in both. We identified 68 TF/TCs whose cycling expression was tightly coupled between the photosynthetic and nonphotosynthetic leaf tissues. These TF/TCs likely coordinate key biological processes in pineapple as we demonstrated that this group is enriched in homologous genes that form the core circadian clock in Arabidopsis and includes a STOP1 homolog. Two lines of evidence support the important role of the STOP1 homolog in regulating CAM photosynthesis in pineapple. First, STOP1 responds to acidic pH and regulates a malate channel in multiple plant species. Second, the cycling expression pattern of the pineapple STOP1 and the diurnal pattern of malate accumulation in pineapple leaf are correlated. We further examined duplicate-gene retention and loss in major known circadian genes and refined their evolutionary relationships between pineapple and other plants. Significant variations in duplicate-gene retention and loss were observed for most clock genes in both monocots and dicots. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  3. A strong anti-inflammatory signature revealed by liver transcription profiling of Tmprss6-/- mice.

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    Michela Riba

    Full Text Available Control of systemic iron homeostasis is interconnected with the inflammatory response through the key iron regulator, the antimicrobial peptide hepcidin. We have previously shown that mice with iron deficiency anemia (IDA-low hepcidin show a pro-inflammatory response that is blunted in iron deficient-high hepcidin Tmprss6 KO mice. The transcriptional response associated with chronic hepcidin overexpression due to genetic inactivation of Tmprss6 is unknown. By using whole genome transcription profiling of the liver and analysis of spleen immune-related genes we identified several functional pathways differentially expressed in Tmprss6 KO mice, compared to IDA animals and thus irrespective of the iron status. In the effort of defining genes potentially targets of Tmprss6 we analyzed liver gene expression changes according to the genotype and independently of treatment. Tmprss6 inactivation causes down-regulation of liver pathways connected to immune and inflammatory response as well as spleen genes related to macrophage activation and inflammatory cytokines production. The anti-inflammatory status of Tmprss6 KO animals was confirmed by the down-regulation of pathways related to immunity, stress response and intracellular signaling in both liver and spleen after LPS treatment. Opposite to Tmprss6 KO mice, Hfe(-/- mice are characterized by iron overload with inappropriately low hepcidin levels. Liver expression profiling of Hfe(-/- deficient versus iron loaded mice show the opposite expression of some of the genes modulated by the loss of Tmprss6. Altogether our results confirm the anti-inflammatory status of Tmprss6 KO mice and identify new potential target pathways/genes of Tmprss6.

  4. Identification of new TSGA10 transcript variants in human testis with conserved regulatory RNA elements in 5'untranslated region and distinct expression in breast cancer.

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    Salehipour, Pouya; Nematzadeh, Mahsa; Mobasheri, Maryam Beigom; Afsharpad, Mandana; Mansouri, Kamran; Modarressi, Mohammad Hossein

    2017-09-01

    Testis specific gene antigen 10 (TSGA10) is a cancer testis antigen involved in the process of spermatogenesis. TSGA10 could also play an important role in the inhibition of angiogenesis by preventing nuclear localization of HIF-1α. Although it has been shown that TSGA10 messenger RNA (mRNA) is mainly expressed in testis and some tumors, the transcription pattern and regulatory mechanisms of this gene remain largely unknown. Here, we report that human TSGA10 comprises at least 22 exons and generates four different transcript variants. It was identified that using two distinct promoters and splicing of exons 4 and 7 produced these transcript variants, which have the same coding sequence, but the sequence of 5'untanslated region (5'UTR) is different between them. This is significant because conserved regulatory RNA elements like upstream open reading frame (uORF) and putative internal ribosome entry site (IRES) were found in this region which have different combinations in each transcript variant and it may influence translational efficiency of them in normal or unusual environmental conditions like hypoxia. To indicate the transcription pattern of TSGA10 in breast cancer, expression of identified transcript variants was analyzed in 62 breast cancer samples. We found that TSGA10 tends to express variants with shorter 5'UTR and fewer uORF elements in breast cancer tissues. Our study demonstrates for the first time the expression of different TSGA10 transcript variants in testis and breast cancer tissues and provides a first clue to a role of TSGA10 5'UTR in regulation of translation in unusual environmental conditions like hypoxia. Copyright © 2017. Published by Elsevier B.V.

  5. Discovery of transcription factors and regulatory regions driving in vivo tumor development by ATAC-seq and FAIRE-seq open chromatin profiling.

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    Kristofer Davie

    2015-02-01

    Full Text Available Genomic enhancers regulate spatio-temporal gene expression by recruiting specific combinations of transcription factors (TFs. When TFs are bound to active regulatory regions, they displace canonical nucleosomes, making these regions biochemically detectable as nucleosome-depleted regions or accessible/open chromatin. Here we ask whether open chromatin profiling can be used to identify the entire repertoire of active promoters and enhancers underlying tissue-specific gene expression during normal development and oncogenesis in vivo. To this end, we first compare two different approaches to detect open chromatin in vivo using the Drosophila eye primordium as a model system: FAIRE-seq, based on physical separation of open versus closed chromatin; and ATAC-seq, based on preferential integration of a transposon into open chromatin. We find that both methods reproducibly capture the tissue-specific chromatin activity of regulatory regions, including promoters, enhancers, and insulators. Using both techniques, we screened for regulatory regions that become ectopically active during Ras-dependent oncogenesis, and identified 3778 regions that become (over-activated during tumor development. Next, we applied motif discovery to search for candidate transcription factors that could bind these regions and identified AP-1 and Stat92E as key regulators. We validated the importance of Stat92E in the development of the tumors by introducing a loss of function Stat92E mutant, which was sufficient to rescue the tumor phenotype. Additionally we tested if the predicted Stat92E responsive regulatory regions are genuine, using ectopic induction of JAK/STAT signaling in developing eye discs, and observed that similar chromatin changes indeed occurred. Finally, we determine that these are functionally significant regulatory changes, as nearby target genes are up- or down-regulated. In conclusion, we show that FAIRE-seq and ATAC-seq based open chromatin profiling

  6. Changes in chromatin state reveal ARNT2 at a node of a tumorigenic transcription factor signature driving glioblastoma cell aggressiveness.

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    Bogeas, Alexandra; Morvan-Dubois, Ghislaine; El-Habr, Elias A; Lejeune, François-Xavier; Defrance, Matthieu; Narayanan, Ashwin; Kuranda, Klaudia; Burel-Vandenbos, Fanny; Sayd, Salwa; Delaunay, Virgile; Dubois, Luiz G; Parrinello, Hugues; Rialle, Stéphanie; Fabrega, Sylvie; Idbaih, Ahmed; Haiech, Jacques; Bièche, Ivan; Virolle, Thierry; Goodhardt, Michele; Chneiweiss, Hervé; Junier, Marie-Pierre

    2018-02-01

    Although a growing body of evidence indicates that phenotypic plasticity exhibited by glioblastoma cells plays a central role in tumor development and post-therapy recurrence, the master drivers of their aggressiveness remain elusive. Here we mapped the changes in active (H3K4me3) and repressive (H3K27me3) histone modifications accompanying the repression of glioblastoma stem-like cells tumorigenicity. Genes with changing histone marks delineated a network of transcription factors related to cancerous behavior, stem state, and neural development, highlighting a previously unsuspected association between repression of ARNT2 and loss of cell tumorigenicity. Immunohistochemistry confirmed ARNT2 expression in cell sub-populations within proliferative zones of patients' glioblastoma. Decreased ARNT2 expression was consistently observed in non-tumorigenic glioblastoma cells, compared to tumorigenic cells. Moreover, ARNT2 expression correlated with a tumorigenic molecular signature at both the tissue level within the tumor core and at the single cell level in the patients' tumors. We found that ARNT2 knockdown decreased the expression of SOX9, POU3F2 and OLIG2, transcription factors implicated in glioblastoma cell tumorigenicity, and repressed glioblastoma stem-like cell tumorigenic properties in vivo. Our results reveal ARNT2 as a pivotal component of the glioblastoma cell tumorigenic signature, located at a node of a transcription factor network controlling glioblastoma cell aggressiveness.

  7. Global transcriptome analysis reveals extensive gene remodeling, alternative splicing and differential transcription profiles in non-seed vascular plant Selaginella moellendorffii.

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    Zhu, Yan; Chen, Longxian; Zhang, Chengjun; Hao, Pei; Jing, Xinyun; Li, Xuan

    2017-01-25

    Selaginella moellendorffii, a lycophyte, is a model plant to study the early evolution and development of vascular plants. As the first and only sequenced lycophyte to date, the genome of S. moellendorffii revealed many conserved genes and pathways, as well as specialized genes different from flowering plants. Despite the progress made, little is known about long noncoding RNAs (lncRNA) and the alternative splicing (AS) of coding genes in S. moellendorffii. Its coding gene models have not been fully validated with transcriptome data. Furthermore, it remains important to understand whether the regulatory mechanisms similar to flowering plants are used, and how they operate in a non-seed primitive vascular plant. RNA-sequencing (RNA-seq) was performed for three S. moellendorffii tissues, root, stem, and leaf, by constructing strand-specific RNA-seq libraries from RNA purified using RiboMinus isolation protocol. A total of 176 million reads (44 Gbp) were obtained from three tissue types, and were mapped to S. moellendorffii genome. By comparing with 22,285 existing gene models of S. moellendorffii, we identified 7930 high-confidence novel coding genes (a 35.6% increase), and for the first time reported 4422 lncRNAs in a lycophyte. Further, we refined 2461 (11.0%) of existing gene models, and identified 11,030 AS events (for 5957 coding genes) revealed for the first time for lycophytes. Tissue-specific gene expression with functional implication was analyzed, and 1031, 554, and 269 coding genes, and 174, 39, and 17 lncRNAs were identified in root, stem, and leaf tissues, respectively. The expression of critical genes for vascular development stages, i.e. formation of provascular cells, xylem specification and differentiation, and phloem specification and differentiation, was compared in S. moellendorffii tissues, indicating a less complex regulatory mechanism in lycophytes than in flowering plants. The results were further strengthened by the evolutionary trend of

  8. Genetic variation in metallothionein and metal-regulatory transcription factor 1 in relation to urinary cadmium, copper, and zinc

    International Nuclear Information System (INIS)

    Adams, Scott V.; Barrick, Brian; Christopher, Emily P.; Shafer, Martin M.; Makar, Karen W.; Song, Xiaoling; Lampe, Johanna W.; Vilchis, Hugo; Ulery, April; Newcomb, Polly A.

    2015-01-01

    Background: Metallothionein (MT) proteins play critical roles in the physiological handling of both essential (Cu and Zn) and toxic (Cd) metals. MT expression is regulated by metal-regulatory transcription factor 1 (MTF1). Hence, genetic variation in the MT gene family and MTF1 might influence excretion of these metals. Methods: 321 women were recruited in Seattle, WA and Las Cruces, NM and provided demographic information, urine samples for measurement of metal concentrations by mass spectrometry and creatinine, and blood or saliva for extraction of DNA. Forty-one single nucleotide polymorphisms (SNPs) within the MTF1 gene region and the region of chromosome 16 encoding the MT gene family were selected for genotyping in addition to an ancestry informative marker panel. Linear regression was used to estimate the association of SNPs with urinary Cd, Cu, and Zn, adjusted for age, urinary creatinine, smoking history, study site, and ancestry. Results: Minor alleles of rs28366003 and rs10636 near the MT2A gene were associated with lower urinary Cd, Cu, and Zn. Minor alleles of rs8044719 and rs1599823, near MT1A and MT1B, were associated with lower urinary Cd and Zn, respectively. Minor alleles of rs4653329 in MTF1 were associated with lower urinary Cd. Conclusions: These results suggest that genetic variation in the MT gene region and MTF1 influences urinary Cd, Cu, and Zn excretion. - Highlights: • Genetic variation in metallothionein (MT) genes was assessed in two diverse populations. • Single nucleotide polymorphisms (SNPs) in MT genes were associated with mean urinary Cd, Cu and Zn. • Genetic variation may influence biomarkers of exposure, and associations of exposure with health.

  9. Genetic variation in metallothionein and metal-regulatory transcription factor 1 in relation to urinary cadmium, copper, and zinc

    Energy Technology Data Exchange (ETDEWEB)

    Adams, Scott V., E-mail: sadams@fhcrc.org [Public Health Sciences Division, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave N, Seattle, WA 98109 (United States); Barrick, Brian [Department of Plant and Environmental Sciences, New Mexico State University, Box 30003 MSC 3Q, Las Cruces, NM 88003 (United States); Christopher, Emily P. [Public Health Sciences Division, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave N, Seattle, WA 98109 (United States); Shafer, Martin M. [Environmental Chemistry and Technology, Wisconsin State Laboratory of Hygiene, University of Wisconsin, 2601 Agriculture Dr., Madison, WI 53718 (United States); Makar, Karen W.; Song, Xiaoling [Public Health Science Biomarker Laboratory, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave N, Seattle, WA 98109 (United States); Lampe, Johanna W. [Public Health Sciences Division, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave N, Seattle, WA 98109 (United States); Vilchis, Hugo [Border Epidemiology and Environmental Health Center, New Mexico State University, Box 30001 MSC 3BEC, Las Cruces, NM 88003 (United States); Ulery, April [Department of Plant and Environmental Sciences, New Mexico State University, Box 30003 MSC 3Q, Las Cruces, NM 88003 (United States); Newcomb, Polly A. [Public Health Sciences Division, Fred Hutchinson Cancer Research Center, 1100 Fairview Ave N, Seattle, WA 98109 (United States)

    2015-12-15

    Background: Metallothionein (MT) proteins play critical roles in the physiological handling of both essential (Cu and Zn) and toxic (Cd) metals. MT expression is regulated by metal-regulatory transcription factor 1 (MTF1). Hence, genetic variation in the MT gene family and MTF1 might influence excretion of these metals. Methods: 321 women were recruited in Seattle, WA and Las Cruces, NM and provided demographic information, urine samples for measurement of metal concentrations by mass spectrometry and creatinine, and blood or saliva for extraction of DNA. Forty-one single nucleotide polymorphisms (SNPs) within the MTF1 gene region and the region of chromosome 16 encoding the MT gene family were selected for genotyping in addition to an ancestry informative marker panel. Linear regression was used to estimate the association of SNPs with urinary Cd, Cu, and Zn, adjusted for age, urinary creatinine, smoking history, study site, and ancestry. Results: Minor alleles of rs28366003 and rs10636 near the MT2A gene were associated with lower urinary Cd, Cu, and Zn. Minor alleles of rs8044719 and rs1599823, near MT1A and MT1B, were associated with lower urinary Cd and Zn, respectively. Minor alleles of rs4653329 in MTF1 were associated with lower urinary Cd. Conclusions: These results suggest that genetic variation in the MT gene region and MTF1 influences urinary Cd, Cu, and Zn excretion. - Highlights: • Genetic variation in metallothionein (MT) genes was assessed in two diverse populations. • Single nucleotide polymorphisms (SNPs) in MT genes were associated with mean urinary Cd, Cu and Zn. • Genetic variation may influence biomarkers of exposure, and associations of exposure with health.

  10. Construction and analysis of the transcription factor-microRNA co-regulatory network response to Mycobacterium tuberculosis: a view from the blood.

    Science.gov (United States)

    Lin, Yan; Duan, Zipeng; Xu, Feng; Zhang, Jiayuan; Shulgina, Marina V; Li, Fan

    2017-01-01

    Mycobacterium tuberculosis ( Mtb ) infection has been regional outbreak, recently. The traditional focus on the patterns of "reductionism" which was associated with single molecular changes has been unable to meet the demand of early diagnosis and clinical application when current tuberculosis infection happened. In this study, we employed a systems biology approach to collect large microarray data sets including mRNAs and microRNAs (miRNAs) to identify the differentially expressed mRNAs and miRNAs in the whole blood of TB patients. The aim was to identify key genes associated with the immune response in the pathogenic process of tuberculosis by analyzing the co-regulatory network that was consisted of transcription factors and miRNAs as well as their target genes. The network along with their co-regulatory genes was analyzed utilizing Transcriptional Regulatory Element Database (TRED) and Database for Annotation, Visualization and Integrated Discovery (DAVID). We got 21 (19 up-regulated and 2 down-regulated) differentially expressed genes that were co-regulated by transcription factors and miRNAs. KEGG pathway enrichment analysis showed that the 21 differentially expressed genes were predominantly involved in Tuberculosis signaling pathway, which may play a major role in tuberculosis biological process. Quantitative real-time PCR was performed to verify the over expression of co-regulatory genes ( FCGR1A and CEBPB ). The genetic expression was correlated with clinicopathological characteristics in TB patients and inferences drawn. Our results suggest the TF-miRNA gene co-regulatory network may help us further understand the molecular mechanism of immune response to tuberculosis and provide us a new angle of future biomarker and therapeutic targets.

  11. Logic programming reveals alteration of key transcription factors in multiple myeloma.

    Science.gov (United States)

    Miannay, Bertrand; Minvielle, Stéphane; Roux, Olivier; Drouin, Pierre; Avet-Loiseau, Hervé; Guérin-Charbonnel, Catherine; Gouraud, Wilfried; Attal, Michel; Facon, Thierry; Munshi, Nikhil C; Moreau, Philippe; Campion, Loïc; Magrangeas, Florence; Guziolowski, Carito

    2017-08-23

    Innovative approaches combining regulatory networks (RN) and genomic data are needed to extract biological information for a better understanding of diseases, such as cancer, by improving the identification of entities and thereby leading to potential new therapeutic avenues. In this study, we confronted an automatically generated RN with gene expression profiles (GEP) from a cohort of multiple myeloma (MM) patients and normal individuals using global reasoning on the RN causality to identify key-nodes. We modeled each patient by his or her GEP, the RN and the possible automatically detected repairs needed to establish a coherent flow of the information that explains the logic of the GEP. These repairs could represent cancer mutations leading to GEP variability. With this reasoning, unmeasured protein states can be inferred, and we can simulate the impact of a protein perturbation on the RN behavior to identify therapeutic targets. We showed that JUN/FOS and FOXM1 activities are altered in almost all MM patients and identified two survival markers for MM patients. Our results suggest that JUN/FOS-activation has a strong impact on the RN in view of the whole GEP, whereas FOXM1-activation could be an interesting way to perturb an MM subgroup identified by our method.

  12. Transcriptional analysis of the HeT-A retrotransposon in mutant and wild type stocks reveals high sequence variability at Drosophila telomeres and other unusual features

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    Piñeyro David

    2011-11-01

    Full Text Available Abstract Background Telomere replication in Drosophila depends on the transposition of a domesticated retroelement, the HeT-A retrotransposon. The sequence of the HeT-A retrotransposon changes rapidly resulting in differentiated subfamilies. This pattern of sequence change contrasts with the essential function with which the HeT-A is entrusted and brings about questions concerning the extent of sequence variability, the telomere contribution of different subfamilies, and whether wild type and mutant Drosophila stocks show different HeT-A scenarios. Results A detailed study on the variability of HeT-A reveals that both the level of variability and the number of subfamilies are higher than previously reported. Comparisons between GIII, a strain with longer telomeres, and its parental strain Oregon-R indicate that both strains have the same set of HeT-A subfamilies. Finally, the presence of a highly conserved splicing pattern only in its antisense transcripts indicates a putative regulatory, functional or structural role for the HeT-A RNA. Interestingly, our results also suggest that most HeT-A copies are actively expressed regardless of which telomere and where in the telomere they are located. Conclusions Our study demonstrates how the HeT-A sequence changes much faster than previously reported resulting in at least nine different subfamilies most of which could actively contribute to telomere extension in Drosophila. Interestingly, the only significant difference observed between Oregon-R and GIII resides in the nature and proportion of the antisense transcripts, suggesting a possible mechanism that would in part explain the longer telomeres of the GIII stock.

  13. Transcriptional profiling reveals the expression of novel genes in response to various stimuli in the human dermatophyte Trichophyton rubrum

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    Aquino-Ferreira Roseli

    2010-02-01

    Full Text Available Abstract Background Cutaneous mycoses are common human infections among healthy and immunocompromised hosts, and the anthropophilic fungus Trichophyton rubrum is the most prevalent microorganism isolated from such clinical cases worldwide. The aim of this study was to determine the transcriptional profile of T. rubrum exposed to various stimuli in order to obtain insights into the responses of this pathogen to different environmental challenges. Therefore, we generated an expressed sequence tag (EST collection by constructing one cDNA library and nine suppression subtractive hybridization libraries. Results The 1388 unigenes identified in this study were functionally classified based on the Munich Information Center for Protein Sequences (MIPS categories. The identified proteins were involved in transcriptional regulation, cellular defense and stress, protein degradation, signaling, transport, and secretion, among other functions. Analysis of these unigenes revealed 575 T. rubrum sequences that had not been previously deposited in public databases. Conclusion In this study, we identified novel T. rubrum genes that will be useful for ORF prediction in genome sequencing and facilitating functional genome analysis. Annotation of these expressed genes revealed metabolic adaptations of T. rubrum to carbon sources, ambient pH shifts, and various antifungal drugs used in medical practice. Furthermore, challenging T. rubrum with cytotoxic drugs and ambient pH shifts extended our understanding of the molecular events possibly involved in the infectious process and resistance to antifungal drugs.

  14. Common and distinct DNA-binding and regulatory activities of the BEN-solo transcription factor family.

    Science.gov (United States)

    Dai, Qi; Ren, Aiming; Westholm, Jakub O; Duan, Hong; Patel, Dinshaw J; Lai, Eric C

    2015-01-01

    Recently, the BEN (BANP, E5R, and NAC1) domain was recognized as a new class of conserved DNA-binding domain. The fly genome encodes three proteins that bear only a single BEN domain ("BEN-solo" factors); namely, Insensitive (Insv), Bsg25A (Elba1), and CG9883 (Elba2). Insv homodimers preferentially bind CCAATTGG palindromes throughout the genome to mediate transcriptional repression, whereas Bsg25A and Elba2 heterotrimerize with their obligate adaptor, Elba3 (i.e., the ELBA complex), to recognize a CCAATAAG motif in the Fab-7 insulator. While these data suggest distinct DNA-binding properties of BEN-solo proteins, we performed reporter assays that indicate that both Bsg25A and Elba2 can individually recognize Insv consensus sites efficiently. We confirmed this by solving the structure of Bsg25A complexed to the Insv site, which showed that key aspects of the BEN:DNA recognition strategy are similar between these proteins. We next show that both Insv and ELBA proteins are competent to mediate transcriptional repression via Insv consensus sequences but that the ELBA complex appears to be selective for the ELBA site. Reciprocally, genome-wide analysis reveals that Insv exhibits significant cobinding to class I insulator elements, indicating that it may also contribute to insulator function. Indeed, we observed abundant Insv binding within the Hox complexes with substantial overlaps with class I insulators, many of which bear Insv consensus sites. Moreover, Insv coimmunoprecipitates with the class I insulator factor CP190. Finally, we observed that Insv harbors exclusive activity among fly BEN-solo factors with respect to regulation of Notch-mediated cell fate choices in the peripheral nervous system. This in vivo activity is recapitulated by BEND6, a mammalian BEN-solo factor that conserves the Notch corepressor function of Insv but not its capacity to bind Insv consensus sites. Altogether, our data define an array of common and distinct biochemical and functional

  15. Alu-miRNA interactions modulate transcript isoform diversity in stress response and reveal signatures of positive selection

    Science.gov (United States)

    Pandey, Rajesh; Bhattacharya, Aniket; Bhardwaj, Vivek; Jha, Vineet; Mandal, Amit K.; Mukerji, Mitali

    2016-09-01

    Primate-specific Alus harbor different regulatory features, including miRNA targets. In this study, we provide evidence for miRNA-mediated modulation of transcript isoform levels during heat-shock response through exaptation of Alu-miRNA sites in mature mRNA. We performed genome-wide expression profiling coupled with functional validation of miRNA target sites within exonized Alus, and analyzed conservation of these targets across primates. We observed that two miRNAs (miR-15a-3p and miR-302d-3p) elevated in stress response, target RAD1, GTSE1, NR2C1, FKBP9 and UBE2I exclusively within Alu. These genes map onto the p53 regulatory network. Ectopic overexpression of miR-15a-3p downregulates GTSE1 and RAD1 at the protein level and enhances cell survival. This Alu-mediated fine-tuning seems to be unique to humans as evident from the absence of orthologous sites in other primate lineages. We further analyzed signatures of selection on Alu-miRNA targets in the genome, using 1000 Genomes Phase-I data. We found that 198 out of 3177 Alu-exonized genes exhibit signatures of selection within Alu-miRNA sites, with 60 of them containing SNPs supported by multiple evidences (global-FST > 0.3, pair-wise-FST > 0.5, Fay-Wu’s H  2.0, high ΔDAF) and implicated in p53 network. We propose that by affecting multiple genes, Alu-miRNA interactions have the potential to facilitate population-level adaptations in response to environmental challenges.

  16. Coxiella burnetii transcriptional analysis reveals serendipity clusters of regulation in intracellular bacteria.

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    Quentin Leroy

    Full Text Available Coxiella burnetii, the causative agent of the zoonotic disease Q fever, is mainly transmitted to humans through an aerosol route. A spore-like form allows C. burnetii to resist different environmental conditions. Because of this, analysis of the survival strategies used by this bacterium to adapt to new environmental conditions is critical for our understanding of C. burnetii pathogenicity. Here, we report the early transcriptional response of C. burnetii under temperature stresses. Our data show that C. burnetii exhibited minor changes in gene regulation under short exposure to heat or cold shock. While small differences were observed, C. burnetii seemed to respond similarly to cold and heat shock. The expression profiles obtained using microarrays produced in-house were confirmed by quantitative RT-PCR. Under temperature stresses, 190 genes were differentially expressed in at least one condition, with a fold change of up to 4. Globally, the differentially expressed genes in C. burnetii were associated with bacterial division, (pppGpp synthesis, wall and membrane biogenesis and, especially, lipopolysaccharide and peptidoglycan synthesis. These findings could be associated with growth arrest and witnessed transformation of the bacteria to a spore-like form. Unexpectedly, clusters of neighboring genes were differentially expressed. These clusters do not belong to operons or genetic networks; they have no evident associated functions and are not under the control of the same promoters. We also found undescribed but comparable clusters of regulation in previously reported transcriptomic analyses of intracellular bacteria, including Rickettsia sp. and Listeria monocytogenes. The transcriptomic patterns of C. burnetii observed under temperature stresses permits the recognition of unpredicted clusters of regulation for which the trigger mechanism remains unidentified but which may be the result of a new mechanism of epigenetic regulation.

  17. Transcriptional changes associated with resistance to inhibitors of epidermal growth factor receptor revealed using metaanalysis

    International Nuclear Information System (INIS)

    Younis, Sidra; Javed, Qamar; Blumenberg, Miroslav

    2015-01-01

    EGFR is important in maintaining metabolic homeostasis in healthy cells, but in tumors it activates downstream signaling pathways, causing proliferation, angiogenesis, invasion and metastasis. Consequently, EGFR is targeted in cancers using reversible, irreversible or antibody inhibitors. Unfortunately, tumors develop inhibitor resistance by mutations or overexpressing EGFR, or its ligand, or activating secondary, EGFR-independent pathways. Here we present a global metaanalysis comparing transcriptional profiles from matched pairs of EGFR inhibitor-sensitive vs. -resistant cell lines, using 15 datasets comprising 274 microarrays. We also analyzed separately pairs of cell lines derived using reversible, irreversible or antibody inhibitors. The metaanalysis identifies commonalities in cell lines resistant to EGFR inhibitors: in sensitive cell lines, the ontological categories involving the ErbB receptors pathways, cell adhesion and lipid metabolism are overexpressed; however, resistance to EGFR inhibitors is associated with overexpression of genes for ErbB receptors-independent oncogenic pathways, regulation of cell motility, energy metabolism, immunity especially inflammatory cytokines biosynthesis, cell cycle and responses to exogenous and endogenous stimuli. Specifically in Gefitinib-resistant cell lines, the immunity-associated genes are overexpressed, whereas in Erlotinib-resistant ones so are the mitochondrial genes and processes. Unexpectedly, lines selected using EGFR-targeting antibodies overexpress different gene ontologies from ones selected using kinase inhibitors. Specifically, they have reduced expression of genes for proliferation, chemotaxis, immunity and angiogenesis. This metaanalysis suggests that ‘combination therapies’ can improve cancer treatment outcomes. Potentially, use of mitochondrial blockers with Erlotinib, immunity blockers with Gefitinib, tyrosine kinase inhibitors with antibody inhibitors, may have better chance of avoiding

  18. ChIPBase v2.0: decoding transcriptional regulatory networks of non-coding RNAs and protein-coding genes from ChIP-seq data.

