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Sample records for putative immunophilin gene

  1. Putative modifier genes in mevalonate kinase deficiency.

    Science.gov (United States)

    Marcuzzi, Annalisa; Vozzi, Diego; Girardelli, Martina; Tricarico, Paola Maura; Knowles, Alessandra; Crovella, Sergio; Vuch, Josef; Tommasini, Alberto; Piscianz, Elisa; Bianco, Anna Monica

    2016-04-01

    Mevalonate kinase deficiency (MKD) is an autosomal recessive auto‑inflammatory disease, caused by impairment of the mevalonate pathway. Although the molecular mechanism remains to be elucidated, there is clinical evidence suggesting that other regulatory genes may be involved in determining the phenotype. The identification of novel target genes may explain non‑homogeneous genotype‑phenotype correlations, and provide evidence in support of the hypothesis that novel regulatory genes predispose or amplify deregulation of the mevalonate pathway in this orphan disease. In the present study, DNA samples were obtained from five patients with MKD, which were then analyzed using whole exome sequencing. A missense variation in the PEX11γ gene was observed in homozygosis in P2, possibly correlating with visual blurring. The UNG rare gene variant was detected in homozygosis in P5, without correlating with a specific clinical phenotype. A number of other variants were found in the five analyzed DNA samples from the MKD patients, however no correlation with the phenotype was established. The results of the presents study suggested that further analysis, using next generation sequencing approaches, is required on a larger sample size of patients with MKD, who share the same MVK mutations and exhibit 'extreme' clinical phenotypes. As MVK mutations may be associated with MKD, the identification of specific modifier genes may assist in providing an earlier diagnosis.

  2. Chromosomal Abnormalities and Putative Susceptibility Genes in Autism Spectrum Disorders

    DEFF Research Database (Denmark)

    Nielsen, Mette Gilling

    Autism spectrum disorders (ASDs) is a heterogeneous group of neurodevelopmental disorders with a significant genetic component as shown by family and twin studies. However, only a few genes have repeatedly been shown to be involved in the development of ASDs. The aim of this study has been...... to identify possible ASD susceptibility genes. Genome screens in ASD patients suggest possible susceptibility gene regions on almost every chromosome. We identified four ASD patients with chromosomal rearrangements, two of which were familial rearrangements involving one of these putative susceptibility gene......) was performed for all four patients. By combination of these methods we identified several putative susceptibility genes for ASDs. Expression patterns were established for several of these genes by Quantitative PCR (Q-PCR) or in situ hybridization and one gene was sequenced in 157 ASD patients. Our results...

  3. Expression and characterization of rice putative PAUSED gene

    Institute of Scientific and Technical Information of China (English)

    Chengguo Yao; Liangfa Ge; Wei Li; Botao Zhao; Chaoqun Li; Kangcheng Ruan; Hongxuan Lin; Youxin Jin

    2008-01-01

    In Arab idopsis, PA USED ( PSD ) encodes the ortholog of loslp/ exportin-t, which mediates the nuclear export of transfer RNA (tRNA) in yeast and mammals. However, in monocot plants such as rice, knowledge of the corresponding ortholog is limited, and its effects on growth development and productivity remain unknown. In this study, we verified a rice transfer-DNA insertional mutantpsd line and analyzed its phenotypes;the mutant displayed severe morphological defects including retarded development and low fertility compared with wild-type rice. Examining intronless tRNA-Tyr and intron-containing pre-tRNA-Ala expression levels in cytoplasmic and nuclear fraction with Northern blot analysis between wild -type and mutant leaf tissue suggested that rice PSD might be involved in tRNA export from the nucleus to the cytoplasm.Additionally, reverse transcription-polymerase chain reaction analysis revealed that PSD transcript was expressed throughout normal rice plant development, and subcellular localization assays showed that rice PSD protein was present in both the nucleus and cytoplasm. In summary, our data implied that the putative PSD gene might be indispensable for normal rice development and its function might be the same as that ofArabidopsis PSD.

  4. Isolation and characterization of 17 different genes encoding putative endopolygalacturonase genes from Rhizopus oryzae

    Science.gov (United States)

    Polygalacturonase enzymes are a valuable aid in the retting of flax for production of linens and, more recently, production of biofuels from citrus wastes. In a search of the recently sequenced Rhizopus oryzae strain 99-880 genome database, 18 putative endopolygalacturonase genes were identified, w...

  5. Putative essential and core-essential genes in Mycoplasma genomes.

    Science.gov (United States)

    Lin, Yan; Zhang, Randy Ren

    2011-01-01

    Mycoplasma, which was used to create the first "synthetic life", has been an important species in the emerging field, synthetic biology. However, essential genes, an important concept of synthetic biology, for both M. mycoides and M. capricolum, as well as 14 other Mycoplasma with available genomes, are still unknown. We have developed a gene essentiality prediction algorithm that incorporates information of biased gene strand distribution, homologous search and codon adaptation index. The algorithm, which achieved an accuracy of 80.8% and 78.9% in self-consistence and cross-validation tests, respectively, predicted 5880 essential genes in the 16 Mycoplasma genomes. The intersection set of essential genes in available Mycoplasma genomes consists of 153 core essential genes. The predicted essential genes (available from pDEG, tubic.tju.edu.cn/pdeg) and the proposed algorithm can be helpful for studying minimal Mycoplasma genomes as well as essential genes in other genomes.

  6. Putative essential and core-essential genes in Mycoplasma genomes

    OpenAIRE

    Lin, Yan; Zhang, Randy Ren

    2011-01-01

    Mycoplasma, which was used to create the first “synthetic life”, has been an important species in the emerging field, synthetic biology. However, essential genes, an important concept of synthetic biology, for both M. mycoides and M. capricolum, as well as 14 other Mycoplasma with available genomes, are still unknown. We have developed a gene essentiality prediction algorithm that incorporates information of biased gene strand distribution, homologous search and codon adaptation index. The al...

  7. Putative resistance genes in the CitEST database

    Directory of Open Access Journals (Sweden)

    Simone Guidetti-Gonzalez

    2007-01-01

    Full Text Available Disease resistance in plants is usually associated with the activation of a wide variety of defense responses to prevent pathogen replication and/or movement. The ability of the host plant to recognize the pathogen and to activate defense responses is regulated by direct or indirect interaction between the products of plant resistance (R and pathogen avirulence (Avr genes. Attempted infection of plants by avirulent pathogens elicits a battery of defenses often followed by the collapse of the challenged host cells. Localized host cell death may help to prevent the pathogen from spreading to uninfected tissues, known as hypersensitive response (HR. When either the plant or the pathogen lacks its cognate gene, activation of the plant’s defense responses fails to occur or is delayed and does not prevent pathogen colonization. In the CitEST database, we identified 1,300 reads related to R genes in Citrus which have been reported in other plant species. These reads were translated in silico, and alignments of their amino acid sequences revealed the presence of characteristic domains and motifs that are specific to R gene classes. The description of the reads identified suggests that they function as resistance genes in citrus.

  8. FKBP immunophilins and Alzheimer's disease: A chaperoned affair

    Indian Academy of Sciences (India)

    Weihuan Cao; Mary Konsolaki

    2011-08-01

    The FK506-binding protein (FKBP) family of immunophilins consists of proteins with a variety of protein–protein interaction domains and versatile cellular functions. Analysis of the functions of immunophilins has been the focus of studies in recent years and has led to the identification of various molecular pathways in which FKBPs play an active role. All FKBPs contain a domain with prolyl cis/trans isomerase (PPIase) activity. Binding of the immunosuppressant molecule FK506 to this domain inhibits their PPIase activity while mediating immune suppression through inhibition of calcineurin. The larger members, FKBP51 and FKBP52, interact with Hsp90 and exhibit chaperone activity that is shown to regulate steroid hormone signalling. From these studies it is clear that FKBP proteins are expressed ubiquitously but show relatively high levels of expression in the nervous system. Consistent with this expression, FKBPs have been implicated with both neuroprotection and neurodegeneration. This review will focus on recent studies involving FKBP immunophilins in Alzheimer’s-disease-related pathways.

  9. Expression of putative expansin genes in phylloxera (Daktulosphaira vitifoliae Fitch) induced root galls of Vitis spp.

    Science.gov (United States)

    Lawo, N C; Griesser, M; Forneck, A

    Grape phylloxera (Daktulosphaira vitifoliae Fitch) is a serious global pest in viticulture. The insects are sedentary feeders and require a gall to feed and reproduce. The insects induce their feeding site within the meristematic zone of the root tip, where they stay attached, feeding both intra- and intercellularly, and causing damage by reducing plant vigour. Several changes in cell structure and composition, including increased cell division and tissue swelling close to the feeding site, cause an organoid gall called a nodosity to develop. Because alpha expansin genes are involved in cell enlargement and cell wall loosening in many plant tissues it may be anticipated that they are also involved in nodosity formation. To identify expansin genes in Vitis vinifera cv. Pinot noir, we mined for orthologues genes in a comparative analysis. Eleven putative expansin genes were identified and shown to be present in the rootstock Teleki 5C (V. berlandieri Planch. x V. riparia Michx.) using specific PCR followed by DNA sequencing. Expression analysis of young and mature nodosities and uninfested root tips were conducted via quantitative real time PCR (qRT-PCR). Up-regulation was measured for three putative expansin genes (VvEXPA15, -A17 and partly -A20) or down-regulation for three other putative genes (VvEXPA7, -A12, -A20) in nodosities. The present study clearly shows the involvement of putative expansin genes in the phylloxera-root interaction.

  10. Putative and unique gene sequence utilization for the design of species specific probes as modeled by Lactobacillus plantarum

    Science.gov (United States)

    The concept of utilizing putative and unique gene sequences for the design of species specific probes was tested. The abundance profile of assigned functions within the Lactobacillus plantarum genome was used for the identification of the putative and unique gene sequence, csh. The targeted gene (cs...

  11. Cloning and characterization of prunus serotina AGAMOUS, a putative flower homeotic gene

    Science.gov (United States)

    Xiaomei Liu; Joseph Anderson; Paula Pijut

    2010-01-01

    Members of the AGAMOUS subfamily of MADS-box transcription factors play an important role in regulating the development of reproductive organs in flowering plants. To help understand the mechanism of floral development in black cherry (Prunus serotina), PsAG (a putative flower homeotic identity gene) was isolated...

  12. Functional characterization of a putative β-lactamase gene in the genome of Zymomonas mobilis.

    Science.gov (United States)

    Rajnish, K Narayanan; Asraf, Sheik Abdul Kader Sheik; Manju, Nagarajan; Gunasekaran, Paramasamy

    2011-12-01

    Zymomonas mobilis ZM4 is resistant to β-lactam antibiotics but there are no reports of a β-lactam resistance gene and its regulation. A putative β-lactamase gene sequence (ZMO0103) in the genome of Z. mobilis showed a 55% amino acid sequence identity with class C β-lactamase genes. qPCR analysis of the β-lactamase transcript indicated a higher level expression of the β-lactamase compared to the relative transcript quantities in antibiotic-susceptible bacteria. The putative β-lactamase gene was cloned, expressed in Escherichia coli BL21 and the product, AmpC, was purified to homogeneity. Its optimal activity was at pH 6 and 30 °C. Further, the β-lactamase had a higher affinity towards penicillins than cephalosporin antibiotics. © Springer Science+Business Media B.V. 2011

  13. Cloning and molecular characterization of a putative voltage-gated sodium channel gene in the crayfish.

    Science.gov (United States)

    Coskun, Cagil; Purali, Nuhan

    2016-06-01

    Voltage-gated sodium channel genes and associated proteins have been cloned and studied in many mammalian and invertebrate species. However, there is no data available about the sodium channel gene(s) in the crayfish, although the animal has frequently been used as a model to investigate various aspects of neural cellular and circuit function. In the present work, by using RNA extracts from crayfish abdominal ganglia samples, the complete open reading frame of a putative sodium channel gene has firstly been cloned and molecular properties of the associated peptide have been analyzed. The open reading frame of the gene has a length of 5793 bp that encodes for the synthesis of a peptide, with 1930 amino acids, that is 82% similar to the α-peptide of a sodium channel in a neighboring species, Cancer borealis. The transmembrane topology analysis of the crayfish peptide indicated a pattern of four folding domains with several transmembrane segments, as observed in other known voltage-gated sodium channels. Upon analysis of the obtained sequence, functional regions of the putative sodium channel responsible for the selectivity filter, inactivation gate, voltage sensor, and phosphorylation have been predicted. The expression level of the putative sodium channel gene, as defined by a qPCR method, was measured and found to be the highest in nervous tissue.

  14. Transcriptome Analysis Reveals Putative Genes Involved in Iridoid Biosynthesis in Rehmannia glutinosa

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    Xianen Li

    2012-10-01

    Full Text Available Rehmannia glutinosa, one of the most widely used herbal medicines in the Orient, is rich in biologically active iridoids. Despite their medicinal importance, no molecular information about the iridoid biosynthesis in this plant is presently available. To explore the transcriptome of R. glutinosa and investigate genes involved in iridoid biosynthesis, we used massively parallel pyrosequencing on the 454 GS FLX Titanium platform to generate a substantial EST dataset. Based on sequence similarity searches against the public sequence databases, the sequences were first annotated and then subjected to Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (KEGG based analysis. Bioinformatic analysis indicated that the 454 assembly contained a set of genes putatively involved in iridoid biosynthesis. Significantly, homologues of the secoiridoid pathway genes that were only identified in terpenoid indole alkaloid producing plants were also identified, whose presence implied that route II iridoids and route I iridoids share common enzyme steps in the early stage of biosynthesis. The gene expression patterns of four prenyltransferase transcripts were analyzed using qRT-PCR, which shed light on their putative functions in tissues of R. glutinosa. The data explored in this study will provide valuable information for further studies concerning iridoid biosynthesis.

  15. Transcriptome outlier analysis implicates schizophrenia susceptibility genes and enriches putatively functional rare genetic variants.

    Science.gov (United States)

    Duan, Jubao; Sanders, Alan R; Moy, Winton; Drigalenko, Eugene I; Brown, Eric C; Freda, Jessica; Leites, Catherine; Göring, Harald H H; Gejman, Pablo V

    2015-08-15

    We searched a gene expression dataset comprised of 634 schizophrenia (SZ) cases and 713 controls for expression outliers (i.e., extreme tails of the distribution of transcript expression values) with SZ cases overrepresented compared with controls. These outlier genes were enriched for brain expression and for genes known to be associated with neurodevelopmental disorders. SZ cases showed higher outlier burden (i.e., total outlier events per subject) than controls for genes within copy number variants (CNVs) associated with SZ or neurodevelopmental disorders. Outlier genes were enriched for CNVs and for rare putative regulatory variants, but this only explained a small proportion of the outlier subjects, highlighting the underlying presence of additional genetic and potentially, epigenetic mechanisms.

  16. Identification of a putative tetrasporophyte-specific gene in Gracilaria lemaneiformis (Gracilariales, Rhodophyte)

    Science.gov (United States)

    Ren, Xueying; Zhang, Xuecheng

    2008-08-01

    A putative tetrasporophyte-specific gene, designated as SSH466 (GenBank accession No. DQ019223), was one of the genes identified in this work using suppression subtractive hybridization (SSH) method in Gracilaria lemaneiformis. The full length of the gene was obtained using SMART RACE strategy. Sequence analysis revealed that the gene had 1 019 nucleotides, including an open reading frame of 498 nucleotides encoding 166 amino acid residues, 158 nucleotides of 5' untranslated region and 363 nucleotides of 3' non-coding region. Protein motif and secondary structure prediction showed that there existed a transmembrane domain with a unique β-sheet. Thus, SSH466 protein might be a cross-membrane protein. Sequence homology search in the public GenBank databases did not reveal any significant match with SSH466. Virtual Northern blot analysis confirmed that it was a tetrasporophyte-specific gene.

  17. Identification of a Putative Tetrasporophyte-Specific Gene in Gracilaria lemaneiformis(Gracilariales, Rhodophyte)

    Institute of Scientific and Technical Information of China (English)

    REN Xueying; ZHANG Xuecheng

    2008-01-01

    A putative tetrasporophyte-specific gene, designated as SSH466 (GenBank accession No. DQ019223), was one of the genes identified in this work using suppression subtractive hybridization (SSH) method in Gracilaria lemaneiformis. The full length of the gene was obtained using SMART RACE strategy. Sequence analysis revealed that the gene had 1 019 nucleotides, including an open reading frame of 498 nucleotides encoding 166 amino acid residues, 158 nucleotides of 5' untranslated region and 363 nucleo- tides of 3' non-coding region. Protein motif and secondary structure prediction showed that there existed a transmembrane domain with a unique β-sheet. Thus, SSH466 protein might be a cross-membrane protein. Sequence homology search in the public GenBank databases did not reveal any significant match with SSH466. Virtual Northern blot analysis confirmed that it was a tetrasporo- phyte-specific gene.

  18. Five putative nucleoside triphosphate diphosphohydrolase genes are expressed in Trichomonas vaginalis.

    Science.gov (United States)

    Frasson, Amanda Piccoli; Dos Santos, Odelta; Meirelles, Lúcia Collares; Macedo, Alexandre José; Tasca, Tiana

    2016-01-01

    Trichomonas vaginalis is a protozoan that parasitizes the human urogenital tract causing trichomoniasis, the most common non-viral sexually transmitted disease. The parasite has unique genomic characteristics such as a large genome size and expanded gene families. Ectonucleoside triphosphate diphosphohydrolase (E-NTPDase) is an enzyme responsible for hydrolyzing nucleoside tri- and diphosphates and has already been biochemically characterized in T. vaginalis. Considering the important role of this enzyme in the production of extracellular adenosine for parasite uptake, we evaluated the gene expression of five putative NTPDases in T. vaginalis. We showed that all five putative TvNTPDase genes (TvNTPDase1-5) were expressed by both fresh clinical and long-term grown isolates. The amino acid alignment predicted the presence of the five crucial apyrase conserved regions, transmembrane domains, signal peptides, phosphorylation and catalytic sites. Moreover, a phylogenetic analysis showed that TvNTPDase sequences make up a clade with NTPDases intracellularly located. Biochemical NTPDase activity (ATP and ADP hydrolysis) is responsive to the serum-restrictive conditions and the gene expression of TvNTPDases was mostly increased, mainly TvNTPDase2 and TvNTPDase4, although there was not a clear pattern of expression among them. In summary, the present report demonstrates the gene expression patterns of predicted NTPDases in T. vaginalis.

  19. Bioinformatic selection of putative epigenetically regulated loci associated with obesity using gene expression data.

    Science.gov (United States)

    Turcot, Valérie; Groom, Alexandra; McConnell, James C; Pearce, Mark S; Potter, Catherine; Embleton, Nicholas D; Swan, Daniel C; Relton, Caroline L

    2012-05-10

    There is considerable interest in defining the relationship between epigenetic variation and the risk of common complex diseases. Strategies which assist in the prioritisation of target loci that have the potential to be epigenetically regulated might provide a useful approach in identifying concrete examples of epigenotype-phenotype associations. Focusing on the postulated role of epigenetic factors in the aetiopathogenesis of obesity this report outlines an approach utilising gene expression data and a suite of bioinformatic tools to prioritise a list of target candidate genes for more detailed experimental scrutiny. Gene expression microarrays were performed using peripheral blood RNA from children aged 11-13years selected from the Newcastle Preterm Birth Growth Study which were grouped by body mass index (BMI). Genes showing ≥2.0 fold differential expression between low and high BMI groups were selected for in silico analysis. Several bioinformatic tools were used for each following step; 1) a literature search was carried out to identify whether the differentially expressed genes were associated with adiposity phenotypes. Of those obesity-candidate genes, putative epigenetically regulated promoters were identified by 2) defining the promoter regions, 3) then by selecting promoters with a CpG island (CGI), 4) and then by identifying any transcription factor binding modules covering CpG sites within the CGI. This bioinformatic processing culminated in the identification of a short list of target obesity-candidate genes putatively regulated by DNA methylation which can be taken forward for experimental analysis. The proposed workflow provides a flexible, versatile and low cost methodology for target gene prioritisation that is applicable to multiple species and disease contexts. Copyright © 2012. Published by Elsevier B.V.

  20. Coleopteran-specific and putative novel cry genes in Iranian native Bacillus thuringiensis collection.

    Science.gov (United States)

    Nazarian, Amin; Jahangiri, Rosa; Jouzani, Gholamreza Salehi; Seifinejad, Ali; Soheilivand, Saeed; Bagheri, Omolbanin; Keshavarzi, Mansoureh; Alamisaeid, Khalil

    2009-10-01

    The characterization of the strains containing Coleopteran-specific and also putative novel cry genes in Iranian native Bacillus thuringiensis collection is presented. Characterization was based on PCR analysis using 31 general and specific primers for cry1B, cry1I, cry3A, cry3B, cry3C, cry7A, cry8A, cry8B, cry8C, cry14, cry18, cry26, cry28, cry34 and cry35 genes, protein band patterns as well as their insecticidal activity on Xanthogaleruca luteola Mull. larvae. Forty six isolates (65.7%) contained minimum one Coleopteran-active cry gene. Based on universal primers, strains containing cry18 and cry26 genes were the most abundant and represent 27.1% and 24% of the isolates, respectively, whereas cry14, cry3, cry28, cry34, cry35, cry7, cry8 genes were less abundant, found in 14.2, 12.5, 10, 7, 7 and 5.6% of the strains, respectively. Based on specific primers, isolates containing cry1I were the most abundant (48.5%). Two strains containing Coleopteran-active cry genes showed higher activity against X. luteola larvae than B. thuringiensis subsp. morrisoni pathovar tenebrionis. Thirty isolates, when assayed for cry1C, cry5, cry6, cry8b, cry9, cry10, cry11, cry18, cry24 and cry35 genes, showed unexpected size bands. Cloning and sequencing of the amplicons allowed both the identification of known cry genes and the detection of putative novel cry1C sequences.

  1. Identification of putative orthologous genes for the phylogenetic reconstruction of temperate woody bamboos (Poaceae: Bambusoideae).

    Science.gov (United States)

    Zhang, Li-Na; Zhang, Xian-Zhi; Zhang, Yu-Xiao; Zeng, Chun-Xia; Ma, Peng-Fei; Zhao, Lei; Guo, Zhen-Hua; Li, De-Zhu

    2014-09-01

    The temperate woody bamboos (Arundinarieae) are highly diverse in morphology but lack a substantial amount of genetic variation. The taxonomy of this lineage is intractable, and the relationships within the tribe have not been well resolved. Recent studies indicated that this tribe could have a complex evolutionary history. Although phylogenetic studies of the tribe have been carried out, most of these phylogenetic reconstructions were based on plastid data, which provide lower phylogenetic resolution compared with nuclear data. In this study, we intended to identify a set of desirable nuclear genes for resolving the phylogeny of the temperate woody bamboos. Using two different methodologies, we identified 209 and 916 genes, respectively, as putative single copy orthologous genes. A total of 112 genes was successfully amplified and sequenced by next-generation sequencing technologies in five species sampled from the tribe. As most of the genes exhibited intra-individual allele heterozygotes, we investigated phylogenetic utility by reconstructing the phylogeny based on individual genes. Discordance among gene trees was observed and, to resolve the conflict, we performed a range of analyses using BUCKy and HybTree. While caution should be taken when inferring a phylogeny from multiple conflicting genes, our analysis indicated that 74 of the 112 investigated genes are potential markers for resolving the phylogeny of the temperate woody bamboos. © 2014 John Wiley & Sons Ltd.

  2. Characterization of Putative cis-Regulatory Elements in Genes Preferentially Expressed in Arabidopsis Male Meiocytes

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    Junhua Li

    2014-01-01

    Full Text Available Meiosis is essential for plant reproduction because it is the process during which homologous chromosome pairing, synapsis, and meiotic recombination occur. The meiotic transcriptome is difficult to investigate because of the size of meiocytes and the confines of anther lobes. The recent development of isolation techniques has enabled the characterization of transcriptional profiles in male meiocytes of Arabidopsis. Gene expression in male meiocytes shows unique features. The direct interaction of transcription factors (TFs with DNA regulatory sequences forms the basis for the specificity of transcriptional regulation. Here, we identified putative cis-regulatory elements (CREs associated with male meiocyte-expressed genes using in silico tools. The upstream regions (1 kb of the top 50 genes preferentially expressed in Arabidopsis meiocytes possessed conserved motifs. These motifs are putative binding sites of TFs, some of which share common functions, such as roles in cell division. In combination with cell-type-specific analysis, our findings could be a substantial aid for the identification and experimental verification of the protein-DNA interactions for the specific TFs that drive gene expression in meiocytes.

  3. Identification and functional analysis of Penicillium digitatum genes putatively involved in virulence towards citrus fruit.

    Science.gov (United States)

    López-Pérez, Mario; Ballester, Ana-Rosa; González-Candelas, Luis

    2015-04-01

    The fungus Penicillium digitatum, the causal agent of green mould rot, is the most destructive post-harvest pathogen of citrus fruit in Mediterranean regions. In order to identify P. digitatum genes up-regulated during the infection of oranges that may constitute putative virulence factors, we followed a polymerase chain reaction (PCR)-based suppression subtractive hybridization and cDNA macroarray hybridization approach. The origin of expressed sequence tags (ESTs) was determined by comparison against the available genome sequences of both organisms. Genes coding for fungal proteases and plant cell wall-degrading enzymes represent the largest categories in the subtracted cDNA library. Northern blot analysis of a selection of P. digitatum genes, including those coding for proteases, cell wall-related enzymes, redox homoeostasis and detoxification processes, confirmed their up-regulation at varying time points during the infection process. Agrobacterium tumefaciens-mediated transformation was used to generate knockout mutants for two genes encoding a pectin lyase (Pnl1) and a naphthalene dioxygenase (Ndo1). Two independent P. digitatum Δndo1 mutants were as virulent as the wild-type. However, the two Δpnl1 mutants analysed were less virulent than the parental strain or an ectopic transformant. Together, these results provide a significant advance in our understanding of the putative determinants of the virulence mechanisms of P. digitatum.

