Nuclear Import of the Parsley bZIP Transcription Factor CPRF2 Is Regulated by Phytochrome Photoreceptors

Science.gov (United States)

Kircher, Stefan; Wellmer, Frank; Nick, Peter; Rügner, Alexander; Schäfer, Eberhard; Harter, Klaus

1999-01-01

In plants, light perception by photoreceptors leads to differential expression of an enormous number of genes. An important step for differential gene expression is the regulation of transcription factor activities. To understand these processes in light signal transduction we analyzed the three well-known members of the common plant regulatory factor (CPRF) family from parsley (Petroselinum crispum). Here, we demonstrate that these CPRFs, which belong to the basic- region leucine-zipper (bZIP) domain-containing transcription factors, are differentially distributed within parsley cells, indicating different regulatory functions within the regulatory networks of the plant cell. In particular, we show by cell fractionation and immunolocalization approaches that CPRF2 is transported from the cytosol into the nucleus upon irradiation due to action of phytochrome photoreceptors. Two NH2-terminal domains responsible for cytoplasmic localization of CPRF2 in the dark were characterized by deletion analysis using a set of CPRF2-green fluorescent protein (GFP) gene fusion constructs transiently expressed in parsley protoplasts. We suggest that light-induced nuclear import of CPRF2 is an essential step in phytochrome signal transduction. PMID:9922448

Arabidopsis IRE1 catalyses unconventional splicing of bZIP60 mRNA to produce the active transcription factor

KAUST Repository

Nagashima, Yukihiro; Mishiba, Kei-ichiro; Suzuki, Eiji; Shimada, Yukihisa; Iwata, Yuji; Koizumi, Nozomu

2011-01-01

-type. Transcriptome analysis revealed that genes whose induction was reduced in ire1a/ire1b largely overlapped those in the bzip60 mutant. We observed that the active form of bZIP60 protein detected in the wild-type was missing in ire1a/ire1b. We further demonstrated

AtMyb7, a subgroup 4 R2R3 Myb, negatively regulates ABA-induced inhibition of seed germination by blocking the expression of the bZIP transcription factor ABI5

KAUST Repository

Kim, Junhyeok; Hyun, Wooyoung; Nguyen, Hoai Nguyen; Jeong, Chanyoung; Xiong, Liming; Hong, Sukwhan; Lee, Hojoung

2014-01-01

Various Myb proteins have been shown to play crucial roles in plants, including primary and secondary metabolism, determination of cell fate and identity, regulation of development and involvement in responses to biotic and abiotic stresses. The 126 R2R3 Myb proteins (with two Myb repeats) have been found in Arabidopsis; however, the functions of most of these proteins remain to be fully elucidated. In the present study, we characterized the function of AtMyb7 using molecular biological and genetic analyses. We used qRT-PCR to determine the levels of stress-response gene transcripts in wild-type and atmyb7 plants. We showed that ArabidopsisAtMyb7 plays a critical role in seed germination. Under abscisic acid (ABA) and high-salt stress conditions, atmyb7 plants showed a lower germination rate than did wild-type plants. Furthermore, AtMyb7 promoter:GUS seeds exhibited different expression patterns in response to variations in the seed imbibition period. AtMyb7 negatively controls the expression of the gene encoding bZIP transcription factor, ABI5, which is a key transcription factor in ABA signalling and serves as a crucial regulator of germination inhibition in Arabidopsis. © 2014 John Wiley & Sons Ltd.

AtMyb7, a subgroup 4 R2R3 Myb, negatively regulates ABA-induced inhibition of seed germination by blocking the expression of the bZIP transcription factor ABI5

KAUST Repository

Kim, Junhyeok

2014-08-27

Various Myb proteins have been shown to play crucial roles in plants, including primary and secondary metabolism, determination of cell fate and identity, regulation of development and involvement in responses to biotic and abiotic stresses. The 126 R2R3 Myb proteins (with two Myb repeats) have been found in Arabidopsis; however, the functions of most of these proteins remain to be fully elucidated. In the present study, we characterized the function of AtMyb7 using molecular biological and genetic analyses. We used qRT-PCR to determine the levels of stress-response gene transcripts in wild-type and atmyb7 plants. We showed that ArabidopsisAtMyb7 plays a critical role in seed germination. Under abscisic acid (ABA) and high-salt stress conditions, atmyb7 plants showed a lower germination rate than did wild-type plants. Furthermore, AtMyb7 promoter:GUS seeds exhibited different expression patterns in response to variations in the seed imbibition period. AtMyb7 negatively controls the expression of the gene encoding bZIP transcription factor, ABI5, which is a key transcription factor in ABA signalling and serves as a crucial regulator of germination inhibition in Arabidopsis. © 2014 John Wiley & Sons Ltd.

bZIP transcription factor SmJLB1 regulates autophagy-related genes Smatg8 and Smatg4 and is required for fruiting-body development and vegetative growth in Sordaria macrospora.

Science.gov (United States)

Voigt, Oliver; Herzog, Britta; Jakobshagen, Antonia; Pöggeler, Stefanie

2013-12-01

Autophagy is a precisely controlled degradation process in eukaryotic cells, during which the bulk of the cytoplasm is engulfed by a double membrane vesicle, the autophagosome. Fusion of the autophagosome with the vacuole leads to breakdown of its contents, such as proteins and organelles, and the recycling of nutrients. Earlier studies of autophagic genes of the core autophagic machinery in the filamentous ascomycete Sordaria macrospora elucidated the impact of autophagy on fungal viability, vegetative growth and fruiting-body development. To gain further knowledge about the regulation of autophagy in S. macrospora, we analyzed the function of the bZIP transcription factor SmJLB1, a homolog of the Podospora anserina basic zipper-type transcription factor induced during incompatibility 4 (IDI-4) and the Aspergillus nidulans transcription factor jun-like bZIP A (JlbA). Generation of the homokaryotic deletion mutant demonstrated S. macrospora Smjlb1 is associated with autophagy-dependent processes. Deletion of Smjlb1 abolished fruiting-body formation and impaired vegetative growth. SmJLB1 is localized to the cytoplasm and to nuclei. Quantitative real-time PCR experiments revealed an upregulated expression of autophagy-related genes Smatg8 and Smatg4 in the Smjlb1 deletion mutant, suggesting a transcriptional repression function of SmJLB1. Copyright © 2013 Elsevier Inc. All rights reserved.

CDK-mediated activation of the SCF(FBXO) (28) ubiquitin ligase promotes MYC-driven transcription and tumourigenesis and predicts poor survival in breast cancer.

Science.gov (United States)

Cepeda, Diana; Ng, Hwee-Fang; Sharifi, Hamid Reza; Mahmoudi, Salah; Cerrato, Vanessa Soto; Fredlund, Erik; Magnusson, Kristina; Nilsson, Helén; Malyukova, Alena; Rantala, Juha; Klevebring, Daniel; Viñals, Francesc; Bhaskaran, Nimesh; Zakaria, Siti Mariam; Rahmanto, Aldwin Suryo; Grotegut, Stefan; Nielsen, Michael Lund; Szigyarto, Cristina Al-Khalili; Sun, Dahui; Lerner, Mikael; Navani, Sanjay; Widschwendter, Martin; Uhlén, Mathias; Jirström, Karin; Pontén, Fredrik; Wohlschlegel, James; Grandér, Dan; Spruck, Charles; Larsson, Lars-Gunnar; Sangfelt, Olle

2013-07-01

SCF (Skp1/Cul1/F-box) ubiquitin ligases act as master regulators of cellular homeostasis by targeting key proteins for ubiquitylation. Here, we identified a hitherto uncharacterized F-box protein, FBXO28 that controls MYC-dependent transcription by non-proteolytic ubiquitylation. SCF(FBXO28) activity and stability are regulated during the cell cycle by CDK1/2-mediated phosphorylation of FBXO28, which is required for its efficient ubiquitylation of MYC and downsteam enhancement of the MYC pathway. Depletion of FBXO28 or overexpression of an F-box mutant unable to support MYC ubiquitylation results in an impairment of MYC-driven transcription, transformation and tumourigenesis. Finally, in human breast cancer, high FBXO28 expression and phosphorylation are strong and independent predictors of poor outcome. In conclusion, our data suggest that SCF(FBXO28) plays an important role in transmitting CDK activity to MYC function during the cell cycle, emphasizing the CDK-FBXO28-MYC axis as a potential molecular drug target in MYC-driven cancers, including breast cancer. © 2013 The Authors. Published by John Wiley and Sons, Ltd on behalf of EMBO.

Bioinformatic Analyses of Subgroup-A Members of the Wheat bZIP Transcription Factor Family and Functional Identification of TabZIP174 Involved in Drought Stress Response

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Xueyin Li

2016-11-01

Full Text Available Extensive studies in Arabidopsis and rice have demonstrated that Subgroup-A members of the bZIP transcription factor family play important roles in plant responses to multiple abiotic stresses. Although common wheat (Triticum aestivum is one of the most widely cultivated and consumed food crops in the world, there are limited investigations into Subgroup A of the bZIP family in wheat. In this study, we performed bioinformatic analyses of the 41 Subgroup-A members of the wheat bZIP family. Phylogenetic and conserved motif analyses showed that most of the Subgroup-A bZIP proteins involved in abiotic stress responses of wheat, Arabidopsis and rice clustered in Clade A1 of the phylogenetic tree, and shared a majority of conserved motifs, suggesting the potential importance of Clade-A1 members in abiotic stress responses. Gene structure analysis showed that TabZIP genes with close phylogenetic relationships tended to possess similar exon-intron compositions, and the positions of introns in the hinge regions of the bZIP domains were highly conserved, whereas introns in the leucine zipper regions were at variable positions. Additionally, eleven groups of homologs and two groups of tandem paralogs were also identified in Subgroup A of the wheat bZIP family. Expression profiling analysis indicated that most Subgroup-A TabZIP genes were responsive to abscisic acid and various abiotic stress treatments. TabZIP27, TabZIP74, TabZIP138 and TabZIP174 proteins were localized in the nucleus of wheat protoplasts, whereas TabZIP9-GFP fusion protein was simultaneously present in the nucleus, cytoplasm and cell membrane. Transgenic Arabidopsis overexpressing TabZIP174 displayed increased seed germination rates and primary root lengths under drought treatments. Overexpression of TabZIP174 in transgenic Arabidopsis conferred enhanced drought tolerance, and transgenic plants exhibited lower water loss rates, higher survival rates, higher proline, soluble sugar and leaf

Bioinformatic Analyses of Subgroup-A Members of the Wheat bZIP Transcription Factor Family and Functional Identification of TabZIP174 Involved in Drought Stress Response

Science.gov (United States)

Li, Xueyin; Feng, Biane; Zhang, Fengjie; Tang, Yimiao; Zhang, Liping; Ma, Lingjian; Zhao, Changping; Gao, Shiqing

2016-01-01

Extensive studies in Arabidopsis and rice have demonstrated that Subgroup-A members of the bZIP transcription factor family play important roles in plant responses to multiple abiotic stresses. Although common wheat (Triticum aestivum) is one of the most widely cultivated and consumed food crops in the world, there are limited investigations into Subgroup A of the bZIP family in wheat. In this study, we performed bioinformatic analyses of the 41 Subgroup-A members of the wheat bZIP family. Phylogenetic and conserved motif analyses showed that most of the Subgroup-A bZIP proteins involved in abiotic stress responses of wheat, Arabidopsis, and rice clustered in Clade A1 of the phylogenetic tree, and shared a majority of conserved motifs, suggesting the potential importance of Clade-A1 members in abiotic stress responses. Gene structure analysis showed that TabZIP genes with close phylogenetic relationships tended to possess similar exon–intron compositions, and the positions of introns in the hinge regions of the bZIP domains were highly conserved, whereas introns in the leucine zipper regions were at variable positions. Additionally, eleven groups of homologs and two groups of tandem paralogs were also identified in Subgroup A of the wheat bZIP family. Expression profiling analysis indicated that most Subgroup-A TabZIP genes were responsive to abscisic acid and various abiotic stress treatments. TabZIP27, TabZIP74, TabZIP138, and TabZIP174 proteins were localized in the nucleus of wheat protoplasts, whereas TabZIP9-GFP fusion protein was simultaneously present in the nucleus, cytoplasm, and cell membrane. Transgenic Arabidopsis overexpressing TabZIP174 displayed increased seed germination rates and primary root lengths under drought treatments. Overexpression of TabZIP174 in transgenic Arabidopsis conferred enhanced drought tolerance, and transgenic plants exhibited lower water loss rates, higher survival rates, higher proline, soluble sugar, and leaf chlorophyll

Salt and drought stress and ABA responses related to bZIP genes from V. radiata and V. angularis.

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Wang, Lanfen; Zhu, Jifeng; Li, Xiaoming; Wang, Shumin; Wu, Jing

2018-04-20

Mung bean and adzuki bean are warm-season legumes widely cultivated in China. However, bean production in major producing regions is limited by biotic and abiotic stress, such as drought and salt stress. Basic leucine zipper (bZIP) genes play key roles in responses to various biotic and abiotic stresses. However, only several bZIP genes involved in drought and salt stress in legumes, especially Vigna radiata and Vigna angularis, have been identified. In this study, we identified 54 and 50 bZIP proteins from whole-genome sequences of V. radiata and V. angularis, respectively. First, we comprehensively surveyed the characteristics of all bZIP genes, including their gene structure, chromosome distribution and motif composition. Phylogenetic trees showed that VrbZIP and VabZIP proteins were divided into ten clades comprising nine known and one unknown subgroup. The results of the nucleotide substitution rate of the orthologous gene pairs showed that bZIP proteins have undergone strong purifying selection: V. radiata and V. angularis diverged 1.25 million years ago (mya) to 9.20 mya (average of 4.95 mya). We also found that many cis-acting regulatory elements (CAREs) involved in abiotic stress and plant hormone responses were detected in the putative promoter regions of the bZIP genes. Finally, using the quantitative real-time PCR (qRT-PCR) method, we performed expression profiling of the bZIP genes in response to drought, salt and abscisic acid (ABA). We identified several bZIP genes that may be involved in drought and salt responses. Generally, our results provided useful and rich resources of VrbZIP and VabZIP genes for the functional characterization and understanding of bZIP transcription factors (TFs) in warm-season legumes. In addition, our results revealed important and interesting data - a subset of VrbZIP and VabZIP gene expression profiles in response to drought, salt and ABA stress. These results provide gene expression evidence for the selection of

SCF, regulated by HIF-1α, promotes pancreatic ductal adenocarcinoma cell progression.

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Chuntao Gao

Full Text Available Stem cell factor (SCF and hypoxia-inducible factor-1α (HIF-1α both have important functions in pancreatic ductal adenocarcinoma (PDAC. This study aims to analyze the expression and clinicopathological significance of SCF and HIF-1α in PDAC specimens and explore the molecular mechanism at PDAC cells in vitro and in vivo. We showed that the expression of SCF was significantly correlated with HIF-1α expression via Western blot, PCR, chromatin immunoprecipitation (ChIP assay, and luciferase assay analysis. The SCF level was also correlated with lymph node metastasis and the pathological tumor node metastasis (pTNM stage in PDAC samples. The SCF higher-expression group had significantly lower survival rates than the SCF lower-expression group (p<0.05. Hypoxia up-regulated the expression of SCF through the hypoxia-inducible factor (HIF-1α in PDAC cells at the protein and RNA levels. When HIF-1α was knocked down by RNA interference, the SCF level decreased significantly. Additionally, ChIP and luciferase results demonstrated that HIF-1α can directly bind to the hypoxia response element (HRE region of the SCF promoter and activate the SCF transcription under hypoxia. The results of colony formation, cell scratch, and transwell migration assay showed that SCF promoted the proliferation and invasion of PANC-1 cells under hypoxia. Furthermore, the down-regulated ability of cell proliferation and invasion following HIF-1α knockdown was rescued by adding exogenous SCF under hypoxia in vitro. Finally, when the HIF-1α expression was inhibited by digoxin, the tumor volume and the SCF level decreased, thereby proving the relationship between HIF-1α and SCF in vivo. In conclusion, SCF is an important factor for the growth of PDAC. In our experiments, we proved that SCF, a downstream gene of HIF-1α, can promote the development of PDAC under hypoxia. Thus, SCF might be a potential therapeutic target for PDAC.

Identification of an Arabidopsis transmembrane bZIP transcription factor involved in the endoplasmic reticulum stress response

International Nuclear Information System (INIS)

Tajima, Hiromi; Iwata, Yuji; Iwano, Megumi; Takayama, Seiji; Koizumi, Nozomu

2008-01-01

Among 75 bZIP transcription factors identified in Arabidopsis, 3 (AtbZIP17, AtbZIP28, and AtbZIP49) possess a putative transmembrane domain (TMD) in addition to AtbZIP60, which was characterized previously. In the present study, cDNAs of AtbZIP17 and AtbZIP28 were isolated. Truncated forms of AtbZIP17 and AtbZIP28 lacking the C-terminal domain including TMD were examined as putative active forms. One of them, AtbZIP28ΔC, activated BiP1 and BiP3 promoters through the cis-elements P-UPRE and ERSE responsible for the ER stress response. Subsequently, a fusion protein of green fluorescent protein (GFP) and AtbZIP28 was expressed in Arabidopsis cultured cells. Under non-stress conditions, GFP fluorescence localization almost overlapped with an ER marker; however, tunicamycin and dithiothreitol treatment clearly increased GFP fluorescence in the nucleus suggesting that the N-terminal fragment of AtbZIP28 translocates to the nucleus in response to ER stress

Isolation and functional characterisation of two new bZIP maize regulators of the ABA responsive gene rab28.

Science.gov (United States)

Nieva, Claudia; Busk, Peter K; Domínguez-Puigjaner, Eva; Lumbreras, Victoria; Testillano, Pilar S; Risueño, Maria-Carmen; Pagès, Montserrat

2005-08-01

The plant hormone abscisic acid regulates gene expression in response to growth stimuli and abiotic stress. Previous studies have implicated members of the bZIP family of transcription factors as mediators of abscisic acid dependent gene expression through the ABRE cis-element. Here, we identify two new maize bZIP transcription factors, EmBP-2 and ZmBZ-1 related to EmBP-1 and OsBZ-8 families. They are differentially expressed during embryo development; EmBP-2 is constitutive, whereas ZmBZ-1 is abscisic acid-inducible and accumulates during late embryogenesis. Both factors are nuclear proteins that bind to ABREs and activate transcription of the abscisic acid-inducible gene rab28 from maize. EmBP-2 and ZmBZ-1 are phosphorylated by protein kinase CK2 and phosphorylation alters their DNA binding properties. Our data suggest that EmBP-2 and ZmBZ-1 are involved in the expression of abscisic acid inducible genes such as rab28 and their activity is modulated by ABA and by phosphorylation.

BACH transcription factors in innate and adaptive immunity.

Science.gov (United States)

Igarashi, Kazuhiko; Kurosaki, Tomohiro; Roychoudhuri, Rahul

2017-07-01

BTB and CNC homology (BACH) proteins are transcriptional repressors of the basic region leucine zipper (bZIP) transcription factor family. Recent studies indicate widespread roles of BACH proteins in controlling the development and function of the innate and adaptive immune systems, including the differentiation of effector and memory cells of the B and T cell lineages, CD4 + regulatory T cells and macrophages. Here, we emphasize similarities at a molecular level in the cell-type-specific activities of BACH factors, proposing that competitive interactions of BACH proteins with transcriptional activators of the bZIP family form a common mechanistic theme underlying their diverse actions. The findings contribute to a general understanding of how transcriptional repressors shape lineage commitment and cell-type-specific functions through repression of alternative lineage programmes.

A highly triflated rare-earth ion in [Eu(O_3SCF_3)_8]"5"-

International Nuclear Information System (INIS)

Bruns, Joern; Kluener, Thorsten; Kraeuter, Jessica; Wickleder, Mathias S.; Krueger, Sascha; Adlung, Matthias; Wickleder, Claudia; Niehaus, Oliver; Poettgen, Rainer

2015-01-01

The reaction of Eu_2O_3 with fuming nitric acid, trifluormethanesulfonic acid, and its anhydride in torch-sealed glass ampoules at 120 C gave the europium compound (NO)_5[Eu(O_3SCF_3)_8] (orthorhombic, Fddd, Z=16, a=1932.69(4), b=2878.44(7), c=2955.12(7) pm, V=16439.7(7) Aa"3). The compound exhibits the [Eu(O_3SCF_3)_8]"5"- anion showing for the first time a lanthanide ion that is exclusively coordinated by eight triflate anions. The anion has been further investigated by DFT calculations, which also allowed clear assignment of the vibrational spectra. Moreover, magnetochemical and luminescence measurements gave additional insight into the properties of this complex. The luminescence spectra revealed that the Eu"3"+ ions are in a pseudo D_4_d symmetric environment. (copyright 2015 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

Characterization of pollen-expressed bZIP protein interactions and the role of ATbZIP18 in the male gametophyte

Czech Academy of Sciences Publication Activity Database

Gibalová, A.; Steinbachová, L.; Hafidh, S.; Bláhová, Veronika; Gadiou, Z.; Michailidis, Ch.; Müller, K.; Pleskot, Roman; Dupľáková, N.; Honys, D.

2017-01-01

Roč. 30, č. 1 (2017), s. 1-17 ISSN 2194-7953 Institutional support: RVO:67985823 ; RVO:61388963 Keywords : bZIP * transcription factors * regulatory network * male gametophyte * Y2H * pollen development Subject RIV: ED - Physiology; CE - Biochemistry (UOCHB-X) OBOR OECD: Physiology (including cytology); Biochemistry and molecular biology (UOCHB-X) Impact factor: 2.629, year: 2016

Characterization of pollen-expressed bZIP protein interactions and the role of ATbZIP18 in the male gametophyte

Czech Academy of Sciences Publication Activity Database

Gibalová, Antónia; Steinbachová, Lenka; Hafidh, Said; Bláhová, Veronika; Gadiou, Zuzana; Michailidis, Christos; Müller, Karel; Pleskot, Roman; Dupľáková, Nikoleta; Honys, David

2017-01-01

Roč. 30, č. 1 (2017), s. 1-17 ISSN 2194-7953 R&D Projects: GA ČR GA15-22720S; GA ČR(CZ) GA14-32292S; GA ČR GP13-41444P; GA MŠk(CZ) LD14109 Institutional support: RVO:61389030 Keywords : reticulum stress-response * transcription factor * arabidopsis-thaliana * molecular-interactions * plant transformation * gene-expression * heterodimerization * dimerization * evolution * specificity * bZIP * Transcription factors * Regulatory network * Male gametophyte * y2h * Pollen development Subject RIV: ED - Physiology OBOR OECD: Plant sciences, botany Impact factor: 2.629, year: 2016

SlbZIP38, a Tomato bZIP Family Gene Downregulated by Abscisic Acid, Is a Negative Regulator of Drought and Salt Stress Tolerance

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Pan, Yanglu; Hu, Xin; Li, Chunyan; Xu, Xing; Su, Chenggang; Li, Jinhua; Song, Hongyuan; Zhang, Xingguo; Pan, Yu

2017-01-01

The basic leucine zipper (bZIP) transcription factors have crucial roles in plant stress responses. In this study, the bZIP family gene SlbZIP38 (GenBank accession No: XM004239373) was isolated from a tomato (Solanum lycopersicum cv. Ailsa Craig) mature leaf cDNA library. The DNA sequence of SlbZIP38 encodes a protein of 484 amino acids, including a highly conserved bZIP DNA-binding domain in the C-terminal region. We found that SlbZIP38 was differentially expressed in various organs of the tomato plant and was downregulated by drought, salt stress, and abscisic acid (ABA). However, overexpression of SlbZIP38 significantly decreased drought and salt stress tolerance in tomatoes (Ailsa Craig). The findings that SlbZIP38 overexpression reduced the chlorophyll and free proline content in leaves but increased the malondialdehyde content may explain the reduced drought and salt tolerance observed in these lines. These results suggest that SlbZIP38 is a negative regulator of drought and salt resistance that acts by modulating ABA signaling. PMID:29261143

SCF(KMD) controls cytokinin signaling by regulating the degradation of type-B response regulators.

Science.gov (United States)

Kim, Hyo Jung; Chiang, Yi-Hsuan; Kieber, Joseph J; Schaller, G Eric

2013-06-11

Cytokinins are plant hormones that play critical roles in growth and development. In Arabidopsis, the transcriptional response to cytokinin is regulated by action of type-B Arabidopsis response regulators (ARRs). Although central elements in the cytokinin signal transduction pathway have been identified, mechanisms controlling output remain to be elucidated. Here we demonstrate that a family of F-box proteins, called the kiss me deadly (KMD) family, targets type-B ARR proteins for degradation. KMD proteins form an S-phase kinase-associated PROTEIN1 (SKP1)/Cullin/F-box protein (SCF) E3 ubiquitin ligase complex and directly interact with type-B ARR proteins. Loss-of-function KMD mutants stabilize type-B ARRs and exhibit an enhanced cytokinin response. In contrast, plants with elevated KMD expression destabilize type-B ARR proteins leading to cytokinin insensitivity. Our results support a model in which an SCF(KMD) complex negatively regulates cytokinin responses by controlling levels of a key family of transcription factors.

Tribbles ortholog NIPI-3 and bZIP transcription factor CEBP-1 regulate a Caenorhabditis elegans intestinal immune surveillance pathway.

Science.gov (United States)

McEwan, Deborah L; Feinbaum, Rhonda L; Stroustrup, Nicholas; Haas, Wilhelm; Conery, Annie L; Anselmo, Anthony; Sadreyev, Ruslan; Ausubel, Frederick M

2016-12-07

Many pathogens secrete toxins that target key host processes resulting in the activation of immune pathways. The secreted Pseudomonas aeruginosa toxin Exotoxin A (ToxA) disrupts intestinal protein synthesis, which triggers the induction of a subset of P. aeruginosa-response genes in the nematode Caenorhabditis elegans. We show here that one ToxA-induced C. elegans gene, the Tribbles pseudokinase ortholog nipi-3, is essential for host survival following exposure to P. aeruginosa or ToxA. We find that NIPI-3 mediates the post-developmental expression of intestinal immune genes and proteins and primarily functions in parallel to known immune pathways, including p38 MAPK signaling. Through mutagenesis screening, we identify mutants of the bZIP C/EBP transcription factor cebp-1 that suppress the hypersusceptibility defects of nipi-3 mutants. NIPI-3 is a negative regulator of CEBP-1, which in turn negatively regulates protective immune mechanisms. This pathway represents a previously unknown innate immune signaling pathway in intestinal epithelial cells that is involved in the surveillance of cellular homeostasis. Because NIPI-3 and CEBP-1 are also essential for C. elegans development, NIPI-3 is analogous to other key innate immune signaling molecules such as the Toll receptors in Drosophila that have an independent role during development.

An active Mitochondrial Complex II Present in Mature Seeds Contains an Embryo-Specific Iron-Sulfur Subunit Regulated by ABA and bZIP53 and Is Involved in Germination and Seedling Establishment.

Science.gov (United States)

Restovic, Franko; Espinoza-Corral, Roberto; Gómez, Isabel; Vicente-Carbajosa, Jesús; Jordana, Xavier

2017-01-01

Complex II (succinate dehydrogenase) is an essential mitochondrial enzyme involved in both the tricarboxylic acid cycle and the respiratory chain. In Arabidopsis thaliana , its iron-sulfur subunit (SDH2) is encoded by three genes, one of them ( SDH2.3 ) being specifically expressed during seed maturation in the embryo. Here we show that seed SDH2.3 expression is regulated by abscisic acid (ABA) and we define the promoter region (-114 to +49) possessing all the cis -elements necessary and sufficient for high expression in seeds. This region includes between -114 and -32 three ABRE (ABA-responsive) elements and one RY-enhancer like element, and we demonstrate that these elements, although necessary, are not sufficient for seed expression, our results supporting a role for the region encoding the 5' untranslated region (+1 to +49). The SDH2.3 promoter is activated in leaf protoplasts by heterodimers between the basic leucine zipper transcription factors bZIP53 (group S1) and bZIP10 (group C) acting through the ABRE elements, and by the B3 domain transcription factor ABA insensitive 3 (ABI3). The in vivo role of bZIP53 is further supported by decreased SDH2.3 expression in a knockdown bzip53 mutant. By using the protein synthesis inhibitor cycloheximide and sdh2 mutants we have been able to conclusively show that complex II is already present in mature embryos before imbibition, and contains mainly SDH2.3 as iron-sulfur subunit. This complex plays a role during seed germination sensu-stricto since we have previously shown that seeds lacking SDH2.3 show retarded germination and now we demonstrate that low concentrations of thenoyltrifluoroacetone, a complex II inhibitor, also delay germination. Furthermore, complex II inhibitors completely block hypocotyl elongation in the dark and seedling establishment in the light, highlighting an essential role of complex II in the acquisition of photosynthetic competence and the transition from heterotrophy to autotrophy.

Characterization of StABF1, a stress-responsive bZIP transcription factor from Solanum tuberosum L. that is phosphorylated by StCDPK2 in vitro.

Science.gov (United States)

Muñiz García, María Noelia; Giammaria, Verónica; Grandellis, Carolina; Téllez-Iñón, María Teresa; Ulloa, Rita María; Capiati, Daniela Andrea

2012-04-01

ABF/AREB bZIP transcription factors mediate plant abiotic stress responses by regulating the expression of stress-related genes. These proteins bind to the abscisic acid (ABA)-responsive element (ABRE), which is the major cis-acting regulatory sequence in ABA-dependent gene expression. In an effort to understand the molecular mechanisms of abiotic stress resistance in cultivated potato (Solanum tuberosum L.), we have cloned and characterized an ABF/AREB-like transcription factor from potato, named StABF1. The predicted protein shares 45-57% identity with A. thaliana ABFs proteins and 96% identity with the S. lycopersicum SlAREB1 and presents all of the distinctive features of ABF/AREB transcription factors. Furthermore, StABF1 is able to bind to the ABRE in vitro. StABF1 gene is induced in response to ABA, drought, salt stress and cold, suggesting that it might be a key regulator of ABA-dependent stress signaling pathways in cultivated potato. StABF1 is phosphorylated in response to ABA and salt stress in a calcium-dependent manner, and we have identified a potato CDPK isoform (StCDPK2) that phosphorylates StABF1 in vitro. Interestingly, StABF1 expression is increased during tuber development and by tuber-inducing conditions (high sucrose/nitrogen ratio) in leaves. We also found that StABF1 calcium-dependent phosphorylation is stimulated by tuber-inducing conditions and inhibited by gibberellic acid, which inhibits tuberization.

Bone marrow adipocytes promote the regeneration of stem cells and hematopoiesis by secreting SCF

Science.gov (United States)

Zhou, Bo O.; Yu, Hua; Yue, Rui; Zhao, Zhiyu; Rios, Jonathan J.; Naveiras, Olaia; Morrison, Sean J.

2017-01-01

Endothelial cells and Leptin Receptor+ (LepR+) stromal cells are critical sources of haematopoietic stem cell (HSC) niche factors, including Stem Cell Factor (SCF), in bone marrow. After irradiation or chemotherapy, these cells are depleted while adipocytes become abundant. We discovered that bone marrow adipocytes synthesize SCF. They arise from Adipoq-Cre/ER+ progenitors, which represent ~5% of LepR+ cells, and proliferate after irradiation. Scf deletion using Adipoq-Cre/ER inhibited hematopoietic regeneration after irradiation or 5-fluorouracil treatment, depleting HSCs and reducing mouse survival. Scf from LepR+ cells, but not endothelial, hematopoietic, or osteoblastic cells, also promoted regeneration. In non-irradiated mice, Scf deletion using Adipoq-Cre/ER did not affect HSC frequency in long bones, which have few adipocytes, but depleted HSCs in tail vertebrae, which have abundant adipocytes. A-ZIP/F1 ‘fatless” mice exhibited delayed hematopoietic regeneration in long bones but not in tail vertebrae, where adipocytes inhibited vascularization. Adipocytes are a niche component that promotes hematopoietic regeneration. PMID:28714970

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

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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

Deregulation of sucrose-controlled translation of a bZIP-type transcription factor results in sucrose accumulation in leaves.

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Sunil Kumar Thalor

Full Text Available Sucrose is known to repress the translation of Arabidopsis thaliana AtbZIP11 transcript which encodes a protein belonging to the group of S (S--stands for small basic region-leucine zipper (bZIP-type transcription factor. This repression is called sucrose-induced repression of translation (SIRT. It is mediated through the sucrose-controlled upstream open reading frame (SC-uORF found in the AtbZIP11 transcript. The SIRT is reported for 4 other genes belonging to the group of S bZIP in Arabidopsis. Tobacco tbz17 is phylogenetically closely related to AtbZIP11 and carries a putative SC-uORF in its 5'-leader region. Here we demonstrate that tbz17 exhibits SIRT mediated by its SC-uORF in a manner similar to genes belonging to the S bZIP group of the Arabidopsis genus. Furthermore, constitutive transgenic expression of tbz17 lacking its 5'-leader region containing the SC-uORF leads to production of tobacco plants with thicker leaves composed of enlarged cells with 3-4 times higher sucrose content compared to wild type plants. Our finding provides a novel strategy to generate plants with high sucrose content.

Deregulation of sucrose-controlled translation of a bZIP-type transcription factor results in sucrose accumulation in leaves.

Science.gov (United States)

Thalor, Sunil Kumar; Berberich, Thomas; Lee, Sung Shin; Yang, Seung Hwan; Zhu, Xujun; Imai, Ryozo; Takahashi, Yoshihiro; Kusano, Tomonobu

2012-01-01

Sucrose is known to repress the translation of Arabidopsis thaliana AtbZIP11 transcript which encodes a protein belonging to the group of S (S--stands for small) basic region-leucine zipper (bZIP)-type transcription factor. This repression is called sucrose-induced repression of translation (SIRT). It is mediated through the sucrose-controlled upstream open reading frame (SC-uORF) found in the AtbZIP11 transcript. The SIRT is reported for 4 other genes belonging to the group of S bZIP in Arabidopsis. Tobacco tbz17 is phylogenetically closely related to AtbZIP11 and carries a putative SC-uORF in its 5'-leader region. Here we demonstrate that tbz17 exhibits SIRT mediated by its SC-uORF in a manner similar to genes belonging to the S bZIP group of the Arabidopsis genus. Furthermore, constitutive transgenic expression of tbz17 lacking its 5'-leader region containing the SC-uORF leads to production of tobacco plants with thicker leaves composed of enlarged cells with 3-4 times higher sucrose content compared to wild type plants. Our finding provides a novel strategy to generate plants with high sucrose content.

A highly triflated rare-earth ion in [Eu(O{sub 3}SCF{sub 3}){sub 8}]{sup 5-}

Energy Technology Data Exchange (ETDEWEB)

Bruns, Joern; Kluener, Thorsten; Kraeuter, Jessica; Wickleder, Mathias S. [Carl von Ossietzky Universitaet Oldenburg, Institut fuer Chemie (Germany); Krueger, Sascha; Adlung, Matthias; Wickleder, Claudia [Universitaet Siegen, Institut fuer Anorganische Chemie (Germany); Niehaus, Oliver; Poettgen, Rainer [Westfaelische Wilhelms Universitaet Muenster, Institut fuer Anorganische und Analytische Chemie (Germany)

2015-08-24

The reaction of Eu{sub 2}O{sub 3} with fuming nitric acid, trifluormethanesulfonic acid, and its anhydride in torch-sealed glass ampoules at 120 C gave the europium compound (NO){sub 5}[Eu(O{sub 3}SCF{sub 3}){sub 8}] (orthorhombic, Fddd, Z=16, a=1932.69(4), b=2878.44(7), c=2955.12(7) pm, V=16439.7(7) Aa{sup 3}). The compound exhibits the [Eu(O{sub 3}SCF{sub 3}){sub 8}]{sup 5-} anion showing for the first time a lanthanide ion that is exclusively coordinated by eight triflate anions. The anion has been further investigated by DFT calculations, which also allowed clear assignment of the vibrational spectra. Moreover, magnetochemical and luminescence measurements gave additional insight into the properties of this complex. The luminescence spectra revealed that the Eu{sup 3+} ions are in a pseudo D{sub 4d} symmetric environment. (copyright 2015 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

Bone marrow adipocytes promote the regeneration of stem cells and haematopoiesis by secreting SCF.

Science.gov (United States)

Zhou, Bo O; Yu, Hua; Yue, Rui; Zhao, Zhiyu; Rios, Jonathan J; Naveiras, Olaia; Morrison, Sean J

2017-08-01

Endothelial cells and leptin receptor + (LepR + ) stromal cells are critical sources of haematopoietic stem cell (HSC) niche factors, including stem cell factor (SCF), in bone marrow. After irradiation or chemotherapy, these cells are depleted while adipocytes become abundant. We discovered that bone marrow adipocytes synthesize SCF. They arise from Adipoq-Cre/ER + progenitors, which represent ∼5% of LepR + cells, and proliferate after irradiation. Scf deletion using Adipoq-Cre/ER inhibited haematopoietic regeneration after irradiation or 5-fluorouracil treatment, depleting HSCs and reducing mouse survival. Scf from LepR + cells, but not endothelial, haematopoietic or osteoblastic cells, also promoted regeneration. In non-irradiated mice, Scf deletion using Adipoq-Cre/ER did not affect HSC frequency in long bones, which have few adipocytes, but depleted HSCs in tail vertebrae, which have abundant adipocytes. A-ZIP/F1 'fatless' mice exhibited delayed haematopoietic regeneration in long bones but not in tail vertebrae, where adipocytes inhibited vascularization. Adipocytes are a niche component that promotes haematopoietic regeneration.

Evolutionary and Expression Analyses of the Apple Basic Leucine Zipper Transcription Factor Family

Science.gov (United States)

Zhao, Jiao; Guo, Rongrong; Guo, Chunlei; Hou, Hongmin; Wang, Xiping; Gao, Hua

2016-01-01

Transcription factors (TFs) play essential roles in the regulatory networks controlling many developmental processes in plants. Members of the basic leucine (Leu) zipper (bZIP) TF family, which is unique to eukaryotes, are involved in regulating diverse processes, including flower and vascular development, seed maturation, stress signaling, and defense responses to pathogens. The bZIP proteins have a characteristic bZIP domain composed of a DNA-binding basic region and a Leu zipper dimerization region. In this study, we identified 112 apple (Malus domestica Borkh) bZIP TF-encoding genes, termed MdbZIP genes. Synteny analysis indicated that segmental and tandem duplication events, as well as whole genome duplication, have contributed to the expansion of the apple bZIP family. The family could be divided into 11 groups based on structural features of the encoded proteins, as well as on the phylogenetic relationship of the apple bZIP proteins to those of the model plant Arabidopsis thaliana (AtbZIP genes). Synteny analysis revealed that several paired MdbZIP genes and AtbZIP gene homologs were located in syntenic genomic regions. Furthermore, expression analyses of group A MdbZIP genes showed distinct expression levels in 10 different organs. Moreover, changes in these expression profiles in response to abiotic stress conditions and various hormone treatments identified MdbZIP genes that were responsive to high salinity and drought, as well as to different phytohormones. PMID:27066030

Evolutionary and Expression Analyses of the Apple Basic Leucine Zipper Transcription Factor Family

Directory of Open Access Journals (Sweden)

Jiao eZhao

2016-03-01

Full Text Available Transcription factors (TFs play essential roles in the regulatory networks controlling many developmental processes in plants. Members of the basic leucine (Leu zipper (bZIP TF family, which is unique to eukaryotes, are involved in regulating diverse processes, including flower and vascular development, seed maturation, stress signaling and defense responses to pathogens. The bZIP proteins have a characteristic bZIP domain composed of a DNA-binding basic region and a Leu zipper dimerization region. In this study, we identified 112 apple (Malus domestica Borkh bZIP TF-encoding genes, termed MdbZIP genes. Synteny analysis indicated that segmental and tandem duplication events, as well as whole genome duplication, have contributed to the expansion of the apple bZIP family. The family could be divided into 11 groups based on structural features of the encoded proteins, as well as on the phylogenetic relationship of the apple bZIP proteins to those of the model plant Arabidopsis thaliana (AtbZIP genes. Synteny analysis revealed that several paired MdbZIP genes and AtbZIP gene homologs were located in syntenic genomic regions. Furthermore, expression analyses of group A MdbZIP genes showed distinct expression levels in ten different organs. Moreover, changes in these expression profiles in response to abiotic stress conditions and various hormone treatments identified MdbZIP genes that were responsive to high salinity and drought, as well as to different phytohormones.

A novel bZIP gene from Tamarix hispida mediates physiological responses to salt stress in tobacco plants.

Science.gov (United States)

Wang, Yucheng; Gao, Caiqiu; Liang, Yenan; Wang, Chao; Yang, Chuanping; Liu, Guifeng

2010-02-15

Basic leucine zipper proteins (bZIPs) are transcription factors that bind abscisic acid (ABA)-responsive elements (ABREs) and enable plants to withstand adverse environmental conditions. In the present study, a novel bZIP gene, ThbZIP1 was cloned from Tamarix hispida. Expression studies in T. hispida showed differential regulation of ThbZIP1 in response to treatment with NaCl, polyethylene glycol (PEG) 6000, NaHCO(3), and CdCl(2), suggesting that ThbZIP1 is involved in abiotic stress responses. To identify the physiological responses mediated by ThbZIP1, transgenic tobacco plants overexpressing exogenous ThbZIP1 were generated. Various physiological parameters related to salt stress were measured and compared between transgenic and wild type (WT) plants. Our results indicate that overexpression of ThbZIP1 can enhance the activity of both peroxidase (POD) and superoxide dismutase (SOD), and increase the content of soluble sugars and soluble proteins under salt stress conditions. These results suggest that ThbZIP1 contributes to salt tolerance by mediating signaling through multiple physiological pathways. Furthermore, ThbZIP1 confers stress tolerance to plants by enhancing reactive oxygen species (ROS) scavenging, facilitating the accumulation of compatible osmolytes, and inducing and/or enhancing the biosynthesis of soluble proteins. Copyright 2009 Elsevier GmbH. All rights reserved.

Starting SCF Calculations by Superposition of Atomic Densities

NARCIS (Netherlands)

van Lenthe, J.H.; Zwaans, R.; van Dam, H.J.J.; Guest, M.F.

2006-01-01

We describe the procedure to start an SCF calculation of the general type from a sum of atomic electron densities, as implemented in GAMESS-UK. Although the procedure is well-known for closed-shell calculations and was already suggested when the Direct SCF procedure was proposed, the general

Regulating expressin of cell and tissue-specific genes by modifying transcription

Energy Technology Data Exchange (ETDEWEB)

Beachy, Roger N. [Donald Danforth Plant Science Center, St. Louis, MO (United States); Dai, Shunhong [Donald Danforth Plant Science Center, St. Louis, MO (United States)

2009-12-15

Transcriptional regulation is the primary step to control gene expression, therefore function. Such regulation is achieved primarily via a combination of the activities of the promoter cis regulatory DNA elements and trans regulatory proteins that function through binding to these DNA elements. Our research supported by this program has led to the identification of rice bZIP transcription factors RF2a, RF2b and RLP1 that play key roles in regulating the activity of a vascular tissue specific promoter isolated from Rice Tungro Bacilliform Virus (RTBV) through their interactions with the Box II essential cis element located in the promoter. RF2a, RF2b and RLP1 possess multiple regulatory domains. Functional characterization reveals that those domains can activate or repress the activity of the RTBV promoter. Studies of transcriptional regulation of the RTBV promoter by this group of bZIP proteins not only provide insights about gene expression in the vascular tissue, but also insights about general mechanisms of transcription activation and repression. The knowledge gained from this research will also enable us to develop a well-described set of tools that can be used to control expression of multiple genes in transgenic plants and to improve biofuel feedstock.

Tautomerism in 5-aminotetrazole investigated by core-level photoelectron spectroscopy and {Delta}SCF calculations

Energy Technology Data Exchange (ETDEWEB)

Pinto, R.M., E-mail: ruipinto@fct.unl.pt [CFA, Centro de Fisica Atomica, Departamento de Fisica, Faculdade de Ciencias e Tecnologia, FCT, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal); Dias, A.A. [CFA, Centro de Fisica Atomica, Departamento de Fisica, Faculdade de Ciencias e Tecnologia, FCT, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal); Coreno, M. [CNR-IMIP, Montelibretti, Rome I-00016 (Italy); Simone, M. de [CNR-IOM, Laboratorio TASC, 34149 Trieste (Italy); Giuliano, B.M. [Departamento de Quimica da Universidade de Coimbra, 3004-535 Coimbra (Portugal); Santos, J.P.; Costa, M.L. [CFA, Centro de Fisica Atomica, Departamento de Fisica, Faculdade de Ciencias e Tecnologia, FCT, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal)

2012-03-15

Highlights: Black-Right-Pointing-Pointer High-resolution XPS of 5-aminotetrazole reveals different tautomers. Black-Right-Pointing-Pointer 5ATZ exists mainly in the 2H-form. Black-Right-Pointing-Pointer Results obtained with DSCF are in good agreement with the observed binding energies. - Abstract: The C 1s and N 1s photoelectron spectra of gas-phase 5-aminotetrazole (5ATZ) were recorded using synchrotron radiation, with the aim of evaluating 1H/2H tautomer population ratios. The core-electron binding energies (CEBEs) were estimated from computational results, using the delta self-consistent-field ({Delta}SCF) approach. Simulated spectra were generated using these CEBEs and the results from GAUSSIAN-n (Gn, n = 1, 2 and 3) and Complete Basis Set (CBS-4M and CBS-Q) methods. Results reveal the almost exclusive predominance of the 2H-tautomer, with a 1H/2H ratio of ca. 0.12/0.88, taken from a gross analysis of the XPS C 1s spectrum, recorded at 365 K.

ABI-like transcription factor gene TaABL1 from wheat improves multiple abiotic stress tolerances in transgenic plants.

Science.gov (United States)

Xu, Dong-Bei; Gao, Shi-Qing; Ma, You-Zhi; Xu, Zhao-Shi; Zhao, Chang-Ping; Tang, Yi-Miao; Li, Xue-Yin; Li, Lian-Cheng; Chen, Yao-Feng; Chen, Ming

2014-12-01

The phytohormone abscisic acid (ABA) plays crucial roles in adaptive responses of plants to abiotic stresses. ABA-responsive element binding proteins (AREBs) are basic leucine zipper transcription factors that regulate the expression of downstream genes containing ABA-responsive elements (ABREs) in promoter regions. A novel ABI-like (ABA-insensitive) transcription factor gene, named TaABL1, containing a conserved basic leucine zipper (bZIP) domain was cloned from wheat. Southern blotting showed that three copies were present in the wheat genome. Phylogenetic analyses indicated that TaABL1 belonged to the AREB subfamily of the bZIP transcription factor family and was most closely related to ZmABI5 in maize and OsAREB2 in rice. Expression of TaABL1 was highly induced in wheat roots, stems, and leaves by ABA, drought, high salt, and low temperature stresses. TaABL1 was localized inside the nuclei of transformed wheat mesophyll protoplast. Overexpression of TaABL1 enhanced responses of transgenic plants to ABA and hastened stomatal closure under stress, thereby improving tolerance to multiple abiotic stresses. Furthermore, overexpression of TaABL1 upregulated or downregulated the expression of some stress-related genes controlling stomatal closure in transgenic plants under ABA and drought stress conditions, suggesting that TaABL1 might be a valuable genetic resource for transgenic molecular breeding.

HTLV-1 bZIP factor induces T-cell lymphoma and systemic inflammation in vivo.

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Yorifumi Satou

2011-02-01

Full Text Available Human T-cell leukemia virus type 1 (HTLV-1 is the causal agent of a neoplastic disease of CD4+ T cells, adult T-cell leukemia (ATL, and inflammatory diseases including HTLV-1 associated myelopathy/tropical spastic paraparesis, dermatitis, and inflammatory lung diseases. ATL cells, which constitutively express CD25, resemble CD25+CD4+ regulatory T cells (T(reg. Approximately 60% of ATL cases indeed harbor leukemic cells that express FoxP3, a key transcription factor for T(reg cells. HTLV-1 encodes an antisense transcript, HTLV-1 bZIP factor (HBZ, which is expressed in all ATL cases. In this study, we show that transgenic expression of HBZ in CD4+ T cells induced T-cell lymphomas and systemic inflammation in mice, resembling diseases observed in HTLV-1 infected individuals. In HBZ-transgenic mice, CD4+Foxp3+ T(reg cells and effector/memory CD4+ T cells increased in vivo. As a mechanism of increased T(reg cells, HBZ expression directly induced Foxp3 gene transcription in T cells. The increased CD4+Foxp3+ T(reg cells in HBZ transgenic mice were functionally impaired while their proliferation was enhanced. HBZ could physically interact with Foxp3 and NFAT, thereby impairing the suppressive function of T(reg cells. Thus, the expression of HBZ in CD4+ T cells is a key mechanism of HTLV-1-induced neoplastic and inflammatory diseases.

N-terminal segments modulate the α-helical propensities of the intrinsically disordered basic regions of bZIP proteins.

Science.gov (United States)

Das, Rahul K; Crick, Scott L; Pappu, Rohit V

2012-02-17

Basic region leucine zippers (bZIPs) are modular transcription factors that play key roles in eukaryotic gene regulation. The basic regions of bZIPs (bZIP-bRs) are necessary and sufficient for DNA binding and specificity. Bioinformatic predictions and spectroscopic studies suggest that unbound monomeric bZIP-bRs are uniformly disordered as isolated domains. Here, we test this assumption through a comparative characterization of conformational ensembles for 15 different bZIP-bRs using a combination of atomistic simulations and circular dichroism measurements. We find that bZIP-bRs have quantifiable preferences for α-helical conformations in their unbound monomeric forms. This helicity varies from one bZIP-bR to another despite a significant sequence similarity of the DNA binding motifs (DBMs). Our analysis reveals that intramolecular interactions between DBMs and eight-residue segments directly N-terminal to DBMs are the primary modulators of bZIP-bR helicities. We test the accuracy of this inference by designing chimeras of bZIP-bRs to have either increased or decreased overall helicities. Our results yield quantitative insights regarding the relationship between sequence and the degree of intrinsic disorder within bZIP-bRs, and might have general implications for other intrinsically disordered proteins. Understanding how natural sequence variations lead to modulation of disorder is likely to be important for understanding the evolution of specificity in molecular recognition through intrinsically disordered regions (IDRs). Copyright © 2011 Elsevier Ltd. All rights reserved.

Identification of the G13 (cAMP-response-element-binding protein-related protein) gene product related to activating transcription factor 6 as a transcriptional activator of the mammalian unfolded protein response.

Science.gov (United States)

Haze, K; Okada, T; Yoshida, H; Yanagi, H; Yura, T; Negishi, M; Mori, K

2001-04-01

Eukaryotic cells control the levels of molecular chaperones and folding enzymes in the endoplasmic reticulum (ER) by a transcriptional induction process termed the unfolded protein response (UPR). The mammalian UPR is mediated by the cis-acting ER stress response element consisting of 19 nt (CCAATN(9)CCACG), the CCACG part of which is considered to provide specificity. We recently identified the basic leucine zipper (bZIP) protein ATF6 as a mammalian UPR-specific transcription factor; ATF6 is activated by ER stress-induced proteolysis and binds directly to CCACG. Here we report that eukaryotic cells express another bZIP protein closely related to ATF6 in both structure and function. This protein encoded by the G13 (cAMP response element binding protein-related protein) gene is constitutively synthesized as a type II transmembrane glycoprotein anchored in the ER membrane and processed into a soluble form upon ER stress as occurs with ATF6. The proteolytic processing of ATF6 and the G13 gene product is accompanied by their relocation from the ER to the nucleus; their basic regions seem to function as a nuclear localization signal. Overexpression of the soluble form of the G13 product constitutively activates the UPR, whereas overexpression of a mutant lacking the activation domain exhibits a strong dominant-negative effect. Furthermore, the soluble forms of ATF6 and the G13 gene product are unable to bind to several point mutants of the cis-acting ER stress response element in vitro that hardly respond to ER stress in vivo. We thus concluded that the two related bZIP proteins are crucial transcriptional regulators of the mammalian UPR, and propose calling the ATF6 gene product ATF6alpha and the G13 gene product ATF6beta.

Multi-time series RNA-seq analysis of Enterobacter lignolyticus SCF1 during growth in lignin-amended medium

Energy Technology Data Exchange (ETDEWEB)

Orellana, Roberto; Chaput, Gina; Markillie, Lye Meng; Mitchell, Hugh; Gaffrey, Matt; Orr, Galya; DeAngelis, Kristen M.; Yang, Shihui

2017-10-19

The production of lignocellulosic-derived biofuels is a highly promising source of alternative energy, but it has been constrained by the lack of a microbial platform capable to efficiently degrade this recalcitrant material and cope with by-products that can be toxic to cells. Species that naturally grow in environments where carbon is mainly available as lignin are promising for finding new ways of removing the lignin that protects cellulose for improved conversion of lignin to fuel precursors. Enterobacter lignolyticus SCF1 is a facultative anaerobic Gammaproteobacteria isolated from tropical rain forest soil collected in El Yunque forest, Puerto Rico under anoxic growth conditions with lignin as sole carbon source. Whole transcriptome analysis of SCF1 during E.lignolyticus SCF1 lignin degradation was conducted on cells grown in the presence (0.1%, w/w) and the absence of lignin, where samples were taken at three different times during growth, beginning of exponential phase, midexponential phase and beginning of stationary phase. Lignin-amended cultures achieved twice the cell biomass as unamended cultures over three days, and in this time degraded 60% of lignin. Transcripts in early exponential phase reflected this accelerated growth. A complement of laccases, aryl-alcohol dehydrogenases, and peroxidases were most up-regulated in lignin amended conditions in mid-exponential and early stationary phases compared to unamended growth. The association of hydrogen production by way of the formate hydrogenlyase complex with lignin degradation suggests a possible value added to lignin degradation in the future.

Multi-time series RNA-seq analysis of Enterobacter lignolyticus SCF1 during growth in lignin-amended medium.

Science.gov (United States)

Orellana, Roberto; Chaput, Gina; Markillie, Lye Meng; Mitchell, Hugh; Gaffrey, Matt; Orr, Galya; DeAngelis, Kristen M

2017-01-01

The production of lignocellulosic-derived biofuels is a highly promising source of alternative energy, but it has been constrained by the lack of a microbial platform capable to efficiently degrade this recalcitrant material and cope with by-products that can be toxic to cells. Species that naturally grow in environments where carbon is mainly available as lignin are promising for finding new ways of removing the lignin that protects cellulose for improved conversion of lignin to fuel precursors. Enterobacter lignolyticus SCF1 is a facultative anaerobic Gammaproteobacteria isolated from tropical rain forest soil collected in El Yunque forest, Puerto Rico under anoxic growth conditions with lignin as sole carbon source. Whole transcriptome analysis of SCF1 during E.lignolyticus SCF1 lignin degradation was conducted on cells grown in the presence (0.1%, w/w) and the absence of lignin, where samples were taken at three different times during growth, beginning of exponential phase, mid-exponential phase and beginning of stationary phase. Lignin-amended cultures achieved twice the cell biomass as unamended cultures over three days, and in this time degraded 60% of lignin. Transcripts in early exponential phase reflected this accelerated growth. A complement of laccases, aryl-alcohol dehydrogenases, and peroxidases were most up-regulated in lignin amended conditions in mid-exponential and early stationary phases compared to unamended growth. The association of hydrogen production by way of the formate hydrogenlyase complex with lignin degradation suggests a possible value added to lignin degradation in the future.

SCF analysis of a pressurized vessel-nozzle intersection with wall thinning damage

International Nuclear Information System (INIS)

Qadir, M.; Redekop, D.

2009-01-01

A three-dimensional finite element analysis is carried out of a pressurized vessel-nozzle intersection (tee joint), with wall thinning damage. A convergence-validation study is first carried out for undamaged intersections, in which comparisons are made with previously published work for the stress concentration factor (SCF), and good agreement is observed. A study is then carried out for specific tee joints to examine the effect on the SCF of varying the extent of the wall thinning damage. Finally, a parametric study is conducted in which the SCF is computed for a wide range of tee joints, initially considered undamaged, and then with wall thinning damage.

Effect of copy number and spacing of the ACGT and GT cis elements ...

Indian Academy of Sciences (India)

Unknown

cognized by transcription factors of the bZIP family. The core ACGT element occurs at different relative positions in one or more copies upstream of the minimal promoter region. Protein-DNA interaction studies have shown that sequences flanking the ACGT core affect bZIP protein binding specificity. The bZIP transcription ...

ATF3, an HTLV-1 bZip factor binding protein, promotes proliferation of adult T-cell leukemia cells

Directory of Open Access Journals (Sweden)

Ohshima Koichi

2011-03-01

Full Text Available Abstract Background Adult T-cell leukemia (ATL is an aggressive malignancy of CD4+ T-cells caused by human T-cell leukemia virus type 1 (HTLV-1. The HTLV-1 bZIP factor (HBZ gene, which is encoded by the minus strand of the viral genome, is expressed as an antisense transcript in all ATL cases. By using yeast two-hybrid screening, we identified activating transcription factor 3 (ATF3 as an HBZ-interacting protein. ATF3 has been reported to be expressed in ATL cells, but its biological significance is not known. Results Immunoprecipitation analysis confirmed that ATF3 interacts with HBZ. Expression of ATF3 was upregulated in ATL cell lines and fresh ATL cases. Reporter assay revealed that ATF3 could interfere with the HTLV-1 Tax's transactivation of the 5' proviral long terminal repeat (LTR, doing so by affecting the ATF/CRE site, as well as HBZ. Suppressing ATF3 expression inhibited proliferation and strongly reduced the viability of ATL cells. As mechanisms of growth-promoting activity of ATF3, comparative expression profiling of ATF3 knockdown cells identified candidate genes that are critical for the cell cycle and cell death, including cell division cycle 2 (CDC2 and cyclin E2. ATF3 also enhanced p53 transcriptional activity, but this activity was suppressed by HBZ. Conclusions Thus, ATF3 expression has positive and negative effects on the proliferation and survival of ATL cells. HBZ impedes its negative effects, leaving ATF3 to promote proliferation of ATL cells via mechanisms including upregulation of CDC2 and cyclin E2. Both HBZ and ATF3 suppress Tax expression, which enables infected cells to escape the host immune system.

Wheat Transcription Factor TaAREB3 Participates in Drought and Freezing Tolerances in Arabidopsis.

Science.gov (United States)

Wang, Jingyi; Li, Qian; Mao, Xinguo; Li, Ang; Jing, Ruilian

2016-01-01

AREB (ABA response element binding) proteins in plants play direct regulatory roles in response to multiple stresses, but their functions in wheat (Triticum aestivum L.) are not clear. In the present study, TaAREB3, a new member of the AREB transcription factor family, was isolated from wheat. Sequence analysis showed that the TaAREB3 protein is composed of three parts, a conserved N-terminal, a variable M region, and a conserved C-terminal with a bZIP domain. It belongs to the group A subfamily of bZIP transcription factors. TaAREB3 was constitutively expressed in stems, leaves, florets, anthers, pistils, seeds, and most highly, in roots. TaAREB3 gene expression was induced with abscisic acid (ABA) and low temperature stress, and its protein was localized in the nucleus when transiently expressed in tobacco epidermal cells and stably expressed in transgenic Arabidopsis. TaAREB3 protein has transcriptional activation activity, and can bind to the ABRE cis-element in vitro. Overexpression of TaAREB3 in Arabidopsis not only enhanced ABA sensitivity, but also strengthened drought and freezing tolerances. TaAREB3 also activated RD29A, RD29B, COR15A, and COR47 by binding to their promoter regions in transgenic Arabidopsis. These results demonstrated that TaAREB3 plays an important role in drought and freezing tolerances in Arabidopsis.

σ-SCF: A direct energy-targeting method to mean-field excited states.

Science.gov (United States)

Ye, Hong-Zhou; Welborn, Matthew; Ricke, Nathan D; Van Voorhis, Troy

2017-12-07

The mean-field solutions of electronic excited states are much less accessible than ground state (e.g., Hartree-Fock) solutions. Energy-based optimization methods for excited states, like Δ-SCF (self-consistent field), tend to fall into the lowest solution consistent with a given symmetry-a problem known as "variational collapse." In this work, we combine the ideas of direct energy-targeting and variance-based optimization in order to describe excited states at the mean-field level. The resulting method, σ-SCF, has several advantages. First, it allows one to target any desired excited state by specifying a single parameter: a guess of the energy of that state. It can therefore, in principle, find all excited states. Second, it avoids variational collapse by using a variance-based, unconstrained local minimization. As a consequence, all states-ground or excited-are treated on an equal footing. Third, it provides an alternate approach to locate Δ-SCF solutions that are otherwise hardly accessible by the usual non-aufbau configuration initial guess. We present results for this new method for small atoms (He, Be) and molecules (H 2 , HF). We find that σ-SCF is very effective at locating excited states, including individual, high energy excitations within a dense manifold of excited states. Like all single determinant methods, σ-SCF shows prominent spin-symmetry breaking for open shell states and our results suggest that this method could be further improved with spin projection.

σ-SCF: A direct energy-targeting method to mean-field excited states

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Ye, Hong-Zhou; Welborn, Matthew; Ricke, Nathan D.; Van Voorhis, Troy

2017-12-01

The mean-field solutions of electronic excited states are much less accessible than ground state (e.g., Hartree-Fock) solutions. Energy-based optimization methods for excited states, like Δ-SCF (self-consistent field), tend to fall into the lowest solution consistent with a given symmetry—a problem known as "variational collapse." In this work, we combine the ideas of direct energy-targeting and variance-based optimization in order to describe excited states at the mean-field level. The resulting method, σ-SCF, has several advantages. First, it allows one to target any desired excited state by specifying a single parameter: a guess of the energy of that state. It can therefore, in principle, find all excited states. Second, it avoids variational collapse by using a variance-based, unconstrained local minimization. As a consequence, all states—ground or excited—are treated on an equal footing. Third, it provides an alternate approach to locate Δ-SCF solutions that are otherwise hardly accessible by the usual non-aufbau configuration initial guess. We present results for this new method for small atoms (He, Be) and molecules (H2, HF). We find that σ-SCF is very effective at locating excited states, including individual, high energy excitations within a dense manifold of excited states. Like all single determinant methods, σ-SCF shows prominent spin-symmetry breaking for open shell states and our results suggest that this method could be further improved with spin projection.

Studies on Transcriptional Incorporation of 5'-N-Triphosphates of 5'-Amino-5'-Deoxyribonucleosides.

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Weronika Kotkowiak

Full Text Available In this study, several RNA polymerases were used for the first time to examine the possibility of transcriptional incorporation of 5'-N-triphosphates of 5'-amino-5'-deoxyribonucleosides (5'NH NTPs. The T3, T7, Sp6 and T7 Y639F RNA polymerases were employed to show that the full-length transcript cannot be synthesized. The results suggest that the application of 5'NH NTPs could decrease transcription reaction rates. What is more, the modification of transcription conditions had no influence on the rate of 5'NH NTPs incorporation. Based on experimental data it is postulated that 5'NH NTPs can be used as potential transcription inhibitors. Our findings expand the knowledge on suitable uses of the 5'-N-triphosphates of 5'-amino-5'-deoxyribonucleoside and the exact mechanism of transcriptional inhibition.

The SCF ubiquitin ligase Slimb controls Nerfin-1 turnover in Drosophila.

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Lin, Xiaohui; Wang, Feng; Li, Yuanpei; Zhai, Chaojun; Wang, Guiping; Zhang, Xiaoting; Gao, Yang; Yi, Tao; Sun, Dan; Wu, Shian

2018-01-01

The C2H2 type zinc-finger transcription factor Nerfin-1 expresses dominantly in Drosophila nervous system and plays an important role in early axon guidance decisions and preventing neurons dedifferentiation. Recently, increasing reports indicated that INSM1 (homologue to nerfin-1 in mammals) is a useful marker for prognosis of neuroendocrine tumors. The dynamic expression of Nerfin-1 is regulated post-transcriptionally by multiple microRNAs; however, its post-translational regulation is still unclear. Here we showed that the protein turnover of Nerfin-1 is regulated by Slimb, the substrate adaptor of SCF Slimb ubiquitin ligase complex. Mechanistically, Slimb associates with Nerfin-1 and promotes it ubiquitination and degradation in Drosophila S2R + cells. Furthermore, we determined that the C-terminal half of Nerfin-1 (Nerfin-1 CT ) is required for its binding to Slimb. Genetic epistasis assays showed that Slimb misexpression antagonizes, while knock-down enhances the activity of Nerfin-1 CT in Drosophila eyes. Our data revealed a new link to understand the underlying mechanism for Nerfin-1 turnover in post-translational level, and provided useful insights in animal development and disease treatment by manipulating the activity of Slimb and Nerfin-1. Copyright © 2017 Elsevier Inc. All rights reserved.

The Multifaceted Roles of HY5 in Plant Growth and Development.