    Science.gov (United States)

    Zhou, Ke-Ren; Liu, Shun; Sun, Wen-Ju; Zheng, Ling-Ling; Zhou, Hui; Yang, Jian-Hua; Qu, Liang-Hu

    2017-01-04

    The abnormal transcriptional regulation of non-coding RNAs (ncRNAs) and protein-coding genes (PCGs) is contributed to various biological processes and linked with human diseases, but the underlying mechanisms remain elusive. In this study, we developed ChIPBase v2.0 (http://rna.sysu.edu.cn/chipbase/) to explore the transcriptional regulatory networks of ncRNAs and PCGs. ChIPBase v2.0 has been expanded with ∼10 200 curated ChIP-seq datasets, which represent about 20 times expansion when comparing to the previous released version. We identified thousands of binding motif matrices and their binding sites from ChIP-seq data of DNA-binding proteins and predicted millions of transcriptional regulatory relationships between transcription factors (TFs) and genes. We constructed 'Regulator' module to predict hundreds of TFs and histone modifications that were involved in or affected transcription of ncRNAs and PCGs. Moreover, we built a web-based tool, Co-Expression, to explore the co-expression patterns between DNA-binding proteins and various types of genes by integrating the gene expression profiles of ∼10 000 tumor samples and ∼9100 normal tissues and cell lines. ChIPBase also provides a ChIP-Function tool and a genome browser to predict functions of diverse genes and visualize various ChIP-seq data. This study will greatly expand our understanding of the transcriptional regulations of ncRNAs and PCGs. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. Menin and RNF20 recruitment is associated with dynamic histone modifications that regulate signal transducer and activator of transcription 1 (STAT1-activated transcription of the interferon regulatory factor 1 gene (IRF1

    Directory of Open Access Journals (Sweden)

    Buro Lauren J

    2010-09-01

    Full Text Available Abstract Background Signal transducer and activator of transcription (STAT activation of gene expression is both rapid and transient, and when properly executed it affects growth, differentiation, homeostasis and the immune response, but when dysregulated it contributes to human disease. Transcriptional activation is regulated by alterations to the chromatin template. However, the role of histone modification at gene loci that are activated for transcription in response to STAT signaling is poorly defined. Results Using chromatin immunoprecipitation, we profiled several histone modifications during STAT1 activation of the interferon regulatory factor 1 gene (IRF1. Methylated lysine histone proteins H3K4me2, H3K4me3, H3K79me3, H3K36me3 and monoubiquitinated histone ubH2B are dynamic and correlate with interferon (IFNγ induction of STAT1 activity. Chemical inhibition of H3K4 methylation downregulates IRF1 transcription and decreases RNA polymerase II (Pol II occupancy at the IRF1 promoter. MEN1, a component of a complex proteins associated with Set1 (COMPASS-like complex and the hBRE1 component, RNF20, are localized to IRF1 in the uninduced state and are further recruited when IRF1 is activated. RNAi-mediated depletion of RNF20 lowers both ubH2B and H3K4me3, but surprisingly, upregulates IFNγ induced IRF1 transcription. The dynamics of phosphorylation in the C-terminal domain (CTD of Pol II are disrupted during gene activation as well. Conclusions H2B monoubiquitination promotes H3K4 methylation, but the E3 ubiquitin ligase, RNF20, is repressive of inducible transcription at the IRF1 gene locus, suggesting that ubH2B can, directly or indirectly, affect Pol II CTD phosphorylation cycling to exert control on ongoing transcription.

  20. Multi-tissue omics analyses reveal molecular regulatory networks for puberty in composite beef cattle

    Science.gov (United States)

    Puberty is a complex physiological event by which animals mature into an adult capable of sexual reproduction. In order to enhance our understanding of the genes and regulatory pathways and networks involved in puberty, we characterized the transcriptome of five reproductive tissues (i.e., hypothal...

  1. CRX ChIP-seq reveals the cis-regulatory architecture of mouse photoreceptors

    NARCIS (Netherlands)

    J.C. Corbo (Joseph); K.A. Lawrence (Karen); M. Karlstetter (Marcus); C.A. Myers (Connie); M. Abdelaziz (Musa); W. Dirkes (William); K. Weigelt (Karin); M. Seifert (Martin); V. Benes (Vladimir); L.G. Fritsche (Lars); B.H.F. Weber (Bernhard); T. Langmann (Thomas)

    2010-01-01

    textabstractApproximately 98% of mammalian DNA is noncoding, yet we understand relatively little about the function of this enigmatic portion of the genome. The cis-regulatory elements that control gene expression reside in noncoding regions and can be identified by mapping the binding sites of

  2. A Proteome-wide, Quantitative Survey of In Vivo Ubiquitylation Sites Reveals Widespread Regulatory Roles

    DEFF Research Database (Denmark)

    Wagner, Sebastian Alexander; Beli, Petra; Weinert, Brian Tate

    2011-01-01

    Post-translational modification of proteins by ubiquitin is a fundamentally important regulatory mechanism. However, proteome-wide analysis of endogenous ubiquitylation remains a challenging task, and almost always has relied on cells expressing affinity tagged ubiquitin. Here we combine single...

  3. A Transcriptional Regulatory Network Containing Nuclear Receptors and Long Noncoding RNAs Controls Basal and Drug-Induced Expression of Cytochrome P450s in HepaRG Cells.

    Science.gov (United States)

    Chen, Liming; Bao, Yifan; Piekos, Stephanie C; Zhu, Kexin; Zhang, Lirong; Zhong, Xiao-Bo

    2018-07-01

    Cytochrome P450 (P450) enzymes are responsible for metabolizing drugs. Expression of P450s can directly affect drug metabolism, resulting in various outcomes in therapeutic efficacy and adverse effects. Several nuclear receptors are transcription factors that can regulate expression of P450s at both basal and drug-induced levels. Some long noncoding RNAs (lncRNAs) near a transcription factor are found to participate in the regulatory functions of the transcription factors. The aim of this study is to determine whether there is a transcriptional regulatory network containing nuclear receptors and lncRNAs controlling both basal and drug-induced expression of P450s in HepaRG cells. Small interfering RNAs or small hairpin RNAs were applied to knock down four nuclear receptors [hepatocyte nuclear factor 1 α (HNF1 α ), hepatocyte nuclear factor 4 α (HNF4 α ), pregnane X receptor (PXR), and constitutive androstane receptor (CAR)] as well as two lncRNAs [HNF1 α antisense RNA 1 (HNF1 α -AS1) and HNF4 α antisense RNA 1 (HNF4 α -AS1)] in HepaRG cells with or without treatment of phenobarbital or rifampicin. Expression of eight P450 enzymes was examined in both basal and drug-induced levels. CAR and PXR mainly regulated expression of specific P450s. HNF1 α and HNF4 α affected expression of a wide range of P450s as well as other transcription factors. HNF1 α and HNF4 α controlled the expression of their neighborhood lncRNAs, HNF1 α -AS1 and HNF4 α -AS1, respectively. HNF1 α -AS1 and HNF4 α -AS1 was also involved in the regulation of P450s and transcription factors in diverse manners. Altogether, our study concludes that a transcription regulatory network containing the nuclear receptors and lncRNAs controls both basal and drug-induced expression of P450s in HepaRG cells. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

  4. Transcriptional response of zebrafish embryos exposed to neurotoxic compounds reveals a muscle activity dependent hspb11 expression.

    Directory of Open Access Journals (Sweden)

    Nils Klüver

    Full Text Available Acetylcholinesterase (AChE inhibitors are widely used as pesticides and drugs. Their primary effect is the overstimulation of cholinergic receptors which results in an improper muscular function. During vertebrate embryonic development nerve activity and intracellular downstream events are critical for the regulation of muscle fiber formation. Whether AChE inhibitors and related neurotoxic compounds also provoke specific changes in gene transcription patterns during vertebrate development that allow them to establish a mechanistic link useful for identification of developmental toxicity pathways has, however, yet not been investigated. Therefore we examined the transcriptomic response of a known AChE inhibitor, the organophosphate azinphos-methyl (APM, in zebrafish embryos and compared the response with two non-AChE inhibiting unspecific control compounds, 1,4-dimethoxybenzene (DMB and 2,4-dinitrophenol (DNP. A highly specific cluster of APM induced gene transcripts was identified and a subset of strongly regulated genes was analyzed in more detail. The small heat shock protein hspb11 was found to be the most sensitive induced gene in response to AChE inhibitors. Comparison of expression in wildtype, ache and sop(fixe mutant embryos revealed that hspb11 expression was dependent on the nicotinic acetylcholine receptor (nAChR activity. Furthermore, modulators of intracellular calcium levels within the whole embryo led to a transcriptional up-regulation of hspb11 which suggests that elevated intracellular calcium levels may regulate the expression of this gene. During early zebrafish development, hspb11 was specifically expressed in muscle pioneer cells and Hspb11 morpholino-knockdown resulted in effects on slow muscle myosin organization. Our findings imply that a comparative toxicogenomic approach and functional analysis can lead to the identification of molecular mechanisms and specific marker genes for potential neurotoxic compounds.

  5. Comparative analyses of six solanaceous transcriptomes reveal a high degree of sequence conservation and species-specific transcripts

    Directory of Open Access Journals (Sweden)

    Ouyang Shu

    2005-09-01

    Full Text Available Abstract Background The Solanaceae is a family of closely related species with diverse phenotypes that have been exploited for agronomic purposes. Previous studies involving a small number of genes suggested sequence conservation across the Solanaceae. The availability of large collections of Expressed Sequence Tags (ESTs for the Solanaceae now provides the opportunity to assess sequence conservation and divergence on a genomic scale. Results All available ESTs and Expressed Transcripts (ETs, 449,224 sequences for six Solanaceae species (potato, tomato, pepper, petunia, tobacco and Nicotiana benthamiana, were clustered and assembled into gene indices. Examination of gene ontologies revealed that the transcripts within the gene indices encode a similar suite of biological processes. Although the ESTs and ETs were derived from a variety of tissues, 55–81% of the sequences had significant similarity at the nucleotide level with sequences among the six species. Putative orthologs could be identified for 28–58% of the sequences. This high degree of sequence conservation was supported by expression profiling using heterologous hybridizations to potato cDNA arrays that showed similar expression patterns in mature leaves for all six solanaceous species. 16–19% of the transcripts within the six Solanaceae gene indices did not have matches among Solanaceae, Arabidopsis, rice or 21 other plant gene indices. Conclusion Results from this genome scale analysis confirmed a high level of sequence conservation at the nucleotide level of the coding sequence among Solanaceae. Additionally, the results indicated that part of the Solanaceae transcriptome is likely to be unique for each species.

  6. Transcriptome Analysis of Three Sheep Intestinal Regions reveals Key Pathways and Hub Regulatory Genes of Large Intestinal Lipid Metabolism.

    Science.gov (United States)

    Chao, Tianle; Wang, Guizhi; Ji, Zhibin; Liu, Zhaohua; Hou, Lei; Wang, Jin; Wang, Jianmin

    2017-07-13

    The large intestine, also known as the hindgut, is an important part of the animal digestive system. Recent studies on digestive system development in ruminants have focused on the rumen and the small intestine, but the molecular mechanisms underlying sheep large intestine metabolism remain poorly understood. To identify genes related to intestinal metabolism and to reveal molecular regulation mechanisms, we sequenced and compared the transcriptomes of mucosal epithelial tissues among the cecum, proximal colon and duodenum. A total of 4,221 transcripts from 3,254 genes were identified as differentially expressed transcripts. Between the large intestine and duodenum, differentially expressed transcripts were found to be significantly enriched in 6 metabolism-related pathways, among which PPAR signaling was identified as a key pathway. Three genes, CPT1A, LPL and PCK1, were identified as higher expression hub genes in the large intestine. Between the cecum and colon, differentially expressed transcripts were significantly enriched in 5 lipid metabolism related pathways, and CEPT1 and MBOAT1 were identified as hub genes. This study provides important information regarding the molecular mechanisms of intestinal metabolism in sheep and may provide a basis for further study.

  7. Characterization of WRKY transcription factors in Solanum lycopersicum reveals collinearity and their expression patterns under cold treatment.

    Science.gov (United States)

    Chen, Lin; Yang, Yang; Liu, Can; Zheng, Yanyan; Xu, Mingshuang; Wu, Na; Sheng, Jiping; Shen, Lin

    2015-08-28

    WRKY transcription factors play an important role in cold defense of plants. However, little information is available about the cold-responsive WRKYs in tomato (Solanum lycopersicum). In the present study, a complete characterization of this gene family was described. Eighty WRKY genes in the tomato genome were identified. Almost all WRKY genes contain putative stress-responsive cis-elements in their promoter regions. Segmental duplications contributed significantly to the expansion of the SlWRKY gene family. Transcriptional analysis revealed notable differential expression in tomato tissues and expression patterns under cold stress, which indicated wide functional divergence in this family. Ten WRKYs in tomato were strongly induced more than 2-fold during cold stress. These genes represented candidate genes for future functional analysis of WRKYs involved in the cold-related signal pathways. Our data provide valuable information about tomato WRKY proteins and form a foundation for future studies of these proteins, especially for those that play an important role in response to cold stress. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. A Knockout Screen of ApiAP2 Genes Reveals Networks of Interacting Transcriptional Regulators Controlling the Plasmodium Life Cycle.

    Science.gov (United States)

    Modrzynska, Katarzyna; Pfander, Claudia; Chappell, Lia; Yu, Lu; Suarez, Catherine; Dundas, Kirsten; Gomes, Ana Rita; Goulding, David; Rayner, Julian C; Choudhary, Jyoti; Billker, Oliver

    2017-01-11

    A family of apicomplexa-specific proteins containing AP2 DNA-binding domains (ApiAP2s) was identified in malaria parasites. This family includes sequence-specific transcription factors that are key regulators of development. However, functions for the majority of ApiAP2 genes remain unknown. Here, a systematic knockout screen in Plasmodium berghei identified ten ApiAP2 genes that were essential for mosquito transmission: four were critical for the formation of infectious ookinetes, and three were required for sporogony. We describe non-essential functions for AP2-O and AP2-SP proteins in blood stages, and identify AP2-G2 as a repressor active in both asexual and sexual stages. Comparative transcriptomics across mutants and developmental stages revealed clusters of co-regulated genes with shared cis promoter elements, whose expression can be controlled positively or negatively by different ApiAP2 factors. We propose that stage-specific interactions between ApiAP2 proteins on partly overlapping sets of target genes generate the complex transcriptional network that controls the Plasmodium life cycle. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  9. Hepatitis C virus nonstructural protein-5A activates sterol regulatory element-binding protein-1c through transcription factor Sp1

    Energy Technology Data Exchange (ETDEWEB)

    Xiang, Zhonghua; Qiao, Ling; Zhou, Yan [Vaccine and Infectious Disease Organization, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E3 (Canada); Babiuk, Lorne A. [University of Alberta, Edmonton, Alberta (Canada); Liu, Qiang, E-mail: qiang.liu@usask.ca [Vaccine and Infectious Disease Organization, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E3 (Canada)

    2010-11-19

    Research highlights: {yields} A chimeric subgenomic HCV replicon expresses HCV-3a NS5A in an HCV-1b backbone. {yields} HCV-3a NS5A increases mature SREBP-1c protein level. {yields} HCV-3a NS5A activates SREBP-1c transcription. {yields} Domain II of HCV-3a NS5A is more effective in SREBP-1c promoter activation. {yields} Transcription factor Sp1 is required for SREBP-1c activation by HCV-3a NS5A. -- Abstract: Steatosis is an important clinical manifestation of hepatitis C virus (HCV) infection. The molecular mechanisms of HCV-associated steatosis are not well understood. Sterol regulatory element-binding protein-1c (SREBP-1c) is a key transcription factor which activates the transcription of lipogenic genes. Here we showed that the nuclear, mature SREBP-1c level increases in the nucleus of replicon cells expressing HCV-3a nonstructural protein-5A (NS5A). We further showed that HCV-3a NS5A up-regulates SREBP-1c transcription. Additional analysis showed that transcriptional factor Sp1 is involved in SREBP-1c activation by HCV-3a NS5A because inhibition of Sp1 activity by mithramycin A or a dominant-negative Sp1 construct abrogated SREBP-1c promoter activation by HCV-3a NS5A. In addition, chromatin immunoprecipitation (ChIP) assay demonstrated enhanced binding of Sp1 on the SREBP-1c promoter in HCV-3a NS5A replicon cells. These results showed that HCV-3a NS5A activates SREBP-1c transcription through Sp1. Taken together, our results suggest that HCV-3a NS5A is a contributing factor for steatosis caused by HCV-3a infection.

  10. Hepatitis C virus nonstructural protein-5A activates sterol regulatory element-binding protein-1c through transcription factor Sp1

    International Nuclear Information System (INIS)

    Xiang, Zhonghua; Qiao, Ling; Zhou, Yan; Babiuk, Lorne A.; Liu, Qiang

    2010-01-01

    Research highlights: → A chimeric subgenomic HCV replicon expresses HCV-3a NS5A in an HCV-1b backbone. → HCV-3a NS5A increases mature SREBP-1c protein level. → HCV-3a NS5A activates SREBP-1c transcription. → Domain II of HCV-3a NS5A is more effective in SREBP-1c promoter activation. → Transcription factor Sp1 is required for SREBP-1c activation by HCV-3a NS5A. -- Abstract: Steatosis is an important clinical manifestation of hepatitis C virus (HCV) infection. The molecular mechanisms of HCV-associated steatosis are not well understood. Sterol regulatory element-binding protein-1c (SREBP-1c) is a key transcription factor which activates the transcription of lipogenic genes. Here we showed that the nuclear, mature SREBP-1c level increases in the nucleus of replicon cells expressing HCV-3a nonstructural protein-5A (NS5A). We further showed that HCV-3a NS5A up-regulates SREBP-1c transcription. Additional analysis showed that transcriptional factor Sp1 is involved in SREBP-1c activation by HCV-3a NS5A because inhibition of Sp1 activity by mithramycin A or a dominant-negative Sp1 construct abrogated SREBP-1c promoter activation by HCV-3a NS5A. In addition, chromatin immunoprecipitation (ChIP) assay demonstrated enhanced binding of Sp1 on the SREBP-1c promoter in HCV-3a NS5A replicon cells. These results showed that HCV-3a NS5A activates SREBP-1c transcription through Sp1. Taken together, our results suggest that HCV-3a NS5A is a contributing factor for steatosis caused by HCV-3a infection.

  11. Mitotic Transcriptional Activation: Clearance of Actively Engaged Pol II via Transcriptional Elongation Control in Mitosis.

    Science.gov (United States)

    Liang, Kaiwei; Woodfin, Ashley R; Slaughter, Brian D; Unruh, Jay R; Box, Andrew C; Rickels, Ryan A; Gao, Xin; Haug, Jeffrey S; Jaspersen, Sue L; Shilatifard, Ali

    2015-11-05

    Although it is established that some general transcription factors are inactivated at mitosis, many details of mitotic transcription inhibition (MTI) and its underlying mechanisms are largely unknown. We have identified mitotic transcriptional activation (MTA) as a key regulatory step to control transcription in mitosis for genes with transcriptionally engaged RNA polymerase II (Pol II) to activate and transcribe until the end of the gene to clear Pol II from mitotic chromatin, followed by global impairment of transcription reinitiation through MTI. Global nascent RNA sequencing and RNA fluorescence in situ hybridization demonstrate the existence of transcriptionally engaged Pol II in early mitosis. Both genetic and chemical inhibition of P-TEFb in mitosis lead to delays in the progression of cell division. Together, our study reveals a mechanism for MTA and MTI whereby transcriptionally engaged Pol II can progress into productive elongation and finish transcription to allow proper cellular division. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Reconstruction and analysis of transcription factor-miRNA co-regulatory feed-forward loops in human cancers using filter-wrapper feature selection.

    Directory of Open Access Journals (Sweden)

    Chen Peng

    Full Text Available BACKGROUND: As one of the most common types of co-regulatory motifs, feed-forward loops (FFLs control many cell functions and play an important role in human cancers. Therefore, it is crucial to reconstruct and analyze cancer-related FFLs that are controlled by transcription factor (TF and microRNA (miRNA simultaneously, in order to find out how miRNAs and TFs cooperate with each other in cancer cells and how they contribute to carcinogenesis. Current FFL studies rely on predicted regulation information and therefore suffer the false positive issue in prediction results. More critically, FFLs generated by existing approaches cannot represent the dynamic and conditional regulation relationship under different experimental conditions. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we proposed a novel filter-wrapper feature selection method to accurately identify co-regulatory mechanism by incorporating prior information from predicted regulatory interactions with parallel miRNA/mRNA expression datasets. By applying this method, we reconstructed 208 and 110 TF-miRNA co-regulatory FFLs from human pan-cancer and prostate datasets, respectively. Further analysis of these cancer-related FFLs showed that the top-ranking TF STAT3 and miRNA hsa-let-7e are key regulators implicated in human cancers, which have regulated targets significantly enriched in cellular process regulations and signaling pathways that are involved in carcinogenesis. CONCLUSIONS/SIGNIFICANCE: In this study, we introduced an efficient computational approach to reconstruct co-regulatory FFLs by accurately identifying gene co-regulatory interactions. The strength of the proposed feature selection method lies in the fact it can precisely filter out false positives in predicted regulatory interactions by quantitatively modeling the complex co-regulation of target genes mediated by TFs and miRNAs simultaneously. Moreover, the proposed feature selection method can be generally applied to

  13. RNA-Seq Reveals Extensive Transcriptional Response to Heat Stress in the Stony Coral Galaxea fascicularis

    Science.gov (United States)

    Hou, Jing; Xu, Tao; Su, Dingjia; Wu, Ying; Cheng, Li; Wang, Jun; Zhou, Zhi; Wang, Yan

    2018-01-01

    Galaxea fascicularis, a stony coral belonging to family Oculinidae, is widely distributed in Red Sea, the Gulf of Aden and large areas of the Indo-Pacific oceans. So far there is a lack of gene expression knowledge concerning this massive coral. In the present study, G. fascicularis was subjected to heat stress at 32.0 ± 0.5°C in the lab, we found that the density of symbiotic zooxanthellae decreased significantly; meanwhile apparent bleaching and tissue lysing were observed at 10 h and 18 h after heat stress. The transcriptome responses were investigated in the stony coral G. fascicularis during heat bleaching using RNA-seq. A total of 42,028 coral genes were assembled from over 439 million reads. Gene expressions were compared at 10 and 18 h after heat stress. The significantly upregulated genes found in the Control_10h vs. Heat_10h comparison, presented mainly in GO terms related with DNA integration and unfolded protein response; and for the Control_18h vs. Heat_18h comparison, the GO terms include DNA integration. In addition, comparison between groups of Control_10h vs. Heat_10h and Control_18h vs. Heat_18h revealed that 125 genes were significantly upregulated in common between the two groups, whereas 21 genes were significantly downregulated in common, all these differentially expressed genes were found to be involved in stress response, DNA integration and unfolded protein response. Taken together, our results suggest that high temperature could activate the stress response at the early stage, and subsequently induce the bleaching and lysing through DNA integration and unfolded protein response, which are able to disrupt the balance of coral-zooxanthella symbiosis in the stony coral G. fascicularis. PMID:29487614

  14. RNA-Seq Reveals Extensive Transcriptional Response to Heat Stress in the Stony Coral Galaxea fascicularis

    Directory of Open Access Journals (Sweden)

    Jing Hou

    2018-02-01

    Full Text Available Galaxea fascicularis, a stony coral belonging to family Oculinidae, is widely distributed in Red Sea, the Gulf of Aden and large areas of the Indo-Pacific oceans. So far there is a lack of gene expression knowledge concerning this massive coral. In the present study, G. fascicularis was subjected to heat stress at 32.0 ± 0.5°C in the lab, we found that the density of symbiotic zooxanthellae decreased significantly; meanwhile apparent bleaching and tissue lysing were observed at 10 h and 18 h after heat stress. The transcriptome responses were investigated in the stony coral G. fascicularis during heat bleaching using RNA-seq. A total of 42,028 coral genes were assembled from over 439 million reads. Gene expressions were compared at 10 and 18 h after heat stress. The significantly upregulated genes found in the Control_10h vs. Heat_10h comparison, presented mainly in GO terms related with DNA integration and unfolded protein response; and for the Control_18h vs. Heat_18h comparison, the GO terms include DNA integration. In addition, comparison between groups of Control_10h vs. Heat_10h and Control_18h vs. Heat_18h revealed that 125 genes were significantly upregulated in common between the two groups, whereas 21 genes were significantly downregulated in common, all these differentially expressed genes were found to be involved in stress response, DNA integration and unfolded protein response. Taken together, our results suggest that high temperature could activate the stress response at the early stage, and subsequently induce the bleaching and lysing through DNA integration and unfolded protein response, which are able to disrupt the balance of coral-zooxanthella symbiosis in the stony coral G. fascicularis.

  15. The hsp 16 Gene of the Probiotic Lactobacillus acidophilus Is Differently Regulated by Salt, High Temperature and Acidic Stresses, as Revealed by Reverse Transcription Quantitative PCR (qRT-PCR Analysis

    Directory of Open Access Journals (Sweden)

    Daniela Fiocco

    2011-08-01

    Full Text Available Small heat shock proteins (sHsps are ubiquitous conserved chaperone-like proteins involved in cellular proteins protection under stressful conditions. In this study, a reverse transcription quantitative PCR (RT-qPCR procedure was developed and used to quantify the transcript level of a small heat shock gene (shs in the probiotic bacterium Lactobacillus acidophilus NCFM, under stress conditions such as heat (45 °C and 53 °C, bile (0.3% w/v, hyperosmosis (1 M and 2.5 M NaCl, and low pH value (pH 4. The shs gene of L. acidophilus NCFM was induced by salt, high temperature and acidic stress, while repression was observed upon bile stress. Analysis of the 5' noncoding region of the hsp16 gene reveals the presence of an inverted repeat (IR sequence (TTAGCACTC-N9-GAGTGCTAA homologue to the controlling IR of chaperone expression (CIRCE elements found in the upstream regulatory region of Gram-positive heat shock operons, suggesting that the hsp16 gene of L. acidophilus might be transcriptionally controlled by HrcA. In addition, the alignment of several small heat shock proteins identified so far in lactic acid bacteria, reveals that the Hsp16 of L. acidophilus exhibits a strong evolutionary relationship with members of the Lactobacillus acidophilus group.

  16. Arabidopsis Pol II-Dependent in Vitro Transcription System Reveals Role of Chromatin for Light-Inducible rbcS Gene Transcription1

    Science.gov (United States)

    Ido, Ayaka; Iwata, Shinya; Iwata, Yuka; Igarashi, Hisako; Hamada, Takahiro; Sonobe, Seiji; Sugiura, Masahiro; Yukawa, Yasushi

    2016-01-01

    In vitro transcription is an essential tool to study the molecular mechanisms of transcription. For over a decade, we have developed an in vitro transcription system from tobacco (Nicotiana tabacum)-cultured cells (BY-2), and this system supported the basic activities of the three RNA polymerases (Pol I, Pol II, and Pol III). However, it was not suitable to study photosynthetic genes, because BY-2 cells have lost their photosynthetic activity. Therefore, Arabidopsis (Arabidopsis thaliana) in vitro transcription systems were developed from green and etiolated suspension cells. Sufficient in vitro Pol II activity was detected after the minor modification of the nuclear soluble extracts preparation method; removal of vacuoles from protoplasts and L-ascorbic acid supplementation in the extraction buffer were particularly effective. Surprisingly, all four Arabidopsis Rubisco small subunit (rbcS-1A, rbcS-1B, rbcS-2B, and rbcS-3B) gene members were in vitro transcribed from the naked DNA templates without any light-dependent manner. However, clear light-inducible transcriptions were observed using chromatin template of rbcS-1A gene, which was prepared with a human nucleosome assembly protein 1 (hNAP1) and HeLa histones. This suggested that a key determinant of light-dependency through the rbcS gene transcription was a higher order of DNA structure (i.e. chromatin). PMID:26662274

  17. Genome-wide Reconstruction of OxyR and SoxRS Transcriptional Regulatory Networks under Oxidative Stress in Escherichia coli K-12 MG1655

    DEFF Research Database (Denmark)

    Seo, Sang Woo; Kim, Donghyuk; Szubin, Richard

    2015-01-01

    Three transcription factors (TFs), OxyR, SoxR, and SoxS, play a critical role in transcriptional regulation of the defense system for oxidative stress in bacteria. However, their full genome-wide regulatory potential is unknown. Here, we perform a genome-scale reconstruction of the OxyR, SoxR, an...