  4. Analysis of the expression of putatively imprinted genes in bovine peri-implantation embryos

    DEFF Research Database (Denmark)

    Tveden-Nyborg, Pernille Yde; Alexopoulos, N.I.; Cooney, M.A.

    2008-01-01

    imprinted genes (Ata3, Dlk1, Gnas, Grb10, Magel2, Mest-1, Ndn and Sgce) in bovine peri-implantation embryos. Two embryonic developmental stages were examined, Day 14 and Day 21. The gene expression pattern of single embryos was recorded for in vivo, in vitro produced (IVP) and parthenogenetic embryos...... (LOS) in bovine embryos resulting in increased embryonic morbidity and mortality. In the bovine, limited numbers of imprinted genes have been studied and studies have primarily been restricted to pre-implantation stages. This study reports original data on the expression pattern of 8 putatively...... procedures, either by in vitro maturation, fertilization or culture. In conclusion, effects of genomic imprinting and of in vitro procedures for embryo production may influence the success of bovine embryo implantation....

  5. Novel mutations in the GH gene (GH1) uncover putative splicing regulatory elements.

    Science.gov (United States)

    Babu, Deepak; Mellone, Simona; Fusco, Ileana; Petri, Antonella; Walker, Gillian E; Bellone, Simonetta; Prodam, Flavia; Momigliano-Richiardi, Patricia; Bona, Gianni; Giordano, Mara

    2014-05-01

    Mutations affecting exon 3 splicing are the main cause of autosomal dominant Isolated GH Deficiency II (IGHDII) by increasing the level of exon 3-skipped mRNA encoding the functionally inactive dominant-negative 17.5-kDa isoform. The exons and introns of the gene encoding GH (GH1) were screened for the presence of mutations in 103 sporadic isolated GH deficiency cases. Four different variations within exon 3 were identified in 3 patients. One carried c.261C>T (p.Pro87Pro) and c.272A>T (p.Glu91Val), the second c.255G>A (p.Pro85Pro) and c.261 C>T, and the third c.246G>C (p.Glu82Asp). All the variants were likely generated by gene conversion from an homologous gene in the GH1 cluster. In silico analysis predicted that positions c.255 and c.272 were included within 2 putative novel exon splicing enhancers (ESEs). Their effect on splicing was confirmed in vitro. Constructs bearing these 2 variants induced consistently higher levels both of transcript and protein corresponding to the 17.5-kDa isoform. When c.255 and c.272 were combined in cis with the c.261 variant, as in our patients, their effect was weaker. In conclusion, we identified 2 variations, c.255G>A and c.272A>T, located in 2 novel putative exon splicing enhancers and affecting GH1 splicing in vitro by increasing the production of alternatively spliced isoforms. The amount of aberrant isoforms is further regulated by the presence in cis of the c.261 variant. Thus, our results evidenced novel putative splicing regulatory elements within exon 3, confirming the crucial role of this exon in mRNA processing.

  6. Two putative-aquaporin genes are differentially expressed during arbuscular mycorrhizal symbiosis in Lotus japonicus

    Science.gov (United States)

    2012-01-01

    Background Arbuscular mycorrhizas (AM) are widespread symbioses that provide great advantages to the plant, improving its nutritional status and allowing the fungus to complete its life cycle. Nevertheless, molecular mechanisms that lead to the development of AM symbiosis are not yet fully deciphered. Here, we have focused on two putative aquaporin genes, LjNIP1 and LjXIP1, which resulted to be upregulated in a transcriptomic analysis performed on mycorrhizal roots of Lotus japonicus. Results A phylogenetic analysis has shown that the two putative aquaporins belong to different functional families: NIPs and XIPs. Transcriptomic experiments have shown the independence of their expression from their nutritional status but also a close correlation with mycorrhizal and rhizobial interaction. Further transcript quantification has revealed a good correlation between the expression of one of them, LjNIP1, and LjPT4, the phosphate transporter which is considered a marker gene for mycorrhizal functionality. By using laser microdissection, we have demonstrated that one of the two genes, LjNIP1, is expressed exclusively in arbuscule-containing cells. LjNIP1, in agreement with its putative role as an aquaporin, is capable of transferring water when expressed in yeast protoplasts. Confocal analysis have demonstrated that eGFP-LjNIP1, under its endogenous promoter, accumulates in the inner membrane system of arbusculated cells. Conclusions Overall, the results have shown different functionality and expression specificity of two mycorrhiza-inducible aquaporins in L. japonicus. One of them, LjNIP1 can be considered a novel molecular marker of mycorrhizal status at different developmental stages of the arbuscule. At the same time, LjXIP1 results to be the first XIP family aquaporin to be transcriptionally regulated during symbiosis. PMID:23046713

  7. Two putative-aquaporin genes are differentially expressed during arbuscular mycorrhizal symbiosis in Lotus japonicus

    Directory of Open Access Journals (Sweden)

    Giovannetti Marco

    2012-10-01

    Full Text Available Abstract Background Arbuscular mycorrhizas (AM are widespread symbioses that provide great advantages to the plant, improving its nutritional status and allowing the fungus to complete its life cycle. Nevertheless, molecular mechanisms that lead to the development of AM symbiosis are not yet fully deciphered. Here, we have focused on two putative aquaporin genes, LjNIP1 and LjXIP1, which resulted to be upregulated in a transcriptomic analysis performed on mycorrhizal roots of Lotus japonicus. Results A phylogenetic analysis has shown that the two putative aquaporins belong to different functional families: NIPs and XIPs. Transcriptomic experiments have shown the independence of their expression from their nutritional status but also a close correlation with mycorrhizal and rhizobial interaction. Further transcript quantification has revealed a good correlation between the expression of one of them, LjNIP1, and LjPT4, the phosphate transporter which is considered a marker gene for mycorrhizal functionality. By using laser microdissection, we have demonstrated that one of the two genes, LjNIP1, is expressed exclusively in arbuscule-containing cells. LjNIP1, in agreement with its putative role as an aquaporin, is capable of transferring water when expressed in yeast protoplasts. Confocal analysis have demonstrated that eGFP-LjNIP1, under its endogenous promoter, accumulates in the inner membrane system of arbusculated cells. Conclusions Overall, the results have shown different functionality and expression specificity of two mycorrhiza-inducible aquaporins in L. japonicus. One of them, LjNIP1 can be considered a novel molecular marker of mycorrhizal status at different developmental stages of the arbuscule. At the same time, LjXIP1 results to be the first XIP family aquaporin to be transcriptionally regulated during symbiosis.

  8. Mutation analysis of novel human liver-related putative tumor suppressor gene in hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Cheng Liao; Tsai-Ping Li; Mu-Jun Zhao; Jing Zhao; Hai Song; Pascal Pineau; Agnès Marchio; Anne Dejean; Pierre Tiollais; Hong-Yang Wang

    2003-01-01

    AIM: To find the point mutations meaningful for inactivationof liver-related putative tumor suppressor gene (LPTS) gene,a human novel liver-related putative tumor suppressor geneand telomerase inhibitor in hepatocellular carcinoma.METHODS: The entire coding sequence of LPTS genewas examined for mutations by single strand conformationpolymorphism (SSCP) assay and PCR products directsequencing in 56 liver cancer cell lines, 7 ovarian cancerand 7 head & neck tumor cell lines and 70 pairs of HCCtissues samples. The cDNA fragment coding for the mostfrequent mutant protein was subcloned into GST fusionexpression vector. The product was expressed in E. coliand purified by glutathione-agarose column. Telomericrepeat amplification protocol (TRAP) assays wereperformed to study the effect of point mutation totelomerase inhibitory activity.RESULTS: SSCP gels showed the abnormal shifting bandsand DNA sequencing found that there were 5 differentmutations and/or polymorphisms in 12 tumor cell lineslocated at exon2, exon5 and exon7. The main alterationswere A(778)A/G and A(880)T in exon7. The change in siteof 778 could not be found in HCC tissue samples, while themutation in position 880 was seen in 7 (10 %) cases. Themutation in the site of 880 had no effect on telomeraseinhibitory activity.CONCLUSION: Alterations identified in this study arepolymorphisms of LPTS gene. LPTS mutations occur in HCCbut are infrequent and of little effect on the telomeraseinhibitory function of the protein. Epigenetics, such asmethylation, acetylation, may play the key role in inactivationof LPTS.

  9. Machine learning techniques to identify putative genes involved in nitrogen catabolite repression in the yeast Saccharomyces cerevisiae

    Science.gov (United States)

    Kontos, Kevin; Godard, Patrice; André, Bruno; van Helden, Jacques; Bontempi, Gianluca

    2008-01-01

    Background Nitrogen is an essential nutrient for all life forms. Like most unicellular organisms, the yeast Saccharomyces cerevisiae transports and catabolizes good nitrogen sources in preference to poor ones. Nitrogen catabolite repression (NCR) refers to this selection mechanism. All known nitrogen catabolite pathways are regulated by four regulators. The ultimate goal is to infer the complete nitrogen catabolite pathways. Bioinformatics approaches offer the possibility to identify putative NCR genes and to discard uninteresting genes. Results We present a machine learning approach where the identification of putative NCR genes in the yeast Saccharomyces cerevisiae is formulated as a supervised two-class classification problem. Classifiers predict whether genes are NCR-sensitive or not from a large number of variables related to the GATA motif in the upstream non-coding sequences of the genes. The positive and negative training sets are composed of annotated NCR genes and manually-selected genes known to be insensitive to NCR, respectively. Different classifiers and variable selection methods are compared. We show that all classifiers make significant and biologically valid predictions by comparing these predictions to annotated and putative NCR genes, and by performing several negative controls. In particular, the inferred NCR genes significantly overlap with putative NCR genes identified in three genome-wide experimental and bioinformatics studies. Conclusion These results suggest that our approach can successfully identify potential NCR genes. Hence, the dimensionality of the problem of identifying all genes involved in NCR is drastically reduced. PMID:19091052

  10. Expression of putative immune response genes during early ontogeny in the coral Acropora millepora.

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    Eneour Puill-Stephan

    Full Text Available BACKGROUND: Corals, like many other marine invertebrates, lack a mature allorecognition system in early life history stages. Indeed, in early ontogeny, when corals acquire and establish associations with various surface microbiota and dinoflagellate endosymbionts, they do not efficiently distinguish between closely and distantly related individuals from the same population. However, very little is known about the molecular components that underpin allorecognition and immunity responses or how they change through early ontogeny in corals. METHODOLOGY/PRINCIPAL FINDINGS: Patterns in the expression of four putative immune response genes (apextrin, complement C3, and two CELIII type lectin genes were examined in juvenile colonies of Acropora millepora throughout a six-month post-settlement period using quantitative real-time PCR (qPCR. Expression of a CELIII type lectin gene peaked in the fourth month for most of the coral juveniles sampled and was significantly higher at this time than at any other sampling time during the six months following settlement. The timing of this increase in expression levels of putative immune response genes may be linked to allorecognition maturation which occurs around this time in A. millepora. Alternatively, the increase may represent a response to immune challenges, such as would be involved in the recognition of symbionts (such as Symbiodinium spp. or bacteria during winnowing processes as symbioses are fine-tuned. CONCLUSIONS/SIGNIFICANCE: Our data, although preliminary, are consistent with the hypothesis that lectins may play an important role in the maturation of allorecognition responses in corals. The co-expression of lectins with apextrin during development of coral juveniles also raises the possibility that these proteins, which are components of innate immunity in other invertebrates, may influence the innate immune systems of corals through a common pathway or system. However, further studies

  11. Putative cross-kingdom horizontal gene transfer in sponge (Porifera mitochondria

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    Ilan Micha

    2006-09-01

    Full Text Available Abstract Background The mitochondrial genome of Metazoa is usually a compact molecule without introns. Exceptions to this rule have been reported only in corals and sea anemones (Cnidaria, in which group I introns have been discovered in the cox1 and nad5 genes. Here we show several lines of evidence demonstrating that introns can also be found in the mitochondria of sponges (Porifera. Results A 2,349 bp fragment of the mitochondrial cox1 gene was sequenced from the sponge Tetilla sp. (Spirophorida. This fragment suggests the presence of a 1143 bp intron. Similar to all the cnidarian mitochondrial introns, the putative intron has group I intron characteristics. The intron is present in the cox1 gene and encodes a putative homing endonuclease. In order to establish the distribution of this intron in sponges, the cox1 gene was sequenced from several representatives of the demosponge diversity. The intron was found only in the sponge order Spirophorida. A phylogenetic analysis of the COI protein sequence and of the intron open reading frame suggests that the intron may have been transmitted horizontally from a fungus donor. Conclusion Little is known about sponge-associated fungi, although in the last few years the latter have been frequently isolated from sponges. We suggest that the horizontal gene transfer of a mitochondrial intron was facilitated by a symbiotic relationship between fungus and sponge. Ecological relationships are known to have implications at the genomic level. Here, an ecological relationship between sponge and fungus is suggested based on the genomic analysis.

  12. Putative cross-kingdom horizontal gene transfer in sponge (Porifera) mitochondria

    Science.gov (United States)

    Rot, Chagai; Goldfarb, Itay; Ilan, Micha; Huchon, Dorothée

    2006-01-01

    Background The mitochondrial genome of Metazoa is usually a compact molecule without introns. Exceptions to this rule have been reported only in corals and sea anemones (Cnidaria), in which group I introns have been discovered in the cox1 and nad5 genes. Here we show several lines of evidence demonstrating that introns can also be found in the mitochondria of sponges (Porifera). Results A 2,349 bp fragment of the mitochondrial cox1 gene was sequenced from the sponge Tetilla sp. (Spirophorida). This fragment suggests the presence of a 1143 bp intron. Similar to all the cnidarian mitochondrial introns, the putative intron has group I intron characteristics. The intron is present in the cox1 gene and encodes a putative homing endonuclease. In order to establish the distribution of this intron in sponges, the cox1 gene was sequenced from several representatives of the demosponge diversity. The intron was found only in the sponge order Spirophorida. A phylogenetic analysis of the COI protein sequence and of the intron open reading frame suggests that the intron may have been transmitted horizontally from a fungus donor. Conclusion Little is known about sponge-associated fungi, although in the last few years the latter have been frequently isolated from sponges. We suggest that the horizontal gene transfer of a mitochondrial intron was facilitated by a symbiotic relationship between fungus and sponge. Ecological relationships are known to have implications at the genomic level. Here, an ecological relationship between sponge and fungus is suggested based on the genomic analysis. PMID:16972986

  13. Identification and characterization of a gene encoding a putative lysophosphatidyl acyltransferase from Arachis hypogaea

    Indian Academy of Sciences (India)

    Si-Long Chen; Jia-Quan Huang; Lei Yong; Yue-Ting Zhang; Xiao-Ping Ren; Yu-Ning Chen; Hui-Fang Jiang; Li-Ying Yan; Yu-Rong Li; Bo-Shou Liao

    2012-12-01

    Lysophosphatidyl acyltransferase (LPAT) is the important enzyme responsible for the acylation of lysophosphatidic acid (LPA), leading to the generation of phosphatidic acid (PA) in plant. Its encoding gene is an essential candidate for oil crops to improve oil composition and increase seed oil content through genetic engineering. In this study, a full-length AhLPAT4 gene was isolated via cDNA library screening and rapid amplification of cDNA ends (RACE); our data demonstrated that AhLPAT4 had 1631 nucleotides, encoding a putative 43.8 kDa protein with 383 amino acid residues. The deduced protein included a conserved acyltransferase domain and four motifs (I–IV) with putative LPA and acyl-CoA catalytic and binding sites. Bioinformatic analysis indicated that AhLPAT4 contained four transmembrane domains (TMDs), localized to the endoplasmic reticulum (ER) membrane; detailed analysis indicated that motif I and motifs II–III in AhLPAT4 were separated by the third TMD, which located on cytosolic and ER luminal side respectively, and hydrophobic residues on the surface of AhLPAT4 protein fold to form a hydrophobic tunnel to accommodate the acyl chain. Subcellular localization analysis confirmed that AhLPAT4 was a cytoplasm protein. Phylogenetic analysis revealed that AhLPAT4 had a high homology (63.7–78.3%) with putative LPAT4 proteins from Glycine max, Arabidopsis thaliana and Ricinus communis. AhLPAT4 was ubiquitously expressed in diverse tissues except in flower, which is almost undetectable. The expression analysis in different developmental stages in peanut seeds indicated that AhLPAT4 did not coincide with oil accumulation.

  14. High-Resolution Genomic and Expression Profiling Reveals 105 Putative Amplification Target Genes in Pancreatic Cancer

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    Eija H. Mahlamaki

    2004-09-01

    Full Text Available Comparative genomic hybridization (CGH studies have provided a wealth of information on common copy number aberrations in pancreatic cancer, but the genes affected by these aberrations are largely unknown. To identify putative amplification target genes in pancreatic cancer, we performed a parallel copy number and expression survey in 13 pancreatic cancer cell lines using a 12,232-clone cDNA microarray, providing an average resolution of 300 kb throughout the human genome. CGH on cDNA microarray allowed highly accurate mapping of copy number increases and resulted in identification of 24 independent amplicons, ranging in size from 130 kb to 11 Mb. Statistical evaluation of gene copy number and expression data across all 13 cell lines revealed a set of 105 genes whose elevated expression levels were directly attributable to increased copy number. These included genes previously reported to be amplified in cancer as well as several novel targets for copy number alterations, such as p21-activated kinase 4 (PAK4, which was previously shown to be involved in cell migration, cell adhesion, and anchorage-independent growth. In conclusion, our results implicate a set of 105 genes that is likely to be actively involved in the development and progression of pancreatic cancer.

  15. Identification of putative candidate genes for juvenile wood density in Pinus radiata.

    Science.gov (United States)

    Li, Xinguo; Wu, Harry X; Southerton, Simon G

    2012-08-01

    Wood formation is a complex developmental process driven by the annual activity of the vascular cambium. Conifers usually produce juvenile wood at young ages followed by mature wood for the rest of their lifetime. Juvenile wood exhibits poorer wood quality (i.e., lower density) compared with mature wood and can account for up to 50% of short-rotation harvested logs, thus representing a major challenge for commercial forestry globally. Wood density is an important quality trait for many timber-related products. Understanding the molecular mechanisms involved in the regulation of juvenile wood density is critical for the improvement of juvenile wood quality via marker-aided selection. A previous study has identified several candidate genes affecting mature wood density in Picea sitchensis (Bong.) Carr.; however, genes associated with juvenile wood density in conifers remain poorly characterized. Here, cDNA microarrays containing 3320 xylem unigenes were used to investigate genes differentially transcribed in juvenile wood with high (HD) and low density (LD) in Pinus radiata D.Don. In total, 814 xylem unigenes with differential transcription were identified in at least one of two microarray experiments and 73 genes (45 for HD, 28 for LD) were identified in both experiments, thus representing putative candidate genes for juvenile wood density. Interestingly, cellulose synthases (PrCesA3, PrCesA11) and sucrose synthase (SuSy), which are involved in secondary cell wall formation, had stronger transcription in juvenile wood with HD, while genes functioning in primary wall formation (pectin synthesis, cell expansion and other modifications) were more transcribed in LD wood. Cell wall genes encoding monolignol biosynthesis enzymes, arabinogalactan proteins, actins and tubulins were differentially transcribed in either HD or LD juvenile wood; however, the latter had exclusively greater transcription of genes involved in monolignol polymerization (laccase and peroxidase). The

  16. Genetic analysis of BIRC4/XIAP as a putative modifier gene of Wilson disease.

    Science.gov (United States)

    Weiss, Karl Heinz; Runz, Heiko; Noe, Barbara; Gotthardt, Daniel Nils; Merle, Uta; Ferenci, Peter; Stremmel, Wolfgang; Füllekrug, Joachim

    2010-12-01

    Wilson disease (WD) is an autosomal-recessive copper overload disorder caused by mutations in the copper-transporting adenosine triphosphatase (ATPase) ATP7B. It presents with a highly variable clinical phenotype ranging from asymptomatic to fulminant hepatic failure or progressive neurological involvement. No clear genotype-phenotype correlation has been established. Thus, variants in modifier genes could have an impact on WD manifestation and severity. Recently, the antiapoptotic protein baculoviral IAP repeat-containing protein 4 BIRC4/XIAP has been suggested as a regulator of copper-induced cell death. With the aim of investigating a putative role of BIRC4/XIAP as modifier gene in individuals with copper overload, we analyzed a WD patient cohort (n = 98) for sequence variants at the BIRC4/XIAP locus. When compared with clinical data, the previously described coding single nucleotide polymorphisms (SNPs) at the BRIC4/XIAP locus (rs28382721, rs28382722, rs28382723, rs5956583, rs28382740, rs12838858, rs28382741) did not correlate with age of onset or clinical presentation in our collective. However, three previously unreported variants in the BIRC4/XIAP gene were identified (c.1-26 T > G; c.1408A > T; p.T470S; c.1019A > G; p.N340S). The two patients with variants leading to amino acid exchanges in the BIRC4/XIAP protein showed a remarkably early disease onset at the age of 5 years. Furthermore, one of these patients was only heterozygous for disease-causing mutations in the ATP7B gene. In summary, these data emphasize the need to further elucidate a role of BIRC4/XIAP variants as putative pathogenetic factors in copper overload disorders.

  17. High amino acid diversity and positive selection at a putative coral immunity gene (tachylectin-2

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    Hellberg Michael E

    2010-05-01

    Full Text Available Abstract Background Genes involved in immune functions, including pathogen recognition and the activation of innate defense pathways, are among the most genetically variable known, and the proteins that they encode are often characterized by high rates of amino acid substitutions, a hallmark of positive selection. The high levels of variation characteristic of immunity genes make them useful tools for conservation genetics. To date, highly variable immunity genes have yet to be found in corals, keystone organisms of the world's most diverse marine ecosystem, the coral reef. Here, we examine variation in and selection on a putative innate immunity gene from Oculina, a coral genus previously used as a model for studies of coral disease and bleaching. Results In a survey of 244 Oculina alleles, we find high nonsynonymous variation and a signature of positive selection, consistent with a putative role in immunity. Using computational protein structure prediction, we generate a structural model of the Oculina protein that closely matches the known structure of tachylectin-2 from the Japanese horseshoe crab (Tachypleus tridentatus, a protein with demonstrated function in microbial recognition and agglutination. We also demonstrate that at least three other genera of anthozoan cnidarians (Acropora, Montastrea and Nematostella possess proteins structurally similar to tachylectin-2. Conclusions Taken together, the evidence of high amino acid diversity, positive selection and structural correspondence to the horseshoe crab tachylectin-2 suggests that this protein is 1 part of Oculina's innate immunity repertoire, and 2 evolving adaptively, possibly under selective pressure from coral-associated microorganisms. Tachylectin-2 may serve as a candidate locus to screen coral populations for their capacity to respond adaptively to future environmental change.