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Gangappa, Sreeramaiah N; Botto, Javier F

2016-10-10

ELONGATED HYPOCOTYL5 (HY5), a member of the bZIP transcription factor family, inhibits hypocotyl growth and lateral root development, and promotes pigment accumulation in a light-dependent manner in Arabidopsis. Recent research on its role in different processes such as hormone, nutrient, abiotic stress (abscisic acid, salt, cold), and reactive oxygen species signaling pathways clearly places HY5 at the center of a transcriptional network hub. HY5 regulates the transcription of a large number of genes by directly binding to cis-regulatory elements. Recently, HY5 has also been shown to activate its own expression under both visible and UV-B light. Moreover, HY5 acts as a signal that moves from shoot to root to promote nitrate uptake and root growth. Here, we review recent advances on HY5 research in diverse aspects of plant development and highlight still open questions that need to be addressed in the near future for a complete understanding of its function in plant signaling and beyond. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

The COP9 signalosome interacts with SCF UFO and participates in Arabidopsis flower development.

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Wang, Xiping; Feng, Suhua; Nakayama, Naomi; Crosby, W L; Irish, Vivian; Deng, Xing Wang; Wei, Ning

2003-05-01

The COP9 signalosome (CSN) is involved in multiple developmental processes. It interacts with SCF ubiquitin ligases and deconjugates Nedd8/Rub1 from cullins (deneddylation). CSN is highly expressed in Arabidopsis floral tissues. To investigate the role of CSN in flower development, we examined the expression pattern of CSN in developing flowers. We report here that two csn1 partially deficient Arabidopsis strains exhibit aberrant development of floral organs, decline of APETALA3 (AP3) expression, and low fertility in addition to defects in shoot and inflorescence meristems. We show that UNUSUAL FLORAL ORGANS (UFO) forms a SCF(UFO) complex, which is associated with CSN in vivo. Genetic interaction analysis indicates that CSN is necessary for the gain-of-function activity of the F-box protein UFO in AP3 activation and in floral organ transformation. Compared with the previously reported csn5 antisense and csn1 null mutants, partial deficiency of CSN1 causes a reduction in the level of CUL1 in the mutant flowers without an obvious defect in CUL1 deneddylation. We conclude that CSN is an essential regulator of Arabidopsis flower development and suggest that CSN regulates Arabidopsis flower development in part by modulating SCF(UFO)-mediated AP3 activation.

The leucine zipper domains of the transcription factors GCN4 and c-Jun have ribonuclease activity.

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Yaroslav Nikolaev

Full Text Available Basic-region leucine zipper (bZIP proteins are one of the largest transcription factor families that regulate a wide range of cellular functions. Owing to the stability of their coiled coil structure leucine zipper (LZ domains of bZIP factors are widely employed as dimerization motifs in protein engineering studies. In the course of one such study, the X-ray structure of the retro-version of the LZ moiety of yeast transcriptional activator GCN4 suggested that this retro-LZ may have ribonuclease activity. Here we show that not only the retro-LZ but also the authentic LZ of GCN4 has weak but distinct ribonuclease activity. The observed cleavage of RNA is unspecific, it is not suppressed by the ribonuclease A inhibitor RNasin and involves the breakage of 3',5'-phosphodiester bonds with formation of 2',3'-cyclic phosphates as the final products as demonstrated by HPLC/electrospray ionization mass spectrometry. Several mutants of the GCN4 leucine zipper are catalytically inactive, providing important negative controls and unequivocally associating the enzymatic activity with the peptide under study. The leucine zipper moiety of the human factor c-Jun as well as the entire c-Jun protein are also shown to catalyze degradation of RNA. The presented data, which was obtained in the test-tube experiments, adds GCN4 and c-Jun to the pool of proteins with multiple functions (also known as moonlighting proteins. If expressed in vivo, the endoribonuclease activity of these bZIP-containing factors may represent a direct coupling between transcription activation and controlled RNA turnover. As an additional result of this work, the retro-leucine zipper of GCN4 can be added to the list of functional retro-peptides.

Whi7 is an unstable cell-cycle repressor of the Start transcriptional program.

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Gomar-Alba, Mercè; Méndez, Ester; Quilis, Inma; Bañó, M Carmen; Igual, J Carlos

2017-08-24

Start is the main decision point in eukaryotic cell cycle in which cells commit to a new round of cell division. It involves the irreversible activation of a transcriptional program by G1 CDK-cyclin complexes through the inactivation of Start transcriptional repressors, Whi5 in yeast or Rb in mammals. Here we provide novel keys of how Whi7, a protein related at sequence level to Whi5, represses Start. Whi7 is an unstable protein, degraded by the SCF Grr1 ubiquitin-ligase, whose stability is cell cycle regulated by CDK1 phosphorylation. Importantly, Whi7 associates to G1/S gene promoters in late G1 acting as a repressor of SBF-dependent transcription. Our results demonstrate that Whi7 is a genuine paralog of Whi5. In fact, both proteins collaborate in Start repression bringing to light that yeast cells, as occurs in mammalian cells, rely on the combined action of multiple transcriptional repressors to block Start transition.The commitment of cells to a new cycle of division involves inactivation of the Start transcriptional repressor Whi5. Here the authors show that the sequence related protein Whi7 associates to G1/S gene promoters in late G1 and collaborates with Whi5 in Start repression.

Hierarchy of stroma-derived factors in supporting growth of stroma-dependent hemopoietic cells: membrane-bound SCF is sufficient to confer stroma competence to epithelial cells.

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Friel, Jutta; Itoh, Katsuhiko; Bergholz, Ulla; Jücker, Manfred; Stocking, Carol; Harrison, Paul; Ostertag, Wolfram

2002-03-01

Hemopoiesis takes place in a microenvironment where hemopoietic cells are closely associated with stroma by various interactions. Stroma coregulates the proliferation and differentiation of hemopoietic cells. Stroma-hemopoietic-cell contact can be supported by locally produced membrane associated growth factors. The stroma derived growth factor, stem cell factor (SCF) is important in hemopoiesis. We examined the different biological interactions of membrane bound and soluble SCF with human hemopoietic cells expressing the SCF receptor, c-kit. To analyze the function of the SCF isoforms in inducing the proliferation of hemopoietic TF1 or Cord blood (CB) CD34+ cells we used stroma cell lines that differ in their presentation of no SCF, membrane SCF, or soluble SCF. We established a new coculture system using an epithelial cell line that excludes potential interfering effects with other known stroma encoded hemopoietic growth factors. We show that soluble SCF, in absence of membrane-bound SCF, inhibits long term clonal growth of primary or established CD34+ hemopoietic cells, whereas membrane-inserted SCF "dominantly" induces long term proliferation of these cells. We demonstrate a hierarchy of these SCF isoforms in the interaction of stroma with hemopoietic TF1 cells. Membrane-bound SCF is "dominant" over soluble SCF, whereas soluble SCF acts epistatically in interacting with hemopoietic cells compared with other stroma derived factors present in SCF deficient stroma. A hierarchy of stroma cell lines can be arranged according to their presentation of membrane SCF or soluble SCF. In our model system, membrane-bound SCF expression is sufficient to confer stroma properties to an epithelial cell line but soluble SCF does not.

Computational chemistry with transputers: A direct SCF program

International Nuclear Information System (INIS)

Wedig, U.; Burkhardt, A.; Schnering, H.G. von

1989-01-01

By using transputers it is possible to build up networks of parallel processors with varying topology. Due to the architecture of the processors it is appropriate to use the MIMD (multiple instruction multiple data) concept of parallel computing. The most suitable programming language is OCCAM. We investigate the use of transputer networks in computational chemistry, starting with the direct SCF method. The most time consuming step, the calculation of the two electron integrals is executed parallelly. Each node in the network calculates whole batches of integrals. The main program is written in OCCAM. For some large-scale arithmetic processes running on a single node, however, we used FORTRAN subroutines out of standard ab-initio programs to reduce the programming effort. Test calculations show, that the integral calculation step can be parallelled very efficiently. We observed a speed-up of almost 8 using eight network processors. Even in consideration of the scalar part of the SCF iteration, the speed-up is not less than 7.1. (orig.)

Isolation of three B-box zinc finger proteins that interact with STF1 and COP1 defines a HY5/COP1 interaction network involved in light control of development in soybean

International Nuclear Information System (INIS)

Shin, Su Young; Kim, Seong Hee; Kim, Hye Jin; Jeon, Su Jeong; Sim, Soon Ae; Ryu, Gyeong Ryul; Yoo, Cheol Min; Cheong, Yong Hwa; Hong, Jong Chan

2016-01-01

LONG HYPOCOTYL5 (HY5) and STF1 (Soybean TGACG-motif binding Factor 1) are two related bZIP transcription factors that play a positive role in photomorphogenesis and hormonal signaling. In this study, we compared full length STF1 and truncated STF1 overexpression lines and found that the C-terminal 133 amino acids (194–306) possess all the HY5-like function in Arabidopsis. The STF1-DC1 mutant (1–306), with a 20 amino acid deletion at the carboxy terminus, failed to complement the hy5 mutant phenotype, which suggests an intact C-terminus is required for STF1 function. To understand the role of the C-terminal domain in photomorphogenesis we used a yeast two-hybrid screen to isolate proteins that bind to the STF1 C-terminus. We isolated three soybean cDNAs encoding the zinc-finger proteins GmSTO, GmSTH, and GmSTH2, which interact with STF1. These proteins belong to a family of B-box zinc finger proteins that include Arabidopsis SALT TOLERANCE (STO) and STO HOMOLOG (STH) and STH2, which play a role in light-dependent development and gene expression. The C-terminal 63 amino acids of STF1, containing a leucine zipper and the two N-terminal B-boxes, contains the domain involved in interactions between STF1 and GmSTO. In addition, we identified an interaction between soybean COP1 (GmCOP1) and GmSTO and GmSTH, as well as STF1, which strongly suggests the presence of a similar regulatory circuit for light signaling in soybean as in Arabidopsis. This study shows that photomorphogenic control requires complex molecular interactions among several different classes of transcription factors such as bZIP, B-box factors, and COP1, a ubiquitin ligase. - Highlights: • STF1 interact with GmSTO, GmSTH and GmSTH2. • The bZIP transcription factor STF1 requires an intact C-terminal domain for STF1 function. • STF1 and GmSTO are nuclear proteins.

Aureochrome 1 illuminated: structural changes of a transcription factor probed by molecular spectroscopy.

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Silke Kerruth

Full Text Available Aureochrome 1 from Vaucheria frigida is a recently identified blue-light receptor that acts as a transcription factor. The protein comprises a photosensitive light-, oxygen- and voltage-sensitive (LOV domain and a basic zipper (bZIP domain that binds DNA rendering aureochrome 1 a prospective optogenetic tool. Here, we studied the photoreaction of full-length aureochrome 1 by molecular spectroscopy. The kinetics of the decay of the red-shifted triplet state and the blue-shifted signaling state were determined by time-resolved UV/Vis spectroscopy. It is shown that the presence of the bZIP domain further prolongs the lifetime of the LOV390 signaling state in comparison to the isolated LOV domain whereas bound DNA does not influence the photocycle kinetics. The light-dark Fourier transform infrared (FTIR difference spectrum shows the characteristic features of the flavin mononucleotide chromophore except that the S-H stretching vibration of cysteine 254, which is involved in the formation of the thio-adduct state, is significantly shifted to lower frequencies compared to other LOV domains. The presence of the target DNA influences the light-induced FTIR difference spectrum of aureochrome 1. Vibrational bands that can be assigned to arginine and lysine side chains as well to the phosphate backbone, indicate crucial changes in interactions between transcription factor and DNA.

Resveratrol Improved the Progression of Chronic Prostatitis via the Downregulation of c-kit/SCF by Activating Sirt1.

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He, Yi; Zeng, Huizhi; Yu, Yang; Zhang, Jiashu; Zeng, Xiaona; Gong, Fengtao; Duan, Xingping; Liu, Qi; Yang, Bo

2017-07-19

The regulation mechanism of inflammation inducing prostate carcinogenesis remains largely unknown. Therefore, we investigated the role of the c-kit/SCF pathway, which has been associated with the control of prostate carcinogenesis, in chronic prostatitis (CP) rats and evaluated the anti-prostatitis effect of resveratrol. We performed hemolysin and eosin staining to evaluate the histopathological changes in prostates. Multiple approaches evaluated the expression levels of c-kit, stem cell factor (SCF), Sirt1, and carcinogenesis-associated proteins. The CP group exhibited severe diffuse chronic inflammation. Meanwhile, the prostate cells appeared atypia; the activity of c-kit/SCF was upregulated, and carcinogenesis-associated proteins are dysregulated significantly in CP rats. Resveratrol treatment significantly improved these factors by Sirt1 activation. In summary, CP could further cause prostate carcinogenesis, which may be associated with activated c-kit/SCF signaling. Resveratrol treatment could improve the progression of CP via the downregulation of c-kit/SCF by activating Sirt1.

The AT-Hook motif as a versatile minor groove anchor for promoting DNA binding of transcription factor fragments? ?Electronic supplementary information (ESI) available: Peptide synthesis, full experimental procedures and analytical data of the peptides and products obtained. See DOI: 10.1039/c5sc01415h Click here for additional data file.

OpenAIRE

Rodr?guez, J?ssica; Mosquera, Jes?s; Couceiro, Jose R.; V?zquez, M. Eugenio; Mascare?as, Jos? L.

2015-01-01

We report the development of chimeric DNA binding peptides comprising a DNA binding fragment of natural transcription factors (the basic region of a bZIP protein or a monomeric zinc finger module) and an AT-Hook peptide motif. The resulting peptide conjugates display high DNA affinity and excellent sequence selectivity. Furthermore, the AT-Hook motif also favors the cell internalization of the conjugates.

Transcription Factor Functional Protein-Protein Interactions in Plant Defense Responses

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Murilo S. Alves

2014-03-01

Full Text Available Responses to biotic stress in plants lead to dramatic reprogramming of gene expression, favoring stress responses at the expense of normal cellular functions. Transcription factors are master regulators of gene expression at the transcriptional level, and controlling the activity of these factors alters the transcriptome of the plant, leading to metabolic and phenotypic changes in response to stress. The functional analysis of interactions between transcription factors and other proteins is very important for elucidating the role of these transcriptional regulators in different signaling cascades. In this review, we present an overview of protein-protein interactions for the six major families of transcription factors involved in plant defense: basic leucine zipper containing domain proteins (bZIP, amino-acid sequence WRKYGQK (WRKY, myelocytomatosis related proteins (MYC, myeloblastosis related proteins (MYB, APETALA2/ ETHYLENE-RESPONSIVE ELEMENT BINDING FACTORS (AP2/EREBP and no apical meristem (NAM, Arabidopsis transcription activation factor (ATAF, and cup-shaped cotyledon (CUC (NAC. We describe the interaction partners of these transcription factors as molecular responses during pathogen attack and the key components of signal transduction pathways that take place during plant defense responses. These interactions determine the activation or repression of response pathways and are crucial to understanding the regulatory networks that modulate plant defense responses.

The dynamic changes of DNA methylation and histone modifications of salt responsive transcription factor genes in soybean.

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Yuguang Song

Full Text Available Epigenetic modification contributes to the regulation of gene expression and plant development under salinity stress. Here we describe the identification of 49 soybean transcription factors by microarray analysis as being inducible by salinity stress. A semi-quantitative RT-PCR-based expression assay confirmed the salinity stress inducibility of 45 of these 49 transcription factors, and showed that ten of them were up-regulated when seedlings were exposed to the demethylation agent 5-aza-2-deoxycytidine. Salinity stress was shown to affect the methylation status of four of these ten transcription factors (one MYB, one b-ZIP and two AP2/DREB family members using a combination of bisulfite sequencing and DNA methylation-sensitive DNA gel blot analysis. ChIP analysis indicated that the activation of three of the four DNA methylated transcription factors was correlated with an increased level of histone H3K4 trimethylation and H3K9 acetylation, and/or a reduced level of H3K9 demethylation in various parts of the promoter or coding regions. Our results suggest a critical role for some transcription factors' activation/repression by DNA methylation and/or histone modifications in soybean tolerance to salinity stress.

Cyclin D3 interacts with human activating transcription factor 5 and potentiates its transcription activity

International Nuclear Information System (INIS)

Liu Wenjin; Sun Maoyun; Jiang Jianhai; Shen Xiaoyun; Sun Qing; Liu Weicheng; Shen Hailian; Gu Jianxin

2004-01-01

The Cyclin D3 protein is a member of the D-type cyclins. Besides serving as cell cycle regulators, D-type cyclins have been reported to be able to interact with several transcription factors and modulate their transcriptional activations. Here we report that human activating transcription factor 5 (hATF5) is a new interacting partner of Cyclin D3. The interaction was confirmed by in vivo coimmunoprecipitation and in vitro binding analysis. Neither interaction between Cyclin D1 and hATF5 nor interaction between Cyclin D2 and hATF5 was observed. Confocal microscopy analysis showed that Cyclin D3 could colocalize with hATF5 in the nuclear region. Cyclin D3 could potentiate hATF5 transcriptional activity independently of its Cdk4 partner. But Cyclin D1 and Cyclin D2 had no effect on hATF5 transcriptional activity. These data provide a new clue to understand the new role of Cyclin D3 as a transcriptional regulator

MNF, an ankyrin repeat protein of myxoma virus, is part of a native cellular SCF complex during viral infection

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Gelfi Jacqueline

2010-03-01

Full Text Available Abstract Myxoma virus (MYXV, a member of the Poxviridae family, is the agent responsible for myxomatosis, a fatal disease in the European rabbit (Oryctolagus cuniculus. Like all poxviruses, MYXV is known for encoding multiple proteins that regulate cellular signaling pathways. Among them, four proteins share the same ANK/PRANC structure: M148R, M149R, MNF (Myxoma Nuclear factor and M-T5, all of them described as virulence factors. This family of poxvirus proteins, recently identified, has drawn considerable attention for its potential role in modulating the host ubiquitin-proteasome system during viral infection. To date, many members of this novel protein family have been shown to interact with SCF components, in vitro. Here, we focus on MNF gene, which has been shown to express a nuclear protein presenting nine ANK repeats, one of which has been identified as a nuclear localization signal. In transfection, MNF has been shown to colocalise with the transcription factor NF-κB in the nucleus of TNFα-stimulated cells. Functionally, MNF is a critical virulence factor since its deletion generates an almost apathogenic virus. In this study, to pursue the investigation of proteins interacting with MNF and of its mechanism of action, we engineered a recombinant MYXV expressing a GFP-linked MNF under the control of MNF native promoter. Infection of rabbits with MYXV-GFPMNF recombinant virus provided the evidence that the GFP fusion does not disturb the main function of MNF. Hence, cells were infected with MYXV-GFPMNF and immunoprecipitation of the GFPMNF fusion protein was performed to identify MNF's partners. For the first time, endogenous components of SCF (Cullin-1 and Skp1 were co-precipitated with an ANK myxoma virus protein, expressed in an infectious context, and without over-expression of any protein.

MNF, an ankyrin repeat protein of myxoma virus, is part of a native cellular SCF complex during viral infection

Science.gov (United States)

2010-01-01

Myxoma virus (MYXV), a member of the Poxviridae family, is the agent responsible for myxomatosis, a fatal disease in the European rabbit (Oryctolagus cuniculus). Like all poxviruses, MYXV is known for encoding multiple proteins that regulate cellular signaling pathways. Among them, four proteins share the same ANK/PRANC structure: M148R, M149R, MNF (Myxoma Nuclear factor) and M-T5, all of them described as virulence factors. This family of poxvirus proteins, recently identified, has drawn considerable attention for its potential role in modulating the host ubiquitin-proteasome system during viral infection. To date, many members of this novel protein family have been shown to interact with SCF components, in vitro. Here, we focus on MNF gene, which has been shown to express a nuclear protein presenting nine ANK repeats, one of which has been identified as a nuclear localization signal. In transfection, MNF has been shown to colocalise with the transcription factor NF-κB in the nucleus of TNFα-stimulated cells. Functionally, MNF is a critical virulence factor since its deletion generates an almost apathogenic virus. In this study, to pursue the investigation of proteins interacting with MNF and of its mechanism of action, we engineered a recombinant MYXV expressing a GFP-linked MNF under the control of MNF native promoter. Infection of rabbits with MYXV-GFPMNF recombinant virus provided the evidence that the GFP fusion does not disturb the main function of MNF. Hence, cells were infected with MYXV-GFPMNF and immunoprecipitation of the GFPMNF fusion protein was performed to identify MNF's partners. For the first time, endogenous components of SCF (Cullin-1 and Skp1) were co-precipitated with an ANK myxoma virus protein, expressed in an infectious context, and without over-expression of any protein. PMID:20211013

Overexpression of the transcription factor NF-YC9 confers abscisic acid hypersensitivity in Arabidopsis.

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Bi, Chao; Ma, Yu; Wang, Xiao-Fang; Zhang, Da-Peng

2017-11-01

Nuclear factor Y (NF-Y) family proteins are involved in many developmental processes and responses to environmental cues in plants, but whether and how they regulate phytohormone abscisic acid (ABA) signaling need further studies. In the present study, we showed that over-expression of the NF-YC9 gene confers ABA hypersensitivity in both the early seedling growth and stomatal response, while down-regulation of NF-YC9 does not affect ABA response in these processes. We also showed that over-expression of the NF-YC9 gene confers salt and osmotic hypersensitivity in early seedling growth, which is likely to be directly associated with the ABA hypersensitivity. Further, we observed that NF-YC9 physically interacts with the ABA-responsive bZIP transcription factor ABA-INSENSITIVE5 (ABI5), and facilitates the function of ABI5 to bind and activate the promoter of a target gene EM6. Additionally, NF-YC9 up-regulates expression of the ABI5 gene in response to ABA. These findings show that NF-YC9 may be involved in ABA signaling as a positive regulator and likely functions redundantly together with other NF-YC members, and support the model that the NF-YC9 mediates ABA signaling via targeting to and aiding the ABA-responsive transcription factors such as ABI5.

Cyanogen Azide. Ionization Potentials and Ab Initio SCF MO Calculation

DEFF Research Database (Denmark)

Bak, Börge; Jansen, Peter; Stafast, Herbert

1975-01-01

The Ne(I) and He(I) photoelectron(PE) spectra of cyanogen azide, NCN3, have been recorded at high resolution. Their interpretation is achieved by comparison with the PE spectrum of HN3 and an ab initio LCGO SCF MO calculation. Deviations from Koopmans' theorem of quite different magnitudes...

Potential Role of Activating Transcription Factor 5 during Osteogenesis

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Luisa Vicari

2016-01-01

Full Text Available Human adipose-derived stem cells are an abundant population of stem cells readily isolated from human adipose tissue that can differentiate into connective tissue lineages including bone, cartilage, fat, and muscle. Activating transcription factor 5 is a transcription factor of the ATF/cAMP response element-binding protein (CREB family. It is transcribed in two types of mRNAs (activating transcription factor 5 isoform 1 and activating transcription factor 5 isoform 2, encoding the same single 30-kDa protein. Although it is well demonstrated that it regulates the proliferation, differentiation, and apoptosis, little is known about its potential role in osteogenic differentiation. The aim of this study was to evaluate the expression levels of the two isoforms and protein during osteogenic differentiation of human adipose-derived stem cells. Our data indicate that activating transcription factor 5 is differentially expressed reaching a peak of expression at the stage of bone mineralization. These findings suggest that activating transcription factor 5 could play an interesting regulatory role during osteogenesis, which would provide a powerful tool to study bone physiology.

Potential Role of Activating Transcription Factor 5 during Osteogenesis.

Science.gov (United States)

Vicari, Luisa; Calabrese, Giovanna; Forte, Stefano; Giuffrida, Raffaella; Colarossi, Cristina; Parrinello, Nunziatina Laura; Memeo, Lorenzo

2016-01-01

Human adipose-derived stem cells are an abundant population of stem cells readily isolated from human adipose tissue that can differentiate into connective tissue lineages including bone, cartilage, fat, and muscle. Activating transcription factor 5 is a transcription factor of the ATF/cAMP response element-binding protein (CREB) family. It is transcribed in two types of mRNAs (activating transcription factor 5 isoform 1 and activating transcription factor 5 isoform 2), encoding the same single 30-kDa protein. Although it is well demonstrated that it regulates the proliferation, differentiation, and apoptosis, little is known about its potential role in osteogenic differentiation. The aim of this study was to evaluate the expression levels of the two isoforms and protein during osteogenic differentiation of human adipose-derived stem cells. Our data indicate that activating transcription factor 5 is differentially expressed reaching a peak of expression at the stage of bone mineralization. These findings suggest that activating transcription factor 5 could play an interesting regulatory role during osteogenesis, which would provide a powerful tool to study bone physiology.

TNF-α inhibits SCF, ghrelin, and substance P expressions through the NF-κB pathway activation in interstitial cells of Cajal.

Science.gov (United States)

Ren, Keyu; Yong, Chunming; Yuan, Hao; Cao, Bin; Zhao, Kun; Wang, Jin

2018-01-01

Ulcerative colitis is a chronic inflammatory disease of the colon where intestinal motility is disturbed. Interstitial cells of Cajal (ICC) are required to maintain normal intestinal motility. In the present study, we assessed the effect of tumor necrosis factor-alpha (TNF-α) on viability and apoptosis of ICC, as well as on the expression of stem cell factor (SCF), ghrelin, and substance P. ICC were derived from the small intestines of Swiss albino mice. Cell viability and apoptosis were measured using CCK-8 assay and flow cytometry, respectively. ELISA was used to measure the concentrations of IL-1β, IL-6, ghrelin, substance P, and endothelin-1. Quantitative RT-PCR was used to measure the expression of SCF. Western blotting was used to measure the expression of apoptosis-related proteins, interleukins, SCF, and NF-κB signaling pathway proteins. TNF-α induced inflammatory injury in ICC by decreasing cell viability and increasing apoptosis and levels of IL-1β and IL-6. TNF-α decreased the levels of SCF, ghrelin, and substance P, but had no effect on endothelin-1. TNF-α down-regulated expressions of SCF, ghrelin, and substance P by activating the NF-κB pathway in ICC. In conclusion, TNF-α down-regulated the expressions of SCF, ghrelin, and substance P via the activation of the NF-κB pathway in ICC.

Localized Symmetry Breaking for Tuning Thermal Expansion in ScF ₃ Nanoscale Frameworks

Energy Technology Data Exchange (ETDEWEB)

Hu, Lei [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Department of Materials Science and Engineering, Northwestern University, Evanston, Illinois 60208, United States; Qin, Feiyu [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Sanson, Andrea [Department of Physics and Astronomy, University of Padova, Padova I-35131, Italy; Huang, Liang-Feng [Department of Materials Science and Engineering, Northwestern University, Evanston, Illinois 60208, United States; Pan, Zhao [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Li, Qiang [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Sun, Qiang [International Laboratory for Quantum Functional Materials of Henan, School of Physics and Engineering, Zhengzhou University, Zhengzhou 450001, China; Wang, Lu [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Guo, Fangmin [X-Ray Science Division, Argonne National Laboratory, Argonne, Illinois 60439, United States; Aydemir, Umut [Department of Materials Science and Engineering, Northwestern University, Evanston, Illinois 60208, United States; Department of Chemistry, Koc University, Sariyer, Istanbul 34450, Turkey; Ren, Yang [X-Ray Science Division, Argonne National Laboratory, Argonne, Illinois 60439, United States; Sun, Chengjun [X-Ray Science Division, Argonne National Laboratory, Argonne, Illinois 60439, United States; Deng, Jinxia [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Aquilanti, Giuliana [Elettra Sincrotrone Trieste, Basovizza, Trieste I-34149, Italy; Rondinelli, James M. [Department of Materials Science and Engineering, Northwestern University, Evanston, Illinois 60208, United States; Chen, Jun [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China; Xing, Xianran [Department of Physical Chemistry, University of Science and Technology Beijing, Beijing 100083, China

2018-03-15

The local symmetry, beyond the averaged crystallographic structure, tends to bring unu-sual performances. Negative thermal expansion is a peculiar physical property of solids. Here, we report the delicate design of the localized symmetry breaking to achieve the controllable thermal expansion in ScF3 nano-scale frameworks. Intriguingly, an isotropic zero thermal expansion is concurrently engi-neered by localized symmetry breaking, with a remarkably low coefficient of thermal expansion of about +4.0×10-8/K up to 675K. This mechanism is investigated by the joint analysis of atomic pair dis-tribution function of synchrotron X-ray total scattering and extended X-ray absorption fine structure spectra. A localized rhombohedral distortion presumably plays a critical role in stiffening ScF3 nano-scale frameworks and concomitantly suppressing transverse thermal vibrations of fluorine atoms. This physical scenario is also theoretically corroborated by the extinction of phonon modes with negative Grüneisen parameters in the rhombohedral ScF3. The present work opens an untraditional chemical modification to achieve controllable thermal expansion by breaking local symmetries of materials.

Plant transducers of the endoplasmic reticulum unfolded protein response

KAUST Repository

Iwata, Yuji; Koizumi, Nozomu

2012-01-01

The unfolded protein response (UPR) activates a set of genes to overcome accumulation of unfolded proteins in the endoplasmic reticulum (ER), a condition termed ER stress, and constitutes an essential part of ER protein quality control that ensures efficient maturation of secretory and membrane proteins in eukaryotes. Recent studies on Arabidopsis and rice identified the signaling pathway in which the ER membrane-localized ribonuclease IRE1 (inositol-requiring enzyme 1) catalyzes unconventional cytoplasmic splicing of mRNA, thereby producing the active transcription factor Arabidopsis bZIP60 (basic leucine zipper 60) and its ortholog in rice. Here we review recent findings identifying the molecular components of the plant UPR, including IRE1/bZIP60 and the membrane-bound transcription factors bZIP17 and bZIP28, and implicating its importance in several physiological phenomena such as pathogen response. © 2012 Elsevier Ltd.

Plant transducers of the endoplasmic reticulum unfolded protein response

KAUST Repository

Iwata, Yuji

2012-12-01

The unfolded protein response (UPR) activates a set of genes to overcome accumulation of unfolded proteins in the endoplasmic reticulum (ER), a condition termed ER stress, and constitutes an essential part of ER protein quality control that ensures efficient maturation of secretory and membrane proteins in eukaryotes. Recent studies on Arabidopsis and rice identified the signaling pathway in which the ER membrane-localized ribonuclease IRE1 (inositol-requiring enzyme 1) catalyzes unconventional cytoplasmic splicing of mRNA, thereby producing the active transcription factor Arabidopsis bZIP60 (basic leucine zipper 60) and its ortholog in rice. Here we review recent findings identifying the molecular components of the plant UPR, including IRE1/bZIP60 and the membrane-bound transcription factors bZIP17 and bZIP28, and implicating its importance in several physiological phenomena such as pathogen response. © 2012 Elsevier Ltd.

Molecular Characterization of PDGFR-α/PDGF-A and c-KIT/SCF in Gliosarcomas

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Rui M. Reis

2005-01-01

Full Text Available Gliosarcomas are rare and poorly characterized malignant brain tumors that exhibit a biphasic tissue pattern with areas of gliomatous and sarcomatous differentiation. These tumors are histological variants of glioblastoma, displaying a similar genetic profile and dismal prognosis. Up-regulation of PDGFR subfamily of tyrosine kinase members, PDGFR-α and c-Kit, and their intracellular effectors RAS/RAF/MAPK has a crucial role in the cancer development. In addition, signal transduction mediated by activating mutations of c-Kit and PDGFR can be effectively blocked by specific tyrosine kinase inhibitors, such as Imatinib mesylate. The aim of this study was to characterize the molecular alterations of PDGFR signaling in gliosarcomas. Six cases were analyzed by immunohistochemistry for the expression of PDGFR-α, c-Kit and their ligands PDGF-A and SCF, respectively. The cases were further evaluated for the presence of activating mutations of PDGFR-α (exons 12 and 18 and c-kit (exons 9, 11, 13, and 17, as well as B-RAF (exons 11 and 15. Expression of PDGF-A was found in all cases and co-expression of PDGFR-α was observed in three cases. Four cases showed expression of SCF, and c-Kit was observed only in one case that also expressed SCF. Generally, immunoreaction predominates in the glial component. The mutational analysis of PDGFR-α showed the presence of an IVS17-50insT intronic insertion in two cases, one of them also with a 2472C > T silent mutation; this silent mutation was also found in another case. Glioma cell line analysis of IVS17-50insT insertion showed no influence on PDGFR-α gene splicing. No mutations were detected in c-kit and B-RAF oncogenes. Our Results indicate that activating mutations of PDGFR-α, c-kit and B-RAF are absent in gliosarcomas. Nevertheless, the presence of a PDGFR-a/PDGFA and c-Kit/SCF autocrine/paracrine stimulation loop in a proportion of cases, supports the potential role of specific tyrosine kinase inhibitors in

Local structure of perovskites ReO3 and ScF3 with negative thermal expansion: interpretation beyond the quasiharmonic approximation

International Nuclear Information System (INIS)

Purans, Juris; Piskunov, Sergei; Bocharov, Dmitry; Kalinko, Aleksandr; Kuzmin, Alexei; Ali, Shehab E.; Rocca, Francesco

2016-01-01

We propose an approach beyond the quasiharmonic approximation for interpretation of EXAFS and XRD data and for ab initio calculations of electronic and vibration properties of materials with negative thermal expansion. Ab initio electronic structure and lattice dynamics calculations for cubic and distorted ScF 3 were performed using the linear combination of atomic orbitals (LCAO) method. The band gap obtained in calculations for ScF 3 is equal to 10.54 eV and agree well with the expected value. The calculated infrared spectra of F displaced (FD) cubic ScF 3 allow us to predict that its mean Sc-F-Sc angle within NTE deviates from 180 degree. (paper)

Resveratrol Improves Cell Cycle Arrest in Chronic Prostatitis Rats, by C-kit/SCF Suppression.

Science.gov (United States)

He, Yi; Zeng, Huizhi; Yu, Yang; Zhang, Jiashu; Zeng, Xiaona; Gong, Fengtao; Liu, Qi; Yang, Bo

2017-08-01

Chronic prostatitis (CP) with complex pathogenesis is difficult for treatment. c-kit has been associated with the control of cell proliferation of prostate cells. This study aims to evaluate the role of resveratrol, an activator of Sirt1, in regulating the expression of c-kit in CP and investigate the consequent effects on cell cycle. Rat model of CP was established through subcutaneous injections of diphtheria-pertussis-tetanus vaccine and subsequently treated with resveratrol. Hematoxylin and eosin staining was performed to identify the histopathological changes in prostates. Western blotting and immunohistochemical staining examined the expression level of c-kit, stem cell factor (SCF), Sirt1, and cell cycle-associated proteins. The model group exhibited severe diffuse chronic inflammation, characterized by leukocyte infiltration and papillary frond protrusion into the gland cavities, and a notable increase in prostatic epithelial height. Gland lumen diameter was also significantly smaller; the activity of c-kit/SCF in the CP rats was increased significantly compared to the control group. Meanwhile, the cell cycle proteins are dysregulated significantly in CP rats. Resveratrol treatment significantly improved these factors by Sirt1 activation. Dysregulation of cell cycle was involved in the pathological processes of CP, which was improved after resveratrol treatment by the downregulation of c-kit/SCF by activating Sirt1.

Transcript profiling of Elf5+/- mammary glands during pregnancy identifies novel targets of Elf5.

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Renee L Rogers

Full Text Available BACKGROUND: Elf5, an epithelial specific Ets transcription factor, plays a crucial role in the pregnancy-associated development of the mouse mammary gland. Elf5(-/- embryos do not survive, however the Elf5(+/- mammary gland displays a severe pregnancy-associated developmental defect. While it is known that Elf5 is crucial for correct mammary development and lactation, the molecular mechanisms employed by Elf5 to exert its effects on the mammary gland are largely unknown. PRINCIPAL FINDINGS: Transcript profiling was used to investigate the transcriptional changes that occur as a result of Elf5 haploinsufficiency in the Elf5(+/- mouse model. We show that the development of the mouse Elf5(+/- mammary gland is delayed at a transcriptional and morphological level, due to the delayed increase in Elf5 protein in these glands. We also identify a number of potential Elf5 target genes, including Mucin 4, whose expression, is directly regulated by the binding of Elf5 to an Ets binding site within its promoter. CONCLUSION: We identify novel transcriptional targets of Elf5 and show that Muc4 is a direct target of Elf5, further elucidating the mechanisms through which Elf5 regulates proliferation and differentiation in the mammary gland.

Hijacking of the host SCF ubiquitin ligase machinery by plant pathogens

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Shimpei eMagori

2011-11-01

Full Text Available The SCF (SKP1-CUL1-F-box protein ubiquitin ligase complex mediates polyubiquitination of proteins targeted for degradation, thereby controlling a plethora of biological processes in eukaryotic cells. Although this ubiquitination machinery is found and functional only in eukaryotes, many non-eukaryotic pathogens also encode F-box proteins, the critical subunits of the SCF complex. Increasing evidence indicates that such non-eukaryotic F-box proteins play an essential role in subverting or exploiting the host ubiquitin/proteasome system for efficient pathogen infection. A recent bioinformatic analysis has identified more than 70 F-box proteins in 22 different bacterial species, suggesting that use of pathogen-encoded F-box effectors in the host cell may be a widespread infection strategy. In this review, we focus on plant pathogen-encoded F-box effectors, such as VirF of Agrobacterium tumefaciens, GALAs of Ralstonia solanacearum, and P0 of Poleroviruses, and discuss the molecular mechanism by which plant pathogens use these factors to manipulate the host cell for their own benefit.

Functional requirements document for the Earth Observing System Data and Information System (EOSDIS) Scientific Computing Facilities (SCF) of the NASA/MSFC Earth Science and Applications Division, 1992

Science.gov (United States)

Botts, Michael E.; Phillips, Ron J.; Parker, John V.; Wright, Patrick D.

1992-01-01

Five scientists at MSFC/ESAD have EOS SCF investigator status. Each SCF has unique tasks which require the establishment of a computing facility dedicated to accomplishing those tasks. A SCF Working Group was established at ESAD with the charter of defining the computing requirements of the individual SCFs and recommending options for meeting these requirements. The primary goal of the working group was to determine which computing needs can be satisfied using either shared resources or separate but compatible resources, and which needs require unique individual resources. The requirements investigated included CPU-intensive vector and scalar processing, visualization, data storage, connectivity, and I/O peripherals. A review of computer industry directions and a market survey of computing hardware provided information regarding important industry standards and candidate computing platforms. It was determined that the total SCF computing requirements might be most effectively met using a hierarchy consisting of shared and individual resources. This hierarchy is composed of five major system types: (1) a supercomputer class vector processor; (2) a high-end scalar multiprocessor workstation; (3) a file server; (4) a few medium- to high-end visualization workstations; and (5) several low- to medium-range personal graphics workstations. Specific recommendations for meeting the needs of each of these types are presented.

SCF Ubiquitin Ligase F-box Protein Fbx15 Controls Nuclear Co-repressor Localization, Stress Response and Virulence of the Human Pathogen Aspergillus fumigatus.

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Bastian Jöhnk

2016-09-01

Full Text Available F-box proteins share the F-box domain to connect substrates of E3 SCF ubiquitin RING ligases through the adaptor Skp1/A to Cul1/A scaffolds. F-box protein Fbx15 is part of the general stress response of the human pathogenic mold Aspergillus fumigatus. Oxidative stress induces a transient peak of fbx15 expression, resulting in 3x elevated Fbx15 protein levels. During non-stress conditions Fbx15 is phosphorylated and F-box mediated interaction with SkpA preferentially happens in smaller subpopulations in the cytoplasm. The F-box of Fbx15 is required for an appropriate oxidative stress response, which results in rapid dephosphorylation of Fbx15 and a shift of the cellular interaction with SkpA to the nucleus. Fbx15 binds SsnF/Ssn6 as part of the RcoA/Tup1-SsnF/Ssn6 co-repressor and is required for its correct nuclear localization. Dephosphorylated Fbx15 prevents SsnF/Ssn6 nuclear localization and results in the derepression of gliotoxin gene expression. fbx15 deletion mutants are unable to infect immunocompromised mice in a model for invasive aspergillosis. Fbx15 has a novel dual molecular function by controlling transcriptional repression and being part of SCF E3 ubiquitin ligases, which is essential for stress response, gliotoxin production and virulence in the opportunistic human pathogen A. fumigatus.

Characterisation of Cdkl5 transcript isoforms in rat.

Science.gov (United States)

Hector, Ralph D; Dando, Owen; Ritakari, Tuula E; Kind, Peter C; Bailey, Mark E S; Cobb, Stuart R

2017-03-01

CDKL5 deficiency is a severe neurological disorder caused by mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5). The predominant human CDKL5 brain isoform is a 9.7kb transcript comprised of 18 exons with a large 6.6kb 3'-untranslated region (UTR). Mammalian models of CDKL5 disorder are currently limited to mouse, and little is known about Cdkl5 in other organisms used to model neurodevelopmental disorders, such as rat. In this study we characterise, both bioinformatically and experimentally, the rat Cdkl5 gene structure and its associated transcript isoforms. New exonic regions, splice sites and UTRs are described, confirming the presence of four distinct transcript isoforms. The predominant isoform in the brain, which we name rCdkl5_1, is orthologous to the human hCDKL5_1 and mouse mCdkl5_1 isoforms and is the most highly expressed isoform across all brain regions tested. This updated gene model of Cdkl5 in rat provides a framework for studies into its protein products and provides a reference for the development of molecular therapies for testing in rat models of CDKL5 disorder. Copyright © 2016 Elsevier B.V. All rights reserved.

5' diversity of human hepatic PXR (NR1I2) transcripts and identification of the major transcription initiation site.

Science.gov (United States)

Kurose, Kouichi; Koyano, Satoru; Ikeda, Shinobu; Tohkin, Masahiro; Hasegawa, Ryuichi; Sawada, Jun-Ichi

2005-05-01

The human pregnane X receptor (PXR) is a crucial regulator of the genes encoding several major cytochrome P450 enzymes and transporters, such as CYP3A4 and MDR1, but its own transcriptional regulation remains unclear. To elucidate the transcriptional mechanisms of human PXR gene, we first endeavored to identify the transcription initiation site of human PXR using 5'-RACE. Five types of 5'-variable transcripts (a, b, c, d, and e) with common exon 2 sequence were found, and comparison of these sequences with the genomic sequence suggested that their 5' diversity is derived from initiation by alternative promoters and alternative splicing. None of the exons found in our study contain any new in-frame coding regions. Newly identified introns IVS-a and IVS-b were found to have CT-AC splice sites that do not follow the GT-AG rule of conventional donor and acceptor splice sites. Of the five types of 5' variable transcripts identified, RT-PCR showed that type-a was the major transcript type. Four transcription initiation sites (A-D) for type-a transcript were identified by 5'-RACE using GeneRacer RACE Ready cDNA (human liver) constructed by the oligo-capping method. Putative TATA boxes were located approximately 30 bp upstream from the transcriptional start sites of the major transcript (C) and the longest minor transcript (A) expressed in the human liver. These results indicate that the initiation of transcription of human PXR is more complex than previously reported.

Characterisation of CDKL5 Transcript Isoforms in Human and Mouse.

Science.gov (United States)

Hector, Ralph D; Dando, Owen; Landsberger, Nicoletta; Kilstrup-Nielsen, Charlotte; Kind, Peter C; Bailey, Mark E S; Cobb, Stuart R

2016-01-01

Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5) cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3'-untranslated region (UTR), which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders.

RHFPPP, SCF-LCAO-MO Calculation for Closed Shell and Open Shell Organic Molecules

International Nuclear Information System (INIS)

Bieber, A.; Andre, J.J.

1987-01-01

1 - Nature of physical problem solved: Complete program performs SCF-LCAO-MO calculations for both closed and open-shell organic pi-molecules. The Pariser-Parr-People approximations are used with- in the framework of the restricted Hartree-Fock method. The SCF calculation is followed, if desired, by a variational configuration interaction (CI) calculation including singly excited configurations. 2 - Method of solution: A standard procedure is used; at each step a real symmetric matrix has to be diagonalized. The self-consistency is checked by comparing the eigenvectors between two consecutive steps. 3 - Restrictions on the complexity of the problem: i) The calculations are restricted to planar molecules. ii) In order to avoid accumulation of round-off errors, in the iterative procedure, double precision arithmetic is used. iii) The program is restricted to systems up to about 16 atoms; however the size of the systems can easily be modified if required

Transcriptomic profiling of Arabidopsis gene expression in response to varying micronutrient zinc supply

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Herlânder Azevedo

2016-03-01

Full Text Available Deficiency of the micronutrient zinc is a widespread condition in agricultural soils, causing a negative impact on crop quality and yield. Nevertheless, there is an insufficient knowledge on the regulatory and molecular mechanisms underlying the plant response to inadequate zinc nutrition [1]. This information should contribute to the development of plant-based solutions with improved nutrient-use-efficiency traits in crops. Previously, the transcription factors bZIP19 and bZIP23 were identified as essential regulators of the response to zinc deficiency in Arabidopsis thaliana [2]. A microarray experiment comparing gene expression between roots of wild-type and the mutant bzip19 bzip23, exposed to zinc deficiency, led to the identification of differentially expressed genes related with zinc homeostasis, namely its transport and plant internal translocation [2]. Here, we provide the detailed methodology, bioinformatics analysis and quality controls related to the microarray gene expression profiling published by Assunção and co-workers [2]. Most significantly, the present dataset comprises new experimental variables, including analysis of shoot tissue, and zinc sufficiency and excess supply. Thus, it expands from 8 to 42 microarrays hybridizations, which have been deposited at the Gene Expression Omnibus (GEO under the accession number GSE77286. Overall, it provides a resource for research on the molecular basis and regulatory events of the plant response to zinc supply, emphasizing the importance of Arabidopsis bZIP19 and bZIP23 transcription factors. Keywords: Microarray, Micronutrient, Zinc deficiency, Arabidopsis, bZIP

ABF2, an ABRE-binding bZIP factor, is an essential component of glucose signaling and its overexpression affects multiple stress tolerance.

Science.gov (United States)

Kim, Sunmi; Kang, Jung-Youn; Cho, Dong-Im; Park, Ji Hye; Kim, Soo Young

2004-10-01

Phytohormone abscisic acid (ABA) regulates stress-responsive gene expression during vegetative growth, which is mediated largely by cis-elements sharing the ACGTGGC consensus. Although many transcription factors are known to bind the elements in vitro, only a few have been demonstrated to have in vivo functions and their specific roles in ABA/stress responses are mostly unknown. Here, we report that ABF2, an ABF subfamily member of bZIP proteins interacting with the ABA-responsive elements, is involved in ABA/stress responses. Its overexpression altered ABA sensitivity, dehydration tolerance, and the expression levels of ABA/stress-regulated genes. Furthermore, ABF2 overexpression promoted glucose-induced inhibition of seedling development, whereas its mutation impaired glucose response. The reduced sugar sensitivity was not observed with mutants of two other ABF family members, ABF3 and ABF4. Instead, these mutants displayed defects in ABA, salt, and dehydration responses, which were not observed with the abf2 mutant. Our data indicate distinct roles of ABF family members: whereas ABF3 and ABF4 play essential roles in ABA/stress responses, ABF2 is required for normal glucose response. We also show that ABF2 overexpression affects multiple stress tolerance.

Characterisation of CDKL5 Transcript Isoforms in Human and Mouse.

Directory of Open Access Journals (Sweden)

Ralph D Hector

Full Text Available Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5 cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3'-untranslated region (UTR, which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders.

MLL-ENL cooperates with SCF to transform primary avian multipotent cells.

Science.gov (United States)

Schulte, Cathleen E; von Lindern, Marieke; Steinlein, Peter; Beug, Hartmut; Wiedemann, Leanne M

2002-08-15

The MLL gene is targeted by chromosomal translocations, which give rise to heterologous MLL fusion proteins and are associated with distinct types of acute lymphoid and myeloid leukaemia. To determine how MLL fusion proteins alter the proliferation and/or differentiation of primary haematopoietic progenitors, we introduced the MLL-AF9 and MLL-ENL fusion proteins into primary chicken bone marrow cells. Both fusion proteins caused the sustained outgrowth of immature haematopoietic cells, which was strictly dependent on stem cell factor (SCF). The renewing cells have a long in vitro lifespan exceeding the Hayflick limit of avian cells. Analysis of clonal cultures identified the renewing cells as immature, multipotent progenitors, expressing erythroid, myeloid, lymphoid and stem cell surface markers. Employing a two-step commitment/differentiation protocol involving the controlled withdrawal of SCF, the MLL-ENL-transformed progenitors could be induced to terminal erythroid or myeloid differentiation. Finally, in cooperation with the weakly leukaemogenic receptor tyrosine kinase v-Sea, the MLL-ENL fusion protein gave rise to multilineage leukaemia in chicks, suggesting that other activated, receptor tyrosine kinases can substitute for ligand-activated c-Kit in vivo.

Multiple promoters and alternative splicing: Hoxa5 transcriptional complexity in the mouse embryo.

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Yan Coulombe

2010-05-01

Full Text Available The genomic organization of Hox clusters is fundamental for the precise spatio-temporal regulation and the function of each Hox gene, and hence for correct embryo patterning. Multiple overlapping transcriptional units exist at the Hoxa5 locus reflecting the complexity of Hox clustering: a major form of 1.8 kb corresponding to the two characterized exons of the gene and polyadenylated RNA species of 5.0, 9.5 and 11.0 kb. This transcriptional intricacy raises the question of the involvement of the larger transcripts in Hox function and regulation.We have undertaken the molecular characterization of the Hoxa5 larger transcripts. They initiate from two highly conserved distal promoters, one corresponding to the putative Hoxa6 promoter, and a second located nearby Hoxa7. Alternative splicing is also involved in the generation of the different transcripts. No functional polyadenylation sequence was found at the Hoxa6 locus and all larger transcripts use the polyadenylation site of the Hoxa5 gene. Some larger transcripts are potential Hoxa6/Hoxa5 bicistronic units. However, even though all transcripts could produce the genuine 270 a.a. HOXA5 protein, only the 1.8 kb form is translated into the protein, indicative of its essential role in Hoxa5 gene function. The Hoxa6 mutation disrupts the larger transcripts without major phenotypic impact on axial specification in their expression domain. However, Hoxa5-like skeletal anomalies are observed in Hoxa6 mutants and these defects can be explained by the loss of expression of the 1.8 kb transcript. Our data raise the possibility that the larger transcripts may be involved in Hoxa5 gene regulation.Our observation that the Hoxa5 larger transcripts possess a developmentally-regulated expression combined to the increasing sum of data on the role of long noncoding RNAs in transcriptional regulation suggest that the Hoxa5 larger transcripts may participate in the control of Hox gene expression.

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

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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.

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.

Localization of RNA transcription sites in insect oocytes using microinjections of 5-bromouridine 5'-triphosphate.

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Dmitry Bogolyubov

2007-06-01

Full Text Available In the present study we used 5-bromouridine 5'-triphosphate (BrUTP microinjections to localize the transcription sites in oocytes of insects with different types of the ovarium structure: panoistic, meroistic polytrophic, and meroistic telotrophic. We found that in an insect with panoistic ovaries (Acheta domesticus, oocyte nuclei maintain their transcription activity during the long period of oocyte growth. In insects with meroistic ovaries (Tenebrio molitor and Panorpa communis, early oocyte chromosomes were found to be transcriptionally active, and some transcription activity still persist while the karyosphere, a compact structure formed by all condensed oocyte chromosomes, begins to develop. At the latest stages of karyosphere development, no anti-Br-RNA signal was registered in the karyosphere.

RNA-Seq for enrichment and analysis of IRF5 transcript expression in SLE.

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Rivka C Stone

Full Text Available Polymorphisms in the interferon regulatory factor 5 (IRF5 gene have been consistently replicated and shown to confer risk for or protection from the development of systemic lupus erythematosus (SLE. IRF5 expression is significantly upregulated in SLE patients and upregulation associates with IRF5-SLE risk haplotypes. IRF5 alternative splicing has also been shown to be elevated in SLE patients. Given that human IRF5 exists as multiple alternatively spliced transcripts with distinct function(s, it is important to determine whether the IRF5 transcript profile expressed in healthy donor immune cells is different from that expressed in SLE patients. Moreover, it is not currently known whether an IRF5-SLE risk haplotype defines the profile of IRF5 transcripts expressed. Using standard molecular cloning techniques, we identified and isolated 14 new differentially spliced IRF5 transcript variants from purified monocytes of healthy donors and SLE patients to generate an IRF5 variant transcriptome. Next-generation sequencing was then used to perform in-depth and quantitative analysis of full-length IRF5 transcript expression in primary immune cells of SLE patients and healthy donors by next-generation sequencing. Evidence for additional alternatively spliced transcripts was obtained from de novo junction discovery. Data from these studies support the overall complexity of IRF5 alternative splicing in SLE. Results from next-generation sequencing correlated with cloning and gave similar abundance rankings in SLE patients thus supporting the use of this new technology for in-depth single gene transcript profiling. Results from this study provide the first proof that 1 SLE patients express an IRF5 transcript signature that is distinct from healthy donors, 2 an IRF5-SLE risk haplotype defines the top four most abundant IRF5 transcripts expressed in SLE patients, and 3 an IRF5 transcript signature enables clustering of SLE patients with the H2 risk haplotype.

The Bphi008a gene interacts with the ethylene pathway and transcriptionally regulates MAPK genes in the response of rice to brown planthopper feeding.

Science.gov (United States)

Hu, Jing; Zhou, Jiangbo; Peng, Xinxin; Xu, Henghao; Liu, Caixiang; Du, Bo; Yuan, Hongyu; Zhu, Lili; He, Guangcun

2011-06-01

We examined ways in which the Brown planthopper induced008a (Bphi008a; AY256682) gene of rice (Oryza sativa) enhances the plant's resistance to a specialist herbivore, the brown planthopper (BPH; Nilaparvata lugens). Measurement of the expression levels of ethylene synthases and of ethylene emissions showed that BPH feeding rapidly initiated the ethylene signaling pathway and up-regulated Bphi008a transcript levels after 6 to 96 h of feeding. In contrast, blocking ethylene transduction (using 1-methylcyclopropene) reduced Bphi008a transcript levels in wild-type plants fed upon by BPH. In vitro kinase assays showed that Bphi008a can be phosphorylated by rice Mitogen-activated Protein Kinase5 (OsMPK5), and yeast two-hybrid assays demonstrated that the carboxyl-terminal proline-rich region of Bphi008a interacts directly with this kinase. Furthermore, bimolecular fluorescence complementation assays showed that this interaction occurs in the nucleus. Subsequently, we found that Bphi008a up-regulation and down-regulation were accompanied by different changes in transcription levels of OsMPK5, OsMPK12, OsMPK13, and OsMPK17 in transgenic plants. Immunoblot analysis also showed that the OsMPK5 protein level increased in overexpressing plants and decreased in RNA interference plants after BPH feeding. In transgenic lines, changes in the expression levels of several enzymes that are important components of the defenses against the BPH were also observed. Finally, yeast two-hybrid screening results showed that Bphi008a is able to interact with a b-ZIP transcription factor (OsbZIP60) and a RNA polymerase polypeptide (SDRP).

Identification of She3 as an SCF(Grr1 substrate in budding yeast.

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Ruiwen Wang

Full Text Available The highly orchestrated progression of the cell cycle depends on the degradation of many regulatory proteins at different cell cycle stages. One of the key cell cycle ubiquitin ligases is the Skp1-cullin-F-box (SCF complex. Acting in concert with the substrate-binding F-box protein Grr1, SCF(Grr1 promotes the degradation of cell cycle regulators as well as various metabolic enzymes. Using a yeast two-hybrid assay with a Grr1 derivative as the bait, we identified She3, which is an adaptor protein in the asymmetric mRNA transport system, as a novel Grr1 substrate. We generated stabilized She3 mutants, which no longer bound to Grr1, and found that the degradation of She3 is not required for regulating asymmetric mRNA transport. However, She3 stabilization leads to slower growth compared to wild-type cells in a co-culture assay, demonstrating that the degradation of She3 by Grr1 is required for optimal cell growth.

Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

Science.gov (United States)

In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

1997-03-01

Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation.

SCF^{Cyclin F}-dependent degradation of CDC6 suppresses DNA re-replication

DEFF Research Database (Denmark)

Walter, David; Hoffmann, Saskia; Komseli, Eirini-Stavroula

2016-01-01

interact through defined sequence motifs that promote CDC6 ubiquitylation and degradation. Absence of Cyclin F or expression of a stable mutant of CDC6 promotes re-replication and genome instability in cells lacking the CDT1 inhibitor Geminin. Together, our work reveals a novel SCF(Cyclin F...

Encapsulation of a trinuclear silver(I) cluster by two imido-nitrido metalloligands [{Ti(eta5-C5Me5)(micro-NH)}3(micro3-N)].

Science.gov (United States)

Martín, Avelino; Martínez-Espada, Noelia; Mena, Miguel; Yélamos, Carlos

2007-07-28

Treatment of the metalloligand [{Ti(eta(5)-C(5)Me(5))(micro-NH)}(3)(micro(3)-N)] with silver(i) trifluoromethanesulfonate in different molar ratios gives the ionic compounds [Ag{(micro(3)-NH)(3)Ti(3)(eta(5)-C(5)Me(5))(3)(micro(3)-N)}(2)][O(3)SCF(3)] and [Ag{(micro(3)-NH)(3)Ti(3)(eta(5)-C(5)Me(5))(3)(micro(3)-N)}][O(3)SCF(3)] or the triangular silver cluster [(CF(3)SO(2)O)(3)Ag(3){(micro(3)-NH)(3)Ti(3)(eta(5)-C(5)Me(5))(3)(micro(3)-N)}(2)] in which each face is capped by a metalloligand.

The bZip transscription factor HY5 mediates CRY1a-induced anthocyanin biosynthesis in tomato.

Science.gov (United States)

Liu, Chao-Chao; Chi, Cheng; Jin, Li-Juan; Zhu, Jianhua; Yu, Jing-Quan; Zhou, Yan-Hong

2018-03-22

The production of anthocyanin is regulated by light and corresponding photoreceptors. In this study, we found that exposure to blue light and overexpression of CRY1a are associated with increased accumulation of anthocyanin in tomato (Solanum lycopersicum L.). These responses are the result of changes in mRNA and the protein levels of SlHY5, a transcription factor. In vitro and in vivo experiments using EMSA and ChIP-qPCR assays revealed that SlHY5 could directly recognize and bind to the G-box and ACE motifs in the promoters of anthocyanin biosynthesis genes, such as CHS1, CHS2 and DFR. Silencing of SlHY5 in OE-CRY1a lines decreased the accumulation of anthocyanin. The findings presented here not only deepened our understanding of how light controls anthocyanin biosynthesis and associated photoprotection in tomato leaves, but also allowed us to explore potential targets for improving pigment production. This article is protected by copyright. All rights reserved.

Transcriptomic profiling of Arabidopsis gene expression in response to varying micronutrient zinc supply

DEFF Research Database (Denmark)

Azevedo, Herlânder; Azinheiro, Sarah Gaspar; Muñoz-Mérida, Antonio

2016-01-01

Deficiency of the micronutrient zinc is a widespread condition in agricultural soils, causing a negative impact on crop quality and yield. Nevertheless, there is an insufficient knowledge on the regulatory and molecular mechanisms underlying the plant response to inadequate zinc nutrition [1......]. This information should contribute to the development of plant-based solutions with improved nutrient-use-efficiency traits in crops. Previously, the transcription factors bZIP19 and bZIP23 were identified as essential regulators of the response to zinc deficiency in Arabidopsis thaliana [2]. A microarray...... experiment comparing gene expression between roots of wild-type and the mutant bzip19 bzip23, exposed to zinc deficiency, led to the identification of differentially expressed genes related with zinc homeostasis, namely its transport and plant internal translocation [2]. Here, we provide the detailed...

A systems biology perspective on the role of WRKY transcription factors in drought responses in plants.

Science.gov (United States)

Tripathi, Prateek; Rabara, Roel C; Rushton, Paul J

2014-02-01

Drought is one of the major challenges affecting crop productivity and yield. However, water stress responses are notoriously multigenic and quantitative with strong environmental effects on phenotypes. It is also clear that water stress often does not occur alone under field conditions but rather in conjunction with other abiotic stresses such as high temperature and high light intensities. A multidisciplinary approach with successful integration of a whole range of -omics technologies will not only define the system, but also provide new gene targets for both transgenic approaches and marker-assisted selection. Transcription factors are major players in water stress signaling and some constitute major hubs in the signaling webs. The main transcription factors in this network include MYB, bHLH, bZIP, ERF, NAC, and WRKY transcription factors. The role of WRKY transcription factors in abiotic stress signaling networks is just becoming apparent and systems biology approaches are starting to define their places in the signaling network. Using systems biology approaches, there are now many transcriptomic analyses and promoter analyses that concern WRKY transcription factors. In addition, reports on nuclear proteomics have identified WRKY proteins that are up-regulated at the protein level by water stress. Interactomics has started to identify different classes of WRKY-interacting proteins. What are often lacking are connections between metabolomics, WRKY transcription factors, promoters, biosynthetic pathways, fluxes and downstream responses. As more levels of the system are characterized, a more detailed understanding of the roles of WRKY transcription factors in drought responses in crops will be obtained.

OsbZIP58, a basic leucine zipper transcription factor, regulates starch biosynthesis in rice endosperm.

Science.gov (United States)

Wang, Jie-Chen; Xu, Heng; Zhu, Ying; Liu, Qiao-Quan; Cai, Xiu-Ling

2013-08-01

Starch composition and the amount in endosperm, both of which contribute dramatically to seed yield, cooking quality, and taste in cereals, are determined by a series of complex biochemical reactions. However, the mechanism regulating starch biosynthesis in cereal seeds is not well understood. This study showed that OsbZIP58, a bZIP transcription factor, is a key transcriptional regulator controlling starch synthesis in rice endosperm. OsbZIP58 was expressed mainly in endosperm during active starch synthesis. osbzip58 null mutants displayed abnormal seed morphology with altered starch accumulation in the white belly region and decreased amounts of total starch and amylose. Moreover, osbzip58 had a higher proportion of short chains and a lower proportion of intermediate chains of amylopectin. Furthermore, OsbZIP58 was shown to bind directly to the promoters of six starch-synthesizing genes, OsAGPL3, Wx, OsSSIIa, SBE1, OsBEIIb, and ISA2, and to regulate their expression. These findings indicate that OsbZIP58 functions as a key regulator of starch synthesis in rice seeds and provide new insights into seed quality control.

Evidence supporting dissimilatory and assimilatory lignin degradation in Enterobacter lignolyticus SCF1

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Kristen M DeAngelis

2013-09-01

Full Text Available The anaerobic isolate Enterobacter lignolyticus SCF1 was initially cultivated based on anaerobic growth on lignin as sole carbon source. The source of the isolated bacteria was from tropical forest soils that decompose litter rapidly with low and fluctuating redox potentials, making it likely that bacteria using oxygen-independent enzymes play an important role in decomposition. We have used transcriptomics and proteomics to examine the increased growth of the anaerobic isolate Enterobacter lignolyticus SCF1 when grown on media amended with lignin compared to unamended growth. Proteomics revealed accelerated xylose uptake and metabolism under lignin-amended growth, and lignin degradation via the 4-hydroxyphenylacetate degradation pathway, catalase/peroxidase enzymes, and the glutathione biosynthesis and glutathione S-transferase proteins. We also observed increased production of NADH-quinone oxidoreductase, other electron transport chain proteins, and ATP synthase and ATP-binding cassette (ABC transporters. We detected significant lignin degradation over time by absorbance, and also used metabolomics to demonstrate increased xylose utilization in lignin-amended compared to unamended growth. Our data shows the advantages of a multi-omics approach, where incomplete pathways identified by genomics were completed, and new observations made on coping with poor carbon availability. The fast growth, high efficiency and specificity of enzymes employed in bacterial anaerobic litter deconstruction makes these soils useful templates for improving biofuel production.

Salicylic acid suppresses jasmonic acid signaling downstream of SCFCOI1-JAZ by targeting GCC promoter motifs via transcription factor ORA59.

Science.gov (United States)

Van der Does, Dieuwertje; Leon-Reyes, Antonio; Koornneef, Annemart; Van Verk, Marcel C; Rodenburg, Nicole; Pauwels, Laurens; Goossens, Alain; Körbes, Ana P; Memelink, Johan; Ritsema, Tita; Van Wees, Saskia C M; Pieterse, Corné M J

2013-02-01

Antagonism between the defense hormones salicylic acid (SA) and jasmonic acid (JA) plays a central role in the modulation of the plant immune signaling network, but the molecular mechanisms underlying this phenomenon are largely unknown. Here, we demonstrate that suppression of the JA pathway by SA functions downstream of the E3 ubiquitin-ligase Skip-Cullin-F-box complex SCF(COI1), which targets JASMONATE ZIM-domain transcriptional repressor proteins (JAZs) for proteasome-mediated degradation. In addition, neither the stability nor the JA-induced degradation of JAZs was affected by SA. In silico promoter analysis of the SA/JA crosstalk transcriptome revealed that the 1-kb promoter regions of JA-responsive genes that are suppressed by SA are significantly enriched in the JA-responsive GCC-box motifs. Using GCC:GUS lines carrying four copies of the GCC-box fused to the β-glucuronidase reporter gene, we showed that the GCC-box motif is sufficient for SA-mediated suppression of JA-responsive gene expression. Using plants overexpressing the GCC-box binding APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) transcription factors ERF1 or ORA59, we found that SA strongly reduces the accumulation of ORA59 but not that of ERF1. Collectively, these data indicate that the SA pathway inhibits JA signaling downstream of the SCF(COI1)-JAZ complex by targeting GCC-box motifs in JA-responsive promoters via a negative effect on the transcriptional activator ORA59.