  18. Modularity of gene-regulatory networks revealed in sea-star development

    Directory of Open Access Journals (Sweden)

    Degnan Bernard M

    2011-01-01

    Full Text Available Abstract Evidence that conserved developmental gene-regulatory networks can change as a unit during deutersostome evolution emerges from a study published in BMC Biology. This shows that genes consistently expressed in anterior brain patterning in hemichordates and chordates are expressed in a similar spatial pattern in another deuterostome, an asteroid echinoderm (sea star, but in a completely different developmental context (the animal-vegetal axis. This observation has implications for hypotheses on the type of development present in the deuterostome common ancestor. See research article: http://www.biomedcentral.com/1741-7007/8/143/abstract

  19. Comparative Bioinformatics Analysis of Transcription Factor Genes Indicates Conservation of Key Regulatory Domains among Babesia bovis, Babesia microti, and Theileria equi.

    Science.gov (United States)

    Alzan, Heba F; Knowles, Donald P; Suarez, Carlos E

    2016-11-01

    Apicomplexa tick-borne hemoparasites, including Babesia bovis, Babesia microti, and Theileria equi are responsible for bovine and human babesiosis and equine theileriosis, respectively. These parasites of vast medical, epidemiological, and economic impact have complex life cycles in their vertebrate and tick hosts. Large gaps in knowledge concerning the mechanisms used by these parasites for gene regulation remain. Regulatory genes coding for DNA binding proteins such as members of the Api-AP2, HMG, and Myb families are known to play crucial roles as transcription factors. Although the repertoire of Api-AP2 has been defined and a HMG gene was previously identified in the B. bovis genome, these regulatory genes have not been described in detail in B. microti and T. equi. In this study, comparative bioinformatics was used to: (i) identify and map genes encoding for these transcription factors among three parasites' genomes; (ii) identify a previously unreported HMG gene in B. microti; (iii) define a repertoire of eight conserved Myb genes; and (iv) identify AP2 correlates among B. bovis and the better-studied Plasmodium parasites. Searching the available transcriptome of B. bovis defined patterns of transcription of these three gene families in B. bovis erythrocyte stage parasites. Sequence comparisons show conservation of functional domains and general architecture in the AP2, Myb, and HMG proteins, which may be significant for the regulation of common critical parasite life cycle transitions in B. bovis, B. microti, and T. equi. A detailed understanding of the role of gene families encoding DNA binding proteins will provide new tools for unraveling regulatory mechanisms involved in B. bovis, B. microti, and T. equi life cycles and environmental adaptive responses and potentially contributes to the development of novel convergent strategies for improved control of babesiosis and equine piroplasmosis.

  20. Comparative Bioinformatics Analysis of Transcription Factor Genes Indicates Conservation of Key Regulatory Domains among Babesia bovis, Babesia microti, and Theileria equi.

    Directory of Open Access Journals (Sweden)

    Heba F Alzan

    2016-11-01

    Full Text Available Apicomplexa tick-borne hemoparasites, including Babesia bovis, Babesia microti, and Theileria equi are responsible for bovine and human babesiosis and equine theileriosis, respectively. These parasites of vast medical, epidemiological, and economic impact have complex life cycles in their vertebrate and tick hosts. Large gaps in knowledge concerning the mechanisms used by these parasites for gene regulation remain. Regulatory genes coding for DNA binding proteins such as members of the Api-AP2, HMG, and Myb families are known to play crucial roles as transcription factors. Although the repertoire of Api-AP2 has been defined and a HMG gene was previously identified in the B. bovis genome, these regulatory genes have not been described in detail in B. microti and T. equi. In this study, comparative bioinformatics was used to: (i identify and map genes encoding for these transcription factors among three parasites' genomes; (ii identify a previously unreported HMG gene in B. microti; (iii define a repertoire of eight conserved Myb genes; and (iv identify AP2 correlates among B. bovis and the better-studied Plasmodium parasites. Searching the available transcriptome of B. bovis defined patterns of transcription of these three gene families in B. bovis erythrocyte stage parasites. Sequence comparisons show conservation of functional domains and general architecture in the AP2, Myb, and HMG proteins, which may be significant for the regulation of common critical parasite life cycle transitions in B. bovis, B. microti, and T. equi. A detailed understanding of the role of gene families encoding DNA binding proteins will provide new tools for unraveling regulatory mechanisms involved in B. bovis, B. microti, and T. equi life cycles and environmental adaptive responses and potentially contributes to the development of novel convergent strategies for improved control of babesiosis and equine piroplasmosis.

  1. The identification of transcription factors expressed in the notochord of Ciona intestinalis adds new potential players to the brachyury gene regulatory network.

    Science.gov (United States)

    José-Edwards, Diana S; Kerner, Pierre; Kugler, Jamie E; Deng, Wei; Jiang, Di; Di Gregorio, Anna

    2011-07-01

    The notochord is the distinctive characteristic of chordates; however, the knowledge of the complement of transcription factors governing the development of this structure is still incomplete. Here we present the expression patterns of seven transcription factor genes detected in the notochord of the ascidian Ciona intestinalis at various stages of embryonic development. Four of these transcription factors, Fos-a, NFAT5, AFF and Klf15, have not been directly associated with the notochord in previous studies, while the others, including Spalt-like-a, Lmx-like, and STAT5/6-b, display evolutionarily conserved expression in this structure as well as in other domains. We examined the hierarchical relationships between these genes and the transcription factor Brachyury, which is necessary for notochord development in all chordates. We found that Ciona Brachyury regulates the expression of most, although not all, of these genes. These results shed light on the genetic regulatory program underlying notochord formation in Ciona and possibly other chordates. Copyright © 2011 Wiley-Liss, Inc.

  2. Nsite, NsiteH and NsiteM Computer Tools for Studying Tran-scription Regulatory Elements

    KAUST Repository

    Shahmuradov, Ilham; Solovyev, Victor

    2015-01-01

    regions. Computer methods for identification of REs remain a widely used tool for studying and understanding transcriptional regulation mechanisms. The Nsite, NsiteH and NsiteM programs perform searches for statistically significant (non-random) motifs

  3. Integration analysis of microRNA and mRNA paired expression profiling identifies deregulated microRNA-transcription factor-gene regulatory networks in ovarian endometriosis.

    Science.gov (United States)

    Zhao, Luyang; Gu, Chenglei; Ye, Mingxia; Zhang, Zhe; Li, Li'an; Fan, Wensheng; Meng, Yuanguang

    2018-01-22

    The etiology and pathophysiology of endometriosis remain unclear. Accumulating evidence suggests that aberrant microRNA (miRNA) and transcription factor (TF) expression may be involved in the pathogenesis and development of endometriosis. This study therefore aims to survey the key miRNAs, TFs and genes and further understand the mechanism of endometriosis. Paired expression profiling of miRNA and mRNA in ectopic endometria compared with eutopic endometria were determined by high-throughput sequencing techniques in eight patients with ovarian endometriosis. Binary interactions and circuits among the miRNAs, TFs, and corresponding genes were identified by the Pearson correlation coefficients. miRNA-TF-gene regulatory networks were constructed using bioinformatic methods. Eleven selected miRNAs and TFs were validated by quantitative reverse transcription-polymerase chain reaction in 22 patients. Overall, 107 differentially expressed miRNAs and 6112 differentially expressed mRNAs were identified by comparing the sequencing of the ectopic endometrium group and the eutopic endometrium group. The miRNA-TF-gene regulatory network consists of 22 miRNAs, 12 TFs and 430 corresponding genes. Specifically, some key regulators from the miR-449 and miR-34b/c cluster, miR-200 family, miR-106a-363 cluster, miR-182/183, FOX family, GATA family, and E2F family as well as CEBPA, SOX9 and HNF4A were suggested to play vital regulatory roles in the pathogenesis of endometriosis. Integration analysis of the miRNA and mRNA expression profiles presents a unique insight into the regulatory network of this enigmatic disorder and possibly provides clues regarding replacement therapy for endometriosis.

  4. The ThPOK transcription factor differentially affects the development and function of self-specific CD8(+) T cells and regulatory CD4(+) T cells.

    Science.gov (United States)

    Twu, Yuh-Ching; Teh, Hung-Sia

    2014-03-01

    The zinc finger transcription factor ThPOK plays a crucial role in CD4 T-cell development and CD4/CD8 lineage decision. In ThPOK-deficient mice, developing T cells expressing MHC class II-restricted T-cell receptors are redirected into the CD8 T-cell lineage. In this study, we investigated whether the ThPOK transgene affected the development and function of two additional types of T cells, namely self-specific CD8 T cells and CD4(+) FoxP3(+) T regulatory cells. Self-specific CD8 T cells are characterized by high expression of CD44, CD122, Ly6C, 1B11 and proliferation in response to either IL-2 or IL-15. The ThPOK transgene converted these self-specific CD8 T cells into CD4 T cells. The converted CD4(+) T cells are no longer self-reactive, lose the characteristics of self-specific CD8 T cells, acquire the properties of conventional CD4 T cells and survive poorly in peripheral lymphoid organs. By contrast, the ThPOK transgene promoted the development of CD4(+) FoxP3(+) regulatory T cells resulting in an increased recovery of CD4(+) FoxP3(+) regulatory T cells that expressed higher transforming growth factor-β-dependent suppressor activity. These studies indicate that the ThPOK transcription factor differentially affects the development and function of self-specific CD8 T cells and CD4(+) FoxP3(+) regulatory T cells. © 2013 John Wiley & Sons Ltd.

  5. Transcriptional regulatory network triggered by oxidative signals configures the early response mechanisms of japonica rice to chilling stress

    KAUST Repository

    Yun, Kil-Young; Park, Myoung Ryoul; Mohanty, Bijayalaxmi; Herath, Venura; Xu, Fuyu; Mauleon, Ramil; Wijaya, Edward; Bajic, Vladimir B.; Bruskiewich, Richard; de los Reyes, Benildo G

    2010-01-01

    -plant level analyses established a holistic view of chilling stress response mechanism of japonica rice. Early response regulatory network triggered by oxidative signals is critical for prolonged survival under sub-optimal temperature. Integration of stress

  6. Global transcriptional profiling of Burkholderia pseudomallei under salt stress reveals differential effects on the Bsa type III secretion system

    Directory of Open Access Journals (Sweden)

    Singsuksawat Ekapot

    2010-06-01

    Full Text Available Abstract Background Burkholderia pseudomallei is the causative agent of melioidosis where the highest reported incidence world wide is in the Northeast of Thailand, where saline soil and water are prevalent. Moreover, recent reports indicate a potential pathogenic role for B. pseudomallei in cystic fibrosis lung disease, where an increased sodium chloride (NaCl concentration in airway surface liquid has been proposed. These observations raise the possibility that high salinity may represent a favorable niche for B. pseudomallei. We therefore investigated the global transcriptional response of B. pseudomallei to increased salinity using microarray analysis. Results Transcriptome analysis of B. pseudomallei under salt stress revealed several genes significantly up-regulated in the presence of 320 mM NaCl including genes associated with the bsa-derived Type III secretion system (T3SS. Microarray data were verified by reverse transcriptase-polymerase chain reactions (RT-PCR. Western blot analysis confirmed the increased expression and secretion of the invasion-associated type III secreted proteins BipD and BopE in B. pseudomallei cultures at 170 and 320 mM NaCl relative to salt-free medium. Furthermore, salt-treated B. pseudomallei exhibited greater invasion efficiency into the lung epithelial cell line A549 in a manner partly dependent on a functional Bsa system. Conclusions B. pseudomallei responds to salt stress by modulating the transcription of a relatively small set of genes, among which is the bsa locus associated with invasion and virulence. Expression and secretion of Bsa-secreted proteins was elevated in the presence of exogenous salt and the invasion efficiency was enhanced. Our data indicate that salinity has the potential to influence the virulence of B. pseudomallei.

  7. Genome-wide analysis of WRKY transcription factors in white pear (Pyrus bretschneideri) reveals evolution and patterns under drought stress.

    Science.gov (United States)

    Huang, Xiaosan; Li, Kongqing; Xu, Xiaoyong; Yao, Zhenghong; Jin, Cong; Zhang, Shaoling

    2015-12-24

    WRKY transcription factors (TFs) constitute one of the largest protein families in higher plants, and its members contain one or two conserved WRKY domains, about 60 amino acid residues with the WRKYGQK sequence followed by a C2H2 or C2HC zinc finger motif. WRKY proteins play significant roles in plant development, and in responses to biotic and abiotic stresses. Pear (Pyrus bretschneideri) is one of the most important fruit crops in the world and is frequently threatened by abiotic stress, such as drought, affecting growth, development and productivity. Although the pear genome sequence has been released, little is known about the WRKY TFs in pear, especially in respond to drought stress at the genome-wide level. We identified a total of 103 WRKY TFs in the pear genome. Based on the structural features of WRKY proteins and topology of the phylogenetic tree, the pear WRKY (PbWRKY) family was classified into seven groups (Groups 1, 2a-e, and 3). The microsyteny analysis indicated that 33 (32%) PbWRKY genes were tandemly duplicated and 57 genes (55.3%) were segmentally duplicated. RNA-seq experiment data and quantitative real-time reverse transcription PCR revealed that PbWRKY genes in different groups were induced by drought stress, and Group 2a and 3 were mainly involved in the biological pathways in response to drought stress. Furthermore, adaptive evolution analysis detected a significant positive selection for Pbr001425 in Group 3, and its expression pattern differed from that of other members in this group. The present study provides a solid foundation for further functional dissection and molecular evolution of WRKY TFs in pear, especially for improving the water-deficient resistance of pear through manipulation of the PbWRKYs.

  8. Heterologous expression and transcript analysis of gibberellin biosynthetic genes of grasses reveals novel functionality in the GA3ox family.

    Science.gov (United States)

    Pearce, Stephen; Huttly, Alison K; Prosser, Ian M; Li, Yi-dan; Vaughan, Simon P; Gallova, Barbora; Patil, Archana; Coghill, Jane A; Dubcovsky, Jorge; Hedden, Peter; Phillips, Andrew L

    2015-06-05

    The gibberellin (GA) pathway plays a central role in the regulation of plant development, with the 2-oxoglutarate-dependent dioxygenases (2-ODDs: GA20ox, GA3ox, GA2ox) that catalyse the later steps in the biosynthetic pathway of particularly importance in regulating bioactive GA levels. Although GA has important impacts on crop yield and quality, our understanding of the regulation of GA biosynthesis during wheat and barley development remains limited. In this study we identified or assembled genes encoding the GA 2-ODDs of wheat, barley and Brachypodium distachyon and characterised the wheat genes by heterologous expression and transcript analysis. The wheat, barley and Brachypodium genomes each contain orthologous copies of the GA20ox, GA3ox and GA2ox genes identified in rice, with the exception of OsGA3ox1 and OsGA2ox5 which are absent in these species. Some additional paralogs of 2-ODD genes were identified: notably, a novel gene in the wheat B genome related to GA3ox2 was shown to encode a GA 1-oxidase, named as TaGA1ox-B1. This enzyme is likely to be responsible for the abundant 1β-hydroxylated GAs present in developing wheat grains. We also identified a related gene in barley, located in a syntenic position to TaGA1ox-B1, that encodes a GA 3,18-dihydroxylase which similarly accounts for the accumulation of unusual GAs in barley grains. Transcript analysis showed that some paralogs of the different classes of 2-ODD were expressed mainly in a single tissue or at specific developmental stages. In particular, TaGA20ox3, TaGA1ox1, TaGA3ox3 and TaGA2ox7 were predominantly expressed in developing grain. More detailed analysis of grain-specific gene expression showed that while the transcripts of biosynthetic genes were most abundant in the endosperm, genes encoding inactivation and signalling components were more highly expressed in the seed coat and pericarp. The comprehensive expression and functional characterisation of the multigene families encoding the 2-ODD

  9. The nitrogen responsive transcriptome in potato (Solanum tuberosum L.) reveals significant gene regulatory motifs.

    Science.gov (United States)

    Gálvez, José Héctor; Tai, Helen H; Lagüe, Martin; Zebarth, Bernie J; Strömvik, Martina V

    2016-05-19

    Nitrogen (N) is the most important nutrient for the growth of potato (Solanum tuberosum L.). Foliar gene expression in potato plants with and without N supplementation at 180 kg N ha(-1) was compared at mid-season. Genes with consistent differences in foliar expression due to N supplementation over three cultivars and two developmental time points were examined. In total, thirty genes were found to be over-expressed and nine genes were found to be under-expressed with supplemented N. Functional relationships between over-expressed genes were found. The main metabolic pathway represented among differentially expressed genes was amino acid metabolism. The 1000 bp upstream flanking regions of the differentially expressed genes were analysed and nine overrepresented motifs were found using three motif discovery algorithms (Seeder, Weeder and MEME). These results point to coordinated gene regulation at the transcriptional level controlling steady state potato responses to N sufficiency.

  10. Solving the influence maximization problem reveals regulatory organization of the yeast cell cycle.

    Directory of Open Access Journals (Sweden)

    David L Gibbs

    2017-06-01

    Full Text Available The Influence Maximization Problem (IMP aims to discover the set of nodes with the greatest influence on network dynamics. The problem has previously been applied in epidemiology and social network analysis. Here, we demonstrate the application to cell cycle regulatory network analysis for Saccharomyces cerevisiae. Fundamentally, gene regulation is linked to the flow of information. Therefore, our implementation of the IMP was framed as an information theoretic problem using network diffusion. Utilizing more than 26,000 regulatory edges from YeastMine, gene expression dynamics were encoded as edge weights using time lagged transfer entropy, a method for quantifying information transfer between variables. By picking a set of source nodes, a diffusion process covers a portion of the network. The size of the network cover relates to the influence of the source nodes. The set of nodes that maximizes influence is the solution to the IMP. By solving the IMP over different numbers of source nodes, an influence ranking on genes was produced. The influence ranking was compared to other metrics of network centrality. Although the top genes from each centrality ranking contained well-known cell cycle regulators, there was little agreement and no clear winner. However, it was found that influential genes tend to directly regulate or sit upstream of genes ranked by other centrality measures. The influential nodes act as critical sources of information flow, potentially having a large impact on the state of the network. Biological events that affect influential nodes and thereby affect information flow could have a strong effect on network dynamics, potentially leading to disease. Code and data can be found at: https://github.com/gibbsdavidl/miergolf.

  11. Solving the influence maximization problem reveals regulatory organization of the yeast cell cycle

    Science.gov (United States)

    Shmulevich, Ilya

    2017-01-01

    The Influence Maximization Problem (IMP) aims to discover the set of nodes with the greatest influence on network dynamics. The problem has previously been applied in epidemiology and social network analysis. Here, we demonstrate the application to cell cycle regulatory network analysis for Saccharomyces cerevisiae. Fundamentally, gene regulation is linked to the flow of information. Therefore, our implementation of the IMP was framed as an information theoretic problem using network diffusion. Utilizing more than 26,000 regulatory edges from YeastMine, gene expression dynamics were encoded as edge weights using time lagged transfer entropy, a method for quantifying information transfer between variables. By picking a set of source nodes, a diffusion process covers a portion of the network. The size of the network cover relates to the influence of the source nodes. The set of nodes that maximizes influence is the solution to the IMP. By solving the IMP over different numbers of source nodes, an influence ranking on genes was produced. The influence ranking was compared to other metrics of network centrality. Although the top genes from each centrality ranking contained well-known cell cycle regulators, there was little agreement and no clear winner. However, it was found that influential genes tend to directly regulate or sit upstream of genes ranked by other centrality measures. The influential nodes act as critical sources of information flow, potentially having a large impact on the state of the network. Biological events that affect influential nodes and thereby affect information flow could have a strong effect on network dynamics, potentially leading to disease. Code and data can be found at: https://github.com/gibbsdavidl/miergolf. PMID:28628618

  12. Solving the influence maximization problem reveals regulatory organization of the yeast cell cycle.

    Science.gov (United States)

    Gibbs, David L; Shmulevich, Ilya

    2017-06-01

    The Influence Maximization Problem (IMP) aims to discover the set of nodes with the greatest influence on network dynamics. The problem has previously been applied in epidemiology and social network analysis. Here, we demonstrate the application to cell cycle regulatory network analysis for Saccharomyces cerevisiae. Fundamentally, gene regulation is linked to the flow of information. Therefore, our implementation of the IMP was framed as an information theoretic problem using network diffusion. Utilizing more than 26,000 regulatory edges from YeastMine, gene expression dynamics were encoded as edge weights using time lagged transfer entropy, a method for quantifying information transfer between variables. By picking a set of source nodes, a diffusion process covers a portion of the network. The size of the network cover relates to the influence of the source nodes. The set of nodes that maximizes influence is the solution to the IMP. By solving the IMP over different numbers of source nodes, an influence ranking on genes was produced. The influence ranking was compared to other metrics of network centrality. Although the top genes from each centrality ranking contained well-known cell cycle regulators, there was little agreement and no clear winner. However, it was found that influential genes tend to directly regulate or sit upstream of genes ranked by other centrality measures. The influential nodes act as critical sources of information flow, potentially having a large impact on the state of the network. Biological events that affect influential nodes and thereby affect information flow could have a strong effect on network dynamics, potentially leading to disease. Code and data can be found at: https://github.com/gibbsdavidl/miergolf.

  13. A complex regulatory network coordinating cell cycles during C. elegans development is revealed by a genome-wide RNAi screen.

    Science.gov (United States)

    Roy, Sarah H; Tobin, David V; Memar, Nadin; Beltz, Eleanor; Holmen, Jenna; Clayton, Joseph E; Chiu, Daniel J; Young, Laura D; Green, Travis H; Lubin, Isabella; Liu, Yuying; Conradt, Barbara; Saito, R Mako

    2014-02-28

    The development and homeostasis of multicellular animals requires precise coordination of cell division and differentiation. We performed a genome-wide RNA interference screen in Caenorhabditis elegans to reveal the components of a regulatory network that promotes developmentally programmed cell-cycle quiescence. The 107 identified genes are predicted to constitute regulatory networks that are conserved among higher animals because almost half of the genes are represented by clear human orthologs. Using a series of mutant backgrounds to assess their genetic activities, the RNA interference clones displaying similar properties were clustered to establish potential regulatory relationships within the network. This approach uncovered four distinct genetic pathways controlling cell-cycle entry during intestinal organogenesis. The enhanced phenotypes observed for animals carrying compound mutations attest to the collaboration between distinct mechanisms to ensure strict developmental regulation of cell cycles. Moreover, we characterized ubc-25, a gene encoding an E2 ubiquitin-conjugating enzyme whose human ortholog, UBE2Q2, is deregulated in several cancers. Our genetic analyses suggested that ubc-25 acts in a linear pathway with cul-1/Cul1, in parallel to pathways employing cki-1/p27 and lin-35/pRb to promote cell-cycle quiescence. Further investigation of the potential regulatory mechanism demonstrated that ubc-25 activity negatively regulates CYE-1/cyclin E protein abundance in vivo. Together, our results show that the ubc-25-mediated pathway acts within a complex network that integrates the actions of multiple molecular mechanisms to control cell cycles during development. Copyright © 2014 Roy et al.

  14. Comprehensive analysis of differential co-expression patterns reveal transcriptional dysregulation mechanism and identify novel prognostic lncRNAs in esophageal squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Li Z

    2017-06-01

    Full Text Available Zhen Li,1 Qianlan Yao,1 Songjian Zhao,1 Yin Wang,2,3 Yixue Li,1,4 Zhen Wang4 1School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 2Shanghai Center for Bioinformation Technology, Shanghai Academy of Science and Technology, 3Collaborative Innovation Center for Genetics and Development, Fudan University, 4Key Laboratory of Computational Biology, CAS-MPG Partner Institute for Computational Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People’s Republic of China Abstract: Esophageal squamous cell carcinoma (ESCC is one of the most common malignancies worldwide and occurs at a relatively high frequency in People’s Republic of China. However, the molecular mechanism underlying ESCC is still unclear. In this study, the mRNA and long non-coding RNA (lncRNA expression profiles of ESCC were downloaded from the Gene Expression Omnibus database, and then differential co-expression analysis was used to reveal the altered co-expression relationship of gene pairs in ESCC tumors. A total of 3,709 mRNAs and 923 lncRNAs were differentially co-expressed between normal and tumor tissues, and we found that most of the gene pairs lost associations in the tumor tissues. The differential regulatory networking approach deciphered that transcriptional dysregulation was ubiquitous in ESCC, and most of the differentially regulated links were modulated by 37 TFs. Our study also found that two novel lncRNAs (ADAMTS9-AS1 and AP000696.2 might be essential in the development of ectoderm and epithelial cells, which could significantly stratify ESCC patients into high-risk and low-risk groups, and were much better than traditional clinical tumor markers. Further inspection of two risk groups showed that the changes in TF-target regulation in the high-risk patients were significantly higher than those in the low-risk patients. In addition, four signal transduction-related DCmRNAs (ERBB3, ENSA, KCNK7, MFSD5

  15. Brain transcriptional responses to high-fat diet in Acads-deficient mice reveal energy sensing pathways.

    Directory of Open Access Journals (Sweden)

    Claudia Kruger

    Full Text Available How signals from fatty acid metabolism are translated into changes in food intake remains unclear. Previously we reported that mice with a genetic inactivation of Acads (acyl-coenzyme A dehydrogenase, short-chain, the enzyme responsible for mitochondrial beta-oxidation of C4-C6 short-chain fatty acids (SCFAs, shift consumption away from fat and toward carbohydrate when offered a choice between diets. In the current study, we sought to indentify candidate genes and pathways underlying the effects of SCFA oxidation deficiency on food intake in Acads-/- mice.We performed a transcriptional analysis of gene expression in brain tissue of Acads-/- and Acads+/+ mice fed either a high-fat (HF or low-fat (LF diet for 2 d. Ingenuity Pathway Analysis revealed three top-scoring pathways significantly modified by genotype or diet: oxidative phosphorylation, mitochondrial dysfunction, and CREB signaling in neurons. A comparison of statistically significant responses in HF Acads-/- vs. HF Acads+/+ (3917 and Acads+/+ HF vs. LF Acads+/+ (3879 revealed 2551 genes or approximately 65% in common between the two experimental comparisons. All but one of these genes were expressed in opposite direction with similar magnitude, demonstrating that HF-fed Acads-deficient mice display transcriptional responses that strongly resemble those of Acads+/+ mice fed LF diet. Intriguingly, genes involved in both AMP-kinase regulation and the neural control of food intake followed this pattern. Quantitative RT-PCR in hypothalamus confirmed the dysregulation of genes in these pathways. Western blotting showed an increase in hypothalamic AMP-kinase in Acads-/- mice and HF diet increased, a key protein in an energy-sensing cascade that responds to depletion of ATP.Our results suggest that the decreased beta-oxidation of short-chain fatty acids in Acads-deficient mice fed HF diet produces a state of energy deficiency in the brain and that AMP-kinase may be the cellular energy

  16. SVD identifies transcript length distribution functions from DNA microarray data and reveals evolutionary forces globally affecting GBM metabolism.

    Directory of Open Access Journals (Sweden)

    Nicolas M Bertagnolli

    Full Text Available To search for evolutionary forces that might act upon transcript length, we use the singular value decomposition (SVD to identify the length distribution functions of sets and subsets of human and yeast transcripts from profiles of mRNA abundance levels across gel electrophoresis migration distances that were previously measured by DNA microarrays. We show that the SVD identifies the transcript length distribution functions as "asymmetric generalized coherent states" from the DNA microarray data and with no a-priori assumptions. Comparing subsets of human and yeast transcripts of the same gene ontology annotations, we find that in both disparate eukaryotes, transcripts involved in protein synthesis or mitochondrial metabolism are significantly shorter than typical, and in particular, significantly shorter than those involved in glucose metabolism. Comparing the subsets of human transcripts that are overexpressed in glioblastoma multiforme (GBM or normal brain tissue samples from The Cancer Genome Atlas, we find that GBM maintains normal brain overexpression of significantly short transcripts, enriched in transcripts that are involved in protein synthesis or mitochondrial metabolism, but suppresses normal overexpression of significantly longer transcripts, enriched in transcripts that are involved in glucose metabolism and brain activity. These global relations among transcript length, cellular metabolism and tumor development suggest a previously unrecognized physical mode for tumor and normal cells to differentially regulate metabolism in a transcript length-dependent manner. The identified distribution functions support a previous hypothesis from mathematical modeling of evolutionary forces that act upon transcript length in the manner of the restoring force of the harmonic oscillator.