  18. An Sp185/333 gene cluster from the purple sea urchin and putative microsatellite-mediated gene diversification

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    Buckley Katherine M

    2010-10-01

    Full Text Available Abstract Background The immune system of the purple sea urchin, Strongylocentrotus purpuratus, is complex and sophisticated. An important component of sea urchin immunity is the Sp185/333 gene family, which is significantly upregulated in immunologically challenged animals. The Sp185/333 genes are less than 2 kb with two exons and are members of a large diverse family composed of greater than 40 genes. The S. purpuratus genome assembly, however, contains only six Sp185/333 genes. This underrepresentation could be due to the difficulties that large gene families present in shotgun assembly, where multiple similar genes can be collapsed into a single consensus gene. Results To understand the genomic organization of the Sp185/333 gene family, a BAC insert containing Sp185/333 genes was assembled, with careful attention to avoiding artifacts resulting from collapse or artificial duplication/expansion of very similar genes. Twelve candidate BAC assemblies were generated with varying parameters and the optimal assembly was identified by PCR, restriction digests, and subclone sequencing. The validated assembly contained six Sp185/333 genes that were clustered in a 34 kb region at one end of the BAC with five of the six genes tightly clustered within 20 kb. The Sp185/333 genes in this cluster were no more similar to each other than to previously sequenced Sp185/333 genes isolated from three different animals. This was unexpected given their proximity and putative effects of gene homogenization in closely linked, similar genes. All six genes displayed significant similarity including both 5' and 3' flanking regions, which were bounded by microsatellites. Three of the Sp185/333 genes and their flanking regions were tandemly duplicated such that each repeated segment consisted of a gene plus 0.7 kb 5' and 2.4 kb 3' of the gene (4.5 kb total. Both edges of the segmental duplications were bounded by different microsatellites. Conclusions The high sequence

  19. Discovery of Putative Herbicide Resistance Genes and Its Regulatory Network in Chickpea Using Transcriptome Sequencing

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    Mir A. Iquebal

    2017-06-01

    Full Text Available Background: Chickpea (Cicer arietinum L. contributes 75% of total pulse production. Being cheaper than animal protein, makes it important in dietary requirement of developing countries. Weed not only competes with chickpea resulting into drastic yield reduction but also creates problem of harboring fungi, bacterial diseases and insect pests. Chemical approach having new herbicide discovery has constraint of limited lead molecule options, statutory regulations and environmental clearance. Through genetic approach, transgenic herbicide tolerant crop has given successful result but led to serious concern over ecological safety thus non-transgenic approach like marker assisted selection is desirable. Since large variability in tolerance limit of herbicide already exists in chickpea varieties, thus the genes offering herbicide tolerance can be introgressed in variety improvement programme. Transcriptome studies can discover such associated key genes with herbicide tolerance in chickpea.Results: This is first transcriptomic studies of chickpea or even any legume crop using two herbicide susceptible and tolerant genotypes exposed to imidazoline (Imazethapyr. Approximately 90 million paired-end reads generated from four samples were processed and assembled into 30,803 contigs using reference based assembly. We report 6,310 differentially expressed genes (DEGs, of which 3,037 were regulated by 980 miRNAs, 1,528 transcription factors associated with 897 DEGs, 47 Hub proteins, 3,540 putative Simple Sequence Repeat-Functional Domain Marker (SSR-FDM, 13,778 genic Single Nucleotide Polymorphism (SNP putative markers and 1,174 Indels. Randomly selected 20 DEGs were validated using qPCR. Pathway analysis suggested that xenobiotic degradation related gene, glutathione S-transferase (GST were only up-regulated in presence of herbicide. Down-regulation of DNA replication genes and up-regulation of abscisic acid pathway genes were observed. Study further reveals

  20. Pyrosequencing of the Camptotheca acuminata transcriptome reveals putative genes involved in camptothecin biosynthesis and transport

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    Sun Yongzhen

    2011-10-01

    Full Text Available Abstract Background Camptotheca acuminata is a Nyssaceae plant, often called the "happy tree", which is indigenous in Southern China. C. acuminata produces the terpenoid indole alkaloid, camptothecin (CPT, which exhibits clinical effects in various cancer treatments. Despite its importance, little is known about the transcriptome of C. acuminata and the mechanism of CPT biosynthesis, as only few nucleotide sequences are included in the GenBank database. Results From a constructed cDNA library of young C. acuminata leaves, a total of 30,358 unigenes, with an average length of 403 bp, were obtained after assembly of 74,858 high quality reads using GS De Novo assembler software. Through functional annotation, a total of 21,213 unigenes were annotated at least once against the NCBI nucleotide (Nt, non-redundant protein (Nr, Uniprot/SwissProt, Kyoto Encyclopedia of Genes and Genomes (KEGG, and Arabidopsis thaliana proteome (TAIR databases. Further analysis identified 521 ESTs representing 20 enzyme genes that are involved in the backbone of the CPT biosynthetic pathway in the library. Three putative genes in the upstream pathway, including genes for geraniol-10-hydroxylase (CaPG10H, secologanin synthase (CaPSCS, and strictosidine synthase (CaPSTR were cloned and analyzed. The expression level of the three genes was also detected using qRT-PCR in C. acuminata. With respect to the branch pathway of CPT synthesis, six cytochrome P450s transcripts were selected as candidate transcripts by detection of transcript expression in different tissues using qRT-PCR. In addition, one glucosidase gene was identified that might participate in CPT biosynthesis. For CPT transport, three of 21 transcripts for multidrug resistance protein (MDR transporters were also screened from the dataset by their annotation result and gene expression analysis. Conclusion This study produced a large amount of transcriptome data from C. acuminata by 454 pyrosequencing. According to

  1. Identification and developmental expression profiling of putative alkaloid biosynthetic genes in Corydalis yanhusuo bulbs.

    Science.gov (United States)

    Liao, Dengqun; Wang, Pengfei; Jia, Chan; Sun, Peng; Qi, Jianjun; Zhou, Lili; Li, Xian'en

    2016-01-18

    Alkaloids in bulbs of Corydalis (C.) yanhusuo are the major pharmacologically active compounds in treatment of blood vessel diseases, tumors and various pains. However, due to the absence of gene sequences in C. yanhusuo, the genes involved in alkaloid biosynthesis and their expression during bulb development remain unknown. We therefore established the first transcriptome database of C. yanhusuo via Illumina mRNA-Sequencing of a RNA composite sample collected at Bulb initiation (Day 0), early enlargement (Day 10) and maturation (Day 30). 25,013,630 clean 90 bp paired-end reads were de novo assembled into 47,081 unigenes with an average length of 489 bp, among which 30,868 unigenes (65.56%) were annotated in four protein databases. Of 526 putative unigenes involved in biosynthesis o f various alkaloids, 187 were identified as the candidate genes involved in the biosynthesis of benzylisoquinoline alkaloids (BIAs), the only alkaloid type reported in C. yanhusuo untill now. BIAs biosynthetic genes were highly upregulated in the overall pathway during bulb development. Identification of alkaloid biosynthetic genes in C. yanhusuo provide insights on pathways and molecular regulation of alkaloid biosynthesis, to initiate metabolic engineering in order to improve the yield of interesting alkaloids and to identify potentially new alkaloids predicted from the transcriptomic information.

  2. ESTs analysis reveals putative genes involved in symbiotic seed germination in Dendrobium officinale.

    Directory of Open Access Journals (Sweden)

    Ming-Ming Zhao

    Full Text Available Dendrobiumofficinale (Orchidaceae is one of the world's most endangered plants with great medicinal value. In nature, D. officinale seeds must establish symbiotic relationships with fungi to germinate. However, the molecular events involved in the interaction between fungus and plant during this process are poorly understood. To isolate the genes involved in symbiotic germination, a suppression subtractive hybridization (SSH cDNA library of symbiotically germinated D. officinale seeds was constructed. From this library, 1437 expressed sequence tags (ESTs were clustered to 1074 Unigenes (including 902 singletons and 172 contigs, which were searched against the NCBI non-redundant (NR protein database (E-value cutoff, e(-5. Based on sequence similarity with known proteins, 579 differentially expressed genes in D. officinale were identified and classified into different functional categories by Gene Ontology (GO, Clusters of orthologous Groups of proteins (COGs and Kyoto Encyclopedia of Genes and Genomes (KEGG pathways. The expression levels of 15 selected genes emblematic of symbiotic germination were confirmed via real-time quantitative PCR. These genes were classified into various categories, including defense and stress response, metabolism, transcriptional regulation, transport process and signal transduction pathways. All transcripts were upregulated in the symbiotically germinated seeds (SGS. The functions of these genes in symbiotic germination were predicted. Furthermore, two fungus-induced calcium-dependent protein kinases (CDPKs, which were upregulated 6.76- and 26.69-fold in SGS compared with un-germinated seeds (UGS, were cloned from D. officinale and characterized for the first time. This study provides the first global overview of genes putatively involved in D. officinale symbiotic seed germination and provides a foundation for further functional research regarding symbiotic relationships in orchids.

  3. Putative DNA G-quadruplex formation within the promoters of Plasmodium falciparum var genes

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    Rowe J

    2009-08-01

    Full Text Available Abstract Background Guanine-rich nucleic acid sequences are capable of folding into an intramolecular four-stranded structure called a G-quadruplex. When found in gene promoter regions, G-quadruplexes can downregulate gene expression, possibly by blocking the transcriptional machinery. Here we have used a genome-wide bioinformatic approach to identify Putative G-Quadruplex Sequences (PQS in the Plasmodium falciparum genome, along with biophysical techniques to examine the physiological stability of P. falciparum PQS in vitro. Results We identified 63 PQS in the non-telomeric regions of the P. falciparum clone 3D7. Interestingly, 16 of these PQS occurred in the upstream region of a subset of the P. falciparum var genes (group B var genes. The var gene family encodes PfEMP1, the parasite's major variant antigen and adhesin expressed at the surface of infected erythrocytes, that plays a key role in malaria pathogenesis and immune evasion. The ability of the PQS found in the upstream regions of group B var genes (UpsB-Q to form stable G-quadruplex structures in vitro was confirmed using 1H NMR, circular dichroism, UV spectroscopy, and thermal denaturation experiments. Moreover, the synthetic compound BOQ1 that shows a higher affinity for DNA forming quadruplex rather than duplex structures was found to bind with high affinity to the UpsB-Q. Conclusion This is the first demonstration of non-telomeric PQS in the genome of P. falciparum that form stable G-quadruplexes under physiological conditions in vitro. These results allow the generation of a novel hypothesis that the G-quadruplex sequences in the upstream regions of var genes have the potential to play a role in the transcriptional control of this major virulence-associated multi-gene family.

  4. Isolation of a rice gene homologous to the human putative tumor suppressor gene QM

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    QM gene was originally isolated from human by Dowdy et al during a search for a wilms′ tumor suppressor gene. Researches of QM gene focused mainly on animals and yeasts, little was known about plant QM gene. For better understanding of QM gene in rice, a QM homologous fragment was used as a probe to screen rice (Oryza sativa subsp. indica c.v. Guanglu′ ai 4) genomic DNA library,and two clones were obtained. One of them, OSQM2, encoded a highly basic protein of 184 amino acids, the sequence was about 3.1 kb long with a very special promoter region compared with other known QM genes. Seven potential G boxes could be found between -690 and -230. G box, which contains a ACGT core motif, had been reported in many plants to act as a cis acting DNA element in the regulation of genes in a variety of environmental conditions, such as ABA regulated gene expression, red light, UV light, anaerobiosis, and wounding etc. Two closely linked DRE related motifs (dehydration responsive element) could also be found between -182 and 173, which had a CCGAC conserved sequence and had been identified in many cold and drought responsive genes in Arabidopsis. Six MYC recognition sequences with the conserved motif NCANNTGN were also presented, which might be essential for ABA and drought responsive expression of the plant genes.

  5. Enrichment of putative PAX8 target genes at serous epithelial ovarian cancer susceptibility loci.

    Science.gov (United States)

    Kar, Siddhartha P; Adler, Emily; Tyrer, Jonathan; Hazelett, Dennis; Anton-Culver, Hoda; Bandera, Elisa V; Beckmann, Matthias W; Berchuck, Andrew; Bogdanova, Natalia; Brinton, Louise; Butzow, Ralf; Campbell, Ian; Carty, Karen; Chang-Claude, Jenny; Cook, Linda S; Cramer, Daniel W; Cunningham, Julie M; Dansonka-Mieszkowska, Agnieszka; Doherty, Jennifer Anne; Dörk, Thilo; Dürst, Matthias; Eccles, Diana; Fasching, Peter A; Flanagan, James; Gentry-Maharaj, Aleksandra; Glasspool, Rosalind; Goode, Ellen L; Goodman, Marc T; Gronwald, Jacek; Heitz, Florian; Hildebrandt, Michelle A T; Høgdall, Estrid; Høgdall, Claus K; Huntsman, David G; Jensen, Allan; Karlan, Beth Y; Kelemen, Linda E; Kiemeney, Lambertus A; Kjaer, Susanne K; Kupryjanczyk, Jolanta; Lambrechts, Diether; Levine, Douglas A; Li, Qiyuan; Lissowska, Jolanta; Lu, Karen H; Lubiński, Jan; Massuger, Leon F A G; McGuire, Valerie; McNeish, Iain; Menon, Usha; Modugno, Francesmary; Monteiro, Alvaro N; Moysich, Kirsten B; Ness, Roberta B; Nevanlinna, Heli; Paul, James; Pearce, Celeste L; Pejovic, Tanja; Permuth, Jennifer B; Phelan, Catherine; Pike, Malcolm C; Poole, Elizabeth M; Ramus, Susan J; Risch, Harvey A; Rossing, Mary Anne; Salvesen, Helga B; Schildkraut, Joellen M; Sellers, Thomas A; Sherman, Mark; Siddiqui, Nadeem; Sieh, Weiva; Song, Honglin; Southey, Melissa; Terry, Kathryn L; Tworoger, Shelley S; Walsh, Christine; Wentzensen, Nicolas; Whittemore, Alice S; Wu, Anna H; Yang, Hannah; Zheng, Wei; Ziogas, Argyrios; Freedman, Matthew L; Gayther, Simon A; Pharoah, Paul D P; Lawrenson, Kate

    2017-02-14

    Genome-wide association studies (GWAS) have identified 18 loci associated with serous ovarian cancer (SOC) susceptibility but the biological mechanisms driving these findings remain poorly characterised. Germline cancer risk loci may be enriched for target genes of transcription factors (TFs) critical to somatic tumorigenesis. All 615 TF-target sets from the Molecular Signatures Database were evaluated using gene set enrichment analysis (GSEA) and three GWAS for SOC risk: discovery (2196 cases/4396 controls), replication (7035 cases/21 693 controls; independent from discovery), and combined (9627 cases/30 845 controls; including additional individuals). The PAX8-target gene set was ranked 1/615 in the discovery (PGSEA<0.001; FDR=0.21), 7/615 in the replication (PGSEA=0.004; FDR=0.37), and 1/615 in the combined (PGSEA<0.001; FDR=0.21) studies. Adding other genes reported to interact with PAX8 in the literature to the PAX8-target set and applying an alternative to GSEA, interval enrichment, further confirmed this association (P=0.006). Fifteen of the 157 genes from this expanded PAX8 pathway were near eight loci associated with SOC risk at P<10(-5) (including six with P<5 × 10(-8)). The pathway was also associated with differential gene expression after shRNA-mediated silencing of PAX8 in HeyA8 (PGSEA=0.025) and IGROV1 (PGSEA=0.004) SOC cells and several PAX8 targets near SOC risk loci demonstrated in vitro transcriptomic perturbation. Putative PAX8 target genes are enriched for common SOC risk variants. This finding from our agnostic evaluation is of particular interest given that PAX8 is well-established as a specific marker for the cell of origin of SOC.

  6. Complete Genome Sequence of the Filamentous Fungus Aspergillus westerdijkiae Reveals the Putative Biosynthetic Gene Cluster of Ochratoxin A

    Science.gov (United States)

    Chakrabortti, Alolika; Li, Jinming

    2016-01-01

    Ochratoxin A (OTA) is a common mycotoxin that contaminates food and agricultural products. Sequencing of the complete genome of Aspergillus westerdijkiae, a major producer of OTA, reveals more than 50 biosynthetic gene clusters, including a putative OTA biosynthetic gene cluster that encodes a dozen of enzymes, transporters, and regulatory proteins. PMID:27635003

  7. Lipooligosaccharide locus classes and putative virulence genes among chicken and human Campylobacter jejuni isolates.

    Science.gov (United States)

    Ellström, Patrik; Hansson, Ingrid; Nilsson, Anna; Rautelin, Hilpi; Olsson Engvall, Eva

    2016-11-21

    Campylobacter cause morbidity and considerable economic loss due to hospitalization and post infectious sequelae such as reactive arthritis, Guillain Barré- and Miller Fischer syndromes. Such sequelae have been linked to C. jejuni harboring sialic acid structures in their lipooligosaccharide (LOS) layer of the cell wall. Poultry is an important source of human Campylobacter infections but little is known about the prevalence of sialylated C. jejuni isolates and the extent of transmission of such isolates to humans. Genotypes of C. jejuni isolates from enteritis patients were compared with those of broiler chicken with pulsed-field gel electrophoresis (PFGE), to study the patterns of LOS biosynthesis genes and other virulence associated genes and to what extent these occur among Campylobacter genotypes found both in humans and chickens. Chicken and human isolates generally had similar distributions of the putative virulence genes and LOS locus classes studied. However, there were significant differences regarding LOS locus class of PFGE types that were overlapping between chicken and human isolates and those that were distinct to each source. The study highlights the prevalence of virulence associated genes among Campylobacter isolates from humans and chickens and suggests possible patterns of transmission between the two species.

  8. The Pun1 gene for pungency in pepper encodes a putative acyltransferase.

    Science.gov (United States)

    Stewart, Charles; Kang, Byoung-Cheorl; Liu, Kede; Mazourek, Michael; Moore, Shanna L; Yoo, Eun Young; Kim, Byung-Dong; Paran, Ilan; Jahn, Molly M

    2005-06-01

    Pungency in Capsicum fruits is due to the accumulation of the alkaloid capsaicin and its analogs. The biosynthesis of capsaicin is restricted to the genus Capsicum and results from the acylation of an aromatic moiety, vanillylamine, by a branched-chain fatty acid. Many of the enzymes involved in capsaicin biosynthesis are not well characterized and the regulation of the pathway is not fully understood. Based on the current pathway model, candidate genes were identified in public databases and the literature, and genetically mapped. A published EST co-localized with the Pun1 locus which is required for the presence of capsaicinoids. This gene, AT3, has been isolated and its nucleotide sequence has been determined in an array of genotypes within the genus. AT3 showed significant similarity to acyltransferases in the BAHD superfamily. The recessive allele at this locus contains a deletion spanning the promoter and first exon of the predicted coding region in every non-pungent accession tested. Transcript and protein expression of AT3 was tissue-specific and developmentally regulated. Virus-induced gene silencing of AT3 resulted in a decrease in the accumulation of capsaicinoids, a phenotype consistent with pun1. In conclusion, gene mapping, allele sequence data, expression profile and silencing analysis collectively indicate that the Pun1 locus in pepper encodes a putative acyltransferase, and the pun1 allele, used in pepper breeding for nearly 50 000 years, results from a large deletion at this locus.

  9. Discovery of putative capsaicin biosynthetic genes by RNA-Seq and digital gene expression analysis of pepper

    Science.gov (United States)

    Zhang, Zi-Xin; Zhao, Shu-Niu; Liu, Gao-Feng; Huang, Zu-Mei; Cao, Zhen-Mu; Cheng, Shan-Han; Lin, Shi-Sen

    2016-01-01

    The Indian pepper ‘Guijiangwang’ (Capsicum frutescens L.), one of the world’s hottest chili peppers, is rich in capsaicinoids. The accumulation of the alkaloid capsaicin and its analogs in the epidermal cells of the placenta contribute to the pungency of Capsicum fruits. To identify putative genes involved in capsaicin biosynthesis, RNA-Seq was used to analyze the pepper’s expression profiles over five developmental stages. Five cDNA libraries were constructed from the total RNA of placental tissue and sequenced using an Illumina HiSeq 2000. More than 19 million clean reads were obtained from each library, and greater than 50% of the reads were assignable to reference genes. Digital gene expression (DGE) profile analysis using Solexa sequencing was performed at five fruit developmental stages and resulted in the identification of 135 genes of known function; their expression patterns were compared to the capsaicin accumulation pattern. Ten genes of known function were identified as most likely to be involved in regulating capsaicin synthesis. Additionally, 20 new candidate genes were identified related to capsaicin synthesis. We use a combination of RNA-Seq and DGE analyses to contribute to the understanding of the biosynthetic regulatory mechanism(s) of secondary metabolites in a nonmodel plant and to identify candidate enzyme-encoding genes. PMID:27756914

  10. A novel putative enterococcal pathogenicity island linked to the esp virulence gene of Enterococcus faecium and associated with epidemicity.

    Science.gov (United States)

    Leavis, Helen; Top, Janetta; Shankar, Nathan; Borgen, Katrine; Bonten, Marc; van Embden, Jan; Willems, Rob J L

    2004-02-01

    Enterococcus faecalis harbors a virulence-associated surface protein encoded by the esp gene. This gene has been shown to be part of a 150-kb putative pathogenicity island. A gene similar to esp has recently been found in Enterococcus faecium isolates recovered from hospitalized patients. In the present study we analyzed the polymorphism in the esp gene of E. faecium, and we investigated the association of esp with neighboring chromosomal genes. The esp gene showed considerable sequence heterogeneity in the regions encoding the nonrepeat N- and C-terminal domains of the Esp protein as well as differences in the number of repeats. DNA sequencing of chromosomal regions flanking the esp gene of E. faecium revealed seven open reading frames, representing putative genes implicated in virulence, regulation of transcription, and antibiotic resistance. These flanking regions were invariably associated with the presence or absence of the esp gene in E. faecium, indicating that esp in E. faecium is part of a distinct genetic element. Because of the presence of virulence genes in this gene cluster, the lower G+C content relative to that of the genome, and the presence of esp in E. faecium isolates associated with nosocomial outbreaks and clinically documented infections, we conclude that this genetic element constitutes a putative pathogenicity island, the first one described in E. faecium. Except for the presence of esp and araC, this pathogenicity island is completely different from the esp-containing pathogenicity island previously disclosed in E. faecalis.

  11. Prevalence of ten putative virulence genes in the emerging foodborne pathogen Arcobacter isolated from food products.

    Science.gov (United States)

    Girbau, Cecilia; Guerra, Cristian; Martínez-Malaxetxebarria, Irati; Alonso, Rodrigo; Fernández-Astorga, Aurora

    2015-12-01

    Arcobacter spp. are considered to be emerging food- and waterborne pathogens for both humans and animals. However, their virulence mechanisms are still poorly understood. In this study the presence of ten virulence genes (cadF, ciaB, cj1349, hecA, hecB, mviN, pldA, irgA, tlyA and iroE) was assessed in a set of 47 strains of Arcobacter butzleri, 10 of Arcobacter cryaerophilus and 1 Arcobacter skirrowii strain recovered from different food products (pork, chicken, beef, milk, clams and mussels). Overall, the genes cadF, ciaB, cj1349, mviN, pldA and tlyA were detected in all A. butzleri and A. skirrowii strains. Lower detection rates were observed for irgA, iroE, hecA and hecB. The genes hecB and iroE were detected neither in A. cryaerophilus nor in A. skirrowii. The genes hecA and irgA were not detected in A. skirrowii. It was noteworthy that the genes hecA and hecB were significantly (P < 0.05) highly detected in A. butzleri strains isolated from clams compared with strains isolated from milk and chicken. Therefore, our findings underline clams as a source of A. butzleri strains with high prevalence of putative virulence genes. This could be hazardous to human health, especially because these bivalves are usually consumed raw or undercooked.

  12. Functional Characterization of a Putative Nitrate Transporter Gene Promoter from Rice

    Institute of Scientific and Technical Information of China (English)

    Ting-Zhang HU; Kai-Ming CAO; Mian XIA; Xi-Ping WANG

    2006-01-01

    Drought is one of the most significant abiotic stresses that influence plant growth and development. Expression analysis revealed that OsNRT1.3, a putative nitrate transporter gene in rice, was induced by drought. To confirm if the OsNRT1.3 promoter can respond to drought stress, a 2019 bp upstream sequence of OsNRT1.3 was cloned. Three OsNRT1.3 promoter fragments were generated by 5'-deletion, and fused to the β-glucuronidase (GUS) gene. The chimeric genes were introduced into rice plants. NRT2019::GUS, NRT1196: :GUS and NRT719::GUS showed similar expression patterns in seeds,roots, leaves and flowers in all transgenic rice, and GUS activity conferred by different OsNRT1.3 promoter fragments was significantly upregulated by drought stress, indicating that OsNRT1.3 promoter responds to drought stress and the 719 bp upstream sequence of OsNRT1.3 contains the drought response elements.

  13. Differential differences in methylation status of putative imprinted genes among cloned swine genomes.

    Directory of Open Access Journals (Sweden)

    Chih-Jie Shen

    Full Text Available DNA methylation is a major epigenetic modification in the mammalian genome that regulates crucial aspects of gene function. Mammalian cloning by somatic cell nuclear transfer (SCNT often results in gestational or neonatal failure with only a small proportion of manipulated embryos producing live births. Many of the embryos that survive to term later succumb to a variety of abnormalities that are likely due to inappropriate epigenetic reprogramming. Aberrant methylation patterns of imprinted genes in cloned cattle and mice have been elucidated, but few reports have analyzed the cloned pig genome. Four surviving cloned sows that were created by ear fibroblast nuclear transfer, each with a different life span and multiple organ defects, such as heart defects and bone growth delay, were used as epigenetic study materials. First, we identified four putative differential methylation regions (DMR of imprinted genes in the wild-type pig genome, including two maternally imprinted loci (INS and IGF2 and two paternally imprinted loci (H19 and IGF2R. Aberrant DNA methylation, either hypermethylation or hypomethylation, commonly appeared in H19 (45% of imprinted loci hypermethylated vs. 30% hypomethylated, IGF2 (40% vs. 0%, INS (50% vs. 5%, and IGF2R (15% vs. 45% in multiple tissues from these four cloned sows compared with wild-type pigs. Our data suggest that aberrant epigenetic modifications occur frequently in the genome of cloned swine. Even with successful production of cloned swine that avoid prenatal or postnatal death, the perturbation of methylation in imprinted genes still exists, which may be one of reason for their adult pathologies and short life. Understanding the aberrant pattern of gene imprinting would permit improvements in future cloning techniques.

  14. Putative sugarcane FT/TFL1 genes delay flowering time and alter reproductive architecture in Arabidopsis.

    Science.gov (United States)

    Coelho, Carla P; Minow, Mark A A; Chalfun-Júnior, Antonio; Colasanti, Joseph

    2014-01-01

    Agriculturally important grasses such as rice, maize, and sugarcane are evolutionarily distant from Arabidopsis, yet some components of the floral induction process are highly conserved. Flowering in sugarcane is an important factor that negatively affects cane yield and reduces sugar/ethanol production from this important perennial bioenergy crop. Comparative studies have facilitated the identification and characterization of putative orthologs of key flowering time genes in sugarcane, a complex polyploid plant whose genome has yet to be sequenced completely. Using this approach we identified phosphatidylethanolamine-binding protein (PEBP) gene family members in sugarcane that are similar to the archetypical FT and TFL1 genes of Arabidopsis that play an essential role in controlling the transition from vegetative to reproductive growth. Expression analysis of ScTFL1, which falls into the TFL1-clade of floral repressors, showed transcripts in developing leaves surrounding the shoot apex but not at the apex itself. ScFT1 was detected in immature leaves and apical regions of vegetatively growing plants and, after the floral transition, expression also occurred in mature leaves. Ectopic over-expression of ScTFL1 in Arabidopsis caused delayed flowering in Arabidopsis, as might be expected for a gene related to TFL1. In addition, lines with the latest flowering phenotype exhibited aerial rosette formation. Unexpectedly, over-expression of ScFT1, which has greatest similarity to the florigen-encoding FT, also caused a delay in flowering. This preliminary analysis of divergent sugarcane FT and TFL1 gene family members from Saccharum spp. suggests that their expression patterns and roles in the floral transition has diverged from the predicted role of similar PEBP family members.