Characterisation of Cdkl5transcript isoforms in rat

OpenAIRE

Hector, Ralph D.; Dando, Owen; Ritakari, Tuula E.; Kind, Peter C.; Bailey, Mark E.S.; Cobb, Stuart R.

2017-01-01

CDKL5 deficiency is a severe neurological disorder caused by mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5). The predominant human CDKL5 brain isoform is a 9.7kb transcript comprised of 18 exons with a large 6.6kb 3'-untranslated region (UTR). Mammalian models of CDKL5 disorder are currently limited to mouse, and little is known about Cdkl5 in other organisms used to model neurodevelopmental disorders, such as rat. In this study we characterise, both bioinformatically a...

The recruitment of the U5 snRNP to nascent transcripts requires internal loop 1 of U5 snRNA.

Science.gov (United States)

Kim, Rebecca; Paschedag, Joshua; Novikova, Natalya; Bellini, Michel

2012-12-01

In this study, we take advantage of the high spatial resolution offered by the nucleus and lampbrush chromosomes of the amphibian oocyte to investigate the mechanisms that regulate the intranuclear trafficking of the U5 snRNP and its recruitment to nascent transcripts. We monitor the fate of newly assembled fluorescent U5 snRNP in Xenopus oocytes depleted of U4 and/or U6 snRNAs and demonstrate that the U4/U6.U5 tri-snRNP is not required for the association of U5 snRNP with Cajal bodies, splicing speckles, and nascent transcripts. In addition, using a mutational analysis, we show that a non-functional U5 snRNP can associate with nascent transcripts, and we further characterize internal loop structure 1 of U5 snRNA as a critical element for licensing U5 snRNP to target both nascent transcripts and splicing speckles. Collectively, our data support the model where the recruitment of snRNPs onto pre-mRNAs is independent of spliceosome assembly and suggest that U5 snRNP may promote the association of the U4/U6.U5 tri-snRNP with nascent transcripts.

Hybrids of the bHLH and bZIP protein motifs display different DNA-binding activities in vivo vs. in vitro.

Directory of Open Access Journals (Sweden)

Hiu-Kwan Chow

Full Text Available Minimalist hybrids comprising the DNA-binding domain of bHLH/PAS (basic-helix-loop-helix/Per-Arnt-Sim protein Arnt fused to the leucine zipper (LZ dimerization domain from bZIP (basic region-leucine zipper protein C/EBP were designed to bind the E-box DNA site, CACGTG, targeted by bHLHZ (basic-helix-loop-helix-zipper proteins Myc and Max, as well as the Arnt homodimer. The bHLHZ-like structure of ArntbHLH-C/EBP comprises the Arnt bHLH domain fused to the C/EBP LZ: i.e. swap of the 330 aa PAS domain for the 29 aa LZ. In the yeast one-hybrid assay (Y1H, transcriptional activation from the E-box was strong by ArntbHLH-C/EBP, and undetectable for the truncated ArntbHLH (PAS removed, as detected via readout from the HIS3 and lacZ reporters. In contrast, fluorescence anisotropy titrations showed affinities for the E-box with ArntbHLH-C/EBP and ArntbHLH comparable to other transcription factors (K(d 148.9 nM and 40.2 nM, respectively, but only under select conditions that maintained folded protein. Although in vivo yeast results and in vitro spectroscopic studies for ArntbHLH-C/EBP targeting the E-box correlate well, the same does not hold for ArntbHLH. As circular dichroism confirms that ArntbHLH-C/EBP is a much more strongly alpha-helical structure than ArntbHLH, we conclude that the nonfunctional ArntbHLH in the Y1H must be due to misfolding, leading to the false negative that this protein is incapable of targeting the E-box. Many experiments, including protein design and selections from large libraries, depend on protein domains remaining well-behaved in the nonnative experimental environment, especially small motifs like the bHLH (60-70 aa. Interestingly, a short helical LZ can serve as a folding- and/or solubility-enhancing tag, an important device given the focus of current research on exploration of vast networks of biomolecular interactions.

A 5' splice site enhances the recruitment of basal transcription initiation factors in vivo

DEFF Research Database (Denmark)

Damgaard, Christian Kroun; Kahns, Søren; Lykke-Andersen, Søren

2008-01-01

RNAs, harboring wild-type or various 5′ splice site mutations, we demonstrate a strong positive correlation between splicing efficiency and transcription activity. Interestingly, a 5′ splice site can stimulate transcription even in the absence of splicing. Chromatin immunoprecipitation experiments show enhanced...... a promoter-proximal 5′ splice site via its U1 snRNA interaction can feed back to stimulate transcription initiation by enhancing preinitiation complex assembly....

A scalable implementation of RI-SCF on parallel computers

International Nuclear Information System (INIS)

Fruechtl, H.A.; Kendall, R.A.; Harrison, R.J.

1996-01-01

In order to avoid the integral bottleneck of conventional SCF calculations, the Resolution of the Identity (RI) method is used to obtain an approximate solution to the Hartree-Fock equations. In this approximation only three-center integrals are needed to build the Fock matrix. It has been implemented as part of the NWChem package of portable and scalable ab initio programs for parallel computers. Utilizing the V-approximation, both the Coulomb and exchange contribution to the Fock matrix can be calculated from a transformed set of three-center integrals which have to be precalculated and stored. A distributed in-core method as well as a disk based implementation have been programmed. Details of the implementation as well as the parallel programming tools used are described. We also give results and timings from benchmark calculations

Functional Analysis of Maize Silk-Specific ZmbZIP25 Promoter

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Wanying Li

2018-03-01

Full Text Available ZmbZIP25 (Zea mays bZIP (basic leucine zipper transcription factor 25 is a function-unknown protein that belongs to the D group of the bZIP transcription factor family. RNA-seq data showed that the expression of ZmbZIP25 was tissue-specific in maize silks, and this specificity was confirmed by RT-PCR (reverse transcription-polymerase chain reaction. In situ RNA hybridization showed that ZmbZIP25 was expressed exclusively in the xylem of maize silks. A 5′ RACE (rapid amplification of cDNA ends assay identified an adenine residue as the transcription start site of the ZmbZIP25 gene. To characterize this silk-specific promoter, we isolated and analyzed a 2450 bp (from −2083 to +367 and a 2600 bp sequence of ZmbZIP25 (from −2083 to +517, the transcription start site was denoted +1. Stable expression assays in Arabidopsis showed that the expression of the reporter gene GUS driven by the 2450 bp ZmbZIP25 5′-flanking fragment occurred exclusively in the papillae of Arabidopsis stigmas. Furthermore, transient expression assays in maize indicated that GUS and GFP expression driven by the 2450 bp ZmbZIP25 5′-flanking sequences occurred only in maize silks and not in other tissues. However, no GUS or GFP expression was driven by the 2600 bp ZmbZIP25 5′-flanking sequences in either stable or transient expression assays. A series of deletion analyses of the 2450 bp ZmbZIP25 5′-flanking sequence was performed in transgenic Arabidopsis plants, and probable elements prediction analysis revealed the possible presence of negative regulatory elements within the 161 bp region from −1117 to −957 that were responsible for the specificity of the ZmbZIP25 5′-flanking sequence.

Evidence supporting dissimilatory and assimilatory lignin degradation in Enterobacter lignolyticus SCF1

Energy Technology Data Exchange (ETDEWEB)

DeAngelis, Kristen M.; Sharma, Deepak; Varney, Rebecca; Simmons, Blake A.; Isern, Nancy G.; Markillie, Lye Meng; Nicora, Carrie D.; Norbeck, Angela D.; Taylor, Ronald C.; Aldrich, Joshua T.; Robinson, Errol W.

2013-08-29

The anaerobic isolate Enterobacter lignolyticus SCF1 was initially cultivated based on anaerobic growth on lignin as sole carbon source. The source of the isolated bacteria was from tropical forest soils that decompose litter rapidly with low and fluctuating redox potentials, making it likely that bacteria using oxygen-independent enzymes play an important role in decomposition. We have examined differential expression of the anaerobic isolate Enterobacter lignolyticus SCF1 during growth on lignin. After 48 hours of growth, we used transcriptomics and proteomics to define the enzymes and other regulatory machinery that these organisms use to degrade lignin, as well as metabolomics to measure lignin degradation and monitor the use of lignin and iron as terminal electron acceptors that facilitate more efficient use of carbon. Proteomics revealed accelerated xylose uptake and metabolism under lignin-amended growth, and lignin degradation via the 4-hydroxyphenylacetate degradation pathway, catalase/peroxidase enzymes, and the glutathione biosynthesis and glutathione S-transferase proteins. We also observed increased production of NADH-quinone oxidoreductase, other electron transport chain proteins, and ATP synthase and ATP-binding cassette (ABC) transporters. Our data shows the advantages of a multi-omics approach, where incomplete pathways identified by genomics were completed, and new observations made on coping with poor carbon availability. The fast growth, high efficiency and specificity of enzymes employed in bacterial anaerobic litter deconstruction makes these soils useful templates for improving biofuel production.

A 5' UTR-Overlapping LncRNA Activates the Male-Determining Gene doublesex1 in the Crustacean Daphnia magna.

Science.gov (United States)

Kato, Yasuhiko; Perez, Christelle Alexa G; Mohamad Ishak, Nur Syafiqah; Nong, Quang D; Sudo, Yuumi; Matsuura, Tomoaki; Wada, Tadashi; Watanabe, Hajime

2018-06-04

Long noncoding RNAs (lncRNAs) are pervasively transcribed in the eukaryotic genome [1] and are important for the control of master regulatory genes that are involved in cell differentiation and development [2, 3]. Here, we show that a 5' UTR-overlapping lncRNA regulates the male-specific expression of the DM-domain gene doublesex1 (dsx1) in the crustacean Daphnia magna, which produces males in response to environmental stimuli. This lncRNA, named doublesex1 alpha promoter-associated long RNA (DAPALR), is transcribed upstream the transcription start site (TSS) in a sense orientation and subjected to 5' end capping and 3' end processing at a stem-loop structure before the dsx1 coding exon. Similar to dsx1, its expression is only activated in males by the juvenile hormone (JH) and basic-leucine zipper (bZIP) transcription factor Vrille (Vri) and is maintained during embryogenesis. Knockdown of DAPALR in males silenced dsx1 and led to feminization, including egg production, whereas ectopic expression of DAPALR in dsx1-silenced females resulted in the de-repression of dsx1. We further demonstrate that the DAPALR transcript overlaps the dsx1 5'-UTR, and this overlapping region is required for dsx1 activation. Our results suggest that DAPALR can transactivate and possibly maintain dsx1 expression. This might be important for converting transient environmental signals into stable male development, controlled by the continuous expression of dsx1. Copyright © 2018 Elsevier Ltd. All rights reserved.

Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

OpenAIRE

In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A

1997-01-01

Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, delet...

Copper-Assisted Oxidative Trifluoromethylthiolation of 2,3-Allenoic Acids with AgSCF3.

Science.gov (United States)

Pan, Shen; Huang, Yangen; Xu, Xiu-Hua; Qing, Feng-Ling

2017-09-01

The oxidative trifluoromethylthiolation of 2,3-allenoic acids with AgSCF 3 in the presence of (NH 4 ) 2 S 2 O 8 and catalytic copper salt was investigated. A series of 4-aryl-2,3-allenoic acids underwent radical trifluoromethylthiolation/intramolecular cyclization to afford β-trifluoromethylthiolated butenolides, which were conveniently transformed into trifluoromethylthiolated furan derivatives. In contrast, 2-monosubstituted 2,3-allenoic acids were converted into the corresponding 3,4-bis(trifluoromethylthio)but-2-enoic-acids under similar reaction conditions.

AtbZIP34 is required for Arabidopsis pollen wall patterning and the control of several metabolic pathways in developing pollen

Czech Academy of Sciences Publication Activity Database

Gibalová, Antónia; Reňák, David; Matczuk, Katarzyna; Dupľáková, Nikoleta; Cháb, David; Twell, D.; Honys, David

2009-01-01

Roč. 70, č. 5 (2009), s. 581-601 ISSN 0167-4412 R&D Projects: GA ČR GA522/06/0896; GA ČR GA522/09/0858; GA MŠk(CZ) LC06004 Institutional research plan: CEZ:AV0Z50380511 Keywords : bZIP transcription factor * AtbZIP34 * Male gametophyte development Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.978, year: 2009

Rice black streaked dwarf virus P7-2 forms a SCF complex through binding to Oryza sativa SKP1-like proteins, and interacts with GID2 involved in the gibberellin pathway.

Science.gov (United States)

Tao, Tao; Zhou, Cui-Ji; Wang, Qian; Chen, Xiang-Ru; Sun, Qian; Zhao, Tian-Yu; Ye, Jian-Chun; Wang, Ying; Zhang, Zong-Ying; Zhang, Yong-Liang; Guo, Ze-Jian; Wang, Xian-Bing; Li, Da-Wei; Yu, Jia-Lin; Han, Cheng-Gui

2017-01-01

As a core subunit of the SCF complex that promotes protein degradation through the 26S proteasome, S-phase kinase-associated protein 1 (SKP1) plays important roles in multiple cellular processes in eukaryotes, including gibberellin (GA), jasmonate, ethylene, auxin and light responses. P7-2 encoded by Rice black streaked dwarf virus (RBSDV), a devastating viral pathogen that causes severe symptoms in infected plants, interacts with SKP1 from different plants. However, whether RBSDV P7-2 forms a SCF complex and targets host proteins is poorly understood. In this study, we conducted yeast two-hybrid assays to further explore the interactions between P7-2 and 25 type I Oryza sativa SKP1-like (OSK) proteins, and found that P7-2 interacted with eight OSK members with different binding affinity. Co-immunoprecipitation assay further confirmed the interaction of P7-2 with OSK1, OSK5 and OSK20. It was also shown that P7-2, together with OSK1 and O. sativa Cullin-1, was able to form the SCF complex. Moreover, yeast two-hybrid assays revealed that P7-2 interacted with gibberellin insensitive dwarf2 (GID2) from rice and maize plants, which is essential for regulating the GA signaling pathway. It was further demonstrated that the N-terminal region of P7-2 was necessary for the interaction with GID2. Overall, these results indicated that P7-2 functioned as a component of the SCF complex in rice, and interaction of P7-2 with GID2 implied possible roles of the GA signaling pathway during RBSDV infection.

Rice black streaked dwarf virus P7-2 forms a SCF complex through binding to Oryza sativa SKP1-like proteins, and interacts with GID2 involved in the gibberellin pathway.

Directory of Open Access Journals (Sweden)

Tao Tao

Full Text Available As a core subunit of the SCF complex that promotes protein degradation through the 26S proteasome, S-phase kinase-associated protein 1 (SKP1 plays important roles in multiple cellular processes in eukaryotes, including gibberellin (GA, jasmonate, ethylene, auxin and light responses. P7-2 encoded by Rice black streaked dwarf virus (RBSDV, a devastating viral pathogen that causes severe symptoms in infected plants, interacts with SKP1 from different plants. However, whether RBSDV P7-2 forms a SCF complex and targets host proteins is poorly understood. In this study, we conducted yeast two-hybrid assays to further explore the interactions between P7-2 and 25 type I Oryza sativa SKP1-like (OSK proteins, and found that P7-2 interacted with eight OSK members with different binding affinity. Co-immunoprecipitation assay further confirmed the interaction of P7-2 with OSK1, OSK5 and OSK20. It was also shown that P7-2, together with OSK1 and O. sativa Cullin-1, was able to form the SCF complex. Moreover, yeast two-hybrid assays revealed that P7-2 interacted with gibberellin insensitive dwarf2 (GID2 from rice and maize plants, which is essential for regulating the GA signaling pathway. It was further demonstrated that the N-terminal region of P7-2 was necessary for the interaction with GID2. Overall, these results indicated that P7-2 functioned as a component of the SCF complex in rice, and interaction of P7-2 with GID2 implied possible roles of the GA signaling pathway during RBSDV infection.

Activation of cellular immunity and marked inhibition of liver cancer in a mouse model following gene therapy and tumor expression of GM-SCF, IL-21, and Rae-1.

Science.gov (United States)

Cheng, Mingrong; Zhi, Kangkang; Gao, Xiaoyan; He, Bing; Li, Yingchun; Han, Jiang; Zhang, Zhiping; Wu, Yan

2013-12-18

Cancer is both a systemic and a genetic disease. The pathogenesis of cancer might be related to dampened immunity. Host immunity recognizes nascent malignant cells - a process referred to as immune surveillance. Augmenting immune surveillance and suppressing immune escape are crucial in tumor immunotherapy. A recombinant plasmid capable of co-expressing granulocyte-macrophage colony- stimulating factor (GM-SCF), interleukin-21 (IL-21), and retinoic acid early transcription factor-1 (Rae-1) was constructed, and its effects determined in a mouse model of subcutaneous liver cancer. Serum specimens were assayed for IL-2 and INF-γ by ELISA. Liver cancer specimens were isolated for Rae-1 expression by RT-PCR and Western blot, and splenocytes were analyzed by flow cytometry. The recombinant plasmid inhibited the growth of liver cancer and prolonged survival of tumor-loaded mice. Activation of host immunity might have contributed to this effect by promoting increased numbers and cytotoxicity of natural killer (NK) cells and cytotoxic T lymphocytes (CTL) following expression of GM-SCF, IL-21, and Rae-1. By contrast, the frequency of regulatory T cells was decreased, Consequently, activated CTL and NK cells enhanced their secretion of INF-γ, which promoted cytotoxicity of NK cells and CTL. Moreover, active CTL showed dramatic secretion of IL-2, which stimulates CTL. The recombinant expression plasmid also augmented Rae-1 expression by liver cancer cells. Rae-1 receptor expressing CTL and NK cells removed liver cancer. The recombinant expression plasmid inhibited liver cancer by a mechanism that involved activation of cell-mediated immunity and Rae-1 in liver cancer.

A central integrator of transcription networks in plant stress and energy signalling.

Science.gov (United States)

Baena-González, Elena; Rolland, Filip; Thevelein, Johan M; Sheen, Jen

2007-08-23

Photosynthetic plants are the principal solar energy converter sustaining life on Earth. Despite its fundamental importance, little is known about how plants sense and adapt to darkness in the daily light-dark cycle, or how they adapt to unpredictable environmental stresses that compromise photosynthesis and respiration and deplete energy supplies. Current models emphasize diverse stress perception and signalling mechanisms. Using a combination of cellular and systems screens, we show here that the evolutionarily conserved Arabidopsis thaliana protein kinases, KIN10 and KIN11 (also known as AKIN10/At3g01090 and AKIN11/At3g29160, respectively), control convergent reprogramming of transcription in response to seemingly unrelated darkness, sugar and stress conditions. Sensing and signalling deprivation of sugar and energy, KIN10 targets a remarkably broad array of genes that orchestrate transcription networks, promote catabolism and suppress anabolism. Specific bZIP transcription factors partially mediate primary KIN10 signalling. Transgenic KIN10 overexpression confers enhanced starvation tolerance and lifespan extension, and alters architecture and developmental transitions. Significantly, double kin10 kin11 deficiency abrogates the transcriptional switch in darkness and stress signalling, and impairs starch mobilization at night and growth. These studies uncover surprisingly pivotal roles of KIN10/11 in linking stress, sugar and developmental signals to globally regulate plant metabolism, energy balance, growth and survival. In contrast to the prevailing view that sucrose activates plant SnRK1s (Snf1-related protein kinases), our functional analyses of Arabidopsis KIN10/11 provide compelling evidence that SnRK1s are inactivated by sugars and share central roles with the orthologous yeast Snf1 and mammalian AMPK in energy signalling.

Specificity determinants for the abscisic acid response element ?

OpenAIRE

Sarkar, Aditya Kumar; Lahiri, Ansuman

2013-01-01

Abscisic acid (ABA) response elements (ABREs) are a group of cis-acting DNA elements that have been identified from promoter analysis of many ABA-regulated genes in plants. We are interested in understanding the mechanism of binding specificity between ABREs and a class of bZIP transcription factors known as ABRE binding factors (ABFs). In this work, we have modeled the homodimeric structure of the bZIP domain of ABRE binding factor 1 from Arabidopsis thaliana (AtABF1) and studied its interac...

Knockdown of SCF(Skp2 function causes double-parked accumulation in the nucleus and DNA re-replication in Drosophila plasmatocytes.

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Paul T Kroeger

Full Text Available In Drosophila, circulating hemocytes are derived from the cephalic mesoderm during the embryonic wave of hematopoiesis. These cells are contributed to the larva and persist through metamorphosis into the adult. To analyze this population of hemocytes, we considered data from a previously published RNAi screen in the hematopoietic niche, which suggested several members of the SCF complex play a role in lymph gland development. eater-Gal4;UAS-GFP flies were crossed to UAS-RNAi lines to knockdown the function of all known SCF complex members in a plasmatocyte-specific fashion, in order to identify which members are novel regulators of plasmatocytes. This specific SCF complex contains five core members: Lin-19-like, SkpA, Skp2, Roc1a and complex activator Nedd8. The complex was identified by its very distinctive large cell phenotype. Furthermore, these large cells stained for anti-P1, a plasmatocyte-specific antibody. It was also noted that the DNA in these cells appeared to be over-replicated. Gamma-tubulin and DAPI staining suggest the cells are undergoing re-replication as they had multiple centrioles and excessive DNA content. Further experimentation determined enlarged cells were BrdU-positive indicating they have progressed through S-phase. To determine how these cells become enlarged and undergo re-replication, cell cycle proteins were analyzed by immunofluorescence. This analysis identified three proteins that had altered subcellular localization in these enlarged cells: Cyclin E, Geminin and Double-parked. Previous research has shown that Double-parked must be degraded to exit S-phase, otherwise the DNA will undergo re-replication. When Double-parked was titrated from the nucleus by an excess of its inhibitor, geminin, the enlarged cells and aberrant protein localization phenotypes were partially rescued. The data in this report suggests that the SCF(Skp2 complex is necessary to ubiquitinate Double-parked during plasmatocyte cell division

A novel transcript of cyclin-dependent kinase-like 5 (CDKL5) has an alternative C-terminus and is the predominant transcript in brain.

Science.gov (United States)

Williamson, Sarah L; Giudici, Laura; Kilstrup-Nielsen, Charlotte; Gold, Wendy; Pelka, Gregory J; Tam, Patrick P L; Grimm, Andrew; Prodi, Dionigio; Landsberger, Nicoletta; Christodoulou, John

2012-02-01

The X-linked cyclin-dependent kinase-like 5 (CDKL5) gene is an important molecular determinant of early-onset intractable seizures with infantile spasms and Rett syndrome-like phenotype. The gene encodes a kinase that may influence components of molecular pathways associated with MeCP2. In humans there are two previously reported splice variants that differ in the 5' untranslated exons and produce the same 115 kDa protein. Furthermore, very recently, a novel transcript including a novel exon (16b) has been described. By aligning both the human and mouse CDKL5 proteins to the orthologs of other species, we identified a theoretical 107 kDa isoform with an alternative C-terminus that terminates in intron 18. In human brain and all other tissues investigated except the testis, this novel isoform is the major CDKL5 transcript. The detailed characterisation of this novel isoform of CDKL5 reveals functional and subcellular localisation attributes that overlap greatly, but not completely, with that of the previously studied human CDKL5 protein. Considering its predominant expression in the human and mouse brain, we believe that this novel isoform is likely to be of primary pathogenic importance in human diseases associated with CDKL5 deficiency, and suggest that screening of the related intronic sequence should be included in the molecular genetic analyses of patients with a suggestive clinical phenotype.

Application of gradient-corrected density functional theory to the structures and thermochemistries of ScF3, TiF4, VF5, and CrF6

International Nuclear Information System (INIS)

Russo, T.V.; Martin, R.L.; Jeffrey Hay, P.

1995-01-01

Density functional theory (DFT) and Hartree--Fock (HF) calculations are reported for the family of transition metal fluorides ScF 3 , TiF 4 , VF 5 , and CrF 6 . Both HF and the local-density approximation (LDA) yield excellent agreement with experimental bond lengths, while the B-LYP gradient-corrected density functional gives bond lengths 0.04-0.05 A too long. An investigation of various combinations of exchange and correlation functionals shows that, for this series, the origin of this behavior lies in the Becke exchange functional. Much improved bond distances are found using the hybrid HF/DFT functional advocated by Becke. This approximation also leads to much improved thermochemistries. The LDA overestimates average bond energies in this series by 30-40 kcal/mol, whereas the B-LYP functional overbinds by only ∼8-12 kcal/mol, and the hybrid HF/DFT method overbinds by only ∼2 kcal/mol. The hybrid method predicts the octahedral isomer of CrF 6 to be more stable than the trigonal prismatic form by 14 kcal/mol. Comparison of theoretical vibrational frequencies with experiment supports the assignment of an octahedral geometry

Expression analysis of OsbZIP transcription factors in resistance ...

African Journals Online (AJOL)

zino

2013-08-21

Aug 21, 2013 ... Plant basic leucine zipper (bZIP) proteins play an essential role in the genes expression and regulation in higher plants. They have been shown to regulate diverse plant specific phenomena, including germination, floral induction and development, seed maturation, photomorphogenesis, biotic and.

The DOF transcription factor Dof5.1 influences leaf axial patterning by promoting Revoluta transcription in Arabidopsis

KAUST Repository

Kim, Hyungsae

2010-10-05

Dof proteins are transcription factors that have a conserved single zinc finger DNA-binding domain. In this study, we isolated an activation tagging mutant Dof5.1-D exhibiting an upward-curling leaf phenotype due to enhanced expression of the REV gene that is required for establishing adaxialabaxial polarity. Dof5.1-D plants also had reduced transcript levels for IAA6 and IAA19 genes, indicating an altered auxin biosynthesis in Dof5.1-D. An electrophoretic mobility shift assay using the Dof5.1 DNA-binding motif and the REV promoter region indicated that the DNA-binding domain of Dof5.1 binds to a TAAAGT motif located in the 5′-distal promoter region of the REV promoter. Further, transient and chromatin immunoprecipitation assays verified binding activity of the Dof5.1 DNA-binding motif with the REV promoter. Consistent with binding assays, constitutive over-expression of the Dof5.1 DNA-binding domain in wild-type plants caused a downward-curling phenotype, whereas crossing Dof5.1-D to a rev mutant reverted the upward-curling phenotype of the Dof5.1-D mutant leaf to the wild-type. These results suggest that the Dof5.1 protein directly binds to the REV promoter and thereby regulates adaxialabaxial polarity. © 2010 Blackwell Publishing Ltd.

The DOF transcription factor Dof5.1 influences leaf axial patterning by promoting Revoluta transcription in Arabidopsis

KAUST Repository

Kim, Hyungsae; Kim, Sungjin; Abbasi, Nazia; Bressan, Ray Anthony; Yun, Daejin; Yoo, Sangdong; Kwon, SukYun; Choi, Sangbong

2010-01-01

Dof proteins are transcription factors that have a conserved single zinc finger DNA-binding domain. In this study, we isolated an activation tagging mutant Dof5.1-D exhibiting an upward-curling leaf phenotype due to enhanced expression of the REV gene that is required for establishing adaxialabaxial polarity. Dof5.1-D plants also had reduced transcript levels for IAA6 and IAA19 genes, indicating an altered auxin biosynthesis in Dof5.1-D. An electrophoretic mobility shift assay using the Dof5.1 DNA-binding motif and the REV promoter region indicated that the DNA-binding domain of Dof5.1 binds to a TAAAGT motif located in the 5′-distal promoter region of the REV promoter. Further, transient and chromatin immunoprecipitation assays verified binding activity of the Dof5.1 DNA-binding motif with the REV promoter. Consistent with binding assays, constitutive over-expression of the Dof5.1 DNA-binding domain in wild-type plants caused a downward-curling phenotype, whereas crossing Dof5.1-D to a rev mutant reverted the upward-curling phenotype of the Dof5.1-D mutant leaf to the wild-type. These results suggest that the Dof5.1 protein directly binds to the REV promoter and thereby regulates adaxialabaxial polarity. © 2010 Blackwell Publishing Ltd.

Protein Kinase R Degradation Is Essential for Rift Valley Fever Virus Infection and Is Regulated by SKP1-CUL1-F-box (SCF)FBXW11-NSs E3 Ligase.

Science.gov (United States)

Mudhasani, Rajini; Tran, Julie P; Retterer, Cary; Kota, Krishna P; Whitehouse, Chris A; Bavari, Sina

2016-02-01

Activated protein kinase R (PKR) plays a vital role in antiviral defense primarily by inhibiting protein synthesis and augmenting interferon responses. Many viral proteins have adopted unique strategies to counteract the deleterious effects of PKR. The NSs (Non-structural s) protein which is encoded by Rift Valley fever virus (RVFV) promotes early PKR proteasomal degradation through a previously undefined mechanism. In this study, we demonstrate that NSs carries out this activity by assembling the SCF (SKP1-CUL1-F-box)(FBXW11) E3 ligase. NSs binds to the F-box protein, FBXW11, via the six amino acid sequence DDGFVE called the degron sequence and recruits PKR through an alternate binding site to the SCF(FBXW11) E3 ligase. We further show that disrupting the assembly of the SCF(FBXW11-NSs) E3 ligase with MLN4924 (a small molecule inhibitor of SCF E3 ligase activity) or NSs degron viral mutants or siRNA knockdown of FBXW11 can block PKR degradation. Surprisingly, under these conditions when PKR degradation was blocked, NSs was essential and sufficient to activate PKR causing potent inhibition of RVFV infection by suppressing viral protein synthesis. These antiviral effects were antagonized by the loss of PKR expression or with a NSs deleted mutant virus. Therefore, early PKR activation by disassembly of SCF(FBXW11-NSs) E3 ligase is sufficient to inhibit RVFV infection. Furthermore, FBXW11 and BTRC are the two homologues of the βTrCP (Beta-transducin repeat containing protein) gene that were previously described to be functionally redundant. However, in RVFV infection, among the two homologues of βTrCP, FBXW11 plays a dominant role in PKR degradation and is the limiting factor in the assembly of the SCF(FBXW11) complex. Thus, FBXW11 serves as a master regulator of RVFV infection by promoting PKR degradation. Overall these findings provide new insights into NSs regulation of PKR activity and offer potential opportunities for therapeutic intervention of RVFV infection.

N-Myc and GCN5 regulate significantly overlapping transcriptional programs in neural stem cells.

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Verónica Martínez-Cerdeño

Full Text Available Here we examine the functions of the Myc cofactor and histone acetyltransferase, GCN5/KAT2A, in neural stem and precursor cells (NSC using a conditional knockout approach driven by nestin-cre. Mice with GCN5-deficient NSC exhibit a 25% reduction in brain mass with a microcephaly phenotype similar to that observed in nestin-cre driven knockouts of c- or N-myc. In addition, the loss of GCN5 inhibits precursor cell proliferation and reduces their populations in vivo, as does loss of N-myc. Gene expression analysis indicates that about one-sixth of genes whose expression is affected by loss of GCN5 are also affected in the same manner by loss of N-myc. These findings strongly support the notion that GCN5 protein is a key N-Myc transcriptional cofactor in NSC, but are also consistent with recruitment of GCN5 by other transcription factors and the use by N-Myc of other histone acetyltransferases. Putative N-Myc/GCN5 coregulated transcriptional pathways include cell metabolism, cell cycle, chromatin, and neuron projection morphogenesis genes. GCN5 is also required for maintenance of histone acetylation both at its putative specific target genes and at Myc targets. Thus, we have defined an important role for GCN5 in NSC and provided evidence that GCN5 is an important Myc transcriptional cofactor in vivo.

HBZ and its roles in HTLV-1 oncogenesis

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Tiejun eZhao

2012-07-01

Full Text Available Human T-cell leukemia virus type 1 (HTLV-1 causes adult T-cell leukemia (ATL. The minus strand of HTLV-1 provirus encodes a bZIP protein donated as HTLV-1 bZIP factor (HBZ. Among the HTLV-1 regulatory and accessory genes, the tax and HBZ genes were thought to play critical roles in oncogenesis. However, HBZ is the only gene that remains intact and is consistently expressed in all ATL cases, while the tax gene is frequently inactivated by epigenetic modifications or deletion of the 5’LTR. HBZ gene promotes the proliferation of ATL cells through its mRNA form. Moreover, HBZ induces T-cell lymphoma and systemic inflammation in vivo. HBZ fulfills its functions mainly through regulating HTLV-1 5’LTR transcription and modulating a variety of cellular signaling pathways which are related with cell growth, immune response and T-cell differentiation. Taken together, the multiple functions of HBZ render its predominant function in leukemogenesis of ATL.

Fungal Morphology, Iron Homeostasis, and Lipid Metabolism Regulated by a GATA Transcription Factor in Blastomyces dermatitidis.

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Amber J Marty

2015-06-01

Full Text Available In response to temperature, Blastomyces dermatitidis converts between yeast and mold forms. Knowledge of the mechanism(s underlying this response to temperature remains limited. In B. dermatitidis, we identified a GATA transcription factor, SREB, important for the transition to mold. Null mutants (SREBΔ fail to fully complete the conversion to mold and cannot properly regulate siderophore biosynthesis. To capture the transcriptional response regulated by SREB early in the phase transition (0-48 hours, gene expression microarrays were used to compare SREB∆ to an isogenic wild type isolate. Analysis of the time course microarray data demonstrated SREB functioned as a transcriptional regulator at 37°C and 22°C. Bioinformatic and biochemical analyses indicated SREB was involved in diverse biological processes including iron homeostasis, biosynthesis of triacylglycerol and ergosterol, and lipid droplet formation. Integration of microarray data, bioinformatics, and chromatin immunoprecipitation identified a subset of genes directly bound and regulated by SREB in vivo in yeast (37°C and during the phase transition to mold (22°C. This included genes involved with siderophore biosynthesis and uptake, iron homeostasis, and genes unrelated to iron assimilation. Functional analysis suggested that lipid droplets were actively metabolized during the phase transition and lipid metabolism may contribute to filamentous growth at 22°C. Chromatin immunoprecipitation, RNA interference, and overexpression analyses suggested that SREB was in a negative regulatory circuit with the bZIP transcription factor encoded by HAPX. Both SREB and HAPX affected morphogenesis at 22°C; however, large changes in transcript abundance by gene deletion for SREB or strong overexpression for HAPX were required to alter the phase transition.

Interaction between C/EBPβ and Tax down-regulates human T-cell leukemia virus type I transcription

International Nuclear Information System (INIS)

Hivin, P.; Gaudray, G.; Devaux, C.; Mesnard, J.-M.

2004-01-01

The human T-cell leukemia virus type I (HTLV-I) Tax protein trans-activates viral transcription through three imperfect tandem repeats of a 21-bp sequence called Tax-responsive element (TxRE). Tax regulates transcription via direct interaction with some members of the activating transcription factor/CRE-binding protein (ATF/CREB) family including CREM, CREB, and CREB-2. By interacting with their ZIP domain, Tax stimulates the binding of these cellular factors to the CRE-like sequence present in the TxREs. Recent observations have shown that CCAAT/enhancer binding protein β (C/EBPβ) forms stable complexes on the CRE site in the presence of CREB-2. Given that C/EBPβ has also been found to interact with Tax, we analyzed the effects of C/EBPβ on viral Tax-dependent transcription. We show here that C/EBPβ represses viral transcription and that Tax is no more able to form a stable complex with CREB-2 on the TxRE site in the presence of C/EBPβ. We also analyzed the physical interactions between Tax and C/EBPβ and found that the central region of C/EBPβ, excluding its ZIP domain, is required for direct interaction with Tax. It is the first time that Tax is described to interact with a basic leucine-zipper (bZIP) factor without recognizing its ZIP domain. Although unexpected, this result explains why C/EBPβ would be unable to form a stable complex with Tax on the TxRE site and could then down-regulate viral transcription. Lastly, we found that C/EBPβ was able to inhibit Tax expression in vivo from an infectious HTLV-I molecular clone. In conclusion, we propose that during cell activation events, which stimulate the Tax synthesis, C/EBPβ may down-regulate the level of HTLV-I expression to escape the cytotoxic-T-lymphocyte response

Electronic structure and related properties of ferrocyanide ion calculated by the SCF Xα-scattered wave method

International Nuclear Information System (INIS)

Guenzburger, D.; Maffeo, B.; Siqueira, M.L. de

1975-08-01

The SCF-XαSW method is used to calculate the electronic structure of the ferrocyanide ion. Optical transitions and X-Ray photoelectron emission are obtained from the energy level scheme and compared with experimental results. The charge density in the Fe nucleus is also computed and the result is correlated with isomer shift measurements made on this and other Fe complexes for which theoretical calculations have been performed

Comprehensive Behavioral Analysis of Activating Transcription Factor 5-Deficient Mice

Directory of Open Access Journals (Sweden)

Mariko Umemura

2017-07-01

Full Text Available Activating transcription factor 5 (ATF5 is a member of the CREB/ATF family of basic leucine zipper transcription factors. We previously reported that ATF5-deficient (ATF5-/- mice demonstrated abnormal olfactory bulb development due to impaired interneuron supply. Furthermore, ATF5-/- mice were less aggressive than ATF5+/+ mice. Although ATF5 is widely expressed in the brain, and involved in the regulation of proliferation and development of neurons, the physiological role of ATF5 in the higher brain remains unknown. Our objective was to investigate the physiological role of ATF5 in the higher brain. We performed a comprehensive behavioral analysis using ATF5-/- mice and wild type littermates. ATF5-/- mice exhibited abnormal locomotor activity in the open field test. They also exhibited abnormal anxiety-like behavior in the light/dark transition test and open field test. Furthermore, ATF5-/- mice displayed reduced social interaction in the Crawley’s social interaction test and increased pain sensitivity in the hot plate test compared with wild type. Finally, behavioral flexibility was reduced in the T-maze test in ATF5-/- mice compared with wild type. In addition, we demonstrated that ATF5-/- mice display disturbances of monoamine neurotransmitter levels in several brain regions. These results indicate that ATF5 deficiency elicits abnormal behaviors and the disturbance of monoamine neurotransmitter levels in the brain. The behavioral abnormalities of ATF5-/- mice may be due to the disturbance of monoamine levels. Taken together, these findings suggest that ATF5-/- mice may be a unique animal model of some psychiatric disorders.

Calculations of chemisorption of hydrogen and nitrogen on titanium using Xα-SW and SCF-CI methods

International Nuclear Information System (INIS)

Hung, S.C.

1983-01-01

Two theoretical studies are used as examples of chemisorption on transition metal surfaces. The first calculation employs the SCF-Sα-SW method to determine the electronic structures of several close-packed Ti-Ti and Ti-H systems. The relative importance of Ti s, p, and electron bonding to its Ti neighbors and to H are reported. The roles of the d electrons in the Ti-Ti and Ti-H bond are discussed. The effect of the presence of an H atom on the first few layers of the metal is also studied. The second set of calculations employs the Hartree-Fock-Roothaan SCF formulation supplemented by configuration-interaction (CI) to determine the energetics of adsorption of an N 2 molecule at a Ti surface. These calculations are made tractable by using a Ti core potential and by a unitary localization transformation to define a local surface region of localized lattice plus adsorbate orbitals. The results show that an underlayer of N atoms forms at the octahedral sites. The calculated adsorption energy of 115 kcal/mole compares favorably with experiment. Binding energy contributions in the several successive steps of the calculations are discussed. Eigenvalue spectra for the selected lattice-adsorbate geometries are compared to the UPS experimental data

SCF MS Xα determination of ionization energies and atomic populations of octaedral TiO6-8 cluster

International Nuclear Information System (INIS)

Michel-Calendini, F.M.; Chermette, H.; Pertosa, P.

1979-02-01

The ionization energies of titanium and oxygen states in BaTiO 3 crystal have been investigated through the self-consistent-field-Xα-scattered-wave (SCF MS Xα) method, with the Slater transition state model, applied to a TiO 6 -8 cluster of octaedral symmetry. Ionization energies and electronic charge distribution are compared to XPS data and related to results obtained from tight-binding band computations

The obesity-associated transcription factor ETV5 modulates circulating glucocorticoids

Science.gov (United States)

Gutierrez-Aguilar, Ruth; Thompson, Abigail; Marchand, Nathalie; Dumont, Patrick; Woods, Stephen C.; de Launoit, Yvan; Seeley, Randy J.; Ulrich-Lai, Yvonne M.

2015-01-01

The transcription factor E-twenty-six version 5 (ETV5) has been linked with obesity in genome-wide association studies. Moreover, ETV5-deficient mice (knockout; KO) have reduced body weight, lower fat mass, and are resistant to diet-induced obesity, directly linking ETV5 to the regulation of energy balance and metabolism. ETV5 is expressed in hypothalamic brain regions that regulate both metabolism and HPA axis activity, suggesting that ETV5 may also modulate HPA axis function. In order to test this possibility, plasma corticosterone levels were measured in ETV5 KO and wildtype (WT) mice before (pre-stress) and after (post-stress) a mild stressor (intraperitoneal injection). ETV5 deficiency increased both pre- and post-stress plasma corticosterone, suggesting that loss of ETV5 elevated glucocorticoid tone. Consistent with this idea, ETV5 KO mice have reduced thymus weight, suggestive of increased glucocorticoid-induced thymic involution. ETV5 deficiency also decreased the mRNA expression of glucocorticoid receptor (GR), mineralocorticoid receptor (MR), and vasopressin receptor 1A in the hypothalamus, without altering vasopressin, corticotropin-releasing hormone, or oxytocin mRNA expression. In order to test whether reduced MR and GR expression affected glucocorticoid negative feedback, a dexamethasone suppression test was performed. Dexamethasone reduced plasma corticosterone in both ETV5 KO and WT mice, suggesting that glucocorticoid negative feedback was unaltered by ETV5 deficiency. In summary, these data suggest that the obesity-associated transcription factor ETV5 normally acts to diminish circulating glucocorticoids. This might occur directly via ETV5 actions on HPA-regulatory brain circuitry, and/or indirectly via ETV5-induced alterations in metabolic factors that then influence the HPA axis. PMID:25813907

Transcript levels, alternative splicing and proteolytic cleavage of TFIIIA control 5S rRNA accumulation during Arabidopsis thaliana development.

Science.gov (United States)

Layat, Elodie; Cotterell, Sylviane; Vaillant, Isabelle; Yukawa, Yasushi; Tutois, Sylvie; Tourmente, Sylvette

2012-07-01

Ribosome biogenesis is critical for eukaryotic cells and requires coordinated synthesis of the protein and rRNA moieties of the ribosome, which are therefore highly regulated. 5S ribosomal RNA, an essential component of the large ribosomal subunit, is transcribed by RNA polymerase III and specifically requires transcription factor IIIA (TFIIIA). To obtain insight into the regulation of 5S rRNA transcription, we have investigated the expression of 5S rRNA and the exon-skipped (ES) and exon-including (EI) TFIIIA transcripts, two transcript isoforms that result from alternative splicing of the TFIIIA gene, and TFIIIA protein amounts with respect to requirements for 5S rRNA during development. We show that 5S rRNA quantities are regulated through distinct but complementary mechanisms operating through transcriptional and post-transcriptional control of TFIIIA transcripts as well as at the post-translational level through proteolytic cleavage of the TFIIIA protein. During the reproductive phase, high expression of the TFIIIA gene together with low proteolytic cleavage contributes to accumulation of functional, full-length TFIIIA protein, and results in 5S rRNA accumulation in the seed. In contrast, just after germination, the levels of TFIIIA-encoding transcripts are low and stable. Full-length TFIIIA protein is undetectable, and the level of 5S rRNA stored in the embryo progressively decreases. After day 4, in correlation with the reorganization of 5S rDNA chromatin to a mature state, full-length TFIIIA protein with transcriptional activity accumulates and permits de novo transcription of 5S rRNA. © 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.

Phylogenetic analysis of F-bZIP transcription factors indicates conservation of the zinc deficiency response across land plants

DEFF Research Database (Denmark)

Castro, Pedro Humberto Araújo R F; Lilay, Grmay Hailu; Muñoz-Mérida, Antonio

2017-01-01

Basic leucine zipper (bZIP) transcription factors control important developmental and physiological processes in plants. In Arabidopsis thaliana, the three gene F-bZIP subfamily has been associated with zinc deficiency and salt stress response. Benefiting from the present abundance of plant genomic...... data, we performed an evolutionary and structural characterization of plant F-bZIPs. We observed divergence during seed plant evolution, into two groups and inferred different selective pressures for each. Group 1 contains AtbZIP19 and AtbZIP23 and appears more conserved, whereas Group 2, containing...... of AtZIP4. A survey of AtZIP4 orthologs promoters across different plant taxa revealed an enrichment of the Zinc Deficiency Response Element (ZDRE) to which both AtbZIP19/23 bind. Overall, our results indicate that while the AtbZIP24 function in the regulation of the salt stress response may...

Pseudosubstrate regulation of the SCF(beta-TrCP) ubiquitin ligase by hnRNP-U

DEFF Research Database (Denmark)

Davis, Matti; Hatzubai, Ada; Andersen, Jens S

2002-01-01

in the nucleus. Here we report the isolation of the major E3RS-associated protein, hnRNP-U, an abundant nuclear phosphoprotein. This protein occupies E3RS in a specific and stoichiometric manner, stabilizes the E3 component, and is likely responsible for its nuclear localization. hnRNP-U binding was abolished....... Consequently, hnRNP-U engages a highly neddylated active SCF(beta-TrCP), which dissociates in the presence of a high-affinity substrate, resulting in ubiquitination of the latter. Our study points to a novel regulatory mechanism, which secures the localization, stability, substrate binding threshold...

Supra-optimal expression of the cold-regulated OsMyb4 transcription factor in transgenic rice changes the complexity of transcriptional network with major effects on stress tolerance and panicle development

KAUST Repository

Park, Myoungryoul

2010-09-28

The R2R3-type OsMyb4 transcription factor of rice has been shown to play a role in the regulation of osmotic adjustment in heterologous overexpression studies. However, the exact composition and organization of its underlying transcriptional network has not been established to be a robust tool for stress tolerance enhancement by regulon engineering. OsMyb4 network was dissected based on commonalities between the global chilling stress transcriptome and the transcriptome configured by OsMyb4 overexpression. OsMyb4 controls a hierarchical network comprised of several regulatory sub-clusters associated with cellular defense and rescue, metabolism and development. It regulates target genes either directly or indirectly through intermediary MYB, ERF, bZIP, NAC, ARF and CCAAT-HAP transcription factors. Regulatory sub-clusters have different combinations of MYB-like, GCC-box-like, ERD1-box-like, ABRE-like, G-box-like, as1/ocs/TGA-like, AuxRE-like, gibberellic acid response element (GARE)-like and JAre-like cis-elements. Cold-dependent network activity enhanced cellular antioxidant capacity through radical scavenging mechanisms and increased activities of phenylpropanoid and isoprenoid metabolic processes involving various abscisic acid (ABA), jasmonic acid (JA), salicylic acid (SA), ethylene and reactive oxygen species (ROS) responsive genes. OsMyb4 network is independent of drought response element binding protein/C-repeat binding factor (DREB/CBF) and its sub-regulons operate with possible co-regulators including nuclear factor-Y. Because of its upstream position in the network hierarchy, OsMyb4 functions quantitatively and pleiotrophically. Supra-optimal expression causes misexpression of alternative targets with costly trade-offs to panicle development. © 2010 Blackwell Publishing Ltd.

Supra-optimal expression of the cold-regulated OsMyb4 transcription factor in transgenic rice changes the complexity of transcriptional network with major effects on stress tolerance and panicle development.

Science.gov (United States)

Park, Myoung-Ryoul; Yun, Kil-Young; Mohanty, Bijayalaxmi; Herath, Venura; Xu, Fuyu; Wijaya, Edward; Bajic, Vladimir B; Yun, Song-Joong; De Los Reyes, Benildo G

2010-12-01

The R2R3-type OsMyb4 transcription factor of rice has been shown to play a role in the regulation of osmotic adjustment in heterologous overexpression studies. However, the exact composition and organization of its underlying transcriptional network has not been established to be a robust tool for stress tolerance enhancement by regulon engineering. OsMyb4 network was dissected based on commonalities between the global chilling stress transcriptome and the transcriptome configured by OsMyb4 overexpression. OsMyb4 controls a hierarchical network comprised of several regulatory sub-clusters associated with cellular defense and rescue, metabolism and development. It regulates target genes either directly or indirectly through intermediary MYB, ERF, bZIP, NAC, ARF and CCAAT-HAP transcription factors. Regulatory sub-clusters have different combinations of MYB-like, GCC-box-like, ERD1-box-like, ABRE-like, G-box-like, as1/ocs/TGA-like, AuxRE-like, gibberellic acid response element (GARE)-like and JAre-like cis-elements. Cold-dependent network activity enhanced cellular antioxidant capacity through radical scavenging mechanisms and increased activities of phenylpropanoid and isoprenoid metabolic processes involving various abscisic acid (ABA), jasmonic acid (JA), salicylic acid (SA), ethylene and reactive oxygen species (ROS) responsive genes. OsMyb4 network is independent of drought response element binding protein/C-repeat binding factor (DREB/CBF) and its sub-regulons operate with possible co-regulators including nuclear factor-Y. Because of its upstream position in the network hierarchy, OsMyb4 functions quantitatively and pleiotrophically. Supra-optimal expression causes misexpression of alternative targets with costly trade-offs to panicle development. © 2010 Blackwell Publishing Ltd.

Mutation in an alternative transcript of CDKL5 in a boy with early-onset seizures.

Science.gov (United States)

Bodian, Dale L; Schreiber, John M; Vilboux, Thierry; Khromykh, Alina; Hauser, Natalie S

2018-06-01

Infantile-onset epilepsies are a set of severe, heterogeneous disorders for which clinical genetic testing yields causative mutations in ∼20%-50% of affected individuals. We report the case of a boy presenting with intractable seizures at 2 wk of age, for whom gene panel testing was unrevealing. Research-based whole-genome sequencing of the proband and four unaffected family members identified a de novo mutation, NM_001323289.1:c.2828_2829delGA in CDKL5, a gene associated with X-linked early infantile epileptic encephalopathy 2. CDKL5 has multiple alternative transcripts, and the mutation lies in an exon in the brain-expressed forms. The mutation was undetected by gene panel sequencing because of its intronic location in the CDKL5 transcript typically used to define the exons of this gene for clinical exon-based tests (NM_003159). This is the first report of a patient with a mutation in an alternative transcript of CDKL5 This finding suggests that incorporating alternative transcripts into the design and variant interpretation of exon-based tests, including gene panel and exome sequencing, could improve the diagnostic yield. © 2018 Bodian et al.; Published by Cold Spring Harbor Laboratory Press.

The 5th Symposium on Post-Transcriptional Regulation of Plant Gene Expression (PTRoPGE)

Energy Technology Data Exchange (ETDEWEB)

Karen S. Browning; Marie Petrocek; Bonnie Bartel

2006-06-01

The 5th Symposium on Post-Transcriptional Regulation of Plant Gene Expression (PTRoPGE) will be held June 8-12, 2005 at the University of Texas at Austin. Exciting new and ongoing discoveries show significant regulation of gene expression occurs after transcription. These post-transcriptional control events in plants range from subtle regulation of transcribed genes and phosphorylation, to the processes of gene regulation through small RNAs. This meeting will focus on the regulatory role of RNA, from transcription, through translation and finally degradation. The cross-disciplinary design of this meeting is necessary to encourage interactions between researchers that have a common interest in post-transcriptional gene expression in plants. By bringing together a diverse group of plant molecular biologist and biochemists at all careers stages from across the world, this meeting will bring about more rapid progress in understanding how plant genomes work and how genes are finely regulated by post-transcriptional processes to ultimately regulate cells.

Transcriptional repression of BODENLOS by HD-ZIP transcription factor HB5 in Arabidopsis thaliana.

NARCIS (Netherlands)

Smet, De I.; Lau, S.; Ehrismann, J.S.; Axiotis, I.; Kolb, M.; Kientz, M.; Weijers, D.; Jürgens, G.

2013-01-01

In Arabidopsis thaliana, the phytohormone auxin is an important patterning agent during embryogenesis and post-embryonic development, exerting effects through transcriptional regulation. The main determinants of the transcriptional auxin response machinery are AUXIN RESPONSE FACTOR (ARF)

Tissue-specific 5' heterogeneity of PPARα transcripts and their differential regulation by leptin.

Directory of Open Access Journals (Sweden)

Emma S Garratt

Full Text Available The genes encoding nuclear receptors comprise multiple 5'untranslated exons, which give rise to several transcripts encoding the same protein, allowing tissue-specific regulation of expression. Both human and mouse peroxisome proliferator activated receptor (PPAR α genes have multiple promoters, although their function is unknown. Here we have characterised the rat PPARα promoter region and have identified three alternative PPARα transcripts, which have different transcription start sites owing to the utilisation of distinct first exons. Moreover these alternative PPARα transcripts were differentially expressed between adipose tissue and liver. We show that while the major adipose (P1 and liver (P2 transcripts were both induced by dexamethasone, they were differentially regulated by the PPARα agonist, clofibric acid, and leptin. Leptin had no effect on the adipose-specific P1 transcript, but induced liver-specific P2 promoter activity via a STAT3/Sp1 mechanism. Moreover in Wistar rats, leptin treatment between postnatal day 3-13 led to an increase in P2 but not P1 transcription in adipose tissue which was sustained into adulthood. This suggests that the expression of the alternative PPARα transcripts are in part programmed by early life exposure to leptin leading to persistent change in adipose tissue fatty acid metabolism through specific activation of a quiescent PPARα promoter. Such complexity in the regulation of PPARα may allow the expression of PPARα to be finely regulated in response to environmental factors.

cAMP response element binding protein (CREB activates transcription via two distinct genetic elements of the human glucose-6-phosphatase gene

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Stefano Luisa

2005-01-01

Full Text Available Abstract Background The enzyme glucose-6-phosphatase catalyzes the dephosphorylation of glucose-6-phosphatase to glucose, the final step in the gluconeogenic and glycogenolytic pathways. Expression of the glucose-6-phosphatase gene is induced by glucocorticoids and elevated levels of intracellular cAMP. The effect of cAMP in regulating glucose-6-phosphatase gene transcription was corroborated by the identification of two genetic motifs CRE1 and CRE2 in the human and murine glucose-6-phosphatase gene promoter that resemble cAMP response elements (CRE. Results The cAMP response element is a point of convergence for many extracellular and intracellular signals, including cAMP, calcium, and neurotrophins. The major CRE binding protein CREB, a member of the basic region leucine zipper (bZIP family of transcription factors, requires phosphorylation to become a biologically active transcriptional activator. Since unphosphorylated CREB is transcriptionally silent simple overexpression studies cannot be performed to test the biological role of CRE-like sequences of the glucose-6-phosphatase gene. The use of a constitutively active CREB2/CREB fusion protein allowed us to uncouple the investigation of target genes of CREB from the variety of signaling pathways that lead to an activation of CREB. Here, we show that this constitutively active CREB2/CREB fusion protein strikingly enhanced reporter gene transcription mediated by either CRE1 or CRE2 derived from the glucose-6-phosphatase gene. Likewise, reporter gene transcription was enhanced following expression of the catalytic subunit of cAMP-dependent protein kinase (PKA in the nucleus of transfected cells. In contrast, activating transcription factor 2 (ATF2, known to compete with CREB for binding to the canonical CRE sequence 5'-TGACGTCA-3', did not transactivate reporter genes containing CRE1, CRE2, or both CREs derived from the glucose-6-phosphatase gene. Conclusions Using a constitutively active CREB2

A broad but restricted requirement for TAF-5 (human TAFII100) for embryonic transcription in Caenorhabditis elegans.

Science.gov (United States)

Walker, Amy K; Blackwell, T Keith

2003-02-21

As conserved components of the transcription factor (TF) IID- and TFTC/SAGA-related complexes, TATA-binding protein-associated factors (TAF(II)s) are important for eukaryotic mRNA transcription. In yeast, genetic analyses suggest that, although some individual TAF(II)s are required for transcription of most genes, others have highly specialized functions. Much less is known about the functions of TAF(II)s in metazoans, which have more complex genomes that include many tissue-specific genes. TAF-5 (human (h) TAF(II)100) is of particular interest because it is predicted to have an important structural role. Here we describe the first genetics-based analysis of TAF-5 in a metazoan. By performing RNA interference in Caenorhabditis elegans embryos, which can survive for several cell generations without transcription, we found that taf-5 is important for a significant fraction of transcription. However, TAF-5 is apparently not essential for the expression of multiple developmental and other metazoan-specific genes. This phenotype remarkably resembles the previously described effects of similarly depleting two C. elegans histone fold TAF(II)s, TAF-9 (hTAF(II)31/32) and TAF-10 (hTAF(II)30), but is distinct from the widespread transcription block caused by TAF-4 (hTAF(II)130) depletion. Our findings suggest that TAF-5, TAF-9, and TAF-10 are part of a functional module of TFIID- and TFTC/SAGA-related complexes that can be bypassed in many metazoan-specific genes.

HOXB5 cooperates with NKX2-1 in the transcription of human RET.

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Jiang Zhu

Full Text Available The enteric nervous system (ENS regulates peristaltic movement of the gut, and abnormal ENS causes Hirschsprung's disease (HSCR in newborns. HSCR is a congenital complex genetic disorder characterised by a lack of enteric ganglia along a variable length of the intestine. The receptor tyrosine kinase gene (RET is the major HSCR gene and its expression is crucial for ENS development. We have previously reported that (i HOXB5 transcription factor mediates RET expression, and (ii mouse with defective HOXB5 activity develop HSCR phenotype. In this study, we (i elucidate the underlying mechanisms that HOXB5 mediate RET expression, and (ii examine the interactions between HOXB5 and other transcription factors implicated in RET expression. We show that human HOXB5 binds to the promoter region 5' upstream of the binding site of NKX2-1 and regulates RET expression. HOXB5 and NKX2-1 form a protein complex and mediate RET expression in a synergistic manner. HSCR associated SNPs at the NKX2-1 binding site (-5G>A rs10900296; -1A>C rs10900297, which reduce NKX2-1 binding, abolish the synergistic trans-activation of RET by HOXB5 and NKX2-1. In contrast to the synergistic activation of RET with NKX2-1, HOXB5 cooperates in an additive manner with SOX10, PAX3 and PHOX2B in trans-activation of RET promoter. Taken together, our data suggests that HOXB5 in coordination with other transcription factors mediates RET expression. Therefore, defects in cis- or trans-regulation of RET by HOXB5 could lead to reduction of RET expression and contribute to the manifestation of the HSCR phenotype.

The Fbw7 tumor suppressor targets KLF5 for ubiquitin-mediated degradation and suppresses breast cell proliferation.

Science.gov (United States)

Zhao, Dong; Zheng, Han-Qiu; Zhou, Zhongmei; Chen, Ceshi

2010-06-01

Fbw7 is a tumor suppressor frequently inactivated in cancers. The KLF5 transcription factor promotes breast cell proliferation and tumorigenesis through upregulating FGF-BP. The KLF5 protein degrades rapidly through the ubiquitin proteasome pathway. Here, we show that the Skp1-CUL1-Fbw7 E3 ubiquitin ligase complex (SCF(Fbw7)) targets KLF5 for ubiquitin-mediated degradation in a GSK3beta-mediated KLF5 phosphorylation-dependent manner. Mutation of the critical S303 residue in the KLF5 Cdc4 phospho-degrons motif ((303)SPPSS) abolishes the protein interaction, ubiquitination, and degradation by Fbw7. Inactivation of endogenous Fbw7 remarkably increases the endogenous KLF5 protein abundances. Endogenous Fbw7 suppresses the FGF-BP gene expression and breast cell proliferation through targeting KLF5 for degradation. These findings suggest that Fbw7 inhibits breast cell proliferation at least partially through targeting KLF5 for proteolysis. This new regulatory mechanism of KLF5 degradation may result in useful diagnostic and therapeutic targets for breast cancer and other cancers. Copyright 2010 AACR.

Quantitative transcriptional profiling of ATDC5 mouse progenitor cells during chondrogenesis

DEFF Research Database (Denmark)

Chen, Li; Fink, Trine; Zhang, Xiao-Yan

2005-01-01

During the differentiation of a mouse chondroprogenitor cell line, ATDC5, an analysis of the transcription cartilage-related genes was carried out using real-time RT-PCR in a semiquantitative fashion. A total number of 104 genes both previously linked to chondrogenesis and hitherto not associated...

Stress-induced alterations in 5-HT1A receptor transcriptional modulators NUDR and Freud-1.

Science.gov (United States)

Szewczyk, Bernadeta; Kotarska, Katarzyna; Daigle, Mireille; Misztak, Paulina; Sowa-Kucma, Magdalena; Rafalo, Anna; Curzytek, Katarzyna; Kubera, Marta; Basta-Kaim, Agnieszka; Nowak, Gabriel; Albert, Paul R

2014-11-01

The effect of stress on the mRNA and protein level of the 5-HT1A receptor and two of its key transcriptional modulators, NUDR and Freud-1, was examined in the prefrontal cortex (PFC) and hippocampus (Hp) using rodent models: olfactory bulbectomy (OB) and prenatal stress (PS) in male and female rats; chronic mild stress in male rats (CMS) and pregnancy stress. In PFC, CMS induced the most widespread changes, with significant reduction in both mRNA and protein levels of NUDR, 5-HT1A receptor and in Freud-1 mRNA; while in Hp 5-HT1A receptor and Freud-1 protein levels were also decreased. In male, but not female OB rats PFC Freud-1 and 5-HT1A receptor protein levels were reduced, while in Hp 5-HT1A receptor, Freud-1 and NUDR mRNA's but not protein were reduced. In PS rats PFC 5-HT1A receptor protein was reduced more in females than males; while in Hp Freud-1 protein was increased in females. In pregnancy stress, PFC NUDR, Freud-1 and 5-HT1A protein receptor levels were reduced, and in HP 5-HT1A receptor protein levels were also reduced; in HP only NUDR and Freud-1 mRNA levels were reduced. Overall, CMS and stress during pregnancy produced the most salient changes in 5-HT1A receptor and transcription factor expression, suggesting a primary role for altered transcription factor expression in chronic regulation of 5-HT1A receptor expression. By contrast, OB (in males) and PS (in females) produced gender-specific reductions in PFC 5-HT1A receptor protein levels, suggesting a role for post-transcriptional regulation. These and previous data suggest that chronic stress might be a key regulator of NUDR/Freud-1 gene expression.