  17. Comparative Genomics Reveals the Regulatory Complexity of Bifidobacterial Arabinose and Arabino-Oligosaccharide Utilization

    Directory of Open Access Journals (Sweden)

    Aleksandr A. Arzamasov

    2018-04-01

    Full Text Available Members of the genus Bifidobacterium are common inhabitants of the human gastrointestinal tract. Previously it was shown that arabino-oligosaccharides (AOS might act as prebiotics and stimulate the bifidobacterial growth in the gut. However, despite the rapid accumulation of genomic data, the precise mechanisms by which these sugars are utilized and associated transcription control still remain unclear. In the current study, we used a comparative genomic approach to reconstruct arabinose and AOS utilization pathways in over 40 bacterial species belonging to the Bifidobacteriaceae family. The results indicate that the gene repertoire involved in the catabolism of these sugars is highly diverse, and even phylogenetically close species may differ in their utilization capabilities. Using bioinformatics analysis we identified potential DNA-binding motifs and reconstructed putative regulons for the arabinose and AOS utilization genes in the Bifidobacteriaceae genomes. Six LacI-family transcriptional factors (named AbfR, AauR, AauU1, AauU2, BauR1 and BauR2 and a TetR-family regulator (XsaR presumably act as local repressors for AOS utilization genes encoding various α- or β-L-arabinofuranosidases and predicted AOS transporters. The ROK-family regulator AraU and the LacI-family regulator AraQ control adjacent operons encoding putative arabinose transporters and catabolic enzymes, respectively. However, the AraQ regulator is universally present in all Bifidobacterium species including those lacking the arabinose catabolic genes araBDA, suggesting its control of other genes. Comparative genomic analyses of prospective AraQ-binding sites allowed the reconstruction of AraQ regulons and a proposed binary repression/activation mechanism. The conserved core of reconstructed AraQ regulons in bifidobacteria includes araBDA, as well as genes from the central glycolytic and fermentation pathways (pyk, eno, gap, tkt, tal, galM, ldh. The current study expands the

  18. Diverse activities of viral cis-acting RNA regulatory elements revealed using multicolor, long-term, single-cell imaging.

    Science.gov (United States)

    Pocock, Ginger M; Zimdars, Laraine L; Yuan, Ming; Eliceiri, Kevin W; Ahlquist, Paul; Sherer, Nathan M

    2017-02-01

    Cis-acting RNA structural elements govern crucial aspects of viral gene expression. How these structures and other posttranscriptional signals affect RNA trafficking and translation in the context of single cells is poorly understood. Herein we describe a multicolor, long-term (>24 h) imaging strategy for measuring integrated aspects of viral RNA regulatory control in individual cells. We apply this strategy to demonstrate differential mRNA trafficking behaviors governed by RNA elements derived from three retroviruses (HIV-1, murine leukemia virus, and Mason-Pfizer monkey virus), two hepadnaviruses (hepatitis B virus and woodchuck hepatitis virus), and an intron-retaining transcript encoded by the cellular NXF1 gene. Striking behaviors include "burst" RNA nuclear export dynamics regulated by HIV-1's Rev response element and the viral Rev protein; transient aggregations of RNAs into discrete foci at or near the nuclear membrane triggered by multiple elements; and a novel, pulsiform RNA export activity regulated by the hepadnaviral posttranscriptional regulatory element. We incorporate single-cell tracking and a data-mining algorithm into our approach to obtain RNA element-specific, high-resolution gene expression signatures. Together these imaging assays constitute a tractable, systems-based platform for studying otherwise difficult to access spatiotemporal features of viral and cellular gene regulation. © 2017 Pocock et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  19. Seed maturation associated transcriptional programs and regulatory networks underlying genotypic difference in seed dormancy and size/weight in wheat (Triticum aestivum L.).

    Science.gov (United States)

    Yamasaki, Yuji; Gao, Feng; Jordan, Mark C; Ayele, Belay T

    2017-09-16

    Maturation forms one of the critical seed developmental phases and it is characterized mainly by programmed cell death, dormancy and desiccation, however, the transcriptional programs and regulatory networks underlying acquisition of dormancy and deposition of storage reserves during the maturation phase of seed development are poorly understood in wheat. The present study performed comparative spatiotemporal transcriptomic analysis of seed maturation in two wheat genotypes with contrasting seed weight/size and dormancy phenotype. The embryo and endosperm tissues of maturing seeds appeared to exhibit genotype-specific temporal shifts in gene expression profile that might contribute to the seed phenotypic variations. Functional annotations of gene clusters suggest that the two tissues exhibit distinct but genotypically overlapping molecular functions. Motif enrichment predicts genotypically distinct abscisic acid (ABA) and gibberellin (GA) regulated transcriptional networks contribute to the contrasting seed weight/size and dormancy phenotypes between the two genotypes. While other ABA responsive element (ABRE) motifs are enriched in both genotypes, the prevalence of G-box-like motif specifically in tissues of the dormant genotype suggests distinct ABA mediated transcriptional mechanisms control the establishment of dormancy during seed maturation. In agreement with this, the bZIP transcription factors that co-express with ABRE enriched embryonic genes differ with genotype. The enrichment of SITEIIATCYTC motif specifically in embryo clusters of maturing seeds irrespective of genotype predicts a tissue specific role for the respective TCP transcription factors with no or minimal contribution to the variations in seed dormancy. The results of this study advance our understanding of the seed maturation associated molecular mechanisms underlying variation in dormancy and weight/size in wheat seeds, which is a critical step towards the designing of molecular strategies

  20. ThDof1.4 and ThZFP1 constitute a transcriptional regulatory cascade involved in salt or osmotic stress in Tamarix hispida.

    Science.gov (United States)

    Zang, Dandan; Wang, Lina; Zhang, Yiming; Zhao, Huimin; Wang, Yucheng

    2017-07-01

    Identification of the upstream regulators of a gene is important to characterize the transcriptional pathway and the function of the gene. Previously, we found that a zinc finger protein (ThZFP1) is involved in abiotic stress tolerance of Tamarix hispida. In the present study, we further investigated the transcriptional pathway of ThZFP1. Dof motifs are abundant in the ThZFP1 promoter; therefore, we used them to screen for transcriptional regulators of ThZFP1. A Dof protein, ThDof1.4, binds to the Dof motif specifically, and was hypothesized as the upstream regulator of ThZFP1. Further study showed that overexpression of ThDof1.4 in T. hispida activated the expression of GUS controlled by the ThZFP1 promoter. In T. hispida, transient overexpression of ThDof1.4 increased the transcripts of ThZFP1; conversely, transient RNAi-silencing of ThDof1.4 reduced the expression of ThZFP1. Chromatin immunoprecipitation indicated that ThDof1.4 binds to the ThZFP1 promoter. Additionally, ThDof1.4 and ThZFP1 share similar expression patterns in response to salt or drought stress. Furthermore, like ThZFP1, ThDof1.4 could increase the proline level and enhance ROS scavenging capability to improve salt and osmotic stress tolerance. Together, these results suggested that ThDof1.4 and ThZFP1 form a transcriptional regulatory cascade involved in abiotic stress resistance in T. hispida.

  1. Comprehensive meta-analysis of Signal Transducers and Activators of Transcription (STAT genomic binding patterns discerns cell-specific cis-regulatory modules

    Directory of Open Access Journals (Sweden)

    Kang Keunsoo

    2013-01-01

    Full Text Available Abstract Background Cytokine-activated transcription factors from the STAT (Signal Transducers and Activators of Transcription family control common and context-specific genetic programs. It is not clear to what extent cell-specific features determine the binding capacity of seven STAT members and to what degree they share genetic targets. Molecular insight into the biology of STATs was gained from a meta-analysis of 29 available ChIP-seq data sets covering genome-wide occupancy of STATs 1, 3, 4, 5A, 5B and 6 in several cell types. Results We determined that the genomic binding capacity of STATs is primarily defined by the cell type and to a lesser extent by individual family members. For example, the overlap of shared binding sites between STATs 3 and 5 in T cells is greater than that between STAT5 in T cells and non-T cells. Even for the top 1,000 highly enriched STAT binding sites, ~15% of STAT5 binding sites in mouse female liver are shared by other STATs in different cell types while in T cells ~90% of STAT5 binding sites are co-occupied by STAT3, STAT4 and STAT6. In addition, we identified 116 cis-regulatory modules (CRM, which are recognized by all STAT members across cell types defining a common JAK-STAT signature. Lastly, in liver STAT5 binding significantly coincides with binding of the cell-specific transcription factors HNF4A, FOXA1 and FOXA2 and is associated with cell-type specific gene transcription. Conclusions Our results suggest that genomic binding of STATs is primarily determined by the cell type and further specificity is achieved in part by juxtaposed binding of cell-specific transcription factors.

  2. Conservation of protein abundance patterns reveals the regulatory architecture of the EGFR-MAPK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Shi, T.; Niepel, M.; McDermott, J. E.; Gao, Y.; Nicora, C. D.; Chrisler, W. B.; Markillie, L. M.; Petyuk, V. A.; Smith, R. D.; Rodland, K. D.; Sorger, P. K.; Qian, W. -J.; Wiley, H. S.

    2016-07-12

    It is not known whether cancer cells generally show quantitative differences in the expression of signaling pathway proteins that could dysregulate signal transduction. To explore this issue, we first defined the primary components of the EGF-MAPK pathway in normal human mammary epithelial cells, identifying 16 core proteins and 10 feedback regulators. We then quantified their absolute abundance across a panel of normal and cancer cell lines. We found that core pathway proteins were expressed at very similar levels across all cell types. In contrast, the EGFR and transcriptionally controlled feedback regulators were expressed at highly variable levels. The absolute abundance of most core pathway proteins was between 50,000- 70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower levels (2,000-5,000 per cell). MAPK signaling showed saturation in all cells between 3,000-10,000 occupied EGFR, consistent with the idea that low adaptor levels limit signaling. Our results suggest that the core MAPK pathway is essentially invariant across different cell types, with cell- specific differences in signaling likely due to variable levels of feedback regulators. The low abundance of adaptors relative to the EGFR could be responsible for previous observation of saturable signaling, endocytosis, and high affinity EGFR.

  3. Lipid engineering reveals regulatory roles for membrane fluidity in yeast flocculation and oxygen-limited growth

    Energy Technology Data Exchange (ETDEWEB)

    Degreif, Daniel [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Technical Univ. of Darmstadt (Germany); de Rond, Tristan [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Univ. of California, Berkeley, CA (United States); Bertl, Adam [Technical Univ. of Darmstadt (Germany); Keasling, Jay D. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Univ. of California, Berkeley, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Technical Univ. of Denmark, Lyngby (Denmark); Budin, Itay [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Univ. of California, Berkeley, CA (United States)

    2017-03-18

    Cells modulate lipid metabolism in order to maintain membrane homeostasis. In this paper, we use a metabolic engineering approach to manipulate the stoichiometry of fatty acid unsaturation, a regulator of cell membrane fluidity, in Saccharomyces cerevisiae. Unexpectedly, reduced lipid unsaturation triggered cell-cell adhesion (flocculation), a phenomenon characteristic of industrial yeast but uncommon in laboratory strains. We find that ER lipid saturation sensors induce expression of FLO1 – encoding a cell wall polysaccharide binding protein – independently of its canonical regulator. In wild-type cells, Flo1p-dependent flocculation occurs under oxygen-limited growth, which reduces unsaturated lipid synthesis and thus serves as the environmental trigger for flocculation. Transcriptional analysis shows that FLO1 is one of the most highly induced genes in response to changes in lipid unsaturation, and that the set of membrane fluidity-sensitive genes is globally activated as part of the cell's long-term response to hypoxia during fermentation. Finally, our results show how the lipid homeostasis machinery of budding yeast is adapted to carry out a broad response to an environmental stimulus important in biotechnology.

  4. Transcriptional regulatory program in wild-type and retinoblastoma gene-deficient mouse embryonic fibroblasts during adipocyte differentiation

    DEFF Research Database (Denmark)

    Hakim-Weber, Robab; Krogsdam, Anne-M; Jørgensen, Claus

    2011-01-01

    Although many molecular regulators of adipogenesis have been identified a comprehensive catalogue of components is still missing. Recent studies showed that the retinoblastoma protein (pRb) was expressed in the cell cycle and late cellular differentiation phase during adipogenesis. To investigate...... this dual role of pRb in the early and late stages of adipogenesis we used microarrays to perform a comprehensive systems-level analysis of the common transcriptional program of the classic 3T3-L1 preadipocyte cell line, wild-type mouse embryonic fibroblasts (MEFs), and retinoblastoma gene-deficient MEFs...... of experimental data and computational analyses pinpointed a feedback-loop between Pparg and Foxo1.To analyze the effects of the retinoblastoma protein at the transcriptional level we chose a perturbated system (Rb-/- MEFs) for comparison to the transcriptional program of wild-type MEFs. Gene ontology analysis...

  5. Comparative metabolomics in primates reveals the effects of diet and gene regulatory variation on metabolic divergence.

    Science.gov (United States)

    Blekhman, Ran; Perry, George H; Shahbaz, Sevini; Fiehn, Oliver; Clark, Andrew G; Gilad, Yoav

    2014-07-28

    Human diets differ from those of non-human primates. Among few obvious differences, humans consume more meat than most non-human primates and regularly cook their food. It is hypothesized that a dietary shift during human evolution has been accompanied by molecular adaptations in metabolic pathways. Consistent with this notion, comparative studies of gene expression levels in primates have found that the regulation of genes with metabolic functions tend to evolve rapidly in the human lineage. The metabolic consequences of these regulatory differences, however, remained unknown. To address this gap, we performed a comparative study using a combination of gene expression and metabolomic profiling in livers from humans, chimpanzees, and rhesus macaques. We show that dietary differences between species have a strong effect on metabolic concentrations. In addition, we found that differences in metabolic concentration across species are correlated with inter-species differences in the expression of the corresponding enzymes, which control the same metabolic reaction. We identified a number of metabolic compounds with lineage-specific profiles, including examples of human-species metabolic differences that may be directly related to dietary differences.

  6. Evolutionary analysis reveals regulatory and functional landscape of coding and non-coding RNA editing.

    Science.gov (United States)

    Zhang, Rui; Deng, Patricia; Jacobson, Dionna; Li, Jin Billy

    2017-02-01

    Adenosine-to-inosine RNA editing diversifies the transcriptome and promotes functional diversity, particularly in the brain. A plethora of editing sites has been recently identified; however, how they are selected and regulated and which are functionally important are largely unknown. Here we show the cis-regulation and stepwise selection of RNA editing during Drosophila evolution and pinpoint a large number of functional editing sites. We found that the establishment of editing and variation in editing levels across Drosophila species are largely explained and predicted by cis-regulatory elements. Furthermore, editing events that arose early in the species tree tend to be more highly edited in clusters and enriched in slowly-evolved neuronal genes, thus suggesting that the main role of RNA editing is for fine-tuning neurological functions. While nonsynonymous editing events have been long recognized as playing a functional role, in addition to nonsynonymous editing sites, a large fraction of 3'UTR editing sites is evolutionarily constrained, highly edited, and thus likely functional. We find that these 3'UTR editing events can alter mRNA stability and affect miRNA binding and thus highlight the functional roles of noncoding RNA editing. Our work, through evolutionary analyses of RNA editing in Drosophila, uncovers novel insights of RNA editing regulation as well as its functions in both coding and non-coding regions.

  7. Specific interactions between transcription factors and the promoter-regulatory region of the human cytomegalovirus major immediate-early gene

    International Nuclear Information System (INIS)

    Ghazal, P.; Lubon, H.; Hennighausen, L.

    1988-01-01

    Repeat sequence motifs as well as unique sequences between nucleotides -150 and -22 of the human cytomegalovirus immediate-early 1 gene interact in vitro with nuclear proteins. The authors show that a transcriptional element between nucleotides -91 and -65 stimulated promoter activity in vivo and in vitro by binding specific cellular transcription factors. Finally, a common sequence motif, (T)TGG/AC, present in 15 of the determined binding sites suggests a particular class of nuclear factors associated with the immediate-early 1 gene

  8. Combined chromatin and expression analysis reveals specific regulatory mechanisms within cytokine genes in the macrophage early immune response.

    Directory of Open Access Journals (Sweden)

    Maria Jesus Iglesias

    Full Text Available Macrophages play a critical role in innate immunity, and the expression of early response genes orchestrate much of the initial response of the immune system. Macrophages undergo extensive transcriptional reprogramming in response to inflammatory stimuli such as Lipopolysaccharide (LPS.To identify gene transcription regulation patterns involved in early innate immune responses, we used two genome-wide approaches--gene expression profiling and chromatin immunoprecipitation-sequencing (ChIP-seq analysis. We examined the effect of 2 hrs LPS stimulation on early gene expression and its relation to chromatin remodeling (H3 acetylation; H3Ac and promoter binding of Sp1 and RNA polymerase II phosphorylated at serine 5 (S5P RNAPII, which is a marker for transcriptional initiation. Our results indicate novel and alternative gene regulatory mechanisms for certain proinflammatory genes. We identified two groups of up-regulated inflammatory genes with respect to chromatin modification and promoter features. One group, including highly up-regulated genes such as tumor necrosis factor (TNF, was characterized by H3Ac, high CpG content and lack of TATA boxes. The second group, containing inflammatory mediators (interleukins and CCL chemokines, was up-regulated upon LPS stimulation despite lacking H3Ac in their annotated promoters, which were low in CpG content but did contain TATA boxes. Genome-wide analysis showed that few H3Ac peaks were unique to either +/-LPS condition. However, within these, an unpacking/expansion of already existing H3Ac peaks was observed upon LPS stimulation. In contrast, a significant proportion of S5P RNAPII peaks (approx 40% was unique to either condition. Furthermore, data indicated a large portion of previously unannotated TSSs, particularly in LPS-stimulated macrophages, where only 28% of unique S5P RNAPII peaks overlap annotated promoters. The regulation of the inflammatory response appears to occur in a very specific manner at

  9. A sparse regulatory network of copy-number driven gene expression reveals putative breast cancer oncogenes.

    Science.gov (United States)

    Yuan, Yinyin; Curtis, Christina; Caldas, Carlos; Markowetz, Florian

    2012-01-01

    Copy number aberrations are recognized to be important in cancer as they may localize to regions harboring oncogenes or tumor suppressors. Such genomic alterations mediate phenotypic changes through their impact on expression. Both cis- and transacting alterations are important since they may help to elucidate putative cancer genes. However, amidst numerous passenger genes, trans-effects are less well studied due to the computational difficulty in detecting weak and sparse signals in the data, and yet may influence multiple genes on a global scale. We propose an integrative approach to learn a sparse interaction network of DNA copy-number regions with their downstream transcriptional targets in breast cancer. With respect to goodness of fit on both simulated and real data, the performance of sparse network inference is no worse than other state-of-the-art models but with the advantage of simultaneous feature selection and efficiency. The DNA-RNA interaction network helps to distinguish copy-number driven expression alterations from those that are copy-number independent. Further, our approach yields a quantitative copy-number dependency score, which distinguishes cis- versus trans-effects. When applied to a breast cancer data set, numerous expression profiles were impacted by cis-acting copy-number alterations, including several known oncogenes such as GRB7, ERBB2, and LSM1. Several trans-acting alterations were also identified, impacting genes such as ADAM2 and BAGE, which warrant further investigation. An R package named lol is available from www.markowetzlab.org/software/lol.html.

  10. Transcriptional profiles of hybrid Eucalyptus genotypes with contrasting lignin content reveal that monolignol biosynthesis-related genes regulate wood composition

    Directory of Open Access Journals (Sweden)

    Tomotaka eShinya

    2016-04-01

    Full Text Available Eucalyptus species constitutes the most widely planted hardwood trees in temperate and subtropical regions. In this study, we compared the transcript levels of genes involved in lignocellulose formation such as cellulose, hemicellulose and lignin biosynthesis in two selected three-year old hybrid Eucalyptus (Eucalyptus urophylla x E. grandis genotypes (AM063 and AM380 that have different lignin content. AM063 and AM380 had 20.2 and 35.5% of Klason lignin content and 59.0% and 48.2%, -cellulose contents, respectively. We investigated the correlation between wood properties and transcript levels of wood formation-related genes using RNA-seq with total RNAs extracted from developing xylem tissues at a breast height. Transcript levels of cell wall construction genes such as cellulose synthase (CesA and sucrose synthase (SUSY were almost the same in both genotypes. However, AM063 exhibited higher transcript levels of UDP-glucose pyrophosphorylase (UGP and xyloglucan endotransglucoxylase (XTH than those in AM380. Most monolignol biosynthesis- related isozyme genes showed higher transcript levels in AM380. These results indicate monolignol biosynthesis-related genes may regulate wood composition in Eucalyptus. Flavonoids contents were also observed at much higher levels in AM380 as a result of the elevated transcript levels of common phenylpropanoid pathway genes, phenylalanine ammonium lyase (PAL, cinnamate-4-hydroxylase (C4H and 4-coumarate-CoA ligase (4CL. Secondary plant cell wall formation is regulated by many transcription factors. We analyzed genes encoding NAC, WRKY, AP2/ERF and KNOX transcription factors and found higher transcript levels of these genes in AM380. We also observed increased transcription of some MYB and LIM domain transcription factors in AM380 compared to AM063. All these results show that genes related to monolignol biosynthesis may regulate the wood composition and help maintain the ratio of cellulose and lignin contents

  11. Regulatory mechanisms for 3'-end alternative splicing and polyadenylation of the Glial Fibrillary Acidic Protein, GFAP, transcript

    DEFF Research Database (Denmark)

    Blechingberg, Jenny; Lykke-Andersen, Søren; Jensen, Torben Heick

    2007-01-01

    (PTB) protein enhanced both exon 7a polyadenylation and exon 7a splicing. Finally, increasing transcription by the VP16 trans-activator did not affect the frequency of use of the exon 7a polyadenylation signal whereas the exon 7a splicing frequency was decreased. Our data suggest a model...

  12. Bicarbonate-mediated transcriptional activation of divergent operons by the virulence regulatory protein, RegA, from Citrobacter rodentium.

    Science.gov (United States)

    Yang, Ji; Hart, Emily; Tauschek, Marija; Price, G Dean; Hartland, Elizabeth L; Strugnell, Richard A; Robins-Browne, Roy M

    2008-04-01

    Regulation of virulence gene expression plays a central role in the pathogenesis of enteric bacteria as they encounter diverse environmental conditions in the gastrointestinal tract of their hosts. In this study, we investigated environmental regulation of two putative virulence determinants adcA and kfc by RegA, an AraC/XylS-like regulator, from Citrobacter rodentium, and identified bicarbonate as the environmental signal which induced transcription of adcA and kfc through RegA. Primer extension experiments showed that adcA and kfc were divergently transcribed from sigma(70) promoters. In vivo and in vitro experiments demonstrated that bicarbonate facilitated and stabilized the binding of RegA to an operator located between the two promoters. The interaction of RegA with its DNA target resulted in the formation of a nucleosome-like structure, which evidently displaced the histone-like proteins, H-NS and StpA, from the adcA and kfc promoter regions, leading to transcriptional derepression. In addition, our results indicated that RegA also behaved as a Class I activator by directly stimulating transcription initiation by RNA polymerase. This is the first report to describe the molecular mechanism by which an environmental chemical stimulates transcription of virulence-associated genes of an enteric pathogen through an AraC/XlyS-like activator.

  13. Dissection of TALE-dependent gene activation reveals that they induce transcription cooperatively and in both orientations.

    Science.gov (United States)

    Streubel, Jana; Baum, Heidi; Grau, Jan; Stuttman, Johannes; Boch, Jens

    2017-01-01

    Plant-pathogenic Xanthomonas bacteria inject transcription activator-like effector proteins (TALEs) into host cells to specifically induce transcription of plant genes and enhance susceptibility. Although the DNA-binding mode is well-understood it is still ambiguous how TALEs initiate transcription and whether additional promoter elements are needed to support this. To systematically dissect prerequisites for transcriptional initiation the activity of one TALE was compared on different synthetic Bs4 promoter fragments. In addition, a large collection of artificial TALEs spanning the OsSWEET14 promoter was compared. We show that the presence of a TALE alone is not sufficient to initiate transcription suggesting the requirement of additional supporting promoter elements. At the OsSWEET14 promoter TALEs can initiate transcription from various positions, in a synergistic manner of multiple TALEs binding in parallel to the promoter, and even by binding in reverse orientation. TALEs are known to shift the transcriptional start site, but our data show that this shift depends on the individual position of a TALE within a promoter context. Our results implicate that TALEs function like classical enhancer-binding proteins and initiate transcription in both orientations which has consequences for in planta target gene prediction and design of artificial activators.

  14. GLOBAL TRANSCRIPTION PROFILING REVEALS DIFFERENTIAL RESPONSES TO CHRONIC NITROGEN STRESS AND PUTATIVE NITROGEN REGULATORY COMPONENTS IN ARABIDOPSIS

    Science.gov (United States)

    Background: A large quantity of nitrogen (N) fertilizer is used for crop production to achieve high yields at a significant economic and environmental cost. Efforts have been directed to understanding the molecular basis of plant responses to N and to identifying N-responsive gen...

  15. Elucidation of the regulatory role of the fructose operon reveals a novel target for enhancing the NADPH supply in Corynebacterium glutamicum.

    Science.gov (United States)

    Wang, Zhihao; Chan, Siu Hung Joshua; Sudarsan, Suresh; Blank, Lars M; Jensen, Peter Ruhdal; Solem, Christian

    2016-11-01

    The performance of Corynebacterium glutamicum cell factories producing compounds which rely heavily on NADPH has been reported to depend on the sugar being metabolized. While some aspects of this phenomenon have been elucidated, there are still many unresolved questions as to how sugar metabolism is linked to redox and to the general metabolism. We here provide new insights into the regulation of the metabolism of this important platform organism by systematically characterizing mutants carrying various lesions in the fructose operon. Initially, we found that a strain where the dedicated fructose uptake system had been inactivated (KO-ptsF) was hampered in growth on sucrose minimal medium, and suppressor mutants appeared readily. Comparative genomic analysis in conjunction with enzymatic assays revealed that suppression was linked to inactivation of the pfkB gene, encoding a fructose-1-phosphate kinase. Detailed characterization of KO-ptsF, KO-pfkB and double knock-out (DKO) derivatives revealed a strong role for sugar-phosphates, especially fructose-1-phosphate (F1P), in governing sugar as well as redox metabolism due to effects on transcriptional regulation of key genes. These findings allowed us to propose a simple model explaining the correlation between sugar phosphate concentration, gene expression and ultimately the observed phenotype. To guide us in our analysis and help us identify bottlenecks in metabolism we debugged an existing genome-scale model onto which we overlaid the transcriptome data. Based on the results obtained we managed to enhance the NADPH supply and transform the wild-type strain into delivering the highest yield of lysine ever obtained on sucrose and fructose, thus providing a good example of how regulatory mechanisms can be harnessed for bioproduction. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  16. An in silico approach reveals associations between genetic and epigenetic factors within regulatory elements in B cells from primary Sjögren’s syndrome patients

    Directory of Open Access Journals (Sweden)

    Orsia D. Konsta

    2015-08-01

    Full Text Available Recent advances in genetics have highlighted several regions and candidate genes associated with primary Sjögren's syndrome (SS, a systemic autoimmune epithelitis that combines exocrine gland dysfunctions, and focal lymphocytic infiltrations. In addition to genetic factors, it is now clear that epigenetic deregulations are present during SS and restricted to specific cell type subsets such as lymphocytes and salivary gland epithelial cells. In this study, 72 single nucleotide polymorphisms (SNPs associated with 43 SS gene risk factors were selected from publicly available and peer reviewed literature for further in silico analysis. SS risk variant location was tested revealing a broad distribution in coding sequences (5.6%, intronic sequences (55.6%, upstream/downstream genic regions (30.5%, and intergenic regions (8.3%. Moreover, a significant enrichment of regulatory motifs (promoter, enhancer, insulator, DNAse peak and eQTL characterizes SS risk variants (94.4%. Next, screening SNPs in high linkage disequilibrium (r2 ≥ 0.8 in Caucasians revealed 645 new variants including 5 SNPs with missense mutations, and indicated an enrichment of transcriptionally active motifs according to the cell type (B cells > monocytes > T cells >> A549. Finally, we looked at SS risk variants for histone markers in B cells (GM12878, monocytes (CD14+ and epithelial cells (A548. Active histone markers were associated with SS risk variants at both promoters and enhancers in B cells, and within enhancers in monocytes. In conclusion and based on the obtained in silico results, that need further confirmation, associations were observed between SS genetic risk factors and epigenetic factors and these associations predominate in B cells such as those observed at the FAM167A-BLK locus.

  17. Integrated analysis of microRNA and gene expression profiles reveals a functional regulatory module associated with liver fibrosis.