  15. Putative sugarcane FT/TFL1 genes delay flowering time and alter reproductive architecture in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Carla P. Coelho

    2014-05-01

    Full Text Available Agriculturally important grasses such as rice, maize and sugarcane are evolutionarily distant from Arabidopsis, yet some components of the floral induction process are highly conserved. Flowering in sugarcane is an important factor that negatively affects cane yield and reduces sugar/ethanol production from this important perennial bioenergy crop. Comparative studies have facilitated the identification and characterization of putative orthologs of key flowering time genes in sugarcane, a complex polyploid plant whose genome has yet to be sequenced completely. Using this approach we identified phosphatidylethanolamine-binding protein (PEBP gene family members in sugarcane that are similar to the archetypical FT and TFL1 genes of Arabidopsis that play an essential role in controlling the transition from vegetative to reproductive growth. Expression analysis of ScTFL1, which falls into the TFL1-clade of floral repressors, showed transcripts in developing leaves surrounding the shoot apex but not at the apex itself. ScFT1 was detected in immature leaves and apical regions of vegetatively growing plants and, after the floral transition, expression also occurred in mature leaves. Ectopic over-expression of ScTFL1 in Arabidopsis caused delayed flowering in Arabidopsis, as might be expected for a gene related to TFL1. In addition, lines with the latest flowering phenotype exhibited aerial rosette formation. Unexpectedly, over-expression of ScFT1, which has greatest similarity to the florigen-encoding FT, also caused a delay in flowering. This preliminary analysis of divergent sugarcane FT and TFL1 gene family members from Saccharum spp. suggests that their expression patterns and roles in the floral transition has diverged from the predicted role of similar PEBP family members.

  16. Putative sugarcane FT/TFL1 genes delay flowering time and alter reproductive architecture in Arabidopsis

    Science.gov (United States)

    Coelho, Carla P.; Minow, Mark A. A.; Chalfun-Júnior, Antonio; Colasanti, Joseph

    2014-01-01

    Agriculturally important grasses such as rice, maize, and sugarcane are evolutionarily distant from Arabidopsis, yet some components of the floral induction process are highly conserved. Flowering in sugarcane is an important factor that negatively affects cane yield and reduces sugar/ethanol production from this important perennial bioenergy crop. Comparative studies have facilitated the identification and characterization of putative orthologs of key flowering time genes in sugarcane, a complex polyploid plant whose genome has yet to be sequenced completely. Using this approach we identified phosphatidylethanolamine-binding protein (PEBP) gene family members in sugarcane that are similar to the archetypical FT and TFL1 genes of Arabidopsis that play an essential role in controlling the transition from vegetative to reproductive growth. Expression analysis of ScTFL1, which falls into the TFL1-clade of floral repressors, showed transcripts in developing leaves surrounding the shoot apex but not at the apex itself. ScFT1 was detected in immature leaves and apical regions of vegetatively growing plants and, after the floral transition, expression also occurred in mature leaves. Ectopic over-expression of ScTFL1 in Arabidopsis caused delayed flowering in Arabidopsis, as might be expected for a gene related to TFL1. In addition, lines with the latest flowering phenotype exhibited aerial rosette formation. Unexpectedly, over-expression of ScFT1, which has greatest similarity to the florigen-encoding FT, also caused a delay in flowering. This preliminary analysis of divergent sugarcane FT and TFL1 gene family members from Saccharum spp. suggests that their expression patterns and roles in the floral transition has diverged from the predicted role of similar PEBP family members. PMID:24904616

  17. Multiple functions of mfa-1, a putative pheromone precursor gene of Neurospora crassa.

    Science.gov (United States)

    Kim, Hyojeong; Metzenberg, Robert L; Nelson, Mary Anne

    2002-12-01

    A putative pheromone precursor gene of Neurospora crassa, mfa-1 (which encodes mating factor a-1), was identified as the most abundant clone in starved mycelial and perithecial cDNA libraries. Northern analysis demonstrated high mfa-1 expression in all mating type a tissues and suggested low expression levels in mat A tissues. The mfa-1 gene was expressed as an approximately 1.2-kb transcript predicted to encode a 24-residue peptide, followed by a long 3' untranslated region (3' UTR). The predicted MFA1 sequence showed 100% sequence identity to PPG2 of Sordaria macrospora and structural similarity (a carboxy-terminal CAAX motif) to many hydrophobic fungal pheromone precursors. Mutants with a disrupted open reading frame (ORF) in which the critical cysteine residue had been changed to a nonprenylatable residue, tyrosine (YAAX mutants), were isolated, as were mfa-1 mutants with intact ORFs but multiple mutations in the 3' noncoding region (CAAX mutants). The 3' UTR is required for the full range of mfa-1 gene activity. Both classes of mutants showed delayed and reduced vegetative growth (which was suppressed by supplementation with a minute amount [30 micro M] of ornithine, citrulline, or arginine), as well as aberrant sexual development. When crossed as female parents to wild-type males, the CAAX and YAAX mutants showed greatly reduced ascospore production. No ascospores were produced in homozygous mfa-1 crosses. As males, YAAX mat a mutants were unable to attract wild-type mat A trichogynes (female-specific hyphae) or to initiate sexual development, while CAAX mat a mutants were able to mate and produce sexual progeny despite their inability to attract mat A trichogynes. In the mat A background, both CAAX and YAAX mutants showed normal male fertility but defective vegetative growth and aberrant female sexual development. Thus, the mfa-1 gene appears to have multiple roles in N. crassa development: (i) it encodes a hydrophobic pheromone with a putative farnesylated

  18. Putative psychosis genes in the prefrontal cortex: combined analysis of gene expression microarrays

    Directory of Open Access Journals (Sweden)

    Yolken Robert H

    2008-11-01

    Full Text Available Abstract Background Recent studies have shown similarities between schizophrenia and bipolar disorder in phenotypes and in genotypes, and those studies have contributed to an ongoing re-evaluation of the traditional dichotomy between schizophrenia and bipolar disorder. Bipolar disorder with psychotic features may be closely related to schizophrenia and therefore, psychosis may be an alternative phenotype compared to the traditional diagnosis categories. Methods We performed a cross-study analysis of 7 gene expression microarrays that include both psychosis and non-psychosis subjects. These studies include over 400 microarray samples (163 individual subjects on 3 different Affymetrix microarray platforms. Results We found that 110 transcripts are differentially regulated (p Conclusion This study demonstrates the advantages of cross-study analysis in detecting consensus changes in gene expression across multiple microarray studies. Differential gene expression between individuals with and without psychosis suggests that psychosis may be a useful phenotypic variable to complement the traditional diagnosis categories.

  19. Expression and localization of the immunophilin FKBP51 in colorectal carcinomas and primary metastases, and alterations following oxaliplatin-based chemotherapy

    Science.gov (United States)

    Rotoli, Deborah; Morales, Manuel; Del Carmen Maeso, María; Del Pino García, María; Morales, Araceli; Ávila, Julio; Martín-Vasallo, Pablo

    2016-01-01

    The immunophilin FK506-binding protein 5 (FKBP51) is a scaffold protein that serves a pivotal role in the regulation of multiple signaling pathways, integrating external and internal stimuli into distinct signal outputs. In a previous study, we identified several genes that are significantly up- or downregulated in the peripheral white cells (PWCs) of colorectal adenocarcinoma (CRC) patients undergoing oxaliplatin-based chemotherapy. In our screening, FKBP51 gene expression was downregulated following chemotherapy. In order to determine whether this alteration in gene expression observed in PWCs may be detected at the protein level in tumors and metastases following the administration of adjuvant chemotherapy, an immunohistochemical analysis of FKBP51 in CRC and primary metastasis tissues was performed. The present study confirmed the downregulation of FKBP51 gene expression elicited by chemotherapy with folinic acid (leucovorin), fluorouracil and oxaliplatin in metastasized liver tissue that had been resected after the oxaliplatin-based chemotherapy, compared with tissue section samples of CRC from patients (prior to antineoplastic treatment). Furthermore, the results indicated that, in CRC tissue sections, the expression of FKBP51 protein is associated with an immature phenotype of stromal fibroblasts and with the epithelial-to-mesenchymal transition (EMT) phenotype, suggesting a role for this protein in the EMT process in CRC. Finally, the observation that only certain cells of the stroma express FKBP51 protein suggests a potential role for this immunophilin as a stroma cell subtype marker. PMID:27446431

  20. Cloning and Characterization of a Putative CTR1 Gene from Wheat

    Institute of Scientific and Technical Information of China (English)

    BI Cai-li; WEN Xiao-jie; ZHANG Xue-yong; LIU Xu

    2010-01-01

    CTR1 is a key negative regulator in ethylene signal transduction.A salt-induced CTR1 like gene(TaCTR1)was cloned from wheat,its expression under abiotie stresses,subcellular localization and the effect of overexpression of TaCTR1 on salt tolerance in tobacco was studied.A putative CTR1 gene was cloned and characterized from wheat via rapid amplification of cDNA ends(RACE)and RT-PCR.TaCTR1 expression under stresses was analyzed using semi-quantitative RT-PCR and the effect of overexpression of TaCTR1 on salt tolerance was conducted in tobacco.The full-length cDNA of TaCTR1is 2635 bp which codes for a polypeptide of 759 amino acids.There is a conserved serine/threonine protein kinase domain at the carboxyl terminus containing an ATP-binding site.Southern blot analysis revealed that TaCTR1 consisted of a gene family in wheat.The amino acid homologies of CTR1 among different organisms share higher similarities.Expression analysis revealed that TaCTR1 was induced by NaCl and drought stress but inhibited by ABA treatment.Transient expression of TaCTR1-GFP in the onion epidermal cells indicated that TaCTR1 was probably targeted to the plasma membrane.Overexpression of TaCTR1 decreased salt tolerance in transgenic tobacco(Nicotiana tabacum L.)plants compared with the control.To our knowledge,TaCTR1 is the first CTR1 gene cloned in wheat and may be involved in various abiotic stresses.Overexpression of TaCTR1 decreased the salt tolerance in tobacco suggested that TaCTR1 may act as a negative regulator of salt stress in plants.

  1. Putative cis-regulatory elements associated with heat shock genes activated during excystation of Cryptosporidium parvum.

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    Benjamin Cohn

    Full Text Available BACKGROUND: Cryptosporidiosis is a ubiquitous infectious disease, caused by the protozoan parasites Cryptosporidium hominis and C. parvum, leading to acute, persistent and chronic diarrhea worldwide. Although the complications of this disease can be serious, even fatal, in immunocompromised patients of any age, they have also been found to lead to long term effects, including growth inhibition and impaired cognitive development, in infected immunocompetent children. The Cryptosporidium life cycle alternates between a dormant stage, the oocyst, and a highly replicative phase that includes both asexual vegetative stages as well as sexual stages, implying fine genetic regulatory mechanisms. The parasite is extremely difficult to study because it cannot be cultured in vitro and animal models are equally challenging. The recent publication of the genome sequence of C. hominis and C. parvum has, however, significantly advanced our understanding of the biology and pathogenesis of this parasite. METHODOLOGY/PRINCIPAL FINDINGS: Herein, our goal was to identify cis-regulatory elements associated with heat shock response in Cryptosporidium using a combination of in silico and real time RT-PCR strategies. Analysis with Gibbs-Sampling algorithms of upstream non-translated regions of twelve genes annotated as heat shock proteins in the Cryptosporidium genome identified a highly conserved over-represented sequence motif in eleven of them. RT-PCR analyses, described herein and also by others, show that these eleven genes bearing the putative element are induced concurrent with excystation of parasite oocysts via heat shock. CONCLUSIONS/SIGNIFICANCE: Our analyses suggest that occurrences of a motif identified in the upstream regions of the Cryptosporidium heat shock genes represent parts of the transcriptional apparatus and function as stress response elements that activate expression of these genes during excystation, and possibly at other stages in the life

  2. A putative plant organelle RNA recognition protein gene is essential for maize kernel development

    Institute of Scientific and Technical Information of China (English)

    Antony M Chettoor; Gibum Yi; Elisa Gomez; Gregorio Hueros; Robert B Meeley; Philip W Becraft

    2015-01-01

    Basal endosperm transfer layer (BETL) cel s are responsible for transferring apoplastic solutes from the maternal pedicel into the endosperm, supplying the grain with compounds required for embryo development and storage reserve accumulation. Here, we analyze the maize (Zea mays L.) empty pericarp6 (emp6) mutant, which causes early arrest in grain development. The Emp6þgene function is required independently in both the embryo and endo-sperm. The emp6 mutant causes a notable effect on the differentiation of BETL cel s; the extensive cel wal ingrowths that distinguish BETL cel s are diminished and BETL marker gene expression is compromised in mutant kernels. Transposon tagging identified the emp6 locus as encoding a putative plant organel e RNA recognition (PORR) protein, 1 of 15 PORR family members in maize. The emp6 transcript is widely detected in plant tissues with highest levels in embryos and developing kernels. EMP6‐green fluorescent protein (GFP) fusion proteins transiently expressed in Nicotiana benthamiana leaves were targeted specifical y to mitochondria. These results suggest that BETL cel differentia-tion might be particularly energy intensive, or alternatively, that mitochondria might confer a developmental function.

  3. Molecular cloning and primary sequence analysis of a gene encoding a putative shitinase gene in Brassica oleracea var.capitata

    Institute of Scientific and Technical Information of China (English)

    TANGGUOQING; YONGYANBAI; 等

    1996-01-01

    Chitinase,which catalyzes the hydrolysis of the β-1,4-acetyl-D-glucosamine linkages of the fungal cell wall polymer chitin,is involved in inducible plants defense system.By construction of cabbage(Brassica oleracea var. capitata) genomic library and screening the library with pRCH8,a probe of rice chitinase gene fragment,a chitinase genomic sequence was isolated.The complete uncleotide sequence of the putative cabbage chitinase gene (cabch29) was determined,with its longest open reading frame (ORF) encoding a polypeptide of 413 aa.This polypeptide consists of a 21 aa N-terminal signal peptide,two chitin-binding domains different from those of other classes of plant chitinases,and a catalytic domain.Homology analysis illustrated that this cabch29 gene has 58.8% identity at the nucleotide level with the pRCH8 ORF probe and has 50% identity at the amino acid level tiwh the catalytic domains of chitinase from bean,maize and sugar beet.Meanwhile,several kinds of cis-elements,such as TATA box,CAAT box,GATA motif,ASF-1 binding site,wound-response elements and AATAAA,have also been discovered in the flanking region of cabch29 gene.

  4. Characterization of Two Putative Protein Phosphatase Genes and Their Involvement in Phosphorus Efficiency in Phaseolus vulgari

    Institute of Scientific and Technical Information of China (English)

    Cui-Yue Liang; Zhi-Jian Chen; Zhu-Fang Yao; Jiang Tian; Hong Liao

    2012-01-01

    Protein dephosphorylation mediated by protein phosphatases plays a major role in signal transduction of plant responses to environmental stresses.In this study,two putative protein phosphatases,PvPS2:1 and PvPS2:2 were identified and characterized in bean (Phaseolus vulgaris).The two PvPS2 members were found to be localized to the plasma membrane and the nucleus by transient expression of PvPS2:GFP in onion epidermal cells.Transcripts of the two PvPS2 genes were significantly increased by phosphate (Pi) starvation in the two bean genotypes,G19833 (a P-efficient genotype) and DOR364 (a P-inefficient genotype).However,G19833 exhibited higher PvPS2:1 expression levels than DOR364 in both leaves and roots during P1 starvation.Increased transcription of PvPS2:1 in response to Pi starvation was further verified through histochemical analysis of PvPS2:1 promoter fusion β-glucuronidase (GUS) in transgenic Arabidopsis plants.Analysis of PvPS2∶1 overexpression lines in bean hairy roots and Arabidopsis showed that PvS2:1 was involved in root growth and P accumulation.Furthermore,expression levels of two P(1) starvation responsive genes were upregulated and the APase activities were enhanced in the overexpressing PvPS2∶1 Arabidopsis lines.Taken together,our results strongly suggested that PvPS2∶1positively regulated plant responses to P1 starvation,and could be further targeted as a candidate gene to improve crop P efficiency.

  5. Isolation and characterization of a new chemokine receptor gene, the putative chicken CXCR1.

    Science.gov (United States)

    Li, Q J; Lu, S; Ye, R D; Martins-Green, M

    2000-10-31

    This study delineates the isolation and characterization of a novel chemokine receptor gene, the putative chicken CXC receptor 1 (cCXCR1). Using a human CXCR1 probe, we isolated several positive clones from a chicken genomic library. One of the clones contained a fragment of approximately 5000bp that hybridized strongly with the hCXCR1 probe. This fragment was sequenced and subjected to a variety of computer analyses. The open reading frame for this gene predicts a seven transmembrane domain protein with all the characteristics of a chemokine receptor and with 67% sequence homology to hCXCR1, 65% to hCXCR2 and also with considerable sequence homology to other human chemokine receptors such as hCXCR4 (50%), hCCR2 (49%) and hCCR1 (49%). However, the homology to a previously isolated potential G-protein-coupled receptor for chickens (AvCRL1) is only 47%. Using 5' RACE, two transcription initiation sites were identified suggesting the potential for the expression of two protein isoforms (I and II) in vivo. The promoter for the putative cCXCR1 contains a variety of consensus transcription factor binding elements that can potentially be involved in the expression of this chicken receptor upon stimulation by stress-inducing agents. RT-PCR analysis was used to determine the pattern of expression of the larger isoform (I) of this receptor in a variety of tissues. This form of the receptor is expressed primarily in the organs of the gastrointestinal tract, tissues that are frequently exposed to stress-inducing agents, but not in the central nervous system, tissues that are protected from insult by the blood barrier. Using the same RT-PCR approach we show that stress-inducing agents, such as 'first-hand' and 'second-hand' cigarette smoke components, tumor promoters and thrombin, differentially stimulate the expression of the isoform I in primary fibroblasts. Thrombin is an enzyme that plays many important roles in thrombosis, angiogenesis and wound healing and exposure to

  6. Insilico Analysis unveils Putative Metabolic Pathways and Essential Genes within Leishmania donovani ‘Orfeome’

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    Nithin eRavooru

    2014-08-01

    Full Text Available Leishmaniasis is a parasitic disease caused by the protozoan Leishmania, which is active in two broad forms namely, Visceral Leishmaniasis (VL or Kala Azar and Cutaneous Leishmaniasis (CL. The disease is most prevalent in the tropical regions and poses a threat to more than 70 countries across the globe. In the Indian subcontinent, about 200 million people are estimated to be at risk of developing VL and this area harbours an estimated 67% of the global VL disease burden. The state of Bihar alone has captured almost 50% of the total cases in the Indian region. While no vaccination exists, several pentavalent antimonials and drugs like Paromomycin, Amphotericin, Miltefosine etc., are used in the treatment of Leishmaniasis. However, due to low efficacy of these drugs and the resistance developed by the bug to these medications, there is an urgent need to look into species specific targets. The proteome information available suggests that among the 7960 proteins, a staggering 65% of it remains to be annotated with clarity.Hence, in the present study, we have demonstrated a protocol to integrate the seqeunce and functional information from various databases, such as GO, PFAM, KEGG, String DB, COG and DEG, to assign putative functions to many of the hypothetical seqeucences present in this proteome. These crucial information related to pathways and essential genes show promise for exploring the design strategies towards developing drugs, to tackle this notorious parasitic disease.

  7. Occurrence of putative virulence genes in arcobacter species isolated from humans and animals.

    Science.gov (United States)

    Douidah, Laid; de Zutter, Lieven; Baré, Julie; De Vos, Paul; Vandamme, Peter; Vandenberg, Olivier; Van den Abeele, Anne-Marie; Houf, Kurt

    2012-03-01

    Interest in arcobacters in veterinary and human public health has increased since the first report of the isolation of arcobacters from food of animal origin. Since then, studies worldwide have reported the occurrence of arcobacters on food and in food production animals and have highlighted possible transmission, especially of Arcobacter butzleri, to the human population. In humans, arcobacters are associated with enteritis and septicemia. To assess their clinical relevance for humans and animals, evaluation of potential virulence factors is required. However, up to now, little has been known about the mechanisms of pathogenicity. Because of their close phylogenetic affiliation to the food-borne pathogen Campylobacter and their similar clinical manifestations, the presence of nine putative Campylobacter virulence genes (cadF, ciaB, cj1349, hecA, hecB, irgA, mviN, pldA, and tlyA) previously identified in the recent Arcobacter butzleri ATCC 49616 genome sequence was determined in a large set of human and animal Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii strains after the development of rapid and accurate PCR assays and confirmed by sequencing and dot blot hybridization.

  8. Linkage mapping of putative regulator genes of barley grain development characterized by expression profiling

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    Wobus Ulrich

    2009-01-01

    Full Text Available Abstract Background Barley (Hordeum vulgare L. seed development is a highly regulated process with fine-tuned interaction of various tissues controlling distinct physiological events during prestorage, storage and dessication phase. As potential regulators involved within this process we studied 172 transcription factors and 204 kinases for their expression behaviour and anchored a subset of them to the barley linkage map to promote marker-assisted studies on barley grains. Results By a hierachical clustering of the expression profiles of 376 potential regulatory genes expressed in 37 different tissues, we found 50 regulators preferentially expressed in one of the three grain tissue fractions pericarp, endosperm and embryo during seed development. In addition, 27 regulators found to be expressed during both seed development and germination and 32 additional regulators are characteristically expressed in multiple tissues undergoing cell differentiation events during barley plant ontogeny. Another 96 regulators were, beside in the developing seed, ubiquitously expressed among all tissues of germinating seedlings as well as in reproductive tissues. SNP-marker development for those regulators resulted in anchoring 61 markers on the genetic linkage map of barley and the chromosomal assignment of another 12 loci by using wheat-barley addition lines. The SNP frequency ranged from 0.5 to 1.0 SNP/kb in the parents of the various mapping populations and was 2.3 SNP/kb over all eight lines tested. Exploration of macrosynteny to rice revealed that the chromosomal orders of the mapped putative regulatory factors were predominantly conserved during evolution. Conclusion We identified expression patterns of major transcription factors and signaling related genes expressed during barley ontogeny and further assigned possible functions based on likely orthologs functionally well characterized in model plant species. The combined linkage map and reference

  9. Identification of putative methanol dehydrogenase (moxF) structural genes in methylotrophs and cloning of moxF genes from Methylococcus capsulatus bath and Methylomonas albus BG8

    OpenAIRE

    Stephens, R. L.; Haygood, M G; Lidstrom, M. E.

    1988-01-01

    An open-reading-frame fragment of a Methylobacterium sp. strain AM1 gene (moxF) encoding a portion of the methanol dehydrogenase structural protein has been used as a hybridization probe to detect similar sequences in a variety of methylotrophic bacteria. This hybridization was used to isolate clones containing putative moxF genes from two obligate methanotrophic bacteria, Methylococcus capsulatus Bath and Methylomonas albus BG8. The identity of these genes was confirmed in two ways. A T7 exp...

  10. De Novo assembly of the Japanese flounder (Paralichthys olivaceus spleen transcriptome to identify putative genes involved in immunity.

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    Lin Huang

    Full Text Available Japanese flounder (Paralichthys olivaceus is an economically important marine fish in Asia and has suffered from disease outbreaks caused by various pathogens, which requires more information for immune relevant genes on genome background. However, genomic and transcriptomic data for Japanese flounder remain scarce, which limits studies on the immune system of this species. In this study, we characterized the Japanese flounder spleen transcriptome using an Illumina paired-end sequencing platform to identify putative genes involved in immunity.A cDNA library from the spleen of P. olivaceus was constructed and randomly sequenced using an Illumina technique. The removal of low quality reads generated 12,196,968 trimmed reads, which assembled into 96,627 unigenes. A total of 21,391 unigenes (22.14% were annotated in the NCBI Nr database, and only 1.1% of the BLASTx top-hits matched P. olivaceus protein sequences. Approximately 12,503 (58.45% unigenes were categorized into three Gene Ontology groups, 19,547 (91.38% were classified into 26 Cluster of Orthologous Groups, and 10,649 (49.78% were assigned to six Kyoto Encyclopedia of Genes and Genomes pathways. Furthermore, 40,928 putative simple sequence repeats and 47, 362 putative single nucleotide polymorphisms were identified. Importantly, we identified 1,563 putative immune-associated unigenes that mapped to 15 immune signaling pathways.The P. olivaceus transciptome data provides a rich source to discover and identify new genes, and the immune-relevant sequences identified here will facilitate our understanding of the mechanisms involved in the immune response. Furthermore, the plentiful potential SSRs and SNPs found in this study are important resources with respect to future development of a linkage map or marker assisted breeding programs for the flounder.

  11. Transcriptome-Wide Survey and Expression Profile Analysis of Putative Chrysanthemum HD-Zip I and II Genes

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    Aiping Song

    2016-05-01

    Full Text Available The homeodomain-leucine zipper (HD-Zip transcription factor family is a key transcription factor family and unique to the plant kingdom. It consists of a homeodomain and a leucine zipper that serve in combination as a dimerization motif. The family can be classified into four subfamilies, and these subfamilies participate in the development of hormones and mediation of hormone action and are involved in plant responses to environmental conditions. However, limited information on this gene family is available for the important chrysanthemum ornamental species (Chrysanthemum morifolium. Here, we characterized 17 chrysanthemum HD-Zip genes based on transcriptome sequences. Phylogenetic analyses revealed that 17 CmHB genes were distributed in the HD-Zip subfamilies I and II and identified two pairs of putative orthologous proteins in Arabidopsis and chrysanthemum and four pairs of paralogous proteins in chrysanthemum. The software MEME was used to identify 7 putative motifs with E values less than 1e-3 in the chrysanthemum HD-Zip factors, and they can be clearly classified into two groups based on the composition of the motifs. A bioinformatics analysis predicted that 8 CmHB genes could be targeted by 10 miRNA families, and the expression of these 17 genes in response to phytohormone treatments and abiotic stresses was characterized. The results presented here will promote research on the various functions of the HD-Zip gene family members in plant hormones and stress responses.