Study on the binding sites of radiosensitivity associated transcription factor in the promoter region of Ier5 gene

International Nuclear Information System (INIS)

Cui Wei; Yin Lingling; Dong Lingyue

2012-01-01

Objective: To clarify the mechanism of immediate early response gene 5 (Ier5) transcription induced by radiation. Methods: Deletant construction, site-specific mutagenesis,electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) were used to forecast the promoter region, binding sites and transcription factors of Ier5 gene in HeLa cells. Results: The promoter region of Ier5 gene might be in the region of Ier5 -8 deletant (-408 - -238 bp). The Ier5 gene had two transcription factors of GCF and NFI, and GCF had two binding sites located in the region of -388 - -382 bp and -274 - -270 bp of Ier5 promoter. The binding site of NFI was located in -362 - -357 bp of Ier5 promoter. GCF could inhibit the expression of Ier5 gene and this inhibition was diminished when the radiation dose increased. In contrast, NFI increased the expression of Ier5. Conclusions: The most possible region of Ier5 promoter is from -408 to -238 bp which has two binding sites for the radiosensitivity transcription factors of GCF and NFI that could negatively and positively regulate the expression of Ier5 respectively. (authors)

Modulation of Epstein–Barr Virus Nuclear Antigen 2-dependent transcription by protein arginine methyltransferase 5

International Nuclear Information System (INIS)

Liu, Cheng-Der; Cheng, Chi-Ping; Fang, Jia-Shih; Chen, Ling-Chih; Zhao, Bo; Kieff, Elliott; Peng, Chih-Wen

2013-01-01

Highlights: ► Catalytic active PRMT5 substantially binds to the EBNA2 RG domain. ► PRMT5 augments the EBNA2-dependent transcription. ► PRMT5 triggers the symmetric dimethylation of the EBNA2 RG domain. ► PRMT5 enhances the promoter occupancy of EBNA2 on its target promoters. -- Abstract: Epstein–Barr Virus Nuclear Antigen (EBNA) 2 features an Arginine–Glycine repeat (RG) domain at amino acid positions 335–360, which is a known target for protein arginine methyltransferaser 5 (PRMT5). In this study, we performed protein affinity pull-down assays to demonstrate that endogenous PRMT5 derived from lymphoblastoid cells specifically associated with the protein bait GST-E2 RG. Transfection of a plasmid expressing PRMT5 induced a 2.5- to 3-fold increase in EBNA2-dependent transcription of both the LMP1 promoter in AKATA cells, which contain the EBV genome endogenously, and a Cp-Luc reporter plasmid in BJAB cells, which are EBV negative. Furthermore, we showed that there was a 2-fold enrichment of EBNA2 occupancy in target promoters in the presence of exogenous PRMT5. Taken together, we show that PRMT5 triggers the symmetric dimethylation of EBNA2 RG domain to coordinate with EBNA2-mediated transcription. This modulation suggests that PRMT5 may play a role in latent EBV infection

Modulation of Epstein–Barr Virus Nuclear Antigen 2-dependent transcription by protein arginine methyltransferase 5

Energy Technology Data Exchange (ETDEWEB)

Liu, Cheng-Der; Cheng, Chi-Ping; Fang, Jia-Shih; Chen, Ling-Chih [Department of Life Sciences, Tzu-Chi University, 701 Chung-Yang Rd. Sec 3, Hualien 97004, Taiwan (China); Zhao, Bo; Kieff, Elliott [Department of Medicine and Microbiology and Molecular Genetics, Channing Laboratory, Brigham and Women’s Hospital and Harvard Medical School, 181 Longwood Ave., Boston 02115, MA (United States); Peng, Chih-Wen, E-mail: pengcw@mail.tcu.edu.tw [Department of Life Sciences, Tzu-Chi University, 701 Chung-Yang Rd. Sec 3, Hualien 97004, Taiwan (China)

2013-01-18

Highlights: ► Catalytic active PRMT5 substantially binds to the EBNA2 RG domain. ► PRMT5 augments the EBNA2-dependent transcription. ► PRMT5 triggers the symmetric dimethylation of the EBNA2 RG domain. ► PRMT5 enhances the promoter occupancy of EBNA2 on its target promoters. -- Abstract: Epstein–Barr Virus Nuclear Antigen (EBNA) 2 features an Arginine–Glycine repeat (RG) domain at amino acid positions 335–360, which is a known target for protein arginine methyltransferaser 5 (PRMT5). In this study, we performed protein affinity pull-down assays to demonstrate that endogenous PRMT5 derived from lymphoblastoid cells specifically associated with the protein bait GST-E2 RG. Transfection of a plasmid expressing PRMT5 induced a 2.5- to 3-fold increase in EBNA2-dependent transcription of both the LMP1 promoter in AKATA cells, which contain the EBV genome endogenously, and a Cp-Luc reporter plasmid in BJAB cells, which are EBV negative. Furthermore, we showed that there was a 2-fold enrichment of EBNA2 occupancy in target promoters in the presence of exogenous PRMT5. Taken together, we show that PRMT5 triggers the symmetric dimethylation of EBNA2 RG domain to coordinate with EBNA2-mediated transcription. This modulation suggests that PRMT5 may play a role in latent EBV infection.

The bZIP protein from Tamarix hispida, ThbZIP1, is ACGT elements binding factor that enhances abiotic stress signaling in transgenic Arabidopsis.

Science.gov (United States)

Ji, Xiaoyu; Liu, Guifeng; Liu, Yujia; Zheng, Lei; Nie, Xianguang; Wang, Yucheng

2013-10-04

Tamarix spp. are woody halophyte, which are very tolerant to abiotic stresses such as salinity and drought, but little is known about their specific stress response systems. Basic leucine zipper proteins (bZIPs) play important roles in the ability of plants to withstand adverse environmental conditions. However, their exact roles in abiotic stress tolerance are still not fully known. In the current study, we functionally characterized a bZIP gene (ThbZIP1) from Tamarix hispida in response to abiotic stresses. We addressed the regulatory network of ThbZIP1 in three levels, i.e. its upstream regulators, the cis-acting elements recognized by ThbZIP1, and its downstream target genes. Two MYCs were found to bind to E-box, in the promoter of ThbZIP1 to activate its expression. Expression of ThbZIP1 is induced by ABA, salt, drought, methyl viologen and cold. ThbZIP1 can specifically bind to ACGT elements, with the highest binding affinity to the C-box, followed by the G-box and lastly the A-box. Compared with wild-type (Col-0) Arabidopsis, transgenic plants expressing ThbZIP1 had an increased tolerance to drought and salt, but had an increased sensitivity to ABA during seed germination and root growth; meanwhile, ROS level, cell death and water loss rate in transgenic plants were significantly reduced. Microarray analyses showed that many ROS scavenging genes were up-regulated by ThbZIP1 under salt stress conditions. Based on these data, we suggest that ThbZIP1 confers abiotic stress tolerance through activating stress tolerance genes to modulate ROS scavenging ability and other physiological changes involved in stress tolerance, and plays an important role in the ABA-mediated stress response of T. hispida.

The effect of stem cell factor on proliferation of human endometrial CD146+ cells

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Mehri Fayazi

2016-07-01

Full Text Available Background: Stem cell factor (SCF is a transcriptional factor which plays crucial roles in normal proliferation, differentiation and survival in a range of stem cells. Objective: The aim of the present study was to examine the proliferation effect of different concentrations of SCF on expansion of human endometrial CD146+ cells. Materials and Methods: In this experimental study, total populations of isolated human endometrial suspensions after fourth passage were isolated by magnetic activated cell sorting (MACS into CD146+ cells. Human endometrial CD146+ cells were karyotyped and tested for the effect of SCF on proliferation of CD146+ cells, then different concentrations of 0, 12.5, 25, 50 and 100 ng/ml was carried out and mitogens-stimulated endometrial CD146+ cells proliferation was assessed by MTT assay. Results: Chromosomal analysis showed a normal metaphase spread and 46XX karyotype. The proliferation rate of endometrial CD146P + P cells in the presence of 0, 12.5, 25, 50 and 100 ng/ml SCF were 0.945±0.094, 0.962±0.151, 0.988±0.028, 1.679±0.012 and 1.129±0.145 respectively. There was a significant increase in stem/ stromal cell proliferation following in vitro treatment by 50 ng/ml than other concentrations of SCF (p=0.01. Conclusion: The present study suggests that SCF could have effect on the proliferation and cell survival of human endometrial CD146P+P cells and it has important implications for medical sciences and cell therapies

A p300 and SIRT1 Regulated Acetylation Switch of C/EBPα Controls Mitochondrial Function

NARCIS (Netherlands)

Zaini, Mohamad A; Müller, Christine; de Jong, Tristan V; Ackermann, Tobias; Hartleben, Götz; Kortman, Gertrud; Gührs, Karl-Heinz; Fusetti, Fabrizia; Krämer, Oliver H; Guryev, Victor; Calkhoven, Cornelis F

2018-01-01

Cellular metabolism is a tightly controlled process in which the cell adapts fluxes through metabolic pathways in response to changes in nutrient supply. Among the transcription factors that regulate gene expression and thereby cause changes in cellular metabolism is the basic leucine-zipper (bZIP)

An ABRE promoter sequence is involved in osmotic stress-responsive expression of the DREB2A gene, which encodes a transcription factor regulating drought-inducible genes in Arabidopsis.

Science.gov (United States)

Kim, June-Sik; Mizoi, Junya; Yoshida, Takuya; Fujita, Yasunari; Nakajima, Jun; Ohori, Teppei; Todaka, Daisuke; Nakashima, Kazuo; Hirayama, Takashi; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

2011-12-01

In plants, osmotic stress-responsive transcriptional regulation depends mainly on two major classes of cis-acting elements found in the promoter regions of stress-inducible genes: ABA-responsive elements (ABREs) and dehydration-responsive elements (DREs). ABRE has been shown to perceive ABA-mediated osmotic stress signals, whereas DRE is known to be involved in an ABA-independent pathway. Previously, we reported that the transcription factor DRE-BINDING PROTEIN 2A (DREB2A) regulates DRE-mediated transcription of target genes under osmotic stress conditions in Arabidopsis (Arabidopsis thaliana). However, the transcriptional regulation of DREB2A itself remains largely uncharacterized. To elucidate the transcriptional mechanism associated with the DREB2A gene under osmotic stress conditions, we generated a series of truncated and base-substituted variants of the DREB2A promoter and evaluated their transcriptional activities individually. We found that both ABRE and coupling element 3 (CE3)-like sequences located approximately -100 bp from the transcriptional initiation site are necessary for the dehydration-responsive expression of DREB2A. Coupling our transient expression analyses with yeast one-hybrid and chromatin immunoprecipitation (ChIP) assays indicated that the ABRE-BINDING PROTEIN 1 (AREB1), AREB2 and ABRE-BINDING FACTOR 3 (ABF3) bZIP transcription factors can bind to and activate the DREB2A promoter in an ABRE-dependent manner. Exogenous ABA application induced only a modest accumulation of the DREB2A transcript when compared with the osmotic stress treatment. However, the osmotic stress-induced DREB2A expression was found to be markedly impaired in several ABA-deficient and ABA-insensitive mutants. These results suggest that in addition to an ABA-independent pathway, the ABA-dependent pathway plays a positive role in the osmotic stress-responsive expression of DREB2A.

Tissue-Specific 5′ Heterogeneity of PPARα Transcripts and Their Differential Regulation by Leptin

Science.gov (United States)

Garratt, Emma S.; Vickers, Mark H.; Gluckman, Peter D.; Hanson, Mark A.

2013-01-01

The genes encoding nuclear receptors comprise multiple 5′untranslated exons, which give rise to several transcripts encoding the same protein, allowing tissue-specific regulation of expression. Both human and mouse peroxisome proliferator activated receptor (PPAR) α genes have multiple promoters, although their function is unknown. Here we have characterised the rat PPARα promoter region and have identified three alternative PPARα transcripts, which have different transcription start sites owing to the utilisation of distinct first exons. Moreover these alternative PPARα transcripts were differentially expressed between adipose tissue and liver. We show that while the major adipose (P1) and liver (P2) transcripts were both induced by dexamethasone, they were differentially regulated by the PPARα agonist, clofibric acid, and leptin. Leptin had no effect on the adipose-specific P1 transcript, but induced liver-specific P2 promoter activity via a STAT3/Sp1 mechanism. Moreover in Wistar rats, leptin treatment between postnatal day 3–13 led to an increase in P2 but not P1 transcription in adipose tissue which was sustained into adulthood. This suggests that the expression of the alternative PPARα transcripts are in part programmed by early life exposure to leptin leading to persistent change in adipose tissue fatty acid metabolism through specific activation of a quiescent PPARα promoter. Such complexity in the regulation of PPARα may allow the expression of PPARα to be finely regulated in response to environmental factors. PMID:23825665

Isolation and expression analysis of EcbZIP17 from different finger millet genotypes shows conserved nature of the gene.

Science.gov (United States)

Chopperla, Ramakrishna; Singh, Sonam; Mohanty, Sasmita; Reddy, Nanja; Padaria, Jasdeep C; Solanke, Amolkumar U

2017-10-01

Basic leucine zipper (bZIP) transcription factors comprise one of the largest gene families in plants. They play a key role in almost every aspect of plant growth and development and also in biotic and abiotic stress tolerance. In this study, we report isolation and characterization of EcbZIP17 , a group B bZIP transcription factor from a climate smart cereal, finger millet ( Eleusine coracana L.). The genomic sequence of EcbZIP17 is 2662 bp long encompassing two exons and one intron with ORF of 1722 bp and peptide length of 573 aa. This gene is homologous to AtbZIP17 ( Arabidopsis ), ZmbZIP17 (maize) and OsbZIP60 (rice) which play a key role in endoplasmic reticulum (ER) stress pathway. In silico analysis confirmed the presence of basic leucine zipper (bZIP) and transmembrane (TM) domains in the EcbZIP17 protein. Allele mining of this gene in 16 different genotypes by Sanger sequencing revealed no variation in nucleotide sequence, including the 618 bp long intron. Expression analysis of EcbZIP17 under heat stress exhibited similar pattern of expression in all the genotypes across time intervals with highest upregulation after 4 h. The present study established the conserved nature of EcbZIP17 at nucleotide and expression level.

Epigenetic mediated transcriptional activation of WNT5A participates in arsenical-associated malignant transformation

International Nuclear Information System (INIS)

Jensen, Taylor J.; Wozniak, Ryan J.; Eblin, Kylee E.; Wnek, Sean M.; Gandolfi, A. Jay; Futscher, Bernard W.

2009-01-01

Arsenic is a human carcinogen with exposure associated with cancer of the lung, skin, and bladder. Many potential mechanisms have been implicated as playing a role in the process of arsenical-induced malignancy including the perturbation of signaling pathways and aberrant epigenetic regulation. We initiated studies to examine the role of a member of the non-canonical WNT signaling pathway, WNT5A, in UROtsa cells and arsenite [URO-ASSC] and monomethylarsonous acid [URO-MSC] malignantly transformed variants. We present data herein that suggest that WNT5A is transcriptionally activated during arsenical-induced malignant transformation. This WNT5A transcriptional activation is correlated with the enrichment of permissive histone modifications and the reduction of repressive modifications in the WNT5A promoter region. The epigenetic activation of WNT5A expression and acetylation of its promoter remain after the removal of the arsenical, consistent with the maintenance of an anchorage independent growth phenotype in these cells. Additionally, treatment with epigenetic modifying drugs supports a functional role for these epigenetic marks in controlling gene expression. Reduction of WNT5A using lentiviral shRNA greatly attenuated the ability of these cells to grow in an anchorage independent fashion. Extension of our model into human bladder cancer cell lines indicates that each of the cell lines examined also express WNT5A. Taken together, these data suggest that the epigenetic remodeling of the WNT5A promoter is correlated with its transcriptional activation and this upregulation likely participates in arsenical-induced malignant transformation

Genome-wide investigation of transcription factors provides insights into transcriptional regulation in Plutella xylostella.

Science.gov (United States)

Zhao, Qian; Ma, Dongna; Huang, Yuping; He, Weiyi; Li, Yiying; Vasseur, Liette; You, Minsheng

2018-04-01

Transcription factors (TFs), which play a vital role in regulating gene expression, are prevalent in all organisms and characterization of them may provide important clues for understanding regulation in vivo. The present study reports a genome-wide investigation of TFs in the diamondback moth, Plutella xylostella (L.), a worldwide pest of crucifers. A total of 940 TFs distributed among 133 families were identified. Phylogenetic analysis of insect species showed that some of these families were found to have expanded during the evolution of P. xylostella or Lepidoptera. RNA-seq analysis showed that some of the TF families, such as zinc fingers, homeobox, bZIP, bHLH, and MADF_DNA_bdg genes, were highly expressed in certain tissues including midgut, salivary glands, fat body, and hemocytes, with an obvious sex-biased expression pattern. In addition, a number of TFs showed significant differences in expression between insecticide susceptible and resistant strains, suggesting that these TFs play a role in regulating genes related to insecticide resistance. Finally, we identified an expansion of the HOX cluster in Lepidoptera, which might be related to Lepidoptera-specific evolution. Knockout of this cluster using CRISPR/Cas9 showed that the egg cannot hatch, indicating that this cluster may be related to egg development and maturation. This is the first comprehensive study on identifying and characterizing TFs in P. xylostella. Our results suggest that some TF families are expanded in the P. xylostella genome, and these TFs may have important biological roles in growth, development, sexual dimorphism, and resistance to insecticides. The present work provides a solid foundation for understanding regulation via TFs in P. xylostella and insights into the evolution of the P. xylostella genome.

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.

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.

Identification of SFBB-containing canonical and noncanonical SCF complexes in pollen of apple (Malus × domestica).

Science.gov (United States)

Minamikawa, Mai F; Koyano, Ruriko; Kikuchi, Shinji; Koba, Takato; Sassa, Hidenori

2014-01-01

Gametophytic self-incompatibility (GSI) of Rosaceae, Solanaceae and Plantaginaceae is controlled by a single polymorphic S locus. The S locus contains at least two genes, S-RNase and F-box protein encoding gene SLF/SFB/SFBB that control pistil and pollen specificity, respectively. Generally, the F-box protein forms an E3 ligase complex, SCF complex with Skp1, Cullin1 (CUL1) and Rbx1, however, in Petunia inflata, SBP1 (S-RNase binding protein1) was reported to play the role of Skp1 and Rbx1, and form an SCFSLF-like complex for ubiquitination of non-self S-RNases. On the other hand, in Petunia hybrida and Petunia inflata of Solanaceae, Prunus avium and Pyrus bretschneideri of Rosaceae, SSK1 (SLF-interacting Skp1-like protein1) is considered to form the SCFSLF/SFB complex. Here, we isolated pollen-expressed apple homologs of SSK1 and CUL1, and named MdSSK1, MdCUL1A and MdCUL1B. MdSSK1 was preferentially expressed in pollen, but weakly in other organs analyzed, while, MdCUL1A and MdCUL1B were almost equally expressed in all the organs analyzed. MdSSK1 transcript abundance was significantly (>100 times) higher than that of MdSBP1. In vitro binding assays showed that MdSSK1 and MdSBP1 interacted with MdSFBB1-S9 and MdCUL1, and MdSFBB1-S9 interacted more strongly with MdSSK1 than with MdSBP1. The results suggest that both MdSSK1-containing SCFSFBB1 and MdSBP1-containing SCFSFBB1-like complexes function in pollen of apple, and the former plays a major role.

Model of how plants sense zinc deficiency

DEFF Research Database (Denmark)

Assuncao, Ana G.L.; Persson, Daniel Olof; Husted, Søren

2013-01-01

to develop plant-based solutions addressing nutrient-use-efficiency and adaptation to nutrient-limited or -toxic soils. Recently two transcription factors of the bZIP family (basic-region leucine zipper) have been identified in Arabidopsis and shown to be pivotal in the adaptation response to zinc deficiency...

Identification of the Drosophila skpA gene as a novel target of the transcription factor DREF

International Nuclear Information System (INIS)

Dang Thi Phuong Thao; Ida, Hiroyuki; Yoshida, Hideki; Yamaguchi, Masamitsu

2006-01-01

SKPa is component of a Drosophila SCF complex that functions in combination with the ubiquitin-conjugating enzyme UbcD1. skpA null mutation results in centrosome overduplication, unusual chromatin condensation, defective endoreduplication and cell-cycle progression. While the molecular mechanisms that regulate expression of the skpA gene are poorly understood, the DNA replication-related element (DRE) and the DRE-binding factor (DREF) play important roles in regulating proliferation-related genes in Drosophila and DRE (5'-TATCGATA) and DRE-like (5'-CATCGATT) sequences were here found to be involved in skpA promoter activity. Thus both luciferase transient expression assays in cultured Drosophila S2 cells using skpA promoter-luciferase fusion plasmids and anti-lacZ immunostaining of various tissues from transgenic third instar larvae carrying the skpA promoter-lacZ fusion genes provided supportive evidence. Furthermore, anti-SKPa immunostaining of eye imaginal discs from flies overexpressing DREF showed ectopic expression of protein in the region posterior to the morphogenetic furrow where DREF is overexpressed. Knockdown of DREF in some tissues where SKPa distribution is well known almost completely abrogated the skpA gene expression. These findings, taken together, indicate that the Drosophila skpA gene is a novel target of the transcription factor DREF

Activator Protein-1: redox switch controlling structure and DNA-binding

Energy Technology Data Exchange (ETDEWEB)

Yin, Zhou; Machius, Mischa; Nestler, Eric J.; Rudenko, Gabby (Texas-MED); (Icahn)

2017-09-07

The transcription factor, activator protein-1 (AP-1), binds to cognate DNA under redox control; yet, the underlying mechanism has remained enigmatic. A series of crystal structures of the AP-1 FosB/JunD bZIP domains reveal ordered DNA-binding regions in both FosB and JunD even in absence DNA. However, while JunD is competent to bind DNA, the FosB bZIP domain must undergo a large conformational rearrangement that is controlled by a ‘redox switch’ centered on an inter-molecular disulfide bond. Solution studies confirm that FosB/JunD cannot undergo structural transition and bind DNA when the redox-switch is in the ‘OFF’ state, and show that the mid-point redox potential of the redox switch affords it sensitivity to cellular redox homeostasis. The molecular and structural studies presented here thus reveal the mechanism underlying redox-regulation of AP-1 Fos/Jun transcription factors and provide structural insight for therapeutic interventions targeting AP-1 proteins.

Relação da expressão de fatores de crescimento celular (IGF-1 e (SCF com fatores prognósticos e o alvo da rapamicina em mamíferos (m-TOR em mastocitomas cutâneos caninos IGF-1 and SCF protein expression in cutaneous mast cell tumors in dogs and relation to prognostic factors and mammalian target of rapamycin (m-TOR

Directory of Open Access Journals (Sweden)

Raquel B. Ferioli

2013-04-01

Full Text Available O mastocitoma cutâneo (MTC é a neoplasia maligna mais comum na pele dos cães e seu comportamento biológico é muito variável. Dentre os fatores prognósticos estudados nos MTCs, a classificação histopatológica, o índice proliferativo e o padrão de expressão doc-KIT são os que apresentam uma associação mais relevante com o provável prognóstico deste tumor. O objetivo deste trabalho foi avaliar a expressão proteica de fator de crescimento semelhante à insulina tipo 1 (IGF-1, fator de célula tronco (SCF e sua relação com o receptor tirosina quinase (c-KIT, alvo da rapamicina em mamíferos (m-TOR, grau histológico, índice proliferativo pelo KI-67e o número de figuras de mitose (IM com dados clínicos de cães com MTCs . Foram utilizadas 133 amostras de MTCs, provenientes de 133 cães, dispostas em lâminas de microarranjo de tecidos (TMA. A técnica de imuno-histoquímica foi utilizada para a avaliação destas proteínas. Observou-se associação entre SCF e, a graduação histopatológica proposta em 2011, índice mitótico, proliferação celular (KI-67, escore de IGF-1, local da lesão, idade dos animais e padrão imuno-histoquímico do receptor c-KIT. A relação de dependência também foi observada entre IGF-1 e o porte dos animais, IM, m-TOR e c-KIT. A expressão de SCF teve relacção com a agressividade dos MTCs caninos, uma vez que foi mais freqüente em MTCs com c-KIT citoplasmático. A relação entre a expressão de IGF-1, SCF, c-KIT e m-TOR pode estar associada à integralização de suas vias de ação. A expressão de IGF-1 está associada à MTCs em cães de porte grande.Cutaneous mast cell tumor (MCT is one of the most common neoplasms in the skin of dogs and express variable biological behavior. Among the MTC aspects studied, histological classification, proliferative index and protein expression of c-KIT show the most defined connection with the tumor prognostic. The aim of this study was to evaluate the

The positive transcription factor of the 5S RNA gene proteolyses during direct exchange between 5S DNA sites

OpenAIRE

1986-01-01

We have examined the association, dissociation, and exchange of the 5S specific transcription factor (TFIIIA) with somatic- and oocyte-type 5S DNA. The factor associates faster with somatic than with oocyte 5S DNA, and the rate of complex formation is accelerated by vector DNA. Once formed, the TFIIIA-5S DNA complex is stable for greater than 4 h in the absence of free 5S DNA, and its dissociation is identical for somatic and for oocyte 5S DNA. In the presence of free 5S DNA, the factor trans...

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

CNDO/2-SCF and PCILO (MO) calculations on the 1-butene/NA/and (charge-transfer) complex

Energy Technology Data Exchange (ETDEWEB)

Lochmann, R; Meiler, W

1977-01-01

CNDO/2-SCF and PCILO (MO) calculations on the 1-Butene/Na/sup +/ (charge-transfer) complex involving the olefinic m electrons were made in connection with butene adsorption in zeolites, including the effect of the cation on the conformation of the butene in the zeolite cavity. Calculations were made of rotational energy barriers, preferred cation arrangements with respect to the butene molecule, and charge distributions by both methods. Taking into account systematic errors with the two methods, it is concluded that the PCILO method, which predicts a stabilization of the skew over the cis conformation by the cation, gives closer agreement with experiment. Graph, tables, diagrams, and 19 references.

Methylation associated transcriptional repression of ELOVL5 in novel colorectal cancer cell lines.

Directory of Open Access Journals (Sweden)

Arnoud Boot

Full Text Available Genetic and epigenetic alterations mark colorectal cancer (CRC. Global hypomethylation is observed in nearly all CRC, but a distinct subset of CRC show the CpG Island Methylator Phenotype (CIMP. These tumors show DNA hypermethylation of a specific subset of CpG islands, resulting in transcriptional downregulation of nearby genes. Recently we reported the establishment of novel CRC cell lines derived from primary and metastatic CRC tissues. In this study we describe the DNA methylation profiling of these low passage CRC cell lines. We generated global DNA methylation profiles with Infinium HumanMethylation450 BeadChips and analysed them in conjunction with matching gene expression profiles. Multidimensional scaling of the DNA methylation and gene expression datasets showed that BRAF mutated cell lines form a distinct group. In this group we investigated the 706 loci which we have previously identified to be hypermethylated in BRAF mutant CRC. We validated the significant findings in the The Cancer Genome Atlas colon adenocarcinoma dataset. Our analysis identified ELOVL5, FAM127B, MTERF1, ZNF606 to be subject to transcriptional downregulation through DNA hypermethylation in CRC. We further investigated ELOVL5 with qPCR and immunohistochemical staining, validating our results, but did not find a clear relation between ELOVL5 expression and tumor stage or relapse free survival. ELOVL5, FAM127B, MTERF1, ZNF606 are involved in important cellular processes such as apoptosis, lipogenesis and the downstream transcriptional effect of the MAPK-pathway. We have identified a DNA methylation profile regulating key cellular processes in CRC, resulting in a growth advantage to the tumor cells.

Regulation of stem cell factor expression in inflammation and asthma

Directory of Open Access Journals (Sweden)

Carla A Da Silva

2005-03-01

Full Text Available Stem cell factor (SCF is a major mast cell growth factor, which could be involved in the local increase of mast cell number in the asthmatic airways. In vivo, SCF expression increases in asthmatic patients and this is reversed after treatment with glucocorticoids. In vitro in human lung fibroblasts in culture, IL-1beta, a pro-inflammatory cytokine, confirms this increased SCF mRNA and protein expression implying the MAP kinases p38 and ERK1/2 very early post-treatment, and glucocorticoids confirm this decrease. Surprisingly, glucocorticoids potentiate the IL-1beta-enhanced SCF expression at short term treatment, implying increased SCF mRNA stability and SCF gene transcription rate. This potentiation involves p38 and ERK1/2. Transfection experiments with the SCF promoter including intron1 also confirm this increase and decrease of SCF expression by IL-1beta and glucocorticoids, and the potentiation by glucocorticoids of the IL-1beta-induced SCF expression. Deletion of the GRE or kappaB sites abolishes this potentiation, and the effect of IL-1beta or glucocorticoids alone. DNA binding of GR and NF-kappaB are also demonstrated for these effects. In conclusion, this review concerns new mechanisms of regulation of SCF expression in inflammation that could lead to potential therapeutic strategy allowing to control mast cell number in the asthmatic airways.

Involvement of phosphatidylinositol 4,5-bisphosphate in RNA polymerase I transcription

Czech Academy of Sciences Publication Activity Database

Yildirim, Sukriye; Castano, Enrique; Sobol, Margaryta; Philimonenko, Vlada; Dzijak, Rastislav; Venit, Tomáš; Hozák, Pavel

2013-01-01

Roč. 126, č. 12 (2013), s. 2730-2739 ISSN 0021-9533 R&D Projects: GA ČR GAP305/11/2232; GA ČR(CZ) GD204/09/H084; GA MŠk LC545; GA MŠk(CZ) LC06063 Grant - others:CONACYT(MX) 176598 Institutional support: RVO:68378050 Keywords : nucleolus * transcription * PIP2 * UBF * fibrillarin Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.325, year: 2013

Differential transcription and message stability of two genes encoding soybean ribulose 1,5-bisphosphate carboxylase small subunit

International Nuclear Information System (INIS)

Shirley, B.W.; Berry-Lowe, S.L.; Grandbastien, M.A.; Zurfluh, L.L.; Shah, D.M.; Meagher, R.B.

1987-01-01

The expression of two closely related soybean ribulose bisphosphate carboxylase small subunit (Rubisco ss) genes, SRS1 and SRS4, has been compared. These genes account for approximately 2-4% of the total transcription in light grown leaves, SRS4 being twice as transcriptionally active as SRS1. The transcription of these genes is reduced more than 30 fold after a pulse of far-red light or extended periods of darkness. When etiolated seedlings are shifted to the light the transcription of both genes increases 30-50 fold. Despite this 30-fold range in transcriptional expression the steady state mRNA levels in light and dark grown tissue differ by less than 8 fold. This suggests that the mRNAs are less stable in light grown tissue. 38 refs., 5 figs

Direct Regulation of tRNA and 5S rRNA Gene Transcription by Polo-like Kinase 1

NARCIS (Netherlands)

Fairley, Jennifer A.; Mitchell, Louise E.; Berg, Tracy; Kenneth, Niall S.; von Schubert, Conrad; Sillje, Herman H. W.; Medema, Rene H.; Nigg, Erich A.; White, Robert J.

2012-01-01

Polo-like kinase Plk1 controls numerous aspects of cell-cycle progression. We show that it associates with tRNA and 5S rRNA genes and regulates their transcription by RNA polymerase Ill (pol Ill) through direct binding and phosphorylation of transcription factor Brit During interphase, Plk1 promotes

LnqR, a TetR-family transcriptional regulator, positively regulates lacticin Q production in Lactococcus lactis QU 5.

Science.gov (United States)

Iwatani, Shun; Ishibashi, Naoki; Flores, Floirendo P; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

2016-09-01

Lacticin Q is an unmodified leaderless bacteriocin produced by Lactococcus lactis QU 5. It has been revealed that the production and self-immunity of lacticin Q are facilitated by a gene cluster lnqQBCDEF The gene for a putative TetR-family transcriptional regulator, termed lnqR, was found nearby the lnqQBCDEF cluster, but its involvement in lacticin Q biosynthesis remained unknown. In this study, we created an LnqR-overexpressing QU 5 recombinant by using lactococcal constitutive promoter P32 The recombinant QU 5 showed enhanced production of and self-immunity to lacticin Q. RT-PCR analysis has revealed that an overexpression of LnqR increases the amounts of lnqQBCDEF transcripts, and these six genes are transcribed as an operon in a single transcriptional unit. Interestingly, LnqR expression and thus lacticin Q production by L. lactis QU 5 was found temperature dependent, while LnzR, an LnqR-homologue, in L. lactis QU 14 was expressed in a similar but not identical manner to LnqR, resulting in dissimilar bacteriocin productivities by these strains. This report demonstrates LnqR as the first TetR-family transcriptional regulator involved in LAB bacteriocin biosynthesis and that, as an exceptional case of TetR-family regulators, LnqR positively regulates the transcription of these biosynthetic genes. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Wnt-mediated down-regulation of Sp1 target genes by a transcriptional repressor Sp5

Czech Academy of Sciences Publication Activity Database

Fujimura, Naoko; Vacík, Tomáš; Machoň, Ondřej; Vlček, Čestmír; Scalabrin, S.; Speth, M.; Diep, D.; Krauss, S.; Kozmik, Zbyněk

2007-01-01

Roč. 282, č. 2 (2007), s. 1225-1237 ISSN 0021-9258 Institutional research plan: CEZ:AV0Z50520514 Keywords : Wnt -mediated signaling * Sp5 transcription factor * Sp1 target genes Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.581, year: 2007

RNF11 is a multifunctional modulator of growth factor receptor signalling and transcriptional regulation.

Science.gov (United States)

Azmi, Peter; Seth, Arun

2005-11-01

Our laboratory has found that the 154aa RING finger protein 11 (RNF11), has modular domains and motifs including a RING-H2 finger domain, a PY motif, an ubiquitin interacting motif (UIM), a 14-3-3 binding sequence and an AKT phosphorylation site. RNF11 represents a unique protein with no other known immediate family members yet described. Comparative genetic analysis has shown that RNF11 is highly conserved throughout evolution. This may indicate a conserved and non-redundant role for the RNF11 protein. Molecular binding assays using RNF11 have shown that RNF11 has important roles in growth factor signalling, ubiquitination and transcriptional regulation. RNF11 has been shown to interact with HECT-type E3 ubiquitin ligases Nedd4, AIP4, Smurf1 and Smurf2, as well as with Cullin1, the core protein in the multi-subunit SCF E3 ubiquitin ligase complex. Work done in our laboratory has shown that RNF11 is capable of antagonizing Smurf2-mediated inhibition of TGFbeta signalling. Furthermore, RNF11 is capable of degrading AMSH, a positive regulator of both TGFbeta and EGFR signalling pathways. Recently, we have found that RNF11 can directly enhance TGFbeta signalling through a direct association with Smad4, the common signal transducer and transcription factor in the TGFbeta, BMP, and Activin pathways. Through its association with Smad4 and other transcription factors, RNF11 may have a role in direct transcriptional regulation. Our laboratory and others have found nearly 80 protein interactions for RNF11, placing RNF11 at the cross-roads of cell signalling and transcriptional regulation. RNF11 is highly expressed in breast tumours. Deregulation of RNF11 function may prove to be harmful to patient therapeutic outcomes. RNF11 may therefore provide a novel target for cancer therapeutics. The purpose of this review is to discuss the role of RNF11 in cell signalling and transcription factor modulation with special attention given to the ubiquitin-proteasomal pathway, TGFbeta

Characterization and functional analysis of four HYH splicing variants in Arabidopsis hypocotyl elongation.

Science.gov (United States)

Li, Chen; Zheng, Lanlan; Zhang, Jingxuan; Lv, Yanxia; Liu, Jianping; Wang, Xuanbin; Palfalvi, Gergo; Wang, Guodong; Zhang, Yonghong

2017-07-01

Arabidopsis thaliana LONG HYPOCOTYL5 (HY5) is a positive regulator of the light signaling pathway. The hy5 mutant has an elongated hypocotyl in all light conditions, whereas the hy5 homolog (hyh) mutant has a very weak phenotype, but only in blue light. However, overexpression of HYH rescues the elongated hypocotyl phenotype in the hy5 null mutant. Here, we report the identification of four HYH splicing variants in Arabidopsis. Alternative splicing in the 5' region of the HYH gene occurred such that the proteins encoded by all four HYH variants retained their bZIP domain. In hypocotyl tissue, transcript levels of HYH.2, HYH.3, and HYH.4 were higher than those of HYH.1. Like HY5, all HYH variants were induced by light. Functional analysis of the four HYH variants, based on their abilities to complement the hy5 mutant, indicated that they have similar roles in hypocotyl development, and may function redundantly with HY5. Our results indicate that the bZIP domain in HYH is critical for the function of four variants in the compensation of hy5 mutant in hypocotyl development. Additionally, while HY5/HYH is found in plant species ranging from green algae to flowering plants, the potential alternative splicing events are distinct in different species, with certain HYH variants found with greater frequency in some species than others. Copyright © 2017. Published by Elsevier B.V.

Activator Protein-1: redox switch controlling structure and DNA-binding.

Science.gov (United States)

Yin, Zhou; Machius, Mischa; Nestler, Eric J; Rudenko, Gabby

2017-11-02

The transcription factor, activator protein-1 (AP-1), binds to cognate DNA under redox control; yet, the underlying mechanism has remained enigmatic. A series of crystal structures of the AP-1 FosB/JunD bZIP domains reveal ordered DNA-binding regions in both FosB and JunD even in absence DNA. However, while JunD is competent to bind DNA, the FosB bZIP domain must undergo a large conformational rearrangement that is controlled by a 'redox switch' centered on an inter-molecular disulfide bond. Solution studies confirm that FosB/JunD cannot undergo structural transition and bind DNA when the redox-switch is in the 'OFF' state, and show that the mid-point redox potential of the redox switch affords it sensitivity to cellular redox homeostasis. The molecular and structural studies presented here thus reveal the mechanism underlying redox-regulation of AP-1 Fos/Jun transcription factors and provide structural insight for therapeutic interventions targeting AP-1 proteins. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Pulse radiolysis and ab initio SCF MO studies of hydroxyl radical reactions with 2,2'-bipyridine and its complexes with transition metal ions

Energy Technology Data Exchange (ETDEWEB)

Maliyachel, A C

1984-01-01

In the present study, reactions of hydroxyl radical with 2,2'-bipyridine (bpy) and complexes of iron(II) and cobalt(III) containing 2,2'-bipyridine and/or cyanide as ligands have been investigated by pulse radiolysis and also by ab initio self-consistent field, molecular orbital (SCF MO) theoretical techniques for 2,2'-bipyridine and pyridines. In the pulse radiolysis experiments, the nascent products of hydroxyl radical reactions with these compounds have been characterized through their spectral and kinetic properties. All these reactions occur at near diffusion controlled rates to give transient products having absorption in the ultraviolet, visible and, in some cases, near-IR region. The primary reactions of OH are considered to take place by addition mechanisms in the cases of 2,2'-bipyridine, (Fe(bpy)/sub 3/)/sup 2 +/, (Fe(DMbpy)/sub 3/)/sup 2 +/ and (Co(bpy)/sub 3/)/sup 3 +/. With (Fe(pby)/sub 2/(CN)/sub 2/) and (Fe(bpy)(CN)/sub 4/)/sup 2 -/, both addition and charge transfer processes occur. The present study indicates that hydroxyl radical reactions with 2,2'-bipyridine can be considerably altered by complexation with metal ions such as iron(II) and cobalt(III), and the factors associated with this are discussed. In the second part of this work, ab initio SCF MO calculations have been performed for the reactions of OH with pyridine, pyridinium ion and 2,2'-bipyridine. Based on the calculated total energies for the various hydroxy radical products, the relative stability of OH addition products are found to be for pyridine, meta-C > N >> para-C > ortho-C; for pyridinium ion, meta-C >> para-C > ortho-C > N, and for 2,2'- bipyridine, C/sub 5/ > C/sub 6/ > C/sub 3/ > C/sub 4/ > N.

Abscisic acid affects transcription of chloroplast genes via protein phosphatase 2C-dependent activation of nuclear genes: repression by guanosine-3'-5'-bisdiphosphate and activation by sigma factor 5.

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Yamburenko, Maria V; Zubo, Yan O; Börner, Thomas

2015-06-01

Abscisic acid (ABA) represses the transcriptional activity of chloroplast genes (determined by run-on assays), with the exception of psbD and a few other genes in wild-type Arabidopsis seedlings and mature rosette leaves. Abscisic acid does not influence chloroplast transcription in the mutant lines abi1-1 and abi2-1 with constitutive protein phosphatase 2C (PP2C) activity, suggesting that ABA affects chloroplast gene activity by binding to the pyrabactin resistance (PYR)/PYR1-like or regulatory component of ABA receptor protein family (PYR/PYL/RCAR) and signaling via PP2Cs and sucrose non-fermenting protein-related kinases 2 (SnRK2s). Further we show by quantitative PCR that ABA enhances the transcript levels of RSH2, RSH3, PTF1 and SIG5. RelA/SpoT homolog 2 (RSH2) and RSH3 are known to synthesize guanosine-3'-5'-bisdiphosphate (ppGpp), an inhibitor of the plastid-gene-encoded chloroplast RNA polymerase. We propose, therefore, that ABA leads to an inhibition of chloroplast gene expression via stimulation of ppGpp synthesis. On the other hand, sigma factor 5 (SIG5) and plastid transcription factor 1 (PTF1) are known to be necessary for the transcription of psbD from a specific light- and stress-induced promoter (the blue light responsive promoter, BLRP). We demonstrate that ABA activates the psbD gene by stimulation of transcription initiation at BLRP. Taken together, our data suggest that ABA affects the transcription of chloroplast genes by a PP2C-dependent activation of nuclear genes encoding proteins involved in chloroplast transcription. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

Bacteriophage T5 encodes a homolog of the eukaryotic transcription coactivator PC4 implicated in recombination-dependent DNA replication.

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Steigemann, Birthe; Schulz, Annina; Werten, Sebastiaan

2013-11-15

The RNA polymerase II cofactor PC4 globally regulates transcription of protein-encoding genes through interactions with unwinding DNA, the basal transcription machinery and transcription activators. Here, we report the surprising identification of PC4 homologs in all sequenced representatives of the T5 family of bacteriophages, as well as in an archaeon and seven phyla of eubacteria. We have solved the crystal structure of the full-length T5 protein at 1.9Å, revealing a striking resemblance to the characteristic single-stranded DNA (ssDNA)-binding core domain of PC4. Intriguing novel structural features include a potential regulatory region at the N-terminus and a C-terminal extension of the homodimerisation interface. The genome organisation of T5-related bacteriophages points at involvement of the PC4 homolog in recombination-dependent DNA replication, strongly suggesting that the protein corresponds to the hitherto elusive replicative ssDNA-binding protein of the T5 family. Our findings imply that PC4-like factors intervene in multiple unwinding-related processes by acting as versatile modifiers of nucleic acid conformation and raise the possibility that the eukaryotic transcription coactivator derives from ancestral DNA replication, recombination and repair factors. © 2013.

Salidroside Inhibits Myogenesis by Modulating p-Smad3-Induced Myf5 Transcription

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Peng Zhang

2018-03-01

Full Text Available Aim: Salidroside is an active compound extracted from Rhodiola rosea which is used to alleviate fatigue and enhance endurance in high altitude regions. Some studies have demonstrated that salidroside can affect precursor cell differentiation in hematopoietic stem cells, erythrocytes, and osteoblasts. The aim of this study was to investigate the effect of salidroside on myoblast differentiation and to explore the underlying molecular mechanisms of this effect.Methods: C2C12 myoblast cells were treated with different concentrations of salidroside in differentiation media. Real-time PCR, Western blotting, and immunofluorescence assay were employed to evaluate the effects of salidroside on C2C12 differentiation. RNA interference was used to reveal the important role of Myf5 in myogenesis inhibited by salidroside. Chromatin Immunoprecipitation and dual-luciferase reporter assay were utilized to explore the underlying mechanisms of salidroside-induced upregulation of Myf5.Results: We found that salidroside inhibits myogenesis by downregulating MyoD and myogenin, preserves undifferentiated reserve cell pools by upregulating Myf5. Knocking down Myf5 expression significantly rescued the myogenesis inhibited by salidroside. The effect of salidroside on myogenesis was associated with increased phosphorylated Smad3 (p-Smad3. Both SIS3 (Specific inhibitor of p-Smad3 and dominant negative Smad3 plasmid (DN-Smad3 attenuated the inhibitory effect of salidroside on C2C12 differentiation. Moreover, the induction of Myf5 transcription by salidroside was dependent on a Smad-binding site in the promoter region of Myf5 gene.Conclusion and Implications: Our findings identify a novel role and mechanism for salidroside in regulating myogenesis through p-Smad3-induced Myf5 transcription, which may have implications for its further application in combating degenerative muscular diseases caused by depletion of muscle stem cells, such as Duchenne muscular dystrophy or

Production of the 2400 kb Duchenne muscular dystrophy (DMD) gene transcript; transcription time and cotranscriptional splicing

Energy Technology Data Exchange (ETDEWEB)

Tennyson, C.N.; Worton, R.G. [Univ. of Toronto and the Hospital for Sick Children, Ontario (Canada)

1994-09-01

The largest known gene in any organism is the human DMD gene which has 79 exons that span 2400 kb. The extreme nature of the DMD gene raises questions concerning the time required for transcription and whether splicing begins before transcription is complete. DMD gene transcription is induced as cultured human myoblasts differentiate to form multinucleated myotubes, providing a system for studying the kinetics of transcription and splicing. Using quantitative RT-PCR, transcript accumulation was monitored from four different regions within the gene following induction of expression. By comparing the accumulation of transcripts from the 5{prime} and 3{prime} ends of the gene we have shown that approximately 12 hours are required to transcribe 1770 kb of the gene, extrapolating to a time of 16 hours for the transcription unit expressed in muscle. Comparison of accumulation profiles for spliced and total transcript demonstrated that transcripts are spliced at the 5{prime} end before transcription is complete, providing strong evidence for cotranscriptional splicing of DMD gene transcripts. Finally, the rate of transcript accumulation was reduced at the 3{prime} end of the gene relative to the 5{prime} end, perhaps due to premature termination of transcription complexes as they traverse this enormous transcription unit. The lag between transcription initiation and the appearance of complete transcripts could be important in limiting transcript production in dividing cells and to the timing of mRNA appearance in differentiating muscle.

[4-t-butylphenyl]-N-(4-imidazol-1-yl phenyl)sulfonamide (ISCK03) inhibits SCF/c-kit signaling in 501mel human melanoma cells and abolishes melanin production in mice and brownish guinea pigs.

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Na, Yong Joo; Baek, Heung Su; Ahn, Soo Mi; Shin, Hyun Jung; Chang, Ih-Seop; Hwang, Jae Sung

2007-09-01

It is well known that c-kit is related to pigmentation as well as to the oncology target protein. The objective of this study was to discover a skin-whitening agent that regulates c-kit activity. We have developed a high-throughput screening system using recombinant human c-kit protein. Approximately 10,000 synthetic compounds were screened for their effect on c-kit activity. Phenyl-imidazole sulfonamide derivatives showed inhibitory activity on c-kit phosphorylation in vitro. The effects of one derivative, [4-t-butylphenyl]-N-(4-imidazol-1-yl phenyl)sulfonamide (ISCK03), on stem-cell factor (SCF)/c-kit cellular signaling in 501mel human melanoma cells were examined further. Pretreatment of 501mel cells with ISCK03 inhibited SCF-induced c-kit phosphorylation dose dependently. ISCK03 also inhibited p44/42 ERK mitogen-activated protein kinase (MAPK) phosphorylation, which is known to be involved in SCF/c-kit downstream signaling. However ISCK03 did not inhibit hepatocyte growth factor (HGF)-induced phosphorylation of p44/42 ERK proteins. To determine the in vivo potency of ISCK03, it was orally administered to depilated C57BL/6 mice. Interestingly, oral administration of ISCK03 induced the dose-dependent depigmentation of newly regrown hair, and this was reversed with cessation of ISCK03 treatment. Finally, to investigate whether the inhibitory effect of ISCK03 on SCF/c-kit signaling abolished UV-induced pigmentation, ISCK03 was applied to UV-induced pigmented spots on brownish guinea pig skin. The topical application of ISCK03 promoted the depigmentation of UV-induced hyperpigmented spots. Fontana-Masson staining analysis showed epidermal melanin was diminished in spots treated with ISCK03. These results indicate that phenyl-imidazole sulfonamide derivatives are potent c-kit inhibitors and might be used as skin-whitening agents.

Uridine 5'-Monophosphate Synthase Is Transcriptionally Regulated by Pyrimidine Levels in Nicotiana plumbaginifolia

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Santoso; Thornburg

1998-02-01

To understand the regulation and expression of pyrimidine biosynthesis in plants, we have examined the effect of the metabolic inhibitor 5-fluoroorotic acid (FOA) on uridine-5'-monophosphate synthase (UMPSase) expression in cell cultures of Nicotiana plumbaginifolia. UMPSase is the rate-limiting step of pyrimidine biosynthesis in plants. Addition of FOA causes an up-regulation of UMPSase enzyme activity in cell cultures after a lag phase of several days. Western-blot analysis demonstrated that the up-regulation in enzyme activity was caused by increased expression of the UMPSase protein. Northern-blot analysis demonstrated a higher level of UMPSase mRNA in the FOA-induced tissues than in control tissues. Run-on transcriptional assays showed that the UMPSase gene was transcriptionally activated after FOA treatment. The mechanism of toxicity of FOA is through thymine starvation. We found that addition of thymine abrogated the FOA-mediated up-regulation of UMPSase. In addition, methotrexate and aminopterin, which affect thymine levels by inhibiting dihydrofolate reductase, also up-regulate UMPSase in N. plumbaginifolia cells.

Luminescence properties of Y3Al5O12:Ce nanoceramics

International Nuclear Information System (INIS)

Zorenko, Yu.; Voznyak, T.; Gorbenko, V.; Zych, E.; Nizankovski, S.; Dan'ko, A.; Puzikov, V.

2011-01-01

Comparative analysis of the luminescent properties of Y 3 Al 5 O 12 :Ce (YAG:Ce) transparent optical ceramics (OS) with those of single crystal (SC) and single crystalline film (SCF) analogues has been performed under excitation by pulsed synchrotron radiation in the fundamental absorption range of YAG host. It has been shown that the properties of YAG:Ce OC are closer to the properties of the SCF counterpart, where Y Al antisite defects are completely absent, rather than to the properties of SC of this garnet with large concentration of Y Al antisite defects. At the same time, the luminescence spectra of YAG:Ce OC show weak emission bands in the 200-470 nm range related to Y Al antisite defects and charged oxygen vacancies (F + and F centers). YAG:Ce OS also possesses significantly larger contribution of slow components in the Ce 3+ luminescence decay under high-energy excitation in comparison with SC and SCF of this garnet due to the involvement of antisite defects, charged oxygen vacancies as well as boundaries of grains in the energy transfer processes from the host to the Ce 3+ ions.

CDK-mediated activation of the SCFFBXO28 ubiquitin ligase promotes MYC-driven transcription and tumourigenesis and predicts poor survival in breast cancer

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Cepeda, Diana; Ng, Hwee-Fang; Sharifi, Hamid Reza; Mahmoudi, Salah; Cerrato, Vanessa Soto; Fredlund, Erik; Magnusson, Kristina; Nilsson, Helén; Malyukova, Alena; Rantala, Juha; Klevebring, Daniel; Viñals, Francesc; Bhaskaran, Nimesh; Zakaria, Siti Mariam; Rahmanto, Aldwin Suryo; Grotegut, Stefan; Nielsen, Michael Lund; Szigyarto, Cristina Al-Khalili; Sun, Dahui; Lerner, Mikael; Navani, Sanjay; Widschwendter, Martin; Uhlén, Mathias; Jirström, Karin; Pontén, Fredrik; Wohlschlegel, James; Grandér, Dan; Spruck, Charles; Larsson, Lars-Gunnar; Sangfelt, Olle

2013-01-01

SCF (Skp1/Cul1/F-box) ubiquitin ligases act as master regulators of cellular homeostasis by targeting key proteins for ubiquitylation. Here, we identified a hitherto uncharacterized F-box protein, FBXO28 that controls MYC-dependent transcription by non-proteolytic ubiquitylation. SCFFBXO28 activity and stability are regulated during the cell cycle by CDK1/2-mediated phosphorylation of FBXO28, which is required for its efficient ubiquitylation of MYC and downsteam enhancement of the MYC pathway. Depletion of FBXO28 or overexpression of an F-box mutant unable to support MYC ubiquitylation results in an impairment of MYC-driven transcription, transformation and tumourigenesis. Finally, in human breast cancer, high FBXO28 expression and phosphorylation are strong and independent predictors of poor outcome. In conclusion, our data suggest that SCFFBXO28 plays an important role in transmitting CDK activity to MYC function during the cell cycle, emphasizing the CDK-FBXO28-MYC axis as a potential molecular drug target in MYC-driven cancers, including breast cancer. PMID:23776131

Luminescent and scintillation properties of the Ce3+ doped Y3−xLuxAl5O12:Ce single crystalline films

International Nuclear Information System (INIS)

Zorenko, Yu.; Gorbenko, V.; Zorenko, T.; Popielarski, P.; Mosińska, L.; Fedorov, A.

2016-01-01

The work is related to the investigation of scintillation and luminescent properties of single crystalline films (SCF) of solid solutions of Ce 3+ doped Y 3−x Lu x Al 5 O 12 :Ce garnets with x value in the 0–3 range. We have shown a possibility of realization of high-energy shift of the Ce 3+ ion emission spectrum in these garnets up to 22 nm. We have also found that the light yield of the radioluminescence under α-particle excitation of LuAG:Ce SCF can exceed by 1.3 times the corresponding values for the YAG:Ce SCF counterpart. For investigation of the luminescent properties of Y 3−x Lu x Al 5 O 12 :Ce SCF at different x values the luminescent spectroscopy of these SCFs under excitation by synchrotron radiation in the VUV range was performed. - Highlights: • Single crystalline films of Y 3−x Lu x Al 5 O 12 garnets at x=0–3.0 were grown by LPE method onto YAG substrates. • Lattice constant of Y 3−x Lu x Al 5 O 12 :Ce film and the film/substrate misfit changed linearly with increasing of Lu content in the x=0–3.0 range. • High-energy shift of the Ce 3+ emission up to 22 nm in Y 3−x Lu x Al 5 O 12 film with increasing of Lu content in the x=0–3.0 range. • Light yield of Y 3−x Lu x Al 5 O 12 :Ce film decreases in the x=0–1.8 range and increases in the x=1.8–3.0 range. • Scintillation LY of Lu 3 Al 5 O 12 :Ce film can exceed by 1.3 times the LY for YAG:Ce film counterpart.

bZIP transcription factor CgAP1 is essential for oxidative stress tolerance and full virulence of the poplar anthracnose fungus Colletotrichum gloeosporioides.

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Sun, Yingjiao; Wang, Yonglin; Tian, Chengming

2016-10-01

Yeast AP1 transcription factor is a regulator of oxidative stress response. Here, we report the identification and characterization of CgAP1, an ortholog of YAP1 in poplar anthracnose fungus Colletotrichum gloeosporioides. The expression of CgAP1 was highly induced by reactive oxygen species. CgAP1 deletion mutants displayed enhanced sensitivity to oxidative stress compared with the wild-type strain, and their poplar leaf virulence was obviously reduced. However, the mutants exhibited no obvious defects in aerial hyphal growth, conidia production, and appressoria formation. CgAP1::eGFP fusion protein localized to the nucleus after TBH (tert-Butyl hydroperoxide) treatment, suggesting that CgAP1 functions as a redox sensor in C. gloeosporioides. In addition, CgAP1 prevented the accumulation of ROS during early stages of biotrophic growth. CgAP1 also acted as a positive regulator of several ROS-related genes (i.e., Glr1, Hyr1, and Cyt1) involved in the antioxidative response. These results highlight the key regulatory role of CgAP1 transcription factor in oxidative stress response and provide insights into the function of ROS detoxification in virulence of C. gloeosporioides. Copyright © 2016 Elsevier Inc. All rights reserved.

Multiple 5' ends of human cytomegalovirus UL57 transcripts identify a complex, cycloheximide-resistant promoter region that activates oriLyt

International Nuclear Information System (INIS)

Kiehl, Anita; Huang, Lili; Franchi, David; Anders, David G.

2003-01-01

The human cytomegalovirus (HCMV) UL57 gene lies adjacent to HCMV oriLyt, from which it is separated by an organizationally conserved, mostly noncoding region that is thought to both regulate UL57 expression and activate oriLyt function. However, the UL57 promoter has not been studied. We determined the 5' ends of UL57 transcripts toward an understanding of the potential relationship between UL57 expression and oriLyt activation. The results presented here identified three distinct 5' ends spread over 800 bp, at nt 90302, 90530, and 91138; use of these sites exhibited differential sensitivity to phosphonoformic acid treatment. Interestingly, a 10-kb UL57 transcript accumulated in cycloheximide-treated infected cells, even though other early transcripts were not detectable. However, the 10-kb transcript did not accumulate in cells treated with the more stringent translation inhibitor anisomycin. Consistent with the notion that the identified 5' ends arise from distinct transcription start sites, the sequences upstream of sites I and II functioned as promoters responsive to HCMV infection in transient assays. However, the origin-proximal promoter region III required downstream sequences for transcriptional activity. Mutation of candidate core promoter elements suggested that promoter III is regulated by an initiator region (Inr) and a downstream promoter element. Finally, a 42-bp sequence containing the candidate Inr activated a minimal oriLyt core construct in transient replication assays. Thus, these studies showed that a large, complex promoter region with novel features controls UL57 expression, and identified a sequence that regulates both UL57 transcription and oriLyt activation

SCF(JFK) is a bona fide E3 ligase for ING4 and a potent promoter of the angiogenesis and metastasis of breast cancer.

Science.gov (United States)

Yan, Ruorong; He, Lin; Li, Zhongwu; Han, Xiao; Liang, Jing; Si, Wenzhe; Chen, Zhe; Li, Lei; Xie, Guojia; Li, Wanjin; Wang, Peiyan; Lei, Liandi; Zhang, Hongquan; Pei, Fei; Cao, Dengfeng; Sun, Luyang; Shang, Yongfeng

2015-03-15

Loss of function/dysregulation of inhibitor of growth 4 (ING4) and hyperactivation of NF-κB are frequent events in many types of human malignancies. However, the molecular mechanisms underlying these remarkable aberrations are not understood. Here, we report that ING4 is physically associated with JFK. We demonstrated that JFK targets ING4 for ubiquitination and degradation through assembly of an Skp1-Cul1-F-box (SCF) complex. We showed that JFK-mediated ING4 destabilization leads to the hyperactivation of the canonical NF-κB pathway and promotes angiogenesis and metastasis of breast cancer. Significantly, the expression of JFK is markedly up-regulated in breast cancer, and the level of JFK is negatively correlated with that of ING4 and positively correlated with an aggressive clinical behavior of breast carcinomas. Our study identified SCF(JFK) as a bona fide E3 ligase for ING4 and unraveled the JFK-ING4-NF-κB axis as an important player in the development and progression of breast cancer, supporting the pursuit of JFK as a potential target for breast cancer intervention. © 2015 Yan et al.; Published by Cold Spring Harbor Laboratory Press.

Investigation of the Semicoa SCF9550 and the International Rectifier IRHM57260SE for Single-Event Gate Rapture and Single-Event Burnout : NASA Electronic Parts and Packaging (NEPP) Program Office of Safety and Mission Assurance

Science.gov (United States)

Scheick, Leif

2011-01-01

Single-event-effect test results for hi-rel total-dose-hardened power MOSFETs are presented in this report. TheSCF9550 from Semicoa and the IRHM57260SE from International Rectifier were tested to NASA test condition/standards and requirements.The IRHM57260SE performed much better when compared to previous testing. These initial results confirm that parts from the Temecula line are marginally comparable to the El Segundo line. The SCF9550 from Semicoa was also tested and represents the initial parts offering from this vendor. Both parts experienced single-event gate rupture (SEGR) and single-event burnout (SEB). All of the SEGR was from gate to drain.

Identification of drought, cadmium and root-lesion nematode infection stress-responsive transcription factors in ramie

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Zheng Xia

2016-01-01

Full Text Available Drought, cadmium (Cd stress, and root lesion nematode (RLN infection are three of the most important stresses affecting ramie growth and development; therefore, ramie breeding programs focus on their management more than on any other abiotic or biotic stresses. The fact that only a small number of stress-responsive transcription factors (TFs have been identified so far is a major obstacle in the elucidation of mechanisms regulating the response to these three stresses in ramie. In this study, in order to uncover more stress-responsive TFs, a total of 179 nonredundant genes with full-length open reading frames from the MYB, AP2/ERF, bZIP, HD-ZIP, and COL families were obtained by searching for against the ramie transcriptome. Expression pattern analysis demonstrated that most of these genes showed relatively higher expression in the stem xylem and bast than in other tissues. Among these genes, 96 genes were found to be involved in responses to drought, Cd exposure, or RLN-infection. The expression of 54 of these genes was regulated by at least two stresses. These stress-responsive TFs probably have roles in the regulation of stress tolerance. The discovery of these stress-responsive TFs will be helpful for furthering our understanding of the mechanisms that regulate stress responses in ramie.

MiT/TFE transcription factors are activated during mitophagy downstream of Parkin and Atg5.

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Nezich, Catherine L; Wang, Chunxin; Fogel, Adam I; Youle, Richard J

2015-08-03

The kinase PINK1 and ubiquitin ligase Parkin can regulate the selective elimination of damaged mitochondria through autophagy (mitophagy). Because of the demand on lysosomal function by mitophagy, we investigated a role for the transcription factor EB (TFEB), a master regulator of lysosomal biogenesis, in this process. We show that during mitophagy TFEB translocates to the nucleus and displays transcriptional activity in a PINK1- and Parkin-dependent manner. MITF and TFE3, homologues of TFEB belonging to the same microphthalmia/transcription factor E (MiT/TFE) family, are similarly regulated during mitophagy. Unlike TFEB translocation after starvation-induced mammalian target of rapamycin complex 1 inhibition, Parkin-mediated TFEB relocalization required Atg9A and Atg5 activity. However, constitutively active Rag guanosine triphosphatases prevented TFEB translocation during mitophagy, suggesting cross talk between these two MiT/TFE activation pathways. Analysis of clustered regularly interspaced short palindromic repeats-generated TFEB/MITF/TFE3/TFEC single, double, and triple knockout cell lines revealed that these proteins partly facilitate Parkin-mediated mitochondrial clearance. These results illuminate a pathway leading to MiT/TFE transcription factor activation, distinct from starvation-induced autophagy, which occurs during mitophagy.

Transcription factor Nrf1 is topologically repartitioned across membranes to enable target gene transactivation through its acidic glucose-responsive domains.

Science.gov (United States)

Zhang, Yiguo; Ren, Yonggang; Li, Shaojun; Hayes, John D

2014-01-01

The membrane-bound Nrf1 transcription factor regulates critical homeostatic and developmental genes. The conserved N-terminal homology box 1 (NHB1) sequence in Nrf1 targets the cap'n'collar (CNC) basic basic-region leucine zipper (bZIP) factor to the endoplasmic reticulum (ER), but it is unknown how its activity is controlled topologically within membranes. Herein, we report a hitherto unknown mechanism by which the transactivation activity of Nrf1 is controlled through its membrane-topology. Thus after Nrf1 is anchored within ER membranes, its acidic transactivation domains (TADs), including the Asn/Ser/Thr-rich (NST) glycodomain situated between acidic domain 1 (AD1) and AD2, are transiently translocated into the lumen of the ER, where NST is glycosylated in the presence of glucose to yield an inactive 120-kDa Nrf1 glycoprotein. Subsequently, portions of the TADs partially repartition across membranes into the cyto/nucleoplasmic compartments, whereupon an active 95-kDa form of Nrf1 accumulates, a process that is more obvious in glucose-deprived cells and may involve deglycosylation. The repartitioning of Nrf1 out of membranes is monitored within this protein by its acidic-hydrophobic amphipathic glucose-responsive domains, particularly the Neh5L subdomain within AD1. Therefore, the membrane-topological organization of Nrf1 dictates its post-translational modifications (i.e. glycosylation, the putative deglycosylation and selective proteolysis), which together control its ability to transactivate target genes.

Histone H1 and Chromosomal Protein HMGN2 Regulate Prolactin-induced STAT5 Transcription Factor Recruitment and Function in Breast Cancer Cells.

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Schauwecker, Suzanne M; Kim, J Julie; Licht, Jonathan D; Clevenger, Charles V

2017-02-10

The hormone prolactin (PRL) contributes to breast cancer pathogenesis through various signaling pathways, one of the most notable being the JAK2/signal transducer and activator of transcription 5 (STAT5) pathway. PRL-induced activation of the transcription factor STAT5 results in the up-regulation of numerous genes implicated in breast cancer pathogenesis. However, the molecular mechanisms that enable STAT5 to access the promoters of these genes are not well understood. Here, we show that PRL signaling induces chromatin decompaction at promoter DNA, corresponding with STAT5 binding. The chromatin-modifying protein high mobility group nucleosomal binding domain 2 (HMGN2) specifically promotes STAT5 accessibility at promoter DNA by facilitating the dissociation of the linker histone H1 in response to PRL. Knockdown of H1 rescues the decrease in PRL-induced transcription following HMGN2 knockdown, and it does so by allowing increased STAT5 recruitment. Moreover, H1 and STAT5 are shown to function antagonistically in regulating PRL-induced transcription as well as breast cancer cell biology. While reduced STAT5 activation results in decreased PRL-induced transcription and cell proliferation, knockdown of H1 rescues both of these effects. Taken together, we elucidate a novel mechanism whereby the linker histone H1 prevents STAT5 binding at promoter DNA, and the PRL-induced dissociation of H1 mediated by HMGN2 is necessary to allow full STAT5 recruitment and promote the biological effects of PRL signaling. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Kctd10 regulates heart morphogenesis by repressing the transcriptional activity of Tbx5a in zebrafish

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Tong, Xiangjun; Zu, Yao; Li, Zengpeng; Li, Wenyuan; Ying, Lingxiao; Yang, Jing; Wang, Xin; He, Shuonan; Liu, Da; Zhu, Zuoyan; Chen, Jianming; Lin, Shuo; Zhang, Bo

2014-01-01

The T-box transcription factor Tbx5 (Tbx5a in zebrafish) plays a crucial role in the formation of cardiac chambers in a dose-dependent manner. Its deregulation leads to congenital heart disease. However, little is known regarding its regulation. Here we isolate a zebrafish mutant with heart malformations, called 34c. The affected gene is identified as kctd10, a member of the potassium channel tetramerization domain (KCTD)-containing family. In the mutant, the expressions of the atrioventricular canal marker genes, such as tbx2b, hyaluronan synthase 2 (has2), notch1b and bmp4, are changed. The knockdown of tbx5 rescues the ectopic expression of has2, and knockdown of either tbx5a or has2 alleviates the heart defects. We show that Kctd10 directly binds to Tbx5 to repress its transcriptional activity. Our results reveal a new essential factor for cardiac development and suggest that KCTD10 could be considered as a new causative gene of congenital heart disease.