    Science.gov (United States)

    Chen, Wei; Zhao, Wenshan; Yang, Aiting; Xu, Anjian; Wang, Huan; Cong, Min; Liu, Tianhui; Wang, Ping; You, Hong

    2017-12-15

    Liver fibrosis, characterized with the excessive accumulation of extracellular matrix (ECM) proteins, represents the final common pathway of chronic liver inflammation. Ever-increasing evidence indicates microRNAs (miRNAs) dysregulation has important implications in the different stages of liver fibrosis. However, our knowledge of miRNA-gene regulation details pertaining to such disease remains unclear. The publicly available Gene Expression Omnibus (GEO) datasets of patients suffered from cirrhosis were extracted for integrated analysis. Differentially expressed miRNAs (DEMs) and genes (DEGs) were identified using GEO2R web tool. Putative target gene prediction of DEMs was carried out using the intersection of five major algorithms: DIANA-microT, TargetScan, miRanda, PICTAR5 and miRWalk. Functional miRNA-gene regulatory network (FMGRN) was constructed based on the computational target predictions at the sequence level and the inverse expression relationships between DEMs and DEGs. DAVID web server was selected to perform KEGG pathway enrichment analysis. Functional miRNA-gene regulatory module was generated based on the biological interpretation. Internal connections among genes in liver fibrosis-related module were determined using String database. MiRNA-gene regulatory modules related to liver fibrosis were experimentally verified in recombinant human TGFβ1 stimulated and specific miRNA inhibitor treated LX-2 cells. We totally identified 85 and 923 dysregulated miRNAs and genes in liver cirrhosis biopsy samples compared to their normal controls. All evident miRNA-gene pairs were identified and assembled into FMGRN which consisted of 990 regulations between 51 miRNAs and 275 genes, forming two big sub-networks that were defined as down-network and up-network, respectively. KEGG pathway enrichment analysis revealed that up-network was prominently involved in several KEGG pathways, in which "Focal adhesion", "PI3K-Akt signaling pathway" and "ECM

  18. Regulatory complexity revealed by integrated cytological and RNA-seq analyses of meiotic substages in mouse spermatocytes.

    Science.gov (United States)

    Ball, Robyn L; Fujiwara, Yasuhiro; Sun, Fengyun; Hu, Jianjun; Hibbs, Matthew A; Handel, Mary Ann; Carter, Gregory W

    2016-08-12

    The continuous and non-synchronous nature of postnatal male germ-cell development has impeded stage-specific resolution of molecular events of mammalian meiotic prophase in the testis. Here the juvenile onset of spermatogenesis in mice is analyzed by combining cytological and transcriptomic data in a novel computational analysis that allows decomposition of the transcriptional programs of spermatogonia and meiotic prophase substages. Germ cells from testes of individual mice were obtained at two-day intervals from 8 to 18 days post-partum (dpp), prepared as surface-spread chromatin and immunolabeled for meiotic stage-specific protein markers (STRA8, SYCP3, phosphorylated H2AFX, and HISTH1T). Eight stages were discriminated cytologically by combinatorial antibody labeling, and RNA-seq was performed on the same samples. Independent principal component analyses of cytological and transcriptomic data yielded similar patterns for both data types, providing strong evidence for substage-specific gene expression signatures. A novel permutation-based maximum covariance analysis (PMCA) was developed to map co-expressed transcripts to one or more of the eight meiotic prophase substages, thereby linking distinct molecular programs to cytologically defined cell states. Expression of meiosis-specific genes is not substage-limited, suggesting regulation of substage transitions at other levels. This integrated analysis provides a general method for resolving complex cell populations. Here it revealed not only features of meiotic substage-specific gene expression, but also a network of substage-specific transcription factors and relationships to potential target genes.

  19. A model of gene expression based on random dynamical systems reveals modularity properties of gene regulatory networks.

    Science.gov (United States)

    Antoneli, Fernando; Ferreira, Renata C; Briones, Marcelo R S

    2016-06-01

    Here we propose a new approach to modeling gene expression based on the theory of random dynamical systems (RDS) that provides a general coupling prescription between the nodes of any given regulatory network given the dynamics of each node is modeled by a RDS. The main virtues of this approach are the following: (i) it provides a natural way to obtain arbitrarily large networks by coupling together simple basic pieces, thus revealing the modularity of regulatory networks; (ii) the assumptions about the stochastic processes used in the modeling are fairly general, in the sense that the only requirement is stationarity; (iii) there is a well developed mathematical theory, which is a blend of smooth dynamical systems theory, ergodic theory and stochastic analysis that allows one to extract relevant dynamical and statistical information without solving the system; (iv) one may obtain the classical rate equations form the corresponding stochastic version by averaging the dynamic random variables (small noise limit). It is important to emphasize that unlike the deterministic case, where coupling two equations is a trivial matter, coupling two RDS is non-trivial, specially in our case, where the coupling is performed between a state variable of one gene and the switching stochastic process of another gene and, hence, it is not a priori true that the resulting coupled system will satisfy the definition of a random dynamical system. We shall provide the necessary arguments that ensure that our coupling prescription does indeed furnish a coupled regulatory network of random dynamical systems. Finally, the fact that classical rate equations are the small noise limit of our stochastic model ensures that any validation or prediction made on the basis of the classical theory is also a validation or prediction of our model. We illustrate our framework with some simple examples of single-gene system and network motifs. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. Complex and extensive post-transcriptional regulation revealed by integrative proteomic and transcriptomic analysis of metabolite stress response in Clostridium acetobutylicum.

    Science.gov (United States)

    Venkataramanan, Keerthi P; Min, Lie; Hou, Shuyu; Jones, Shawn W; Ralston, Matthew T; Lee, Kelvin H; Papoutsakis, E Terry

    2015-01-01

    Clostridium acetobutylicum is a model organism for both clostridial biology and solvent production. The organism is exposed to its own toxic metabolites butyrate and butanol, which trigger an adaptive stress response. Integrative analysis of proteomic and RNAseq data may provide novel insights into post-transcriptional regulation. The identified iTRAQ-based quantitative stress proteome is made up of 616 proteins with a 15 % genome coverage. The differentially expressed proteome correlated poorly with the corresponding differential RNAseq transcriptome. Up to 31 % of the differentially expressed proteins under stress displayed patterns opposite to those of the transcriptome, thus suggesting significant post-transcriptional regulation. The differential proteome of the translation machinery suggests that cells employ a different subset of ribosomal proteins under stress. Several highly upregulated proteins but with low mRNA levels possessed mRNAs with long 5'UTRs and strong RBS scores, thus supporting the argument that regulatory elements on the long 5'UTRs control their translation. For example, the oxidative stress response rubrerythrin was upregulated only at the protein level up to 40-fold without significant mRNA changes. We also identified many leaderless transcripts, several displaying different transcriptional start sites, thus suggesting mRNA-trimming mechanisms under stress. Downregulation of Rho and partner proteins pointed to changes in transcriptional elongation and termination under stress. The integrative proteomic-transcriptomic analysis demonstrated complex expression patterns of a large fraction of the proteome. Such patterns could not have been detected with one or the other omic analyses. Our analysis proposes the involvement of specific molecular mechanisms of post-transcriptional regulation to explain the observed complex stress response.

  1. Mapping the transcription start points of the Staphylococcus aureus eap, emp, and vwb promoters reveals a conserved octanucleotide sequence that is essential for expression of these genes.

    Science.gov (United States)

    Harraghy, Niamh; Homerova, Dagmar; Herrmann, Mathias; Kormanec, Jan

    2008-01-01

    Mapping the transcription start points of the eap, emp, and vwb promoters revealed a conserved octanucleotide sequence (COS). Deleting this sequence abolished the expression of eap, emp, and vwb. However, electrophoretic mobility shift assays gave no evidence that this sequence was a binding site for SarA or SaeR, known regulators of eap and emp.

  2. Quantitative Proteomics Reveals the Regulatory Networks of Circular RNA CDR1as in Hepatocellular Carcinoma Cells.

    Science.gov (United States)

    Yang, Xue; Xiong, Qian; Wu, Ying; Li, Siting; Ge, Feng

    2017-10-06

    Circular RNAs (circRNAs), a class of widespread endogenous RNAs, play crucial roles in diverse biological processes and are potential biomarkers in diverse human diseases and cancers. Cerebellar-degeneration-related protein 1 antisense RNA (CDR1as), an oncogenic circRNA, is involved in human tumorigenesis and is dysregulated in hepatocellular carcinoma (HCC). However, the molecular mechanisms underlying CDR1as functions in HCC remain unclear. Here we explored the functions of CDR1as and searched for CDR1as-regulated proteins in HCC cells. A quantitative proteomics strategy was employed to globally identify CDR1as-regulated proteins in HCC cells. In total, we identified 330 differentially expressed proteins (DEPs) upon enhanced CDR1as expression in HepG2 cells, indicating that they could be proteins regulated by CDR1as. Bioinformatic analysis revealed that many DEPs were involved in cell proliferation and the cell cycle. Further functional studies of epidermal growth factor receptor (EGFR) found that CDR1as exerts its effects on cell proliferation at least in part through the regulation of EGFR expression. We further confirmed that CDR1as could inhibit the expression of microRNA-7 (miR-7). EGFR is a validated target of miR-7; therefore, CDR1as may exert its function by regulating EGFR expression via targeting miR-7 in HCC cells. Taken together, we revealed novel functions and underlying mechanisms of CDR1as in HCC cells. This study serves as the first proteome-wide analysis of a circRNA-regulated protein in cells and provides a reliable and highly efficient method for globally identifying circRNA-regulated proteins.

  3. Foot-and-mouth disease virus leader proteinase inhibits dsRNA-induced type I interferon transcription by decreasing interferon regulatory factor 3/7 in protein levels

    International Nuclear Information System (INIS)

    Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun; Xiao, Shaobo

    2010-01-01

    Research highlights: → FMDV L pro inhibits poly(I:C)-induced IFN-α1/β mRNA expression. → L pro inhibits MDA5-mediated activation of the IFN-α1/β promoter. → L pro significantly reduced the transcription of multiple IRF-responsive genes. → L pro inhibits IFN-α1/β promoter activation by decreasing IRF-3/7 in protein levels. → The ability to process eIF-4G of L pro is not necessary to inhibit IFN-α1/β activation. -- Abstract: The leader proteinase (L pro ) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-β (IFN-β) antagonist that disrupts the integrity of transcription factor nuclear factor κB (NF-κB). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-α1/β expression caused by L pro was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-α/β. Furthermore, overexpression of L pro significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L pro mutants indicated that the ability to process eIF-4G of L pro is not required for suppressing dsRNA-induced activation of the IFN-α1/β promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-κB, L pro also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

  4. Foot-and-mouth disease virus leader proteinase inhibits dsRNA-induced type I interferon transcription by decreasing interferon regulatory factor 3/7 in protein levels

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Dang; Fang, Liurong; Luo, Rui; Ye, Rui; Fang, Ying; Xie, Lilan; Chen, Huanchun [Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); Xiao, Shaobo, E-mail: shaoboxiao@yahoo.com [Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China)

    2010-08-13

    Research highlights: {yields} FMDV L{sup pro} inhibits poly(I:C)-induced IFN-{alpha}1/{beta} mRNA expression. {yields} L{sup pro} inhibits MDA5-mediated activation of the IFN-{alpha}1/{beta} promoter. {yields} L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes. {yields} L{sup pro} inhibits IFN-{alpha}1/{beta} promoter activation by decreasing IRF-3/7 in protein levels. {yields} The ability to process eIF-4G of L{sup pro} is not necessary to inhibit IFN-{alpha}1/{beta} activation. -- Abstract: The leader proteinase (L{sup pro}) of foot-and-mouth disease virus (FMDV) has been identified as an interferon-{beta} (IFN-{beta}) antagonist that disrupts the integrity of transcription factor nuclear factor {kappa}B (NF-{kappa}B). In this study, we showed that the reduction of double stranded RNA (dsRNA)-induced IFN-{alpha}1/{beta} expression caused by L{sup pro} was also associated with a decrease of interferon regulatory factor 3/7 (IRF-3/7) in protein levels, two critical transcription factors for activation of IFN-{alpha}/{beta}. Furthermore, overexpression of L{sup pro} significantly reduced the transcription of multiple IRF-responsive genes including 2',5'-OAS, ISG54, IP-10, and RANTES. Screening L{sup pro} mutants indicated that the ability to process eIF-4G of L{sup pro} is not required for suppressing dsRNA-induced activation of the IFN-{alpha}1/{beta} promoter and decreasing IRF-3/7 expression. Taken together, our results demonstrate that, in addition to disrupting NF-{kappa}B, L{sup pro} also decreases IRF-3/7 expression to suppress dsRNA-induced type I IFN production, suggesting multiple strategies used by FMDV to counteract the immune response to viral infection.

  5. Crystal structures of the transcriptional repressor RolR reveals a novel recognition mechanism between inducer and regulator.

    Directory of Open Access Journals (Sweden)

    De-Feng Li

    Full Text Available Many members of the TetR family control the transcription of genes involved in multidrug resistance and pathogenicity. RolR (ResorcinolRegulator, the recently reported TetR-type regulator for aromatic catabolism from Corynebacterium glutamicum, distinguishes itself by low sequence similarities and different regulation from the previously known members of the TetR family. Here we report the crystal structures of RolR in its effector-bound (with resorcinol and aop- forms at 2.5 Å and 3.6 Å, respectively. The structure of resorcinol-RolR complex reveal that the hydrogen-bonded network mediated by the four-residue motif (Asp94- Arg145- Arg148- Asp149 with two water molecules and the hydrophobic interaction via five residues (Phe107, Leu111, Leu114, Leu142, and Phe172 are the key factors for the recognition and binding between the resorcinol and RolR molecules. The center-to-center separation of the recognition helices h3-h3' is decreased upon effector-binding from 34.9 Å to 30.4 Å. This structural change results in that RolR was unsuitable for DNA binding. Those observations are distinct from that in other TetR members. Structure-based mutagenesis on RolR was carried out and the results confirmed the critical roles of the above mentioned residues for effector-binding specificity and affinity. Similar sequence searches and sequence alignments identified 29 RolR homologues from GenBank, and all the above mentioned residues are highly conserved in the homologues. Based on these structural and other functional investigations, it is proposed that RolR may represent a new subfamily of TetR proteins that are invovled in aromatic degradation and sharing common recognition mode as for RolR.

  6. Single-Cell RNA-Seq Reveals the Transcriptional Landscape and Heterogeneity of Aortic Macrophages in Murine Atherosclerosis.

    Science.gov (United States)

    Cochain, Clément; Vafadarnejad, Ehsan; Arampatzi, Panagiota; Jaroslav, Pelisek; Winkels, Holger; Ley, Klaus; Wolf, Dennis; Saliba, Antoine-Emmanuel; Zernecke, Alma

    2018-03-15

    Rationale: It is assumed that atherosclerotic arteries contain several macrophage subsets endowed with specific functions. The precise identity of these subsets is poorly characterized as they ha ve been defined by the expression of a restricted number of markers. Objective: We have applied single-cell RNA-seq as an unbiased profiling strategy to interrogate and classify aortic macrophage heterogeneity at the single-cell level in atherosclerosis. Methods and Results: We performed single-cell RNA sequencing of total aortic CD45 + cells extracted from the non-diseased (chow fed) and atherosclerotic (11 weeks of high fat diet) aorta of Ldlr -/- mice. Unsupervised clustering singled out 13 distinct aortic cell clusters. Among the myeloid cell populations, Resident-like macrophages with a gene expression profile similar to aortic resident macrophages were found in healthy and diseased aortae, whereas monocytes, monocyte-derived dendritic cells (MoDC), and two populations of macrophages were almost exclusively detectable in atherosclerotic aortae, comprising Inflammatory macrophages showing enrichment in I l1b , and previously undescribed TREM2 hi macrophages. Differential gene expression and gene ontology enrichment analyses revealed specific gene expression patterns distinguishing these three macrophage subsets and MoDC, and uncovered putative functions of each cell type. Notably, TREM2 hi macrophages appeared to be endowed with specialized functions in lipid metabolism and catabolism, and presented a gene expression signature reminiscent of osteoclasts, suggesting a role in lesion calcification. TREM2 expression was moreover detected in human lesional macrophages. Importantly, these macrophage populations were present also in advanced atherosclerosis and in Apoe -/- aortae, indicating relevance of our findings in different stages of atherosclerosis and mouse models. Conclusions: These data unprecedentedly uncovered the transcriptional landscape and phenotypic

  7. Meta-analysis reveals an association between signal transducer and activator of transcription-4 polymorphism and hepatocellular carcinoma risk.

    Science.gov (United States)

    Zhang, Li; Xu, Kuihua; Liu, Chuanmiao; Chen, Jiasheng

    2017-03-01

    Hepatocellular carcinoma (HCC) is one of the most common causes of cancer-related mortality worldwide. Signal transducer and activator of transcription (STAT) proteins play a multitude of important functions in liver pathophysiology. Recent studies have indicated associations of rs7574865 single nucleotide polymorphism (SNP) in the STAT4 gene with various autoimmune diseases. The association between STAT4 polymorphism and the risk of HCC has been analyzed in several studies, but results remain inconsistent. This study used a meta-analysis approach to comprehensively investigate the correlation between STAT4 polymorphism and HCC risk based on previously published reports. Studies were searched from the databases of PubMed, EMBase, Web of Science, and the Chinese National Knowledge Infrastructure up to 31 December 2015. The meta-analysis was carried out based on the statement of Preferred Reporting Items for Systematic Reviews and Meta-Analyses. Eight published studies, consisting of 7503 HCC patients (cases) and 13 831 individuals without HCC (controls), were included in the present study. Meta-analysis of the included studies revealed that STAT4 rs7574865 polymorphism contributed to the risk of HCC under all four genetic models, consisting of the allelic model (G vs. T: odds ratio [OR], 1.25; 95% confidence interval [CI], 1.19-1.30), the dominant effect model (GG + GT vs. TT: OR, 1.52; 95% CI, 1.26-1.84), the recessive effect model (GG vs. GT + TT: OR, 1.35; 95% CI, 1.21-1.50), and the co-dominant effect model (GG vs.. TT: OR, 1.72; 95% CI, 1.42-2.10) comparisons. No publication bias was indicated from either visualization of the funnel plot or Egger's test. A significantly increased risk of HCC associated with the rs7574865 G was found. The rs7574865 polymorphism might be used as one risk factor for HCC. © 2016 The Japan Society of Hepatology.

  8. A functional screen reveals an extensive layer of transcriptional and splicing control underlying RAS/MAPK signaling in Drosophila.

    Directory of Open Access Journals (Sweden)

    Dariel Ashton-Beaucage

    2014-03-01

    Full Text Available The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway--including a new protein complex modulating RAF activation--we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing

  9. Genome-Wide Spectra of Transcription Insertions and Deletions Reveal That Slippage Depends on RNA:DNA Hybrid Complementarity.

    Science.gov (United States)

    Traverse, Charles C; Ochman, Howard

    2017-08-29

    Advances in sequencing technologies have enabled direct quantification of genome-wide errors that occur during RNA transcription. These errors occur at rates that are orders of magnitude higher than rates during DNA replication, but due to technical difficulties such measurements have been limited to single-base substitutions and have not yet quantified the scope of transcription insertions and deletions. Previous reporter gene assay findings suggested that transcription indels are produced exclusively by elongation complex slippage at homopolymeric runs, so we enumerated indels across the protein-coding transcriptomes of Escherichia coli and Buchnera aphidicola , which differ widely in their genomic base compositions and incidence of repeat regions. As anticipated from prior assays, transcription insertions prevailed in homopolymeric runs of A and T; however, transcription deletions arose in much more complex sequences and were rarely associated with homopolymeric runs. By reconstructing the relocated positions of the elongation complex as inferred from the sequences inserted or deleted during transcription, we show that continuation of transcription after slippage hinges on the degree of nucleotide complementarity within the RNA:DNA hybrid at the new DNA template location. IMPORTANCE The high level of mistakes generated during transcription can result in the accumulation of malfunctioning and misfolded proteins which can alter global gene regulation and in the expenditure of energy to degrade these nonfunctional proteins. The transcriptome-wide occurrence of base substitutions has been elucidated in bacteria, but information on transcription insertions and deletions-errors that potentially have more dire effects on protein function-is limited to reporter gene constructs. Here, we capture the transcriptome-wide spectrum of insertions and deletions in Escherichia coli and Buchnera aphidicola and show that they occur at rates approaching those of base substitutions

  10. Structure reveals regulatory mechanisms of a MaoC-like hydratase from Phytophthora capsici involved in biosynthesis of polyhydroxyalkanoates (PHAs).

    Science.gov (United States)

    Wang, Huizheng; Zhang, Kai; Zhu, Jie; Song, Weiwei; Zhao, Li; Zhang, Xiuguo

    2013-01-01

    Polyhydroxyalkanoates (PHAs) have attracted increasing attention as "green plastic" due to their biodegradable, biocompatible, thermoplastic, and mechanical properties, and considerable research has been undertaken to develop low cost/high efficiency processes for the production of PHAs. MaoC-like hydratase (MaoC), which belongs to (R)-hydratase involved in linking the β-oxidation and the PHA biosynthetic pathways, has been identified recently. Understanding the regulatory mechanisms of (R)-hydratase catalysis is critical for efficient production of PHAs that promise synthesis an environment-friendly plastic. We have determined the crystal structure of a new MaoC recognized from Phytophthora capsici. The crystal structure of the enzyme was solved at 2.00 Å resolution. The structure shows that MaoC has a canonical (R)-hydratase fold with an N-domain and a C-domain. Supporting its dimerization observed in structure, MaoC forms a stable homodimer in solution. Mutations that disrupt the dimeric MaoC result in a complete loss of activity toward crotonyl-CoA, indicating that dimerization is required for the enzymatic activity of MaoC. Importantly, structure comparison reveals that a loop unique to MaoC interacts with an α-helix that harbors the catalytic residues of MaoC. Deletion of the loop enhances the enzymatic activity of MaoC, suggesting its inhibitory role in regulating the activity of MaoC. The data in our study reveal the regulatory mechanism of an (R)-hydratase, providing information on enzyme engineering to produce low cost PHAs.

  11. Structure reveals regulatory mechanisms of a MaoC-like hydratase from Phytophthora capsici involved in biosynthesis of polyhydroxyalkanoates (PHAs.

    Directory of Open Access Journals (Sweden)

    Huizheng Wang

    Full Text Available Polyhydroxyalkanoates (PHAs have attracted increasing attention as "green plastic" due to their biodegradable, biocompatible, thermoplastic, and mechanical properties, and considerable research has been undertaken to develop low cost/high efficiency processes for the production of PHAs. MaoC-like hydratase (MaoC, which belongs to (R-hydratase involved in linking the β-oxidation and the PHA biosynthetic pathways, has been identified recently. Understanding the regulatory mechanisms of (R-hydratase catalysis is critical for efficient production of PHAs that promise synthesis an environment-friendly plastic.We have determined the crystal structure of a new MaoC recognized from Phytophthora capsici. The crystal structure of the enzyme was solved at 2.00 Å resolution. The structure shows that MaoC has a canonical (R-hydratase fold with an N-domain and a C-domain. Supporting its dimerization observed in structure, MaoC forms a stable homodimer in solution. Mutations that disrupt the dimeric MaoC result in a complete loss of activity toward crotonyl-CoA, indicating that dimerization is required for the enzymatic activity of MaoC. Importantly, structure comparison reveals that a loop unique to MaoC interacts with an α-helix that harbors the catalytic residues of MaoC. Deletion of the loop enhances the enzymatic activity of MaoC, suggesting its inhibitory role in regulating the activity of MaoC.The data in our study reveal the regulatory mechanism of an (R-hydratase, providing information on enzyme engineering to produce low cost PHAs.

  12. Knock-down of transcript abundance of a family of Kunitz proteinase inhibitor genes in white clover (Trifolium repens) reveals a redundancy and diversity of gene function.

    Science.gov (United States)

    Islam, Afsana; Leung, Susanna; Burgess, Elisabeth P J; Laing, William A; Richardson, Kim A; Hofmann, Rainer W; Dijkwel, Paul P; McManus, Michael T

    2015-12-01

    The transcriptional regulation of four phylogenetically distinct members of a family of Kunitz proteinase inhibitor (KPI) genes isolated from white clover (Trifolium repens; designated Tr-KPI1, Tr-KPI2, Tr-KPI4 and Tr-KPI5) has been investigated to determine their wider functional role. The four genes displayed differential transcription during seed germination, and in different tissues of the mature plant, and transcription was also ontogenetically regulated. Heterologous over-expression of Tr-KPI1, Tr-KPI2, Tr-KPI4 and Tr-KPI5 in Nicotiana tabacum retarded larval growth of the herbivore Spodoptera litura, and an increase in the transcription of the pathogenesis-related genes PR1 and PR4 was observed in the Tr-KPI1 and Tr-KPI4 over-expressing lines. RNA interference (RNAi) knock-down lines in white clover displayed significantly altered vegetative growth phenotypes with inhibition of shoot growth and a stimulation of root growth, while knock-down of Tr-KPI1, Tr-KPI2 and Tr-KPI5 transcript abundance also retarded larval growth of S. litura. Examination of these RNAi lines revealed constitutive stress-associated phenotypes as well as altered transcription of cellular signalling genes. These results reveal a functional redundancy across members of the KPI gene family. Further, the regulation of transcription of at least one member of the family, Tr-KPI2, may occupy a central role in the maintenance of a cellular homeostasis. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

  13. Integration of transcriptomic and metabolic data reveals hub transcription factors involved in drought stress response in sunflower (Helianthus annuus L.).

    Science.gov (United States)

    Moschen, Sebastián; Di Rienzo, Julio A; Higgins, Janet; Tohge, Takayuki; Watanabe, Mutsumi; González, Sergio; Rivarola, Máximo; García-García, Francisco; Dopazo, Joaquin; Hopp, H Esteban; Hoefgen, Rainer; Fernie, Alisdair R; Paniego, Norma; Fernández, Paula; Heinz, Ruth A

    2017-07-01

    By integration of transcriptional and metabolic profiles we identified pathways and hubs transcription factors regulated during drought conditions in sunflower, useful for applications in molecular and/or biotechnological breeding. Drought is one of the most important environmental stresses that effects crop productivity in many agricultural regions. Sunflower is tolerant to drought conditions but the mechanisms involved in this tolerance remain unclear at the molecular level. The aim of this study was to characterize and integrate transcriptional and metabolic pathways related to drought stress in sunflower plants, by using a system biology approach. Our results showed a delay in plant senescence with an increase in the expression level of photosynthesis related genes as well as higher levels of sugars, osmoprotectant amino acids and ionic nutrients under drought conditions. In addition, we identified transcription factors that were upregulated during drought conditions and that may act as hubs in the transcriptional network. Many of these transcription factors belong to families implicated in the drought response in model species. The integration of transcriptomic and metabolomic data in this study, together with physiological measurements, has improved our understanding of the biological responses during droughts and contributes to elucidate the molecular mechanisms involved under this environmental condition. These findings will provide useful biotechnological tools to improve stress tolerance while maintaining crop yield under restricted water availability.

  14. Regulatory Interactions of Csr Components: the RNA Binding Protein CsrA Activates csrB Transcription in Escherichia coli

    OpenAIRE

    Gudapaty, Seshagirirao; Suzuki, Kazushi; Wang, Xin; Babitzke, Paul; Romeo, Tony

    2002-01-01

    The global regulator CsrA (carbon storage regulator) of Escherichia coli is a small RNA binding protein that represses various metabolic pathways and processes that are induced in the stationary phase of growth, while it activates certain exponential phase functions. Both repression and activation by CsrA involve posttranscriptional mechanisms, in which CsrA binding to mRNA leads to decreased or increased transcript stability, respectively. CsrA also binds to a small untranslated RNA, CsrB, f...

  15. Time-resolved transcriptome and proteome landscape of human regulatory T cell (Treg) differentiation reveals novel regulators of FOXP3

    KAUST Repository

    Schmidt, Angelika

    2018-04-27

    BackgroundRegulatory T cells (Tregs) expressing the transcription factor FOXP3 are crucial mediators of self-tolerance, preventing autoimmune diseases but possibly hampering tumor rejection. Clinical manipulation of Tregs is of great interest, and first-in-man trials of Treg transfer have achieved promising outcomes. Yet, the mechanisms governing induced Treg (iTreg) differentiation and the regulation of FOXP3 are incompletely understood.ResultsTo gain a comprehensive and unbiased molecular understanding of FOXP3 induction, we performed time-series RNA sequencing (RNA-Seq) and proteomics profiling on the same samples during human iTreg differentiation. To enable the broad analysis of universal FOXP3-inducing pathways, we used five differentiation protocols in parallel. Integrative analysis of the transcriptome and proteome confirmed involvement of specific molecular processes, as well as overlap of a novel iTreg subnetwork with known Treg regulators and autoimmunity-associated genes. Importantly, we propose 37 novel molecules putatively involved in iTreg differentiation. Their relevance was validated by a targeted shRNA screen confirming a functional role in FOXP3 induction, discriminant analyses classifying iTregs accordingly, and comparable expression in an independent novel iTreg RNA-Seq dataset.ConclusionThe data generated by this novel approach facilitates understanding of the molecular mechanisms underlying iTreg generation as well as of the concomitant changes in the transcriptome and proteome. Our results provide a reference map exploitable for future discovery of markers and drug candidates governing control of Tregs, which has important implications for the treatment of cancer, autoimmune, and inflammatory diseases.