  12. Transcriptome-Wide Survey and Expression Profile Analysis of Putative Chrysanthemum HD-Zip I and II Genes.

    Science.gov (United States)

    Song, Aiping; Li, Peiling; Xin, Jingjing; Chen, Sumei; Zhao, Kunkun; Wu, Dan; Fan, Qingqing; Gao, Tianwei; Chen, Fadi; Guan, Zhiyong

    2016-05-17

    The homeodomain-leucine zipper (HD-Zip) transcription factor family is a key transcription factor family and unique to the plant kingdom. It consists of a homeodomain and a leucine zipper that serve in combination as a dimerization motif. The family can be classified into four subfamilies, and these subfamilies participate in the development of hormones and mediation of hormone action and are involved in plant responses to environmental conditions. However, limited information on this gene family is available for the important chrysanthemum ornamental species (Chrysanthemum morifolium). Here, we characterized 17 chrysanthemum HD-Zip genes based on transcriptome sequences. Phylogenetic analyses revealed that 17 CmHB genes were distributed in the HD-Zip subfamilies I and II and identified two pairs of putative orthologous proteins in Arabidopsis and chrysanthemum and four pairs of paralogous proteins in chrysanthemum. The software MEME was used to identify 7 putative motifs with E values less than 1e-3 in the chrysanthemum HD-Zip factors, and they can be clearly classified into two groups based on the composition of the motifs. A bioinformatics analysis predicted that 8 CmHB genes could be targeted by 10 miRNA families, and the expression of these 17 genes in response to phytohormone treatments and abiotic stresses was characterized. The results presented here will promote research on the various functions of the HD-Zip gene family members in plant hormones and stress responses.

  13. Comparative Investigation of Copper Tolerance and Identification of Putative Tolerance Related Genes in Tardigrades

    Science.gov (United States)

    Hygum, Thomas L.; Fobian, Dannie; Kamilari, Maria; Jørgensen, Aslak; Schiøtt, Morten; Grosell, Martin; Møbjerg, Nadja

    2017-01-01

    Tardigrades are microscopic aquatic animals renowned for their tolerance toward extreme environmental conditions. The current study is the first to investigate their tolerance toward heavy metals and we present a novel tardigrade toxicant tolerance assay based on activity assessments as a measure of survival. Specifically, we compare tolerance toward copper in four species representing different evolutionary lineages, habitats and adaptation strategies, i.e., a marine heterotardigrade, Echiniscoides sigismundi, a limno-terrestrial heterotardigrade, Echiniscus testudo, a limno-terrestrial eutardigrade, Ramazzottius oberhaeuseri, and a marine eutardigrade, Halobiotus crispae. The latter was sampled at a time of year, when the population is predominantly represented by aberrant P1 cysts, while the other species were in normal active states prior to exposure. Based on volume measurements and a general relation between body mass and copper tolerance, expected tardigrade EC50 values were estimated at 0.5–2 μg l−1. Following 24 h of exposure, tolerance was high with no apparent link to lineage or habitat. EC50s (95% CI), 24 h after exposure, were estimated at 178 (168–186) and 310 (295–328) μg l−1, respectively, for E. sigismundi and R. oberhaeuseri, whereas E. testudo and H. crispae were less affected. Highest tolerance was observed in H. crispae with a mean ± s.e.m. activity of 77 ± 2% (n = 3) 24 h after removal from ~3 mg l−1 copper, suggesting that tardigrade cysts have increased tolerance toward toxicants. In order to identify putative tolerance related genes, an E. sigismundi transcriptome was searched for key enzymes involved in osmoregulation, antioxidant defense and copper metabolism. We found high expression of Na/K ATPase and carbonic anhydrase, known targets for copper. Our transcriptome, furthermore, revealed high expression of antioxidant enzymes, copper transporters, ATOX1, and a Cu-ATPase. In summary, our results indicate that tardigrades

  14. Validation of candidate genes putatively associated with resistance to SCMV and MDMV in maize (Zea mays L.) by expression profiling.

    Science.gov (United States)

    Uzarowska, Anna; Dionisio, Giuseppe; Sarholz, Barbara; Piepho, Hans-Peter; Xu, Mingliang; Ingvardsen, Christina Rønn; Wenzel, Gerhard; Lübberstedt, Thomas

    2009-02-02

    The potyviruses sugarcane mosaic virus (SCMV) and maize dwarf mosaic virus (MDMV) are major pathogens of maize worldwide. Two loci, Scmv1 and Scmv2, have ealier been shown to confer complete resistance to SCMV. Custom-made microarrays containing previously identified SCMV resistance candidate genes and resistance gene analogs were utilised to investigate and validate gene expression and expression patterns of isogenic lines under pathogen infection in order to obtain information about the molecular mechanisms involved in maize-potyvirus interactions. By employing time course microarray experiments we identified 68 significantly differentially expressed sequences within the different time points. The majority of differentially expressed genes differed between the near-isogenic line carrying Scmv1 resistance locus at chromosome 6 and the other isogenic lines. Most differentially expressed genes in the SCMV experiment (75%) were identified one hour after virus inoculation, and about one quarter at multiple time points. Furthermore, most of the identified mapped genes were localised outside the Scmv QTL regions. Annotation revealed differential expression of promising pathogenesis-related candidate genes, validated by qRT-PCR, coding for metallothionein-like protein, S-adenosylmethionine synthetase, germin-like protein or 26S ribosomal RNA. Our study identified putative candidate genes and gene expression patterns related to resistance to SCMV. Moreover, our findings support the effectiveness and reliability of the combination of different expression profiling approaches for the identification and validation of candidate genes. Genes identified in this study represent possible future targets for manipulation of SCMV resistance in maize.

  15. A putative MYB35 ortholog is a candidate for the sex-determining genes in Asparagus officinalis.

    Science.gov (United States)

    Tsugama, Daisuke; Matsuyama, Kohei; Ide, Mayui; Hayashi, Masato; Fujino, Kaien; Masuda, Kiyoshi

    2017-02-08

    Asparagus officinalis (garden asparagus) is a dioecious perennial crop. For agricultural production of A. officinalis, male plants have advantages over female plants. The dioecism of A. officinalis is determined by the single dominant masculinizing M locus, which is involved in tapetal cell development in stamens, but thus far no specific M locus genes have been identified. We re-analyzed previously published RNA-Seq data for the A. officinalis transcriptome, cloned some genes, and discovered that a putative ortholog of MYB35, which is indispensable for tapetal cell development in Arabidopsis thaliana, is absent in the genome of female plants in A. officinalis. In a reverse transcription-PCR analysis, this gene (AoMYB35) exhibited strong expression in stamens in male flowers at an early developmental stage. In an in situ hybridization analysis, AoMYB35 mRNA was detected in tapetal cells in young male flowers. GFP-fused AoMYB35 was detected in the nucleus when expressed in onion epidermal cells. These results suggest that AoMYB35 is a male-specific gene encoding a putative transcription factor that acts in tapetal cells at an early stage of flower development in A. officinalis. Together, the results support the idea that AoMYB35 is a candidate for one of the M locus genes in A. officinalis.

  16. Application of a Colorimetric Assay to Identify Putative Ribofuranosylaminobenzene 5'-Phosphate Synthase Genes Expressed with Activity in Escherichia coli

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    Bechard Matthew E.

    2003-01-01

    Full Text Available Tetrahydromethanopterin (H4MPT is a tetrahydrofolate analog originally discovered in methanogenic archaea, but later found in other archaea and bacteria. The extent to which H4MPT occurs among living organisms is unknown. The key enzyme which distinguishes the biosynthetic pathways of H4MPT and tetrahydrofolate is ribofuranosylaminobenzene 5'-phosphate synthase (RFAP synthase. Given the importance of RFAP synthase in H4MPT biosynthesis, the identification of putative RFAP synthase genes and measurement of RFAP synthase activity would provide an indication of the presence of H4MPT in untested microorganisms. Investigation of putative archaeal RFAP synthase genes has been hampered by the tendency of the resulting proteins to form inactive inclusion bodies in Escherichia coli. The current work describes a colorimetric assay for measuring RFAP synthase activity, and two modified procedures for expressing recombinant RFAP synthase genes to produce soluble, active enzyme. By lowering the incubation temperature during expression, RFAP synthase from Archaeoglobus fulgidus was produced in E. coli and purified to homogeneity. The production of active RFAP synthase from Methanothermobacter thermautotrophicus was achieved by coexpression of the gene MTH0830 with a molecular chaperone. This is the first direct biochemical identification of a methanogen gene that codes for an active RFAP synthase.

  17. DNA and RNA from Uninfected Vertebrate Cells Contain Nucleotide Sequences Related to the Putative Transforming Gene of Avian Myelocytomatosis Virus

    Science.gov (United States)

    Sheiness, Diana; Bishop, J. Michael

    1979-01-01

    The avian carcinoma virus MC29 (MC29V) contains a sequence of approximately 1,500 nucleotides which may represent a gene responsible for tumorigenesis by MC29V. We present evidence that MC29V has acquired this nucleotide sequence from the DNA of its host. The host sequence which has been incorporated by MC29V is transcribed into RNA in uninfected chicken cells and thus probably encodes a cellular gene. We have prepared radioactive DNA complementary to the putative MC29V transforming gene (cDNAmc29) and have found that sequences homologous to cDNAmc29 are present in the genomes of several uninfected vertebrate species. The DNA of chicken, the natural host for MC29V, contains at least 90% of the sequences represented by cDNAmc29. DNAs from other animals show significant but decreasing amounts of complementarity to cDNAmc29 in accordance with their evolutionary divergence from chickens; the thermal stabilities of duplexes formed between cDNAmc29 and avian DNAs also reflect phylogenetic divergence. Sequences complementary to cDNAmc29 are transcribed into approximately 10 copies per cell of polyadenylated RNA in uninfected chicken fibroblasts. Thus, the vertebrate homolog of cDNAmc29 may be a gene which has been conserved throughout vertebrate evolution and which served as a progenitor for the putative transforming gene of MC29V. Recent experiments suggest that the putative transforming gene of avian erythroblastosis virus, like that of MC29V, may have arisen by incorporation of a host gene (Stehelin et al., personal communication). These findings for avian erythroblastosis virus and MC29V closely parallel previous results, suggesting a host origin for src (D. H. Spector, B. Baker, H. E. Varmus, and J. M. Bishop, Cell 13:381-386, 1978; D. H. Spector, K. Smith, T. Padgett, P. McCombe, D. Roulland-Dussoix, C. Moscovici, H. E. Varmus, and J. M. Bishop, Cell 13:371-379, 1978; D. H. Spector, H. E. Varmus, and J. M. Bishop, Proc. Natl. Acad. Sci. U.S.A. 75:4102-4106, 1978; D

  18. Extra- and intracellular lactose catabolism in Penicillium chrysogenum: phylogenetic and expression analysis of the putative permease and hydrolase genes.

    Science.gov (United States)

    Jónás, Ágota; Fekete, Erzsébet; Flipphi, Michel; Sándor, Erzsébet; Jäger, Szilvia; Molnár, Ákos P; Szentirmai, Attila; Karaffa, Levente

    2014-07-01

    Penicillium chrysogenum is used as an industrial producer of penicillin. We investigated its catabolism of lactose, an abundant component of whey used in penicillin fermentation, comparing the type strain NRRL 1951 with the high producing strain AS-P-78. Both strains grew similarly on lactose as the sole carbon source under batch conditions, exhibiting almost identical time profiles of sugar depletion. In silico analysis of the genome sequences revealed that P. chrysogenum features at least five putative β-galactosidase (bGal)-encoding genes at the annotated loci Pc22g14540, Pc12g11750, Pc16g12750, Pc14g01510 and Pc06g00600. The first two proteins appear to be orthologs of two Aspergillus nidulans family 2 intracellular glycosyl hydrolases expressed on lactose. The latter three P. chrysogenum proteins appear to be distinct paralogs of the extracellular bGal from A. niger, LacA, a family 35 glycosyl hydrolase. The P. chrysogenum genome also specifies two putative lactose transporter genes at the annotated loci Pc16g06850 and Pc13g08630. These are orthologs of paralogs of the gene encoding the high-affinity lactose permease (lacpA) in A. nidulans for which P. chrysogenum appears to lack the ortholog. Transcript analysis of Pc22g14540 showed that it was expressed exclusively on lactose, whereas Pc12g11750 was weakly expressed on all carbon sources tested, including D-glucose. Pc16g12750 was co-expressed with the two putative intracellular bGal genes on lactose and also responded on L-arabinose. The Pc13g08630 transcript was formed exclusively on lactose. The data strongly suggest that P. chrysogenum exhibits a dual assimilation strategy for lactose, simultaneously employing extracellular and intracellular hydrolysis, without any correlation to the penicillin-producing potential of the studied strains.

  19. Identification of putative effector genes and their transcripts in three strains related to 'Candidatus Phytoplasma aurantifolia'.

    Science.gov (United States)

    Anabestani, Ameneh; Izadpanah, Keramat; Abbà, Simona; Galetto, Luciana; Ghorbani, Abozar; Palmano, Sabrina; Siampour, Majid; Veratti, Flavio; Marzachì, Cristina

    2017-06-01

    Molecular mechanisms underlying phytoplasma interactions with host plants are largely unknown. In this study attempts were made to identify effectors of three phytoplasma strains related to 'Ca. P. aurantifolia', crotalaria phyllody (CrP), faba bean phyllody (FBP), and witches' broom disease of lime (WBDL), using information from draft genome of peanut witches' broom phytoplasma. Seven putative effectors were identified in WBDL genome (SAP11, SAP21, Eff64, Eff115, Eff197, Eff211 and EffSAP67), five (SAP11, SAP21, Eff64, Eff99 and Eff197) in CrP and two (SAP11, Eff64) in FBP. No homologs to Eff64, Eff197 and Eff211 in phytoplasmas of other phylogenetic groups were found. SAP11 and Eff64 homologs of 'Ca. P. aurantifolia' strains shared at least 95.9% identity and were detected in the three phytoplasmas, supporting their role within the group. Five of the putative effectors (SAP11, SAP21, Eff64, Eff115, and Eff99) were transcribed from total RNA extracts of periwinkle plants infected with these phytoplasmas. Transcription profiles of selected putative effectors of CrP, FBP and WBDL indicated that SAP11 transcripts were the most abundant in the three phytoplasmas. SAP21 transcript levels were comparable to those of SAP11 for CrP and not measurable for the other phytoplasmas. Eff64 had the lowest transcription level irrespective of sampling date and phytoplasma isolate. Eff115 transcript levels were the highest in WBDL infected plants. This work reports the first sequence information for 14 putative effectors in three strains related to 'Ca. P. aurantifolia', and offers novel insight into the transcription profile of five of them during infection of periwinkle. Copyright © 2017 Elsevier GmbH. All rights reserved.

  20. Bioinformatic Analysis of Putative Gene Products Encoded in SARS-HCoV Genome

    Institute of Scientific and Technical Information of China (English)

    赵心刚; 韩敬东; 宁元亨; 孟安明; 陈晔光

    2003-01-01

    The cause of severe acute respiratory syndrome (SARS) has been identified as a new coronavirus named as SARS-HCoV.Using bioinformatic methods, we have performed a detailed domain search.In addition to the viral structure proteins, we have found that several putative polypeptides share sequence similarity to known domains or proteins.This study may provide a basis for future studies on the infection and replication process of this notorious virus.

  1. Prevalence of Putative Virulence Genes in Campylobacter and Arcobacter Species Isolated from Poultry and Poultry By-Products in Tunisia.

    Science.gov (United States)

    Jribi, Hela; Sellami, Hanen; Hassena, Amal Ben; Gdoura, Radhouane

    2017-10-01

    Campylobacter and Arcobacter spp. are common causes of gastroenteritis in humans; these infections are commonly due to undercooked poultry. However, their virulence mechanism is still poorly understood. The aim of this study was to evaluate the presence of genotypic virulence markers in Campylobacter and Arcobacter species using PCR. The prevalence of virulence and cytolethal distending toxin (CDT) genes was estimated in 71 Campylobacteraceae isolates. PCR was used to detect the presence of virulence genes (iam, cadF, virB1, flaA, cdtA, cdtB, and cdtC) using specific primers for a total of 45 Campylobacter isolates, including 37 C. jejuni and 8 C. coli. All the Campylobacter isolates were positive for the cadF gene. The plasmid gene virB11 was not detected in any strain. The invasion associated marker was not detected in C. jejuni. Lower detection rates were observed for flaA, cdtA, cdtB, and cdtC. The presence of nine putative Arcobacter virulence genes (cadF, ciaB, cj1349, mviN, pldA, tlyA, irgA, hecA, and hecB) was checked in a set of 22 Arcobacter butzleri and 4 Arcobacter cryaerophilus isolates. The pldA and mviN genes were predominant (88.64%). Lower detection rates were observed for tlyA (84.76%), ciaB (84.61%), cadF and cj1349 (76.92%), IrgA and hecA (61.53%), and hecB (57.69%). The findings revealed that a majority of the Campylobacteraceae strains have these putative virulence genes that may lead to pathogenic effects in humans.

  2. Whole-genome survey of the putative ATP-binding cassette transporter family genes in Vitis vinifera.

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    Birsen Çakır

    Full Text Available The ATP-binding cassette (ABC protein superfamily constitutes one of the largest protein families known in plants. In this report, we performed a complete inventory of ABC protein genes in Vitis vinifera, the whole genome of which has been sequenced. By comparison with ABC protein members of Arabidopsis thaliana, we identified 135 putative ABC proteins with 1 or 2 NBDs in V. vinifera. Of these, 120 encode intrinsic membrane proteins, and 15 encode proteins missing TMDs. V. vinifera ABC proteins can be divided into 13 subfamilies with 79 "full-size," 41 "half-size," and 15 "soluble" putative ABC proteins. The main feature of the Vitis ABC superfamily is the presence of 2 large subfamilies, ABCG (pleiotropic drug resistance and white-brown complex homolog and ABCC (multidrug resistance-associated protein. We identified orthologs of V. vinifera putative ABC transporters in different species. This work represents the first complete inventory of ABC transporters in V. vinifera. The identification of Vitis ABC transporters and their comparative analysis with the Arabidopsis counterparts revealed a strong conservation between the 2 species. This inventory could help elucidate the biological and physiological functions of these transporters in V. vinifera.

  3. Whole-genome survey of the putative ATP-binding cassette transporter family genes in Vitis vinifera.

    Science.gov (United States)

    Çakır, Birsen; Kılıçkaya, Ozan

    2013-01-01

    The ATP-binding cassette (ABC) protein superfamily constitutes one of the largest protein families known in plants. In this report, we performed a complete inventory of ABC protein genes in Vitis vinifera, the whole genome of which has been sequenced. By comparison with ABC protein members of Arabidopsis thaliana, we identified 135 putative ABC proteins with 1 or 2 NBDs in V. vinifera. Of these, 120 encode intrinsic membrane proteins, and 15 encode proteins missing TMDs. V. vinifera ABC proteins can be divided into 13 subfamilies with 79 "full-size," 41 "half-size," and 15 "soluble" putative ABC proteins. The main feature of the Vitis ABC superfamily is the presence of 2 large subfamilies, ABCG (pleiotropic drug resistance and white-brown complex homolog) and ABCC (multidrug resistance-associated protein). We identified orthologs of V. vinifera putative ABC transporters in different species. This work represents the first complete inventory of ABC transporters in V. vinifera. The identification of Vitis ABC transporters and their comparative analysis with the Arabidopsis counterparts revealed a strong conservation between the 2 species. This inventory could help elucidate the biological and physiological functions of these transporters in V. vinifera.

  4. Transcriptome Analysis of Mango (Mangifera indica L.) Fruit Epidermal Peel to Identify Putative Cuticle-Associated Genes.

    Science.gov (United States)

    Tafolla-Arellano, Julio C; Zheng, Yi; Sun, Honghe; Jiao, Chen; Ruiz-May, Eliel; Hernández-Oñate, Miguel A; González-León, Alberto; Báez-Sañudo, Reginaldo; Fei, Zhangjun; Domozych, David; Rose, Jocelyn K C; Tiznado-Hernández, Martín E

    2017-04-20

    Mango fruit (Mangifera indica L.) are highly perishable and have a limited shelf life, due to postharvest desiccation and senescence, which limits their global distribution. Recent studies of tomato fruit suggest that these traits are influenced by the expression of genes that are associated with cuticle metabolism. However, studies of these phenomena in mango fruit are limited by the lack of genome-scale data. In order to gain insight into the mango cuticle biogenesis and identify putative cuticle-associated genes, we analyzed the transcriptomes of peels from ripe and overripe mango fruit using RNA-Seq. Approximately 400 million reads were generated and de novo assembled into 107,744 unigenes, with a mean length of 1,717 bp and with this information an online Mango RNA-Seq Database (http://bioinfo.bti.cornell.edu/cgi-bin/mango/index.cgi) which is a valuable genomic resource for molecular research into the biology of mango fruit was created. RNA-Seq analysis suggested that the pathway leading to biosynthesis of the cuticle component, cutin, is up-regulated during overripening. This data was supported by analysis of the expression of several putative cuticle-associated genes and by gravimetric and microscopic studies of cuticle deposition, revealing a complex continuous pattern of cuticle deposition during fruit development and involving substantial accumulation during ripening/overripening.

  5. Transcriptome Analysis of Mango (Mangifera indica L.) Fruit Epidermal Peel to Identify Putative Cuticle-Associated Genes

    Science.gov (United States)

    Tafolla-Arellano, Julio C.; Zheng, Yi; Sun, Honghe; Jiao, Chen; Ruiz-May, Eliel; Hernández-Oñate, Miguel A.; González-León, Alberto; Báez-Sañudo, Reginaldo; Fei, Zhangjun; Domozych, David; Rose, Jocelyn K. C.; Tiznado-Hernández, Martín E.

    2017-04-01

    Mango fruit (Mangifera indica L.) are highly perishable and have a limited shelf life, due to postharvest desiccation and senescence, which limits their global distribution. Recent studies of tomato fruit suggest that these traits are influenced by the expression of genes that are associated with cuticle metabolism. However, studies of these phenomena in mango fruit are limited by the lack of genome-scale data. In order to gain insight into the mango cuticle biogenesis and identify putative cuticle-associated genes, we analyzed the transcriptomes of peels from ripe and overripe mango fruit using RNA-Seq. Approximately 400 million reads were generated and de novo assembled into 107,744 unigenes, with a mean length of 1,717 bp and with this information an online Mango RNA-Seq Database (http://bioinfo.bti.cornell.edu/cgi-bin/mango/index.cgi) which is a valuable genomic resource for molecular research into the biology of mango fruit was created. RNA-Seq analysis suggested that the pathway leading to biosynthesis of the cuticle component, cutin, is up-regulated during overripening. This data was supported by analysis of the expression of several putative cuticle-associated genes and by gravimetric and microscopic studies of cuticle deposition, revealing a complex continuous pattern of cuticle deposition during fruit development and involving substantial accumulation during ripening/overripening.

  6. The presence of the putative Gardnerella vaginalis sialidase A gene in vaginal specimens is associated with bacterial vaginosis biofilm

    Science.gov (United States)

    Jespers, Vicky; Van den Bulck, Magelien; Buyze, Jozefien; Mwambarangwe, Lambert; Musengamana, Viateur; Vaneechoutte, Mario; Crucitti, Tania

    2017-01-01

    Bacterial vaginosis (BV) is a difficult-to-treat recurrent condition in which health-associated lactobacilli are outnumbered by other anaerobic bacteria, such as Gardnerella vaginalis. Certain genotypes of G. vaginalis can produce sialidase, while others cannot. Sialidase is known to facilitate the destruction of the protective mucus layer on the vaginal epithelium by hydrolysis of sialic acid on the glycans of mucous membranes. This process possibly facilitates adhesion of bacterial cells on the epithelium since it has been linked with the development of biofilm in other pathogenic conditions. Although it has not been demonstrated yet, it is probable that G. vaginalis benefits from this mechanism by attaching to the vaginal epithelium to initiate biofilm development. In this study, using vaginal specimens of 120 women enrolled in the Ring Plus study, we assessed the association between the putative G. vaginalis sialidase A gene by quantitative polymerase chain reaction (qPCR), the diagnosis of BV according to Nugent score, and the occurrence of a BV-associated biofilm dominated by G. vaginalis by fluorescence in situ hybridisation (FISH). We detected the putative sialidase A gene in 75% of the G. vaginalis-positive vaginal specimens and found a strong association (p<0.001) between the presence of a G. vaginalis biofilm, the diagnosis of BV according to Nugent and the detection of high loads of the G. vaginalis sialidase A gene in the vaginal specimens. These results could redefine diagnosis of BV, and in addition might guide research for new treatment. PMID:28241058

  7. Sequence analysis and gene expression of putative exo- and endo-glucanases from oil palm (Elaeis guineensis) during fungal infection.

    Science.gov (United States)

    Yeoh, Keat-Ai; Othman, Abrizah; Meon, Sariah; Abdullah, Faridah; Ho, Chai-Ling

    2012-10-15

    Glucanases are enzymes that hydrolyze a variety β-d-glucosidic linkages. Plant β-1,3-glucanases are able to degrade fungal cell walls; and promote the release of cell-wall derived fungal elicitors. In this study, three full-length cDNA sequences encoding oil palm (Elaeis guineensis) glucanases were analyzed. Sequence analyses of the cDNA sequences suggested that EgGlc1-1 is a putative β-d-glucan exohydolase belonging to glycosyl hydrolase (GH) family 3 while EgGlc5-1 and EgGlc5-2 are putative glucan endo-1,3-β-glucosidases belonging to GH family 17. The transcript abundance of these genes in the roots and leaves of oil palm seedlings treated with Ganoderma boninense and Trichoderma harzianum was profiled to investigate the involvement of these glucanases in oil palm during fungal infection. The gene expression of EgGlc1-1 in the root of oil palm seedlings was increased by T. harzianum but suppressed by G. boninense; while the gene expression of both EgGlc5-1 and EgGlc5-2 in the roots of oil palm seedlings was suppressed by G. boninense or/and T. harzianum. Copyright © 2012 Elsevier GmbH. All rights reserved.