The chimeric transcript RUNX1-GLRX5: a biomarker for good postoperative prognosis in Stage IA non-small-cell lung cancer.

Science.gov (United States)

Ishikawa, Rie; Amano, Yosuke; Kawakami, Masanori; Sunohara, Mitsuhiro; Watanabe, Kousuke; Kage, Hidenori; Ohishi, Nobuya; Yatomi, Yutaka; Nakajima, Jun; Fukayama, Masashi; Nagase, Takahide; Takai, Daiya

2016-02-01

Stage IA non-small-cell lung cancer cases have been recognized as having a low risk of relapse; however, occasionally, relapse may occur. To predict clinical outcome in Stage IA non-small-cell lung cancer patients, we searched for chimeric transcripts that can be used as biomarkers and identified a novel chimeric transcript, RUNX1-GLRX5, comprising RUNX1, a transcription factor, and GLRX5. This chimera was detected in approximately half of the investigated Stage IA non-small-cell lung cancer patients (44/104 cases, 42.3%). Although there was no significant difference in the overall survival rate between RUNX1-GLRX5-positive and -negative cases (P = 0.088), a significantly lower relapse rate was observed in the RUNX1-GLRX5-positive cases (P = 0.039), indicating that this chimera can be used as a biomarker for good prognosis in Stage IA patients. Detection of the RUNX1-GLRX5 chimeric transcript may therefore be useful for the determination of a postoperative treatment plan for Stage IA non-small-cell lung cancer patients. © The Author 2015. Published by Oxford University Press.

PRMT5 restricts hepatitis B virus replication through epigenetic repression of covalently closed circular DNA transcription and interference with pregenomic RNA encapsidation.

Science.gov (United States)

Zhang, Wen; Chen, Jieliang; Wu, Min; Zhang, Xiaonan; Zhang, Min; Yue, Lei; Li, Yaming; Liu, Jiangxia; Li, Baocun; Shen, Fang; Wang, Yang; Bai, Lu; Protzer, Ulrike; Levrero, Massimo; Yuan, Zhenghong

2017-08-01

Chronic hepatitis B virus (HBV) infection remains a major health problem worldwide. The covalently closed circular DNA (cccDNA) minichromosome, which serves as the template for the transcription of viral RNAs, plays a key role in viral persistence. While accumulating evidence suggests that cccDNA transcription is regulated by epigenetic machinery, particularly the acetylation of cccDNA-bound histone 3 (H3) and H4, the potential contributions of histone methylation and related host factors remain obscure. Here, by screening a series of methyltransferases and demethylases, we identified protein arginine methyltransferase 5 (PRMT5) as an effective restrictor of HBV transcription and replication. In cell culture-based models for HBV infection and in liver tissues of patients with chronic HBV infection, we found that symmetric dimethylation of arginine 3 on H4 on cccDNA was a repressive marker of cccDNA transcription and was regulated by PRMT5 depending on its methyltransferase domain. Moreover, PRMT5-triggered symmetric dimethylation of arginine 3 on H4 on the cccDNA minichromosome involved an interaction with the HBV core protein and the Brg1-based human SWI/SNF chromatin remodeler, which resulted in down-regulation of the binding of RNA polymerase II to cccDNA. In addition to the inhibitory effect on cccDNA transcription, PRMT5 inhibited HBV core particle DNA production independently of its methyltransferase activity. Further study revealed that PRMT5 interfered with pregenomic RNA encapsidation by preventing its interaction with viral polymerase protein through binding to the reverse transcriptase-ribonuclease H region of polymerase, which is crucial for the polymerase-pregenomic RNA interaction. PRMT5 restricts HBV replication through a two-part mechanism including epigenetic suppression of cccDNA transcription and interference with pregenomic RNA encapsidation; these findings improve the understanding of epigenetic regulation of HBV transcription and host

Influence of thermal treatment and γ-radiation on absorption, luminescence and scintillation properties of Lu3Al5O12:Ce single crystalline films

International Nuclear Information System (INIS)

Zorenko, Yu.; Pavlyk, B.; Turchak, R.; Zorenko, T.; Gorbenko, V.; Konstankevych, I.; Savchyn, V.; Voznyak, T.

2007-01-01

The influence of thermal treatment (TT) and irradiation by γ-quanta ( 60 Co, 1.25 MeV) in the range of absorbed doses 10 6 -10 8 R on the absorption, luminescence and light yield (LY) of scintillators based on Lu 3 Al 5 O 12 :Ce (LuAG:Ce) single crystalline films (SCF) grown by LPE from the Pb-based flux is studied. It has been shown that transformation of the Fe 3+ ↔Fe 2+ and Pb 2+ ↔Pb 4+ valent states of iron and lead trace impurities cause the change in the optical absorption of SCF induced by the TT in air and vacuum in the range of annealing temperatures of 800-1200 0 C. Possible types of radiation defects in LuAG:Ce SCF induced by γ-radiation are analyzed

The regulatory effect of SC-236 (4-[5-(4-chlorophenyl)-3-(trifluoromethyl)-1-pyrazol-1-l] benzenesulfonamide) on stem cell factor induced migration of mast cells

International Nuclear Information System (INIS)

Kim, Su-Jin; Jeong, Hyun-Ja; Park, Rae-Kil; Lee, Kang-Min; Kim, Hyung-Min; Um, Jae-Young; Hong, Seung-Heon

2007-01-01

SC-236 (4-[5-(4-chlorophenyl)-3-(trifluoromethyl)-1-pyrazol-1-]benzenesulfonamide; C 16 H 11 ClF 3 N 3 O 2 S), is a highly selective cyclooxygenase (COX)-2 inhibitor. Recently, there have been reports that SC-236 protects against cartilage damage in addition to reducing inflammation and pain in osteoarthritis. However, the mechanism involved in the inflammatory allergic reaction has not been examined. Mast cells accumulation can be related to inflammatory conditions, including allergic rhinitis, asthma, and rheumatoid arthritis. The aim of the present study is to investigate the effects of SC-236 on stem cell factor (SCF)-induced migration, morphological alteration, and cytokine production of rat peritoneal mast cells (RPMCs). We observed that SCF significantly induced the migration and morphological alteration. The ability of SCF to enhance migration and morphological alteration was abolished by treatment with SC-236. In addition, production of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and vascular endothelial growth factor (VEGF) production induced by SCF was significantly inhibited by treatment with SC-236. Previous work has demonstrated that SCF-induced migration and cytokine production of mast cells require p38 MAPK activation. We also showed that SC-236 suppresses the SCF-induced p38 MAPK activation in RPMCs. These data suggest that SC-236 inhibits migration and cytokine production through suppression of p38 MAPK activation. These results provided new insight into the pharmacological actions of SC-236 and its potential therapeutic role in the treatment of inflammatory allergic diseases

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.

Elevated expression and potential roles of human Sp5, a member of Sp transcription factor family, in human cancers

International Nuclear Information System (INIS)

Chen Yongxin; Guo Yingqiu; Ge Xijin; Itoh, Hirotaka; Watanabe, Akira; Fujiwara, Takeshi; Kodama, Tatsuhiko; Aburatani, Hiroyuki

2006-01-01

In this report, we describe the expression and function of human Sp5, a member of the Sp family of zinc finger transcription factors. Like other family members, the Sp5 protein contains a Cys2His2 zinc finger DNA binding domain at the C-terminus. Our experiments employing Gal4-Sp5 fusion proteins reveal multiple transcriptional domains, including a N-terminal activity domain, an intrinsic repressive element, and a C-terminal synergistic domain. Elevated expression of Sp5 was noted in several human tumors including hepatocellular carcinoma, gastric cancer, and colon cancer. To study the effects of the Sp5 protein on growth properties of human cancer cells and facilitate the identification of its downstream genes, we combined an inducible gene expression system with microarray analysis to screen for its transcriptional targets. Transfer of Sp5 into MCF-7 cells that expressed no detectable endogenous Sp5 protein elicited significant growth promotion activity. Several of the constitutively deregulated genes have been associated with tumorigenesis (CDC25C, CEACAM6, TMPRSS2, XBP1, MYBL1, ABHD2, and CXCL12) and Wnt/β-Catenin signaling pathways (BAMBI, SIX1, IGFBP5, AES, and p21 WAF1 ). This information could be utilized for further mechanistic research and for devising optimized therapeutic strategies against human cancers

miR-17-5p targets the p300/CBP-associated factor and modulates androgen receptor transcriptional activity in cultured prostate cancer cells

International Nuclear Information System (INIS)

Gong, Ai-Yu; Eischeid, Alex N; Xiao, Jing; Zhao, Jian; Chen, Dongqing; Wang, Zhao-Yi; Young, Charles YF; Chen, Xian-Ming

2012-01-01

Androgen receptor (AR) signalling is critical to the initiation and progression of prostate cancer (PCa). Transcriptional activity of AR involves chromatin recruitment of co-activators, including the p300/CBP-associated factor (PCAF). Distinct miRNA expression profiles have been identified in PCa cells during the development and progression of the disease. Whether miRNAs regulate PCAF expression in PCa cells to regulate AR transcriptional activity is still unclear. Expression of PCAF was investigated in several PCa cell lines by qRT-PCR, Western blot, and immunocytochemistry. The effects of PCAF expression on AR-regulated transcriptional activity and cell growth in PCa cells were determined by chromatin immunoprecipitation, reporter gene construct analysis, and MTS assay. Targeting of PCAF by miR-17-5p was evaluated using the luciferase reporter assay. PCAF was upregulated in several PCa cell lines. Upregulation of PCAF promoted AR transcriptional activation and cell growth in cultured PCa cells. Expression of PCAF in PCa cells was associated with the downregulation of miR-17-5p. Targeting of the 3’-untranslated region of PCAF mRNA by miR-17-5p caused translational suppression and RNA degradation, and, consequently, modulation of AR transcriptional activity in PCa cells. PCAF is upregulated in cultured PCa cells, and upregulation of PCAF is associated with the downregulation of miR-17-5p. Targeting of PCAF by miR-17-5p modulates AR transcriptional activity and cell growth in cultured PCa cells

Interaction of higher plant ribosomal 5S RNAs with ''Xenopus laevis'' transcriptional factor IIIA

International Nuclear Information System (INIS)

Barciszewska, M.Z.

1994-01-01

In this paper transcriptional factor IIIA (TFIIIA) has been used as a probe for identity of three-dimensional-structure of eukaryotic 5S rRNAs. I was interested in finding a common motif in plant and ''Xenopus'' 5S rRNAs for TFIIIA recognition. I found that the two eukaryotic 5S rRNAs (from wheat germ and lupin seeds) are recognized by ''X. laevis'' TFIIIA and the data clearly suggest that these 5S rRNAs have very similar if not identical three-dimensional structures. Also effects of various conditions on stability of these complexes have been studied. (author). 30 refs, 6 figs, 1 tab

The Arabidopsis Phytocystatin AtCYS5 Enhances Seed Germination and Seedling Growth under Heat Stress Conditions.

Science.gov (United States)

Song, Chieun; Kim, Taeyoon; Chung, Woo Sik; Lim, Chae Oh

2017-08-01

Phytocystatins (PhyCYSs) are plant-specific proteinaceous inhibitors that are implicated in protein turnover and stress responses. Here, we characterized a PhyCYS from Arabidopsis thaliana , which was designated AtCYS5. RT-qPCR analysis showed that the expression of AtCYS5 in germinating seeds was induced by heat stress (HS) and exogenous abscisic acid (ABA) treatment. Analysis of the expression of the β -glucuronidase reporter gene under the control of the AtCYS5 promoter showed that AtCYS5 expression during seed germination was induced by HS and ABA. Constitutive overexpression of AtCYS5 driven by the cauliflower mosaic virus 35S promoter led to enhanced HS tolerance in transgenic Arabidopsis , which was characterized by higher fresh weight and root length compared to wild-type (WT) and knockout ( cys5 ) plants grown under HS conditions. The HS tolerance of At-CYS5 -overexpressing transgenic plants was associated with increased insensitivity to exogenous ABA during both seed germination and post-germination compared to WT and cys5 . Although no HS elements were identified in the 5'-flanking region of AtCYS5 , canonical ABA-responsive elements (ABREs) were detected. AtCYS5 was upregulated in ABA-treated protoplasts transiently co-expressing this gene and genes encoding bZIP ABRE-binding factors (ABFs and AREB3). In the absence of ABA, ABF1 and ABF3 directly bound to the ABREs in the AtCYS5 promoter, which activated the transcription of this gene in the presence of ABA. These results suggest that an ABA-dependent pathway plays a positive role in the HS-responsive expression of AtCYS5 during seed germination and post-germination growth.

Stimulation of Pol III-dependent 5S rRNA and U6 snRNA gene expression by AP-1 transcription factors.

Science.gov (United States)

Ahuja, Richa; Kumar, Vijay

2017-07-01

RNA polymerase III transcribes structurally diverse group of essential noncoding RNAs including 5S ribosomal RNA (5SrRNA) and U6 snRNA. These noncoding RNAs are involved in RNA processing and ribosome biogenesis, thus, coupling Pol III activity to the rate of protein synthesis, cell growth, and proliferation. Even though a few Pol II-associated transcription factors have been reported to participate in Pol III-dependent transcription, its activation by activator protein 1 (AP-1) factors, c-Fos and c-Jun, has remained unexplored. Here, we show that c-Fos and c-Jun bind to specific sites in the regulatory regions of 5S rRNA (type I) and U6 snRNA (type III) gene promoters and stimulate their transcription. Our chromatin immunoprecipitation studies suggested that endogenous AP-1 factors bind to their cognate promoter elements during the G1/S transition of cell cycle apparently synchronous with Pol III transcriptional activity. Furthermore, the interaction of c-Jun with histone acetyltransferase p300 promoted the recruitment of p300/CBP complex on the promoters and facilitated the occupancy of Pol III transcriptional machinery via histone acetylation and chromatin remodeling. The findings of our study, together, suggest that AP-1 factors are novel regulators of Pol III-driven 5S rRNA and U6 snRNA expression with a potential role in cell proliferation. © 2017 Federation of European Biochemical Societies.

Citrus PH5-like H+-ATPase genes: identification and transcript analysis to investigate their possible relationship with citrate accumulation in fruits

Directory of Open Access Journals (Sweden)

Cai-Yun eShi

2015-03-01

Full Text Available PH5 is a petunia gene that encodes a plasma membrane H+-ATPase and determines the vacuolar pH. The citrate content of fruit cell vacuoles influences citrus organoleptic qualities. Although citrus could have PH5-like homologs that are involved in citrate accumulation, the details are still unknown. In this study, extensive data-mining with the PH5 sequence and PCR amplification confirmed that there are at least eight PH5-like genes (CsPH1-8 in the citrus genome. CsPHs have a molecular mass of approximately 100 kDa, and they have high similarity to PhPH5, AtAHA10 or AtAHA2 (from 64.6% to 80.9%. They contain 13-21 exons and 12-20 introns and were evenly distributed into four subgroups of the P3A-subfamily (CsPH1, CsPH2, and CsPH3 in Group I, CsPH4 and CsPH5 in Group II, CsPH6 in Group IV, and CsPH7 and CsPH8 in Group III together with PhPH5. A transcript analysis showed that CsPH1, 3, and 4 were predominantly expressed in mature leaves, whereas CsPH2 and 7 were predominantly expressed in roots, CsPH5 and 6 were predominantly expressed in flowers, and CsPH8 was predominantly expressed in fruit juice sacs. Moreover, the CsPH transcript profiles differed between orange and pummelo, as well as between high-acid and low-acid cultivars. The low-acid orange ‘Honganliu’ exhibits low transcript levels of CsPH3, CsPH4, CsPH5, and CsPH8, whereas the acid-free pummelo has only a low transcript level of CsPH8. In addition, ABA injection increased the citrate content significantly, which was accompanied by the obvious induction of CsPH2, 6, 7, and 8 transcript levels. Taken together, we suggest that CsPH8 seems likely to regulate citrate accumulation in the citrus fruit vacuole.

ADA5/SPT20 links the ADA and SPT genes, which are involved in yeast transcription.

OpenAIRE

Marcus, G A; Horiuchi, J; Silverman, N; Guarente, L

1996-01-01

In this report we described the cloning and characterization of ADA5, a gene identified by resistance to GAL4-VP16-mediated toxicity. ADA5 binds directly to the VP16 activation domain but not to a transcriptionally defective VP16 double point mutant. Double mutants with mutations in ada5 and other genes (ada2 or ada3) isolated by resistance to GAL4-VP16 grow like ada5 single mutants, suggesting that ADA5 is in the same pathway as the other ADA genes. Further, ADA5 cofractionates and coprecipi...

Luminescence of Bi3+ ions in Y3Al5O12:Bi single crystalline films

International Nuclear Information System (INIS)

Zorenko, Yu.; Gorbenko, V.; Voznyak, T.; Vistovsky, V.; Nedilko, S.; Nikl, M.

2007-01-01

The absorption and cathodoluminescence spectra of single crystalline films (SCF) of Y 3 Al 5 O 12 :Bi garnet depending on Bi concentration were analyzed. For consideration of the nature of the UV and visible Bi-related emission bands the time-resolved luminescence of Bi 3+ (ns 2 ) ions in YAG:Bi SCF was studied at 10 K under excitation by synchrotron radiation. The difference in the excitation spectra and emission decay of the UV and visible bands has been explained via radiative relaxation from the 3 P 1,0 excited states to the 1 S 0 ground state of the isolated and pair/clustered Bi 3+ emission centers in the garnet lattice, respectively

Genome-Wide Identification and Analysis of Drought-Responsive Genes and MicroRNAs in Tobacco

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Fuqiang Yin

2015-03-01

Full Text Available Drought stress response is a complex trait regulated at transcriptional and post-transcriptional levels in tobacco. Since the 1990s, many studies have shown that miRNAs act in many ways to regulate target expression in plant growth, development and stress response. The recent draft genome sequence of Nicotiana benthamiana has provided a framework for Digital Gene Expression (DGE and small RNA sequencing to understand patterns of transcription in the context of plant response to environmental stress. We sequenced and analyzed three Digital Gene Expression (DGE libraries from roots of normal and drought-stressed tobacco plants, and four small RNA populations from roots, stems and leaves of control or drought-treated tobacco plants, respectively. We identified 276 candidate drought responsive genes (DRGs with sequence similarities to 64 known DRGs from other model plant crops, 82 were transcription factors (TFs including WRKY, NAC, ERF and bZIP families. Of these tobacco DRGs, 54 differentially expressed DRGs included 21 TFs, which belonged to 4 TF families such as NAC (6, MYB (4, ERF (10, and bZIP (1. Additionally, we confirmed expression of 39 known miRNA families (122 members and five conserved miRNA families, which showed differential regulation under drought stress. Targets of miRNAs were further surveyed based on a recently published study, of which ten targets were DRGs. An integrated gene regulatory network is proposed for the molecular mechanisms of tobacco root response to drought stress using differentially expressed DRGs, the changed expression profiles of miRNAs and their target transcripts. This network analysis serves as a reference for future studies on tobacco response stresses such as drought, cold and heavy metals.

Repression of Meiotic Genes by Antisense Transcription and by Fkh2 Transcription Factor in Schizosaccharomyces pombe

OpenAIRE

Chen, Huei-Mei; Rosebrock, Adam P.; Khan, Sohail R.; Futcher, Bruce; Leatherwood, Janet K.

2012-01-01

In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription ...

Transcription Factor KLF5 Binds a Cyclin E1 Polymorphic Intronic Enhancer to Confer Increased Bladder Cancer Risk

Science.gov (United States)

Pattison, Jillian M.; Posternak, Valeriya; Cole, Michael D.

2016-01-01

It is well established that environmental toxins, such as exposure to arsenic, are risk factors in the development of urinary bladder cancer, yet recent genome-wide association studies (GWAS) provide compelling evidence that there is a strong genetic component associated with disease predisposition. A single nucleotide polymorphism (SNP), rs8102137, was identified on chromosome 19q12, residing 6 kb upstream of the important cell cycle regulator and proto-oncogene, Cyclin E1 (CCNE1). However, the functional role of this variant in bladder cancer predisposition has been unclear since it lies within a non-coding region of the genome. Here, it is demonstrated that bladder cancer cells heterozygous for this SNP exhibit biased allelic expression of CCNE1 with 1.5-fold more transcription occurring from the risk allele. Furthermore, using chromatin immunoprecipitation assays, a novel enhancer element was identified within the first intron of CCNE1 that binds Kruppel-like Factor 5 (KLF5), a known transcriptional activator in bladder cancer. Moreover, the data reveal that the presence of rs200996365, a SNP in high linkage disequilibrium with rs8102137 residing in the center of a KLF5 motif, alters KLF5 binding to this genomic region. Through luciferase assays and CRISPR-Cas9 genome editing, a novel polymorphic intronic regulatory element controlling CCNE1 transcription is characterized. These studies uncover how a cancer-associated polymorphism mechanistically contributes to an increased predisposition for bladder cancer development. Implications A polymorphic KLF5 binding site near the CCNE1 gene explains genetic risk identified through genome wide association studies. PMID:27514407

16 CFR 5.63 - Evidence; transcript; in camera orders; proposed findings of fact and conclusions of law.

Science.gov (United States)

2010-01-01

... 16 Commercial Practices 1 2010-01-01 2010-01-01 false Evidence; transcript; in camera orders; proposed findings of fact and conclusions of law. 5.63 Section 5.63 Commercial Practices FEDERAL TRADE... findings of fact and conclusions of law. Sections 3.43, 3.44, 3.45, and 3.46 of the Commission's Rules of...

Reciprocal occupancy of BCL6 and STAT5 on Growth Hormone target genes: contrasting transcriptional outcomes and promoter-specific roles of p300 and HDAC3.

Science.gov (United States)

Lin, Grace; LaPensee, Christopher R; Qin, Zhaohui S; Schwartz, Jessica

2014-09-01

Expression of the Growth Hormone (GH)-stimulated gene Socs2 (Suppressor of Cytokine Signaling 2) is mediated by the transcription activator STAT5 (Signal Transducer and Activator of Transcription 5) and the transcription repressor BCL6 (B-Cell Lymphoma 6). ChIP-Sequencing identified Cish (Cytokine-Inducible SH2-containing protein) and Bcl6 as having similar patterns of reciprocal occupancy by BCL6 and STAT5 in response to GH, though GH stimulates Cish and inhibits Bcl6 expression. The co-activator p300 occupied Socs2, Cish and Bcl6 promoters, and enhanced STAT5-mediated activation of Socs2 and Cish. In contrast, on Bcl6, p300 functioned as a repressor and inhibited in conjunction with STAT5 or BCL6. The co-repressor HDAC3 (Histone deacetylase 3) inhibited the Socs2, Cish and Bcl6 promoters in the presence of STAT5. Thus transcriptional outcomes on GH-regulated genes occupied by BCL6 and STAT5 are determined in a promoter-specific fashion by co-regulatory proteins which mediate the distinction between activating and repressive transcription factors. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Uridine 5′-Monophosphate Synthase Is Transcriptionally Regulated by Pyrimidine Levels in Nicotiana plumbaginifolia1

Science.gov (United States)

Santoso, Djoko; Thornburg, Robert

1998-01-01

To understand the regulation and expression of pyrimidine biosynthesis in plants, we have examined the effect of the metabolic inhibitor 5-fluoroorotic acid (FOA) on uridine-5′-monophosphate synthase (UMPSase) expression in cell cultures of Nicotiana plumbaginifolia. UMPSase is the rate-limiting step of pyrimidine biosynthesis in plants. Addition of FOA causes an up-regulation of UMPSase enzyme activity in cell cultures after a lag phase of several days. Western-blot analysis demonstrated that the up-regulation in enzyme activity was caused by increased expression of the UMPSase protein. Northern-blot analysis demonstrated a higher level of UMPSase mRNA in the FOA-induced tissues than in control tissues. Run-on transcriptional assays showed that the UMPSase gene was transcriptionally activated after FOA treatment. The mechanism of toxicity of FOA is through thymine starvation. We found that addition of thymine abrogated the FOA-mediated up-regulation of UMPSase. In addition, methotrexate and aminopterin, which affect thymine levels by inhibiting dihydrofolate reductase, also up-regulate UMPSase in N. plumbaginifolia cells. PMID:9490773

Signal transducer and activator of transcription 5 activation is sufficient to drive transcriptional induction of cyclin D2 gene and proliferation of rat pancreatic beta-cells

DEFF Research Database (Denmark)

Friedrichsen, Birgitte N; Richter, Henrijette E; Hansen, Johnny A

2003-01-01

in a time-dependent manner by hGH in INS-1 cells. Inhibition of protein synthesis by coincubation with cycloheximide did not affect the hGH-induced increase of cyclin D2 mRNA levels at 4 h. Expression of a dominant negative STAT5 mutant, STAT5aDelta749, partially inhibited cyclin D2 protein levels. INS-1...... cells transiently transfected with a cyclin D2 promoter-reporter construct revealed a 3- to 5-fold increase of transcriptional activity in response to hGH stimulation. Furthermore, coexpression of a constitutive active STAT5 mutant (either CA-STAT5a or CA-STAT5b) was sufficient to drive transactivation...

Nucleoside Triphosphate Phosphohydrolase I (NPH I) Functions as a 5′ to 3′ Translocase in Transcription Termination of Vaccinia Early Genes*

Science.gov (United States)

Hindman, Ryan; Gollnick, Paul

2016-01-01

Vaccinia virus early genes are transcribed immediately upon infection. Nucleoside triphosphate phosphohydrolase I (NPH I) is an essential component of the early gene transcription complex. NPH I hydrolyzes ATP to release transcripts during transcription termination. The ATPase activity of NPH I requires single-stranded (ss) DNA as a cofactor; however, the source of this cofactor within the transcription complex is not known. Based on available structures of transcription complexes it has been hypothesized that the ssDNA cofactor is obtained from the unpaired non-template strand within the transcription bubble. In vitro transcription on templates that lack portions of the non-template strand within the transcription bubble showed that the upstream portion of the transcription bubble is required for efficient NPH I-mediated transcript release. Complementarity between the template and non-template strands in this region is also required for NPH I-mediated transcript release. This observation complicates locating the source of the ssDNA cofactor within the transcription complex because removal of the non-template strand also disrupts transcription bubble reannealing. Prior studies have shown that ssRNA binds to NPH I, but it does not activate ATPase activity. Chimeric transcription templates with RNA in the non-template strand confirm that the source of the ssDNA cofactor for NPH I is the upstream portion of the non-template strand in the transcription bubble. Consistent with this conclusion we also show that isolated NPH I acts as a 5′ to 3′ translocase on single-stranded DNA. PMID:27189950

Kub5-Hera, the human Rtt103 homolog, plays dual functional roles in transcription termination and DNA repair.

Science.gov (United States)

Morales, Julio C; Richard, Patricia; Rommel, Amy; Fattah, Farjana J; Motea, Edward A; Patidar, Praveen L; Xiao, Ling; Leskov, Konstantin; Wu, Shwu-Yuan; Hittelman, Walter N; Chiang, Cheng-Ming; Manley, James L; Boothman, David A

2014-04-01

Functions of Kub5-Hera (In Greek Mythology Hera controlled Artemis) (K-H), the human homolog of the yeast transcription termination factor Rtt103, remain undefined. Here, we show that K-H has functions in both transcription termination and DNA double-strand break (DSB) repair. K-H forms distinct protein complexes with factors that repair DSBs (e.g. Ku70, Ku86, Artemis) and terminate transcription (e.g. RNA polymerase II). K-H loss resulted in increased basal R-loop levels, DSBs, activated DNA-damage responses and enhanced genomic instability. Significantly lowered Artemis protein levels were detected in K-H knockdown cells, which were restored with specific K-H cDNA re-expression. K-H deficient cells were hypersensitive to cytotoxic agents that induce DSBs, unable to reseal complex DSB ends, and showed significantly delayed γ-H2AX and 53BP1 repair-related foci regression. Artemis re-expression in K-H-deficient cells restored DNA-repair function and resistance to DSB-inducing agents. However, R loops persisted consistent with dual roles of K-H in transcription termination and DSB repair.

RFX Transcription Factor DAF-19 Regulates 5-HT and Innate Immune Responses to Pathogenic Bacteria in Caenorhabditis elegans

Science.gov (United States)

Choi, Sunju; Xu, Lu; Sze, Ji Ying

2013-01-01

In Caenorhabditis elegans the Toll-interleukin receptor domain adaptor protein TIR-1 via a conserved mitogen-activated protein kinase (MAPK) signaling cascade induces innate immunity and upregulates serotonin (5-HT) biosynthesis gene tph-1 in a pair of ADF chemosensory neurons in response to infection. Here, we identify transcription factors downstream of the TIR-1 signaling pathway. We show that common transcription factors control the innate immunity and 5-HT biosynthesis. We demonstrate that a cysteine to tyrosine substitution in an ARM motif of the HEAT/Arm repeat region of the TIR-1 protein confers TIR-1 hyperactivation, leading to constitutive tph-1 upregulation in the ADF neurons, increased expression of intestinal antimicrobial genes, and enhanced resistance to killing by the human opportunistic pathogen Pseudomonas aeruginosa PA14. A forward genetic screen for suppressors of the hyperactive TIR-1 led to the identification of DAF-19, an ortholog of regulatory factor X (RFX) transcription factors that are required for human adaptive immunity. We show that DAF-19 concerts with ATF-7, a member of the activating transcription factor (ATF)/cAMP response element-binding B (CREB) family of transcription factors, to regulate tph-1 and antimicrobial genes, reminiscent of RFX-CREB interaction in human immune cells. daf-19 mutants display heightened susceptibility to killing by PA14. Remarkably, whereas the TIR-1-MAPK-DAF-19/ATF-7 pathway in the intestinal immunity is regulated by DKF-2/protein kinase D, we found that the regulation of tph-1 expression is independent of DKF-2 but requires UNC-43/Ca2+/calmodulin-dependent protein kinase (CaMK) II. Our results suggest that pathogenic cues trigger a common core-signaling pathway via tissue-specific mechanisms and demonstrate a novel role for RFX factors in neuronal and innate immune responses to infection. PMID:23505381

Dialogue between E. coli free radical pathways and the mitochondria of C. elegans.

Science.gov (United States)

Govindan, J Amaranath; Jayamani, Elamparithi; Zhang, Xinrui; Mylonakis, Eleftherios; Ruvkun, Gary

2015-10-06

The microbial world presents a complex palette of opportunities and dangers to animals, which have developed surveillance and response strategies to hints of microbial intent. We show here that the mitochondrial homeostatic response pathway of the nematode Caenorhabditis elegans responds to Escherichia coli mutations that activate free radical detoxification pathways. Activation of C. elegans mitochondrial responses could be suppressed by additional mutations in E. coli, suggesting that C. elegans responds to products of E. coli to anticipate challenges to its mitochondrion. Out of 50 C. elegans gene inactivations known to mediate mitochondrial defense, we found that 7 genes were required for C. elegans response to a free radical producing E. coli mutant, including the bZip transcription factor atfs-1 (activating transcription factor associated with stress). An atfs-1 loss-of-function mutant was partially resistant to the effects of free radical-producing E. coli mutant, but a constitutively active atfs-1 mutant growing on wild-type E. coli inappropriately activated the pattern of mitochondrial responses normally induced by an E. coli free radical pathway mutant. Carbonylated proteins from free radical-producing E. coli mutant may directly activate the ATFS-1/bZIP transcription factor to induce mitochondrial stress response: feeding C. elegans with H2O2-treated E. coli induces the mitochondrial unfolded protein response, and inhibition of a gut peptide transporter partially suppressed C. elegans response to free radical damaged E. coli.

SIRT1 deacetylates RFX5 and antagonizes repression of collagen type I (COL1A2) transcription in smooth muscle cells

Energy Technology Data Exchange (ETDEWEB)

Xia, Jun [Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University (China); Department of Respiratory Medicine, Jiangsu Provincial Hospital of Chinese Traditional Medicine (China); Wu, Xiaoyan; Yang, Yuyu; Zhao, Yuhao [Atherosclerosis Research Center, Key Laboratory of Cardiovascular Disease and Molecular Intervention, Department of Pathophysiology, Nanjing Medical University (China); Fang, Mingming [Jiangsu Jiankang Vocational Institute (China); Xie, Weiping, E-mail: wpxienjmu@gmail.com [Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University (China); Wang, Hong, E-mail: hwangnjmu@gmail.com [Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University (China); Xu, Yong [Atherosclerosis Research Center, Key Laboratory of Cardiovascular Disease and Molecular Intervention, Department of Pathophysiology, Nanjing Medical University (China)

2012-11-16

Highlights: Black-Right-Pointing-Pointer SIRT1 interacts with and deacetylates RFX5. Black-Right-Pointing-Pointer SIRT1 activation attenuates whereas SIRT1 inhibition enhances collagen repression by RFX5 in vascular smooth muscle cells. Black-Right-Pointing-Pointer SIRT1 promotes cytoplasmic localization and proteasomal degradation of RFX5 and cripples promoter recruitment of RFX5. Black-Right-Pointing-Pointer IFN-{gamma} represses SIRT1 expression in vascular smooth muscle cells. Black-Right-Pointing-Pointer SIRT1 agonist alleviates collagen repression by IFN-{gamma} in vascular smooth muscle cells. -- Abstract: Decreased expression of collagen by vascular smooth muscle cells (SMCs) within the atherosclerotic plaque contributes to the thinning of the fibrous cap and poses a great threat to plaque rupture. Elucidation of the mechanism underlying repressed collagen type I (COL1A2) gene would potentially provide novel solutions that can prevent rupture-induced complications. We have previously shown that regulatory factor for X-box (RFX5) binds to the COL1A2 transcription start site and represses its transcription. Here we report that SIRT1, an NAD-dependent, class III deacetylase, forms a complex with RFX5. Over-expression of SIRT1 or NAMPT, which synthesizes NAD+ to activate SIRT1, or treatment with the SIRT1 agonist resveratrol decreases RFX5 acetylation and disrupts repression of the COL1A2 promoter activity by RFX5. On the contrary, knockdown of SIRT1 or treatment with SIRT1 inhibitors induces RFX5 acetylation and enhances the repression of collagen transcription. SIRT1 antagonizes RFX5 activity by promoting its nuclear expulsion and proteasomal degradation hence dampening its binding to the COL1A2 promoter. The pro-inflammatory cytokine IFN-{gamma} represses COL1A2 transcription by down-regulating SIRT1 expression in SMCs. Therefore, our data have identified as novel pathway whereby SIRT1 maintains collagen synthesis in SMCs by modulating RFX5 activity.

Two Novel Variants Affecting CDKL5 Transcript Associated with Epileptic Encephalopathy.

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Neupauerová, Jana; Štěrbová, Katalin; Vlčková, Markéta; Sebroňová, Věra; Maříková, Tat'ána; Krůtová, Marcela; David, Staněk; Kršek, Pavel; Žaliová, Markéta; Seeman, Pavel; Laššuthová, Petra

2017-10-01

Variants in the human X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been reported as being etiologically associated with early infantile epileptic encephalopathy type 2 (EIEE2). We report on two patients, a boy and a girl, with EIEE2 that present with early onset epilepsy, hypotonia, severe intellectual disability, and poor eye contact. Massively parallel sequencing (MPS) of a custom-designed gene panel for epilepsy and epileptic encephalopathy containing 112 epilepsy-related genes was performed. Sanger sequencing was used to confirm the novel variants. For confirmation of the functional consequence of an intronic CDKL5 variant in patient 2, an RNA study was done. DNA sequencing revealed de novo variants in CDKL5, a c.2578C>T (p. Gln860*) present in a hemizygous state in a 3-year-old boy, and a potential splice site variant c.463+5G>A in heterozygous state in a 5-year-old girl. Multiple in silico splicing algorithms predicted a highly reduced splice site score for c.463+5G>A. A subsequent mRNA study confirmed an aberrant shorter transcript lacking exon 7. Our data confirmed that variants in the CDKL5 are associated with EIEE2. There is credible evidence that the novel identified variants are pathogenic and, therefore, are likely the cause of the disease in the presented patients. In one of the patients a stop codon variant is predicted to produce a truncated protein, and in the other patient an intronic variant results in aberrant splicing.

Dose-dependent and gender-related radiation-induced transcription alterations of Gadd45a and Ier5 in human lymphocytes exposed to gamma ray emitted by 60Co

International Nuclear Information System (INIS)

Tavakoli, H.; Manoochehri, M.; Mosalla, S. M. M.; Ghafori, M.; Karimi, A. A.

2013-01-01

Growth arrest DNA damage-inducible 45a gene (Gadd45a) and immediate early response gene 5 (Ier5) have been emphasised as ideal radiation bio-markers in several reports. However, some aspects of radiation-induced transcriptional alterations of these genes are unknown. In this study, gender-dependency and dose-dependency as two factors that may affect radiation induced transcription of Gadd45a and Ier5 genes were investigated. Human lymphocyte cells from six healthy voluntary blood donors (three women and three men) were irradiated in vitro with doses of 0.5-4.0 Gy from a 60 Co source and RNA isolated 4 h later using the High Pure RNA Isolation Kit. Dose and gender dependency of radiation-induced transcriptional alterations of Gadd45a and Ier5 genes were studied by quantitative real-time polymerase chain reaction. The results showed that as a whole, Gadd45a and Ier5 gave responses to gamma rays, while the responses were independent of radiation doses. Therefore, regardless of radiation dose, Gadd45a and Ier5 can be considered potential radiation bio-markers. Besides, although radiation-induced transcriptional alterations of Gadd45a in female and male lymphocyte samples were insignificant at 0.5 Gy, at other doses, their quantities in female samples were at a significantly higher level than in male samples. Radiation induced transcription of Ier5 of females samples had a reduction in comparison with male samples at 1 and 2 Gy, but at doses of 0.5 and 4 Gy, females were significantly more susceptible to radiation-induced transcriptional alteration of Ier5. (authors)

The membrane tethered transcription factor EcbZIP17 from finger millet promotes plant growth and enhances tolerance to abiotic stresses.

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Ramakrishna, Chopperla; Singh, Sonam; Raghavendrarao, Sangala; Padaria, Jasdeep C; Mohanty, Sasmita; Sharma, Tilak Raj; Solanke, Amolkumar U

2018-02-01

The occurrence of various stresses, as the outcome of global climate change, results in the yield losses of crop plants. Prospecting of genes in stress tolerant plant species may help to protect and improve their agronomic performance. Finger millet (Eleusine coracana L.) is a valuable source of superior genes and alleles for stress tolerance. In this study, we isolated a novel endoplasmic reticulum (ER) membrane tethered bZIP transcription factor from finger millet, EcbZIP17. Transgenic tobacco plants overexpressing this gene showed better vegetative growth and seed yield compared with wild type (WT) plants under optimal growth conditions and confirmed upregulation of brassinosteroid signalling genes. Under various abiotic stresses, such as 250 mM NaCl, 10% PEG6000, 400 mM mannitol, water withdrawal, and heat stress, the transgenic plants showed higher germination rate, biomass, primary and secondary root formation, and recovery rate, compared with WT plants. The transgenic plants exposed to an ER stress inducer resulted in greater leaf diameter and plant height as well as higher expression of the ER stress-responsive genes BiP, PDIL, and CRT1. Overall, our results indicated that EcbZIP17 improves plant growth at optimal conditions through brassinosteroid signalling and provide tolerance to various environmental stresses via ER signalling pathways.

THAP5 is a DNA-binding transcriptional repressor that is regulated in melanoma cells during DNA damage-induced cell death

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Balakrishnan, Meenakshi P.; Cilenti, Lucia; Ambivero, Camilla [Biomolecular Science Center, Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL (United States); Goto, Yamafumi [Department of Dermatology, Shinshu University School of Medicine, Matsumoto (Japan); Takata, Minoru [Department of Dermatology, Okayama University Graduate School of Medical Dentistry and Pharmaceutical Sciences, Okayama (Japan); Turkson, James; Li, Xiaoman Shawn [Biomolecular Science Center, Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL (United States); Zervos, Antonis S., E-mail: azervos@mail.ucf.edu [Biomolecular Science Center, Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL (United States)

2011-01-07

Research highlights: {yields} THAP5 is a DNA-binding protein and a transcriptional repressor. {yields} THAP5 is induced in melanoma cells upon exposure to UV or treatment with cisplatin. {yields} THAP5 induction correlates with the degree of apoptosis in melanoma cell population. {yields} THAP5 is a pro-apoptotic protein involved in melanoma cell death. -- Abstract: THAP5 was originally isolated as a specific interactor and substrate of the mitochondrial pro-apoptotic Omi/HtrA2 protease. It is a human zinc finger protein characterized by a restricted pattern of expression and the lack of orthologs in mouse and rat. The biological function of THAP5 is unknown but our previous studies suggest it could regulate G2/M transition in kidney cells and could be involved in human cardiomyocyte cell death associated with coronary artery disease (CAD). In this report, we expanded our studies on the properties and function of THAP5 in human melanoma cells. THAP5 was expressed in primary human melanocytes as well as in all melanoma cell lines that were tested. THAP5 protein level was significantly induced by UV irradiation or cisplatin treatment, conditions known to cause DNA damage. The induction of THAP5 correlated with a significant increase in apoptotic cell death. In addition, we show that THAP5 is a nuclear protein that could recognize and bind a specific DNA motif. THAP5 could also repress the transcription of a reporter gene in a heterologous system. Our work suggests that THAP5 is a DNA-binding protein and a transcriptional repressor. Furthermore, THAP5 has a pro-apoptotic function and it was induced in melanoma cells under conditions that promoted cell death.

THAP5 is a DNA-binding transcriptional repressor that is regulated in melanoma cells during DNA damage-induced cell death

International Nuclear Information System (INIS)

Balakrishnan, Meenakshi P.; Cilenti, Lucia; Ambivero, Camilla; Goto, Yamafumi; Takata, Minoru; Turkson, James; Li, Xiaoman Shawn; Zervos, Antonis S.

2011-01-01

Research highlights: → THAP5 is a DNA-binding protein and a transcriptional repressor. → THAP5 is induced in melanoma cells upon exposure to UV or treatment with cisplatin. → THAP5 induction correlates with the degree of apoptosis in melanoma cell population. → THAP5 is a pro-apoptotic protein involved in melanoma cell death. -- Abstract: THAP5 was originally isolated as a specific interactor and substrate of the mitochondrial pro-apoptotic Omi/HtrA2 protease. It is a human zinc finger protein characterized by a restricted pattern of expression and the lack of orthologs in mouse and rat. The biological function of THAP5 is unknown but our previous studies suggest it could regulate G2/M transition in kidney cells and could be involved in human cardiomyocyte cell death associated with coronary artery disease (CAD). In this report, we expanded our studies on the properties and function of THAP5 in human melanoma cells. THAP5 was expressed in primary human melanocytes as well as in all melanoma cell lines that were tested. THAP5 protein level was significantly induced by UV irradiation or cisplatin treatment, conditions known to cause DNA damage. The induction of THAP5 correlated with a significant increase in apoptotic cell death. In addition, we show that THAP5 is a nuclear protein that could recognize and bind a specific DNA motif. THAP5 could also repress the transcription of a reporter gene in a heterologous system. Our work suggests that THAP5 is a DNA-binding protein and a transcriptional repressor. Furthermore, THAP5 has a pro-apoptotic function and it was induced in melanoma cells under conditions that promoted cell death.

Transcriptional dysregulation of 5-HT1A autoreceptors in mental illness

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Albert Paul R

2011-05-01

Full Text Available Abstract The serotonin-1A (5-HT1A receptor is among the most abundant and widely distributed 5-HT receptors in the brain, but is also expressed on serotonin neurons as an autoreceptor where it plays a critical role in regulating the activity of the entire serotonin system. Over-expression of the 5-HT1A autoreceptor has been implicated in reducing serotonergic neurotransmission, and is associated with major depression and suicide. Extensive characterization of the transcriptional regulation of the 5-HT1A gene (HTR1A using cell culture systems has revealed a GC-rich "housekeeping" promoter that non-selectively drives its expression; this is flanked by a series of upstream repressor elements for REST, Freud-1/CC2D1A and Freud-2/CC2D1B factors that not only restrict its expression to neurons, but may also regulate the level of expression of 5-HT1A receptors in various subsets of neurons, including serotonergic neurons. A separate set of allele-specific factors, including Deaf1, Hes1 and Hes5 repress at the HTR1A C(-1019G (rs6295 polymorphism in serotonergic neurons in culture, as well as in vivo. Pet1, an obligatory enhancer for serotonergic differentiation, has been identified as a potent activator of 5-HT1A autoreceptor expression. Taken together, these results highlight an integrated regulation of 5-HT1A autoreceptors that differs in several aspects from regulation of post-synaptic 5-HT1A receptors, and could be selectively targeted to enhance serotonergic neurotransmission.

Glucose transformation to 5-hydroxymethylfurfural in acidic ionic liquid: A quantum mechanical study.

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Arifin; Puripat, Maneeporn; Yokogawa, Daisuke; Parasuk, Vudhichai; Irle, Stephan

2016-01-30

Isomerization and transformation of glucose and fructose to 5-hydroxymethylfurfural (HMF) in both ionic liquids (ILs) and water has been studied by the reference interaction site model self-consistent field spatial electron density distribution (RISM-SCF-SEDD) method coupled with ab initio electronic structure theory, namely coupled cluster single, double, and perturbative triple excitation (CCSD(T)). Glucose isomerization to fructose has been investigated via cyclic and open chain mechanisms. In water, the calculations support the cyclic mechanism of glucose isomerization; with the predicted activation free energy is 23.8 kcal mol(-1) at experimental condition. Conversely, open ring mechanism is more favorable in ILs with the energy barrier is 32.4 kcal mol(-1) . Moreover, the transformation of fructose into HMF via cyclic mechanism is reasonable; the calculated activation barriers are 16.0 and 21.5 kcal mol(-1) in aqueous and ILs solutions, respectively. The solvent effects of ILs could be explained by the decomposition of free energies and radial distribution functions of solute-solvent that are produced by RISM-SCF-SEDD. © 2015 Wiley Periodicals, Inc.

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.

The transcription factor VvMYB5b contributes to the regulation of anthocyanin and proanthocyanidin biosynthesis in developing grape berries.

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Deluc, Laurent; Bogs, Jochen; Walker, Amanda R; Ferrier, Thilia; Decendit, Alain; Merillon, Jean-Michel; Robinson, Simon P; Barrieu, François

2008-08-01

Among the dramatic changes occurring during grape berry (Vitis vinifera) development, those affecting the flavonoid pathway have provoked a number of investigations in the last 10 years. In addition to producing several compounds involved in the protection of the berry and the dissemination of the seeds, final products of this pathway also play a critical role in berry and wine quality. In this article, we describe the cloning and functional characterization of VvMYB5b, a cDNA isolated from a grape berry (V. vinifera 'Cabernet Sauvignon') library. VvMYB5b encodes a protein belonging to the R2R3-MYB family of transcription factors and displays significant similarity with VvMYB5a, another MYB factor recently shown to regulate flavonoid synthesis in grapevine. The ability of VvMYB5a and VvMYB5b to activate the grapevine promoters of several structural genes of the flavonoid pathway was confirmed by transient expression of the corresponding cDNAs in grape cells. Overexpression of VvMYB5b in tobacco (Nicotiana tabacum) leads to an up-regulation of genes encoding enzymes of the flavonoid pathway and results in the accumulation of anthocyanin- and proanthocyanidin-derived compounds. The ability of VvMYB5b to regulate particularly the anthocyanin and the proanthocyanidin pathways is discussed in relation to other recently characterized MYB transcription factors in grapevine. Taken together, data presented in this article give insight into the transcriptional mechanisms associated with the regulation of the flavonoid pathway throughout grape berry development.

Specificity determinants for the abscisic acid response element.

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Sarkar, Aditya Kumar; Lahiri, Ansuman

2013-01-01

Abscisic acid (ABA) response elements (ABREs) are a group of cis-acting DNA elements that have been identified from promoter analysis of many ABA-regulated genes in plants. We are interested in understanding the mechanism of binding specificity between ABREs and a class of bZIP transcription factors known as ABRE binding factors (ABFs). In this work, we have modeled the homodimeric structure of the bZIP domain of ABRE binding factor 1 from Arabidopsis thaliana (AtABF1) and studied its interaction with ACGT core motif-containing ABRE sequences. We have also examined the variation in the stability of the protein-DNA complex upon mutating ABRE sequences using the protein design algorithm FoldX. The high throughput free energy calculations successfully predicted the ability of ABF1 to bind to alternative core motifs like GCGT or AAGT and also rationalized the role of the flanking sequences in determining the specificity of the protein-DNA interaction.

The Polycomb Group Protein L3MBTL1 Represses a SMAD5-Mediated Hematopoietic Transcriptional Program in Human Pluripotent Stem Cells

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Fabiana Perna

2015-04-01

Full Text Available Epigenetic regulation of key transcriptional programs is a critical mechanism that controls hematopoietic development, and, thus, aberrant expression patterns or mutations in epigenetic regulators occur frequently in hematologic malignancies. We demonstrate that the Polycomb protein L3MBTL1, which is monoallelically deleted in 20q- myeloid malignancies, represses the ability of stem cells to drive hematopoietic-specific transcriptional programs by regulating the expression of SMAD5 and impairing its recruitment to target regulatory regions. Indeed, knockdown of L3MBTL1 promotes the development of hematopoiesis and impairs neural cell fate in human pluripotent stem cells. We also found a role for L3MBTL1 in regulating SMAD5 target gene expression in mature hematopoietic cell populations, thereby affecting erythroid differentiation. Taken together, we have identified epigenetic priming of hematopoietic-specific transcriptional networks, which may assist in the development of therapeutic approaches for patients with anemia.

Hoxa5 overexpression correlates with IGFBP1 upregulation and postnatal dwarfism: evidence for an interaction between Hoxa5 and Forkhead box transcription factors.

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Foucher, Isabelle; Volovitch, Michel; Frain, Monique; Kim, J Julie; Souberbielle, Jean-Claude; Gan, Lixia; Unterman, Terry G; Prochiantz, Alain; Trembleau, Alain

2002-09-01

Transgenic mice expressing the homeobox gene Hoxa5 under the control of Hoxb2 regulatory elements present a growth arrest during weeks two and three of postnatal development, resulting in proportionate dwarfism. These mice present a liver phenotype illustrated by a 12-fold increase in liver insulin-like growth factor binding protein 1 (IGFBP1) mRNA and a 50% decrease in liver insulin-like growth factor 1 (IGF1) mRNA correlated with a 50% decrease in circulating IGF1. We show that the Hoxa5 transgene is expressed in the liver of these mice, leading to an overexpression of total (endogenous plus transgene) Hoxa5 mRNA in this tissue. We have used several cell lines to investigate a possible physiological interaction of Hoxa5 with the main regulator of IGFBP1 promoter activity, the Forkhead box transcription factor FKHR. In HepG2 cells, Hoxa5 has little effect by itself but inhibits the FKHR-dependent activation of the IGFBP1 promoter. In HuF cells, Hoxa5 cooperates with FKHR to dramatically enhance IGFBP1 promoter activity. This context-dependent physiological interaction probably corresponds to the existence of a direct interaction between Hoxa5 and FKHR and FoxA2/HNF3beta, as demonstrated by pull-down experiments achieved either in vitro or after cellular co-expression. In conclusion, we propose that the impaired growth observed in this transgenic line relates to a liver phenotype best explained by a direct interaction between Hoxa5 and liver-specific Forkhead box transcription factors, in particular FKHR but also Foxa2/HNF3beta. Because Hoxa5 and homeogenes of the same paralog group are normally expressed in the liver, the present results raise the possibility that homeoproteins, in addition to their established role during early development, regulate systemic physiological functions.

Recent advances in utilizing transcription factors to improve plant abiotic stress tolerance by transgenic technology

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Hongyan eWang

2016-02-01

Full Text Available Agricultural production and quality are adversely affected by various abiotic stresses worldwide and this will be exacerbated by the deterioration of global climate. To feed a growing world population, it is very urgent to breed stress-tolerant crops with higher yields and improved qualities against multiple environmental stresses. Since conventional breeding approaches had marginal success due to the complexity of stress tolerance traits, the transgenic approach is now being popularly used to breed stress-tolerant crops. So identifying and characterizing the the critical genes involved in plant stress responses is an essential prerequisite for engineering stress-tolerant crops. Far beyond the manipulation of single functional gene, engineering certain regulatory genes has emerged as an effective strategy now for controlling the expression of many stress-responsive genes. Transcription factors (TFs are good candidates for genetic engineering to breed stress-tolerant crop because of their role as master regulators of many stress-responsive genes. Many TFs belonging to families AP2/EREBP, MYB, WRKY, NAC, bZIP have been found to be involved in various abiotic stresses and some TF genes have also been engineered to improve stress tolerance in model and crop plants. In this review, we take five large families of TFs as examples and review the recent progress of TFs involved in plant abiotic stress responses and their potential utilization to improve multiple stress tolerance of crops in the field conditions.

A single amino acid change within the R2 domain of the VvMYB5b transcription factor modulates affinity for protein partners and target promoters selectivity

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Granier Thierry

2011-08-01

Full Text Available Abstract Background Flavonoid pathway is spatially and temporally controlled during plant development and the transcriptional regulation of the structural genes is mostly orchestrated by a ternary protein complex that involves three classes of transcription factors (R2-R3-MYB, bHLH and WDR. In grapevine (Vitis vinifera L., several MYB transcription factors have been identified but the interactions with their putative bHLH partners to regulate specific branches of the flavonoid pathway are still poorly understood. Results In this work, we describe the effects of a single amino acid substitution (R69L located in the R2 domain of VvMYB5b and predicted to affect the formation of a salt bridge within the protein. The activity of the mutated protein (name VvMYB5bL, the native protein being referred as VvMYB5bR was assessed in different in vivo systems: yeast, grape cell suspensions, and tobacco. In the first two systems, VvMYB5bL exhibited a modified trans-activation capability. Moreover, using yeast two-hybrid assay, we demonstrated that modification of VvMYB5b transcriptional properties impaired its ability to correctly interact with VvMYC1, a grape bHLH protein. These results were further substantiated by overexpression of VvMYB5bR and VvMYB5bL genes in tobacco. Flowers from 35S::VvMYB5bL transgenic plants showed a distinct phenotype in comparison with 35S::VvMYB5bR and the control plants. Finally, significant differences in transcript abundance of flavonoid metabolism genes were observed along with variations in pigments accumulation. Conclusions Taken together, our findings indicate that VvMYB5bL is still able to bind DNA but the structural consequences linked to the mutation affect the capacity of the protein to activate the transcription of some flavonoid genes by modifying the interaction with its co-partner(s. In addition, this study underlines the importance of an internal salt bridge for protein conformation and thus for the establishment

BCR-crosslinking induces a transcription of protein phosphatase component G5PR that is required for mature B-cell survival

International Nuclear Information System (INIS)

Huq Ronny, Faisal Mahmudul; Igarashi, Hideya; Sakaguchi, Nobuo

2006-01-01

BCR-crosslinking triggers activation-induced cell death (AICD) selectively in the restricted stage of B-cell differentiation. We examined the transcription of a protein phosphatase subunit G5PR in immature and mature B-cells, because absence of this factor augmented cell sensitivity to AICD, associated with increased activation of JNK and Bim. BCR-crosslinking-induced G5pr transcription in AICD-resistant mature splenic IgM lo IgD hi B-cells but not in AICD susceptible immature IgM hi IgD lo B-cells. Thus, G5pr induction correlated with the prevention of AICD; High in mature splenic CD23 hi B-cells but low in immature B-cells of neonatal mice, sub-lethally irradiated mice, or xid mice. Lack of G5pr upregulation was associated with the prolonged activation of JNK. The G5pr cDNA transfection protected an immature B-cell line WEHI-231 from BCR-mediated AICD. The differential expression of G5PR might be responsible for the antigen-dependent selection of B-cells

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

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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.

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.

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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.

Effects of stem cell factor on hypoxia-inducible factor 1 alpha accumulation in human acute myeloid leukaemia and LAD2 mast cells.

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Bernhard F Gibbs

Full Text Available Stem cell factor (SCF is a hematopoietic growth factor that exerts its activity by signalling through the tyrosine kinase receptor known as Kit or CD117. SCF-Kit signalling is crucial for the survival, proliferation and differentiation of hematopoietic cells of myeloid lineage. Furthermore, since myeloid leukaemia cells express the Kit receptor, SCF may play an important role in myeloid leukaemia progression too. However, the mechanisms of this pathophysiological effect remain unclear. Recent evidence shows that SCF triggers accumulation of the inducible alpha subunit of hypoxia-inducible factor 1 (HIF-1 in hematopoietic cells--a transcription complex that plays a pivotal role in cellular adaptation to low oxygen availability. However, it is unknown how SCF impacts on HIF-1α accumulation in human myeloid leukaemia and mast cells. Here we show that SCF induces HIF-1α accumulation in THP-1 human myeloid leukaemia cells but not in LAD2 mast cells. We demonstrated that LAD2 cells have a more robust glutathione (GSH-dependent antioxidative system compared to THP-1 cells and are therefore protected against the actions of ROS generated in an SCF-dependent manner. BSO-induced GSH depletion led to a significant decrease in HIF-1α prolyl hydroxylase (PHD activity in THP-1 cells and to near attenuation of it in LAD2 cells. In THP-1 cells, SCF-induced HIF-1α accumulation is controlled via ERK, PI3 kinase/PKC-δ/mTOR-dependent and to a certain extent by redox-dependent mechanisms. These results demonstrate for the first time an important cross-talk of signalling pathways associated with HIF-1 activation--an important stage of the myeloid leukaemia cell life cycle.

Validation of aspirin response-related transcripts in patients with coronary artery disease and preliminary investigation on CMTM5 function.

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Zhang, J W; Liu, T F; Chen, X H; Liang, W Y; Feng, X R; Wang, L; Fu, Sidney W; McCaffrey, Timothy A; Liu, M L

2017-08-15

Aspirin is widely used in the prevention of cardiovascular diseases, but the antiplatelet responses vary from one patient to another. To validate aspirin response related transcripts and illustrate their roles in predicting cardiovascular events, we have quantified the relative expression of 14 transcripts previously identified as related to high on-aspirin platelet reactivity (HAPR) in 223 patients with coronary artery disease (CAD) on regular aspirin treatment. All patients were followed up regularly for cardiovascular events (CVE). The mean age of our enrolled population was 75.80±8.57years. HAPR patients showed no significant differences in terms of co-morbidities and combined drugs. Besides, the relative expression of HLA-DQA1 was significantly lower in low on-aspirin platelet reactivity (LAPR) patients, when compared with HAPR and high normal (HN) group (p=0.028). What's more, the number of arteries involved, HAPR status and the relative expression of CLU, CMTM5 and SPARC were independent risk factors for CVE during follow up (p<0.05). In addition, overexpression of CMTM5 attenuated endothelial cells (ECs) migration and proliferation, with significantly decreased phosphorylated-Akt levels, while its inhibition promoted these processes in vitro (p<0.05).Our study provides evidence that circulating transcripts might be potential biomarkers in predicting cardiovascular events. CMTM5 might exert anti-atherosclerotic effects via suppressing migration and proliferation in the vessel wall. Nevertheless, larger-scale and long-term studies are still needed. Copyright © 2017 Elsevier B.V. All rights reserved.

Synthesis of in vitro Co1E1 transcripts with 5'-terminal ribonucleotides that exhibit noncomplementarity with the DNA template

International Nuclear Information System (INIS)

Parker, R.C.

1986-01-01

A region that forms the S1 nuclease site in Co1E1 DNA is shown to code for an in vitro transcript, called S1 RNA-B, which contains a 5'-terminal GTP residue that exhibits noncomplementarity with the template's DNA sequence. The synthesis of S1 RNA-B initiates four bases upstream from the start point for S1 RNA-C. The initial four bases in S1 RNA-B and S1 RNA-C are identical. The relative synthesis of S1 RNA-B to S1 RNA-C is sensitive to the concentration of GTP, a substrate that is required for elongation past the +4 position in S1 RNA-C. Dinucleotides that are expected to only initiate synthesis of S1 RNA-C yield two transcripts that appear to initiate from the S1 RNA-C and S1 RNA-B start sites. In vitro studies involving other Co1E1 transcripts, RNA-B and RNA-C, provide similar observations concerning the noncomplementary initiation phenomenon. A model involving transcriptional slippage is suggested to explain the noncomplementary initiation phenomenon. The model proposes that the cycling reaction of Escherichia coli RNA polymerase produces tetranucleotides that are transposed to nearby upstream sequences for priming transcription

DOT/FAA Human Factors Workshop on Aviation (5th). Transcript.

Science.gov (United States)

1982-01-01

This document is a verbatim transcript of the proceedings of the Fifth Human Factors Workshop held at the Mike Monroney Aeronautical Center in Oklahoma City, Oklahoma, on July 7-9, 1981. The Sixth Human Factors Workshop was held at the same facility ...

17β-estradiol-induced regulation of the novel 5-HT1A-related transcription factors NUDR and Freud-1 in SH SY5Y cells.

Science.gov (United States)

Adeosun, Samuel O; Albert, Paul R; Austin, Mark C; Iyo, Abiye H

2012-05-01

Nuclear deformed epidermal autoregulatory factor-1 (NUDR/Deaf-1) and five prime repressor element under dual repression (Freud-1) are novel transcriptional regulators of the 5-HT(1A) receptor, a receptor that has been implicated in the pathophysiology of various psychiatric illnesses. The antidepressant effect of 17β-Estradiol (17βE(2)) is purported to involve the downregulation of this receptor. We investigated the possible role of NUDR and Freud-1 in 17βE(2)-induced downregulation of the 5-HT(1A) receptor in the neuroblastoma cell line SH SY5Y. Cells were treated with 10 nM of 17βE(2) for 3 or 48 h, followed by a 24-h withdrawal period. Proteins were isolated and analyzed by western blotting. 17βE(2) treatment increased NUDR immunoreactivity while Freud-1 and the 5-HT(1A) receptor showed significant decreases. Upon withdrawal of 17βE(2), protein expression returned to control levels, except for NUDR, which remained significantly elevated in the 3-h treatment. Taken together, these data support a non-genomic downregulation of 5-HT(1A) receptor protein by 17βE(2), which does not involve NUDR and Freud-1. Rather, changes in both transcription factors seem to be compensatory/homeostatic responses to changes in 5-HT(1A) receptor induced by 17βE(2). These observations further highlight the importance of NUDR and Freud-1 in regulating 5-HT(1A) receptor expression.

A nitrogen response pathway regulates virulence functions in Fusarium oxysporum via the protein kinase TOR and the bZIP protein MeaB.

Science.gov (United States)

López-Berges, Manuel S; Rispail, Nicolas; Prados-Rosales, Rafael C; Di Pietro, Antonio

2010-07-01

During infection, fungal pathogens activate virulence mechanisms, such as host adhesion, penetration and invasive growth. In the vascular wilt fungus Fusarium oxysporum, the mitogen-activated protein kinase Fmk1 is required for plant infection and controls processes such as cellophane penetration, vegetative hyphal fusion, or root adhesion. Here, we show that these virulence-related functions are repressed by the preferred nitrogen source ammonium and restored by treatment with l-methionine sulfoximine or rapamycin, two specific inhibitors of Gln synthetase and the protein kinase TOR, respectively. Deletion of the bZIP protein MeaB also resulted in nitrogen source-independent activation of virulence mechanisms. Activation of these functions did not require the global nitrogen regulator AreA, suggesting that MeaB-mediated repression of virulence functions does not act through inhibition of AreA. Tomato plants (Solanum lycopersicum) supplied with ammonium rather than nitrate showed a significant reduction in vascular wilt symptoms when infected with the wild type but not with the DeltameaB strain. Nitrogen source also affected invasive growth in the rice blast fungus Magnaporthe oryzae and the wheat head blight pathogen Fusarium graminearum. We propose that a conserved nitrogen-responsive pathway might operate via TOR and MeaB to control virulence in plant pathogenic fungi.

Non-canonical transcription initiation: the expanding universe of transcription initiating substrates.

Science.gov (United States)

Barvík, Ivan; Rejman, Dominik; Panova, Natalya; Šanderová, Hana; Krásný, Libor

2017-03-01

RNA polymerase (RNAP) is the central enzyme of transcription of the genetic information from DNA into RNA. RNAP recognizes four main substrates: ATP, CTP, GTP and UTP. Experimental evidence from the past several years suggests that, besides these four NTPs, other molecules can be used to initiate transcription: (i) ribooligonucleotides (nanoRNAs) and (ii) coenzymes such as NAD+, NADH, dephospho-CoA and FAD. The presence of these molecules at the 5΄ ends of RNAs affects the properties of the RNA. Here, we discuss the expanding portfolio of molecules that can initiate transcription, their mechanism of incorporation, effects on RNA and cellular processes, and we present an outlook toward other possible initiation substrates. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

All 17 S-locus F-box proteins of the S2 - and S3 -haplotypes of Petunia inflata are assembled into similar SCF complexes with a specific function in self-incompatibility.