  16. Functional Analysis of In-frame Indel ARID1A Mutations Reveals New Regulatory Mechanisms of Its Tumor Suppressor Functions

    Directory of Open Access Journals (Sweden)

    Bin Guan

    2012-10-01

    Full Text Available AT-rich interactive domain 1A (ARID1A has emerged as a new tumor suppressor in which frequent somatic mutations have been identified in several types of human cancers. Although most ARID1A somatic mutations are frame-shift or nonsense mutations that contribute to mRNA decay and loss of protein expression, 5% of ARID1A mutations are in-frame insertions or deletions (indels that involve only a small stretch of peptides. Naturally occurring in-frame indel mutations provide unique and useful models to explore the biology and regulatory role of ARID1A. In this study, we analyzed indel mutations identified in gynecological cancers to determine how these mutations affect the tumor suppressor function of ARID1A. Our results demonstrate that all in-frame mutants analyzed lost their ability to inhibit cellular proliferation or activate transcription of CDKN1A, which encodes p21, a downstream effector of ARID1A. We also showed that ARID1A is a nucleocytoplasmic protein whose stability depends on its subcellular localization. Nuclear ARID1A is less stable than cytoplasmic ARID1A because ARID1A is rapidly degraded by the ubiquitin-proteasome system in the nucleus. In-frame deletions affecting the consensus nuclear export signal reduce steady-state protein levels of ARID1A. This defect in nuclear exportation leads to nuclear retention and subsequent degradation. Our findings delineate a mechanism underlying the regulation of ARID1A subcellular distribution and protein stability and suggest that targeting the nuclear ubiquitin-proteasome system can increase the amount of the ARID1A protein in the nucleus and restore its tumor suppressor functions.

  17. The transcriptional regulatory network in the drought response and its crosstalk in abiotic stress responses including drought, cold and heat

    Directory of Open Access Journals (Sweden)

    Kazuo eNakashima

    2014-05-01

    Full Text Available Drought negatively impacts plant growth and the productivity of crops around the world. Understanding the molecular mechanisms in the drought response is important for improvement of drought tolerance using molecular techniques. In plants, abscisic acid (ABA is accumulated under osmotic stress conditions caused by drought, and has a key role in stress responses and tolerance. Comprehensive molecular analyses have shown that ABA regulates the expression of many genes under osmotic stress conditions, and the ABA-responsive element (ABRE is the major cis-element for ABA-responsive gene expression. Transcription factors (TFs are master regulators of gene expression. ABRE-binding protein (AREB and ABRE-binding factor (ABF TFs control gene expression in an ABA-dependent manner. SNF1-related protein kinases 2, group A 2C-type protein phosphatases, and ABA receptors were shown to control the ABA signaling pathway. ABA-independent signaling pathways such as dehydration-responsive element-binding protein (DREB TFs and NAC TFs are also involved in stress responses including drought, heat and cold. Recent studies have suggested that there are interactions between the major ABA signaling pathway and other signaling factors in stress responses. The important roles of these transcription factors in crosstalk among abiotic stress responses will be discussed. Control of ABA or stress signaling factor expression can improve tolerance to environmental stresses. Recent studies using crops have shown that stress-specific overexpression of TFs improves drought tolerance and grain yield compared with controls in the field.

  18. Regulatory Shifts in Plastid Transcription Play a Key Role in Morphological Conversions of Plastids during Plant Development.

    Science.gov (United States)

    Liebers, Monique; Grübler, Björn; Chevalier, Fabien; Lerbs-Mache, Silva; Merendino, Livia; Blanvillain, Robert; Pfannschmidt, Thomas

    2017-01-01

    Plastids display a high morphological and functional diversity. Starting from an undifferentiated small proplastid, these plant cell organelles can develop into four major forms: etioplasts in the dark, chloroplasts in green tissues, chromoplasts in colored flowers and fruits and amyloplasts in roots. The various forms are interconvertible into each other depending on tissue context and respective environmental condition. Research of the last two decades uncovered that each plastid type contains its own specific proteome that can be highly different from that of the other types. Composition of these proteomes largely defines the enzymatic functionality of the respective plastid. The vast majority of plastid proteins is encoded in the nucleus and must be imported from the cytosol. However, a subset of proteins of the photosynthetic and gene expression machineries are encoded on the plastid genome and are transcribed by a complex transcriptional apparatus consisting of phage-type nuclear-encoded RNA polymerases and a bacterial-type plastid-encoded RNA polymerase. Both types recognize specific sets of promoters and transcribe partly over-lapping as well as specific sets of genes. Here we summarize the current knowledge about the sequential activity of these plastid RNA polymerases and their relative activities in different types of plastids. Based on published plastid gene expression profiles we hypothesize that each conversion from one plastid type into another is either accompanied or even preceded by significant changes in plastid transcription suggesting that these changes represent important determinants of plastid morphology and protein composition and, hence, the plastid type.

  19. Analysis of transcriptional regulatory pathways of photoreceptor genes by expression profiling of the Otx2-deficient retina.

    Science.gov (United States)

    Omori, Yoshihiro; Katoh, Kimiko; Sato, Shigeru; Muranishi, Yuki; Chaya, Taro; Onishi, Akishi; Minami, Takashi; Fujikado, Takashi; Furukawa, Takahisa

    2011-01-01

    In the vertebrate retina, the Otx2 transcription factor plays a crucial role in the cell fate determination of both rod and cone photoreceptors. We previously reported that Otx2 conditional knockout (CKO) mice exhibited a total absence of rods and cones in the retina due to their cell fate conversion to amacrine-like cells. In order to investigate the entire transcriptome of the Otx2 CKO retina, we compared expression profile of Otx2 CKO and wild-type retinas at P1 and P12 using microarray. We observed that expression of 101- and 1049-probe sets significantly decreased in the Otx2 CKO retina at P1 and P12, respectively, whereas, expression of 3- and 4149-probe sets increased at P1 and P12, respectively. We found that expression of genes encoding transcription factors involved in photoreceptor development, including Crx, Nrl, Nr2e3, Esrrb, and NeuroD, was markedly down-regulated in the Otx2 CKO at both P1 and P12. Furthermore, we identified three human retinal disease loci mapped in close proximity to certain down-regulated genes in the Otx2 CKO retina including Ccdc126, Tnfsf13 and Pitpnm1, suggesting that these genes are possibly responsible for these diseases. These transcriptome data sets of the Otx2 CKO retina provide a resource on developing rods and cones to further understand the molecular mechanisms underlying photoreceptor development, function and disease.

  20. Analysis of transcriptional regulatory pathways of photoreceptor genes by expression profiling of the Otx2-deficient retina.

    Directory of Open Access Journals (Sweden)

    Yoshihiro Omori

    Full Text Available In the vertebrate retina, the Otx2 transcription factor plays a crucial role in the cell fate determination of both rod and cone photoreceptors. We previously reported that Otx2 conditional knockout (CKO mice exhibited a total absence of rods and cones in the retina due to their cell fate conversion to amacrine-like cells. In order to investigate the entire transcriptome of the Otx2 CKO retina, we compared expression profile of Otx2 CKO and wild-type retinas at P1 and P12 using microarray. We observed that expression of 101- and 1049-probe sets significantly decreased in the Otx2 CKO retina at P1 and P12, respectively, whereas, expression of 3- and 4149-probe sets increased at P1 and P12, respectively. We found that expression of genes encoding transcription factors involved in photoreceptor development, including Crx, Nrl, Nr2e3, Esrrb, and NeuroD, was markedly down-regulated in the Otx2 CKO at both P1 and P12. Furthermore, we identified three human retinal disease loci mapped in close proximity to certain down-regulated genes in the Otx2 CKO retina including Ccdc126, Tnfsf13 and Pitpnm1, suggesting that these genes are possibly responsible for these diseases. These transcriptome data sets of the Otx2 CKO retina provide a resource on developing rods and cones to further understand the molecular mechanisms underlying photoreceptor development, function and disease.

  1. Proteome-wide Structural Analysis of PTM Hotspots Reveals Regulatory Elements Predicted to Impact Biological Function and Disease*

    Science.gov (United States)

    Dewhurst, Henry; Sundararaman, Niveda

    2016-01-01

    Post-translational modifications (PTMs) regulate protein behavior through modulation of protein-protein interactions, enzymatic activity, and protein stability essential in the translation of genotype to phenotype in eukaryotes. Currently, less than 4% of all eukaryotic PTMs are reported to have biological function - a statistic that continues to decrease with an increasing rate of PTM detection. Previously, we developed SAPH-ire (Structural Analysis of PTM Hotspots) - a method for the prioritization of PTM function potential that has been used effectively to reveal novel PTM regulatory elements in discrete protein families (Dewhurst et al., 2015). Here, we apply SAPH-ire to the set of eukaryotic protein families containing experimental PTM and 3D structure data - capturing 1,325 protein families with 50,839 unique PTM sites organized into 31,747 modified alignment positions (MAPs), of which 2010 (∼6%) possess known biological function. Here, we show that using an artificial neural network model (SAPH-ire NN) trained to identify MAP hotspots with biological function results in prediction outcomes that far surpass the use of single hotspot features, including nearest neighbor PTM clustering methods. We find the greatest enhancement in prediction for positions with PTM counts of five or less, which represent 98% of all MAPs in the eukaryotic proteome and 90% of all MAPs found to have biological function. Analysis of the top 1092 MAP hotspots revealed 267 of truly unknown function (containing 5443 distinct PTMs). Of these, 165 hotspots could be mapped to human KEGG pathways for normal and/or disease physiology. Many high-ranking hotspots were also found to be disease-associated pathogenic sites of amino acid substitution despite the lack of observable PTM in the human protein family member. Taken together, these experiments demonstrate that the functional relevance of a PTM can be predicted very effectively by neural network models, revealing a large but testable

  2. Proteome-wide Structural Analysis of PTM Hotspots Reveals Regulatory Elements Predicted to Impact Biological Function and Disease.

    Science.gov (United States)

    Torres, Matthew P; Dewhurst, Henry; Sundararaman, Niveda

    2016-11-01

    Post-translational modifications (PTMs) regulate protein behavior through modulation of protein-protein interactions, enzymatic activity, and protein stability essential in the translation of genotype to phenotype in eukaryotes. Currently, less than 4% of all eukaryotic PTMs are reported to have biological function - a statistic that continues to decrease with an increasing rate of PTM detection. Previously, we developed SAPH-ire (Structural Analysis of PTM Hotspots) - a method for the prioritization of PTM function potential that has been used effectively to reveal novel PTM regulatory elements in discrete protein families (Dewhurst et al., 2015). Here, we apply SAPH-ire to the set of eukaryotic protein families containing experimental PTM and 3D structure data - capturing 1,325 protein families with 50,839 unique PTM sites organized into 31,747 modified alignment positions (MAPs), of which 2010 (∼6%) possess known biological function. Here, we show that using an artificial neural network model (SAPH-ire NN) trained to identify MAP hotspots with biological function results in prediction outcomes that far surpass the use of single hotspot features, including nearest neighbor PTM clustering methods. We find the greatest enhancement in prediction for positions with PTM counts of five or less, which represent 98% of all MAPs in the eukaryotic proteome and 90% of all MAPs found to have biological function. Analysis of the top 1092 MAP hotspots revealed 267 of truly unknown function (containing 5443 distinct PTMs). Of these, 165 hotspots could be mapped to human KEGG pathways for normal and/or disease physiology. Many high-ranking hotspots were also found to be disease-associated pathogenic sites of amino acid substitution despite the lack of observable PTM in the human protein family member. Taken together, these experiments demonstrate that the functional relevance of a PTM can be predicted very effectively by neural network models, revealing a large but testable

  3. Positive- and negative-acting regulatory elements contribute to the tissue-specific expression of INNER NO OUTER, a YABBY-type transcription factor gene in Arabidopsis

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    Simon Marissa K

    2012-11-01

    Full Text Available Abstract Background The INNER NO OUTER (INO gene, which encodes a YABBY-type transcription factor, specifies and promotes the growth of the outer integument of the ovule in Arabidopsis. INO expression is limited to the abaxial cell layer of the developing outer integument of the ovule and is regulated by multiple regions of the INO promoter, including POS9, a positive element that when present in quadruplicate can produce low-level expression in the normal INO pattern. Results Significant redundancy in activity between different regions of the INO promoter is demonstrated. For specific regulatory elements, multimerization or the addition of the cauliflower mosaic virus 35S general enhancer was able to activate expression of reporter gene constructs that were otherwise incapable of expression on their own. A new promoter element, POS6, is defined and is shown to include sufficient positive regulatory information to reproduce the endogenous pattern of expression in ovules, but other promoter regions are necessary to fully suppress expression outside of ovules. The full-length INO promoter, but not any of the INO promoter deletions tested, is able to act as an enhancer-blocking insulator to prevent the ectopic activation of expression by the 35S enhancer. Sequence conservation between the promoter regions of Arabidopsis thaliana, Brassica oleracea and Brassica rapa aligns closely with the functional definition of the POS6 and POS9 regions, and with a defined INO minimal promoter. The B. oleracea INO promoter is sufficient to promote a similar pattern and level of reporter gene expression in Arabidopsis to that observed for the Arabidopsis promoter. Conclusions At least two independent regions of the INO promoter contain sufficient regulatory information to direct the specific pattern but not the level of INO gene expression. These regulatory regions act in a partially redundant manner to promote the expression in a specific pattern in the ovule and

  4. The transcriptional programme of Salmonella enterica serovar Typhimurium reveals a key role for tryptophan metabolism in biofilms.

    LENUS (Irish Health Repository)

    Hamilton, Shea

    2009-12-11

    Abstract Background Biofilm formation enhances the capacity of pathogenic Salmonella bacteria to survive stresses that are commonly encountered within food processing and during host infection. The persistence of Salmonella within the food chain has become a major health concern, as biofilms can serve as a reservoir for the contamination of food products. While the molecular mechanisms required for the survival of bacteria on surfaces are not fully understood, transcriptional studies of other bacteria have demonstrated that biofilm growth triggers the expression of specific sets of genes, compared with planktonic cells. Until now, most gene expression studies of Salmonella have focused on the effect of infection-relevant stressors on virulence or the comparison of mutant and wild-type bacteria. However little is known about the physiological responses taking place inside a Salmonella biofilm. Results We have determined the transcriptomic and proteomic profiles of biofilms of Salmonella enterica serovar Typhimurium. We discovered that 124 detectable proteins were differentially expressed in the biofilm compared with planktonic cells, and that 10% of the S. Typhimurium genome (433 genes) showed a 2-fold or more change in the biofilm compared with planktonic cells. The genes that were significantly up-regulated implicated certain cellular processes in biofilm development including amino acid metabolism, cell motility, global regulation and tolerance to stress. We found that the most highly down-regulated genes in the biofilm were located on Salmonella Pathogenicity Island 2 (SPI2), and that a functional SPI2 secretion system regulator (ssrA) was required for S. Typhimurium biofilm formation. We identified STM0341 as a gene of unknown function that was needed for biofilm growth. Genes involved in tryptophan (trp) biosynthesis and transport were up-regulated in the biofilm. Deletion of trpE led to decreased bacterial attachment and this biofilm defect was restored by

  5. The lncRNA Malat1 Is Dispensable for Mouse Development but Its Transcription Plays a cis-Regulatory Role in the Adult

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    Bin Zhang

    2012-07-01

    Full Text Available Genome-wide studies have identified thousands of long noncoding RNAs (lncRNAs lacking protein-coding capacity. However, most lncRNAs are expressed at a very low level, and in most cases there is no genetic evidence to support their in vivo function. Malat1 (metastasis associated lung adenocarcinoma transcript 1 is among the most abundant and highly conserved lncRNAs, and it exhibits an uncommon 3′-end processing mechanism. In addition, its specific nuclear localization, developmental regulation, and dysregulation in cancer are suggestive of it having a critical biological function. We have characterized a Malat1 loss-of-function genetic model that indicates that Malat1 is not essential for mouse pre- and postnatal development. Furthermore, depletion of Malat1 does not affect global gene expression, splicing factor level and phosphorylation status, or alternative pre-mRNA splicing. However, among a small number of genes that were dysregulated in adult Malat1 knockout mice, many were Malat1 neighboring genes, thus indicating a potential cis-regulatory role of Malat1 gene transcription.

  6. Transcriptional Profiling of Saccharomyces cerevisiae Reveals the Impact of Variation of a Single Transcription Factor on Differential Gene Expression in 4NQO, Fermentable, and Nonfermentable Carbon Sources

    Directory of Open Access Journals (Sweden)

    Xiaoqing Rong-Mullins

    2018-02-01

    Full Text Available Cellular metabolism can change the potency of a chemical’s tumorigenicity. 4-nitroquinoline-1-oxide (4NQO is a tumorigenic drug widely used on animal models for cancer research. Polymorphisms of the transcription factor Yrr1 confer different levels of resistance to 4NQO in Saccharomyces cerevisiae. To study how different Yrr1 alleles regulate gene expression leading to resistance, transcriptomes of three isogenic S. cerevisiae strains carrying different Yrr1 alleles were profiled via RNA sequencing (RNA-Seq and chromatin immunoprecipitation coupled with sequencing (ChIP-Seq in the presence and absence of 4NQO. In response to 4NQO, all alleles of Yrr1 drove the expression of SNQ2 (a multidrug transporter, which was highest in the presence of 4NQO resistance-conferring alleles, and overexpression of SNQ2 alone was sufficient to overcome 4NQO-sensitive growth. Using shape metrics to refine the ChIP-Seq peaks, Yrr1 strongly associated with three loci including SNQ2. In addition to a known Yrr1 target SNG1, Yrr1 also bound upstream of RPL35B; however, overexpression of these genes did not confer 4NQO resistance. RNA-Seq data also implicated nucleotide synthesis pathways including the de novo purine pathway, and the ribonuclease reductase pathways were downregulated in response to 4NQO. Conversion of a 4NQO-sensitive allele to a 4NQO-resistant allele by a single point mutation mimicked the 4NQO-resistant allele in phenotype, and while the 4NQO resistant allele increased the expression of the ADE genes in the de novo purine biosynthetic pathway, the mutant Yrr1 increased expression of ADE genes even in the absence of 4NQO. These same ADE genes were only increased in the wild-type alleles in the presence of 4NQO, indicating that the point mutation activated Yrr1 to upregulate a pathway normally only activated in response to stress. The various Yrr1 alleles also influenced growth on different carbon sources by altering the function of the mitochondria

  7. Differential proteomic analysis reveals sequential heat stress-responsive regulatory network in radish (Raphanus sativus L.) taproot.

    Science.gov (United States)

    Wang, Ronghua; Mei, Yi; Xu, Liang; Zhu, Xianwen; Wang, Yan; Guo, Jun; Liu, Liwang

    2018-05-01

    Differential abundance protein species (DAPS) involved in reducing damage and enhancing thermotolerance in radish were firstly identified. Proteomic analysis and omics association analysis revealed a HS-responsive regulatory network in radish. Heat stress (HS) is a major destructive factor influencing radish production and supply in summer, for radish is a cool season vegetable crop being susceptible to high temperature. In this study, the proteome changes of radish taproots under 40 °C treatment at 0 h (Control), 12 h (Heat12) and 24 h (Heat24) were analyzed using iTRAQ (Isobaric Tag for Relative and Absolute Quantification) approach. In total, 2258 DAPS representing 1542 differentially accumulated uniprotein species which respond to HS were identified. A total of 604, 910 and 744 DAPS was detected in comparison of Control vs. Heat12, Control vs. Heat24, and Heat12 vs. Heat24, respectively. Gene ontology and pathway analysis showed that annexin, ubiquitin-conjugating enzyme, ATP synthase, heat shock protein (HSP) and other stress-related proteins were predominately enriched in signal transduction, stress and defense pathways, photosynthesis and energy metabolic pathways, working cooperatively to reduce stress-induced damage in radish. Based on iTRAQ combined with the transcriptomics analysis, a schematic model of a sequential HS-responsive regulatory network was proposed. The initial sensing of HS occurred at the plasma membrane, and then key components of stress signal transduction triggered heat-responsive genes in the plant protective metabolism to re-establish homeostasis and enhance thermotolerance. These results provide new insights into characteristics of HS-responsive DAPS and facilitate dissecting the molecular mechanisms underlying heat tolerance in radish and other root crops.

  8. Pathway-based analysis of genome-wide siRNA screens reveals the regulatory landscape of APP processing.

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    Luiz Miguel Camargo

    Full Text Available The progressive aggregation of Amyloid-β (Aβ in the brain is a major trait of Alzheimer's Disease (AD. Aβ is produced as a result of proteolytic processing of the β-amyloid precursor protein (APP. Processing of APP is mediated by multiple enzymes, resulting in the production of distinct peptide products: the non-amyloidogenic peptide sAPPα and the amyloidogenic peptides sAPPβ, Aβ40, and Aβ42. Using a pathway-based approach, we analyzed a large-scale siRNA screen that measured the production of different APP proteolytic products. Our analysis identified many of the biological processes/pathways that are known to regulate APP processing and have been implicated in AD pathogenesis, as well as revealing novel regulatory mechanisms. Furthermore, we also demonstrate that some of these processes differentially regulate APP processing, with some mechanisms favouring production of certain peptide species over others. For example, synaptic transmission having a bias towards regulating Aβ40 production over Aβ42 as well as processes involved in insulin and pancreatic biology having a bias for sAPPβ production over sAPPα. In addition, some of the pathways identified as regulators of APP processing contain genes (CLU, BIN1, CR1, PICALM, TREM2, SORL1, MEF2C, DSG2, EPH1A recently implicated with AD through genome wide association studies (GWAS and associated meta-analysis. In addition, we provide supporting evidence and a deeper mechanistic understanding of the role of diabetes in AD. The identification of these processes/pathways, their differential impact on APP processing, and their relationships to each other, provide a comprehensive systems biology view of the "regulatory landscape" of APP.

  9. Hepatitis B virus nuclear export elements: RNA stem-loop α and β, key parts of the HBV post-transcriptional regulatory element.

    Science.gov (United States)

    Lim, Chun Shen; Brown, Chris M

    2016-09-01

    Many viruses contain RNA elements that modulate splicing and/or promote nuclear export of their RNAs. The RNAs of the major human pathogen, hepatitis B virus (HBV) contain a large (~600 bases) composite cis-acting 'post-transcriptional regulatory element' (PRE). This element promotes expression from these naturally intronless transcripts. Indeed, the related woodchuck hepadnavirus PRE (WPRE) is used to enhance expression in gene therapy and other expression vectors. These PRE are likely to act through a combination of mechanisms, including promotion of RNA nuclear export. Functional components of both the HBV PRE and WPRE are 2 conserved RNA cis-acting stem-loop (SL) structures, SLα and SLβ. They are within the coding regions of polymerase (P) gene, and both P and X genes, respectively. Based on previous studies using mutagenesis and/or nuclear magnetic resonance (NMR), here we propose 2 covariance models for SLα and SLβ. The model for the 30-nucleotide SLα contains a G-bulge and a CNGG(U) apical loop of which the first and the fourth loop residues form a CG pair and the fifth loop residue is bulged out, as observed in the NMR structure. The model for the 23-nucleotide SLβ contains a 7-base-pair stem and a 9-nucleotide loop. Comparison of the models with other RNA structural elements, as well as similarity searches of human transcriptome and viral genomes demonstrate that SLα and SLβ are specific to HBV transcripts. However, they are well conserved among the hepadnaviruses of non-human primates, the woodchuck and ground squirrel.

  10. Systematic analysis of phloem-feeding insect-induced transcriptional reprogramming in Arabidopsis highlights common features and reveals distinct responses to specialist and generalist insects.

    Science.gov (United States)

    Foyer, Christine H; Verrall, Susan R; Hancock, Robert D

    2015-02-01

    Phloem-feeding insects (PFIs), of which aphids are the largest group, are major agricultural pests causing extensive damage to crop plants. In contrast to chewing insects, the nature of the plant response to PFIs remains poorly characterized. Scrutiny of the literature concerning transcriptional responses of model and crop plant species to PFIs reveals surprisingly little consensus with respect to the transcripts showing altered abundance following infestation. Nevertheless, core features of the transcriptional response to PFIs can be defined in Arabidopsis thaliana. This comparison of the PFI-associated transcriptional response observed in A. thaliana infested by the generalists Myzus persicae and Bemisia tabaci with the specialist Brevicoryne brassicae highlights the importance of calcium-dependent and receptor kinase-associated signalling. We discuss these findings within the context of the complex cross-talk between the different hormones regulating basal immune response mechanisms in plants. We identify PFI-responsive genes, highlighting the importance of cell wall-associated kinases in plant-PFI interactions, as well as the significant role of kinases containing the domain of unknown function 26. A common feature of plant-PFI interaction is enhanced abundance of transcripts encoding WRKY transcription factors. However, significant divergence was observed with respect to secondary metabolism dependent upon the insect attacker. Transcripts encoding enzymes and proteins associated with glucosinolate metabolism were decreased following attack by the generalist M. persicae but not by the specialist B. brassicae. This analysis provides a comprehensive overview of the molecular patterns associated with the plant response to PFIs and suggests that plants recognize and respond to perturbations in the cell wall occurring during PFI infestation. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights

  11. Revealing genome-scale transcriptional regulatory landscape of OmpR highlights its expanded regulatory roles under osmotic stress in Escherichia coli K-12 MG1655

    DEFF Research Database (Denmark)

    Seo, Sang Woo; Gao, Ye; Kim, Donghyuk

    2017-01-01

    ) belong to OmpR regulon. Among them, 26 genes show more than two-fold changes in expression level in an OmpR knock-out strain. Specifically, we find that: 1) OmpR regulates mostly membrane-located gene products involved in diverse fundamental biological processes, such as narU (encoding nitrate/nitrite...

  12. Screening of the transcriptional regulatory regions of vascular endothelial growth factor receptor 2 (VEGFR2 in amyotrophic lateral sclerosis

    Directory of Open Access Journals (Sweden)

    Hartley Judith

    2007-04-01

    Full Text Available Abstract Background Vascular endothelial growth factor (VEGF has neurotrophic activity which is mediated by its main agonist receptor, VEGFR2. Dysregulation of VEGF causes motor neurone degeneration in a mouse model of amyotrophic lateral sclerosis (ALS, and expression of VEGFR2 is reduced in motor neurones and spinal cord of patients with ALS. Methods We have screened the promoter region and 4 exonic regions of functional significance of the VEGFR2 gene in a UK population of patients with ALS, for mutations and polymorphisms that may affect expression or function of this VEGF receptor. Results No mutations were identified in the VEGFR2 gene. We found no association between polymorphisms in the regulatory regions of the VEGFR2 gene and ALS. Conclusion Mechanisms other than genetic variation may downregulate expression or function of the VEGFR2 receptor in patients with ALS.

  13. Single-Cell RNA-Seq Reveals Transcriptional Heterogeneity in Latent and Reactivated HIV-Infected Cells.

    Science.gov (United States)

    Golumbeanu, Monica; Cristinelli, Sara; Rato, Sylvie; Munoz, Miguel; Cavassini, Matthias; Beerenwinkel, Niko; Ciuffi, Angela

    2018-04-24

    Despite effective treatment, HIV can persist in latent reservoirs, which represent a major obstacle toward HIV eradication. Targeting and reactivating latent cells is challenging due to the heterogeneous nature of HIV-infected cells. Here, we used a primary model of HIV latency and single-cell RNA sequencing to characterize transcriptional heterogeneity during HIV latency and reactivation. Our analysis identified transcriptional programs leading to successful reactivation of HIV expression. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  14. Multiple regulatory roles of the mouse transmembrane adaptor protein NTAL in gene transcription and mast cell physiology.