  8. Expression of putative zinc-finger protein lcn61 gene in lymphocystis disease virus China (LCDV-cn) genome

    Institute of Scientific and Technical Information of China (English)

    YAN Xiuying; SUN Xiuqin

    2009-01-01

    An open reading frame (lcn61) of iymphocystis disease virus China (LCDV-cn), probably responsible for encoding putative zinc-finger proteins was amplified and inserted into pET24a (+) vector.Then it expressed in E. coli BL21 (DE3), and His-tag fusion protein of high yield was obtained. It was found that the fusion protein existed in E. coli mainly as inclusion bodies. The bioinformatics analysis indicates that LCN61 is C2H2 type zinc-finger protein containing four C2H2 zinc-finger motifs. This work provides a theory for functional research of lcn61 gene.

  9. Quantitative Trait Locus and Genetical Genomics Analysis Identifies Putatively Causal Genes for Fecundity and Brooding in the Chicken

    Directory of Open Access Journals (Sweden)

    Martin Johnsson

    2016-02-01

    Full Text Available Life history traits such as fecundity are important to evolution because they make up components of lifetime fitness. Due to their polygenic architectures, such traits are difficult to investigate with genetic mapping. Therefore, little is known about their molecular basis. One possible way toward finding the underlying genes is to map intermediary molecular phenotypes, such as gene expression traits. We set out to map candidate quantitative trait genes for egg fecundity in the chicken by combining quantitative trait locus mapping in an advanced intercross of wild by domestic chickens with expression quantitative trait locus mapping in the same birds. We measured individual egg fecundity in 232 intercross chickens in two consecutive trials, the second one aimed at measuring brooding. We found 12 loci for different aspects of egg fecundity. We then combined the genomic confidence intervals of these loci with expression quantitative trait loci from bone and hypothalamus in the same intercross. Overlaps between egg loci and expression loci, and trait–gene expression correlations identify 29 candidates from bone and five from hypothalamus. The candidate quantitative trait genes include fibroblast growth factor 1, and mitochondrial ribosomal proteins L42 and L32. In summary, we found putative quantitative trait genes for egg traits in the chicken that may have been affected by regulatory variants under chicken domestication. These represent, to the best of our knowledge, some of the first candidate genes identified by genome-wide mapping for life history traits in an avian species.

  10. Quantitative Trait Locus and Genetical Genomics Analysis Identifies Putatively Causal Genes for Fecundity and Brooding in the Chicken.

    Science.gov (United States)

    Johnsson, Martin; Jonsson, Kenneth B; Andersson, Leif; Jensen, Per; Wright, Dominic

    2015-12-04

    Life history traits such as fecundity are important to evolution because they make up components of lifetime fitness. Due to their polygenic architectures, such traits are difficult to investigate with genetic mapping. Therefore, little is known about their molecular basis. One possible way toward finding the underlying genes is to map intermediary molecular phenotypes, such as gene expression traits. We set out to map candidate quantitative trait genes for egg fecundity in the chicken by combining quantitative trait locus mapping in an advanced intercross of wild by domestic chickens with expression quantitative trait locus mapping in the same birds. We measured individual egg fecundity in 232 intercross chickens in two consecutive trials, the second one aimed at measuring brooding. We found 12 loci for different aspects of egg fecundity. We then combined the genomic confidence intervals of these loci with expression quantitative trait loci from bone and hypothalamus in the same intercross. Overlaps between egg loci and expression loci, and trait-gene expression correlations identify 29 candidates from bone and five from hypothalamus. The candidate quantitative trait genes include fibroblast growth factor 1, and mitochondrial ribosomal proteins L42 and L32. In summary, we found putative quantitative trait genes for egg traits in the chicken that may have been affected by regulatory variants under chicken domestication. These represent, to the best of our knowledge, some of the first candidate genes identified by genome-wide mapping for life history traits in an avian species.

  11. Validation of candidate genes putatively associated with resistance to SCMV and MDMV in maize (Zea mays L. by expression profiling

    Directory of Open Access Journals (Sweden)

    Wenzel Gerhard

    2009-02-01

    Full Text Available Abstract Background The potyviruses sugarcane mosaic virus (SCMV and maize dwarf mosaic virus (MDMV are major pathogens of maize worldwide. Two loci, Scmv1 and Scmv2, have ealier been shown to confer complete resistance to SCMV. Custom-made microarrays containing previously identified SCMV resistance candidate genes and resistance gene analogs were utilised to investigate and validate gene expression and expression patterns of isogenic lines under pathogen infection in order to obtain information about the molecular mechanisms involved in maize-potyvirus interactions. Results By employing time course microarray experiments we identified 68 significantly differentially expressed sequences within the different time points. The majority of differentially expressed genes differed between the near-isogenic line carrying Scmv1 resistance locus at chromosome 6 and the other isogenic lines. Most differentially expressed genes in the SCMV experiment (75% were identified one hour after virus inoculation, and about one quarter at multiple time points. Furthermore, most of the identified mapped genes were localised outside the Scmv QTL regions. Annotation revealed differential expression of promising pathogenesis-related candidate genes, validated by qRT-PCR, coding for metallothionein-like protein, S-adenosylmethionine synthetase, germin-like protein or 26S ribosomal RNA. Conclusion Our study identified putative candidate genes and gene expression patterns related to resistance to SCMV. Moreover, our findings support the effectiveness and reliability of the combination of different expression profiling approaches for the identification and validation of candidate genes. Genes identified in this study represent possible future targets for manipulation of SCMV resistance in maize.

  12. Identification of multiple putative S-layer genes partly expressed by Lysinibacillus sphaericus JG-B53.

    Science.gov (United States)

    Lederer, Franziska L; Weinert, Ulrike; Günther, Tobias J; Raff, Johannes; Weiß, Stephan; Pollmann, Katrin

    2013-06-01

    Lysinibacillus sphaericus JG-B53 was isolated from the uranium mining waste pile Haberland near Johanngeorgenstadt, Germany. Previous studies have shown that many bacteria that have been isolated from these heavy metal contaminated environments possess surface layer (S-layer) proteins that enable the bacteria to survive by binding metals with high affinity. Conversely, essential trace elements are able to cross the filter layer and reach the interior of the cell. This is especially true of the S-layer of L. sphaericus JG-B53, which possesses outstanding recrystallization and metal-binding properties. In this study, S-layer protein gene sequences encoded in the genome of L. sphaericus JG-B53 were identified using next-generation sequencing technology followed by bioinformatic analyses. The genome of L. sphaericus JG-B53 encodes at least eight putative S-layer protein genes with distinct differences. Using mRNA analysis the expression of the putative S-layer protein genes was studied. The functional S-layer protein B53 Slp1 was identified as the dominantly expressed S-layer protein in L. sphaericus JG-B53 by mRNA studies, SDS-PAGE and N-terminal sequencing. B53 Slp1 is characterized by square lattice symmetry and a molecular mass of 116 kDa. The S-layer protein B53 Slp1 shows a high similarity to the functional S-layer protein of L. sphaericus JG-A12, which was isolated from the same uranium mining waste pile Haberland and has been described by previous research. These similarities indicate horizontal gene transfer and DNA rearrangements between these bacteria. The presence of multiple S-layer gene copies may enable the bacterial strains to quickly adapt to changing environments.

  13. Identification of putative regulatory motifs in the upstream regions of co-expressed functional groups of genes in Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Joshi NV

    2009-01-01

    Full Text Available Abstract Background Regulation of gene expression in Plasmodium falciparum (Pf remains poorly understood. While over half the genes are estimated to be regulated at the transcriptional level, few regulatory motifs and transcription regulators have been found. Results The study seeks to identify putative regulatory motifs in the upstream regions of 13 functional groups of genes expressed in the intraerythrocytic developmental cycle of Pf. Three motif-discovery programs were used for the purpose, and motifs were searched for only on the gene coding strand. Four motifs – the 'G-rich', the 'C-rich', the 'TGTG' and the 'CACA' motifs – were identified, and zero to all four of these occur in the 13 sets of upstream regions. The 'CACA motif' was absent in functional groups expressed during the ring to early trophozoite transition. For functional groups expressed in each transition, the motifs tended to be similar. Upstream motifs in some functional groups showed 'positional conservation' by occurring at similar positions relative to the translational start site (TLS; this increases their significance as regulatory motifs. In the ribonucleotide synthesis, mitochondrial, proteasome and organellar translation machinery genes, G-rich, C-rich, CACA and TGTG motifs, respectively, occur with striking positional conservation. In the organellar translation machinery group, G-rich motifs occur close to the TLS. The same motifs were sometimes identified for multiple functional groups; differences in location and abundance of the motifs appear to ensure different modes of action. Conclusion The identification of positionally conserved over-represented upstream motifs throws light on putative regulatory elements for transcription in Pf.

  14. Characterization of a putative pollen-specific arabinogalactan protein gene, BcMF8, from Brassica campestris ssp. chinensis.

    Science.gov (United States)

    Huang, Li; Cao, Jia-Shu; Zhang, Ai-Hong; Ye, Yi-Qun

    2008-12-01

    The BcMF8 (Brassica campestris male fertility 8) gene, possessing the features of 'classical' arabinogalactan protein (AGP) was isolated from Brassica campestris L. ssp. chinensis, Makino syn. B. rapa L. ssp. chinensis. This gene was highly abundant in the fertile flower buds but silenced in the sterile ones of genic male sterile A/B line ('ZUBajh97-01A/B') in B. campestris. Expression patterns analysis suggested BcMF8 was a pollen-specific gene, whose transcript started to be expressed at the uninucleate stage and maintained throughout to the pollen at pollination stage. BcMF8 is highly homologous to the known pollen-specific AGP genes Sta 39-4 and Sta 39-3 from B. napus. Isolation and multiple alignment of the homologs of BcMF8 gene in the family Cruciferae indicated that BcMF8 was highly conserved in this family, which reflect the conservation in biological function and importance of this putative AGP gene in plant development. Similarity analysis also demonstrated Sta 39-4 and Sta 39-3 may originate from different genomes.

  15. Spatial patterns of diversity at the putative recognition domain of resistance gene candidates in wild bean populations.

    Science.gov (United States)

    de Meaux, J; Neema, C

    2003-01-01

    Leucine Rich Repeats (LRR) domains have been identified on most known plant resistance genes and appear to be involved in the specific recognition of pathogen strains. Here we explore the processes which may drive the evolution of this putative recognition domain. We developed AFLP markers specifically situated in the LRR domain of members of the PRLJ1 complex Resistance Gene Candidate (RGC) family identified in common bean (Phaseolus vulgaris). Diversity for these markers was assessed in ten wild populations of P. vulgaris and compared to locally co-occurring pathogen populations of Colletotrichum lindemuthianum. Nine PRLJ1 LRR specific markers were obtained. Marker sequences revealed that RGC diversity at PRLJ1 is similar to that at other complex R-loci. Wild bean populations showed contrasting levels of PRLJ1 LRR diversity and were all significantly differentiated. We could not detect an effect of local C. lindemuthianum population diversity on the spatial distribution of P. vulgaris PRLJ1 diversity. However, host populations have been previously assessed for neutral (RAPD) markers and for resistance phenotypes to six strains of C. lindemuthianum isolated from cultivated bean fields. A comparative analysis of PRLJ1 LRR diversity and host diversity for resistance phenotypes indicated that evolutionary processes related to the antagonistic C. lindemuthianum/P. vulgaris interaction are likely to have shaped molecular diversity of the putative recognition domains of the PRLJ1 RGC family members.

  16. Gene transcript accumulation and in situ mRNA hybridization of two putative glutamate dehydrogenase genes in etiolated Glycine max seedlings.

    Science.gov (United States)

    Dimou, M; Tsaniklidis, G; Aivalakis, G; Katinakis, P

    2015-01-01

    Glutamate dehydrogenase (EC 1.4.1.2) is a multimeric enzyme that catalyzes the reversible amination of α-ketoglutarate to form glutamate. We characterized cDNA clones of two Glycine max sequences, GmGDH1 and GmGDH2, that code for putative α- and β-subunits, respectively, of the NADH dependent enzyme. Temporal and spatial gene transcript accumulation studies using semiquantitative RT-PCR and in situ hybridization have shown an overlapping gene transcript accumulation pattern with differences in relative gene transcript accumulation in the organs examined. Detection of NADH-dependent glutamate dehydrogenase activity in situ using a histochemical method showed concordance with the spatial gene transcript accumulation patterns. Our findings suggest that although the two gene transcripts are co-localized in roots of etiolated soybean seedlings, the ratio of the two subunits of the active holoenzyme may vary among tissues.

  17. Classification of genes and putative biomarker identification using distribution metrics on expression profiles.

    Directory of Open Access Journals (Sweden)

    Hung-Chung Huang

    Full Text Available BACKGROUND: Identification of genes with switch-like properties will facilitate discovery of regulatory mechanisms that underlie these properties, and will provide knowledge for the appropriate application of Boolean networks in gene regulatory models. As switch-like behavior is likely associated with tissue-specific expression, these gene products are expected to be plausible candidates as tissue-specific biomarkers. METHODOLOGY/PRINCIPAL FINDINGS: In a systematic classification of genes and search for biomarkers, gene expression profiles (GEPs of more than 16,000 genes from 2,145 mouse array samples were analyzed. Four distribution metrics (mean, standard deviation, kurtosis and skewness were used to classify GEPs into four categories: predominantly-off, predominantly-on, graded (rheostatic, and switch-like genes. The arrays under study were also grouped and examined by tissue type. For example, arrays were categorized as 'brain group' and 'non-brain group'; the Kolmogorov-Smirnov distance and Pearson correlation coefficient were then used to compare GEPs between brain and non-brain for each gene. We were thus able to identify tissue-specific biomarker candidate genes. CONCLUSIONS/SIGNIFICANCE: The methodology employed here may be used to facilitate disease-specific biomarker discovery.

  18. Characterization of five putative aspartate aminotransferase genes in the N2-fixing heterocystous cyanobacterium Anabaena sp. strain PCC 7120.

    Science.gov (United States)

    Xu, Xinyi; Gu, Liping; He, Ping; Zhou, Ruanbao

    2015-06-01

    Aspartate and glutamate are two key amino acids used in biosynthesis of many amino acids that play vital role in cellular metabolism. Aspartate aminotransferases (AspATs) are required for channelling nitrogen (N(2)) between Glu and Asp in all life forms. Biochemical and genetic characterization of AspATs have been lacking in N(2)-fixing cyanobacteria. In this report, five putative AspAT genes (alr1039, all2340, alr2765, all4327 and alr4853) were identified in the N(2)-fixing heterocystous cyanobacterium Anabaena sp. PCC 7120. Five recombinant C-terminal hexahistidine-tagged AspATs (AspAT-H(6)) were overexpressed in Escherichia coli and purified to homogeneity. Biochemical analysis demonstrated that these five putative AspATs have authentic AspAT activity in vitro using aspartate as an amino donor. However, the enzymic activities of the five AspATs differed in vitro. Alr4853-H(6) showed the highest AspAT activity, while the enzymic activity for the other four AspATs ranged from 6.5 to 53.7 % activity compared to Alr4853 (100 %). Genetic characterization of the five AspAT genes was also performed by inactivating each individual gene. All of the five AspAT knockout mutants exhibited reduced diazotrophic growth, and alr4853 was further identified to be a Fox gene (requiring fixed N(2) for growth in the presence of oxygen). Four out of five P(aspAT)-gfp transcriptional fusions were constitutively expressed in both diazotrophic and nitrate-dependent growth conditions. Quantitative reverse transcriptase PCR showed that alr4853 expression was increased by 2.3-fold after 24 h of N(2) deprivation. Taken together, these findings add to our understanding of the role of AspATs in N(2)-fixing within heterocystous cyanobacteria.

  19. The gene corresponding to the putative Goodpasture antigen is present in Alport's syndrome.

    Science.gov (United States)

    Savige, J A

    1991-08-01

    Alport's syndrome is a heterogeneous group of inherited abnormalities of basement membranes that may result in progressive renal failure, defective hearing and lens abnormalities. The glomerular basement membrane (GBM) characteristically has areas of reduplication, lamellation and attenuation on electron microscopic examination. In the majority of affected males and some females, there is reduced or variable binding of serum from patients with anti-GBM disease (Goodpasture's syndrome) to these basement membranes. These sera contain antibodies directed against the Goodpasture antigen which has been thought to be located in the non-collagenous domain of the alpha3 chain of type IV collagen and is presumed to be important in cross-linking of the collagen molecules. The reduced staining for the Goodpasture antigen suggests that this structure is either absent or masked in Alport's syndrome. We have tested DNA from six unrelated individuals with Alport's syndrome. All had been transplanted for renal failure. The diagnosis of Alport's syndrome was made on the characteristic electron microscopic appearance of the renal basement membranes (n = 4), the presence of sensori-neural deafness (n = 4), a family history of Alport's syndrome (n = 5) and the presence of circulating inhibitable anti-GBM antibody activity post-transplant (n = 2). Oligonucleotides (20mers) corresponding to the 5' and 3' ends of the known 25 amino acid sequence for the putative Goodpasture antigen were used as primers for amplification of genomic DNA. The products were then blotted and probed with an intermediate 19-mer DNA. All Alport's patients contained a 75-bp fragment corresponding to the published peptide sequence for the non-collagenous domain of the alpha 3 chain of type IV collagen, suggesting that a large deletion of this region, the putative Goodpasture antigen, is unlikely to account for the defect in Alport's syndrome.

  20. SlDEAD31, a Putative DEAD-Box RNA Helicase Gene, Regulates Salt and Drought Tolerance and Stress-Related Genes in Tomato

    OpenAIRE

    Mingku Zhu; Guoping Chen; Tingting Dong; Lingling Wang; Jianling Zhang; Zhiping Zhao; Zongli Hu

    2015-01-01

    The DEAD-box RNA helicases are involved in almost every aspect of RNA metabolism, associated with diverse cellular functions including plant growth and development, and their importance in response to biotic and abiotic stresses is only beginning to emerge. However, none of DEAD-box genes was well characterized in tomato so far. In this study, we reported on the identification and characterization of two putative DEAD-box RNA helicase genes, SlDEAD30 and SlDEAD31 from tomato, which were class...

  1. Identification of putative noncoding RNA genes in the Burkholderia cenocepacia J2315 genome

    DEFF Research Database (Denmark)

    Coenye, T.; Drevinek, P.; Mahenthiralingam, E.

    2007-01-01

    Noncoding RNA (ncRNA) genes are not involved in the production of mRNA and proteins, but produce transcripts that function directly as structural or regulatory RNAs. In the present study, the presence of ncRNA genes in the genome of Burkholderia cenocepacia J2315 was evaluated by combining compar...

  2. Digital transcriptome analysis of putative sex-determination genes in papaya (Carica papaya.

    Directory of Open Access Journals (Sweden)

    Naoya Urasaki

    Full Text Available Papaya (Carica papaya is a trioecious plant species that has male, female and hermaphrodite flowers on different plants. The primitive sex chromosomes genetically determine the sex of the papaya. Although draft sequences of the papaya genome are already available, the genes for sex determination have not been identified, likely due to the complicated structure of its sex-chromosome sequences. To identify the candidate genes for sex determination, we conducted a transcriptome analysis of flower samples from male, female and hermaphrodite plants using high-throughput SuperSAGE for digital gene expression analysis. Among the short sequence tags obtained from the transcripts, 312 unique tags were specifically mapped to the primitive sex chromosome (X or Y(h sequences. An annotation analysis revealed that retroelements are the most abundant sequences observed in the genes corresponding to these tags. The majority of tags on the sex chromosomes were located on the X chromosome, and only 30 tags were commonly mapped to both the X and Y(h chromosome, implying a loss of many genes on the Y(h chromosome. Nevertheless, candidate Y(h chromosome-specific female determination genes, including a MADS-box gene, were identified. Information on these sex chromosome-specific expressed genes will help elucidating sex determination in the papaya.

  3. Digital transcriptome analysis of putative sex-determination genes in papaya (Carica papaya).

    Science.gov (United States)

    Urasaki, Naoya; Tarora, Kazuhiko; Shudo, Ayano; Ueno, Hiroki; Tamaki, Moritoshi; Miyagi, Norimichi; Adaniya, Shinichi; Matsumura, Hideo

    2012-01-01

    Papaya (Carica papaya) is a trioecious plant species that has male, female and hermaphrodite flowers on different plants. The primitive sex chromosomes genetically determine the sex of the papaya. Although draft sequences of the papaya genome are already available, the genes for sex determination have not been identified, likely due to the complicated structure of its sex-chromosome sequences. To identify the candidate genes for sex determination, we conducted a transcriptome analysis of flower samples from male, female and hermaphrodite plants using high-throughput SuperSAGE for digital gene expression analysis. Among the short sequence tags obtained from the transcripts, 312 unique tags were specifically mapped to the primitive sex chromosome (X or Y(h)) sequences. An annotation analysis revealed that retroelements are the most abundant sequences observed in the genes corresponding to these tags. The majority of tags on the sex chromosomes were located on the X chromosome, and only 30 tags were commonly mapped to both the X and Y(h) chromosome, implying a loss of many genes on the Y(h) chromosome. Nevertheless, candidate Y(h) chromosome-specific female determination genes, including a MADS-box gene, were identified. Information on these sex chromosome-specific expressed genes will help elucidating sex determination in the papaya.

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

    Science.gov (United States)

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

    2011-07-01

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

  5. Genome-wide digital transcript analysis of putative fruitlet abscission related genes regulated by ethephon in litchi

    Directory of Open Access Journals (Sweden)

    Caiqin eLi

    2015-07-01

    Full Text Available The high level of physiological fruitlet abscission in litchi (Litchi chinensis Sonn. causes severe yield loss. Cell separation occurs at the fruit abscission zone (FAZ and can be triggered by ethylene. However, a deep knowledge of the molecular events occurring in the FAZ is still unknown. Here, genome-wide digital transcript abundance (DTA analysis of putative fruit abscission related genes regulated by ethephon in litchi were studied. More than 81 million high quality reads from seven ethephon treated and untreated control libraries were obtained by high-throughput sequencing. Through DTA profile analysis in combination with Gene Ontology and KEGG pathway enrichment analyses, a total of 2,730 statistically significant candidate genes were involved in the ethephon-promoted litchi fruitlet abscission. Of these, there were 1,867 early-responsive genes whose expressions were up- or down-regulated from 0 to 1 d after treatment. The most affected genes included those related to ethylene biosynthesis and signaling, auxin transport and signaling, transcription factors, protein ubiquitination, ROS response, calcium signal transduction and cell wall modification. These genes could be clustered into 4 groups and 13 subgroups according to their similar expression patterns. qRT-PCR displayed the expression pattern of 41 selected candidate genes, which proved the accuracy of our DTA data. Ethephon treatment significantly increased fruit abscission and ethylene production of fruitlet. The possible molecular events to control the ethephon-promoted litchi fruitlet abscission were prompted out. The increased ethylene evolution in fruitlet would suppress the synthesis and polar transport of auxin and trigger abscission signaling. To the best of our knowledge, it is the first time to monitor the gene expression profile occurring in the FAZ-enriched pedicel during litchi fruit abscission induced by ethephon on the genome-wide level. This study will contribute to

  6. Differential expression of putative drug resistance genes in Mycobacterium tuberculosis clinical isolates.

    Science.gov (United States)

    González-Escalante, Laura; Peñuelas-Urquides, Katia; Said-Fernández, Salvador; Silva-Ramírez, Beatriz; Bermúdez de León, Mario

    2015-12-01

    Understanding drug resistance in Mycobacterium tuberculosis requires an integrated analysis of strain lineages, mutations and gene expression. Previously, we reported the differential expression of esxG, esxH, infA, groES, rpmI, rpsA and lipF genes in a sensitive M. tuberculosis strain and in a multidrug-resistant clinical isolate. Here, we have evaluated the expression of these genes in 24 clinical isolates that belong to different lineages and have different drug resistance profiles. In vitro, growth kinetics analysis showed no difference in the growth of the clinical isolates, and thus drug resistance occurred without a fitness cost. However, a quantitative reverse transcription PCR analysis of gene expression revealed high variability among the clinical isolates, including those with similar drug resistance profiles. Due to the complexity of gene regulation pathways and the wide diversity of M. tuberculosis lineages, the use of gene expression as a molecular signature for drug resistance is not straightforward. Therefore, we recommend that the expression of M. tuberculosis genes be performed individually, and baseline expression levels should be verified among several different clinical isolates, before any further applications of these findings.

  7. An analysis of sequence variability in eight genes putatively involved in drought response in sunflower (Helianthus annuus L.).