Science.gov (United States)

Li, Shu; Williams, Justin S; Sun, Penglin; Kao, Teh-Hui

2016-09-01

The collaborative non-self-recognition model for S-RNase-based self-incompatibility predicts that multiple S-locus F-box proteins (SLFs) produced by pollen of a given S-haplotype collectively mediate ubiquitination and degradation of all non-self S-RNases, but not self S-RNases, in the pollen tube, thereby resulting in cross-compatible pollination but self-incompatible pollination. We had previously used pollen extracts containing GFP-fused S2 -SLF1 (SLF1 with an S2 -haplotype) of Petunia inflata for co-immunoprecipitation (Co-IP) and mass spectrometry (MS), and identified PiCUL1-P (a pollen-specific Cullin1), PiSSK1 (a pollen-specific Skp1-like protein) and PiRBX1 (a conventional Rbx1) as components of the SCF(S) (2-) (SLF) (1) complex. Using pollen extracts containing PiSSK1:FLAG:GFP for Co-IP/MS, we identified two additional SLFs (SLF4 and SLF13) that were assembled into SCF(SLF) complexes. As 17 SLF genes (SLF1 to SLF17) have been identified in S2 and S3 pollen, here we examined whether all 17 SLFs are assembled into similar complexes and, if so, whether these complexes are unique to SLFs. We modified the previous Co-IP/MS procedure, including the addition of style extracts from four different S-genotypes to pollen extracts containing PiSSK1:FLAG:GFP, to perform four separate experiments. The results taken together show that all 17 SLFs and an SLF-like protein, SLFLike1 (encoded by an S-locus-linked gene), co-immunoprecipitated with PiSSK1:FLAG:GFP. Moreover, of the 179 other F-box proteins predicted by S2 and S3 pollen transcriptomes, only a pair with 94.9% identity and another pair with 99.7% identity co-immunoprecipitated with PiSSK1:FLAG:GFP. These results suggest that SCF(SLF) complexes have evolved specifically to function in self-incompatibility. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Expression, purification, crystallization and preliminary X-ray analysis of a C-terminal fragment of the Epstein–Barr virus ZEBRA protein

Energy Technology Data Exchange (ETDEWEB)

Morand, Patrice [European Molecular Biology Laboratory, Grenoble Outstation, BP 181, 38042 Grenoble CEDEX 9 (France); Laboratoire de Virologie Moléculaire et Structurale, EA 2939, Université Joseph Fourier, Grenoble (France); Budayova-Spano, Monika [European Molecular Biology Laboratory, Grenoble Outstation, BP 181, 38042 Grenoble CEDEX 9 (France); Perrissin, Monique [Laboratoire de Virologie Moléculaire et Structurale, EA 2939, Université Joseph Fourier, Grenoble (France); Müller, Christoph W., E-mail: mueller@embl-grenoble.fr; Petosa, Carlo [European Molecular Biology Laboratory, Grenoble Outstation, BP 181, 38042 Grenoble CEDEX 9 (France)

2006-03-01

A C-terminal fragment of the Epstein–Barr virus lytic switch protein ZEBRA has been crystallized in complex with DNA. A C-terminal fragment of the Epstein–Barr virus immediate-early transcription factor ZEBRA has been expressed as a recombinant protein in Escherichia coli and purified to homogeneity. The fragment behaves as a dimer in solution, consistent with the presence of a basic region leucine-zipper (bZIP) domain. Crystals of the fragment in complex with a DNA duplex were grown by the hanging-drop vapour-diffusion technique using polyethylene glycol 4000 and magnesium acetate as crystallization agents. Crystals diffract to better than 2.5 Å resolution using synchrotron radiation (λ = 0.976 Å). Crystals belong to space group C2, with unit-cell parameters a = 94.2, b = 26.5, c = 98.1 Å, β = 103.9°.

Roles of PU.1 in monocyte- and mast cell-specific gene regulation: PU.1 transactivates CIITA pIV in cooperation with IFN-gamma.

Science.gov (United States)

Ito, Tomonobu; Nishiyama, Chiharu; Nakano, Nobuhiro; Nishiyama, Makoto; Usui, Yoshihiko; Takeda, Kazuyoshi; Kanada, Shunsuke; Fukuyama, Kanako; Akiba, Hisaya; Tokura, Tomoko; Hara, Mutsuko; Tsuboi, Ryoji; Ogawa, Hideoki; Okumura, Ko

2009-07-01

Over-expression of PU.1, a myeloid- and lymphoid-specific transcription factor belonging to the Ets family, induces monocyte-specific gene expression in mast cells. However, the effects of PU.1 on each target gene and the involvement of cytokine signaling in PU.1-mediated gene expression are largely unknown. In the present study, PU.1 was over-expressed in two different types of bone marrow-derived cultured mast cells (BMMCs): BMMCs cultured with IL-3 plus stem cell factor (SCF) and BMMCs cultured with pokeweed mitogen-stimulated spleen-conditioned medium (PWM-SCM). PU.1 over-expression induced expression of MHC class II, CD11b, CD11c and F4/80 on PWM-SCM-cultured BMMCs, whereas IL-3/SCF-cultured BMMCs expressed CD11b and F4/80, but not MHC class II or CD11c. When IFN-gamma was added to the IL-3/SCF-based medium, PU.1 transfectant acquired MHC class II expression, which was abolished by antibody neutralization or in Ifngr(-/-) BMMCs, through the induction of expression of the MHC class II transactivator, CIITA. Real-time PCR detected CIITA mRNA driven by the fourth promoter, pIV, and chromatin immunoprecipitation indicated direct binding of PU.1 to pIV in PU.1-over-expressing BMMCs. PU.1-over-expressing cells showed a marked increase in IL-6 production in response to LPS stimulation in both IL-3/SCF and PWM-SCM cultures. These results suggest that PU.1 overproduction alone is sufficient for both expression of CD11b and F4/80 and for amplification of LPS-induced IL-6 production. However, IFN-gamma stimulation is essential for PU.1-mediated transactivation of CIITA pIV. Reduced expression of mast cell-related molecules and transcription factors GATA-1/2 and up-regulation of C/EBPalpha in PU.1 transfectants indicate that enforced PU.1 suppresses mast cell-specific gene expression through these transcription factors.

Repression of meiotic genes by antisense transcription and by Fkh2 transcription factor in Schizosaccharomyces pombe.

Science.gov (United States)

Chen, Huei-Mei; Rosebrock, Adam P; Khan, Sohail R; Futcher, Bruce; Leatherwood, Janet K

2012-01-01

In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s) of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the "unspliced" signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression.

Tautomerism, ionization, and bond dissociations of 5-nitro-2,4-dihydro-3H-1,2,4-triazolone.

NARCIS (Netherlands)

Harris, N.J.; Lammertsma, K.

1996-01-01

Tautomerization, ionization, and bond dissociations of the insensitive high-energy explosive 5-nitro-2,4-dihydro-3H-1,2,4-triazolone (NTO) were studied by molecular orbital SCF and MP2 theories and with the Becke3LYP hybrid density functional using the 6-31+G* and 6-311+G** basis sets. Energies

A nitrogen response pathway regulates virulence in plant pathogenic fungi: role of TOR and the bZIP protein MeaB.

Science.gov (United States)

López-Berges, Manuel S; Rispail, Nicolas; Prados-Rosales, Rafael C; Di Pietro, Antonio

2010-12-01

Virulence in plant pathogenic fungi is controlled through a variety of cellular pathways in response to the host environment. Nitrogen limitation has been proposed to act as a key signal to trigger the in planta expression of virulence genes. Moreover, a conserved Pathogenicity mitogen activated protein kinase (MAPK) cascade is strictly required for plant infection in a wide range of pathogens. We investigated the relationship between nitrogen signaling and the Pathogenicity MAPK cascade in controlling infectious growth of the vascular wilt fungus Fusarium oxysporum. Several MAPK-activated virulence functions such as invasive growth, vegetative hyphal fusion and host adhesion were strongly repressed in the presence of the preferred nitrogen source ammonium. Repression of these functions by ammonium was abolished by L-Methionine sulfoximine (MSX) or rapamycin, two specific inhibitors of Gln synthetase and the protein kinase TOR (Target Of Rapamycin), respectively, and was dependent on the bZIP protein MeaB. Supplying tomato plants with ammonium rather than nitrate resulted in a significant delay of vascular wilt symptoms caused by the F. oxysporum wild type strain, but not by the ΔmeaB mutant. Ammonium also repressed invasive growth in two other pathogens, the rice blast fungus Magnaporthe oryzae and the wheat head blight pathogen Fusarium graminearum. Our results suggest the presence of a conserved nitrogen-responsive pathway that operates via TOR and MeaB to control infectious growth in plant pathogenic fungi.

Insulin increases transcription of rat gene 33 through cis-acting elements in 5[prime]-flanking DNA

Energy Technology Data Exchange (ETDEWEB)

Cadilla, C.; Isham, K.R.; Lee, K.L.; Ch' ang, L.Y.; Kenney, F.T. (Oak Ridge National Lab., TN (United States)); Johnson, A.C. (National Cancer Institute, Bethesda, MD (United States). Lab. of Molecular Biology)

1992-01-01

Gene 33 is a multihormonally-regulated rat gene whose transcription is rapidly and markedly enhanced by insulin in liver and cultured hepatoma cells. To examine the mechanism by which insulin regulates transcription, the authors have constructed chimeric plasmids in which expression of the bacterial cat gene, encoding chloramphenicol acetyltransferase (CAT), is governed by gene 33 promoter elements and contiguous sequence in DNA flanking the transcription start point (tsp). When transfected into H4IIE hepatoma cells, these constructs gave rise to stably transformed cell lines producing the bacterial CAT enzyme. This expression was increased by insulin treatment in a fashion resembling the effect of this hormone on transcription of the native gene. In vitro transcription assays in nuclear extracts also revealed increased transcription of the chimeric plasmids when the extracts were prepared from insulin-treated rat hepatoma cells. The results demonstrate that induction by insulin is mediated by cis-acting nucleotide sequences located between bp [minus]480 to +27 relative to the tsp.

The mGluR5 antagonist AFQ056 does not affect methylation and transcription of the mutant FMR1 gene in vitro

Directory of Open Access Journals (Sweden)

Tabolacci Elisabetta

2012-03-01

Full Text Available Abstract Background Fragile X syndrome (FXS, the leading cause of inherited mental retardation, is due to expansion and methylation of a CGG sequence in the FMR1 gene, which result in its silencing and consequent absence of FMRP protein. This absence causes loss of repression of metabotropic glutamate receptor 5 (mGluR5-mediated pathways resulting in the behavioral and cognitive impairments associated with FXS. In a randomized, double-blind trial it was recently demonstrated a beneficial effect of AFQ056, a selective inhibitor of metabotrobic glutamate receptor type 5 (mGluR5, on fully methylated FXS patients respect to partially methylated FXS ones. Methods To determine whether AFQ056 may have secondary effects on the methylation and transcription of FMR1, here we treated three FXS lymphoblastoid cell lines and one normal control male line. A quantitative RT-PCR was performed to assess transcriptional reactivation of the FMR1 gene. To assess the methylation status of the FMR1 gene promoter it was carried out a bisulphite sequencing analysis. Results Both FMR1-mRNA levels and DNA methylation were unmodified with respect to untreated controls. Conclusions These results demonstrate that the AFQ056 effect on fully methylated FXS patients is not due to a secondary effect on DNA methylation and consequent transcriptional activation of FMR1.

A Herpesviral Immediate Early Protein Promotes Transcription Elongation of Viral Transcripts.

Science.gov (United States)

Fox, Hannah L; Dembowski, Jill A; DeLuca, Neal A

2017-06-13

Herpes simplex virus 1 (HSV-1) genes are transcribed by cellular RNA polymerase II (RNA Pol II). While four viral immediate early proteins (ICP4, ICP0, ICP27, and ICP22) function in some capacity in viral transcription, the mechanism by which ICP22 functions remains unclear. We observed that the FACT complex (comprised of SSRP1 and Spt16) was relocalized in infected cells as a function of ICP22. ICP22 was also required for the association of FACT and the transcription elongation factors SPT5 and SPT6 with viral genomes. We further demonstrated that the FACT complex interacts with ICP22 throughout infection. We therefore hypothesized that ICP22 recruits cellular transcription elongation factors to viral genomes for efficient transcription elongation of viral genes. We reevaluated the phenotype of an ICP22 mutant virus by determining the abundance of all viral mRNAs throughout infection by transcriptome sequencing (RNA-seq). The accumulation of almost all viral mRNAs late in infection was reduced compared to the wild type, regardless of kinetic class. Using chromatin immunoprecipitation sequencing (ChIP-seq), we mapped the location of RNA Pol II on viral genes and found that RNA Pol II levels on the bodies of viral genes were reduced in the ICP22 mutant compared to wild-type virus. In contrast, the association of RNA Pol II with transcription start sites in the mutant was not reduced. Taken together, our results indicate that ICP22 plays a role in recruiting elongation factors like the FACT complex to the HSV-1 genome to allow for efficient viral transcription elongation late in viral infection and ultimately infectious virion production. IMPORTANCE HSV-1 interacts with many cellular proteins throughout productive infection. Here, we demonstrate the interaction of a viral protein, ICP22, with a subset of cellular proteins known to be involved in transcription elongation. We determined that ICP22 is required to recruit the FACT complex and other transcription

Repression of Meiotic Genes by Antisense Transcription and by Fkh2 Transcription Factor in Schizosaccharomyces pombe

Science.gov (United States)

Chen, Huei-Mei; Rosebrock, Adam P.; Khan, Sohail R.; Futcher, Bruce; Leatherwood, Janet K.

2012-01-01

In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s) of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the “unspliced” signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression. PMID:22238674

Repression of meiotic genes by antisense transcription and by Fkh2 transcription factor in Schizosaccharomyces pombe.

Directory of Open Access Journals (Sweden)

Huei-Mei Chen

Full Text Available In S. pombe, about 5% of genes are meiosis-specific and accumulate little or no mRNA during vegetative growth. Here we use Affymetrix tiling arrays to characterize transcripts in vegetative and meiotic cells. In vegetative cells, many meiotic genes, especially those induced in mid-meiosis, have abundant antisense transcripts. Disruption of the antisense transcription of three of these mid-meiotic genes allowed vegetative sense transcription. These results suggest that antisense transcription represses sense transcription of meiotic genes in vegetative cells. Although the mechanism(s of antisense mediated transcription repression need to be further explored, our data indicates that RNAi machinery is not required for repression. Previously, we and others used non-strand specific methods to study splicing regulation of meiotic genes and concluded that 28 mid-meiotic genes are spliced only in meiosis. We now demonstrate that the "unspliced" signal in vegetative cells comes from the antisense RNA, not from unspliced sense RNA, and we argue against the idea that splicing regulates these mid-meiotic genes. Most of these mid-meiotic genes are induced in mid-meiosis by the forkhead transcription factor Mei4. Interestingly, deletion of a different forkhead transcription factor, Fkh2, allows low levels of sense expression of some mid-meiotic genes in vegetative cells. We propose that vegetative expression of mid-meiotic genes is repressed at least two independent ways: antisense transcription and Fkh2 repression.

Ubiquitin fusion constructs allow the expression and purification of multi-KOW domain complexes of the Saccharomyces cerevisiae transcription elongation factor Spt4/5.

Science.gov (United States)

Blythe, Amanda; Gunasekara, Sanjika; Walshe, James; Mackay, Joel P; Hartzog, Grant A; Vrielink, Alice

2014-08-01

Spt4/5 is a hetero-dimeric transcription elongation factor that can both inhibit and promote transcription elongation by RNA polymerase II (RNAPII). However, Spt4/5's mechanism of action remains elusive. Spt5 is an essential protein and the only universally-conserved RNAP-associated transcription elongation factor. The protein contains multiple Kyrpides, Ouzounis and Woese (KOW) domains. These domains, in other proteins, are thought to bind RNA although there is little direct evidence in the literature to support such a function in Spt5. This could be due, at least in part, to difficulties in expressing and purifying recombinant Spt5. When expressed in Escherichia coli (E. coli), Spt5 is innately insoluble. Here we report a new approach for the successful expression and purification of milligram quantities of three different multi-KOW domain complexes of Saccharomyces cerevisiae Spt4/5 for use in future functional studies. Using the E. coli strain Rosetta2 (DE3) we have developed strategies for co-expression of Spt4 and multi-KOW domain Spt5 complexes from the bi-cistronic pET-Duet vector. In a second strategy, Spt4/5 was expressed via co-transformation of Spt4 in the vector pET-M11 with Spt5 ubiquitin fusion constructs in the vector pHUE. We characterized the multi-KOW domain Spt4/5 complexes by Western blot, limited proteolysis, circular dichroism, SDS-PAGE and size exclusion chromatography-multiangle light scattering and found that the proteins are folded with a Spt4:Spt5 hetero-dimeric stoichiometry of 1:1. These expression constructs encompass a larger region of Spt5 than has previously been reported, and will provide the opportunity to elucidate the biological function of the multi-KOW containing Spt5. Copyright © 2014 Elsevier Inc. All rights reserved.

The N-Terminus of the Floral Arabidopsis TGA Transcription Factor PERIANTHIA Mediates Redox-Sensitive DNA-Binding.

Directory of Open Access Journals (Sweden)

Nora Gutsche

Full Text Available The Arabidopsis TGA transcription factor (TF PERIANTHIA (PAN regulates the formation of the floral organ primordia as revealed by the pan mutant forming an abnormal pentamerous arrangement of the outer three floral whorls. The Arabidopsis TGA bZIP TF family comprises 10 members, of which PAN and TGA9/10 control flower developmental processes and TGA1/2/5/6 participate in stress-responses. For the TGA1 protein it was shown that several cysteines can be redox-dependently modified. TGA proteins interact in the nucleus with land plant-specific glutaredoxins, which may alter their activities posttranslationally. Here, we investigated the DNA-binding of PAN to the AAGAAT motif under different redox-conditions. The AAGAAT motif is localized in the second intron of the floral homeotic regulator AGAMOUS (AG, which controls stamen and carpel development as well as floral determinacy. Whereas PAN protein binds to this regulatory cis-element under reducing conditions, the interaction is strongly reduced under oxidizing conditions in EMSA studies. The redox-sensitive DNA-binding is mediated via a special PAN N-terminus, which is not present in other Arabidopsis TGA TFs and comprises five cysteines. Two N-terminal PAN cysteines, Cys68 and Cys87, were shown to form a disulfide bridge and Cys340, localized in a C-terminal putative transactivation domain, can be S-glutathionylated. Comparative land plant analyses revealed that the AAGAAT motif exists in asterid and rosid plant species. TGA TFs with N-terminal extensions of variable length were identified in all analyzed seed plants. However, a PAN-like N-terminus exists only in the rosids and exclusively Brassicaceae homologs comprise four to five of the PAN N-terminal cysteines. Redox-dependent modifications of TGA cysteines are known to regulate the activity of stress-related TGA TFs. Here, we show that the N-terminal PAN cysteines participate in a redox-dependent control of the PAN interaction with a highly

Multiplex Reverse Transcription-Polymerase Chain Reaction untuk Deteksi Cepat Virus Flu Burung H5N1 (MULTIPLEX REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION FOR RAPID DETECTION OF H5N1 AVIAN INFLUENZA VIRUS

Directory of Open Access Journals (Sweden)

Raden Wasito

2015-05-01

Full Text Available Avian influenza virus subtype H5N1 (AIV H5N1 is highly pathogenic and fatal in poultry. The virusis still endemic with low virulence rate, although it may play a critical role in causing high morbidity andmortality rates in poultry in Indonesia. In general, diagnostic approach for AIV H5N1 is based onconventional serological and viral isolation methods that have the potential to produce consumings oftime and relatively expensive cost within the laboratory without compromising test utility. Thus, amolecular approach of multiplex reverse transcription-polymerase chain reaction (mRT-PCR was developedand applied for the detection of matrix gene type A influenza viruses, AIV subtype subtype H5hemagglutinin gene with simultaneous detection of N1 nucleoprotein gene. Thirty sera specimens fromthe diseased commercial chickens that were specifically amplified positive-RT-PCR for AIV H5N1 wereselected for mRT-PCR. The mRT-PCR products were visualized by agarose gel electrophoresis and consistedof DNA fragments of AIV of 245 bp, 545 bp and 343 bp for M, H5 and N1 genes, respectively. Thus, themRT-PCR that can rapidly differentiate simultaneously between these genes is very important for thecontrol and even eradication of AIV transmission in poultry in Indonesia.

Trans-activation of the 5' to 3' viral DNA strand transfer by nucleocapsid protein during reverse transcription of HIV1 RNA.

Science.gov (United States)

Darlix, J L; Vincent, A; Gabus, C; de Rocquigny, H; Roques, B

1993-08-01

Two DNA strand transfer reactions take place during reverse transcription of the retroviral genome. The first transfer, that of the minus-strand strong stop DNA from the 5' end of the viral RNA to the 3' end, has been studied in vitro with two RNAs mimicking the 5' and 3' regions of the HIV1 genome and with nucleocapsid protein, NCp7, and reverse transcriptase. The results show that NCp7 strongly activates the 5' to 3' DNA strand transfer during reverse transcription while a basic peptide resembling NCp7 is inactive. Activation of the first transfer by several NCp7 derived peptides and the influence of the terminal redundancies (R) present at the 5' and 3' ends of HIV1 RNA were also examined. The first transfer is optimal in the presence of intact NCp7 and necessitates R on both the 5' and 3' RNAs. Sequencing of full length viral DNA products reveals approximately 40% misincorporations at the first nucleotide beyond the transfer point. If such base misincorporations occur during proviral DNA synthesis with possible homologous recombinations it may well contribute to the high level of genetic variability of HIV.

Virulence factor NSs of rift valley fever virus recruits the F-box protein FBXO3 to degrade subunit p62 of general transcription factor TFIIH.

Science.gov (United States)

Kainulainen, Markus; Habjan, Matthias; Hubel, Philipp; Busch, Laura; Lau, Simone; Colinge, Jacques; Superti-Furga, Giulio; Pichlmair, Andreas; Weber, Friedemann

2014-03-01

The nonstructural protein NSs is the main virulence factor of Rift Valley fever virus (RVFV; family Bunyaviridae, genus Phlebovirus), a serious pathogen of livestock and humans in Africa. RVFV NSs blocks transcriptional upregulation of antiviral type I interferons (IFN) and destroys the general transcription factor TFIIH subunit p62 via the ubiquitin/proteasome pathway. Here, we identified a subunit of E3 ubiquitin ligases, F-box protein FBXO3, as a host cell interactor of NSs. Small interfering RNA (siRNA)-mediated depletion of FBXO3 rescued p62 protein levels in RVFV-infected cells and elevated IFN transcription by 1 order of magnitude. NSs interacts with the full-length FBXO3 protein as well as with a truncated isoform that lacks the C-terminal acidic and poly(R)-rich domains. These isoforms are present in both the nucleus and the cytoplasm. NSs exclusively removes the nuclear pool of full-length FBXO3, likely due to consumption during the degradation process. F-box proteins form the variable substrate recognition subunit of the so-called SCF ubiquitin ligases, which also contain the constant components Skp1, cullin 1 (or cullin 7), and Rbx1. siRNA knockdown of Skp1 also protected p62 from degradation, suggesting involvement in NSs action. However, knockdown of cullin 1, cullin 7, or Rbx1 could not rescue p62 degradation by NSs. Our data show that the enzymatic removal of p62 via the host cell factor FBXO3 is a major mechanism of IFN suppression by RVFV. Rift Valley fever virus is a serious emerging pathogen of animals and humans. Its main virulence factor, NSs, enables unhindered virus replication by suppressing the antiviral innate immune system. We identified the E3 ubiquitin ligase FBXO3 as a novel host cell interactor of NSs. NSs recruits FBXO3 to destroy the general host cell transcription factor TFIIH-p62, resulting in suppression of the transcriptional upregulation of innate immunity.

A Nitrogen Response Pathway Regulates Virulence Functions in Fusarium oxysporum via the Protein Kinase TOR and the bZIP Protein MeaB[C][W

Science.gov (United States)

López-Berges, Manuel S.; Rispail, Nicolas; Prados-Rosales, Rafael C.; Di Pietro, Antonio

2010-01-01

During infection, fungal pathogens activate virulence mechanisms, such as host adhesion, penetration and invasive growth. In the vascular wilt fungus Fusarium oxysporum, the mitogen-activated protein kinase Fmk1 is required for plant infection and controls processes such as cellophane penetration, vegetative hyphal fusion, or root adhesion. Here, we show that these virulence-related functions are repressed by the preferred nitrogen source ammonium and restored by treatment with l-methionine sulfoximine or rapamycin, two specific inhibitors of Gln synthetase and the protein kinase TOR, respectively. Deletion of the bZIP protein MeaB also resulted in nitrogen source–independent activation of virulence mechanisms. Activation of these functions did not require the global nitrogen regulator AreA, suggesting that MeaB-mediated repression of virulence functions does not act through inhibition of AreA. Tomato plants (Solanum lycopersicum) supplied with ammonium rather than nitrate showed a significant reduction in vascular wilt symptoms when infected with the wild type but not with the ΔmeaB strain. Nitrogen source also affected invasive growth in the rice blast fungus Magnaporthe oryzae and the wheat head blight pathogen Fusarium graminearum. We propose that a conserved nitrogen-responsive pathway might operate via TOR and MeaB to control virulence in plant pathogenic fungi. PMID:20639450

A Herpesviral Immediate Early Protein Promotes Transcription Elongation of Viral Transcripts

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Hannah L. Fox

2017-06-01

Full Text Available Herpes simplex virus 1 (HSV-1 genes are transcribed by cellular RNA polymerase II (RNA Pol II. While four viral immediate early proteins (ICP4, ICP0, ICP27, and ICP22 function in some capacity in viral transcription, the mechanism by which ICP22 functions remains unclear. We observed that the FACT complex (comprised of SSRP1 and Spt16 was relocalized in infected cells as a function of ICP22. ICP22 was also required for the association of FACT and the transcription elongation factors SPT5 and SPT6 with viral genomes. We further demonstrated that the FACT complex interacts with ICP22 throughout infection. We therefore hypothesized that ICP22 recruits cellular transcription elongation factors to viral genomes for efficient transcription elongation of viral genes. We reevaluated the phenotype of an ICP22 mutant virus by determining the abundance of all viral mRNAs throughout infection by transcriptome sequencing (RNA-seq. The accumulation of almost all viral mRNAs late in infection was reduced compared to the wild type, regardless of kinetic class. Using chromatin immunoprecipitation sequencing (ChIP-seq, we mapped the location of RNA Pol II on viral genes and found that RNA Pol II levels on the bodies of viral genes were reduced in the ICP22 mutant compared to wild-type virus. In contrast, the association of RNA Pol II with transcription start sites in the mutant was not reduced. Taken together, our results indicate that ICP22 plays a role in recruiting elongation factors like the FACT complex to the HSV-1 genome to allow for efficient viral transcription elongation late in viral infection and ultimately infectious virion production.

On the electronic structure of 5g1 complexes of element 125: a quasi - relativistic MS-Xα study

International Nuclear Information System (INIS)

Makhyoun, M.A.

1988-01-01

Quasi-relativistic SCF Xα calculations are reported for the hypothetical complexes E0 2 2+ , EF 6 (E = element 125) and the 5f 1 ion Np0 2 2 + . The calculations indicate that the E complexes have a 5g 1 outer electronic configuration with good agreement with previous predictions. The ligand field energy diagram for G 1 system in 0 h symmetry in discussed in relation to the obtained X α results

Interferon-γ Promotes Inflammation and Development of T-Cell Lymphoma in HTLV-1 bZIP Factor Transgenic Mice.

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Yu Mitagami

2015-08-01

Full Text Available Human T-cell leukemia virus type 1 (HTLV-1 is an etiological agent of several inflammatory diseases and a T-cell malignancy, adult T-cell leukemia (ATL. HTLV-1 bZIP factor (HBZ is the only viral gene that is constitutively expressed in HTLV-1-infected cells, and it has multiple functions on T-cell signaling pathways. HBZ has important roles in HTLV-1-mediated pathogenesis, since HBZ transgenic (HBZ-Tg mice develop systemic inflammation and T-cell lymphomas, which are similar phenotypes to HTLV-1-associated diseases. We showed previously that in HBZ-Tg mice, HBZ causes unstable Foxp3 expression, leading to an increase in regulatory T cells (Tregs and the consequent induction of IFN-γ-producing cells, which in turn leads to the development of inflammation in the mice. In this study, we show that the severity of inflammation is correlated with the development of lymphomas in HBZ-Tg mice, suggesting that HBZ-mediated inflammation is closely linked to oncogenesis in CD4+ T cells. In addition, we found that IFN-γ-producing cells enhance HBZ-mediated inflammation, since knocking out IFN-γ significantly reduced the incidence of dermatitis as well as lymphoma. Recent studies show the critical roles of the intestinal microbiota in the development of Tregs in vivo. We found that even germ-free HBZ-Tg mice still had an increased number of Tregs and IFN-γ-producing cells, and developed dermatitis, indicating that an intrinsic activity of HBZ evokes aberrant T-cell differentiation and consequently causes inflammation. These results show that immunomodulation by HBZ is implicated in both inflammation and oncogenesis, and suggest a causal connection between HTLV-1-associated inflammation and ATL.

Relação da expressão de fatores de crescimento celular (IGF-1 e (SCF com fatores prognósticos e o alvo da rapamicina em mamíferos (m-TOR em mastocitomas cutâneos caninos

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Raquel B. Ferioli

2013-04-01

Full Text Available O mastocitoma cutâneo (MTC é a neoplasia maligna mais comum na pele dos cães e seu comportamento biológico é muito variável. Dentre os fatores prognósticos estudados nos MTCs, a classificação histopatológica, o índice proliferativo e o padrão de expressão doc-KIT são os que apresentam uma associação mais relevante com o provável prognóstico deste tumor. O objetivo deste trabalho foi avaliar a expressão proteica de fator de crescimento semelhante à insulina tipo 1 (IGF-1, fator de célula tronco (SCF e sua relação com o receptor tirosina quinase (c-KIT, alvo da rapamicina em mamíferos (m-TOR, grau histológico, índice proliferativo pelo KI-67e o número de figuras de mitose (IM com dados clínicos de cães com MTCs . Foram utilizadas 133 amostras de MTCs, provenientes de 133 cães, dispostas em lâminas de microarranjo de tecidos (TMA. A técnica de imuno-histoquímica foi utilizada para a avaliação destas proteínas. Observou-se associação entre SCF e, a graduação histopatológica proposta em 2011, índice mitótico, proliferação celular (KI-67, escore de IGF-1, local da lesão, idade dos animais e padrão imuno-histoquímico do receptor c-KIT. A relação de dependência também foi observada entre IGF-1 e o porte dos animais, IM, m-TOR e c-KIT. A expressão de SCF teve relacção com a agressividade dos MTCs caninos, uma vez que foi mais freqüente em MTCs com c-KIT citoplasmático. A relação entre a expressão de IGF-1, SCF, c-KIT e m-TOR pode estar associada à integralização de suas vias de ação. A expressão de IGF-1 está associada à MTCs em cães de porte grande.

Dynamical behavior of psb gene transcripts in greening wheat seedlings. I. Time course of accumulation of the pshA through psbN gene transcripts during light-induced greening.

Science.gov (United States)

Kawaguchi, H; Fukuda, I; Shiina, T; Toyoshima, Y

1992-11-01

The time course of the accumulation of the transcripts from 13 psb genes encoding a major part of the proteins composing photosystem II during light-induced greening of dark-grown wheat seedlings was examined focusing on early stages of plastid development (0.5 h through 72 h). The 13 genes can be divided into three groups. (1) The psbA gene is transcribed as a single transcript of 1.3 kb in the dark-grown seedlings, but its level increases 5- to 7-fold in response to light due to selective increase in RNA stability as well as in transcription activity. (2) The psbE-F-L-J operon, psbM and psbN genes are transcribed as a single transcript of 1.1 kb, two transcripts of 0.5 and 0.7 kb and a single transcript of 0.3 kb, respectively, in the dark-grown seedlings. The levels of accumulation of every transcript remain unchanged or rather decrease during plastid development under illumination. (3) The psbK-I-D-C gene cluster and psbB-H operon exhibit fairly complicated northern hybridization patterns during the greening process. When a psbC or psbD gene probe was used for northern hybridization, five transcripts differing in length were detected in the etioplasts from 5-day old dark-grown seedlings. After 2 h illumination, two new transcripts of different length appeared. Light induction of new transcripts was also observed in the psbB-H operon.

Arabidopsis MADS-Box Transcription Factor AGL21 Acts as Environmental Surveillance of Seed Germination by Regulating ABI5 Expression.

Science.gov (United States)

Yu, Lin-Hui; Wu, Jie; Zhang, Zi-Sheng; Miao, Zi-Qing; Zhao, Ping-Xia; Wang, Zhen; Xiang, Cheng-Bin

2017-06-05

Seed germination is a crucial checkpoint for plant survival under unfavorable environmental conditions. Abscisic acid (ABA) signaling plays a vital role in integrating environmental information to regulate seed germination. It has been well known that MCM1/AGAMOUS/DEFICIENS/SRF (MADS)-box transcription factors are key regulators of seed and flower development in Arabidopsis. However, little is known about their functions in seed germination. Here we report that MADS-box transcription factor AGL21 is a negative regulator of seed germination and post-germination growth by controlling the expression of ABA-INSENSITIVE 5 (ABI5) in Arabidopsis. The AGL21-overexpressing plants were hypersensitive to ABA, salt, and osmotic stresses during seed germination and early post-germination growth, whereas agl21 mutants were less sensitive. We found that AGL21 positively regulated ABI5 expression in seeds. Consistently, genetic analyses showed that AGL21 is epistatic to ABI5 in controlling seed germination. Chromatin immunoprecipitation assays further demonstrated that AGL21 could directly bind to the ABI5 promoter in plant cells. Moreover, we found that AGL21 responded to multiple environmental stresses and plant hormones during seed germination. Taken together, our results suggest that AGL21 acts as a surveillance integrator that incorporates environmental cues and endogenous hormonal signals into ABA signaling to regulate seed germination and early post-germination growth. Copyright © 2017 The Author. Published by Elsevier Inc. All rights reserved.

Light-dependent changes in psbD and psbC transcripts of barley chloroplasts: accumulation of two transcripts maintains psbD and psbC translation capability in mature chloroplasts.

OpenAIRE

Gamble, P E; Sexton, T B; Mullet, J E

1988-01-01

The psbD and psbC genes encode two polypeptides of Photosystem II. These genes are adjacent in the barley chloroplast genome and are part of a 5.7 kbp transcription unit. In dark-grown barley, four large transcripts hybridize to psbD and psbC; two additional transcripts hybridize to psbC. Illumination of 4.5-day-old dark-grown seedlings causes a decrease in the six psbD--psbC transcripts found in etioplasts and the accumulation of two different transcripts of 4.0 and 3.2 kb which hybridize to...

Molecular characterization of 5S ribosomal RNA genes and transcripts in the protozoan parasite Leishmania major.

Science.gov (United States)

Moreno-Campos, Rodrigo; Florencio-Martínez, Luis E; Nepomuceno-Mejía, Tomás; Rojas-Sánchez, Saúl; Vélez-Ramírez, Daniel E; Padilla-Mejía, Norma E; Figueroa-Angulo, Elisa; Manning-Cela, Rebeca; Martínez-Calvillo, Santiago

2016-12-01

Eukaryotic 5S rRNA, synthesized by RNA polymerase III (Pol III), is an essential component of the large ribosomal subunit. Most organisms contain hundreds of 5S rRNA genes organized into tandem arrays. However, the genome of the protozoan parasite Leishmania major contains only 11 copies of the 5S rRNA gene, which are interspersed and associated with other Pol III-transcribed genes. Here we report that, in general, the number and order of the 5S rRNA genes is conserved between different species of Leishmania. While in most organisms 5S rRNA genes are normally associated with the nucleolus, combined fluorescent in situ hybridization and indirect immunofluorescence experiments showed that 5S rRNA genes are mainly located at the nuclear periphery in L. major. Similarly, the tandemly repeated 5S rRNA genes in Trypanosoma cruzi are dispersed throughout the nucleus. In contrast, 5S rRNA transcripts in L. major were localized within the nucleolus, and scattered throughout the cytoplasm, where mature ribosomes are located. Unlike other rRNA species, stable antisense RNA complementary to 5S rRNA is not detected in L. major.

Overlapping ETS and CRE Motifs (G/CCGGAAGTGACGTCA) Preferentially Bound by GABPα and CREB Proteins

Science.gov (United States)

Chatterjee, Raghunath; Zhao, Jianfei; He, Ximiao; Shlyakhtenko, Andrey; Mann, Ishminder; Waterfall, Joshua J.; Meltzer, Paul; Sathyanarayana, B. K.; FitzGerald, Peter C.; Vinson, Charles

2012-01-01

Previously, we identified 8-bps long DNA sequences (8-mers) that localize in human proximal promoters and grouped them into known transcription factor binding sites (TFBS). We now examine split 8-mers consisting of two 4-mers separated by 1-bp to 30-bps (X4-N1-30-X4) to identify pairs of TFBS that localize in proximal promoters at a precise distance. These include two overlapping TFBS: the ETS⇔ETS motif (C/GCCGGAAGCGGAA) and the ETS⇔CRE motif (C/GCGGAAGTGACGTCAC). The nucleotides in bold are part of both TFBS. Molecular modeling shows that the ETS⇔CRE motif can be bound simultaneously by both the ETS and the B-ZIP domains without protein-protein clashes. The electrophoretic mobility shift assay (EMSA) shows that the ETS protein GABPα and the B-ZIP protein CREB preferentially bind to the ETS⇔CRE motif only when the two TFBS overlap precisely. In contrast, the ETS domain of ETV5 and CREB interfere with each other for binding the ETS⇔CRE. The 11-mer (CGGAAGTGACG), the conserved part of the ETS⇔CRE motif, occurs 226 times in the human genome and 83% are in known regulatory regions. In vivo GABPα and CREB ChIP-seq peaks identified the ETS⇔CRE as the most enriched motif occurring in promoters of genes involved in mRNA processing, cellular catabolic processes, and stress response, suggesting that a specific class of genes is regulated by this composite motif. PMID:23050235

Cooperation between RUNX1-ETO9a and novel transcriptional partner KLF6 in upregulation of Alox5 in acute myeloid leukemia.

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Russell C DeKelver

Full Text Available Fusion protein RUNX1-ETO (AML1-ETO, RUNX1-RUNX1T1 is expressed as the result of the 8q22;21q22 translocation [t(8;21], which is one of the most common chromosomal abnormalities found in acute myeloid leukemia. RUNX1-ETO is thought to promote leukemia development through the aberrant regulation of RUNX1 (AML1 target genes. Repression of these genes occurs via the recruitment of the corepressors N-COR and SMRT due to their interaction with ETO. Mechanisms of RUNX1-ETO target gene upregulation remain less well understood. Here we show that RUNX1-ETO9a, the leukemogenic alternatively spliced transcript expressed from t(8;21, upregulates target gene Alox5, which is a gene critically required for the promotion of chronic myeloid leukemia development by BCR-ABL. Loss of Alox5 expression reduces activity of RUNX1-ETO9a, MLL-AF9 and PML-RARα in vitro. However, Alox5 is not essential for the induction of leukemia by RUNX1-ETO9a in vivo. Finally, we demonstrate that the upregulation of Alox5 by RUNX1-ETO9a occurs via the C₂H₂ zinc finger transcription factor KLF6, a protein required for early hematopoiesis and yolk sac development. Furthermore, KLF6 is specifically upregulated by RUNX1-ETO in human leukemia cells. This identifies KLF6 as a novel mediator of t(8;21 target gene regulation, providing a new mechanism for RUNX1-ETO transcriptional control.

Assembly of proteins and 5 S rRNA to transcripts of the major structural domains of 23 S rRNA

DEFF Research Database (Denmark)

Ostergaard, P; Phan, H; Johansen, L B

1998-01-01

The six major structural domains of 23 S rRNA from Escherichia coli, and all combinations thereof, were synthesized as separate T7 transcripts and reconstituted with total 50 S subunit proteins. Analysis by one and two-dimensional gel electrophoresis demonstrated the presence of at least one prim...... approach was used to map the putative binding regions on domain V of protein L9 and the 5 S RNA-L5-L18 complex....

Structural, functional and evolutionary characterization of major drought transcription factors families in maize

Science.gov (United States)

Mittal, Shikha; Banduni, Pooja; Mallikarjuna, Mallana G.; Rao, Atmakuri R.; Jain, Prashant A.; Dash, Prasanta K.; Thirunavukkarasu, Nepolean

2018-05-01

Drought is one of the major threats to maize production. In order to improve the production and to breed tolerant hybrids, understanding the genes and regulatory mechanisms during drought stress is important. Transcription factors (TFs) play a major role in gene regulation and many TFs have been identified in response to drought stress. In our experiment, a set of 15 major TF families comprising 1436 genes was structurally and functionally characterized using in-silico tools and a gene expression assay. All 1436 genes were mapped on 10 chromosome of maize. The functional annotation indicated the involvement of these genes in ABA signaling, ROS scavenging, photosynthesis, stomatal regulation, and sucrose metabolism. Duplication was identified as the primary force in divergence and expansion of TF families. Phylogenetic relationship was developed individually for each TF family as well as combined TF families. Phylogenetic analysis grouped the TF family of genes into TF-specific and mixed groups. Phylogenetic analysis of genes belonging to various TF families suggested that the origin of TFs occurred in the lineage of maize evolution. Gene structure analysis revealed that more number of genes were intron-rich as compared to intronless genes. Drought-responsive CRE’s such as ABREA, ABREB, DRE1 and DRECRTCOREAT have been identified. Expression and interaction analyses identified leaf-specific bZIP TF, GRMZM2G140355, as a potential contributor toward drought tolerance in maize. We also analyzed protein-protein interaction network of 269 drought-responsive genes belonging to different drought-related TFs. The information generated on structural and functional characteristics, expression and interaction of the drought-related TF families will be useful to decipher the drought tolerance mechanisms and to derive drought-tolerant genotypes in maize.

Xenobiotic metabolism in the fourth dimension: PARtners in time.

Science.gov (United States)

Green, Carla B; Takahashi, Joseph S

2006-07-01

A significant portion of the transcriptome in mammals, including the PAR bZIP transcription factors DBP, HLF, and TEF, is under circadian clock control. In this issue of Cell Metabolism, Gachon and colleagues (Gachon et al., 2006) show that disruption of these three genes in mice alters gene expression patterns of many proteins involved in drug metabolism and in liver and kidney responses to xenobiotic agents. Triple mutant mice have severe physiological deficits, including increased hypersensitivity to xenobiotic agents and premature aging, highlighting the profound effect the circadian clock has on this important response system.

Luminescent and scintillation properties of Sc3+and La3+doped Y2SiO5 powders and single crystalline films

International Nuclear Information System (INIS)

Zorenko, Yu.; Gorbenko, V.; Zorenko, T.; Voznyak, T.; Voloshynovskii, A.; Vistovskiy, V.; Paprocki, K.; Mosińska, L.; Bilski, P.; Twardak, A.; Fedorov, A.; Nikl, M.; Mares, J.A.

2016-01-01

The paper is dedicated to the investigation of the luminescence of Sc 3+ and La 3+ isoelectronic impurities in Y 2 SiO 5 (YSO) single crystalline films (SCF), grown by the liquid phase epitaxy (LPE) method, and in the powder analogs of these compounds prepared using the ceramic technology. The Sc 3+ and La 3+ dopants replacing the Y 3+ cations in Y1and Y2 positions of YSO host introduce the strong complex emission bands in the UV range peaked at 330 and 345 nm, respectively. The Sc 3+ and La 3+ dopant in YSO matrix yields also the strong TSL peaks at 400 and 405 K related to the ScY and ScLa centers formation, respectively. The luminescence and scintillation properties of YSO SCFs doped with Sc 3+ and La 3+ ions and co-doped with Ce 3+ ions on the trace impurity level have been also studied in our work. We have found that the light yield (LY) of these YSO:Sc and YSO:La SCFs can reach 50–65% of LY in reference YSO:Ce SCF due to strong quenching influence of Pb 2+ ions. Finally, the potential of Sc 3+ and La 3+ doped SCF of orthosilicates for creation of heavy scintillation screens, emitting in the UV range, is discussed.

The Pseudomonas transcriptional regulator AlgR controls LipA expression via the noncoding RNA RsmZ in Pseudomonas protegens Pf-5.

Science.gov (United States)

Li, Menggang; Yan, Jinyong; Yan, Yunjun

2017-05-20

Pseudomonas lipases are well studied enzymes. However, few studies have been conducted to explore the mechanism underlying the regulation of lipases expression. AlgR, a global regulator, controls the expression of multiple genes, regulates bacterial peristalsis, and participates in the regulation of quorum-sensing (QS) system, and so on. In this study, the effect of AlgR on lipase expression was investigated by knocking out the algR and rsmZ genes or overexpressing them. It is found out that AlgR can regulate the expression of lipA at both transcriptional and translational levels, but the transcriptional level was dominant. AlgR is also able to regulate the expression of rsmX/rsmY/rsmZ. Additionally, using algR/rsmZ double gene knock-out, it showed that AlgR could directly bind to the promoter sequence of rsmZ to regulate lipA activity. In conclusion, this study for the first time indicates that AlgR directly binds to rsmZ to regulates the expression of lipA via regulating transcription of rsmZ, and mainly regulates the expression of lipA at transcriptional level in P. protegens Pf-5. Copyright © 2017 Elsevier Inc. All rights reserved.

The ETS-5 transcription factor regulates activity states in Caenorhabditis elegans by controlling satiety

DEFF Research Database (Denmark)

Juozaityte, Vaida; Pladevall-Morera, David; Podolska, Agnieszka

2017-01-01

Animal behavior is shaped through interplay among genes, the environment, and previous experience. As in mammals, satiety signals induce quiescence in Caenorhabditis elegans Here we report that the C. elegans transcription factor ETS-5, an ortholog of mammalian FEV/Pet1, controls satiety......-induced quiescence. Nutritional status has a major influence on C. elegans behavior. When foraging, food availability controls behavioral state switching between active (roaming) and sedentary (dwelling) states; however, when provided with high-quality food, C. elegans become sated and enter quiescence. We show......-regulated behavioral state switching. Taken together, our results identify a neuronal mechanism for controlling intestinal fat stores and organismal behavioral states in C. elegans, and establish a paradigm for the elucidation of obesity-relevant mechanisms....

The transcriptional landscape

DEFF Research Database (Denmark)

Nielsen, Henrik

2011-01-01

The application of new and less biased methods to study the transcriptional output from genomes, such as tiling arrays and deep sequencing, has revealed that most of the genome is transcribed and that there is substantial overlap of transcripts derived from the two strands of DNA. In protein coding...... regions, the map of transcripts is very complex due to small transcripts from the flanking ends of the transcription unit, the use of multiple start and stop sites for the main transcript, production of multiple functional RNA molecules from the same primary transcript, and RNA molecules made...... by independent transcription from within the unit. In genomic regions separating those that encode proteins or highly abundant RNA molecules with known function, transcripts are generally of low abundance and short-lived. In most of these cases, it is unclear to what extent a function is related to transcription...

The putative bZIP transcription factor BzpN slows proliferation and functions in the regulation of cell density by autocrine signals in Dictyostelium.

Directory of Open Access Journals (Sweden)

Jonathan E Phillips

Full Text Available The secreted proteins AprA and CfaD function as autocrine signals that inhibit cell proliferation in Dictyostelium discoideum, thereby regulating cell numbers by a negative feedback mechanism. We report here that the putative basic leucine zipper transcription factor BzpN plays a role in the inhibition of proliferation by AprA and CfaD. Cells lacking BzpN proliferate more rapidly than wild-type cells but do not reach a higher stationary density. Recombinant AprA inhibits wild-type cell proliferation but does not inhibit the proliferation of cells lacking BzpN. Recombinant CfaD also inhibits wild-type cell proliferation, but promotes the proliferation of cells lacking BzpN. Overexpression of BzpN results in a reduced cell density at stationary phase, and this phenotype requires AprA, CfaD, and the kinase QkgA. Conditioned media from high-density cells stops the proliferation of wild-type but not bzpN(- cells and induces a nuclear localization of a BzpN-GFP fusion protein, though this localization does not require AprA or CfaD. Together, the data suggest that BzpN is necessary for some but not all of the effects of AprA and CfaD, and that BzpN may function downstream of AprA and CfaD in a signal transduction pathway that inhibits proliferation.

The putative bZIP transcription factor BzpN slows proliferation and functions in the regulation of cell density by autocrine signals in Dictyostelium.

Science.gov (United States)

Phillips, Jonathan E; Huang, Eryong; Shaulsky, Gad; Gomer, Richard H

2011-01-01

The secreted proteins AprA and CfaD function as autocrine signals that inhibit cell proliferation in Dictyostelium discoideum, thereby regulating cell numbers by a negative feedback mechanism. We report here that the putative basic leucine zipper transcription factor BzpN plays a role in the inhibition of proliferation by AprA and CfaD. Cells lacking BzpN proliferate more rapidly than wild-type cells but do not reach a higher stationary density. Recombinant AprA inhibits wild-type cell proliferation but does not inhibit the proliferation of cells lacking BzpN. Recombinant CfaD also inhibits wild-type cell proliferation, but promotes the proliferation of cells lacking BzpN. Overexpression of BzpN results in a reduced cell density at stationary phase, and this phenotype requires AprA, CfaD, and the kinase QkgA. Conditioned media from high-density cells stops the proliferation of wild-type but not bzpN(-) cells and induces a nuclear localization of a BzpN-GFP fusion protein, though this localization does not require AprA or CfaD. Together, the data suggest that BzpN is necessary for some but not all of the effects of AprA and CfaD, and that BzpN may function downstream of AprA and CfaD in a signal transduction pathway that inhibits proliferation.

Dicty_cDB: CHN523 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available -09 5 CV164410 |CV164410.1 rsmsxl_003361.y1.scf cDNA Library of Salvia miltiorrhiza Salvia miltiorrhiza cDNA... 5', mRNA sequence. 54 5e-09 3 CV172465 |CV172465.1 rsmsxlre_008717.z1.scf cDNA L

Overexpression of GmERF5, a new member of the soybean EAR motif-containing ERF transcription factor, enhances resistance to Phytophthora sojae in soybean.

Science.gov (United States)

Dong, Lidong; Cheng, Yingxin; Wu, Junjiang; Cheng, Qun; Li, Wenbin; Fan, Sujie; Jiang, Liangyu; Xu, Zhaolong; Kong, Fanjiang; Zhang, Dayong; Xu, Pengfei; Zhang, Shuzhen

2015-05-01

Phytophthora root and stem rot of soybean [Glycine max (L.) Merr.], caused by Phytophthora sojae Kaufmann and Gerdemann, is a destructive disease throughout the soybean planting regions in the world. Here, we report insights into the function and underlying mechanisms of a novel ethylene response factor (ERF) in soybean, namely GmERF5, in host responses to P. sojae. GmERF5-overexpressing transgenic soybean exhibited significantly enhanced resistance to P. sojae and positively regulated the expression of the PR10, PR1-1, and PR10-1 genes. Sequence analysis suggested that GmERF5 contains an AP2/ERF domain of 58 aa and a conserved ERF-associated amphiphilic repression (EAR) motif in its C-terminal region. Following stress treatments, GmERF5 was significantly induced by P. sojae, ethylene (ET), abscisic acid (ABA), and salicylic acid (SA). The activity of the GmERF5 promoter (GmERF5P) was upregulated in tobacco leaves with ET, ABA, Phytophthora nicotianae, salt, and drought treatments, suggesting that GmERF5 could be involved not only in the induced defence response but also in the ABA-mediated pathway of salt and drought tolerance. GmERF5 could bind to the GCC-box element and act as a repressor of gene transcription. It was targeted to the nucleus when transiently expressed in Arabidopsis protoplasts. GmERF5 interacted with a basic helix-loop-helix transcription factor (GmbHLH) and eukaryotic translation initiation factor (GmEIF) both in yeast cells and in planta. To the best of our knowledge, GmERF5 is the first soybean EAR motif-containing ERF transcription factor demonstrated to be involved in the response to pathogen infection. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

mRNA Transcript Diversity Creates New Opportunities for Pharmacological Intervention

OpenAIRE

Barrie, Elizabeth S.; Smith, Ryan M.; Sanford, Jonathan C.; Sadee, Wolfgang

2012-01-01

Most protein coding genes generate multiple RNA transcripts through alternative splicing, variable 3′ and 5′UTRs, and RNA editing. Although drug design typically targets the main transcript, alternative transcripts can have profound physiological effects, encoding proteins with distinct functions or regulatory properties. Formation of these alternative transcripts is tissue-selective and context-dependent, creating opportunities for more effective and targeted therapies with reduced adverse e...

Pregnane and Xenobiotic Receptor gene expression in liver cells is modulated by Ets-1 in synchrony with transcription factors Pax5, LEF-1 and c-jun

Energy Technology Data Exchange (ETDEWEB)

Kumari, Sangeeta; Saradhi, Mallampati; Rana, Manjul; Chatterjee, Swagata [Special Centre for Molecular Medicine, Jawaharlal Nehru University, New Delhi 110067 (India); Aumercier, Marc [IRI, CNRS USR 3078, Université de Lille-Nord de France, Parc CNRS de la Haute Borne, 50 Avenue de Halley, BP 70478, 59658 Villeneuve d’Ascq Cedex (France); Mukhopadhyay, Gauranga [Special Centre for Molecular Medicine, Jawaharlal Nehru University, New Delhi 110067 (India); Tyagi, Rakesh K., E-mail: rktyagi@yahoo.com [Special Centre for Molecular Medicine, Jawaharlal Nehru University, New Delhi 110067 (India)

2015-01-15

Nuclear receptor PXR is predominantly expressed in liver and intestine. Expression of PXR is observed to be dysregulated in various metabolic disorders indicating its involvement in disease development. However, information available on mechanisms of PXR self-regulation is fragmentary. The present investigation identifies some of the regulatory elements responsible for its tight regulation and low cellular expression. Here, we report that the PXR-promoter is a target for some key transcription factors like PU.1/Ets-1, Pax5, LEF-1 and c-Jun. Interestingly, we observed that PXR-promoter responsiveness to Pax5, LEF-1 and c-Jun, is considerably enhanced by Ets transcription factors (PU.1 and Ets-1). Co-transfection of cells with Ets-1, LEF-1 and c-Jun increased PXR-promoter activity by 5-fold and also induced expression of endogenous human PXR. Site-directed mutagenesis and transfection studies revealed that two Ets binding sites and two of the three LEF binding sites in the PXR-promoter are functional and have a positive effect on PXR transcription. Results suggest that expression of Ets family members, in conjunction with Pax5, LEF-1 and c-Jun, lead to coordinated up-regulation of PXR gene transcription. Insights obtained on the regulation of PXR gene have relevance in offering important cues towards normal functioning as well as development of several metabolic disorders via PXR signaling. - Highlights: • The study identified cis-regulatory elements in the nuclear receptor PXR promoter. • Several trans-acting factors modulating the PXR-promoter have been identified. • PU.1/Ets-1, Pax5, LEF-1, c-Jun, LyF-VI and NF-1 act as modulators of the PXR-promoter. • Ets-1 in conjunction with LEF-1 and c-Jun exhibit 5-fold activation of the PXR-promoter. • Insights into PXR-regulation have relevance in normal and pathological conditions.

Cooperation of three WRKY-domain transcription factors WRKY18, WRKY40, and WRKY60 in repressing two ABA-responsive genes ABI4 and ABI5 in Arabidopsis

OpenAIRE

Liu, Zhi-Qiang; Yan, Lu; Wu, Zhen; Mei, Chao; Lu, Kai; Yu, Yong-Tao; Liang, Shan; Zhang, Xiao-Feng; Wang, Xiao-Fang; Zhang, Da-Peng

2012-01-01

Three evolutionarily closely related WRKY-domain transcription factors WRKY18, WRKY40, and WRKY60 in Arabidopsis were previously identified as negative abscisic acid (ABA) signalling regulators, of which WRKY40 regulates ABI4 and ABI5 expression, but it remains unclear whether and how the three transcription factors cooperate to regulate expression of ABI4 and ABI5. In the present experiments, it was shown that WRKY18 and WRKY60, like WRKY40, interact with the W-box in the promoters of ABI4 a...

Murine Leukemia Virus Uses TREX Components for Efficient Nuclear Export of Unspliced Viral Transcripts

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Toshie Sakuma

2014-03-01

Full Text Available Previously we reported that nuclear export of both unspliced and spliced murine leukemia virus (MLV transcripts depends on the nuclear export factor (NXF1 pathway. Although the mRNA export complex TREX, which contains Aly/REF, UAP56, and the THO complex, is involved in the NXF1-mediated nuclear export of cellular mRNAs, its contribution to the export of MLV mRNA transcripts remains poorly understood. Here, we studied the involvement of TREX components in the export of MLV transcripts. Depletion of UAP56, but not Aly/REF, reduced the level of both unspliced and spliced viral transcripts in the cytoplasm. Interestingly, depletion of THO components, including THOC5 and THOC7, affected only unspliced viral transcripts in the cytoplasm. Moreover, the RNA immunoprecipitation assay showed that only the unspliced viral transcript interacted with THOC5. These results imply that MLV requires UAP56, THOC5 and THOC7, in addition to NXF1, for nuclear export of viral transcripts. Given that naturally intronless mRNAs, but not bulk mRNAs, require THOC5 for nuclear export, it is plausible that THOC5 plays a key role in the export of unspliced MLV transcripts.

The Not5 subunit of the ccr4-not complex connects transcription and translation.

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Zoltan Villanyi

2014-10-01

Full Text Available Recent studies have suggested that a sub-complex of RNA polymerase II composed of Rpb4 and Rpb7 couples the nuclear and cytoplasmic stages of gene expression by associating with newly made mRNAs in the nucleus, and contributing to their translation and degradation in the cytoplasm. Here we show by yeast two hybrid and co-immunoprecipitation experiments, followed by ribosome fractionation and fluorescent microscopy, that a subunit of the Ccr4-Not complex, Not5, is essential in the nucleus for the cytoplasmic functions of Rpb4. Not5 interacts with Rpb4; it is required for the presence of Rpb4 in polysomes, for interaction of Rpb4 with the translation initiation factor eIF3 and for association of Rpb4 with mRNAs. We find that Rpb7 presence in the cytoplasm and polysomes is much less significant than that of Rpb4, and that it does not depend upon Not5. Hence Not5-dependence unlinks the cytoplasmic functions of Rpb4 and Rpb7. We additionally determine with RNA immunoprecipitation and native gel analysis that Not5 is needed in the cytoplasm for the co-translational assembly of RNA polymerase II. This stems from the importance of Not5 for the association of the R2TP Hsp90 co-chaperone with polysomes translating RPB1 mRNA to protect newly synthesized Rpb1 from aggregation. Hence taken together our results show that Not5 interconnects translation and transcription.

Dissection of Ire1 functions reveals stress response mechanisms uniquely evolved in Candida glabrata.

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Taiga Miyazaki

2013-01-01

Full Text Available Proper protein folding in the endoplasmic reticulum (ER is vital in all eukaryotes. When misfolded proteins accumulate in the ER lumen, the transmembrane kinase/endoribonuclease Ire1 initiates splicing of HAC1 mRNA to generate the bZIP transcription factor Hac1, which subsequently activates its target genes to increase the protein-folding capacity of the ER. This cellular machinery, called the unfolded protein response (UPR, is believed to be an evolutionarily conserved mechanism in eukaryotes. In this study, we comprehensively characterized mutant phenotypes of IRE1 and other related genes in the human fungal pathogen Candida glabrata. Unexpectedly, Ire1 was required for the ER stress response independently of Hac1 in this fungus. C. glabrata Ire1 did not cleave mRNAs encoding Hac1 and other bZIP transcription factors identified in the C. glabrata genome. Microarray analysis revealed that the transcriptional response to ER stress is not mediated by Ire1, but instead is dependent largely on calcineurin signaling and partially on the Slt2 MAPK pathway. The loss of Ire1 alone did not confer increased antifungal susceptibility in C. glabrata contrary to UPR-defective mutants in other fungi. Taken together, our results suggest that the canonical Ire1-Hac1 UPR is not conserved in C. glabrata. It is known in metazoans that active Ire1 nonspecifically cleaves and degrades a subset of ER-localized mRNAs to reduce the ER load. Intriguingly, this cellular response could occur in an Ire1 nuclease-dependent fashion in C. glabrata. We also uncovered the attenuated virulence of the C. glabrata Δire1 mutant in a mouse model of disseminated candidiasis. This study has unveiled the unique evolution of ER stress response mechanisms in C. glabrata.

Interplay between the HTLV-2 Tax and APH-2 proteins in the regulation of the AP-1 pathway

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Marban Céline

2012-12-01

Full Text Available Abstract Background In contrast with human T-cell leukemia virus type 1 (HTLV-1 that causes ATL (adult T-cell leukemia, HTLV-2 has not been causally linked to malignant disease. The minus strand of the HTLV genomes encode the regulatory proteins HTLV-1 bZIP factor (HBZ for HTLV-1 and antisense protein of HTLV-2 (APH-2 for HTLV-2. Unlike the viral proteins Tax1 and Tax2, both HBZ and APH-2 are constitutively expressed in infected cells suggesting that they may play important roles in the pathogenesis of these viruses. To date, very little is known about the function of APH-2 except that it inhibits Tax2-mediated transcription of HTLV-2 genes. In the present study, we investigated the role of APH-2 in basal and Tax2B-mediated activation of the AP-1 pathway. Results We demonstrate that, unlike HBZ, APH-2 stimulates basal AP-1 transcription by interacting with c-Jun and JunB through its non-conventional bZIP domain. In addition, when Tax2 and APH-2 are co-expressed, they physically interact in vivo and in vitro and APH-2 acts as an inhibitor of Tax2-mediated activation of AP-1 transcription. Conclusions This report is the first to document that HTLV-2 can modulate the AP-1 pathway. Altogether our results reveal that, in contrast with HBZ, APH-2 regulates AP-1 activity in a Tax2-dependant manner. As the AP-1 pathway is involved in numerous cellular functions susceptible to affect the life cycle of the virus, these distinct biological properties between HBZ and APH-2 may contribute to the differential pathogenic potential of HTLV-1 and HTLV-2.

The formation of an aberrant PAX5 transcript in a patient with mixed phenotype acute leukemia harboring der(9t(7;9(q11.2;p13

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Jun Amaki

2016-01-01

Full Text Available We experienced the case of a 56-year-old male with B-lymphoid/myeloid lineage mixed phenotype acute leukemia (MPAL. A cytogenetic analysis of the patient's bone marrow revealed a complex karyotype, including der(9t(7;9(q11.2;p13. We identified an aberrant PAX5 transcript, including the exons 1A to 5 and the contiguous intron 5/6 sequence using the 3′ rapid amplification of cDNA ends-polymerase chain reaction method, and confirmed their expression in the leukemic cells. Our case suggests that der(9t(7;9(q11.2;p13 can cause the truncation of the PAX5 transcript, which is supposed to contribute to the generation of MPAL, in addition to three previously reported types of PAX5 fusion.

Transcriptional regulation by competing transcription factor modules.

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Rutger Hermsen

2006-12-01

Full Text Available Gene regulatory networks lie at the heart of cellular computation. In these networks, intracellular and extracellular signals are integrated by transcription factors, which control the expression of transcription units by binding to cis-regulatory regions on the DNA. The designs of both eukaryotic and prokaryotic cis-regulatory regions are usually highly complex. They frequently consist of both repetitive and overlapping transcription factor binding sites. To unravel the design principles of these promoter architectures, we have designed in silico prokaryotic transcriptional logic gates with predefined input-output relations using an evolutionary algorithm. The resulting cis-regulatory designs are often composed of modules that consist of tandem arrays of binding sites to which the transcription factors bind cooperatively. Moreover, these modules often overlap with each other, leading to competition between them. Our analysis thus identifies a new signal integration motif that is based upon the interplay between intramodular cooperativity and intermodular competition. We show that this signal integration mechanism drastically enhances the capacity of cis-regulatory domains to integrate signals. Our results provide a possible explanation for the complexity of promoter architectures and could be used for the rational design of synthetic gene circuits.

Transcriptional regulation of genes related to progesterone production.