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    Iva Polakovicova

    Full Text Available Non-T cell activation linker (NTAL; also called LAB or LAT2 is a transmembrane adaptor protein that is expressed in a subset of hematopoietic cells, including mast cells. There are conflicting reports on the role of NTAL in the high affinity immunoglobulin E receptor (FcεRI signaling. Studies carried out on mast cells derived from mice with NTAL knock out (KO and wild type mice suggested that NTAL is a negative regulator of FcεRI signaling, while experiments with RNAi-mediated NTAL knockdown (KD in human mast cells and rat basophilic leukemia cells suggested its positive regulatory role. To determine whether different methodologies of NTAL ablation (KO vs KD have different physiological consequences, we compared under well defined conditions FcεRI-mediated signaling events in mouse bone marrow-derived mast cells (BMMCs with NTAL KO or KD. BMMCs with both NTAL KO and KD exhibited enhanced degranulation, calcium mobilization, chemotaxis, tyrosine phosphorylation of LAT and ERK, and depolymerization of filamentous actin. These data provide clear evidence that NTAL is a negative regulator of FcεRI activation events in murine BMMCs, independently of possible compensatory developmental alterations. To gain further insight into the role of NTAL in mast cells, we examined the transcriptome profiles of resting and antigen-activated NTAL KO, NTAL KD, and corresponding control BMMCs. Through this analysis we identified several genes that were differentially regulated in nonactivated and antigen-activated NTAL-deficient cells, when compared to the corresponding control cells. Some of the genes seem to be involved in regulation of cholesterol-dependent events in antigen-mediated chemotaxis. The combined data indicate multiple regulatory roles of NTAL in gene expression and mast cell physiology.

  15. A compact, in vivo screen of all 6-mers reveals drivers of tissue-specific expression and guides synthetic regulatory element design.

    Science.gov (United States)

    Smith, Robin P; Riesenfeld, Samantha J; Holloway, Alisha K; Li, Qiang; Murphy, Karl K; Feliciano, Natalie M; Orecchia, Lorenzo; Oksenberg, Nir; Pollard, Katherine S; Ahituv, Nadav

    2013-07-18

    Large-scale annotation efforts have improved our ability to coarsely predict regulatory elements throughout vertebrate genomes. However, it is unclear how complex spatiotemporal patterns of gene expression driven by these elements emerge from the activity of short, transcription factor binding sequences. We describe a comprehensive promoter extension assay in which the regulatory potential of all 6 base-pair (bp) sequences was tested in the context of a minimal promoter. To enable this large-scale screen, we developed algorithms that use a reverse-complement aware decomposition of the de Bruijn graph to design a library of DNA oligomers incorporating every 6-bp sequence exactly once. Our library multiplexes all 4,096 unique 6-mers into 184 double-stranded 15-bp oligomers, which is sufficiently compact for in vivo testing. We injected each multiplexed construct into zebrafish embryos and scored GFP expression in 15 tissues at two developmental time points. Twenty-seven constructs produced consistent expression patterns, with the majority doing so in only one tissue. Functional sequences are enriched near biologically relevant genes, match motifs for developmental transcription factors, and are required for enhancer activity. By concatenating tissue-specific functional sequences, we generated completely synthetic enhancers for the notochord, epidermis, spinal cord, forebrain and otic lateral line, and show that short regulatory sequences do not always function modularly. This work introduces a unique in vivo catalog of short, functional regulatory sequences and demonstrates several important principles of regulatory element organization. Furthermore, we provide resources for designing compact, reverse-complement aware k-mer libraries.

  16. Untangling the Effect of Fatty Acid Addition at Species Level Revealed Different Transcriptional Responses of the Biogas Microbial Community Members

    DEFF Research Database (Denmark)

    Treu, Laura; Campanaro, Stefano; Kougias, Panagiotis

    2016-01-01

    In the present study, RNA-sequencing was used to elucidate the change of anaerobic digestion metatranscriptome after long chain fatty acids (oleate) exposure. To explore the general transcriptional behavior of the microbiome, the analysis was first performed on shotgun reads without considering...

  17. Network-directed cis-mediator analysis of normal prostate tissue expression profiles reveals downstream regulatory associations of prostate cancer susceptibility loci.

    Science.gov (United States)

    Larson, Nicholas B; McDonnell, Shannon K; Fogarty, Zach; Larson, Melissa C; Cheville, John; Riska, Shaun; Baheti, Saurabh; Weber, Alexandra M; Nair, Asha A; Wang, Liang; O'Brien, Daniel; Davila, Jaime; Schaid, Daniel J; Thibodeau, Stephen N

    2017-10-17

    Large-scale genome-wide association studies have identified multiple single-nucleotide polymorphisms associated with risk of prostate cancer. Many of these genetic variants are presumed to be regulatory in nature; however, follow-up expression quantitative trait loci (eQTL) association studies have to-date been restricted largely to cis -acting associations due to study limitations. While trans -eQTL scans suffer from high testing dimensionality, recent evidence indicates most trans -eQTL associations are mediated by cis -regulated genes, such as transcription factors. Leveraging a data-driven gene co-expression network, we conducted a comprehensive cis -mediator analysis using RNA-Seq data from 471 normal prostate tissue samples to identify downstream regulatory associations of previously identified prostate cancer risk variants. We discovered multiple trans -eQTL associations that were significantly mediated by cis -regulated transcripts, four of which involved risk locus 17q12, proximal transcription factor HNF1B , and target trans -genes with known HNF response elements ( MIA2 , SRC , SEMA6A , KIF12 ). We additionally identified evidence of cis -acting down-regulation of MSMB via rs10993994 corresponding to reduced co-expression of NDRG1 . The majority of these cis -mediator relationships demonstrated trans -eQTL replicability in 87 prostate tissue samples from the Gene-Tissue Expression Project. These findings provide further biological context to known risk loci and outline new hypotheses for investigation into the etiology of prostate cancer.

  18. The transcriptional regulatory network in the drought response and its crosstalk in abiotic stress responses including drought, cold, and heat.

    Science.gov (United States)

    Nakashima, Kazuo; Yamaguchi-Shinozaki, Kazuko; Shinozaki, Kazuo

    2014-01-01

    Drought negatively impacts plant growth and the productivity of crops around the world. Understanding the molecular mechanisms in the drought response is important for improvement of drought tolerance using molecular techniques. In plants, abscisic acid (ABA) is accumulated under osmotic stress conditions caused by drought, and has a key role in stress responses and tolerance. Comprehensive molecular analyses have shown that ABA regulates the expression of many genes under osmotic stress conditions, and the ABA-responsive element (ABRE) is the major cis-element for ABA-responsive gene expression. Transcription factors (TFs) are master regulators of gene expression. ABRE-binding protein and ABRE-binding factor TFs control gene expression in an ABA-dependent manner. SNF1-related protein kinases 2, group A 2C-type protein phosphatases, and ABA receptors were shown to control the ABA signaling pathway. ABA-independent signaling pathways such as dehydration-responsive element-binding protein TFs and NAC TFs are also involved in stress responses including drought, heat, and cold. Recent studies have suggested that there are interactions between the major ABA signaling pathway and other signaling factors in stress responses. The important roles of these TFs in crosstalk among abiotic stress responses will be discussed. Control of ABA or stress signaling factor expression can improve tolerance to environmental stresses. Recent studies using crops have shown that stress-specific overexpression of TFs improves drought tolerance and grain yield compared with controls in the field.

  19. Integrated analysis of hematopoietic differentiation outcomes and molecular characterization reveals unbiased differentiation capacity and minor transcriptional memory in HPC/HSC-iPSCs.

    Science.gov (United States)

    Gao, Shuai; Hou, Xinfeng; Jiang, Yonghua; Xu, Zijian; Cai, Tao; Chen, Jiajie; Chang, Gang

    2017-01-23

    Transcription factor-mediated reprogramming can reset the epigenetics of somatic cells into a pluripotency compatible state. Recent studies show that induced pluripotent stem cells (iPSCs) always inherit starting cell-specific characteristics, called epigenetic memory, which may be advantageous, as directed differentiation into specific cell types is still challenging; however, it also may be unpredictable when uncontrollable differentiation occurs. In consideration of biosafety in disease modeling and personalized medicine, the availability of high-quality iPSCs which lack a biased differentiation capacity and somatic memory could be indispensable. Herein, we evaluate the hematopoietic differentiation capacity and somatic memory state of hematopoietic progenitor and stem cell (HPC/HSC)-derived-iPSCs (HPC/HSC-iPSCs) using a previously established sequential reprogramming system. We found that HPC/HSCs are amenable to being reprogrammed into iPSCs with unbiased differentiation capacity to hematopoietic progenitors and mature hematopoietic cells. Genome-wide analyses revealed that no global epigenetic memory was detectable in HPC/HSC-iPSCs, but only a minor transcriptional memory of HPC/HSCs existed in a specific tetraploid complementation (4 N)-incompetent HPC/HSC-iPSC line. However, the observed minor transcriptional memory had no influence on the hematopoietic differentiation capacity, indicating the reprogramming of the HPC/HSCs was nearly complete. Further analysis revealed the correlation of minor transcriptional memory with the aberrant distribution of H3K27me3. This work provides a comprehensive framework for obtaining high-quality iPSCs from HPC/HSCs with unbiased hematopoietic differentiation capacity and minor transcriptional memory.

  20. Multi-Omics and Integrated Network Analyses Reveal New Insights into the Systems Relationships between Metabolites, Structural Genes, and Transcriptional Regulators in Developing Grape Berries (Vitis vinifera L. Exposed to Water Deficit

    Directory of Open Access Journals (Sweden)

    Stefania Savoi

    2017-07-01

    Full Text Available Grapes are one of the major fruit crops and they are cultivated in many dry environments. This study comprehensively characterizes the metabolic response of grape berries exposed to water deficit at different developmental stages. Increases of proline, branched-chain amino acids, phenylpropanoids, anthocyanins, and free volatile organic compounds have been previously observed in grape berries exposed to water deficit. Integrating RNA-sequencing analysis of the transcriptome with large-scale analysis of central and specialized metabolites, we reveal that these increases occur via a coordinated regulation of key structural pathway genes. Water deficit-induced up-regulation of flavonoid genes is also coordinated with the down-regulation of many stilbene synthases and a consistent decrease in stilbenoid concentration. Water deficit activated both ABA-dependent and ABA-independent signal transduction pathways by modulating the expression of several transcription factors. Gene-gene and gene-metabolite network analyses showed that water deficit-responsive transcription factors such as bZIPs, AP2/ERFs, MYBs, and NACs are implicated in the regulation of stress-responsive metabolites. Enrichment of known and novel cis-regulatory elements in the promoters of several ripening-specific/water deficit-induced modules further affirms the involvement of a transcription factor cross-talk in the berry response to water deficit. Together, our integrated approaches show that water deficit-regulated gene modules are strongly linked to key fruit-quality metabolites and multiple signal transduction pathways may be critical to achieve a balance between the regulation of the stress-response and the berry ripening program. This study constitutes an invaluable resource for future discoveries and comparative studies, in grapes and other fruits, centered on reproductive tissue metabolism under abiotic stress.

  1. Multi-Omics and Integrated Network Analyses Reveal New Insights into the Systems Relationships between Metabolites, Structural Genes, and Transcriptional Regulators in Developing Grape Berries (Vitis vinifera L.) Exposed to Water Deficit.

    Science.gov (United States)

    Savoi, Stefania; Wong, Darren C J; Degu, Asfaw; Herrera, Jose C; Bucchetti, Barbara; Peterlunger, Enrico; Fait, Aaron; Mattivi, Fulvio; Castellarin, Simone D

    2017-01-01

    Grapes are one of the major fruit crops and they are cultivated in many dry environments. This study comprehensively characterizes the metabolic response of grape berries exposed to water deficit at different developmental stages. Increases of proline, branched-chain amino acids, phenylpropanoids, anthocyanins, and free volatile organic compounds have been previously observed in grape berries exposed to water deficit. Integrating RNA-sequencing analysis of the transcriptome with large-scale analysis of central and specialized metabolites, we reveal that these increases occur via a coordinated regulation of key structural pathway genes. Water deficit-induced up-regulation of flavonoid genes is also coordinated with the down-regulation of many stilbene synthases and a consistent decrease in stilbenoid concentration. Water deficit activated both ABA-dependent and ABA-independent signal transduction pathways by modulating the expression of several transcription factors. Gene-gene and gene-metabolite network analyses showed that water deficit-responsive transcription factors such as bZIPs, AP2/ERFs, MYBs, and NACs are implicated in the regulation of stress-responsive metabolites. Enrichment of known and novel cis -regulatory elements in the promoters of several ripening-specific/water deficit-induced modules further affirms the involvement of a transcription factor cross-talk in the berry response to water deficit. Together, our integrated approaches show that water deficit-regulated gene modules are strongly linked to key fruit-quality metabolites and multiple signal transduction pathways may be critical to achieve a balance between the regulation of the stress-response and the berry ripening program. This study constitutes an invaluable resource for future discoveries and comparative studies, in grapes and other fruits, centered on reproductive tissue metabolism under abiotic stress.

  2. The crystal structures of apo and cAMP-bound GlxR from Corynebacterium glutamicum reveal structural and dynamic changes upon cAMP binding in CRP/FNR family transcription factors.

    Directory of Open Access Journals (Sweden)

    Philip D Townsend

    Full Text Available The cyclic AMP-dependent transcriptional regulator GlxR from Corynebacterium glutamicum is a member of the super-family of CRP/FNR (cyclic AMP receptor protein/fumarate and nitrate reduction regulator transcriptional regulators that play central roles in bacterial metabolic regulatory networks. In C. glutamicum, which is widely used for the industrial production of amino acids and serves as a non-pathogenic model organism for members of the Corynebacteriales including Mycobacterium tuberculosis, the GlxR homodimer controls the transcription of a large number of genes involved in carbon metabolism. GlxR therefore represents a key target for understanding the regulation and coordination of C. glutamicum metabolism. Here we investigate cylic AMP and DNA binding of GlxR from C. glutamicum and describe the crystal structures of apo GlxR determined at a resolution of 2.5 Å, and two crystal forms of holo GlxR at resolutions of 2.38 and 1.82 Å, respectively. The detailed structural analysis and comparison of GlxR with CRP reveals that the protein undergoes a distinctive conformational change upon cyclic AMP binding leading to a dimer structure more compatible to DNA-binding. As the two binding sites in the GlxR homodimer are structurally identical dynamic changes upon binding of the first ligand are responsible for the allosteric behavior. The results presented here show how dynamic and structural changes in GlxR lead to optimization of orientation and distance of its two DNA-binding helices for optimal DNA recognition.

  3. Transcript levels of several epigenome regulatory genes in bovine somatic donor cells are not correlated with their cloning efficiency.

    Science.gov (United States)

    Zhou, Wenli; Sadeghieh, Sanaz; Abruzzese, Ronald; Uppada, Subhadra; Meredith, Justin; Ohlrichs, Charletta; Broek, Diane; Polejaeva, Irina

    2009-09-01

    Among many factors that potentially affect somatic cell nuclear transfer (SCNT) embryo development is the donor cell itself. Cloning potentials of somatic donor cells vary greatly, possibly because the cells have different capacities to be reprogrammed by ooplasma. It is therefore intriguing to identify factors that regulate the reprogrammability of somatic donor cells. Gene expression analysis is a widely used tool to investigate underlying mechanisms of various phenotypes. In this study, we conducted a retrospective analysis investigating whether donor cell lines with distinct cloning efficiencies express different levels of genes involved in epigenetic reprogramming including histone deacetylase-1 (HDAC1), -2 (HDAC2); DNA methyltransferase-1 (DNMT1), -3a (DNMT3a),-3b (DNMT3b), and the bovine homolog of yeast sucrose nonfermenting-2 (SNF2L), a SWI/SNF family of ATPases. Cell samples from 12 bovine donor cell lines were collected at the time of nuclear transfer experiments and expression levels of the genes were measured using quantitative polymerase chain reaction (PCR). Our results show that there are no significant differences in expression levels of these genes between donor cell lines of high and low cloning efficiency defined as live calving rates, although inverse correlations are observed between in vitro embryo developmental rates and expression levels of HDAC2 and SNF2L. We also show that selection of stable reference genes is important for relative quantification, and different batches of cells can have different gene expression patterns. In summary, we demonstrate that expression levels of these epigenome regulatory genes in bovine donor cells are not correlated with cloning potential. The experimental design and data analysis method reported here can be applied to study any genes expressed in donor cells.

  4. Transcriptional program of ciliated epithelial cells reveals new cilium and centrosome components and links to human disease.

    Directory of Open Access Journals (Sweden)

    Ramona A Hoh

    Full Text Available Defects in the centrosome and cilium are associated with a set of human diseases having diverse phenotypes. To further characterize the components that define the function of these organelles we determined the transcriptional profile of multiciliated tracheal epithelial cells. Cultures of mouse tracheal epithelial cells undergoing differentiation in vitro were derived from mice expressing GFP from the ciliated-cell specific FOXJ1 promoter (FOXJ1:GFP. The transcriptional profile of ciliating GFP+ cells from these cultures was defined at an early and a late time point during differentiation and was refined by subtraction of the profile of the non-ciliated GFP- cells. We identified 649 genes upregulated early, when most cells were forming basal bodies, and 73 genes genes upregulated late, when most cells were fully ciliated. Most, but not all, of known centrosome proteins are transcriptionally upregulated early, particularly Plk4, a master regulator of centriole formation. We found that three genes associated with human disease states, Mdm1, Mlf1, and Dyx1c1, are upregulated during ciliogenesis and localize to centrioles and cilia. This transcriptome for mammalian multiciliated epithelial cells identifies new candidate centrosome and cilia proteins, highlights similarities between components of motile and primary cilia, and identifies new links between cilia proteins and human disease.

  5. Cloning and characterization of the mouse Mcoln1 gene reveals an alternatively spliced transcript not seen in humans

    Directory of Open Access Journals (Sweden)

    Stahl Stefanie

    2002-02-01

    Full Text Available Abstract Background Mucolipidosis type IV (MLIV is an autosomal recessive lysosomal storage disorder characterized by severe neurologic and ophthalmologic abnormalities. Recently the MLIV gene, MCOLN1, has been identified as a new member of the transient receptor potential (TRP cation channel superfamily. Here we report the cloning and characterization of the mouse homologue, Mcoln1, and report a novel splice variant that is not seen in humans. Results The human and mouse genes display a high degree of synteny. Mcoln1 shows 91% amino acid and 86% nucleotide identity to MCOLN1. Also, Mcoln1 maps to chromosome 8 and contains an open reading frame of 580 amino acids, with a transcript length of approximately 2 kb encoded by 14 exons, similar to its human counterpart. The transcript that results from murine specific alternative splicing encodes a 611 amino acid protein that differs at the c-terminus. Conclusions Mcoln1 is highly similar to MCOLN1, especially in the transmembrane domains and ion pore region. Also, the late endosomal/lysosomal targeting signal is conserved, supporting the hypothesis that the protein is localized to these vesicle membranes. To date, there are very few reports describing species-specific splice variants. While identification of Mcoln1 is crucial to the development of mouse models for MLIV, the fact that there are two transcripts in mice suggests an additional or alternate function of the gene that may complicate phenotypic assessment.

  6. Gene Expression Profiling Reveals a Massive, Aneuploidy-Dependent Transcriptional Deregulation and Distinct Differences between Lymph Node–Negative and Lymph Node–Positive Colon Carcinomas

    Science.gov (United States)

    Grade, Marian; Hörmann, Patrick; Becker, Sandra; Hummon, Amanda B.; Wangsa, Danny; Varma, Sudhir; Simon, Richard; Liersch, Torsten; Becker, Heinz; Difilippantonio, Michael J.; Ghadimi, B. Michael; Ried, Thomas

    2016-01-01

    To characterize patterns of global transcriptional deregulation in primary colon carcinomas, we did gene expression profiling of 73 tumors [Unio Internationale Contra Cancrum stage II (n = 33) and stage III (n = 40)] using oligonucleotide microarrays. For 30 of the tumors, expression profiles were compared with those from matched normal mucosa samples. We identified a set of 1,950 genes with highly significant deregulation between tumors and mucosa samples (P 5-fold average expression difference between normal colon mucosa and carcinomas, including up-regulation of MYC and of HMGA1, a putative oncogene. Furthermore, we identified 68 genes that were significantly differentially expressed between lymph node–negative and lymph node–positive tumors (P deregulated genes were validated using quantitative real-time reverse transcription-PCR in >40 tumor and normal mucosa samples with good concordance between the techniques. Finally, we established a relationship between specific genomic imbalances, which were mapped for 32 of the analyzed colon tumors by comparative genomic hybridization, and alterations of global transcriptional activity. Previously, we had conducted a similar analysis of primary rectal carcinomas. The systematic comparison of colon and rectal carcinomas revealed a significant overlap of genomic imbalances and transcriptional deregulation, including activation of the Wnt/β-catenin signaling cascade, suggesting similar pathogenic pathways. PMID:17210682

  7. Regulatory motifs for CREB-binding protein and Nfe2l2 transcription factors in the upstream enhancer of the mitochondrial uncoupling protein 1 gene.

    Science.gov (United States)

    Rim, Jong S; Kozak, Leslie P

    2002-09-13

    Thermogenesis against cold exposure in mammals occurs in brown adipose tissue (BAT) through mitochondrial uncoupling protein (UCP1). Expression of the Ucp1 gene is unique in brown adipocytes and is regulated tightly. The 5'-flanking region of the mouse Ucp1 gene contains cis-acting elements including PPRE, TRE, and four half-site cAMP-responsive elements (CRE) with BAT-specific enhancer elements. In the course of analyzing how these half-site CREs are involved in Ucp1 expression, we found that a DNA regulatory element for NF-E2 overlaps CRE2. Electrophoretic mobility shift assay and competition assays with the CRE2 element indicates that nuclear proteins from BAT, inguinal fat, and retroperitoneal fat tissue interact with the CRE2 motif (CGTCA) in a specific manner. A supershift assay using an antibody against the CRE-binding protein (CREB) shows specific affinity to the complex from CRE2 and nuclear extract of BAT. Additionally, Western blot analysis for phospho-CREB/ATF1 shows an increase in phosphorylation of CREB/ATF1 in HIB-1B cells after norepinephrine treatment. Transient transfection assay using luciferase reporter constructs also indicates that the two half-site CREs are involved in transcriptional regulation of Ucp1 in response to norepinephrine and cAMP. We also show that a second DNA regulatory element for NF-E2 is located upstream of the CRE2 region. This element, which is found in a similar location in the 5'-flanking region of the human and rodent Ucp1 genes, shows specific binding to rat and human NF-E2 by electrophoretic mobility shift assay with nuclear extracts from brown fat. Co-transfections with an Nfe2l2 expression vector and a luciferase reporter construct of the Ucp1 enhancer region provide additional evidence that Nfe2l2 is involved in the regulation of Ucp1 by cAMP-mediated signaling.

  8. A new in vivo model of pantothenate kinase-associated neurodegeneration reveals a surprising role for transcriptional regulation in pathogenesis.

    Directory of Open Access Journals (Sweden)

    Varun ePandey

    2013-09-01

    Full Text Available Pantothenate Kinase-Associated Neurodegeneration (PKAN is a neurodegenerative disorder with a poorly understood molecular mechanism. It is caused by mutations in Pantothenate Kinase, the first enzyme in the Coenzyme A (CoA biosynthetic pathway. Here, we developed a Drosophila model of PKAN (tim-fbl flies that allows us to continuously monitor the modeled disease in the brain. In tim-fbl flies, downregulation of fumble, the Drosophila PanK homologue in the cells containing a circadian clock results in characteristic features of PKAN such as developmental lethality, hypersensitivity to oxidative stress, and diminished life span. Despite quasi-normal circadian transcriptional rhythms, tim-fbl flies display brain-specific aberrant circadian locomotor rhythms, and a unique transcriptional signature. Comparison with expression data from flies exposed to paraquat demonstrates that, as previously suggested, pathways others than oxidative stress are affected by PANK downregulation. Surprisingly we found a significant decrease in the expression of key components of the photoreceptor recycling pathways, which could lead to retinal degeneration, a hallmark of PKAN. Importantly, these defects are not accompanied by changes in structural components in eye genes suggesting that changes in gene expression in the eye precede and may cause the retinal degeneration. Indeed tim-fbl flies have diminished response to light transitions, and their altered day/night patterns of activity demonstrates defects in light perception. This suggest that retinal lesions are not solely due to oxidative stress and demonstrates a role for the transcriptional response to CoA deficiency underlying the defects observed in dPanK deficient flies. Moreover, in the present study we developed a new fly model that can be applied to other diseases and that allows the assessment of neurodegeneration in the brains of living flies.

  9. High-throughput screening reveals alsterpaullone, 2-cyanoethyl as a potent p27Kip1 transcriptional inhibitor.

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    Brandon J Walters

    Full Text Available p27Kip1 is a cell cycle inhibitor that prevents cyclin dependent kinase (CDK/cyclin complexes from phosphorylating their targets. p27Kip1 is a known tumor suppressor, as the germline loss of p27Kip1 results in sporadic pituitary formation in aged rodents, and its presence in human cancers is indicative of a poor prognosis. In addition to its role in cancer, loss of p27Kip1 results in regenerative phenotypes in some tissues and maintenance of stem cell pluripotency, suggesting that p27Kip1 inhibitors could be beneficial for tissue regeneration. Because p27Kip1 is an intrinsically disordered protein, identifying direct inhibitors of the p27Kip1 protein is difficult. Therefore, we pursued a high-throughput screening strategy to identify novel p27Kip1 transcriptional inhibitors. We utilized a luciferase reporter plasmid driven by the p27Kip1 promoter to transiently transfect HeLa cells and used cyclohexamide as a positive control for non-specific inhibition. We screened a "bioactive" library consisting of 8,904 (4,359 unique compounds, of which 830 are Food and Drug Administration (FDA approved. From this screen, we successfully identified 111 primary hits with inhibitory effect against the promoter of p27Kip1. These hits were further refined using a battery of secondary screens. Here we report four novel p27Kip1 transcriptional inhibitors, and further demonstrate that our most potent hit compound (IC50 = 200 nM Alsterpaullone 2-cyanoethyl, inhibits p27Kip1 transcription by preventing FoxO3a from binding to the p27Kip1 promoter. This screen represents one of the first attempts to identify inhibitors of p27Kip1 and may prove useful for future tissue regeneration studies.

  10. Transcriptome analysis of paired primary colorectal carcinoma and liver metastases reveals fusion transcripts and similar gene expression profiles in primary carcinoma and liver metastases

    International Nuclear Information System (INIS)

    Lee, Ja-Rang; Kwon, Chae Hwa; Choi, Yuri; Park, Hye Ji; Kim, Hyun Sung; Jo, Hong-Jae; Oh, Nahmgun; Park, Do Youn

    2016-01-01

    Despite the clinical significance of liver metastases, the difference between molecular and cellular changes in primary colorectal cancers (CRC) and matched liver metastases is poorly understood. In order to compare gene expression patterns and identify fusion genes in these two types of tumors, we performed high-throughput transcriptome sequencing of five sets of quadruple-matched tissues (primary CRC, liver metastases, normal colon, and liver). The gene expression patterns in normal colon and liver were successfully distinguished from those in CRCs; however, RNA sequencing revealed that the gene expression between primary CRCs and their matched liver metastases is highly similar. We identified 1895 genes that were differentially expressed in the primary carcinoma and liver metastases, than that in the normal colon tissues. A major proportion of the transcripts, identified by gene expression profiling as significantly enriched in the primary carcinoma and metastases, belonged to gene ontology categories involved in the cell cycle, mitosis, and cell division. Furthermore, we identified gene fusion events in primary carcinoma and metastases, and the fusion transcripts were experimentally confirmed. Among these, a chimeric transcript resulting from the fusion of RNF43 and SUPT4H1 was found to occur frequently in primary colorectal carcinoma. In addition, knockdown of the expression of this RNF43-SUPT4H1 chimeric transcript was found to have a growth-inhibitory effect in colorectal cancer cells. The present study reports a high concordance of gene expression in the primary carcinoma and liver metastases, and reveals potential new targets, such as fusion genes, against primary and metastatic colorectal carcinoma. The online version of this article (doi:10.1186/s12885-016-2596-3) contains supplementary material, which is available to authorized users

  11. A Novel Sucrose-Regulatory MADS-Box Transcription Factor GmNMHC5 Promotes Root Development and Nodulation in Soybean (Glycine max [L.] Merr.

    Directory of Open Access Journals (Sweden)

    Wei Liu

    2015-08-01

    Full Text Available The MADS-box protein family includes many transcription factors that have a conserved DNA-binding MADS-box domain. The proteins in this family were originally recognized to play prominent roles in floral development. Recent findings, especially with regard to the regulatory roles of the AGL17 subfamily in root development, have greatly broadened their known functions. In this study, a gene from soybean (Glycine max [L.] Merr., GmNMHC5, was cloned from the Zigongdongdou cultivar and identified as a member of the AGL17 subfamily. Real-time fluorescence quantitative PCR analysis showed that GmNMHC5 was expressed at much higher levels in roots and nodules than in other organs. The activation of expression was first examined in leaves and roots, followed by shoot apexes. GmNMHC5 expression levels rose sharply when the plants were treated under short-day conditions (SD and started to pod, whereas low levels were maintained in non-podding plants under long-day conditions (LD. Furthermore, overexpression of GmNMHC5 in transgenic soybean significantly promoted lateral root development and nodule building. Moreover, GmNMHC5 is upregulated by exogenous sucrose. These results indicate that GmNMHC5 can sense the sucrose signal and plays significant roles in lateral root development and nodule building.