    Science.gov (United States)

    Giordani, T; Buti, M; Natali, L; Pugliesi, C; Cattonaro, F; Morgante, M; Cavallini, A

    2011-04-01

    With the aim to study variability in genes involved in ecological adaptations, we have analysed sequence polymorphisms of eight unique genes putatively involved in drought response by isolation and analysis of allelic sequences in eight inbred lines of sunflower of different origin and phenotypic characters and showing different drought response in terms of leaf relative water content (RWC). First, gene sequences were amplified by PCR on genomic DNA from a highly inbred line and their products were directly sequenced. In the absence of single nucleotide polymorphisms, the gene was considered as unique. Then, the same PCR reaction was performed on genomic DNAs of eight inbred lines to isolate allelic variants to be compared. The eight selected genes encode a dehydrin, a heat shock protein, a non-specific lipid transfer protein, a z-carotene desaturase, a drought-responsive-element-binding protein, a NAC-domain transcription regulator, an auxin-binding protein, and an ABA responsive-C5 protein. Nucleotide diversity per synonymous and non-synonymous sites was calculated for each gene sequence. The π (a)/π (s) ratio range was usually very low, indicating strong purifying selection, though with locus-to-locus differences. As far as non-coding regions, the intron showed a larger variability than the other regions only in the case of the dehydrin gene. In the other genes tested, in which one or more introns occur, variability in the introns was similar or even lower than in the other regions. On the contrary, 3'-UTRs were usually more variable than the coding regions. Linkage disequilibrium in the selected genes decayed on average within 1,000 bp, with large variation among genes. A pairwise comparison between genetic distances calculated on the eight genes and the difference in RWC showed a significant correlation in the first phases of drought stress. The results are discussed in relation to the function of analysed genes, i.e. involved in gene regulation and signal

  8. A putative gene cluster from a Lyngbya wollei bloom that encodes paralytic shellfish toxin biosynthesis.

    Directory of Open Access Journals (Sweden)

    Troco K Mihali

    Full Text Available Saxitoxin and its analogs cause the paralytic shellfish-poisoning syndrome, adversely affecting human health and coastal shellfish industries worldwide. Here we report the isolation, sequencing, annotation, and predicted pathway of the saxitoxin biosynthetic gene cluster in the cyanobacterium Lyngbya wollei. The gene cluster spans 36 kb and encodes enzymes for the biosynthesis and export of the toxins. The Lyngbya wollei saxitoxin gene cluster differs from previously identified saxitoxin clusters as it contains genes that are unique to this cluster, whereby the carbamoyltransferase is truncated and replaced by an acyltransferase, explaining the unique toxin profile presented by Lyngbya wollei. These findings will enable the creation of toxin probes, for water monitoring purposes, as well as proof-of-concept for the combinatorial biosynthesis of these natural occurring alkaloids for the production of novel, biologically active compounds.

  9. Nucleotide diversity and linkage disequilibrium in five Lolium perenne genes with putative role in shoot branching

    DEFF Research Database (Denmark)

    Brazauskas, Gintaras; Pašakinskienė, Izolda; Asp, Torben

    2010-01-01

    .59, respectively. No evidence of selection was found for LpIAA1, LpRUB1, LpSHOOT1 and LpTB1 genes, however, a significant and negative Tajima's D as well as high average LD in LpBRI1 suggest a role of this gene in shaping ryegrass shoot morphology during development of elite germplasm.......Knowledge on nucleotide diversity and linkage disequilibrium (LD) patterns is prerequisite for association analyses. However, little is known about the nucleotide diversity in the evolutionary important ryegrass shoot morphology genes. Five candidate genes, LpIAA1, LpRUB1, LpBRI1, LpSHOOT1 and Lp...... detected. On average, one SNP was present every 94 bp between two randomly selected sequences for the five genes. As expected, the number of synonymous substitutions was much higher compared to the number of non-synonymous mutations for most of the genes. However, six non-synonymous and only two synonymous...

  10. Cloning and characterization of two putative seven-transmembrane receptor genes from cotton

    Institute of Scientific and Technical Information of China (English)

    Peng Gao; Piming Zhao; Juan Wang; Haiyun Wang; Guiling Wang; Guixian Xia

    2008-01-01

    Using rapid amplification of cDNA ends (RACE)-PCR,two full-length cDNAs encoding putative seven-transmembrane receptors (designated Gh7TMpR1 and Gh7TMpR2) were cloned from cotton plants.Southern blot and an ApaLl restriction site polymorphism analyses revealed that GhTTMpR1 was derived from the ancestral A diploid genome,while Gh7TMpR2 was from the D subgenome.Northern blot hybridization indicated that both Gh7TMpR1 and Gh7TMpR2 were expressed preferentially in the elongation phase of fiber development.Majority of the Gh7TMpR1 proteins were located within the membrane structure and displayed a punctuate pattern of distribution.Overexpression of Gh7TMpR1 in fission yeast disrupted the polar growth and caused the formation of rounded cells.These results suggest that GhT7MpRI may play a critical role in cotton fiber development,perhaps as a signaling receptor that is involved in controlling fiber elongation.

  11. Identification of a Putative Quantitative Trait Gene for Resistance to Obesity in Mice Using Transcriptome Analysis and Causal Inference Tests.

    Science.gov (United States)

    Ishikawa, Akira

    2017-01-01

    It is still challenging to identify causal genes governing obesity. Pbwg1.5, a quantitative trait locus (QTL) for resistance to obesity, was previously discovered from wild Mus musculus castaneus mice and was fine-mapped to a 2.1-Mb genomic region of mouse chromosome 2, where no known gene with an effect on white adipose tissue (WAT) has been reported. The aim of this study was to identify a strong candidate gene for Pbwg1.5 by an integration approach of transcriptome analysis (RNA-sequencing followed by real-time PCR analysis) and the causal inference test (CIT), a statistical method to infer causal relationships between diplotypes, gene expression and trait values. Body weight, body composition and biochemical traits were measured in F2 mice obtained from an intercross between the C57BL/6JJcl strain and a congenic strain carrying Pbwg1.5 on the C57BL/6JJcl background. The F2 mice showed significant diplotype differences in 12 traits including body weight, WAT weight and serum cholesterol/triglyceride levels. The transcriptome analysis revealed that Ly75, Pla2r1, Fap and Gca genes were differentially expressed in the liver and that Fap, Ifih1 and Grb14 were differentially expressed in WAT. However, CITs indicated statistical evidence that only the liver Ly75 gene mediated between genotype and WAT. Ly75 expression was negatively associated with WAT weight. The results suggested that Ly75 is a putative quantitative trait gene for the obesity-resistant Pbwg1.5 QTL discovered from the wild M. m. castaneus mouse. The finding provides a novel insight into a better understanding of the genetic basis for prevention of obesity.

  12. Cloning and Expression Pattern of a Gene Encoding a Putative Plastidic ATP/ADP Transporter from Helianthus tuberosus L.

    Institute of Scientific and Technical Information of China (English)

    Kun MENG; Tuan-Jie CHANG; Xiang LIU; Song-Biao CHEN; Yong-Qin WANG; Ai-Jun SUN; Hong-Lin XU; Xiao-Li WEI; Zhen ZHU

    2005-01-01

    Herein, we report the cloning and molecular characterization of a full cDNA encoding a putative plastidic ATP/ADP transporter, designated HtAATP, for Helianthus tuberosus L. The ATP/ADP translocator protein was isolated from the tuber-cDNA library of H. tuberosus for the first time. The predicted HtAATP protein was judged as a plastidic ATP/ADP translocator protein from its high homology at the amino acid sequence level to the two Arabidopsis thaliana plastidic ATP/ADP translocator proteins AATP1 and AATP2 (84.8% and 79.9% identity, respectively). Amino acid sequence analysis of the primary structure of HtAATP revealed that it belonged to the plastidic ATP/ADP transporter family. Hydropathy prediction indicated that HtAATP gene product is a highly hydrophobic membrane protein that contains 10 transmembrane domains to form a spanning topology. Southern blotting analysis showed that the HtAATP gene is a single-copy gene in the H. tuberosus genome. Tissue distribution analysis showed that the HtAATP gene is prominently expressed in sink tissues. A stable expression pattern in tubers at different developmental stages implies an active involvement of HtAATP during carbohydrate formation.

  13. Cladosporium cladosporioides LPSC 1088 produces the 1,8-dihydroxynaphthalene-melanin-like compound and carries a putative pks gene.

    Science.gov (United States)

    Llorente, Carla; Bárcena, Alejandra; Vera Bahima, José; Saparrat, Mario C N; Arambarri, Angélica M; Rozas, M Fernanda; Mirífico, María V; Balatti, Pedro A

    2012-12-01

    Cladosporium cladosporioides is a dematiaceous fungus with coloured mycelia and conidia due to the presence of dark pigments. The purpose of this study was to characterize the dark pigments synthetized by Cladosporium sp. LPSC no. 1088 and also to identify the putative polyketide synthase (pks) gene that might be involved in the pigment biosynthesis. Morphological as well as molecular features like the ITS sequence confirmed that LPSC 1088 is Cladosporium cladosporioides. UV-visible, Fourier Transform Infrared (FTIR) and Electron Spin Resonance (ESR) spectroscopy analysis as well as melanin inhibitors suggest that the main dark pigment of the isolate was 1,8 dihydroxynaphthalene (DHN)-melanin-type compound. Two commercial fungicides, Difenoconazole and Chlorothalonil, inhibited fungal growth as well as increased pigmentation of the colonies suggesting that melanin might protect the fungus against chemical stress. The pigment is most probably synthetized by means of a pentaketide pathway since the sequence of a 651 bp fragment, coding for a putative polyketide synthase, is highly homologous to pks sequences from other fungi.

  14. Identification and characterization of three putative genes for 1-aminocyclopropane-1-carboxylate synthase from etiolated mung bean hypocotyl segments.

    Science.gov (United States)

    Botella, J R; Schlagnhaufer, C D; Arteca, R N; Phillips, A T

    1992-02-01

    The polymerase chain reaction (PCR) was used to produce 3 putative clones for ACC synthase from etiolated mung bean (Vigna radiata Rwilcz cv. Berken) hypocotyls. This was accomplished by utilizing genomic DNA from mung bean and degenerate primers made from information derived from highly conserved regions of ACC synthase from different plant tissues. The total length of pMAC-1, pMAC-2 and pMAC-3 are 308, 321, and 326 bp, respectively, all of which code for 68 amino acids. The introns for pMAC-1, pMAC-2 and pMAC-3 are 92, 105, and 110 bp, respectively. The degrees of homology at the DNA level for each of these clones is ca. 80% in their coding region and ca. 50% in their respective introns. This is the first report providing evidence that there are at least 3 genes for ACC synthase in etiolated mung bean.

  15. Occurrence of putative virulence genes on Arcobacter butzleri isolated from three different environmental sites throughout the dairy chain.

    Science.gov (United States)

    Piva, S; Gariano, G R; Bonilauri, P; Giacometti, F; Decastelli, L; Florio, D; Massella, E; Serraino, A

    2017-04-01

    This comparative study investigated the occurrence of cadF, cj1349, ciaB, pldA, tlyA, hecA, hecB, mviN, irgA and IroE genes in 212 Arcobacter butzleri isolated from three different environmental sites linked to the dairy chain (farms, industrial and artisanal dairy plants) located in three Italian regions (Lombardy, Emilia-Romagna and Calabria). According to the presence of these genes, different pathotypes (P-types) were determined. The main genes detected were ciaB, mviN, tlyA, cj1349, pldA and cadF, while the least common genes were iroE, hecA, hecB and irgA. TlyA, irgA, hecA, hecB and iroE, which were significantly more frequent in isolates recovered in industrial dairy plants. Twelve P-types were detected. The occurrence of the most frequently detected P-types (P-types 1, 2, 3 and 5) differed significantly (P genes and virulence genotype variability depending on the environmental site and geographical origin of the isolates. The present study provides insights into the similar distribution of putative virulence genes in a dairy chain and other sources' isolates and also into a geographical distribution of some P-types. We have shown that industrial dairy plants may represent an environmental site favouring a selection of the isolates with a higher pathogenetic pattern. © 2017 The Society for Applied Microbiology.

  16. Transcriptomics Analysis Reveals Putative Genes Involved in Biofilm Formation and Biofilm-associated Drug Resistance of Enterococcus faecalis.

    Science.gov (United States)

    Seneviratne, Chaminda J; Suriyanarayanan, Tanujaa; Swarup, Sanjay; Chia, Kuan Hui Burton; Nagarajan, Niranjan; Zhang, Chengfei

    2017-06-01

    Enterococcus faecalis is a gram-positive bacterium associated with endodontic infections and is capable of forming biofilms that can confer drug resistance to the bacterium, resulting in treatment failure. Current knowledge on E. faecalis drug resistance is of a limited and conflicting nature. The present study examined the genetic basis of E. faecalis biofilm formation and drug resistance using a RNA sequencing (RNA-Seq)-based transcriptome approach. Eighteen clinical isolates of E. faecalis were screened for their biofilm formation abilities using the crystal violet assay, colony counting, and confocal imaging. Selected isolates were then evaluated for antibiotic susceptibility in planktonic and biofilm growth modes followed by RNA-Seq analysis of E. faecalis planktonic, biofilm, and vancomycin-treated biofilm samples and Kyoto Encyclopedia of Genes and Genomes mapping in order to identify genes associated with biofilm formation and drug resistance of E. faecalis. All 18 clinical isolates retained biofilm formation ability and were classified as strong, weak, or laboratory American Type Culture Collection strainlike biofilm formers. Interestingly, both the strong and weak biofilm-forming isolates were uniformly resistant to ampicillin and vancomycin at the treated concentrations (256-4096 μg/mL). RNA-Seq analysis of these isolates identified a total of 163 and 101 differentially regulated genes in planktonic versus biofilm and vancomycin-treated biofilm versus biofilm comparisons, respectively, with significant differences in arsenic resistance operon genes arsR and arsD, sporulation regulatory gene paiA, ABC drug transporter classes, and penicillin-binding proteins. The present transcriptomic study revealed putative genes associated with E. faecalis biofilm formation and drug resistance, which will provide a foundation for improved therapeutic strategies against E. faecalis infections in the future. Copyright © 2017 American Association of Endodontists

  17. The Drosophila gene brainiac encodes a glycosyltransferase putatively involved in glycosphingolipid synthesis

    DEFF Research Database (Denmark)

    Schwientek, Tilo; Keck, Birgit; Levery, Steven B

    2002-01-01

    The Drosophila genes fringe and brainiac exhibit sequence similarities to glycosyltransferases. Drosophila and mammalian fringe homologs encode UDP-N-acetylglucosamine:fucose-O-Ser beta1,3-N-acetylglucosaminyltransferases that modulate the function of Notch family receptors. The biological function...

  18. Cloning and sequence analysis of putative type II fatty acid synthase genes from Arachis hypogaea L.

    Indian Academy of Sciences (India)

    Meng-Jun Li; Ai-Qin Li; Han Xia; Chuan-Zhi Zhao; Chang-Sheng Li; Shu-Bo Wan; Yu-Ping Bi; Xing-Jun Wang

    2009-06-01

    The cultivated peanut is a valuable source of dietary oil and ranks fifth among the world oil crops. Plant fatty acid biosynthesis is catalysed by type II fatty acid synthase (FAS) in plastids and mitochondria. By constructing a full-length cDNA library derived from immature peanut seeds and homology-based cloning, candidate genes of acyl carrier protein (ACP), malonyl-CoA:ACP transacylase, -ketoacyl-ACP synthase (I, II, III), -ketoacyl-ACP reductase, -hydroxyacyl-ACP dehydrase and enoyl-ACP reductase were isolated. Sequence alignments revealed that primary structures of type II FAS enzymes were highly conserved in higher plants and the catalytic residues were strictly conserved in Escherichia coli and higher plants. Homologue numbers of each type II FAS gene expressing in developing peanut seeds varied from 1 in KASII, KASIII and HD to 5 in ENR. The number of single-nucleotide polymorphisms (SNPs) was quite different in each gene. Peanut type II FAS genes were predicted to target plastids except ACP2 and ACP3. The results suggested that peanut may contain two type II FAS systems in plastids and mitochondria. The type II FAS enzymes in higher plants may have similar functions as those in E. coli.

  19. Enrichment of putative PAX8 target genes at serous epithelial ovarian cancer susceptibility loci

    DEFF Research Database (Denmark)

    Kar, Siddhartha P; Adler, Emily; Tyrer, Jonathan

    2017-01-01

    BACKGROUND: Genome-wide association studies (GWAS) have identified 18 loci associated with serous ovarian cancer (SOC) susceptibility but the biological mechanisms driving these findings remain poorly characterised. Germline cancer risk loci may be enriched for target genes of transcription facto...

  20. Localization of candidate allergen genes on the apple (Malus domestica) genome and their putative allergenicity

    NARCIS (Netherlands)

    Gao Zhongshan,

    2005-01-01

    Apple is generally considered as a healthy food, but 2-3% European people can not eat this fruit because it provokes allergy reaction. Four classes of apple allergen genes have been identified, they are Mal d 1, Mal d 2, Mal d 3 and Mal d 4 . This thesis focuses on the genomic characterization of th

  1. Cloning and sequence analysis of putative type II fatty acid synthase genes from Arachis hypogaea L.

    Science.gov (United States)

    Li, Meng-Jun; Li, Ai-Qin; Xia, Han; Zhao, Chuan-Zhi; Li, Chang-Sheng; Wan, Shu-Bo; Bi, Yu-Ping; Wang, Xing-Jun

    2009-06-01

    The cultivated peanut is a valuable source of dietary oil and ranks fifth among the world oil crops. Plant fatty acid biosynthesis is catalysed by type II fatty acid synthase (FAS) in plastids and mitochondria. By constructing a full-length cDNA library derived from immature peanut seeds and homology-based cloning, candidate genes of acyl carrier protein (ACP), malonyl-CoA:ACP transacylase, beta-ketoacyl-ACP synthase (I, II, III), beta-ketoacyl-ACP reductase, beta-hydroxyacyl-ACP dehydrase and enoyl-ACP reductase were isolated. Sequence alignments revealed that primary structures of type II FAS enzymes were highly conserved in higher plants and the catalytic residues were strictly conserved in Escherichia coli and higher plants. Homologue numbers of each type II FAS gene expressing in developing peanut seeds varied from 1 in KASII, KASIII and HD to 5 in ENR. The number of single-nucleotide polymorphisms (SNPs) was quite different in each gene. Peanut type II FAS genes were predicted to target plastids except ACP2 and ACP3. The results suggested that peanut may contain two type II FAS systems in plastids and mitochondria. The type II FAS enzymes in higher plants may have similar functions as those in E. coli.

  2. Identification of the putative specific pathogenic genes of Porphyromonas gingivalis with type II fimbriae.

    Science.gov (United States)

    Gao, Li; Xu, Yi; Meng, Shu; Wu, Yafei; Huang, Haiyun; Su, Ruiying; Zhao, Lei

    2012-06-01

    Porphyromonas gingivalis, the key etiologic agent of periodontitis, can be classified into six types (I to V and Ib) based on the fimA genes that encode FimA (a subunit of fimbriae). Accumulated evidence indicates that P. gingivalis expressing Type II fimbriae (Pg-II) is the most frequent isolate from severe periodontitis cases and is more virulent than other types of P. gingivalis. However, during the Pg-II infection process, which specific virulence factors play the key role is still unclear. In this study, we examined the capabilities of three Pg-II strains to invade and modulate the inflammatory cytokine expression of human gingival epithelial cells (GECs) compared to two Pg-I strains. P. gingivalis oligo microarrays were used to compare gene expression profiles of Pg-II strains that invade GECs with Pg-I strains. The differential gene expression of Pg-II was confirmed by quantitative reverse transcription-polymerase chain reaction. Our results showed that all of the Pg-II strains could induce interleukin (IL)-1β and IL-6 secretion significantly when compared to Pg-I strains. Thirty-seven genes that were specifically expressed during the pathogenic process of Pg-II were identified by a microarray assay. These findings provide a new insight at the molecular level to explain the specific pathogenic mechanism of Pg-II strains.

  3. A PUTATIVE BETA-GLUCANASE PSEUDOGENE BEHIND THE POTATO GBSS GENE

    NARCIS (Netherlands)

    VANDERLEIJ, FR; ABELN, ECA; HESSELINGMEINDERS, A; FEENSTRA, WJ

    1993-01-01

    We identified an open reading frame (ORF) which is located closely behind the gene encoding granule-bound starch synthase (GBSS) of potato (Solanum tuberosum L.). The ORF ends with a perfect 43 bp direct repeat, which carries the stop triplet precisely at the beginning of the second repeat. The dedu

  4. Influence of putative exopolysaccharide genes on Pseudomonas putida KT2440 biofilm stability

    DEFF Research Database (Denmark)

    Nilsson, Martin; Chiang, Wen-Chi; Fazli, Mustafa;

    2011-01-01

    . putida KT2440 biofilm. The gene clusters alg and bcs, which code for proteins mediating alginate and cellulose biosynthesis, were found to play minor roles in P. putida KT2440 biofilm formation and stability under the conditions tested. A P. putida KT2440 derivative devoid of any identifiable...

  5. Analysis of the transcriptome of Panax notoginseng root uncovers putative triterpene saponin-biosynthetic genes and genetic markers

    Directory of Open Access Journals (Sweden)

    Luo Hongmei

    2011-12-01

    Full Text Available Abstract Background Panax notoginseng (Burk F.H. Chen is important medicinal plant of the Araliacease family. Triterpene saponins are the bioactive constituents in P. notoginseng. However, available genomic information regarding this plant is limited. Moreover, details of triterpene saponin biosynthesis in the Panax species are largely unknown. Results Using the 454 pyrosequencing technology, a one-quarter GS FLX titanium run resulted in 188,185 reads with an average length of 410 bases for P. notoginseng root. These reads were processed and assembled by 454 GS De Novo Assembler software into 30,852 unique sequences. A total of 70.2% of unique sequences were annotated by Basic Local Alignment Search Tool (BLAST similarity searches against public sequence databases. The Kyoto Encyclopedia of Genes and Genomes (KEGG assignment discovered 41 unique sequences representing 11 genes involved in triterpene saponin backbone biosynthesis in the 454-EST dataset. In particular, the transcript encoding dammarenediol synthase (DS, which is the first committed enzyme in the biosynthetic pathway of major triterpene saponins, is highly expressed in the root of four-year-old P. notoginseng. It is worth emphasizing that the candidate cytochrome P450 (Pn02132 and Pn00158 and UDP-glycosyltransferase (Pn00082 gene most likely to be involved in hydroxylation or glycosylation of aglycones for triterpene saponin biosynthesis were discovered from 174 cytochrome P450s and 242 glycosyltransferases by phylogenetic analysis, respectively. Putative transcription factors were detected in 906 unique sequences, including Myb, homeobox, WRKY, basic helix-loop-helix (bHLH, and other family proteins. Additionally, a total of 2,772 simple sequence repeat (SSR were identified from 2,361 unique sequences, of which, di-nucleotide motifs were the most abundant motif. Conclusion This study is the first to present a large-scale EST dataset for P. notoginseng root acquired by next

  6. Putative compensatory mutations in the rpoC gene of rifampin-resistant Mycobacterium tuberculosis are associated with ongoing transmission.

    Science.gov (United States)

    de Vos, M; Müller, B; Borrell, S; Black, P A; van Helden, P D; Warren, R M; Gagneux, S; Victor, T C

    2013-02-01

    Rifampin resistance in clinical isolates of Mycobacterium tuberculosis arises primarily through the selection of bacterial variants harboring mutations in the 81-bp rifampin resistance-determining region of the rpoB gene. While these mutations were shown to infer a fitness cost in the absence of antibiotic pressure, compensatory mutations in rpoA and rpoC were identified which restore the fitness of rifampin-resistant bacteria carrying mutations in rpoB. To investigate the epidemiological relevance of these compensatory mutations, we analyzed 286 drug-resistant and 54 drug-susceptible clinical M. tuberculosis isolates from the Western Cape, South Africa, a high-incidence setting of multidrug-resistant tuberculosis. Sequencing of a portion of the RpoA-RpoC interaction region of the rpoC gene revealed that 23.5% of all rifampin-resistant isolates tested carried a nonsynonymous mutation in this region. These putative compensatory mutations in rpoC were associated with transmission, as 30.8% of all rifampin-resistant isolates with an IS6110 restriction fragment length polymorphism (RFLP) pattern belonging to a recognized RFLP cluster harbored putative rpoC mutations. Such mutations were present in only 9.4% of rifampin-resistant isolates with unique RFLP patterns (P mutations were associated with specific strain genotypes and the rpoB S531L rifampin resistance mutation. Among isolates harboring this rpoB mutation, 44.1% also harbored rpoC mutations, while only 4.1% of the isolates with other rpoB mutations exhibited mutations in rpoC (P mutations in the transmission of multidrug-resistant tuberculosis and illustrates how epistatic interactions between drug resistance-conferring mutations, compensatory mutations, and different strain genetic backgrounds might influence compensatory evolution in drug-resistant M. tuberculosis.

  7. Mutational studies of putative biosynthetic genes for the cyanobacterial sunscreen scytonemin in Nostoc punctiforme ATCC 29133

    Directory of Open Access Journals (Sweden)

    Daniela eFerreira

    2016-05-01

    Full Text Available The heterocyclic indole-alkaloid scytonemin is a sunscreen found exclusively among cyanobacteria. An 18-gene cluster is responsible for scytonemin production in Nostoc punctiforme ATCC 29133. The upstream genes scyABCDEF in the cluster are proposed to be responsible for scytonemin biosynthesis from aromatic amino acid substrates. In vitro studies of ScyA, ScyB and ScyC proved that these enzymes indeed catalyze initial pathway reactions. Here we characterize the role of ScyD, ScyE and ScyF, which were logically predicted to be responsible for late biosynthetic steps, in the biological context of N. punctiforme. In-frame deletion mutants of each were constructed (∆scyD, ∆scyE and ∆scyF and their phenotypes studied. Expectedly, ∆scyE presents a scytoneminless phenotype, but no accumulation of the predicted intermediaries. Surprisingly, ∆scyD retains scytonemin production, implying that it is not required for biosynthesis. Indeed, scyD presents an interesting evolutionary paradox: it likely originated in a duplication event from scyE, and unlike other genes in the operon, it has not been subjected to purifying selection. This would suggest that it is a pseudogene, and yet scyD is highly conserved in the scytonemin operon of cyanobacteria. ∆scyF also retains scytonemin production, albeit exhibiting a reduction of the production yield compared with the wild-type. This indicates that ScyF is not essential but may play an adjuvant role for scytonemin synthesis. Altogether, our findings suggest that these downstream genes are not responsible, as expected, for the late steps of scytonemin synthesis and we must look for those functions elsewhere. These findings are particularly important for biotechnological production of this sunscreen through heterologous expression of its genes in more tractable organisms.