Science.gov (United States)

Mizutani, Tetsuya; Ishikane, Shin; Kawabe, Shinya; Umezawa, Akihiro; Miyamoto, Kaoru

2015-01-01

Steroid hormones are synthesized from cholesterol in various tissues, mainly in the adrenal glands and gonads. Because these lipid-soluble steroid hormones immediately diffuse through the cells in which they are produced, their secretion directly reflects the activity of the genes related to their production. Progesterone is important not only for luteinization and maintenance of pregnancy, but also as a substrate for most other steroids. Steroidogenic acute regulatory protein (STAR), cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), and 3β-hydroxysteroid dehydrogenase/Δ(5)-Δ(4) isomerase (3β-HSD) are well-known proteins essential for progesterone production. In addition to them, glutathione S-transferase A1-1 and A3-3 are shown to exert Δ(5)-Δ(4) isomerization activity to produce progesterone in a cooperative fashion with 3β-HSD. 5-Aminolevulinic acid synthase 1, ferredoxin 1, and ferredoxin reductase also play a role in steroidogenesis as accessory factors. Members of the nuclear receptor 5A (NR5A) family (steroidogenic factor 1 and liver receptor homolog 1) play a crucial role in the transcriptional regulation of these genes. The NR5A family activates these genes by binding to NR5A responsive elements present within their promoter regions, as well as to the elements far from their promoters. In addition, various NR5A-interacting proteins including peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), nuclear receptor subfamily 0, group B, member 1 (DAX-1), and CCAAT/enhancer-binding proteins (C/EBP) are involved in the transcription of NR5A target genes and regulate the transcription either positively or negatively under both basal and tropic hormone-stimulated conditions. In this review, we describe the transcriptional regulation of genes related to progesterone production.

Identification of tyrosine residues in the intracellular domain of the growth hormone receptor required for transcriptional signaling and Stat5 activation

DEFF Research Database (Denmark)

Hansen, L. H.; Wang, X.; Kopchick, J J

1996-01-01

The binding of growth hormone (GH) to its receptor results in its dimerization followed by activation of Jak2 kinase and tyrosine phosphorylation of the GH receptor itself, as well as Jak2 and the transcription factors Stat1, -3, and -5. In order to study the role of GH receptor tyrosine phosphor...

RECQ5 helicase associates with the C-terminal repeat domain of RNA polymerase II during productive elongation phase of transcription

Czech Academy of Sciences Publication Activity Database

Kanagaraj, R.; Huehn, D.; Mackellar, A.; Menigatti, M.; Zheng, L.; Urban, Václav; Shevelev, Igor; Greenleaf, A.L.; Janščák, Pavel

2010-01-01

Roč. 38, č. 22 (2010), s. 8131-8140 ISSN 0305-1048 R&D Projects: GA ČR GA204/09/0565 Grant - others:SNSF(CH) 3100A0-116008; NIH(US) GM040505 Institutional research plan: CEZ:AV0Z50520514 Keywords : RECQ5 DNA helicase * transcription * genome stability Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.836, year: 2010

Identification and characterization of finger millet OPAQUE2 transcription factor gene under different nitrogen inputs for understanding their role during accumulation of prolamin seed storage protein.

Science.gov (United States)

Gaur, Vikram Singh; Kumar, Lallan; Gupta, Supriya; Jaiswal, J P; Pandey, Dinesh; Kumar, Anil

2018-03-01

In this study, we report the isolation and characterization of the mRNA encoding OPAQUE2 (O2) like TF of finger millet (FM) ( Eleusine coracana) ( EcO2 ). Full-length EcO2 mRNA was isolated using conserved primers designed by aligning O2 mRNAs of different cereals followed by 3' and 5' RACE (Rapid Amplification of cDNA Ends). The assembled full-length EcO2 mRNA was found to contain an ORF of 1248-nt coding the 416 amino acids O2 protein. Domain analysis revealed the presence of the BLZ and bZIP-C domains which is a characteristic feature of O2 proteins. Phylogenetic analysis of EcO2 protein with other bZIP proteins identified using finger millet transcriptome data and O2 proteins of other cereals showed that EcO2 shared high sequence similarity with barley BLZ1 protein. Transcripts of EcO2 were detected in root, stem, leaves, and seed development stages. Furthermore, to investigate nitrogen responsiveness and the role of EcO2 in regulating seed storage protein gene expression, the expression profiles of EcO2 along with an α-prolamin gene were studied during the seed development stages of two FM genotypes (GE-3885 and GE-1437) differing in grain protein content (13.8 and 6.2%, respectively) grown under increasing nitrogen inputs. Compared to GE-1437, the EcO2 was relatively highly expressed during the S2 stage of seed development which further increased as nitrogen input was increased. The Ecα - prolamin gene was strongly induced in the high protein genotype (GE-3885) at all nitrogen inputs. These results indicate the presence of nitrogen responsiveness regulatory elements which might play an important role in accumulating protein in FM genotypes through modulating EcO2 expression by sensing plant nitrogen status.

TRE5-A retrotransposition profiling reveals putative RNA polymerase III transcription complex binding sites on the Dictyostelium extrachromosomal rDNA element.

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Thomas Spaller

Full Text Available The amoeba Dictyostelium discoideum has a haploid genome in which two thirds of the DNA encodes proteins. Consequently, the space available for selfish mobile elements to expand without excess damage to the host genome is limited. The non-long terminal repeat retrotransposon TRE5-A maintains an active population in the D. discoideum genome and apparently adapted to this gene-dense environment by targeting positions ~47 bp upstream of tRNA genes that are devoid of protein-coding regions. Because only ~24% of tRNA genes are associated with a TRE5-A element in the reference genome, we evaluated whether TRE5-A retrotransposition is limited to this subset of tRNA genes. We determined that a tagged TRE5-A element (TRE5-Absr integrated at 384 of 405 tRNA genes, suggesting that expansion of the current natural TRE5-A population is not limited by the availability of targets. We further observed that TRE5-Absr targets the ribosomal 5S gene on the multicopy extrachromosomal DNA element that carries the ribosomal RNA genes, indicating that TRE5-A integration may extend to the entire RNA polymerase III (Pol III transcriptome. We determined that both natural TRE5-A and cloned TRE5-Absr retrotranspose to locations on the extrachromosomal rDNA element that contain tRNA gene-typical A/B box promoter motifs without displaying any other tRNA gene context. Based on previous data suggesting that TRE5-A targets tRNA genes by locating Pol III transcription complexes, we propose that A/B box loci reflect Pol III transcription complex assembly sites that possess a function in the biology of the extrachromosomal rDNA element.

Transcript structure and domain display: a customizable transcript visualization tool.

Science.gov (United States)

Watanabe, Kenneth A; Ma, Kaiwang; Homayouni, Arielle; Rushton, Paul J; Shen, Qingxi J

2016-07-01

Transcript Structure and Domain Display (TSDD) is a publicly available, web-based program that provides publication quality images of transcript structures and domains. TSDD is capable of producing transcript structures from GFF/GFF3 and BED files. Alternatively, the GFF files of several model organisms have been pre-loaded so that users only needs to enter the locus IDs of the transcripts to be displayed. Visualization of transcripts provides many benefits to researchers, ranging from evolutionary analysis of DNA-binding domains to predictive function modeling. TSDD is freely available for non-commercial users at http://shenlab.sols.unlv.edu/shenlab/software/TSD/transcript_display.html : jeffery.shen@unlv.nevada.edu. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Novel mechanism for Fc epsilon RI-mediated signal transducer and activator of transcription 5 (STAT5) tyrosine phosphorylation and the selective influence of STAT5B over mast cell cytokine production

Czech Academy of Sciences Publication Activity Database

Pullen, N.A.; Barnstein, B.O.; Falanga, Y.T.; Wang, Z.Q.; Suzuki, R.; Tamang, T.D.L.; Khurana, M.C.; Harry, E.A.; Dráber, Petr; Bunting, K.D.; Mizuno, K.; Wilson, B.S.; Ryan, J.J.

2012-01-01

Roč. 287, č. 3 (2012), s. 2045-2054 ISSN 0021-9258 R&D Projects: GA MŠk 1M0506; GA ČR GAP302/10/1759; GA ČR GA301/09/1826 Grant - others:NIH(US) 1R01AI59638; NIH(US) R01DK059380; NIH(US) AI051575; NIH(US) GM065794; VCU(US) U19A1077435 Institutional support: RVO:68378050 Keywords : cytokine induction * STAT transcription factor * mast cell * Fyn kinase * STAT5 Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.651, year: 2012

Beyond ubiquitination: the atypical functions of Fbxo7 and other F-box proteins.

Science.gov (United States)

Nelson, David E; Randle, Suzanne J; Laman, Heike

2013-10-09

F-box proteins (FBPs) are substrate-recruiting subunits of Skp1-cullin1-FBP (SCF)-type E3 ubiquitin ligases. To date, 69 FBPs have been identified in humans, but ubiquitinated substrates have only been identified for a few, with the majority of FBPs remaining 'orphans'. In recent years, a growing body of work has identified non-canonical, SCF-independent roles for about 12% of the human FBPs. These atypical FBPs affect processes as diverse as transcription, cell cycle regulation, mitochondrial dynamics and intracellular trafficking. Here, we provide a general review of FBPs, with a particular emphasis on these expanded functions. We review Fbxo7 as an exemplar of this special group as it has well-defined roles in both SCF and non-SCF complexes. We review its function as a cell cycle regulator, via its ability to stabilize p27 protein and Cdk6 complexes, and as a proteasome regulator, owing to its high affinity binding to PI31. We also highlight recent advances in our understanding of Fbxo7 function in Parkinson's disease, where it functions in the regulation of mitophagy with PINK1 and Parkin. We postulate that a few extraordinary FBPs act as platforms that seamlessly segue their canonical and non-canonical functions to integrate different cellular pathways and link their regulation.

Separation of replication and transcription domains in nucleoli.

Science.gov (United States)

Smirnov, E; Borkovec, J; Kováčik, L; Svidenská, S; Schröfel, A; Skalníková, M; Švindrych, Z; Křížek, P; Ovesný, M; Hagen, G M; Juda, P; Michalová, K; Cardoso, M C; Cmarko, D; Raška, I

2014-12-01

In mammalian cells, active ribosomal genes produce the 18S, 5.8S and 28S RNAs of ribosomal particles. Transcription levels of these genes are very high throughout interphase, and the cell needs a special strategy to avoid collision of the DNA polymerase and RNA polymerase machineries. To investigate this problem, we measured the correlation of various replication and transcription signals in the nucleoli of HeLa, HT-1080 and NIH 3T3 cells using a specially devised software for analysis of confocal images. Additionally, to follow the relationship between nucleolar replication and transcription in living cells, we produced a stable cell line expressing GFP-RPA43 (subunit of RNA polymerase I, pol I) and RFP-PCNA (the sliding clamp protein) based on human fibrosarcoma HT-1080 cells. We found that replication and transcription signals are more efficiently separated in nucleoli than in the nucleoplasm. In the course of S phase, separation of PCNA and pol I signals gradually increased. During the same period, separation of pol I and incorporated Cy5-dUTP signals decreased. Analysis of single molecule localization microscopy (SMLM) images indicated that transcriptionally active FC/DFC units (i.e. fibrillar centers with adjacent dense fibrillar components) did not incorporate DNA nucleotides. Taken together, our data show that replication of the ribosomal genes is spatially separated from their transcription, and FC/DFC units may provide a structural basis for that separation. Copyright © 2014 Elsevier Inc. All rights reserved.

Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes.

Science.gov (United States)

Riechmann, J L; Heard, J; Martin, G; Reuber, L; Jiang, C; Keddie, J; Adam, L; Pineda, O; Ratcliffe, O J; Samaha, R R; Creelman, R; Pilgrim, M; Broun, P; Zhang, J Z; Ghandehari, D; Sherman, B K; Yu, G

2000-12-15

The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription.

Dicty_cDB: SHA788 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available . 52 6e-09 5 CV164410 |CV164410.1 rsmsxl_003361.y1.scf cDNA Library of Salvia mil...tiorrhiza Salvia miltiorrhiza cDNA 5', mRNA sequence. 54 6e-09 3 CV172465 |CV172465.1 rsmsxlre_008717.z1.scf

Luminescent and scintillation properties of the Pr"3"+ doped single crystalline films of Lu_3Al_5_−_xGa_xO_1_2 garnet

International Nuclear Information System (INIS)

Gorbenko, V.; Zorenko, Yu; Zorenko, T.; Voznyak, T.; Paprocki, K.; Fabisiak, K.; Fedorov, A.; Bilski, P.; Twardak, A.; Zhusupkalieva, G.

2016-01-01

The Pr"3"+ d–f luminescence was investigated in the single crystalline films (SCF) of Lu_3Al_5_−_xGa_xO_1_2:Pr garnet solid solution at x = 1–3, grown by the liquid phase epitaxy (LPE) method from the melt-solution based on the PbO–B_2O_3 flux. The shape of CL spectra and decay kinetics of Pr"3"+ ions in Lu_3Al_5_−_xGa_xO_1_2 SCFs strongly depend on the total gallium concentration x and distribution of Ga"3"+ ions between the tetrahedral and octahedral position of the garnet host. The best scintillation properties of Lu_3Al_5_−_xGa_xO_1_2:Pr SCF are achieved at the nominal Ga content in melt-solution in the x = 2–2.5 range. - Highlights: • Single crystalline films of Lu_3Al_5_−_x Ga_xO_1_2:Pr garnet at x = 1–3 were grown by the LPE method. • Pr"3"+ emission spectra, light yield and decay time of films show strong dependence on Ga content. • The maximal light yield of Lu_3Al_5_−_x Ga_xO_1_2:Pr film is observed at Ga content x = 2.0–2.5.

Identification of novel microRNAs in post-transcriptional control of Nrf2 expression and redox homeostasis in neuronal, SH-SY5Y cells.

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Madhusudhanan Narasimhan

Full Text Available Nuclear factor-erythroid 2-related factor 2 (Nrf2/NFE2L2, a redox-sensitive transcription factor plays a critical role in adaptation to cellular stress and affords cellular defense by initiating transcription of antioxidative and detoxification genes. While a protein can be regulated at multiple levels, control of Nrf2 has been largely studied at post-translational regulation points by Keap1. Importantly, post-transcriptional/translational based regulation of Nrf2 is less understood and to date there are no reports on such mechanisms in neuronal systems. In this context, studies involving the role of microRNAs (miRs which are normally considered as fine tuning regulators of protein production through translation repression and/or post-transcriptional alterations, are in place. In the current study, based on in-silico analysis followed by immunoblotting and real time analysis, we have identified and validated for the first time that human NFE2L2 could be targeted by miR153/miR27a/miR142-5p/miR144 in neuronal, SH-SY5Y cells. Co-transfection studies with individual miR mimics along with either WT 3' UTR of human Nrf2 or mutated miRNA targeting seed sequence within Nrf2 3' UTR, demonstrated that Nrf2 is a direct regulatory target of these miRs. In addition, ectopic expression of miR153/miR27a/miR142-5p/miR144 affected Nrf2 mRNA abundance and nucleo-cytoplasmic concentration of Nrf2 in a Keap1 independent manner resulting in inefficient transactivating ability of Nrf2. Furthermore, forced expression of miRs diminished GCLC and GSR expression resulting in alteration of Nrf2 dependent redox homeostasis. Finally, bioinformatics based miRNA-disease network analysis (MDN along with extended computational network analysis of Nrf2 associated pathologic processes suggests that if in a particular cellular scenario where any of these miR153/miR27a/miR142-5p/miR144 either individually or as a group is altered, it could affect Nrf2 thus triggering and

Typing of Poultry Influenza Virus (H5 and H7 by Reverse Transcription- Polymerase Chain Reaction

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Cesare Bonacina

2010-01-01

Full Text Available The ability of the influenza Orthomixovirus to undergo to continually antigenically changes that can affect its pathogenicity and its diffusion, explains the growing seriousness of this disease and the recent epizoozies in various parts of the world. There have been 15 HA and 9 NA type A sub-types of the influenza virus identified all of which are present in birds. Until now the very virulent avian influenza viruses identified were all included to the H5 and H7 sub-types. We here show that is possible to identify the H5 and H7 sub-types with reverse transcription-polymerase chain reaction (RT-PCR by using a set of specific primers for each HA sub-type. The RT-PCR is a quick and sensitive method of identifying the HA sub-types of the influenza virus directly from homogenised organs.

Evaluation of Long Stress-Induced Non-coding Transcripts 5 Polymorphism in Iranian Patients with Bladder Cancer

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Mahla Nazari

2016-07-01

Full Text Available Background: Bladder cancer (BC is the most commonly diagnosed genitourinary cancer in Iran, presented in both men and women. BC is a multifactorial trait resulting from the complex interaction between several genes and environmental factors. Long stress-induced non-coding transcript 5 (LSINCT5, a member of the long non-coding RNAs, is abundantly expressed in high proliferative cells, as well as the cells vulnerable to cellular stress in response to chemical carcinogens. This case-control study aimed to determine any association between LSINCT5 rs2962586 polymorphism and bladder cancer. Materials and Methods: A group of 150 patients with BC were compared with 143 subjects as a control group. Genotyping of the rs2962586 polymorphism was done using tetra- primer amplification refractory mutation system-polymerase chain reaction (T-ARMS PCR method. Results: Genotype and allele distribution were not significantly different between the case and control groups. Smoking was found to be the confounding risk factor for bladder cancer. Conclusion: Considering the result of our analyses, it seems that LSINCT5 could not affect individual susceptibility to BC among Iranian patients, however, it can be considered as a disease predictor among smokers.

5-HT2A receptor deficiency alters the metabolic and transcriptional, but not the behavioral, consequences of chronic unpredictable stress

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Minal Jaggar

2017-12-01

Full Text Available Chronic stress enhances risk for psychiatric disorders, and in animal models is known to evoke depression-like behavior accompanied by perturbed neurohormonal, metabolic, neuroarchitectural and transcriptional changes. Serotonergic neurotransmission, including serotonin2A (5-HT2A receptors, have been implicated in mediating specific aspects of stress-induced responses. Here we investigated the influence of chronic unpredictable stress (CUS on depression-like behavior, serum metabolic measures, and gene expression in stress-associated neurocircuitry of the prefrontal cortex (PFC and hippocampus in 5-HT2A receptor knockout (5-HT2A−/− and wild-type mice of both sexes. While 5-HT2A−/− male and female mice exhibited a baseline reduced anxiety-like state, this did not alter the onset or severity of behavioral despair during and at the cessation of CUS, indicating that these mice can develop stress-evoked depressive behavior. Analysis of metabolic parameters in serum revealed a CUS-evoked dyslipidemia, which was abrogated in 5-HT2A−/− female mice with a hyperlipidemic baseline phenotype. 5-HT2A−/− male mice in contrast did not exhibit such a baseline shift in their serum lipid profile. Specific stress-responsive genes (Crh, Crhr1, Nr3c1, and Nr3c2, trophic factors (Bdnf, Igf1 and immediate early genes (IEGs (Arc, Fos, Fosb, Egr1-4 in the PFC and hippocampus were altered in 5-HT2A−/− mice both under baseline and CUS conditions. Our results support a role for the 5-HT2A receptor in specific metabolic and transcriptional, but not behavioral, consequences of CUS, and highlight that the contribution of the 5-HT2A receptor to stress-evoked changes is sexually dimorphic. Keywords: 5-HT2A−/− mice, Prefrontal cortex, Hippocampus, Gene expression, Sexual dimorphism, Despair

In vitro effects of recombinant human stem cell factor on hematopoietic cells from patients with acute radiation sickness

International Nuclear Information System (INIS)

Li Chuansheng; Cheng Tao; Xu Yanqun

1994-01-01

The effects of rhSCF, rhPIXY 321, rhGM-CSF and rhIL-3 on clonal proliferation of hematopoietic cells from five cases of acute radiation sickness were studied. The results showed that rhSCF could stimulate clonal proliferation of normal hematopoietic cells and the best results were obtained when the concentration of rhSCF was 5 x 10 4 ng/L. Clonal proliferation of hematopoietic cells from four cases of acute radiation sickness was stimulated while that from one case was inhibited. Moreover, the responsiveness of cells to rhSCF was correlated with the doses of radiation. Analysis of cell surface antigen, cell morphology and histochemistry revealed that rhSCF promoted predominantly the proliferation of granulocyte-macrophage lineage. rhSCF in combination with other three factors could further enhance the clonal proliferation of hematopoietic cells. The effects of rhPIXY 321, a fusion protein of GM-CSF and IL-3, were also analysed and found it to be a novel valuable hematopoietic growth factor

Interplay among Drosophila transcription factors Ets21c, Fos and Ftz-F1 drives JNK-mediated tumor malignancy

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Eva Külshammer

2015-10-01

Full Text Available Cancer initiation and maintenance of the transformed cell state depend on altered cellular signaling and aberrant activities of transcription factors (TFs that drive pathological gene expression in response to cooperating genetic lesions. Deciphering the roles of interacting TFs is therefore central to understanding carcinogenesis and for designing cancer therapies. Here, we use an unbiased genomic approach to define a TF network that triggers an abnormal gene expression program promoting malignancy of clonal tumors, generated in Drosophila imaginal disc epithelium by gain of oncogenic Ras (RasV12 and loss of the tumor suppressor Scribble (scrib1. We show that malignant transformation of the rasV12scrib1 tumors requires TFs of distinct families, namely the bZIP protein Fos, the ETS-domain factor Ets21c and the nuclear receptor Ftz-F1, all acting downstream of Jun-N-terminal kinase (JNK. Depleting any of the three TFs improves viability of tumor-bearing larvae, and this positive effect can be enhanced further by their combined removal. Although both Fos and Ftz-F1 synergistically contribute to rasV12scrib1 tumor invasiveness, only Fos is required for JNK-induced differentiation defects and Matrix metalloprotease (MMP1 upregulation. In contrast, the Fos-dimerizing partner Jun is dispensable for JNK to exert its effects in rasV12scrib1 tumors. Interestingly, Ets21c and Ftz-F1 are transcriptionally induced in these tumors in a JNK- and Fos-dependent manner, thereby demonstrating a hierarchy within the tripartite TF network, with Fos acting as the most upstream JNK effector. Of the three TFs, only Ets21c can efficiently substitute for loss of polarity and cooperate with RasV12 in inducing malignant clones that, like rasV12scrib1 tumors, invade other tissues and overexpress MMP1 and the Drosophila insulin-like peptide 8 (Dilp8. While rasV12ets21c tumors require JNK for invasiveness, the JNK activity is dispensable for their growth. In conclusion, our

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.

The T-cell receptor beta chain CDR3 region of BV8S1/BJ1S5 transcripts in type 1 diabetes.

Science.gov (United States)

Naserke, H E; Durinovic-Bellò, I; Seidel, D; Ziegler, A G

1996-01-01

We recently described the T-cell receptor (TCR) beta chain CDR3 motif S-SDRLG-NQPQH (BV8S1-BJ1S5) in an islet-specific T-cell clone (K2.12) from a type 1 diabetic patient (AS). A similar motif (RLGNQ) was also reported in a T-cell clone of non-obese diabetic (NOD) mice by others. In order to determine the frequency of our motif in selected and unselected T-cell populations, we cloned and sequenced the CDR3 region of BV8S1-BJ1S5 transcripts. These transcripts were derived from unstimulated peripheral blood T lymphocytes from two type 1 diabetic patients (AS and FS) and their non-diabetic sibling (WS), as well as from an islet-specific T-cell line of one of the patients. In addition, we compared the structure and composition of the CDR3 region in BV8S1-BJ1S5 transcripts from peripheral blood T cells between the patients and their non-diabetic sibling (>50 sequences each). We found that 30% of the islet-specific T-cell line cDNA clones expressed the entire sequence-motif, whereas it was absent in the clones of unstimulated peripheral blood T cells from both patients and their non-diabetic sibling. The average length of the CDR3 region was shorter in the patients (mean AS 9.9, FS 9.9, versus WS 10.7, p = 0.0037) and the number of inserted nucleotides in N nucleotide addition at the DJ-junction lower (mean AS 3.5, FS 3. 2, versus WS 5.2, P = diabetic sibling. Moreover, the pattern of amino acid usage in the CDR3 region was dissimilar at positions 5 and 6, where polar amino acids predominated in both diabetic siblings. In contrast, basic amino acids are preferentially used at position 5 in the clones of the non-diabetic sibling. These data provide information on the general structure of the TCR(BV8S1-BJ1S5) CDR3 region in type 1 diabetes and may indicate differences in the amino and nucleic acid composition of the TCR beta chain CDR3 region between two type 1 diabetic patients and their non-diabetic sibling.

In vitro synthesis of biologically active transcripts of tomato black ring virus satellite RNA.

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Greif, C; Hemmer, O; Demangeat, G; Fritsch, C

1990-04-01

Synthetic transcripts of tomato black ring virus satellite RNA (TBRV satRNA), isolate L, were prepared from cDNA cloned in the Bluescribe transcription vector. Transcripts with 49 (T49L) or two (T2GL) extra nucleotides at their 5' ends and 42 extra nucleotides at their 3' ends were able to induce, but to different extents, the synthesis in vitro of the satRNA-encoded 48K protein. However, when inoculated into Chenopodium quinoa together with TBRV L genomic RNAs, only T2GL was biologically active, in the presence or absence of a 5' cap analogue in the transcription reactions. Analysis of the 5' and 3' termini of the satRNA isolated from plants showed that nonviral extensions were not maintained in the transcript progeny.

High Power 1443.5 nm Laser with Nd:YAG Single Crystal Fiber

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Han Rao

2017-07-01

Full Text Available A high-power eye-safe 1443.5 nm laser was demonstrated with an Nd:YAG single crystal fiber (SCF as the gain medium. For continuous wave (CW operation, a maximum output power of 13.3 W was obtained under an absorbed pump power of 95.0 W, corresponding to an optical-to-optical conversion efficiency of 14.0%. For acousto-optically (AO Q-switched regime, an output power of 1.95 W was obtained at a pulse repetition frequency (PRF of 10 kHz. The pulse duration was 69.5 ns. The pulse energy and peak power were calculated to be 195 µJ and 2.81 kW, respectively.

Directing traffic on DNA-How transcription factors relieve or induce transcriptional interference.

Science.gov (United States)

Hao, Nan; Palmer, Adam C; Dodd, Ian B; Shearwin, Keith E

2017-03-15

Transcriptional interference (TI) is increasingly recognized as a widespread mechanism of gene control, particularly given the pervasive nature of transcription, both sense and antisense, across all kingdoms of life. Here, we discuss how transcription factor binding kinetics strongly influence the ability of a transcription factor to relieve or induce TI.

Stem cell factor enhances the survival of murine intestinal stem cells after photon irradiation

International Nuclear Information System (INIS)

Leigh, B.R.; Khan, W.; Hancock, S.L.

1995-01-01

Recombinant rat stem cell factor (SCF) has been shown to decrease lethality in mice exposed to total-body irradiation (TBI) in the lower range of lethality through radioprotection of hematopoietic stem cells and acceleration of bone marrow repopulation. This study evaluates the effect of SCF on the survival of the intestinal mucosal stem cell after TBI. This non-hematopoietic cell is clinically relevant. Gastrointestinal toxicity is common during and after abdominal and pelvic radiation therapy and limits the radiation dose in these regions. As observed with bone marrow, the administration of SCF to mice prior to TBI enhanced the survival of mouse duodenal crypt stem cells. The maximum enhancement of survival was seen when 100 μ/kg of SCF was given intraperitoneally 8 h before irradiation. This regimen increased the survival of duodenal crypt stem cells after 12.0 Gy TBI from 22.5 ± 0.7 per duodenal cross section for controls to 30.0 ± 1.7 after treatment with SCF (P=0.03). The TBI dose producing 50% mortality of 6 days (LD 50/6 ) was increased from 14.9 Gy for control mice to 19.0 Gy for mice treated with SCF (dose modification factor = 1.28). These findings demonstrate that SCF (dose modification factor = 1.28). These findings demonstrate that SCF has radioprotective effects on a non-hematopoietic stem cell population and suggest that SCF may be of clinical value in preventing radiation injury to the intestine. 29 refs., 4 figs

Hypertonic-induced lamin A/C synthesis and distribution to nucleoplasmic speckles is mediated by TonEBP/NFAT5 transcriptional activator

International Nuclear Information System (INIS)

Favale, Nicolas O.; Sterin Speziale, Norma B.; Fernandez Tome, Maria C.

2007-01-01

Lamin A/C is the most studied nucleoskeletal constituent. Lamin A/C expression indicates cell differentiation and is also a structural component of nuclear speckles, which are involved in gene expression regulation. Hypertonicity has been reported to induce renal epithelial cell differentiation and expression of TonEBP (NFAT5), a transcriptional activator of hypertonicity-induced gene transcription. In this paper, we investigate the effect of hypertonicity on lamin A/C expression in MDCK cells and the involvement of TonEBP. Hypertonicity increased lamin A/C expression and its distribution to nucleoplasm with speckled pattern. Microscopy showed codistribution of TonEBP and lamin A/C in nucleoplasmic speckles, and immunoprecipitation demonstrated their interaction. TonEBP silencing caused lamin A/C redistribution from nucleoplasmic speckles to the nuclear rim, followed by lamin decrease, thus showing that hypertonicity induces lamin A/C speckles through a TonEBP-dependent mechanism. We suggest that lamin A/C speckles could serve TonEBP as scaffold thus favoring its role in hypertonicity

Dicty_cDB: SHL126 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available ce. 52 6e-09 5 CV164410 |CV164410.1 rsmsxl_003361.y1.scf cDNA Library of Salvia miltiorrhiza Salvia miltiorr...hiza cDNA 5', mRNA sequence. 54 7e-09 3 CV172465 |CV172465.1 rsmsxlre_008717.z1.scf cDNA Library of Salvia m

Interaction of Arabidopsis Trihelix-Domain Transcription Factors VFP3 and VFP5 with Agrobacterium Virulence Protein VirF.

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Elena García-Cano

Full Text Available Agrobacterium is a natural genetic engineer of plants that exports several virulence proteins into host cells in order to take advantage of the cell machinery to facilitate transformation and support bacterial growth. One of these effectors is the F-box protein VirF, which presumably uses the host ubiquitin/proteasome system (UPS to uncoat the packaging proteins from the invading bacterial T-DNA. By analogy to several other bacterial effectors, VirF most likely has several functions in the host cell and, therefore, several interacting partners among host proteins. Here we identify one such interactor, an Arabidopsis trihelix-domain transcription factor VFP3, and further show that its very close homolog VFP5 also interacted with VirF. Interestingly, interactions of VirF with either VFP3 or VFP5 did not activate the host UPS, suggesting that VirF might play other UPS-independent roles in bacterial infection. To better understand the potential scope of VFP3 function, we used RNAi to reduce expression of the VFP3 gene. Transcriptome profiling of these VFP3-silenced plants using high-throughput cDNA sequencing (RNA-seq revealed that VFP3 substantially affected plant gene expression; specifically, 1,118 genes representing approximately 5% of all expressed genes were significantly either up- or down-regulated in the VFP3 RNAi line compared to wild-type Col-0 plants. Among the 507 up-regulated genes were genes implicated in the regulation of transcription, protein degradation, calcium signaling, and hormone metabolism, whereas the 611 down-regulated genes included those involved in redox regulation, light reactions of photosynthesis, and metabolism of lipids, amino acids, and cell wall. Overall, this pattern of changes in gene expression is characteristic of plants under stress. Thus, VFP3 likely plays an important role in controlling plant homeostasis.

Interaction of Arabidopsis Trihelix-Domain Transcription Factors VFP3 and VFP5 with Agrobacterium Virulence Protein VirF

Science.gov (United States)

García-Cano, Elena; Magori, Shimpei; Sun, Qi; Ding, Zehong; Lazarowitz, Sondra G.; Citovsky, Vitaly

2015-01-01

Agrobacterium is a natural genetic engineer of plants that exports several virulence proteins into host cells in order to take advantage of the cell machinery to facilitate transformation and support bacterial growth. One of these effectors is the F-box protein VirF, which presumably uses the host ubiquitin/proteasome system (UPS) to uncoat the packaging proteins from the invading bacterial T-DNA. By analogy to several other bacterial effectors, VirF most likely has several functions in the host cell and, therefore, several interacting partners among host proteins. Here we identify one such interactor, an Arabidopsis trihelix-domain transcription factor VFP3, and further show that its very close homolog VFP5 also interacted with VirF. Interestingly, interactions of VirF with either VFP3 or VFP5 did not activate the host UPS, suggesting that VirF might play other UPS-independent roles in bacterial infection. To better understand the potential scope of VFP3 function, we used RNAi to reduce expression of the VFP3 gene. Transcriptome profiling of these VFP3-silenced plants using high-throughput cDNA sequencing (RNA-seq) revealed that VFP3 substantially affected plant gene expression; specifically, 1,118 genes representing approximately 5% of all expressed genes were significantly either up- or down-regulated in the VFP3 RNAi line compared to wild-type Col-0 plants. Among the 507 up-regulated genes were genes implicated in the regulation of transcription, protein degradation, calcium signaling, and hormone metabolism, whereas the 611 down-regulated genes included those involved in redox regulation, light reactions of photosynthesis, and metabolism of lipids, amino acids, and cell wall. Overall, this pattern of changes in gene expression is characteristic of plants under stress. Thus, VFP3 likely plays an important role in controlling plant homeostasis. PMID:26571494

Metagenomic screening for aromatic compound-responsive transcriptional regulators.

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Taku Uchiyama

Full Text Available We applied a metagenomics approach to screen for transcriptional regulators that sense aromatic compounds. The library was constructed by cloning environmental DNA fragments into a promoter-less vector containing green fluorescence protein. Fluorescence-based screening was then performed in the presence of various aromatic compounds. A total of 12 clones were isolated that fluoresced in response to salicylate, 3-methyl catechol, 4-chlorocatechol and chlorohydroquinone. Sequence analysis revealed at least 1 putative transcriptional regulator, excluding 1 clone (CHLO8F. Deletion analysis identified compound-specific transcriptional regulators; namely, 8 LysR-types, 2 two-component-types and 1 AraC-type. Of these, 9 representative clones were selected and their reaction specificities to 18 aromatic compounds were investigated. Overall, our transcriptional regulators were functionally diverse in terms of both specificity and induction rates. LysR- and AraC- type regulators had relatively narrow specificities with high induction rates (5-50 fold, whereas two-component-types had wide specificities with low induction rates (3 fold. Numerous transcriptional regulators have been deposited in sequence databases, but their functions remain largely unknown. Thus, our results add valuable information regarding the sequence-function relationship of transcriptional regulators.

Light-harvesting complex gene expression is controlled by both transcriptional and post-transcriptional mechanisms during photoacclimation in Chlamydomonas reinhardtii

CERN Document Server

Durnford Dion, G; McKim, Sarah M; Sarchfield, Michelle L

2003-01-01

To compensate for increases in photon flux density (PFD), photosynthetic organisms possess mechanisms for reversibly modulating their photosynthetic apparatus to minimize photodamage. The photoacclimation response in Chlamydomonas reinhardtii was assessed following a 10-fold increase in PFD over 24h. In addition to a 50% reduction in the amount of chlorophyll and light-harvesting complexes (LHC) per cell, the expression of genes encoding polypeptides of the light-harvesting antenna were also affected. The abundance of Lhcb (a LHCH gene), Lhcb4 (a CP29-like gene), and Lhca (a LHCI gene) transcripts were reduced by 65 to 80%, within 1-2 h; however, the RNA levels of all three genes recovered to their low-light (LL) concentrations within 6-8 h. To determine the role of transcript turnover in this transient decline in abundance, the stability of all transcripts was measured. Although there was no change in the Lhcb or Lhca transcript turnover time, the Lhcb4 mRNA stability decreased 2.5-fold immediately following...

RNA Pol II promotes transcription of centromeric satellite DNA in beetles.

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Zeljka Pezer

Full Text Available Transcripts of centromeric satellite DNAs are known to play a role in heterochromatin formation as well as in establishment of the kinetochore. However, little is known about basic mechanisms of satellite DNA expression within constitutive heterochromatin and its regulation. Here we present comprehensive analysis of transcription of abundant centromeric satellite DNA, PRAT from beetle Palorus ratzeburgii (Coleoptera. This satellite is characterized by preservation and extreme sequence conservation among evolutionarily distant insect species. PRAT is expressed in all three developmental stages: larvae, pupae and adults at similar level. Transcripts are abundant comprising 0.033% of total RNA and are heterogeneous in size ranging from 0.5 kb up to more than 5 kb. Transcription proceeds from both strands but with 10 fold different expression intensity and transcripts are not processed into siRNAs. Most of the transcripts (80% are not polyadenylated and remain in the nucleus while a small portion is exported to the cytoplasm. Multiple, irregularly distributed transcription initiation sites as well as termination sites have been mapped within the PRAT sequence using primer extension and RLM-RACE. The presence of cap structure as well as poly(A tails in a portion of the transcripts indicate RNA polymerase II-dependent transcription and a putative polymerase II promoter site overlaps the most conserved part of the PRAT sequence. The treatment of larvae with alpha-amanitin decreases the level of PRAT transcripts at concentrations that selectively inhibit pol II activity. In conclusion, stable, RNA polymerase II dependant transcripts of abundant centromeric satellite DNA, not regulated by RNAi, have been identified and characterized. This study offers a basic understanding of expression of highly abundant heterochromatic DNA which in beetle species constitutes up to 50% of the genome.

Blending of soluble corn fiber with pullulan, sorbitol, or fructose attenuates glycemic and insulinemic responses in the dog and affects hydrolytic digestion in vitro.

Science.gov (United States)

de Godoy, M R C; Knapp, B K; Bauer, L L; Swanson, K S; Fahey, G C

2013-08-01

The objective of these experiments was to measure in vitro hydrolytic digestion and glycemic and insulinemic responses of select carbohydrate blends, all containing the novel carbohydrate soluble corn fiber (SCF). Two SCF that varied in their method of production were used to formulate the carbohydrate blends. One set of blends contained a SCF that was spray dried (SCFsd) and then blended with different amounts of either pullulan, sorbitol, or fructose. The other set of blends contained a SCF produced using longer evaporation time (SCF) and then blended with different ratios of pullulan, sorbitol, and fructose. Free sugar concentrations found in the individual SCFsd and SCF substrates were low but varied. Spray-dried soluble corn fiber had a reduced free sugar concentration compared with SCF (2.8 vs. 14.2%). Glucose was the main free sugar found in both SCFsd and SCF but at different concentrations (2.7 vs. 12.7%, respectively). The majority of the SCFsd blends were completely hydrolyzed to their monosaccharide components. Glucose accounted for most of the hydrolyzed monosaccharides for SCFsd and all the SCFsd blends. Hydrolyzed monosaccharide concentrations for the SCF:pullulan:sorbitol:fructose blends followed similar trends to the SCFsd blends where greater percentages of fructose and sorbitol resulted in decreased (P sorbitol. Total released monosaccharides were high in SCFsd blends containing either 50% fructose or sorbitol, but the combination resulted in reduced concentrations of glucose released (P sorbitol:fructose blends also had intermediate to high released monosaccharides as a result of in vitro hydrolytic digestion. All SCF blends resulted in decreased glycemic and insulinemic responses compared with the maltodextrin control (P sorbitol in the blends had the greatest impact on glycemic and insulinemic responses, even at concentrations as low as 5% of the blends. Overall, SCF and their blends may prove beneficial as components of low glycemic

Functional comparison of antisense proteins of HTLV-1 and HTLV-2 in viral pathogenesis

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Benoit eBarbeau

2013-08-01

Full Text Available The production of antisense transcripts from the 3’ long terminal repeat (LTR in human T-lymphotropic retroviruses has now been clearly demonstrated. After the identification of the antisense strand-encoded HTLV-1 bZIP (HBZ factor, we reported that HBZ could interact with CREB transcription factors and consequently turn off the important activating potential of the viral Tax protein on HTLV-1 5’ LTR promoter activity. We have recently accumulated new results demonstrating that antisense transcripts also exist in HTLV-2, -3 and -4. Furthermore, our data have confirmed the existence of encoded proteins from these antisense transcripts (termed antisense proteins of HTLVs or APHs. APHs are also involved in the down-regulation of Tax-dependent viral transcription. In this review, we will focus on the different molecular mechanisms used by HBZ and APH-2 to control viral expression. While HBZ interacts with CREB through its basic zipper domain, APH-2 binds to this cellular factor through a five amino acid motif localized in its carboxyl terminus. Moreover, unlike APH-2, HBZ possesses an N-terminal activation domain that also contributes to the inhibition of the viral transcription by interacting with the KIX domain of p300/CBP. On the other hand, HBZ was found to induce T-cell proliferation while APH-2 was unable to promote such proliferation. Interestingly, HTLV-2 has not been causally linked to human T-cell leukemia, while HTLV-1 is responsible for the development of the Adult T-cell Leukemia/Lymphoma (ATLL. We will further discuss the possible role played by antisense proteins in the establishment of pathologies induced by viral infection.

Identification of herpes simplex virus type 1 proteins encoded within the first 1.5 kb of the latency-associated transcript.

Science.gov (United States)

Henderson, Gail; Jaber, Tareq; Carpenter, Dale; Wechsler, Steven L; Jones, Clinton

2009-09-01

Expression of the first 1.5 kb of the latency-associated transcript (LAT) that is encoded by herpes simplex virus type 1 (HSV-1) is sufficient for wild-type (wt) levels of reactivation from latency in small animal models. Peptide-specific immunoglobulin G (IgG) was generated against open reading frames (ORFs) that are located within the first 1.5 kb of LAT coding sequences. Cells stably transfected with LAT or trigeminal ganglionic neurons of mice infected with a LAT expressing virus appeared to express the L2 or L8 ORF. Only L2 ORF expression was readily detected in trigeminal ganglionic neurons of latently infected mice.

Transcriptional regulation and DNA methylation in plastids during transitional conversion of chloroplasts to chromoplasts.

Science.gov (United States)

Kobayashi, H; Ngernprasirtsiri, J; Akazawa, T

1990-01-01

During transitional conversion of chloroplasts to chromoplasts in ripening tomato (Lycopersicon esculentum) fruits, transcripts for several plastid genes for photosynthesis decreased to undetectable levels. Run-on transcription of plastids indicated that transcriptional regulation operated as a predominant factor. We found that most of the genes in chloroplasts were actively transcribed in vitro by Escherichia coli and soluble plastid RNA polymerases, but some genes in chromoplasts seemed to be silent when assayed by the in vitro systems. The regulatory step, therefore, was ascribed to DNA templates. The analysis of modified base composition revealed the presence of methylated bases in chromoplast DNA, in which 5-methylcytosine was most abundant. The presence of 5-methylcytosine detected by isoschizomeric endonucleases and Southern hybridization was correlated with the undetectable transcription activity of each gene in the run-on assay and in vitro transcription experiments. It is thus concluded that the suppression of transcription mediated by DNA methylation is one of the mechanisms governing gene expression in plastids converting from chloroplasts to chromoplasts. Images Fig. 1 Fig. 2 Fig. 3. Fig. 4. Fig. 5. PMID:2303026

Transcription of minute virus of mice, an autonomous parvovirus, may be regulated by attenuation

International Nuclear Information System (INIS)

Ben-Asher, E.; Aloni, Y.

1984-01-01

To characterize the transcriptional organization and regulation of minute virus of mice, an autonomous parvovirus, viral transcriptional complexes were isolated and cleaved with restriction enzymes. The in vivo preinitiated nascent RNA was elongated in vitro in the presence of [alpha- 32 P]UTP to generate runoff transcripts. The lengths of the runoff transcripts were analyzed by gel electrophoresis under denaturing conditions. On the basis of the map locations of the restriction sites and the lengths of the runoff transcripts, the in vivo initiation sites were determined. Two major initiation sites having similar activities were thus identified at residues 201 +/- 5 and 2005 +/- 5; both of them were preceded by a TATAA sequence. When uncleaved viral transcriptional complexes or isolated nuclei were incubated in vitro in the presence of [alpha- 32 P]UTP or [alpha- 32 P]CTP, they synthesized labeled RNA that, as determined by polyacrylamide gel electrophoresis, contained a major band of 142 nucleotides. The RNA of the major band was mapped between the initiation site at residue 201 +/- 5 and residue 342. We noticed the potential of forming two mutually exclusive stem-and-loop structures in the 142-nucleotide RNA; one of them is followed by a string of uridylic acid residues typical of a procaryotic transcription termination signal. We propose that, as in the transcription of simian virus 40, RNA transcription in minute virus of mice may be regulated by attenuation and may involve eucaryotic polymerase B, which can respond to a transcription termination signal similar to that of the procaryotic polymerase

Transcripts of mobile element MDG1 during ontogenesis of Drosophila melanogaster

International Nuclear Information System (INIS)

Kuvakina, A.I.; Nurminskii, D.I.; Kogan, G.L.; Gvozdev, V.A.

1989-01-01

It has been demonstrated by Northern hybridization using a single-stranded labeled probes that the number of MDG1 transcripts as well as their size change during ontogenesis of Drosophila. The transcripts of MDG1 were not found in unfertilized eggs. The full-length transcript of MDG1 (about 7 kb long) appears in the embryonic and larval cells, and its quantity sharply increases in pupae and adults. A transcript of about 5 kb length is also found in the pupae and adults. Another, about 2 kb long transcript forms in the embryos, pupae and adults, which is absent in larvae. The main transcript in the larval cells, complementary to the inner part of the body of MDG1, is about 1 kb long. The transcription level of MDG1 and the mobile element copia do not change under heat shock at adult stage

WRKY transcription factors

Science.gov (United States)

Bakshi, Madhunita; Oelmüller, Ralf

2014-01-01

WRKY transcription factors are one of the largest families of transcriptional regulators found exclusively in plants. They have diverse biological functions in plant disease resistance, abiotic stress responses, nutrient deprivation, senescence, seed and trichome development, embryogenesis, as well as additional developmental and hormone-controlled processes. WRKYs can act as transcriptional activators or repressors, in various homo- and heterodimer combinations. Here we review recent progress on the function of WRKY transcription factors in Arabidopsis and other plant species such as rice, potato, and parsley, with a special focus on abiotic, developmental, and hormone-regulated processes. PMID:24492469

Widespread anti-sense transcription in apple is correlated with siRNA production and indicates a large potential for transcriptional and/or post-transcriptional control.

Science.gov (United States)

Celton, Jean-Marc; Gaillard, Sylvain; Bruneau, Maryline; Pelletier, Sandra; Aubourg, Sébastien; Martin-Magniette, Marie-Laure; Navarro, Lionel; Laurens, François; Renou, Jean-Pierre

2014-07-01

Characterizing the transcriptome of eukaryotic organisms is essential for studying gene regulation and its impact on phenotype. The realization that anti-sense (AS) and noncoding RNA transcription is pervasive in many genomes has emphasized our limited understanding of gene transcription and post-transcriptional regulation. Numerous mechanisms including convergent transcription, anti-correlated expression of sense and AS transcripts, and RNAi remain ill-defined. Here, we have combined microarray analysis and high-throughput sequencing of small RNAs (sRNAs) to unravel the complexity of transcriptional and potential post-transcriptional regulation in eight organs of apple (Malus × domestica). The percentage of AS transcript expression is higher than that identified in annual plants such as rice and Arabidopsis thaliana. Furthermore, we show that a majority of AS transcripts are transcribed beyond 3'UTR regions, and may cover a significant portion of the predicted sense transcripts. Finally we demonstrate at a genome-wide scale that anti-sense transcript expression is correlated with the presence of both short (21-23 nt) and long (> 30 nt) siRNAs, and that the sRNA coverage depth varies with the level of AS transcript expression. Our study provides a new insight on the functional role of anti-sense transcripts at the genome-wide level, and a new basis for the understanding of sRNA biogenesis in plants. © 2014 INRA. New Phytologist © 2014 New Phytologist Trust.

Requirements for DNA strand transfer during reverse transcription in mutant HIV-1 virions

NARCIS (Netherlands)

Berkhout, B.; van Wamel, J.; Klaver, B.

1995-01-01

Retroviruses convert their RNA genome into a DNA form by means of reverse transcription. According to the current model of reverse transcription, two strand transfer reactions are needed to synthesize a full-length DNA genome. Because reverse transcription is initiated close to the 5' end of the RNA

Specific transcripts are elevated in Saccharomyces cerevisiae in response to DNA damage

International Nuclear Information System (INIS)

McClanahan, T.; McEntee, K.

1984-01-01

Differential hybridization has been used to identify genes in Saccharomyces cerevisiae displaying increased transcript levels after treatment of cells with UV irradiation or with the mutagen/carcinogen 4-nitroquinoline-1-oxide (NQO). The authors describe the isolation and characterization of four DNA damage responsive genes obtained from screening ca. 9000 yeast genomic clones. Two of these clones, lambda 78A and pBR178C, contain repetitive elements in the yeast genome as shown by Southern hybridization analysis. Although the genomic hybridization pattern is distinct for each of these two clones, both of these sequences hybridize to large polyadenylated transcripts ca. 5 kilobases in length. Two other DNA damage responsive sequences, pBRA2 and pBR3016B, are single-copy genes and hybridize to 0.5- and 3.2-kilobase transcripts, respectively. Kinetic analysis of the 0.5-kilobase transcript homologous to pBRA2 indicates that the level of this RNA increases more than 15-fold within 20 min after exposure to 4-nitroquinoline-1-oxide. Moreover, the level of this transcript is significantly elevated in cells containing the rad52-1 mutation which are deficient in DNA strand break repair and gene conversion. These results provide some of the first evidence that DNA damage stimulates transcription of specific genes in eucaryotic cells

Frequency of BCR-ABL Transcript Types in Syrian CML Patients

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Sulaf Farhat-Maghribi

2016-01-01

Full Text Available Background. In Syria, CML patients are started on tyrosine kinase inhibitors (TKIs and monitored until complete molecular response is achieved. BCR-ABL mRNA transcript type is not routinely identified, contrary to the recommendations. In this study we aimed to identify the frequency of different BCR-ABL transcripts in Syrian CML patients and highlight their significance on monitoring and treatment protocols. Methods. CML patients positive for BCR-ABL transcripts by quantitative RT-PCR were enrolled. BCR-ABL transcript types were investigated using a home-made PCR method that was adapted from published protocols and optimized. The transcript types were then confirmed using a commercially available research kit. Results. Twenty-four transcripts were found in 21 patients. The most common was b2a2, followed by b3a2, b3a3, and e1a3 present solely in 12 (57.1%, 3 (14.3%, 2 (9.5%, and 1 (4.8%, respectively. Three samples (14.3% contained dual transcripts. While b3a2 transcript was apparently associated with warning molecular response to imatinib treatment, b2a2, b3a3, and e1a3 transcripts collectively proved otherwise (P=0.047. Conclusion. It might be advisable to identify the BCR-ABL transcript type in CML patients at diagnosis, using an empirically verified method, in order to link the detected transcript with the clinical findings, possible resistance to treatment, and appropriate monitoring methods.

Expression of the potential therapeutic target CXXC5 in primary acute myeloid leukemia cells - high expression is associated with adverse prognosis as well as altered intracellular signaling and transcriptional regulation.

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Bruserud, Øystein; Reikvam, Håkon; Fredly, Hanne; Skavland, Jørn; Hagen, Karen-Marie; van Hoang, Tuyen Thy; Brenner, Annette K; Kadi, Amir; Astori, Audrey; Gjertsen, Bjørn Tore; Pendino, Frederic

2015-02-20

The CXXC5 gene encodes a transcriptional activator with a zinc-finger domain, and high expression in human acute myeloid leukemia (AML) cells is associated with adverse prognosis. We now characterized the biological context of CXXC5 expression in primary human AML cells. The global gene expression profile of AML cells derived from 48 consecutive patients was analyzed; cells with high and low CXXC5 expression then showed major differences with regard to extracellular communication and intracellular signaling. We observed significant differences in the phosphorylation status of several intracellular signaling mediators (CREB, PDK1, SRC, STAT1, p38, STAT3, rpS6) that are important for PI3K-Akt-mTOR signaling and/or transcriptional regulation. High CXXC5 expression was also associated with high mRNA expression of several stem cell-associated transcriptional regulators, the strongest associations being with WT1, GATA2, RUNX1, LYL1, DNMT3, SPI1, and MYB. Finally, CXXC5 knockdown in human AML cell lines caused significantly increased expression of the potential tumor suppressor gene TSC22 and genes encoding the growth factor receptor KIT, the cytokine Angiopoietin 1 and the selenium-containing glycoprotein Selenoprotein P. Thus, high CXXC5 expression seems to affect several steps in human leukemogenesis, including intracellular events as well as extracellular communication.

Transcriptional regulation of long-term memory in the marine snail Aplysia

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Lee Yong-Seok

2008-06-01

Full Text Available Abstract Whereas the induction of short-term memory involves only covalent modifications of constitutively expressed preexisting proteins, the formation of long-term memory requires gene expression, new RNA, and new protein synthesis. On the cellular level, transcriptional regulation is thought to be the starting point for a series of molecular steps necessary for both the initiation and maintenance of long-term synaptic facilitation (LTF. The core molecular features of transcriptional regulation involved in the long-term process are evolutionally conserved in Aplysia, Drosophila, and mouse, and indicate that gene regulation by the cyclic AMP response element binding protein (CREB acting in conjunction with different combinations of transcriptional factors is critical for the expression of many forms of long-term memory. In the marine snail Aplysia, the molecular mechanisms that underlie the storage of long-term memory have been extensively studied in the monosynaptic connections between identified sensory neuron and motor neurons of the gill-withdrawal reflex. One tail shock or one pulse of serotonin (5-HT, a modulatory transmitter released by tail shocks, produces a transient facilitation mediated by the cAMP-dependent protein kinase leading to covalent modifications in the sensory neurons that results in an enhancement of transmitter release and a strengthening of synaptic connections lasting minutes. By contrast, repeated pulses of 5-hydroxytryptamine (5-HT induce a transcription- and translation-dependent long-term facilitation (LTF lasting more than 24 h and trigger the activation of a family of transcription factors in the presynaptic sensory neurons including ApCREB1, ApCREB2 and ApC/EBP. In addition, we have recently identified novel transcription factors that modulate the expression of ApC/EBP and also are critically involved in LTF. In this review, we examine the roles of these transcription factors during consolidation of LTF induced

Transcriptional regulation of long-term memory in the marine snail Aplysia.

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Lee, Yong-Seok; Bailey, Craig H; Kandel, Eric R; Kaang, Bong-Kiun

2008-06-17

Whereas the induction of short-term memory involves only covalent modifications of constitutively expressed preexisting proteins, the formation of long-term memory requires gene expression, new RNA, and new protein synthesis. On the cellular level, transcriptional regulation is thought to be the starting point for a series of molecular steps necessary for both the initiation and maintenance of long-term synaptic facilitation (LTF). The core molecular features of transcriptional regulation involved in the long-term process are evolutionally conserved in Aplysia, Drosophila, and mouse, and indicate that gene regulation by the cyclic AMP response element binding protein (CREB) acting in conjunction with different combinations of transcriptional factors is critical for the expression of many forms of long-term memory. In the marine snail Aplysia, the molecular mechanisms that underlie the storage of long-term memory have been extensively studied in the monosynaptic connections between identified sensory neuron and motor neurons of the gill-withdrawal reflex. One tail shock or one pulse of serotonin (5-HT), a modulatory transmitter released by tail shocks, produces a transient facilitation mediated by the cAMP-dependent protein kinase leading to covalent modifications in the sensory neurons that results in an enhancement of transmitter release and a strengthening of synaptic connections lasting minutes. By contrast, repeated pulses of 5-hydroxytryptamine (5-HT) induce a transcription- and translation-dependent long-term facilitation (LTF) lasting more than 24 h and trigger the activation of a family of transcription factors in the presynaptic sensory neurons including ApCREB1, ApCREB2 and ApC/EBP. In addition, we have recently identified novel transcription factors that modulate the expression of ApC/EBP and also are critically involved in LTF. In this review, we examine the roles of these transcription factors during consolidation of LTF induced by different

Transcriptional Programs Controlling Perinatal Lung Maturation

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Xu, Yan; Wang, Yanhua; Besnard, Valérie; Ikegami, Machiko; Wert, Susan E.; Heffner, Caleb; Murray, Stephen A.; Donahue, Leah Rae; Whitsett, Jeffrey A.

2012-01-01

The timing of lung maturation is controlled precisely by complex genetic and cellular programs. Lung immaturity following preterm birth frequently results in Respiratory Distress Syndrome (RDS) and Broncho-Pulmonary Dysplasia (BPD), which are leading causes of mortality and morbidity in preterm infants. Mechanisms synchronizing gestational length and lung maturation remain to be elucidated. In this study, we designed a genome-wide mRNA expression time-course study from E15.5 to Postnatal Day 0 (PN0) using lung RNAs from C57BL/6J (B6) and A/J mice that differ in gestational length by ∼30 hr (B6controlling lung maturation. We identified both temporal and strain dependent gene expression patterns during lung maturation. For time dependent changes, cell adhesion, vasculature development, and lipid metabolism/transport were major bioprocesses induced during the saccular stage of lung development at E16.5–E17.5. CEBPA, PPARG, VEGFA, CAV1 and CDH1 were found to be key signaling and transcriptional regulators of these processes. Innate defense/immune responses were induced at later gestational ages (E18.5–20.5), STAT1, AP1, and EGFR being important regulators of these responses. Expression of RNAs associated with the cell cycle and chromatin assembly was repressed during prenatal lung maturation and was regulated by FOXM1, PLK1, chromobox, and high mobility group families of transcription factors. Strain dependent lung mRNA expression differences peaked at E18.5. At this time, mRNAs regulating surfactant and innate immunity were more abundantly expressed in lungs of B6 (short gestation) than in A/J (long gestation) mice, while expression of genes involved in chromatin assembly and histone modification were expressed at lower levels in B6 than in A/J mice. The present study systemically mapped key regulators, bioprocesses, and transcriptional networks controlling lung maturation, providing the basis for new therapeutic strategies to enhance lung function in preterm

SEASTAR: systematic evaluation of alternative transcription start sites in RNA.

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Qin, Zhiyi; Stoilov, Peter; Zhang, Xuegong; Xing, Yi

2018-05-04

Alternative first exons diversify the transcriptomes of eukaryotes by producing variants of the 5' Untranslated Regions (5'UTRs) and N-terminal coding sequences. Accurate transcriptome-wide detection of alternative first exons typically requires specialized experimental approaches that are designed to identify the 5' ends of transcripts. We developed a computational pipeline SEASTAR that identifies first exons from RNA-seq data alone then quantifies and compares alternative first exon usage across multiple biological conditions. The exons inferred by SEASTAR coincide with transcription start sites identified directly by CAGE experiments and bear epigenetic hallmarks of active promoters. To determine if differential usage of alternative first exons can yield insights into the mechanism controlling gene expression, we applied SEASTAR to an RNA-seq dataset that tracked the reprogramming of mouse fibroblasts into induced pluripotent stem cells. We observed dynamic temporal changes in the usage of alternative first exons, along with correlated changes in transcription factor expression. Using a combined sequence motif and gene set enrichment analysis we identified N-Myc as a regulator of alternative first exon usage in the pluripotent state. Our results demonstrate that SEASTAR can leverage the available RNA-seq data to gain insights into the control of gene expression and alternative transcript variation in eukaryotic transcriptomes.

Dual Regulation of Bacillus subtilis kinB Gene Encoding a Sporulation Trigger by SinR through Transcription Repression and Positive Stringent Transcription Control.

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Fujita, Yasutaro; Ogura, Mitsuo; Nii, Satomi; Hirooka, Kazutake

2017-01-01

It is known that transcription of kinB encoding a trigger for Bacillus subtilis sporulation is under repression by SinR, a master repressor of biofilm formation, and under positive stringent transcription control depending on the adenine species at the transcription initiation nucleotide (nt). Deletion and base substitution analyses of the kinB promoter (P kinB ) region using lacZ fusions indicated that either a 5-nt deletion (Δ5, nt -61/-57, +1 is the transcription initiation nt) or the substitution of G at nt -45 with A (G-45A) relieved kinB repression. Thus, we found a pair of SinR-binding consensus sequences (GTTCTYT; Y is T or C) in an inverted orientation (SinR-1) between nt -57/-42, which is most likely a SinR-binding site for kinB repression. This relief from SinR repression likely requires SinI, an antagonist of SinR. Surprisingly, we found that SinR is essential for positive stringent transcription control of P kinB . Electrophoretic mobility shift assay (EMSA) analysis indicated that SinR bound not only to SinR-1 but also to SinR-2 (nt -29/-8) consisting of another pair of SinR consensus sequences in a tandem repeat arrangement; the two sequences partially overlap the '-35' and '-10' regions of P kinB . Introduction of base substitutions (T-27C C-26T) in the upstream consensus sequence of SinR-2 affected positive stringent transcription control of P kinB , suggesting that SinR binding to SinR-2 likely causes this positive control. EMSA also implied that RNA polymerase and SinR are possibly bound together to SinR-2 to form a transcription initiation complex for kinB transcription. Thus, it was suggested in this work that derepression of kinB from SinR repression by SinI induced by Spo0A∼P and occurrence of SinR-dependent positive stringent transcription control of kinB might induce effective sporulation cooperatively, implying an intimate interplay by stringent response, sporulation, and biofilm formation.

The bZIP Transcription Factor HAC-1 Is Involved in the Unfolded Protein Response and Is Necessary for Growth on Cellulose in Neurospora crassa.

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Alejandro Montenegro-Montero

Full Text Available High protein secretion capacity in filamentous fungi requires an extremely efficient system for protein synthesis, folding and transport. When the folding capacity of the endoplasmic reticulum (ER is exceeded, a pathway known as the unfolded protein response (UPR is triggered, allowing cells to mitigate and cope with this stress. In yeast, this pathway relies on the transcription factor Hac1, which mediates the up-regulation of several genes required under these stressful conditions. In this work, we identified and characterized the ortholog of the yeast HAC1 gene in the filamentous fungus Neurospora crassa. We show that its mRNA undergoes an ER stress-dependent splicing reaction, which in N. crassa removes a 23 nt intron and leads to a change in the open reading frame. By disrupting the N. crassa hac-1 gene, we determined it to be crucial for activating UPR and for proper growth in the presence of ER stress-inducing chemical agents. Neurospora is naturally found growing on dead plant material, composed primarily by lignocellulose, and is a model organism for the study of plant cell wall deconstruction. Notably, we found that growth on cellulose, a substrate that requires secretion of numerous enzymes, imposes major demands on ER function and is dramatically impaired in the absence of hac-1, thus broadening the range of physiological functions of the UPR in filamentous fungi. Growth on hemicellulose however, another carbon source that necessitates the secretion of various enzymes for its deconstruction, is not impaired in the mutant nor is the amount of proteins secreted on this substrate, suggesting that secretion, as a whole, is unaltered in the absence of hac-1. The characterization of this signaling pathway in N. crassa will help in the study of plant cell wall deconstruction by fungi and its manipulation may result in important industrial biotechnological applications.

Dicty_cDB: SHJ838 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available 4 CV164410 |CV164410.1 rsmsxl_003361.y1.scf cDNA Library of Salvia miltiorrhiza S...alvia miltiorrhiza cDNA 5', mRNA sequence. 54 6e-09 3 CV172465 |CV172465.1 rsmsxlre_008717.z1.scf cDNA Libra

Transcription Factor SOX5 Promotes the Migration and Invasion of Fibroblast-Like Synoviocytes in Part by Regulating MMP-9 Expression in Collagen-Induced Arthritis

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Yumeng Shi

2018-04-01

Full Text Available ObjectivesFibroblast-like synoviocytes (FLS exhibit a unique aggressive phenotype in rheumatoid arthritis (RA. Increased FLS migration and subsequent invasion of the extracellular matrix are essential to joint destruction in RA. Our previous research reported that transcription factor SOX5 was highly expressed in RA-FLS. Here, the effects of SOX5 in RA-FLS migration and invasion will be investigated.MethodsThe migration and invasion of RA-FLS were evaluated using a transwell chamber assay. The expression of several potential SOX5-targeted genes, including matrix metalloproteinases (MMP-1, 2, 3 and 9, chemokines (CCL4, CCL2, CCR5 and CCR2, and pro-inflammatory cytokines (TNF-α and IL-6, were examined in RA-FLS using SOX5 gain- and loss-of-function study. The molecular mechanisms of SOX5-mediated MMP-9 expressions were assayed by luciferase reporter gene and chromatin immunoprecipitation (ChIP studies. The in vivo effect of SOX5 on FLS migration and invasion was examined using collagen-induced arthritis (CIA in DBA/1J mice.ResultsKnockdown SOX5 decreased lamellipodium formation, migration, and invasion of RA-FLS. The expression of MMP-9 was the only gene tested to be concomitantly affected by silencing or overexpressing SOX5. ChIP assay revealed that SOX5 was bound to the MMP-9 promoter in RA-FLS. The overexpression of SOX5 markedly enhanced the MMP-9 promoter activity, and specific deletion of a putative SOX5-binding site in MMP-9 promoter diminished this promoter-driven transcription in FLS. Locally knocked down SOX5 inhibited MMP-9 expression in the joint tissue and reduced pannus migration and invasion into the cartilage in CIA mice.ConclusionSOX5 plays a novel role in mediating migration and invasion of FLS in part by regulating MMP-9 expression in RA.

Transcription Factor SOX5 Promotes the Migration and Invasion of Fibroblast-Like Synoviocytes in Part by Regulating MMP-9 Expression in Collagen-Induced Arthritis.