  12. Senescence-associated barley NAC (NAM, ATAF1,2, CUC) transcription factor interacts with radical-induced cell death 1 through a disordered regulatory domain

    DEFF Research Database (Denmark)

    Kjaersgaard, Trine; Jensen, Michael K; Christiansen, Michael W

    2011-01-01

    as a transcriptional activator suggesting that an involvement of HvNAC013 and HvNAC005 in senescence will be different. HvNAC013 interacted with barley radical-induced cell death 1 (RCD1) via the very C-terminal part of its TRD, outside of the region containing the LP motif. No significant secondary structure...... (NAM, ATAF1,2, CUC) TF family are up-regulated during senescence in barley (Hordeum vulgare). Both HvNAC005 and HvNAC013 bound the conserved NAC DNA target sequence. Computational and biophysical analyses showed that both proteins are intrinsically disordered in their large C-terminal domains, which...... was induced in the HvNAC013 TRD upon interaction with RCD1. RCD1 also interacted with regions dominated by intrinsic disorder in TFs of the MYB and basic helix-loop-helix families. We propose that RCD1 is a regulatory protein capable of interacting with many different TFs by exploiting their intrinsic...

  13. The RNA-mediated, asymmetric ring regulatory mechanism of the transcription termination Rho helicase decrypted by time-resolved nucleotide analog interference probing (trNAIP).

    Science.gov (United States)

    Soares, Emilie; Schwartz, Annie; Nollmann, Marcello; Margeat, Emmanuel; Boudvillain, Marc

    2014-08-01

    Rho is a ring-shaped, ATP-dependent RNA helicase/translocase that dissociates transcriptional complexes in bacteria. How RNA recognition is coupled to ATP hydrolysis and translocation in Rho is unclear. Here, we develop and use a new combinatorial approach, called time-resolved Nucleotide Analog Interference Probing (trNAIP), to unmask RNA molecular determinants of catalytic Rho function. We identify a regulatory step in the translocation cycle involving recruitment of the 2'-hydroxyl group of the incoming 3'-RNA nucleotide by a Rho subunit. We propose that this step arises from the intrinsic weakness of one of the subunit interfaces caused by asymmetric, split-ring arrangement of primary RNA tethers around the Rho hexamer. Translocation is at highest stake every seventh nucleotide when the weak interface engages the incoming 3'-RNA nucleotide or breaks, depending on RNA threading constraints in the Rho pore. This substrate-governed, 'test to run' iterative mechanism offers a new perspective on how a ring-translocase may function or be regulated. It also illustrates the interest and versatility of the new trNAIP methodology to unveil the molecular mechanisms of complex RNA-based systems. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  14. Comparing genomic expression patterns across plant species reveals highly diverged transcriptional dynamics in response to salt stress

    Directory of Open Access Journals (Sweden)

    Close Timothy J

    2009-08-01

    Full Text Available Abstract Background Rice and barley are both members of Poaceae (grass family but have a marked difference in salt tolerance. The molecular mechanism underlying this difference was previously unexplored. This study employs a comparative genomics approach to identify analogous and contrasting gene expression patterns between rice and barley. Results A hierarchical clustering approach identified several interesting expression trajectories among rice and barley genotypes. There were no major conserved expression patterns between the two species in response to salt stress. A wheat salt-stress dataset was queried for comparison with rice and barley. Roughly one-third of the salt-stress responses of barley were conserved with wheat while overlap between wheat and rice was minimal. These results demonstrate that, at transcriptome level, rice is strikingly different compared to the more closely related barley and wheat. This apparent lack of analogous transcriptional programs in response to salt stress is further highlighted through close examination of genes associated with root growth and development. Conclusion The analysis provides support for the hypothesis that conservation of transcriptional signatures in response to environmental cues depends on the genetic similarity among the genotypes within a species, and on the phylogenetic distance between the species.

  15. De Novo Transcriptome Assembly (NGS) of Curcuma longa L. Rhizome Reveals Novel Transcripts Related to Anticancer and Antimalarial Terpenoids

    Science.gov (United States)

    Jayakumar, Vasanthan; Damodaran, Anand C.; Rao, Sudha Narayana; Katta, Mohan A. V. S. K.; Gopinathan, Sreeja; Sarma, Santosh Prasad; Senthilkumar, Vanitha; Niranjan, Vidya; Gopinath, Ashok; Mugasimangalam, Raja C.

    2013-01-01

    Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa. PMID:23468859

  16. De Novo transcriptome assembly (NGS of Curcuma longa L. rhizome reveals novel transcripts related to anticancer and antimalarial terpenoids.

    Directory of Open Access Journals (Sweden)

    Ramasamy S Annadurai

    Full Text Available Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa.

  17. De Novo transcriptome assembly (NGS) of Curcuma longa L. rhizome reveals novel transcripts related to anticancer and antimalarial terpenoids.

    Science.gov (United States)

    Annadurai, Ramasamy S; Neethiraj, Ramprasad; Jayakumar, Vasanthan; Damodaran, Anand C; Rao, Sudha Narayana; Katta, Mohan A V S K; Gopinathan, Sreeja; Sarma, Santosh Prasad; Senthilkumar, Vanitha; Niranjan, Vidya; Gopinath, Ashok; Mugasimangalam, Raja C

    2013-01-01

    Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa.

  18. Transcriptional profiling of primary endometrial epithelial cells following acute HIV-1 exposure reveals gene signatures related to innate immunity.

    Science.gov (United States)

    Zahoor, Muhammad Atif; Woods, Matthew William; Dizzell, Sara; Nazli, Aisha; Mueller, Kristen M; Nguyen, Philip V; Verschoor, Chris P; Kaushic, Charu

    2018-04-01

    Genital epithelial cells (GECs) line the mucosal surface of the female genital tract (FGT) and are the first cells that interface with both commensal microbiota and sexually transmitted pathogens. Despite the protective barrier formed by GECs, the FGT is a major site of HIV-1 infection. This highlights the importance of studying the interaction of HIV-1 and GECs. Using microarray analysis, we characterized the transcriptional profile of primary endometrial GECs grown in the presence or absence of physiological levels of E2 (10 -9  mol/L) or P4 (10 -7  mol/L) following acute exposure to HIV-1 for 6 hours. Acute exposure of primary endometrial GECs to HIV-1 resulted in the expression of genes related to inflammation, plasminogen activation, adhesion and diapedesis and interferon response. Interestingly, exposure to HIV-1 in the presence of E2 and P4 resulted in differential transcriptional profiles, suggesting that the response of primary endometrial GECs to HIV-1 exposure is modulated by female sex hormones. The gene expression signature of endometrial GECs indicates that the response of these cells may be key to determining host susceptibility to HIV-1 and that sex hormones modulate these interactions. This study allows us to explore possible mechanisms that explain the hormone-mediated fluctuation of HIV-1 susceptibility in women. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  19. A Systematic Analysis of Factors Localized to Damaged Chromatin Reveals PARP-Dependent Recruitment of Transcription Factors.

    Science.gov (United States)

    Izhar, Lior; Adamson, Britt; Ciccia, Alberto; Lewis, Jedd; Pontano-Vaites, Laura; Leng, Yumei; Liang, Anthony C; Westbrook, Thomas F; Harper, J Wade; Elledge, Stephen J

    2015-06-09

    Localization to sites of DNA damage is a hallmark of DNA damage response (DDR) proteins. To identify DDR factors, we screened epitope-tagged proteins for localization to sites of chromatin damaged by UV laser microirradiation and found >120 proteins that localize to damaged chromatin. These include the BAF tumor suppressor complex and the amyotrophic lateral sclerosis (ALS) candidate protein TAF15. TAF15 contains multiple domains that bind damaged chromatin in a poly-(ADP-ribose) polymerase (PARP)-dependent manner, suggesting a possible role as glue that tethers multiple PAR chains together. Many positives were transcription factors; > 70% of randomly tested transcription factors localized to sites of DNA damage, and of these, ∼90% were PARP dependent for localization. Mutational analyses showed that localization to damaged chromatin is DNA-binding-domain dependent. By examining Hoechst staining patterns at damage sites, we see evidence of chromatin decompaction that is PARP dependent. We propose that PARP-regulated chromatin remodeling at sites of damage allows transient accessibility of DNA-binding proteins. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  20. A Systematic Analysis of Factors Localized to Damaged Chromatin Reveals PARP-Dependent Recruitment of Transcription Factors

    Directory of Open Access Journals (Sweden)

    Lior Izhar

    2015-06-01

    Full Text Available Localization to sites of DNA damage is a hallmark of DNA damage response (DDR proteins. To identify DDR factors, we screened epitope-tagged proteins for localization to sites of chromatin damaged by UV laser microirradiation and found >120 proteins that localize to damaged chromatin. These include the BAF tumor suppressor complex and the amyotrophic lateral sclerosis (ALS candidate protein TAF15. TAF15 contains multiple domains that bind damaged chromatin in a poly-(ADP-ribose polymerase (PARP-dependent manner, suggesting a possible role as glue that tethers multiple PAR chains together. Many positives were transcription factors; > 70% of randomly tested transcription factors localized to sites of DNA damage, and of these, ∼90% were PARP dependent for localization. Mutational analyses showed that localization to damaged chromatin is DNA-binding-domain dependent. By examining Hoechst staining patterns at damage sites, we see evidence of chromatin decompaction that is PARP dependent. We propose that PARP-regulated chromatin remodeling at sites of damage allows transient accessibility of DNA-binding proteins.

  1. Analysis of temporal transcription expression profiles reveal links between protein function and developmental stages of Drosophila melanogaster.

    Science.gov (United States)

    Wan, Cen; Lees, Jonathan G; Minneci, Federico; Orengo, Christine A; Jones, David T

    2017-10-01

    Accurate gene or protein function prediction is a key challenge in the post-genome era. Most current methods perform well on molecular function prediction, but struggle to provide useful annotations relating to biological process functions due to the limited power of sequence-based features in that functional domain. In this work, we systematically evaluate the predictive power of temporal transcription expression profiles for protein function prediction in Drosophila melanogaster. Our results show significantly better performance on predicting protein function when transcription expression profile-based features are integrated with sequence-derived features, compared with the sequence-derived features alone. We also observe that the combination of expression-based and sequence-based features leads to further improvement of accuracy on predicting all three domains of gene function. Based on the optimal feature combinations, we then propose a novel multi-classifier-based function prediction method for Drosophila melanogaster proteins, FFPred-fly+. Interpreting our machine learning models also allows us to identify some of the underlying links between biological processes and developmental stages of Drosophila melanogaster.

  2. Analysis of temporal transcription expression profiles reveal links between protein function and developmental stages of Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Cen Wan

    2017-10-01

    Full Text Available Accurate gene or protein function prediction is a key challenge in the post-genome era. Most current methods perform well on molecular function prediction, but struggle to provide useful annotations relating to biological process functions due to the limited power of sequence-based features in that functional domain. In this work, we systematically evaluate the predictive power of temporal transcription expression profiles for protein function prediction in Drosophila melanogaster. Our results show significantly better performance on predicting protein function when transcription expression profile-based features are integrated with sequence-derived features, compared with the sequence-derived features alone. We also observe that the combination of expression-based and sequence-based features leads to further improvement of accuracy on predicting all three domains of gene function. Based on the optimal feature combinations, we then propose a novel multi-classifier-based function prediction method for Drosophila melanogaster proteins, FFPred-fly+. Interpreting our machine learning models also allows us to identify some of the underlying links between biological processes and developmental stages of Drosophila melanogaster.

  3. Multiple independent analyses reveal only transcription factors as an enriched functional class associated with microRNAs

    DEFF Research Database (Denmark)

    Croft, Laurence J; Szklarczyk, Damian; Jensen, Lars J

    2012-01-01

    , self-regulatory connectivity between miRNAs and TFs. Here we investigate the connectivity from TFs and miRNAs to other genes and each other using text-mining, TF promoter binding site and 6 different miRNA binding site prediction methods. RESULTS: In the first approach text-mining of PubMed abstracts......RNA targets, appear not to select for real miRNA targets to any degree. Our manually validated text-mining results also suggests that miRNAs may be activated by more TFs than other classes of genes, as 7% of miRNA:TF co-occurrences in the literature were TFs activating miRNAs. This was confirmed when thirdly...

  4. Metabolite Profiling and Transcript Analysis Reveal Specificities in the Response of a Berry Derived Cell Culture to Abiotic Stresses

    Directory of Open Access Journals (Sweden)

    Biruk eAyenew

    2015-09-01

    Full Text Available As climate changes, there is a need to understand the expected effects on viticulture. In nature, stresses exist in a combined manner, hampering the elucidation of the effect of individual cues on grape berry metabolism. Cell suspension culture originated from pea-size Gamy Red grape berry was used to harness metabolic response to high light (2500 µmol m-2s-1, high temperature (40 0C and their combination in comparison to 25 0C and 100 µmol m-2s-1 under controlled condition. When LC-MS and GC-MS based metabolite profiling was implemented and integrated with targeted RT-qPCR transcript analysis specific responses were observed to the different cues. High light enhanced polyphenol metabolism while high temperature and its combination with high light induced amino acid and organic acid metabolism with additional effect on polyphenols. The trend of increment in TCA cycle genes like ATCs, ACo1 and IDH in the combined treatment might support the observed increment in organic acids, GABA shunt, and their derivatives. The apparent phenylalanine reduction with polyphenol increment under high light suggests enhanced fueling of the precursor towards the downstream phenylpropanoid pathway. In the polyphenol metabolism, a differential pattern of expression of flavonoid 3’,5’ hydroxylase and flavonoid 3’ hydroxylase was observed under high light and combined cues which were accompanied by characteristic metabolite profiles. High temperature decreased glycosylated cyanidin and peonidin forms while the combined cues increased acetylated and coumarylated peonidin forms. Transcription factors regulating anthocyanin metabolism and their methylation, MYB, OMT, UFGT and DFR, were expressed differentially among the treatments, overall in agreement with the metabolite profiles. Taken together these data provide insights into the coordination of central and secondary metabolism in relation to multiple abiotic stresses.

  5. Transcript Quantification by RNA-Seq Reveals Differentially Expressed Genes in the Red and Yellow Fruits of Fragaria vesca.

    Directory of Open Access Journals (Sweden)

    Yuchao Zhang

    Full Text Available Fragaria vesca (2n = 2x = 14, the woodland strawberry, is a perennial herbaceous plant with a small sequenced genome (240 Mb. It is commonly used as a genetic model plant for the Fragaria genus and the Rosaceae family. Fruit skin color is one of the most important traits for both the commercial and esthetic value of strawberry. Anthocyanins are the most prominent pigments in strawberry that bring red, pink, white, and yellow hues to the fruits in which they accumulate. In this study, we conducted a de novo assembly of the fruit transcriptome of woodland strawberry and compared the gene expression profiles with yellow (Yellow Wonder, YW and red (Ruegen, RG fruits. De novo assembly yielded 75,426 unigenes, 21.3% of which were longer than 1,000 bp. Among the high-quality unique sequences, 45,387 (60.2% had at least one significant match to an existing gene model. A total of 595 genes, representing 0.79% of total unigenes, were differentially expressed in YW and RG. Among them, 224 genes were up-regulated and 371 genes were down-regulated in the fruit of YW. Particularly, some flavonoid biosynthetic pathway genes, including C4H, CHS, CHI, F3H, DFR and ANS, as well as some transcription factors (TFs, including MYB (putative MYB86 and MYB39, WDR and MADS, were down-regulated in YW fruit, concurrent with a reduction in anthocyanin accumulation in the yellow pigment phenotype, whereas a putative transcription repressor MYB1R was up-regulated in YW fruit. The altered expression levels of the genes encoding flavonoid biosynthetic enzymes and TFs were confirmed by quantitative RT-PCR. Our study provides important insights into the molecular mechanisms underlying the yellow pigment phenotype in F. vesca.

  6. Laser microdissection reveals that transcripts for five plant and one fungal phosphate transporter genes are contemporaneously present in arbusculated cells.

    Science.gov (United States)

    Balestrini, Raffaella; Gómez-Ariza, Jorge; Lanfranco, Luisa; Bonfante, Paola

    2007-09-01

    The establishment of a symbiotic interaction between plant roots and arbuscular mycorrhizal (AM) fungi requires both partners to undergo significant morphological and physiological modifications which eventually lead to reciprocal beneficial effects. Extensive changes in gene expression profiles recently have been described in transcriptomic studies that have analyzed the whole mycorrhizal root. However, because root colonization by AM fungi involves different cell types, a cell-specific gene expression pattern is likely to occur. We have applied the laser microdissection (LMD) technology to investigate expression profiles of both plant and fungal genes in Lycopersicon esculentum roots colonized by Glomus mosseae. A protocol to harvest arbuscule-containing cells from paraffin sections of mycorrhizal roots has been developed using a Leica AS LMD system. RNA of satisfactory quantity and quality has been extracted for molecular analysis. Transcripts for plant phosphate transporters (LePTs), selected as molecular markers for a functional symbiosis, have been detected by reverse-transcriptase polymerase chain reaction assays and associated to distinct cell types, leading to novel insights into the distribution of LePT mRNAs. In fact, the transcripts of the five phosphate transporters (PTs) have been detected contemporaneously in the same arbusculated cell population, unlike from the neighboring noncolonized cells. In addition, fungal H(+)ATPase (GmHA5) and phosphate transporter (GmosPT) mRNAs were found exclusively in arbusculated cells. The discovery that five plant and one fungal PT genes are consistently expressed inside the arbusculated cells provides a new scenario for plant-fungus nutrient exchanges.

  7. Dual RNA-seq reveals no plastic transcriptional response of the coccidian parasite Eimeria falciformis to host immune defenses.

    Science.gov (United States)

    Ehret, Totta; Spork, Simone; Dieterich, Christoph; Lucius, Richard; Heitlinger, Emanuel

    2017-09-05

    Parasites can either respond to differences in immune defenses that exist between individual hosts plastically or, alternatively, follow a genetically canalized ("hard wired") program of infection. Assuming that large-scale functional plasticity would be discernible in the parasite transcriptome we have performed a dual RNA-seq study of the lifecycle of Eimeria falciformis using infected mice with different immune status as models for coccidian infections. We compared parasite and host transcriptomes (dual transcriptome) between naïve and challenge infected mice, as well as between immune competent and immune deficient ones. Mice with different immune competence show transcriptional differences as well as differences in parasite reproduction (oocyst shedding). Broad gene categories represented by differently abundant host genes indicate enrichments for immune reaction and tissue repair functions. More specifically, TGF-beta, EGF, TNF and IL-1 and IL-6 are examples of functional annotations represented differently depending on host immune status. Much in contrast, parasite transcriptomes were neither different between Coccidia isolated from immune competent and immune deficient mice, nor between those harvested from naïve and challenge infected mice. Instead, parasite transcriptomes have distinct profiles early and late in infection, characterized largely by biosynthesis or motility associated functional gene groups, respectively. Extracellular sporozoite and oocyst stages showed distinct transcriptional profiles and sporozoite transcriptomes were found enriched for species specific genes and likely pathogenicity factors. We propose that the niche and host-specific parasite E. falciformis uses a genetically canalized program of infection. This program is likely fixed in an evolutionary process rather than employing phenotypic plasticity to interact with its host. This in turn might limit the potential of the parasite to adapt to new host species or niches, forcing

  8. Transcriptional profiling of human brain endothelial cells reveals key properties crucial for predictive in vitro blood-brain barrier models.

    Directory of Open Access Journals (Sweden)

    Eduard Urich

    Full Text Available Brain microvascular endothelial cells (BEC constitute the blood-brain barrier (BBB which forms a dynamic interface between the blood and the central nervous system (CNS. This highly specialized interface restricts paracellular diffusion of fluids and solutes including chemicals, toxins and drugs from entering the brain. In this study we compared the transcriptome profiles of the human immortalized brain endothelial cell line hCMEC/D3 and human primary BEC. We identified transcriptional differences in immune response genes which are directly related to the immortalization procedure of the hCMEC/D3 cells. Interestingly, astrocytic co-culturing reduced cell adhesion and migration molecules in both BECs, which possibly could be related to regulation of immune surveillance of the CNS controlled by astrocytic cells within the neurovascular unit. By matching the transcriptome data from these two cell lines with published transcriptional data from freshly isolated mouse BECs, we discovered striking differences that could explain some of the limitations of using cultured BECs to study BBB properties. Key protein classes such as tight junction proteins, transporters and cell surface receptors show differing expression profiles. For example, the claudin-5, occludin and JAM2 expression is dramatically reduced in the two human BEC lines, which likely explains their low transcellular electric resistance and paracellular leakiness. In addition, the human BEC lines express low levels of unique brain endothelial transporters such as Glut1 and Pgp. Cell surface receptors such as LRP1, RAGE and the insulin receptor that are involved in receptor-mediated transport are also expressed at very low levels. Taken together, these data illustrate that BECs lose their unique protein expression pattern outside of their native environment and display a more generic endothelial cell phenotype. A collection of key genes that seems to be highly regulated by the local

  9. Global MYCN transcription factor binding analysis in neuroblastoma reveals association with distinct E-box motifs and regions of DNA hypermethylation.

    LENUS (Irish Health Repository)

    Murphy, Derek M

    2009-01-01

    BACKGROUND: Neuroblastoma, a cancer derived from precursor cells of the sympathetic nervous system, is a major cause of childhood cancer related deaths. The single most important prognostic indicator of poor clinical outcome in this disease is genomic amplification of MYCN, a member of a family of oncogenic transcription factors. METHODOLOGY: We applied MYCN chromatin immunoprecipitation to microarrays (ChIP-chip) using MYCN amplified\\/non-amplified cell lines as well as a conditional knockdown cell line to determine the distribution of MYCN binding sites within all annotated promoter regions. CONCLUSION: Assessment of E-box usage within consistently positive MYCN binding sites revealed a predominance for the CATGTG motif (p<0.0016), with significant enrichment of additional motifs CATTTG, CATCTG, CAACTG in the MYCN amplified state. For cell lines over-expressing MYCN, gene ontology analysis revealed enrichment for the binding of MYCN at promoter regions of numerous molecular functional groups including DNA helicases and mRNA transcriptional regulation. In order to evaluate MYCN binding with respect to other genomic features, we determined the methylation status of all annotated CpG islands and promoter sequences using methylated DNA immunoprecipitation (MeDIP). The integration of MYCN ChIP-chip and MeDIP data revealed a highly significant positive correlation between MYCN binding and DNA hypermethylation. This association was also detected in regions of hemizygous loss, indicating that the observed association occurs on the same homologue. In summary, these findings suggest that MYCN binding occurs more commonly at CATGTG as opposed to the classic CACGTG E-box motif, and that disease associated over expression of MYCN leads to aberrant binding to additional weaker affinity E-box motifs in neuroblastoma. The co-localization of MYCN binding and DNA hypermethylation further supports the dual role of MYCN, namely that of a classical transcription factor affecting the

  10. Battles and hijacks: Noncoding transcription in plants

    KAUST Repository

    Ariel, Federico

    2015-06-01

    Noncoding RNAs have emerged as major components of the eukaryotic transcriptome. Genome-wide analyses revealed the existence of thousands of long noncoding RNAs (lncRNAs) in several plant species. Plant lncRNAs are transcribed by the plant-specific RNA polymerases Pol IV and Pol V, leading to transcriptional gene silencing, as well as by Pol II. They are involved in a wide range of regulatory mechanisms impacting on gene expression, including chromatin remodeling, modulation of alternative splicing, fine-tuning of miRNA activity, and the control of mRNA translation or accumulation. Recently, dual noncoding transcription by alternative RNA polymerases was implicated in epigenetic and chromatin conformation dynamics. This review integrates the current knowledge on the regulatory mechanisms acting through plant noncoding transcription. © 2015 Elsevier Ltd.

  11. Preaxial polydactyly/triphalangeal thumb is associated with changed transcription factor-binding affinity in a family with a novel point mutation in the long-range cis-regulatory element ZRS

    DEFF Research Database (Denmark)

    Farooq, Muhammad; Troelsen, Jesper T; Boyd, Mette

    2010-01-01

    A cis-regulatory sequence also known as zone of polarizing activity (ZPA) regulatory sequence (ZRS) located in intron 5 of LMBR1 is essential for expression of sonic hedgehog (SHH) in the developing posterior limb bud mesenchyme. Even though many point mutations causing preaxial duplication defects...... demonstrated a marked difference between wild-type and the mutant probe, which uniquely bound one or several transcription factors extracted from Caco-2 cells. This finding supports a model in which ectopic anterior SHH expression in the developing limb results from abnormal binding of one or more...

  12. Transcription regulation by the Mediator complex.

    Science.gov (United States)

    Soutourina, Julie

    2018-04-01

    Alterations in the regulation of gene expression are frequently associated with developmental diseases or cancer. Transcription activation is a key phenomenon in the regulation of gene expression. In all eukaryotes, mediator of RNA polymerase II transcription (Mediator), a large complex with modular organization, is generally required for transcription by RNA polymerase II, and it regulates various steps of this process. The main function of Mediator is to transduce signals from the transcription activators bound to enhancer regions to the transcription machinery, which is assembled at promoters as the preinitiation complex (PIC) to control transcription initiation. Recent functional studies of Mediator with the use of structural biology approaches and functional genomics have revealed new insights into Mediator activity and its regulation during transcription initiation, including how Mediator is recruited to transcription regulatory regions and how it interacts and cooperates with PIC components to assist in PIC assembly. Novel roles of Mediator in the control of gene expression have also been revealed by showing its connection to the nuclear pore and linking Mediator to the regulation of gene positioning in the nuclear space. Clear links between Mediator subunits and disease have also encouraged studies to explore targeting of this complex as a potential therapeutic approach in cancer and fungal infections.

  13. R-ChIP Using Inactive RNase H Reveals Dynamic Coupling of R-loops with Transcriptional Pausing at Gene Promoters.

    Science.gov (United States)

    Chen, Liang; Chen, Jia-Yu; Zhang, Xuan; Gu, Ying; Xiao, Rui; Shao, Changwei; Tang, Peng; Qian, Hao; Luo, Daji; Li, Hairi; Zhou, Yu; Zhang, Dong-Er; Fu, Xiang-Dong

    2017-11-16

    R-loop, a three-stranded RNA/DNA structure, has been linked to induced genome instability and regulated gene expression. To enable precision analysis of R-loops in vivo, we develop an RNase-H-based approach; this reveals predominant R-loop formation near gene promoters with strong G/C skew and propensity to form G-quadruplex in non-template DNA, corroborating with all biochemically established properties of R-loops. Transcription perturbation experiments further indicate that R-loop induction correlates to transcriptional pausing. Interestingly, we note that most mapped R-loops are each linked to a nearby free RNA end; by using a ribozyme to co-transcriptionally cleave nascent RNA, we demonstrate that such a free RNA end coupled with a G/C-skewed sequence is necessary and sufficient to induce R-loop. These findings provide a topological solution for RNA invasion into duplex DNA and suggest an order for R-loop initiation and elongation in an opposite direction to that previously proposed. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Combined genome-wide expression profiling and targeted RNA interference in primary mouse macrophages reveals perturbation of transcriptional networks associated with interferon signalling

    Directory of Open Access Journals (Sweden)

    Craigon Marie

    2009-08-01

    Full Text Available Abstract Background Interferons (IFNs are potent antiviral cytokines capable of reprogramming the macrophage phenotype through the induction of interferon-stimulated genes (ISGs. Here we have used targeted RNA interference to suppress the expression of a number of key genes associated with IFN signalling in murine macrophages prior to stimulation with interferon-gamma. Genome-wide changes in transcript abundance caused by siRNA activity were measured using exon-level microarrays in the presence or absence of IFNγ. Results Transfection of murine bone-marrow derived macrophages (BMDMs with a non-targeting (control siRNA and 11 sequence-specific siRNAs was performed using a cationic lipid transfection reagent (Lipofectamine2000 prior to stimulation with IFNγ. Total RNA was harvested from cells and gene expression measured on Affymetrix GeneChip Mouse Exon 1.0 ST Arrays. Network-based analysis of these data revealed six siRNAs to cause a marked shift in the macrophage transcriptome in the presence or absence IFNγ. These six siRNAs targeted the Ifnb1, Irf3, Irf5, Stat1, Stat2 and Nfkb2 transcripts. The perturbation of the transcriptome by the six siRNAs was highly similar in each case and affected the expression of over 600 downstream trans