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Shi, Yumeng; Wu, Qin; Xuan, Wenhua; Feng, Xiaoke; Wang, Fang; Tsao, Betty P; Zhang, Miaojia; Tan, Wenfeng

2018-01-01

Fibroblast-like synoviocytes (FLS) exhibit a unique aggressive phenotype in rheumatoid arthritis (RA). Increased FLS migration and subsequent invasion of the extracellular matrix are essential to joint destruction in RA. Our previous research reported that transcription factor SOX5 was highly expressed in RA-FLS. Here, the effects of SOX5 in RA-FLS migration and invasion will be investigated. The migration and invasion of RA-FLS were evaluated using a transwell chamber assay. The expression of several potential SOX5-targeted genes, including matrix metalloproteinases (MMP-1, 2, 3 and 9), chemokines (CCL4, CCL2, CCR5 and CCR2), and pro-inflammatory cytokines (TNF-α and IL-6), were examined in RA-FLS using SOX5 gain- and loss-of-function study. The molecular mechanisms of SOX5-mediated MMP-9 expressions were assayed by luciferase reporter gene and chromatin immunoprecipitation (ChIP) studies. The in vivo effect of SOX5 on FLS migration and invasion was examined using collagen-induced arthritis (CIA) in DBA/1J mice. Knockdown SOX5 decreased lamellipodium formation, migration, and invasion of RA-FLS. The expression of MMP-9 was the only gene tested to be concomitantly affected by silencing or overexpressing SOX5. ChIP assay revealed that SOX5 was bound to the MMP-9 promoter in RA-FLS. The overexpression of SOX5 markedly enhanced the MMP-9 promoter activity, and specific deletion of a putative SOX5-binding site in MMP-9 promoter diminished this promoter-driven transcription in FLS. Locally knocked down SOX5 inhibited MMP-9 expression in the joint tissue and reduced pannus migration and invasion into the cartilage in CIA mice. SOX5 plays a novel role in mediating migration and invasion of FLS in part by regulating MMP-9 expression in RA.

Prognostic significance of signal transducer and activator of transcription 5 and 5b expression in Epstein-Barr virus-positive patients with chronic lymphocytic leukemia.

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Diamantopoulos, Panagiotis T; Sofotasiou, Maria; Georgoussi, Zafiroula; Giannakopoulou, Nefeli; Papadopoulou, Vasiliki; Galanopoulos, Athanasios; Kontandreopoulou, Elina; Zervakis, Panagiotis; Pallaki, Paschalina; Kalala, Fani; Kyrtsonis, Marie-Christine; Dimitrakopoulou, Aglaia; Vassilakopoulos, Theodoros; Angelopoulou, Maria; Spanakis, Nikolaos; Viniou, Nora-Athina

2016-09-01

Signal transducer and activator of transcription (STAT) proteins have been intensively studied in hematologic malignancies, and the efficacy of agents against STATs in lymphomas is already under research. We investigated the expression of total STAT5 and STAT5b in peripheral blood samples of patients with chronic lymphocytic leukemia (CLL) in correlation with the presence of Epstein-Barr Virus (EBV) and its major oncoprotein (latent membrane protein 1, LMP1). The EBV load was measured in the peripheral blood by real-time PCR for the BXLF1 gene and the levels of LMP1 by PCR and ELISA. Western blotting was performed for total STAT5 and STAT5b in protein extracts. STAT5b was only expressed in patients (not in healthy subjects) and STAT5 but particularly STAT5b expression was correlated with the presence of the virus (77.3% vs. 51.2%, P = 0.006 for STAT5b) and to the expression of LMP1 (58.3% vs. 21.6%, P = 0.011 for STAT5b). Moreover, the expression of STAT5b and the presence of EBV and LMP1 were strongly negatively correlated with the overall survival of the patients (log-rank test P = 0.011, 0.015, 0.006, respectively). Double positive (for EBV and STAT5b) patients had the lowest overall survival (log-rank test P = 0.013). This is the first report of a survival disadvantage of EBV+ patients with CLL, and the first time that STAT5b expression is correlated with survival. The correlation of STAT5 expression with the presence of the virus, along with our survival correlations defines a subgroup of patients with CLL that may benefit from anti-STAT agents. © 2016 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

Grr1p is required for transcriptional induction of amino acid permease genes and proper transcriptional regulation of genes in carbon metabolism of Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Eckert-Boulet, Nadine; Regenberg, Birgitte; Nielsen, Jens

2005-01-01

and a grr1 Delta strain and adding citrulline in the exponential phase. Whole-genome transcription analyses were performed on samples from each cultivation, both immediately before and 30 min after citrulline addition. Transcriptional induction of the AAP genes AGP1, BAP2, BAP3, DIP5, GNP1 and TAT1 is fully...

DNA Topoisomerases Maintain Promoters in a State Competent for Transcriptional Activation in Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Pedersen, Jakob Madsen; Fredsøe, Jacob Christian; Rødgaard, Morten Terpager

2012-01-01

To investigate the role of DNA topoisomerases in transcription, we have studied global gene expression in Saccharomyces cerevisiae cells deficient for topoisomerases I and II and performed single-gene analyses to support our findings. The genome-wide studies show a general transcriptional down......-regulation upon lack of the enzymes, which correlates with gene activity but not gene length. Furthermore, our data reveal a distinct subclass of genes with a strong requirement for topoisomerases. These genes are characterized by high transcriptional plasticity, chromatin regulation, TATA box presence......-depth analysis of the inducible PHO5 gene reveals that topoisomerases are essential for binding of the Pho4p transcription factor to the PHO5 promoter, which is required for promoter nucleosome removal during activation. In contrast, topoisomerases are dispensable for constitutive transcription initiation...

Transcriptional network systems in cartilage development and disease.

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Nishimura, Riko; Hata, Kenji; Nakamura, Eriko; Murakami, Tomohiko; Takahata, Yoshifumi

2018-04-01

Transcription factors play important roles in the regulation of cartilage development by controlling the expression of chondrogenic genes. Genetic studies have revealed that Sox9/Sox5/Sox6, Runx2/Runx3 and Osterix in particular are essential for the sequential steps of cartilage development. Importantly, these transcription factors form network systems that are also required for appropriate cartilage development. Molecular cloning approaches have largely contributed to the identification of several transcriptional partners for Sox9 and Runx2 during cartilage development. Although the importance of a negative-feedback loop between Indian hedgehog (Ihh) and parathyroid hormone-related protein (PTHrP) in chondrocyte hypertrophy has been well established, recent studies indicate that several transcription factors interact with the Ihh-PTHrP loop and demonstrated that Ihh has multiple functions in the regulation of cartilage development. The most common cartilage disorder, osteoarthritis, has been reported to result from the pathological action of several transcription factors, including Runx2, C/EBPβ and HIF-2α. On the other hand, NFAT family members appear to play roles in the protection of cartilage from osteoarthritis. It is also becoming important to understand the homeostasis and regulation of articular chondrocytes, because they have different cellular and molecular features from chondrocytes of the growth plate. This review summarizes the regulation and roles of transcriptional network systems in cartilage development and their pathological roles in osteoarthritis.

Dynamic usage of transcription start sites within core promoters

DEFF Research Database (Denmark)

Kawaji, Hideya; Frith, Martin C; Katayama, Shintaro

2006-01-01

BACKGROUND: Mammalian promoters do not initiate transcription at single, well defined base pairs, but rather at multiple, alternative start sites spread across a region. We previously characterized the static structures of transcription start site usage within promoters at the base pair level......, based on large-scale sequencing of transcript 5' ends. RESULTS: In the present study we begin to explore the internal dynamics of mammalian promoters, and demonstrate that start site selection within many mouse core promoters varies among tissues. We also show that this dynamic usage of start sites...... is associated with CpG islands, broad and multimodal promoter structures, and imprinting. CONCLUSION: Our results reveal a new level of biologic complexity within promoters--fine-scale regulation of transcription starting events at the base pair level. These events are likely to be related to epigenetic...

Regulation of expression of two LY-6 family genes by intron retention and transcription induced chimerism

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Mallya Meera

2008-09-01

Full Text Available Abstract Background Regulation of the expression of particular genes can rely on mechanisms that are different from classical transcriptional and translational control. The LY6G5B and LY6G6D genes encode LY-6 domain proteins, whose expression seems to be regulated in an original fashion, consisting of an intron retention event which generates, through an early premature stop codon, a non-coding transcript, preventing expression in most cell lines and tissues. Results The MHC LY-6 non-coding transcripts have shown to be stable and very abundant in the cell, and not subject to Nonsense Mediated Decay (NMD. This retention event appears not to be solely dependent on intron features, because in the case of LY6G5B, when the intron is inserted in the artificial context of a luciferase expression plasmid, it is fully spliced but strongly stabilises the resulting luciferase transcript. In addition, by quantitative PCR we found that the retained and spliced forms are differentially expressed in tissues indicating an active regulation of the non-coding transcript. EST database analysis revealed that these genes have an alternative expression pathway with the formation of Transcription Induced Chimeras (TIC. This data was confirmed by RT-PCR, revealing the presence of different transcripts that would encode the chimeric proteins CSNKβ-LY6G5B and G6F-LY6G6D, in which the LY-6 domain would join to a kinase domain and an Ig-like domain, respectively. Conclusion In conclusion, the LY6G5B and LY6G6D intron-retained transcripts are not subjected to NMD and are more abundant than the properly spliced forms. In addition, these genes form chimeric transcripts with their neighbouring same orientation 5' genes. Of interest is the fact that the 5' genes (CSNKβ or G6F undergo differential splicing only in the context of the chimera (CSNKβ-LY6G5B or G6F-LY6G6C and not on their own.

Luminescent and scintillation properties of Bi{sup 3+} doped Y{sub 2}SiO{sub 5} and Lu{sub 2}SiO{sub 5} single crystalline films

Energy Technology Data Exchange (ETDEWEB)

Zorenko, Yu., E-mail: zorenko@ukw.edu.pl [Institute of Physics, Kazimierz Wielki University in Bydgoszcz, 85-090 Bydgoszcz (Poland); Gorbenko, V.; Zorenko, T. [Institute of Physics, Kazimierz Wielki University in Bydgoszcz, 85-090 Bydgoszcz (Poland); Laboratory for Optoelectronic Materials (LOM), Department of Electronics of Ivan Franko National University of Lviv, 79017 Lviv (Ukraine); Malinowski, P. [Institute of Physics, Kazimierz Wielki University in Bydgoszcz, 85-090 Bydgoszcz (Poland); Jary, V.; Kucerkova, R.; Beitlerova, A.; Mares, J.A.; Nikl, M. [Institute of Physics AS CR, Cukrovarnicka 10, 16253 Prague (Czech Republic); Fedorov, A. [Institute for Single Crystals NAS of Ukraine, 60 Lenin ave., 61001 Kharkiv (Ukraine)

2014-10-15

In this paper we report our follow-up research on the Bi{sup 3+} luminescence in orthosilicate compounds, focusing on absorption, luminescent and scintillation properties of YSO:Bi and LSO:Bi SCFs with the Bi concentration ranging from 0.05 to 0.18 at%. For purpose of this research, single crystalline films (SCF) of Y{sub 2}SiO{sub 5}:Bi and Lu{sub 2}SiO{sub 5}:Bi have been grown by the LPE method onto YSO and LSO substrates from the melt-solution based on Bi{sub 2}O{sub 3} flux. - Highlights: • YSO:Bi and LSO:Bi films have been grown by liquid phase epitaxy. • Bi{sup 3+} absorption and luminescence depends on Bi concentration. • Scintillation properties of YSO:Bi and LSO:Bi films have been studied.

A new method for the construction of a mutant library with a predictable occurrence rate using Poisson distribution.

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Seong, Ki Moon; Park, Hweon; Kim, Seong Jung; Ha, Hyo Nam; Lee, Jae Yung; Kim, Joon

2007-06-01

A yeast transcriptional activator, Gcn4p, induces the expression of genes that are involved in amino acid and purine biosynthetic pathways under amino acid starvation. Gcn4p has an acidic activation domain in the central region and a bZIP domain in the C-terminus that is divided into the DNA-binding motif and dimerization leucine zipper motif. In order to identify amino acids in the DNA-binding motif of Gcn4p which are involved in transcriptional activation, we constructed mutant libraries in the DNA-binding motif through an innovative application of random mutagenesis. Mutant library made by oligonucleotides which were mutated randomly using the Poisson distribution showed that the actual mutation frequency was in good agreement with expected values. This method could save the time and effort to create a mutant library with a predictable mutation frequency. Based on the studies using the mutant libraries constructed by the new method, the specific residues of the DNA-binding domain in Gcn4p appear to be involved in the transcriptional activities on a conserved binding site.

A Tale of Two Transcriptions. Machine-Assisted Transcription of Historical Sources

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Gunnar Thorvaldsen

2015-01-01

Full Text Available This article explains how two projects implement semi-automated transcription routines: for census sheets in Norway and marriage protocols from Barcelona. The Spanish system was created to transcribe the marriage license books from 1451 to 1905 for the Barcelona area; one of the world’s longest series of preserved vital records. Thus, in the Project “Five Centuries of Marriages” (5CofM at the Autonomous University of Barcelona’s Center for Demographic Studies, the Barcelona Historical Marriage Database has been built. More than 600,000 records were transcribed by 150 transcribers working online. The Norwegian material is cross-sectional as it is the 1891 census, recorded on one sheet per person. This format and the underlining of keywords for several variables made it more feasible to semi-automate data entry than when many persons are listed on the same page. While Optical Character Recognition (OCR for printed text is scientifically mature, computer vision research is now focused on more difficult problems such as handwriting recognition. In the marriage project, document analysis methods have been proposed to automatically recognize the marriage licenses. Fully automatic recognition is still a challenge, but some promising results have been obtained. In Spain, Norway and elsewhere the source material is available as scanned pictures on the Internet, opening up the possibility for further international cooperation concerning automating the transcription of historic source materials. Like what is being done in projects to digitize printed materials, the optimal solution is likely to be a combination of manual transcription and machine-assisted recognition also for hand-written sources.

Growth-hormone-induced signal transducer and activator of transcription 5 signaling causes gigantism, inflammation, and premature death but protects mice from aggressive liver cancer.

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Friedbichler, Katrin; Themanns, Madeleine; Mueller, Kristina M; Schlederer, Michaela; Kornfeld, Jan-Wilhelm; Terracciano, Luigi M; Kozlov, Andrey V; Haindl, Susanne; Kenner, Lukas; Kolbe, Thomas; Mueller, Mathias; Snibson, Kenneth J; Heim, Markus H; Moriggl, Richard

2012-03-01

Persistently high levels of growth hormone (GH) can cause liver cancer. GH activates multiple signal-transduction pathways, among them janus kinase (JAK) 2-signal transducer and activator of transcription (STAT) 5 (signal transducer and activator of transcription 5). Both hyperactivation and deletion of STAT5 in hepatocytes have been implicated in the development of hepatocellular carcinoma (HCC); nevertheless, the role of STAT5 in the development of HCC as a result of high GH levels remains enigmatic. Thus, we crossed a mouse model of gigantism and inflammatory liver cancer caused by hyperactivated GH signaling (GH(tg) ) to mice with hepatic deletion of STAT5 (STAT5(Δhep) ). Unlike GH(tg) mice, GH(tg) STAT5(Δhep) animals did not display gigantism. Moreover, the premature mortality, which was associated with chronic inflammation, as well as the pathologic alterations of hepatocytes observed in GH(tg) mice, were not observed in GH(tg) animals lacking STAT5. Strikingly, loss of hepatic STAT5 proteins led to enhanced HCC development in GH(tg) mice. Despite reduced chronic inflammation, GH(tg) STAT5(Δhep) mice displayed earlier and more advanced HCC than GH(tg) animals. This may be attributed to the combination of increased peripheral lipolysis, hepatic lipid synthesis, loss of hepatoprotective mediators accompanied by aberrant activation of tumor-promoting c-JUN and STAT3 signaling cascades, and accumulation of DNA damage secondary to loss of cell-cycle control. Thus, HCC was never observed in STAT5(Δhep) mice. As a result of their hepatoprotective functions, STAT5 proteins prevent progressive fatty liver disease and the formation of aggressive HCC in the setting of hyperactivated GH signaling. At the same time, they play a key role in controlling systemic inflammation and regulating organ and body size. Copyright © 2011 American Association for the Study of Liver Diseases.

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.

A Next-Generation Sequencing Approach Uncovers Viral Transcripts Incorporated in Poxvirus Virions

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Marica Grossegesse

2017-10-01

Full Text Available Transcripts are known to be incorporated in particles of DNA viruses belonging to the families of Herpesviridae and Mimiviridae, but the presence of transcripts in other DNA viruses, such as poxviruses, has not been analyzed yet. Therefore, we first established a next-generation-sequencing (NGS-based protocol, enabling the unbiased identification of transcripts in virus particles. Subsequently, we applied our protocol to analyze RNA in an emerging zoonotic member of the Poxviridae family, namely Cowpox virus. Our results revealed the incorporation of 19 viral transcripts, while host identifications were restricted to ribosomal and mitochondrial RNA. Most viral transcripts had an unknown and immunomodulatory function, suggesting that transcript incorporation may be beneficial for poxvirus immune evasion. Notably, the most abundant transcript originated from the D5L/I1R gene that encodes a viral inhibitor of the host cytoplasmic DNA sensing machinery.

Characterization of herpes simplex virus 2 primary microRNA Transcript regulation.

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Tang, Shuang; Bosch-Marce, Marta; Patel, Amita; Margolis, Todd P; Krause, Philip R

2015-05-01

In order to understand factors that may influence latency-associated transcription and latency-associated transcript (LAT) phenotypes, we studied the expression of the herpes simplex virus 2 (HSV-2) LAT-associated microRNAs (miRNAs). We mapped the transcription initiation sites of all three primary miRNA transcripts and identified the ICP4-binding sequences at the transcription initiation sites of both HSV-2 LAT (pri-miRNA for miR-I and miR-II, which target ICP34.5, and miR-III, which targets ICP0) and L/ST (a pri-miRNA for miR-I and miR-II) but not at that of the primary miR-H6 (for which the target is unknown). We confirmed activity of the putative HSV-2 L/ST promoter and found that ICP4 trans-activates the L/ST promoter when the ICP4-binding site at its transcription initiation site is mutated, suggesting that ICP4 may play a dual role in regulating transcription of L/ST and, consequently, of miR-I and miR-II. LAT exon 1 (containing LAT enhancer sequences), together with the LAT promoter region, comprises a bidirectional promoter required for the expression of both LAT-encoded miRNAs and miR-H6 in latently infected mouse ganglia. The ability of ICP4 to suppress ICP34.5-targeting miRNAs and to activate lytic viral genes suggests that ICP4 could play a key role in the switch between latency and reactivation. The HSV-2 LAT and viral miRNAs expressed in the LAT region are the most abundant viral transcripts during HSV latency. The balance between the expression of LAT and LAT-associated miRNAs and the expression of lytic viral transcripts from the opposite strand appears to influence whether individual HSV-infected neurons will be latently or productively infected. The outcome of neuronal infection may thus depend on regulation of gene expression of the corresponding primary miRNAs. In the present study, we characterize promoter sequences responsible for miRNA expression, including identification of the primary miRNA 5' ends and evaluation of ICP4 response. These

Gladiolus hybridus ABSCISIC ACID INSENSITIVE 5 (GhABI5) is an important transcription factor in ABA signaling that can enhance Gladiolus corm dormancy and Arabidopsis seed dormancy.

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Wu, Jian; Seng, Shanshan; Sui, Juanjuan; Vonapartis, Eliana; Luo, Xian; Gong, Benhe; Liu, Chen; Wu, Chenyu; Liu, Chao; Zhang, Fengqin; He, Junna; Yi, Mingfang

2015-01-01

The phytohormone abscisic acid (ABA) regulates plant development and is crucial for abiotic stress response. In this study, cold storage contributes to reducing endogenous ABA content, resulting in dormancy breaking of Gladiolus. The ABA inhibitor fluridone also promotes germination, suggesting that ABA is an important hormone that regulates corm dormancy. Here, we report the identification and functional characterization of the Gladiolus ABI5 homolog (GhABI5), which is a basic leucine zipper motif transcriptional factor (TF). GhABI5 is expressed in dormant vegetative organs (corm, cormel, and stolon) as well as in reproductive organs (stamen), and it is up-regulated by ABA or drought. Complementation analysis reveals that GhABI5 rescues the ABA insensitivity of abi5-3 during seed germination and induces the expression of downstream ABA response genes in Arabidopsis thaliana (EM1, EM6, and RD29B). Down-regulation of GhABI5 in dormant cormels via virus induced gene silence promotes sprouting and reduces the expression of downstream genes (GhLEA and GhRD29B). The results of this study reveal that GhABI5 regulates bud dormancy (vegetative organ) in Gladiolus in addition to its well-studied function in Arabidopsis seeds (reproductive organ).

Gladiolus hybridus ABSCISIC ACID INSENSITIVE 5 (GhABI5 is an important transcription factor in ABA signaling that can enhance Gladiolus corm dormancy and Arabidopsis seed dormancy.

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Jian eWu

2015-11-01

Full Text Available The phytohormone abscisic acid (ABA regulates plant development and is crucial for abiotic stress response. In this study, cold storage contributes to reducing endogenous ABA content, resulting in dormancy breaking of Gladiolus. The ABA inhibitor fluridone also promotes germination, suggesting that ABA is an important hormone that regulates corm dormancy. Here, we report the identification and functional characterization of the Gladiolus ABI5 homolog (GhABI5, which is a basic leucine zipper motif transcriptional factor (TF. GhABI5 is expressed in dormant vegetative organs (corm, cormel and stolon as well as in reproductive organs (stamen, and it is up-regulated by ABA or drought. Complementation analysis reveals that GhABI5 rescues the ABA insensitivity of abi5-3 during seed germination and induces the expression of downstream ABA response genes in Arabidopsis thaliana (EM1, EM6 and RD29B. Down-regulation of GhABI5 in dormant cormels via Virus Induced Gene Silence (VIGS promotes sprouting and reduces the expression of downstream genes (GhLEA and GhRD29B. The results of this study reveal that GhABI5 regulates bud dormancy (vegetative organ in Gladiolus in addition to its well-studied function in Arabidopsis seeds (reproductive organ.

Real-time observation of the initiation of RNA polymerase II transcription.

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Fazal, Furqan M; Meng, Cong A; Murakami, Kenji; Kornberg, Roger D; Block, Steven M

2015-09-10

Biochemical and structural studies have shown that the initiation of RNA polymerase II transcription proceeds in the following stages: assembly of the polymerase with general transcription factors and promoter DNA in a 'closed' preinitiation complex (PIC); unwinding of about 15 base pairs of the promoter DNA to form an 'open' complex; scanning downstream to a transcription start site; synthesis of a short transcript, thought to be about 10 nucleotides long; and promoter escape. Here we have assembled a 32-protein, 1.5-megadalton PIC derived from Saccharomyces cerevisiae, and observe subsequent initiation processes in real time with optical tweezers. Contrary to expectation, scanning driven by the transcription factor IIH involved the rapid opening of an extended transcription bubble, averaging 85 base pairs, accompanied by the synthesis of a transcript up to the entire length of the extended bubble, followed by promoter escape. PICs that failed to achieve promoter escape nevertheless formed open complexes and extended bubbles, which collapsed back to closed or open complexes, resulting in repeated futile scanning.

Soluble Fiber Dextrin and Soluble Corn Fiber Supplementation Modify Indices of Health in Cecum and Colon of Sprague-Dawley Rats

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Maria R. C. de Godoy

2013-02-01

Full Text Available The objective of this study was to evaluate health outcomes resulting from dietary supplementation of novel, low-digestible carbohydrates in the cecum and colon of Sprague-Dawley rats randomly assigned to one of four treatment groups for 21 days: 5% cellulose (Control, Pectin, soluble fiber dextrin (SFD, or soluble corn fiber (SCF. Rats fed Pectin had a higher average daily food intake, but no differences in final body weights or rates of weight gain among treatments were observed. No differences were observed in total short-chain fatty acid (SCFA or branched-chain fatty acid (BCFA concentrations in the cecum and colon of rats fed either SFD or SCF. The SFD and SCF treatments increased cecal propionate and decreased butyrate concentrations compared to Control or Pectin. Pectin resulted in increased BCFA in the cecum and colon. Supplementation of SFD and SCF had no effect on cecal microbial populations compared to Control. Consumption of SFD and SCF increased total and empty cecal weight but not colon weight. Gut histomorphology was positively affected by SFD and SCF. Increased crypt depth, goblet cell numbers, and acidic mucin were observed in both the cecum and colon of rats supplemented with SFD, SCF, and Pectin. These novel, low-digestible carbohydrates appear to be beneficial in modulating indices of hindgut morphology when supplemented in the diet of the rat.

Dicty_cDB: CHJ706 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available 09 6 CV164410 |CV164410.1 rsmsxl_003361.y1.scf cDNA Library of Salvia miltiorrhiza Salvia miltiorrhiza cDNA ...5', mRNA sequence. 54 2e-09 3 CV172465 |CV172465.1 rsmsxlre_008717.z1.scf cDNA Library of Salvia miltiorrhiz

Transcriptional activity of Pax3 is co-activated by TAZ

International Nuclear Information System (INIS)

Murakami, Masao; Tominaga, Junji; Makita, Ryosuke; Uchijima, Yasunobu; Kurihara, Yukiko; Nakagawa, Osamu; Asano, Tomoichiro; Kurihara, Hiroki

2006-01-01

Pax3 is a transcription factor which functions in embryonic development and human diseases. In a yeast two-hybrid screen with full-length Pax3 as bait, we isolated a clone encoding transcriptional co-activator with PDZ-binding motif (TAZ) from an E10.5 mouse embryo cDNA library. Co-immunoprecipitation and nuclear co-localization of TAZ with Pax3 suggest that their association is functionally relevant. In situ hybridization revealed TAZ and Pax3 expression to partially overlap in the paraxial mesoderm, limb buds, and the neural tube. In C2C12 myoblast cells and NIH3T3 cells, TAZ enhanced the transcriptional activity of Pax3 on artificial and microphthalmia-associated transcription factor promoter-luciferase constructs, suggesting that TAZ can function as a co-activator of Pax3. Functional interaction between Pax3 and TAZ may provide a clue to clarifying the mechanism by which Pax3 serves as a transcriptional activator during embryogenesis

SoyDB: a knowledge database of soybean transcription factors

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Valliyodan Babu

2010-01-01

Full Text Available Abstract Background Transcription factors play the crucial rule of regulating gene expression and influence almost all biological processes. Systematically identifying and annotating transcription factors can greatly aid further understanding their functions and mechanisms. In this article, we present SoyDB, a user friendly database containing comprehensive knowledge of soybean transcription factors. Description The soybean genome was recently sequenced by the Department of Energy-Joint Genome Institute (DOE-JGI and is publicly available. Mining of this sequence identified 5,671 soybean genes as putative transcription factors. These genes were comprehensively annotated as an aid to the soybean research community. We developed SoyDB - a knowledge database for all the transcription factors in the soybean genome. The database contains protein sequences, predicted tertiary structures, putative DNA binding sites, domains, homologous templates in the Protein Data Bank (PDB, protein family classifications, multiple sequence alignments, consensus protein sequence motifs, web logo of each family, and web links to the soybean transcription factor database PlantTFDB, known EST sequences, and other general protein databases including Swiss-Prot, Gene Ontology, KEGG, EMBL, TAIR, InterPro, SMART, PROSITE, NCBI, and Pfam. The database can be accessed via an interactive and convenient web server, which supports full-text search, PSI-BLAST sequence search, database browsing by protein family, and automatic classification of a new protein sequence into one of 64 annotated transcription factor families by hidden Markov models. Conclusions A comprehensive soybean transcription factor database was constructed and made publicly accessible at http://casp.rnet.missouri.edu/soydb/.

The WRKY57 Transcription Factor Affects the Expression of Jasmonate ZIM-Domain Genes Transcriptionally to Compromise Botrytis cinerea Resistance.

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Jiang, Yanjuan; Yu, Diqiu

2016-08-01

Although necrotrophic pathogens cause many devastating plant diseases, our understanding of the plant defense response to them is limited. Here, we found that loss of function of WRKY57 enhanced the resistance of Arabidopsis (Arabidopsis thaliana) against Botrytis cinerea infection. Further investigation suggested that the negative regulation of WRKY57 against B cinerea depends on the jasmonic acid (JA) signaling pathway. Chromatin immunoprecipitation experiments revealed that WRKY57 directly binds to the promoters of JASMONATE ZIM-DOMAIN1 (JAZ1) and JAZ5, encoding two important repressors of the JA signaling pathway, and activates their transcription. In vivo and in vitro experiments demonstrated that WRKY57 interacts with nuclear-encoded SIGMA FACTOR BINDING PROTEIN1 (SIB1) and SIB2. Further experiments display that the same domain, the VQ motif, of SIB1 and SIB2 interact with WRKY33 and WRKY57. Moreover, transient transcriptional activity assays confirmed that WRKY57 and WRKY33 competitively regulate JAZ1 and JAZ5, SIB1 and SIB2 further enhance these competitions of WRKY57 to WRKY33. Therefore, coordinated regulation of Arabidopsis against B cinerea by transcription activators and repressors would benefit plants by allowing fine regulation of defense. © 2016 American Society of Plant Biologists. All Rights Reserved.

Skp2B overexpression alters a prohibitin-p53 axis and the transcription of PAPP-A, the protease of insulin-like growth factor binding protein 4.

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Harish Chander

Full Text Available We previously reported that the degradation of prohibitin by the SCF(Skp2B ubiquitin ligase results in a defect in the activity of p53. We also reported that MMTV-Skp2B transgenic mice develop mammary gland tumors that are characterized by an increased proteolytic cleavage of the insulin-like growth factor binding protein 4 (IGFBP-4, an inhibitor of IGF signaling. However, whether a link exists between a defect in p53 activity and proteolysis of IGFBP-4 was not established.We analyzed the levels of pregnancy-associated plasma protein A (PAPP-A, the protease of IGFBP-4, in MMTV-Skp2B transgenic mice and found that PAPP-A levels are elevated. Further, we found a p53 binding site in intron 1 of the PAPP-A gene and that both wild type and mutant p53 bind to this site. However, binding of wild type p53 results in the transcriptional repression of PAPP-A, while binding of mutant p53 results in the transcriptional activation of PAPP-A. Since MMTV-Skp2B mice express wild type p53 and yet show elevated levels of PAPP-A, at first, these observations appeared contradictory. However, further analysis revealed that the defect in p53 activity in Skp2B overexpressing cells does not only abolish the activity of wild type of p53 but actually mimics that of mutant p53. Our results suggest that in absence of prohibitin, the half-life of p53 is increased and like mutant p53, the conformation of p53 is denatured.These observations revealed a novel function of prohibitin as a chaperone of p53. Further, they suggest that binding of denatured p53 in intron 1 causes an enhancer effect and increases the transcription of PAPP-A. Therefore, these findings indicate that the defect in p53 function and the increased proteolysis of IGFBP-4, we had observed, represent two components of the same pathway, which contributes to the oncogenic function of Skp2B.

A transcription factor active on the epidermal growth factor receptor gene

International Nuclear Information System (INIS)

Kageyama, R.; Merlino, G.T.; Pastan, I.

1988-01-01

The authors have developed an in vitro transcription system for the epidermal growth factor receptor (EGFR) oncogene by using nuclear extracts of A431 human epidermoid carcinoma cells, which overproduce EGFR. They found that a nuclear factor, termed EGFR-specific transcription factor (ETF), specifically stimulated EGFR transcription by 5- to 10-fold. In this report, ETF, purified by using sequence-specific oligonucleotide affinity chromatography, is shown by renaturing material eluted from a NaDodSO 4 /polyacrylamide gel to be a protein with a molecular mass of 120 kDa. ETF binds to the promoter region, as measured by DNase I footprinting and gel-mobility-shift assays, and specifically stimulates the transcription of the EGFR gene in a reconstituted in vitro transcription system. These results suggest that ETF could play a role in the overexpression of the cellular oncogene EGFR

Water deficits and heat shock effects on photosynthesis of a transgenic Arabidopsis thaliana constitutively expressing ABP9, a bZIP transcription factor

DEFF Research Database (Denmark)

Zhang, Xia; Wollenweber, Bernd; Jiang, Dong

2008-01-01

The effects of water deficits (WD), heat shock (HS), and both (HSWD) on photosynthetic carbon- and light-use efficiencies together with leaf ABA content, pigment composition and expressions of stress- and light harvesting-responsive genes were investigated in ABP9 [ABA-responsive-element (ABRE...... transport rate (ETR) in 5P2 plants were depressed under optimal growth conditions (control) in relation to WT, they were enhanced under HS and HSWD. These results indicate that ABP9 transgenic plants are less susceptible to stress than the WT. In addition, the increased ABA contents in both WT and 5P2......, altered expression of stress-regulated or light harvesting-responsive genes was observed. Collectively, our results indicate that constitutive expression of ABP9 improves the photosynthetic capacity of plants under stress by adjusting photosynthetic pigment composition, dissipating excess light energy...

HTLV-1 bZIP Factor Impairs Anti-viral Immunity by Inducing Co-inhibitory Molecule, T Cell Immunoglobulin and ITIM Domain (TIGIT.

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Keiko Yasuma

2016-01-01

Full Text Available Human T-cell leukemia virus type 1 (HTLV-1 infects CD4+ T cells and induces proliferation of infected cells in vivo, which leads to the onset of adult T-cell leukemia (ATL in some infected individuals. The HTLV-1 bZIP factor (HBZ gene, which is encoded in the minus strand of HTLV-1, plays critical roles in pathogenesis. In this study, RNA-seq and ChIP-seq analyses using HBZ transduced T cells revealed that HBZ upregulates the expression and promoter acetylation levels of a co-inhibitory molecule, T cell immunoglobulin and ITIM domain (TIGIT, in addition to those of regulatory T cells related genes, Foxp3 and Ccr4. TIGIT was expressed on CD4+ T cells from HBZ-transgenic (HBZ-Tg mice, and on ATL cells and HTLV-1 infected CD4+ T cells of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP in vivo. Expression of Blimp1 and IL-10 was upregulated in TIGIT+CD4+ cells of HBZ-Tg mice compared with TIGIT-CD4+ T cells, suggesting the correlation between TIGIT expression and IL-10 production. When CD4+ T cells from HBZ-Tg mice were stimulated with TIGIT's ligand, CD155, their production of the inhibitory cytokine IL-10 was enhanced. Furthermore, dendritic cells from HBZ-Tg mice produced high levels of IL-10 after stimulation. These data suggest that HBZ alters immune system to suppressive state via TIGIT and IL-10. Importantly, TIGIT suppressed T-cell responses to another HTLV-1 virus protein, Tax, in vitro. Blocking of TIGIT and PD-1 slightly increased anti-Tax T-cell activity in some HAM/TSP patients. These results suggest that HBZ-induced TIGIT on HTLV-1 infected cells impairs T-cell responses to viral antigens. This study shows that HBZ-induced TIGIT plays a pivotal role in attenuating host immune responses and shaping a microenvironment favorable to HTLV-1.

25-hydroxycholesterol promotes RANKL-induced osteoclastogenesis through coordinating NFATc1 and Sp1 complex in the transcription of miR-139-5p

International Nuclear Information System (INIS)

Zhang, Lishan; Lv, Yinping; Xian, Guozhe; Lin, Yanliang

2017-01-01

25-hydroxycholesterol (25-HC) is implicated in many processes, including lipid metabolism and the immune response. However, the role of 25-HC in RANKL-induced osteoclastogenesis remains largely unknown. Our results showed that 25-HC inhibited miR-139-5p expression in mouse bone marrow macrophages (BMMs) cultured in receptor activator of NF-κB ligand (RANKL) and monocyte macrophage colony-stimulating factor (M-CSF). Further investigation suggested that 25-HC promoted the expression of nuclear factor of activated T cell cytoplasmic 1 (NFATc1) and Sp1, especially in the presence of RANKL and M-CSF. Meanwhile, 25-HC induced nuclear translocation of NFATc1, resulting in the interaction between NFATc1 and Sp1 that was confirmed by co-immunoprecipitation. Chromatin immunoprecipitation assay indicated that Sp1 could bind to miR-139-5p promoter, but NFATc1 had no binding capacity. Although forming NFATc1/Sp1 complex increased its binding to miR-139-5p promoter, the complex inhibited the transcriptional activity of Sp1. Inhibition of NFATc1 increase the expression of miR-139-5p, which might be due to the release of free Sp1 that could bind to the promoter of miR-139-5p. Enforced expression of miR-139-5p impaired osteoclastogenesis induced by co-treatment with 25-HC and RANKL. These results suggested that 25-HC induced the interaction between NFATc1 and Sp1, reducing the level of free Sp1 to inhibit miR-139-5p expression and promote osteoclastogenesis. - Highlights: • 25-hydroxycholesterol inhibited miR-139-5p expression in bone marrow macrophages. • 25-hydroxycholesterol promoted the expression of NFATc1 and Sp1. • 25-hydroxycholesterol induced the interaction between NFATc1 and Sp1. • NFATc1/Sp1 complex inhibited the transcription of miR-139-5p. • MiR-139-5p impaired osteoclastogenesis induced by 25-hydroxycholesterol and RANKL.

Rationally designed, heterologous S. cerevisiae transcripts expose novel expression determinants

Science.gov (United States)

Ben-Yehezkel, Tuval; Atar, Shimshi; Zur, Hadas; Diament, Alon; Goz, Eli; Marx, Tzipy; Cohen, Rafael; Dana, Alexandra; Feldman, Anna; Shapiro, Ehud; Tuller, Tamir

2015-01-01

Deducing generic causal relations between RNA transcript features and protein expression profiles from endogenous gene expression data remains a major unsolved problem in biology. The analysis of gene expression from heterologous genes contributes significantly to solving this problem, but has been heavily biased toward the study of the effect of 5′ transcript regions and to prokaryotes. Here, we employ a synthetic biology driven approach that systematically differentiates the effect of different regions of the transcript on gene expression up to 240 nucleotides into the ORF. This enabled us to discover new causal effects between features in previously unexplored regions of transcripts, and gene expression in natural regimes. We rationally designed, constructed, and analyzed 383 gene variants of the viral HRSVgp04 gene ORF, with multiple synonymous mutations at key positions along the transcript in the eukaryote S. cerevisiae. Our results show that a few silent mutations at the 5′UTR can have a dramatic effect of up to 15 fold change on protein levels, and that even synonymous mutations in positions more than 120 nucleotides downstream from the ORF 5′end can modulate protein levels up to 160%–300%. We demonstrate that the correlation between protein levels and folding energy increases with the significance of the level of selection of the latter in endogenous genes, reinforcing the notion that selection for folding strength in different parts of the ORF is related to translation regulation. Our measured protein abundance correlates notably(correlation up to r = 0.62 (p=0.0013)) with mean relative codon decoding times, based on ribosomal densities (Ribo-Seq) in endogenous genes, supporting the conjecture that translation elongation and adaptation to the tRNA pool can modify protein levels in a causal/direct manner. This report provides an improved understanding of transcript evolution, design principles of gene expression regulation, and suggests simple

Silencing of human T-cell leukemia virus type I gene transcription by epigenetic mechanisms

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Mueller Nancy

2005-10-01

Full Text Available Abstract Background Human T-cell leukemia virus type I (HTLV-I causes adult T-cell leukemia (ATL after a long latent period. Among accessory genes encoded by HTLV-I, the tax gene is thought to play a central role in oncogenesis. However, Tax expression is disrupted by several mechanims including genetic changes of the tax gene, deletion/hypermethylation of 5'-LTR. To clarify the role of epigenetic changes, we analyzed DNA methylation and histone modification in the whole HTLV-I provirus genome. Results The gag, pol and env genes of HTLV-I provirus were more methylated than pX region, whereas methylation of 5'-LTR was variable and 3'-LTR was not methylated at all. In ATL cell lines, complete DNA methylation of 5'-LTR was associated with transcriptional silencing of viral genes. HTLV-I provirus was more methylated in primary ATL cells than in carrier state, indicating the association with disease progression. In seroconvertors, DNA methylation was already observed in internal sequences of provirus just after seroconversion. Taken together, it is speculated that DNA methylation first occurs in the gag, pol and env regions and then extends in the 5' and 3' directions in vivo, and when 5'-LTR becomes methylated, viral transcription is silenced. Analysis of histone modification in the HTLV-I provirus showed that the methylated provirus was associated with hypoacetylation. However, the tax gene transcript could not be detected in fresh ATL cells regardless of hyperacetylated histone H3 in 5'-LTR. The transcription rapidly recovered after in vitro culture in such ATL cells. Conclusion These results showed that epigenetic changes of provirus facilitated ATL cells to evade host immune system by suppressing viral gene transcription. In addition, this study shows the presence of another reversible mechanism that suppresses the tax gene transcription without DNA methylation and hypoacetylated histone.

Dataset of transcriptional landscape of B cell early activation

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Alexander S. Garruss

2015-09-01

Full Text Available Signaling via B cell receptors (BCR and Toll-like receptors (TLRs result in activation of B cells with distinct physiological outcomes, but transcriptional regulatory mechanisms that drive activation and distinguish these pathways remain unknown. At early time points after BCR and TLR ligand exposure, 0.5 and 2 h, RNA-seq was performed allowing observations on rapid transcriptional changes. At 2 h, ChIP-seq was performed to allow observations on important regulatory mechanisms potentially driving transcriptional change. The dataset includes RNA-seq, ChIP-seq of control (Input, RNA Pol II, H3K4me3, H3K27me3, and a separate RNA-seq for miRNA expression, which can be found at Gene Expression Omnibus Dataset GSE61608. Here, we provide details on the experimental and analysis methods used to obtain and analyze this dataset and to examine the transcriptional landscape of B cell early activation.

Elucidation of functional markers from Aspergillus nidulans developmental regulator FlbB and their phylogenetic distribution.

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Marc S Cortese

Full Text Available Aspergillus nidulans is a filamentous fungus widely used as a model for biotechnological and clinical research. It is also used as a platform for the study of basic eukaryotic developmental processes. Previous studies identified and partially characterized a set of proteins controlling cellular transformations in this ascomycete. Among these proteins, the bZip type transcription factor FlbB is a key regulator of reproduction, stress responses and cell-death. Our aim here was the prediction, through various bioinformatic methods, of key functional residues and motifs within FlbB in order to inform the design of future laboratory experiments and further the understanding of the molecular mechanisms that control fungal development. A dataset of FlbB orthologs and those of its key interaction partner FlbE was assembled from 40 members of the Pezizomycotina. Unique features were identified in each of the three structural domains of FlbB. The N-terminal region encoded a bZip transcription factor domain with a novel histidine-containing DNA binding motif while the dimerization determinants exhibited two distinct profiles that segregated by class. The C-terminal region of FlbB showed high similarity with the AP-1 family of stress response regulators but with variable patterns of conserved cysteines that segregated by class and order. Motif conservation analysis revealed that nine FlbB orthologs belonging to the Eurotiales order contained a motif in the central region that could mediate interaction with FlbE. The key residues and motifs identified here provide a basis for the design of follow-up experimental investigations. Additionally, the presence or absence of these residues and motifs among the FlbB orthologs could help explain the differences in the developmental programs among fungal species as well as define putative complementation groups that could serve to extend known functional characterizations to other species.

Transgenic plants expressing GLK1 and CCA1 having increased nitrogen assimilation capacity

Science.gov (United States)

Coruzzi, Gloria [New York, NY; Gutierrez, Rodrigo A [Santiago, CL; Nero, Damion C [Woodside, NY

2012-04-10

Provided herein are compositions and methods for producing transgenic plants. In specific embodiments, transgenic plants comprise a construct comprising a polynucleotide encoding CCA1, GLK1 or bZIP1, operably linked to a plant-specific promote, wherein the CCA1, GLK1 or bZIP1 is ectopically overexpressed in the transgenic plants, and wherein the promoter is optionally a constitutive or inducible promoter. In other embodiments, transgenic plants in which express a lower level of CCA1, GLK1 or bZIP1 are provided. Also provided herein are commercial products (e.g., pulp, paper, paper products, or lumber) derived from the transgenic plants (e.g., transgenic trees) produced using the methods provided herein.

Methods of affecting nitrogen assimilation in plants

Energy Technology Data Exchange (ETDEWEB)

Coruzzi, Gloria; Gutierrez, Rodrigo A.; Nero, Damion C.

2016-10-11

Provided herein are compositions and methods for producing transgenic plants. In specific embodiments, transgenic plants comprise a construct comprising a polynucleotide encoding CCA1, GLK1 or bZIP1, operably linked to a plant-specific promote, wherein the CCA1, GLK1 or bZIP1 is ectopically overexpressed in the transgenic plants, and wherein the promoter is optionally a constitutive or inducible promoter. In other embodiments, transgenic plants in which express a lower level of CCA1, GLK1 or bZIP1 are provided. Also provided herein are commercial products (e.g., pulp, paper, paper products, or lumber) derived from the transgenic plants (e.g., transgenic trees) produced using the methods provided herein.

A long HBV transcript encoding pX is inefficiently exported from the nucleus

International Nuclear Information System (INIS)

Doitsh, Gilad; Shaul, Yosef

2003-01-01

The longest hepatitis B virus transcript is a 3.9-kb mRNA whose function remained unclear. In this study, we wished to identify the translation products and physiological role of this viral transcript. This transcript initiates from the X promoter region ignoring the inefficient and noncanonical viral polyadenylation signal at the first round of transcription. However, an HBV mutant with canonical polyadenylation signal continues, though with lower efficiency, to program the synthesis of this long transcript, indicating that the deviated HBV polyadenylation signal is important but not essential to enable transcription of the 3.9-kb species. The 3.9-kb RNA contains two times the X open reading frame (ORF). The X ORF at the 5'-end is positioned upstream of the CORE gene. By generating an HBV DNA mutant in which the X and Core ORFs are fused, we demonstrated the production of a 40-kDa X-Core fusion protein that must be encoded by the 3.9-kb transcript. Mutagenesis studies revealed that the production of this protein depends on the 5' X ORF ATG, suggesting that the 3.9-kb RNA is active in translation of the X ORF. Based on these features, the 3.9-kb transcript was designated lxRNA for long X RNA. Unlike other HBV transcripts, lxRNA harbors two copies of PRE, the posttranscriptional regulatory element that controls the nuclear export of HBV mRNAs. Unexpectedly, despite the presence of PRE sequences, RNA fractionation analysis revealed that lxRNA barely accumulates in the cytoplasm, suggesting that nuclear export of lxRNA is poor. Collectively, our data suggest that two distinct HBV mRNA species encode pX and that the HBV transcripts are differentially regulated at the level of nuclear export

Nucleocytoplasmic shuttling of transcription factors

DEFF Research Database (Denmark)

Cartwright, P; Helin, K

2000-01-01

To elicit the transcriptional response following intra- or extracellular stimuli, the signals need to be transmitted to their site of action within the nucleus. The nucleocytoplasmic shuttling of transcription factors is a mechanism mediating this process. The activation and inactivation...... of the transcriptional response is essential for cells to progress through the cell cycle in a normal manner. The involvement of cytoplasmic and nuclear accessory molecules, and the general nuclear membrane transport components, are essential for this process. Although nuclear import and export for different...... transcription factor families are regulated by similar mechanisms, there are several differences that allow for the specific activation of each transcription factor. This review discusses the general import and export pathways found to be common amongst many different transcription factors, and highlights...

Controlling cellular P-TEFb activity by the HIV-1 transcriptional transactivator Tat.

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Lisa Muniz

Full Text Available The human immunodeficiency virus 1 (HIV-1 transcriptional transactivator (Tat is essential for synthesis of full-length transcripts from the integrated viral genome by RNA polymerase II (Pol II. Tat recruits the host positive transcription elongation factor b (P-TEFb to the HIV-1 promoter through binding to the transactivator RNA (TAR at the 5'-end of the nascent HIV transcript. P-TEFb is a general Pol II transcription factor; its cellular activity is controlled by the 7SK small nuclear RNA (snRNA and the HEXIM1 protein, which sequester P-TEFb into transcriptionally inactive 7SK/HEXIM/P-TEFb snRNP. Besides targeting P-TEFb to HIV transcription, Tat also increases the nuclear level of active P-TEFb through promoting its dissociation from the 7SK/HEXIM/P-TEFb RNP by an unclear mechanism. In this study, by using in vitro and in vivo RNA-protein binding assays, we demonstrate that HIV-1 Tat binds with high specificity and efficiency to an evolutionarily highly conserved stem-bulge-stem motif of the 5'-hairpin of human 7SK snRNA. The newly discovered Tat-binding motif of 7SK is structurally and functionally indistinguishable from the extensively characterized Tat-binding site of HIV TAR and importantly, it is imbedded in the HEXIM-binding elements of 7SK snRNA. We show that Tat efficiently replaces HEXIM1 on the 7SK snRNA in vivo and therefore, it promotes the disassembly of the 7SK/HEXIM/P-TEFb negative transcriptional regulatory snRNP to augment the nuclear level of active P-TEFb. This is the first demonstration that HIV-1 specifically targets an important cellular regulatory RNA, most probably to promote viral transcription and replication. Demonstration that the human 7SK snRNA carries a TAR RNA-like Tat-binding element that is essential for the normal transcriptional regulatory function of 7SK questions the viability of HIV therapeutic approaches based on small drugs blocking the Tat-binding site of HIV TAR.

Genome-wide transcription analyses in rice using tiling microarrays

DEFF Research Database (Denmark)

Li, Lei; Wang, Xiangfeng; Stolc, Viktor

2006-01-01

. We report here a full-genome transcription analysis of the indica rice subspecies using high-density oligonucleotide tiling microarrays. Our results provided expression data support for the existence of 35,970 (81.9%) annotated gene models and identified 5,464 unique transcribed intergenic regions...... that share similar compositional properties with the annotated exons and have significant homology to other plant proteins. Elucidating and mapping of all transcribed regions revealed an association between global transcription and cytological chromosome features, and an overall similarity of transcriptional......Sequencing and computational annotation revealed several features, including high gene numbers, unusual composition of the predicted genes and a large number of genes lacking homology to known genes, that distinguish the rice (Oryza sativa) genome from that of other fully sequenced model species...

DNA-methylation dependent regulation of embryo-specific 5S ribosomal DNA cluster transcription in adult tissues of sea urchin Paracentrotus lividus.

Science.gov (United States)

Bellavia, Daniele; Dimarco, Eufrosina; Naselli, Flores; Caradonna, Fabio

2013-10-01

We have previously reported a molecular and cytogenetic characterization of three different 5S rDNA clusters in the sea urchin Paracentrotus lividus and recently, demonstrated the presence of high heterogeneity in functional 5S rRNA. In this paper, we show some important distinctive data on 5S rRNA transcription for this organism. Using single strand conformation polymorphism (SSCP) analysis, we demonstrate the existence of two classes of 5S rRNA, one which is embryo-specific and encoded by the smallest (700 bp) cluster and the other which is expressed at every stage and encoded by longer clusters (900 and 950 bp). We also demonstrate that the embryo-specific class of 5S rRNA is expressed in oocytes and embryonic stages and is silenced in adult tissue and that this phenomenon appears to be due exclusively to DNA methylation, as indicated by sensitivity to 5-azacytidine, unlike Xenopus where this mechanism is necessary but not sufficient to maintain the silenced status. © 2013 Elsevier Inc. All rights reserved.

Arabidopsis ABI5 plays a role in regulating ROS homeostasis by activating CATALASE 1 transcription in seed germination.

Science.gov (United States)

Bi, Chao; Ma, Yu; Wu, Zhen; Yu, Yong-Tao; Liang, Shan; Lu, Kai; Wang, Xiao-Fang

2017-05-01

It has been known that ABA INSENSITIVE 5 (ABI5) plays a vital role in regulating seed germination. In the present study, we showed that inhibition of the catalase activity with 3-amino-1,2,4-triazole (3-AT) inhibits seed germination of Col-0, abi5 mutants and ABI5-overexpression transgenic lines. Compared with Col-0, the seeds of abi5 mutants showed more sensitive to 3-AT during seed germination, while the seeds of ABI5-overexpression transgenic lines showed more insensitive. H 2 O 2 showed the same effect on seed germination of Col-0, abi5 mutants and ABI5-overexpression transgenic lines as 3-AT. These results suggest that ROS is involved in the seed germination mediated by ABI5. Further, we observed that T-DNA insertion mutants of the three catalase members in Arabidopsis displayed 3-AT-insensitive or -hypersensitive phenotypes during seed germination, suggesting that these catalase members regulate ROS homeostasis in a highly complex way. ABI5 affects reactive oxygen species (ROS) homeostasis by affecting CATALASE expression and catalase activity. Furthermore, we showed that ABI5 directly binds to the CAT1 promoter and activates CAT1 expression. Genetic evidence supports the idea that CAT1 functions downstream of ABI5 in ROS signaling during seed germination. RNA-sequencing analysis indicates that the transcription of the genes involved in ROS metabolic process or genes responsive to ROS stress is impaired in abi5-1 seeds. Additionally, expression changes in some genes correlative to seed germination were showed due to the change in ABI5 expression under 3-AT treatment. Together, all the findings suggest that ABI5 regulates seed germination at least partly by affecting ROS homeostasis.

Aqp5 is a new transcriptional target of Dot1a and a regulator of Aqp2.

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Hongyu Wu

Full Text Available Dot1l encodes histone H3 K79 methyltransferase Dot1a. Mice with Dot1l deficiency in renal Aqp2-expressing cells (Dot1l(AC develop polyuria by unknown mechanisms. Here, we report that Aqp5 links Dot1l deletion to polyuria through Aqp2. cDNA array analysis revealed and real-time RT-qPCR validated Aqp5 as the most upregulated gene in Dot1l(AC vs. control mice. Aqp5 protein is barely detectable in controls, but robustly expressed in the Dot1l(AC kidneys, where it colocalizes with Aqp2. The upregulation of Aqp5 is coupled with reduced association of Dot1a and H3 dimethyl K79 with specific subregions in Aqp5 5' flanking region in Dot1l(AC vs. control mice. In vitro studies in IMCD3, MLE-15 and 293Tcells using multiple approaches including real-time RT-qPCR, luciferase reporter assay, cell surface biotinylation assay, colocalization, and co-immunoprecipitation uncovered that Dot1a represses Aqp5. Human AQP5 interacts with AQP2 and impairs its cell surface localization. The AQP5/AQP2 complex partially resides in the ER/Golgi. Consistently, AQP5 is expressed in none of 15 normal controls, but in all of 17 kidney biopsies from patients with diabetic nephropathy. In the patients with diabetic nephropathy, AQP5 colocalizes with AQP2 in the perinuclear region and AQP5 expression is associated with impaired cellular H3 dimethyl K79. Taken together, these data for the first time identify Aqp5 as a Dot1a potential transcriptional target, and an Aqp2 binding partner and regulator, and suggest that the upregulated Aqp5 may contribute to polyuria, possibly by impairing Aqp2 membrane localization, in Dot1l(AC mice and in patients with diabetic nephropathy.

Mitochondrial Reactive Oxygen Species Trigger Hypoxia-Induced Transcription

Science.gov (United States)

Chandel, N. S.; Maltepe, E.; Goldwasser, E.; Mathieu, C. E.; Simon, M. C.; Schumacker, P. T.

1998-09-01

Transcriptional activation of erythropoietin, glycolytic enzymes, and vascular endothelial growth factor occurs during hypoxia or in response to cobalt chloride (CoCl2) in Hep3B cells. However, neither the mechanism of cellular O2 sensing nor that of cobalt is fully understood. We tested whether mitochondria act as O2 sensors during hypoxia and whether hypoxia and cobalt activate transcription by increasing generation of reactive oxygen species (ROS). Results show (i) wild-type Hep3B cells increase ROS generation during hypoxia (1.5% O2) or CoCl2 incubation, (ii) Hep3B cells depleted of mitochondrial DNA (ρ 0 cells) fail to respire, fail to activate mRNA for erythropoietin, glycolytic enzymes, or vascular endothelial growth factor during hypoxia, and fail to increase ROS generation during hypoxia; (iii) ρ 0 cells increase ROS generation in response to CoCl2 and retain the ability to induce expression of these genes; and (iv) the antioxidants pyrrolidine dithiocarbamate and ebselen abolish transcriptional activation of these genes during hypoxia or CoCl2 in wild-type cells, and abolish the response to CoCl2 in ρ 0 cells. Thus, hypoxia activates transcription via a mitochondria-dependent signaling process involving increased ROS, whereas CoCl2 activates transcription by stimulating ROS generation via a mitochondria-independent mechanism.

LL5beta: a regulator of postsynaptic differentiation identified in a screen for synaptically enriched transcripts at the neuromuscular junction.

Science.gov (United States)

Kishi, Masashi; Kummer, Terrance T; Eglen, Stephen J; Sanes, Joshua R

2005-04-25

In both neurons and muscle fibers, specific mRNAs are concentrated beneath and locally translated at synaptic sites. At the skeletal neuromuscular junction, all synaptic RNAs identified to date encode synaptic components. Using microarrays, we compared RNAs in synapse-rich and -free regions of muscles, thereby identifying transcripts that are enriched near synapses and that encode soluble membrane and nuclear proteins. One gene product, LL5beta, binds to both phosphoinositides and a cytoskeletal protein, filamin, one form of which is concentrated at synaptic sites. LL5beta is itself associated with the cytoplasmic face of the postsynaptic membrane; its highest levels border regions of highest acetylcholine receptor (AChR) density, which suggests a role in "corraling" AChRs. Consistent with this idea, perturbing LL5beta expression in myotubes inhibits AChR aggregation. Thus, a strategy designed to identify novel synaptic components led to identification of a protein required for assembly of the postsynaptic apparatus.

High-level expression of human stem cell factor fused with erythropoietin mimetic peptide in Escherichia coli.

Science.gov (United States)

Su, Lin; Chen, Song-Sen; Yang, Ke-Gong; Liu, Chang-Zheng; Zhang, Yan-Li; Liang, Zhi-Quan

2006-06-01

Stem cell factor (SCF) and erythropoietin are essential for normal erythropoiesis and induce proliferation and differentiation synergistically for erythroid progenitor cells. Here, we report our work on construction of SCF/erythropoietin mimetic peptide (EMP) fusion protein gene, in which human SCF cDNA (1-165aa) and EMP sequence (20aa) were connected using a short (GGGGS) or long (GGGGSGGGGGS) linker sequence. The SCF/EMP gene was cloned into the pBV220 vector and expressed in the Escherichia coli DH5alpha strain. The expression level of the fusion protein was about 30% of total cell protein. The resulting inclusion bodies were solubilized with 8 M urea, followed by dilution refolding. The renatured protein was subsequently purified by Q-Sepharose FF column. The final product was >95% pure by SDS-PAGE and the yield of fusion protein was about 40 mg/L of culture. UT-7 cell proliferation and human cord blood cell colony-forming assays showed that the fusion proteins exhibited more potent activity than recombinant human SCF, suggesting a new strategy to enhance biological activities of growth factors.

HAfTs are novel lncRNA transcripts from aflatoxin exposure.

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B Alex Merrick

Full Text Available The transcriptome can reveal insights into precancer biology. We recently conducted RNA-Seq analysis on liver RNA from male rats exposed to the carcinogen, aflatoxin B1 (AFB1, for 90 days prior to liver tumor onset. Among >1,000 differentially expressed transcripts, several novel, unannotated Cufflinks-assembled transcripts, or HAfTs (Hepatic Aflatoxin Transcripts were found. We hypothesized PCR-cloning and RACE (rapid amplification of cDNA ends could further HAfT identification. Sanger data was obtained for 6 transcripts by PCR and 16 transcripts by 5'- and 3'-RACE. BLAST alignments showed, with two exceptions, HAfT transcripts were lncRNAs, >200nt without apparent long open reading frames. Six rat HAfT transcripts were classified as 'novel' without RefSeq annotation. Sequence alignment and genomic synteny showed each rat lncRNA had a homologous locus in the mouse genome and over half had homologous loci in the human genome, including at least two loci (and possibly three others that were previously unannotated. While HAfT functions are not yet clear, coregulatory roles may be possible from their adjacent orientation to known coding genes with altered expression that include 8 HAfT-gene pairs. For example, a unique rat HAfT, homologous to Pvt1, was adjacent to known genes controlling cell proliferation. Additionally, PCR and RACE Sanger sequencing showed many alternative splice variants and refinements of exon sequences compared to Cufflinks assembled transcripts and gene prediction algorithms. Presence of multiple splice variants and short tandem repeats found in some HAfTs may be consequential for secondary structure, transcriptional regulation, and function. In summary, we report novel, differentially expressed lncRNAs after exposure to the genotoxicant, AFB1, prior to neoplastic lesions. Complete cloning and sequencing of such transcripts could pave the way for a new set of sensitive and early prediction markers for chemical

Cdc15 Phosphorylates the C-terminal Domain of RNA Polymerase II for Transcription during Mitosis.

Science.gov (United States)

Singh, Amit Kumar; Rastogi, Shivangi; Shukla, Harish; Asalam, Mohd; Rath, Srikanta Kumar; Akhtar, Md Sohail

2017-03-31

In eukaryotes, the basal transcription in interphase is orchestrated through the regulation by kinases (Kin28, Bur1, and Ctk1) and phosphatases (Ssu72, Rtr1, and Fcp1), which act through the post-translational modification of the C-terminal domain (CTD) of the largest subunit of RNA polymerase II. The CTD comprises the repeated Tyr-Ser-Pro-Thr-Ser-Pro-Ser motif with potential epigenetic modification sites. Despite the observation of transcription and periodic expression of genes during mitosis with entailing CTD phosphorylation and dephosphorylation, the associated CTD specific kinase(s) and its role in transcription remains unknown. Here we have identified Cdc15 as a potential kinase phosphorylating Ser-2 and Ser-5 of CTD for transcription during mitosis in the budding yeast. The phosphorylation of CTD by Cdc15 is independent of any prior Ser phosphorylation(s). The inactivation of Cdc15 causes reduction of global CTD phosphorylation during mitosis and affects the expression of genes whose transcript levels peak during mitosis. Cdc15 also influences the complete transcription of clb2 gene and phosphorylates Ser-5 at the promoter and Ser-2 toward the 3' end of the gene. The observation that Cdc15 could phosphorylate Ser-5, as well as Ser-2, during transcription in mitosis is in contrast to the phosphorylation marks put by the kinases in interphase (G 1 , S, and G 2 ), where Cdck7/Kin28 phosphorylates Ser-5 at promoter and Bur1/Ctk1 phosphorylates Ser-2 at the 3' end of the genes. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Microarray and cDNA sequence analysis of transcription during nerve-dependent limb regeneration

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Bryant Susan V

2009-01-01

Full Text Available Abstract Background Microarray analysis and 454 cDNA sequencing were used to investigate a centuries-old problem in regenerative biology: the basis of nerve-dependent limb regeneration in salamanders. Innervated (NR and denervated (DL forelimbs of Mexican axolotls were amputated and transcripts were sampled after 0, 5, and 14 days of regeneration. Results Considerable similarity was observed between NR and DL transcriptional programs at 5 and 14 days post amputation (dpa. Genes with extracellular functions that are critical to wound healing were upregulated while muscle-specific genes were downregulated. Thus, many processes that are regulated during early limb regeneration do not depend upon nerve-derived factors. The majority of the transcriptional differences between NR and DL limbs were correlated with blastema formation; cell numbers increased in NR limbs after 5 dpa and this yielded distinct transcriptional signatures of cell proliferation in NR limbs at 14 dpa. These transcriptional signatures were not observed in DL limbs. Instead, gene expression changes within DL limbs suggest more diverse and protracted wound-healing responses. 454 cDNA sequencing complemented the microarray analysis by providing deeper sampling of transcriptional programs and associated biological processes. Assembly of new 454 cDNA sequences with existing expressed sequence tag (EST contigs from the Ambystoma EST database more than doubled (3935 to 9411 the number of non-redundant human-A. mexicanum orthologous sequences. Conclusion Many new candidate gene sequences were discovered for the first time and these will greatly enable future studies of wound healing, epigenetics, genome stability, and nerve-dependent blastema formation and outgrowth using the axolotl model.

Gibberellic acid and cGMP-dependent transcriptional regulation in arabidopsis thaliana

KAUST Repository

Bastian, René

2010-03-01

An ever increasing amount of transcriptomic data and analysis tools provide novel insight into complex responses of biological systems. Given these resources we have undertaken to review aspects of transcriptional regulation in response to the plant hormone gibberellic acid (GA) and its second messenger guanosine 3\\',5\\'-cyclic monophosphate (cGMP) in Arabidopsis thaliana, both wild type and selected mutants. Evidence suggests enrichment of GA-responsive (GARE) elements in promoters of genes that are transcriptionally upregulated in response to cGMP but downregulated in a GA insensitive mutant (ga1-3). In contrast, in the genes upregulated in the mutant, no enrichment in the GARE is observed suggesting that GARE motifs are diagnostic for GA-induced and cGMP-dependent transcriptional upregulation. Further, we review how expression studies of GA-dependent transcription factors and transcriptional networks based on common promoter signatures derived from ab initio analyses can contribute to our understanding of plant responses at the systems level. © 2010 Landes Bioscience.