  8. The Emerging Role of TPR-Domain Immunophilins in the Mechanism of Action of Steroid Receptors

    Directory of Open Access Journals (Sweden)

    G. I. Mazaira

    2014-11-01

    Full Text Available In the absence of ligand, some members of nuclear receptor family such as corticosteroid receptors are primarily located in the cytoplasm, and they rapidly accumulate in the nucleus upon ligand-binding. Other members of the family such as the estrogen receptor are mostly nuclear. Regardless of their primary location, these oligomeric proteins undergo a dynamic nuclear-cytoplasmic shuttling, and their transport through the cytoplasmic compartment has always been assumed to occur in a stochastic manner by simple diffusion. Although heuristic, this oversimplified model has never been demonstrated. Moreover, it has always been assumed that the first step related to receptor activation is the dissociation of the Hsp90-based heterocomplex, a process referred to as `transformation.' Nonetheless, recent experimental evidence indicates that the chaperone machinery is required for the retrotransport of the receptor throughout the cytoplasm and facilitates its active passage through the nuclear pore. Therefore, transformation is actually a nuclear event. A group of Hsp90-binding cochaperones belonging to the immunophilin family plays a cardinal role not only in the mechanism for receptor movement, but also in nuclear events leading to interactions with nuclear sites of action and the regulation of transcriptional activity. In this article we analyze the importance of molecular chaperones and TPR-domain immunophilins in the molecular mechanism of action of steroid receptors.

  9. Epigenetic silencing of MAL, a putative tumor suppressor gene, can contribute to human epithelium cell carcinoma

    Directory of Open Access Journals (Sweden)

    Zhang Jun

    2010-11-01

    Full Text Available Abstract Background To identify new and useful candidate biomarkers in head and neck squamous cell carcinoma (HNSCC, we performed a genome-wide survey and found that Myelin and lymphocyte-associated protein (MAL was a gene that was markedly down-regulated in HNSCC. Hence, we investigated the mechanism of MAL silencing and the effects of MAL on the proliferation, invasion, and apoptotic potential in HNSCC. Results MAL was significantly down-regulated in 91.7% of HNSCC specimens at the mRNA level as compared with adjacent normal tissues (P = 0.0004. Moreover, the relative transcript levels of the MAL gene were remarkably decreased by five-fold in nine HNSCC cell lines as compared with normal head and neck epithelium cells. MAL gene expression was restored in 44%, 67%, and 89% in HNSCC cell lines treated with TSA, 5-Aza-dC, and TSA plus 5-Aza-dC, respectively. Furthermore, bisulfate-treated DNA sequencing demonstrated that the two CpG islands (that is, M1 and M2 located in MAL promoter region were completely methylated in the HNSCC cell lines (CpG methylated ratio was more than 90%, and only one CpG island (that is, M1 was partially methylated in HNSCC tissues (CpG methylated ratio between 20% and 90%. A significant reduction in cell proliferation and a change in the cell cycle profile were also observed in MAL transfectants. Matrigel assay demonstrated that the invasiveness of HNSCC cells significantly decreased. A significant increase in the population of apoptotic cells was observed in MAL transfected cells. The exogenous expression of the MAL gene suppressed malignant phenotypes, while the cell death induced by MAL gene transfer was a result of apoptosis as demonstrated by the induction of cleavage of the poly (that is, ADP-ribose polymerase. Additionally, tumor growth was suppressed in cells expressing MAL as compared with cells not expressing MAL. Conclusion Our data suggest that the epigenetic inactivation of MAL, as a candidate tumor

  10. Molecular chaperone activity and biological regulatory actions of the TPR-domain immunophilins FKBP51 and FKBP52.

    Science.gov (United States)

    Erlejman, Alejandra G; Lagadari, Mariana; Harris, Diondra C; Cox, Marc B; Galigniana, Mario D

    2014-05-01

    Immunophilins comprise a family of intracellular proteins with peptidyl-prolyl-(cis/trans)-isomerase activity. These foldases are abundant, ubiquitous, and able to bind immunosuppressant drugs, from which the term immunophilin derives. Family members are found in abundance in virtually all organisms and subcellular compartments, and their amino acid sequences are conserved phylogenetically. Immunophilins possess the ability to function as molecular chaperones favoring the proper folding and biological regulation of their biological actions. Their ability to interact via their TPR domains with the 90-kDa heat-shock protein, and through this chaperone, with several signalling cascade factors is of particular importance. Among the family members, the highly homologous proteins FKBP51 and FKBP52 were first characterized due to their ability to interact with steroid hormone receptors. Since then, much progress has been made in understanding the mechanisms by which they regulate receptor signaling and the resulting roles they play not only in endocrine processes, but also in cell architecture, neurodifferentiation, and tumor progression. In this article we review the most relevant features of these two immunophilins and their potential as pharmacologic targets.

  11. Genetic Characterization of Plasmodium Putative Pantothenate Kinase Genes Reveals Their Essential Role in Malaria Parasite Transmission to the Mosquito.

    Science.gov (United States)

    Hart, Robert J; Cornillot, Emmanuel; Abraham, Amanah; Molina, Emily; Nation, Catherine S; Ben Mamoun, Choukri; Aly, Ahmed S I

    2016-09-20

    The metabolic machinery for the biosynthesis of Coenzyme A (CoA) from exogenous pantothenic acid (Vitamin B5) has long been considered as an excellent target for the development of selective antimicrobials. Earlier studies in the human malaria parasite Plasmodium falciparum have shown that pantothenate analogs interfere with pantothenate phosphorylation and block asexual blood stage development. Although two eukaryotic-type putative pantothenate kinase genes (PanK1 and PanK2) have been identified in all malaria parasite species, their role in the development of Plasmodium life cycle stages remains unknown. Here we report on the genetic characterization of PanK1 and PanK2 in P. yoelii. We show that P. yoelii parasites lacking either PanK1 or PanK2 undergo normal asexual stages development and sexual stages differentiation, however they are severely deficient in ookinete, oocyst and sporozoite formation inside the mosquito vector. Quantitative transcriptional analyses in wild-type and knockout parasites demonstrate an important role for these genes in the regulation of expression of other CoA biosynthesis genes. Together, our data provide the first genetic evidence for the importance of the early steps of pantothenate utilization in the regulation of CoA biosynthesis and malaria parasite transmission to Anopheles mosquitoes.

  12. Genetic Characterization of Plasmodium Putative Pantothenate Kinase Genes Reveals Their Essential Role in Malaria Parasite Transmission to the Mosquito

    Science.gov (United States)

    Hart, Robert J.; Cornillot, Emmanuel; Abraham, Amanah; Molina, Emily; Nation, Catherine S.; Ben Mamoun, Choukri; Aly, Ahmed S. I.

    2016-01-01

    The metabolic machinery for the biosynthesis of Coenzyme A (CoA) from exogenous pantothenic acid (Vitamin B5) has long been considered as an excellent target for the development of selective antimicrobials. Earlier studies in the human malaria parasite Plasmodium falciparum have shown that pantothenate analogs interfere with pantothenate phosphorylation and block asexual blood stage development. Although two eukaryotic-type putative pantothenate kinase genes (PanK1 and PanK2) have been identified in all malaria parasite species, their role in the development of Plasmodium life cycle stages remains unknown. Here we report on the genetic characterization of PanK1 and PanK2 in P. yoelii. We show that P. yoelii parasites lacking either PanK1 or PanK2 undergo normal asexual stages development and sexual stages differentiation, however they are severely deficient in ookinete, oocyst and sporozoite formation inside the mosquito vector. Quantitative transcriptional analyses in wild-type and knockout parasites demonstrate an important role for these genes in the regulation of expression of other CoA biosynthesis genes. Together, our data provide the first genetic evidence for the importance of the early steps of pantothenate utilization in the regulation of CoA biosynthesis and malaria parasite transmission to Anopheles mosquitoes. PMID:27644319

  13. Grouping and characterization of putative glycosyltransferase genes from Panax ginseng Meyer.

    Science.gov (United States)

    Khorolragchaa, Altanzul; Kim, Yu-Jin; Rahimi, Shadi; Sukweenadhi, Johan; Jang, Moon-Gi; Yang, Deok-Chun

    2014-02-15

    Glycosyltransferases are members of the multigene family of plants that can transfer single or multiple activated sugars to a range of plant molecules, resulting in the glycosylation of plant compounds. Although the activities of many glycosyltransferases and their products have been recognized for a long time, only in recent years were some glycosyltransferase genes identified and few have been functionally characterized in detail. Korean ginseng (Panax ginseng Meyer), belonging to Araliaceae, has been well known as a popular mysterious medicinal herb in East Asia for over 2,000 years. A total of 704 glycosyltransferase unique sequences have been found from a ginseng expressed sequence tag (EST) library, and these sequences encode enzymes responsible for the secondary metabolite biosynthesis. Finally, twelve UDP glycosyltransferases (UGTs) were selected as the candidates most likely to be involved in triterpenoid synthesis. In this study, we classified the candidate P. ginseng UGTs (PgUGTs) into proper families and groups, which resulted in eight UGT families and six UGT groups. We also investigated those gene candidates encoding for glycosyltransferases by analysis of gene expression in methyl jasmonate (MeJA)-treated ginseng adventitious roots and different tissues from four-year-old ginseng using quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). For organ-specific expression, most of PgUGT transcription levels were higher in leaves and roots compared with flower buds and stems. The transcription of PgUGTs in adventitious roots treated with MeJA increased as compared with the control. PgUGT1 and PgUGT2, which belong to the UGT71 family genes expressed in MeJA-treated adventitious roots, were especially sensitive, showing 33.32 and 38.88-fold expression increases upon 24h post-treatments, respectively. © 2013 Elsevier B.V. All rights reserved.

  14. Cloning of a Putative Pectate Lyase Gene Expressed in the Subventral Esophageal Glands of Heterodera glycines.

    Science.gov (United States)

    De Boer, J M; Davis, E L; Hussey, R S; Popeijus, H; Smant, G; Baum, T J

    2002-03-01

    We report the cloning of a Heterodera glycines cDNA that has 72% identity at the amino acid level to a pectate lyase from Globodera rostochiensis. In situ hybridizations showed that the corresponding gene (Hg-pel-1) is expressed in the subventral esophageal gland cells of second-stage juveniles. The deduced amino acid sequence of the H. glycines cDNA shows homology to class III pectate lyases of bacterial and fungal origin.

  15. Putative pyrophosphate phosphofructose 1-kinase genes identified in sugar cane may be getting energy from pyrophosphate.

    Science.gov (United States)

    Suzuki, J; Mutton, M A; Ferro, M I T; Lemos, M V F; Pizauro, J M; Mutton, M J R; Di Mauro, S M Z

    2003-12-30

    Pyrophosphate-dependent phosphofructokinase (PPi-PFK) has been detected in several types of plant cells, but the gene has not been reported in sugar cane. Using Citrus paradisi PPi-PFK gene (AF095520 and AF095521) sequences to search the sugar cane EST database, we have identified both the alpha and beta subunits of this enzyme. The deduced amino acid sequences showed 76 and 80% similarity with the corresponding alpha and beta subunits of C. paradisi. A high degree of similarity was also observed among the PFK b subunits when the alignment of the sugar cane sequences was compared to those of Ricinus communis and Solanum tuberosum. It appears that alpha and beta are two distinct subunits; they were found at different concentrations in several sugar cane tissues. It remains to be determined if the different gene expression levels have some physiological importance and how they affect sucrose synthesis, export, and storage in vacuoles. A comparison between the amino acid sequences of b PFKs from a variety of organisms allowed us to identify the two critical Asp residues typical of this enzyme's activity site and the other binding sites; these residues are tightly conserved in all members of this protein family. Apparently, there are catalytic residues on the b subunit of the pyrophosphate-dependent enzyme.

  16. The Novel Neuronal Ceroid Lipofuscinosis Gene MFSD8 Encodes a Putative Lysosomal Transporter

    Science.gov (United States)

    Siintola, Eija ; Topcu, Meral ; Aula, Nina ; Lohi, Hannes ; Minassian, Berge A. ; Paterson, Andrew D. ; Liu, Xiao-Qing ; Wilson, Callum ; Lahtinen, Ulla ; Anttonen, Anna-Kaisa ; Lehesjoki, Anna-Elina 

    2007-01-01

    The late-infantile–onset forms are the most genetically heterogeneous group among the autosomal recessively inherited neurodegenerative disorders, the neuronal ceroid lipofuscinoses (NCLs). The Turkish variant was initially considered to be a distinct genetic entity, with clinical presentation similar to that of other forms of late-infantile–onset NCL (LINCL), including age at onset from 2 to 7 years, epileptic seizures, psychomotor deterioration, myoclonus, loss of vision, and premature death. However, Turkish variant LINCL was recently found to be genetically heterogeneous, because mutations in two genes, CLN6 and CLN8, were identified to underlie the disease phenotype in a subset of patients. After a genomewide scan with single-nucleotide–polymorphism markers and homozygosity mapping in nine Turkish families and one Indian family, not linked to any of the known NCL loci, we mapped a novel variant LINCL locus to chromosome 4q28.1-q28.2 in five families. We identified six different mutations in the MFSD8 gene (previously denoted “MGC33302”), which encodes a novel polytopic 518–amino acid membrane protein that belongs to the major facilitator superfamily of transporter proteins. MFSD8 is expressed ubiquitously, with several alternatively spliced variants. Like the majority of the previously identified NCL proteins, MFSD8 localizes mainly to the lysosomal compartment. However, the function of MFSD8 remains to be elucidated. Analysis of the genome-scan data suggests the existence of at least three more genes in the remaining five families, further corroborating the great genetic heterogeneity of LINCLs. PMID:17564970

  17. Cloning the putative gene of vinyl phenol reductase of Dekkera bruxellensis in Saccharomyces cerevisiae.

    Science.gov (United States)

    Romano, Diego; Valdetara, Federica; Zambelli, Paolo; Galafassi, Silvia; De Vitis, Valerio; Molinari, Francesco; Compagno, Concetta; Foschino, Roberto; Vigentini, Ileana

    2017-05-01

    Vinylphenol reductase of Dekkera bruxellensis, the characteristic enzyme liable for "Brett" sensory modification of wine, has been recently recognized to belong to the short chain dehydrogenases/reductases family. Indeed, a preliminary biochemical characterisation has conferred to the purified protein a dual significance acting as superoxide dismutase and as a NADH-dependent reductase. The present study aimed for providing a certain identification of the enzyme by cloning the VPR gene in S. cerevisiae, a species not producing ethyl phenols. Transformed clones of S. cerevisiae resulted capable of expressing a biologically active form of the heterologous protein, proving its role in the conversion of 4-vinyl guaiacol to 4-ethyl guaiacol. A VPR specific protein activity of 9 ± 0.6 mU/mg was found in crude extracts of S. cerevisiae recombinant strain. This result was confirmed in activity trials carried out with the protein purified from transformant cells of S. cerevisiae by a his-tag purification approach; in particular, VPR-enriched fractions showed a specific activity of 1.83 ± 0.03 U/mg at pH 6.0. Furthermore, in agreement with literature, the purified protein behaves like a SOD, with a calculated specific activity of approximatively 3.41 U/mg. The comparative genetic analysis of the partial VPR gene sequences from 17 different D. bruxellesis strains suggested that the observed polymorphism (2.3%) and the allelic heterozygosity state of the gene do not justify the well described strain-dependent character in producing volatile phenols of this species. Actually, no correlation exists between genotype membership of the analysed strains and their capability to release off-flavours. This work adds valuable knowledge to the study of D. bruxellensis wine spoilage and prepare the ground for interesting future industrial applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Mass spectrometric analysis of putative capa-gene products in Musca domestica and Neobellieria bullata.

    Science.gov (United States)

    Predel, Reinhard; Russell, William K; Tichy, Shane E; Russell, David H; Nachman, Ronald J

    2003-10-01

    Neuropeptides of the capa-gene are typical of the abdominal neurosecretory system of insects. In this study, we investigated these peptides in two widely distributed and large pest flies, namely Musca domestica and Neobellieria bullata. Using a combination of MALDI-TOF and ESI-QTOF mass spectrometry, periviscerokinins and a pyrokinin were analyzed from single perisympathetic organ preparations. The species-specific peptide sequences differ remarkably between the related dipteran species. These differences could make it possible to develop peptide-analogs with group- or species-specific efficacy.

  19. Identification of putative PPAR response elements in and around the murine UCP3 gene

    DEFF Research Database (Denmark)

    Siersbæk, Majken

    in thermogenesis in unerring, the physiological functions of UCP2 and UCP3 are at present not fully understood. Synthetic agonists for the peroxisome proliferator-activated receptors (PPARs) as well as fatty acids have been shown to increase murine UCP2 and UCP3 mRNA expression but response elements and mechanisms...... differentiation. Results from sequencing of chromatin immunoprecipitated (ChIP) material from 3T3-L1 adipocytes revealed three PPAR and retinoid X receptor (RXR) binding sites in and around the murine UCP3 gene. The recruitment of PPAR and RXR to the three potential PPREs was determined by ChIP combined...

  20. Computational identification of putative miRNAs and their target genes in pathogenic amoeba Naegleria fowleri.

    Science.gov (United States)

    Padmashree, Dyavegowda; Swamy, Narayanaswamy Ramachandra

    2015-01-01

    Naegleria fowleri is a parasitic unicellular free living eukaryotic amoeba. The parasite spreads through contaminated water and causes primary amoebic meningoencephalitis (PAM). Therefore, it is of interest to understand its molecular pathogenesis. Hence, we analyzed the parasite genome for miRNAs (microRNAs) that are non-coding, single stranded RNA molecules. We identified 245 miRNAs using computational methods in N. fowleri, of which five miRNAs are conserved. The predicted miRNA targets were analyzed by using miRanda (software) and further studied the functions by subsequently annotating using AmiGo (a gene ontology web tool).

  1. Involvement of a Gene Encoding Putative Acetate Kinase in Magnetosome Synthesis in Magnetospirillum magneticum AMB-1

    Directory of Open Access Journals (Sweden)

    ARIS TRI WAHYUDI

    2006-03-01

    Full Text Available A nonmagnetic mutant of Magnetospirillum magneticum AMB-1, designated NMA40, was constructed by mini-Tn5 transposon mutagenesis to identify genes involved in magnetosome synthesis. Transposon delivery was carried out through conjugation between M. magneticum AMB-1 as a recipient and Escherichia coli S17-1 (λ pir carrying pUTmini-Tn5Km1 as a donor strain. NAM40 did not respond to the magnetic fields and completely lacked of magnetosome in the cell. DNA sequence/gen interrupted by transposon (called flanking DNA was isolated by inverse PCR and cloned into pGEM-T Easy. Alignment of the DNA sequence of the flanking DNA allowed the isolation of an open reading frame (ORF2 within an operon consisting of three genes. The amino acid sequence deduced from ORF2 showed homology with acetate kinase from Sinorhizobium meliloti (50% identity and 67% similarity, which function for acetate metabolism. Further analysis revealed that upstream of ORF2 is ORF1, had homology with phosphotransacetylase of S. meliloti (67% identity, 77% similarity, and ORF3 located downstream of ORF2, had homology with hypothetical protein of Thermotoga maritima (30% identity, 60% similarity. ORF2 was subsequently isolated, cloned, and overexpressed in Escherichia coli BL21 (DE3 pLysS as an ORF2-Histag fusion polypeptide.

  2. Identification of Putative Ortholog Gene Blocks Involved in Gestant and Lactating Mammary Gland Development: A Rodent Cross-Species Microarray Transcriptomics Approach

    Science.gov (United States)

    Rodríguez-Cruz, Maricela; Coral-Vázquez, Ramón M.; Hernández-Stengele, Gabriel; Sánchez, Raúl; Salazar, Emmanuel; Sanchez-Muñoz, Fausto; Encarnación-Guevara, Sergio; Ramírez-Salcedo, Jorge

    2013-01-01

    The mammary gland (MG) undergoes functional and metabolic changes during the transition from pregnancy to lactation, possibly by regulation of conserved genes. The objective was to elucidate orthologous genes, chromosome clusters and putative conserved transcriptional modules during MG development. We analyzed expression of 22,000 transcripts using murine microarrays and RNA samples of MG from virgin, pregnant, and lactating rats by cross-species hybridization. We identified 521 transcripts differentially expressed; upregulated in early (78%) and midpregnancy (89%) and early lactation (64%), but downregulated in mid-lactation (61%). Putative orthologous genes were identified. We mapped the altered genes to orthologous chromosomal locations in human and mouse. Eighteen sets of conserved genes associated with key cellular functions were revealed and conserved transcription factor binding site search entailed possible coregulation among all eight block sets of genes. This study demonstrates that the use of heterologous array hybridization for screening of orthologous gene expression from rat revealed sets of conserved genes arranged in chromosomal order implicated in signaling pathways and functional ontology. Results demonstrate the utilization power of comparative genomics and prove the feasibility of using rodent microarrays to identification of putative coexpressed orthologous genes involved in the control of human mammary gland development. PMID:24288657

  3. Identification of Putative Ortholog Gene Blocks Involved in Gestant and Lactating Mammary Gland Development: A Rodent Cross-Species Microarray Transcriptomics Approach

    Directory of Open Access Journals (Sweden)

    Maricela Rodríguez-Cruz

    2013-01-01

    Full Text Available The mammary gland (MG undergoes functional and metabolic changes during the transition from pregnancy to lactation, possibly by regulation of conserved genes. The objective was to elucidate orthologous genes, chromosome clusters and putative conserved transcriptional modules during MG development. We analyzed expression of 22,000 transcripts using murine microarrays and RNA samples of MG from virgin, pregnant, and lactating rats by cross-species hybridization. We identified 521 transcripts differentially expressed; upregulated in early (78% and midpregnancy (89% and early lactation (64%, but downregulated in mid-lactation (61%. Putative orthologous genes were identified. We mapped the altered genes to orthologous chromosomal locations in human and mouse. Eighteen sets of conserved genes associated with key cellular functions were revealed and conserved transcription factor binding site search entailed possible coregulation among all eight block sets of genes. This study demonstrates that the use of heterologous array hybridization for screening of orthologous gene expression from rat revealed sets of conserved genes arranged in chromosomal order implicated in signaling pathways and functional ontology. Results demonstrate the utilization power of comparative genomics and prove the feasibility of using rodent microarrays to identification of putative coexpressed orthologous genes involved in the control of human mammary gland development.

  4. Isolation and Characterization of a Putative Class E Gene from Taihangia rupestris

    Institute of Scientific and Technical Information of China (English)

    Yong-Qiang Wang; Hui-Yu Tian; Xiao-Qiu Du; Shan-Hua Lü; Wen-Liang Lu; Kang Chong; Zheng Meng

    2007-01-01

    Studies in model plants showed that SEPALLATA (SEP) genes are required for the identification of floral organs and the determination of floral meristems in Arabldopsls. In this paper a SEP homolog, TrSEP3, was Isolated from a SEP3-clade of SEP (prevlous AGL2) subfamily. In sltu hybridization was used to reveal the potential functional specification, and the results showed that TrSEP3 expression was first observed in floral meristems and then confined to the floral primordia of the three inner whorls. In the matured flower, TrSEP3 was strongly expressed in the tips of pistils and weak in stamens and petals. The evolution force analysis shows that TrSEP3 might undergo a relaxed negative selection. These results suggested that TrSEP3 may not only function in determining the identity of floral meristems and the primordia of three inner whorls, but also function in matured reproductive organs.

  5. Characterization of a putative 3-deoxy-D-manno-2-octulosonic acid (Kdo) transferase gene from Arabidopsis thaliana.

    Science.gov (United States)

    Séveno, Martial; Séveno-Carpentier, Emilie; Voxeur, Aline; Menu-Bouaouiche, Laurence; Rihouey, Christophe; Delmas, Frédéric; Chevalier, Christian; Driouich, Azeddine; Lerouge, Patrice

    2010-05-01

    The structures of the pectic polysaccharide rhamnogalacturonan II (RG-II) pectin constituent are remarkably evolutionary conserved in all plant species. At least 12 different glycosyl residues are present in RG-II. Among them is the seldom eight-carbon sugar 3-deoxy-d-manno-octulosonic acid (Kdo) whose biosynthetic pathway has been shown to be conserved between plants and Gram-negative bacteria. Kdo is formed in the cytosol by the condensation of phosphoenol pyruvate with d-arabinose-5-P and then activated by coupling to cytidine monophosphate (CMP) prior to its incorporation in the Golgi apparatus by a Kdo transferase (KDTA) into the nascent polysaccharide RG-II. To gain new insight into RG-II biosynthesis and function, we isolated and characterized null mutants for the unique putative KDTA (AtKDTA) encoded in the Arabidopsis genome. We provide evidence that, in contrast to mutants affecting the RG-II biosynthesis, the extinction of the AtKDTA gene expression does not result in any developmental phenotype in the AtkdtA plants. Furthermore, the structure of RG-II from the null mutants was not altered and contained wild-type amount of Rha-alpha(1-5)Kdo side chain. The cellular localization of AtKDTA was investigated by using laser scanning confocal imaging of the protein fused to green fluorescent protein. In agreement with its cellular prediction, the fusion protein was demonstrated to be targeted to the mitochondria. These data, together with data deduced from sequence analyses of higher plant genomes, suggest that AtKDTA encodes a putative KDTA involved in the synthesis of a mitochondrial not yet identified lipid A-like molecule rather than in the synthesis of the cell wall RG-II.

  6. The choC gene encoding a putative phospholipid methyltransferase is essential for growth and development in Aspergillus nidulans.

    Science.gov (United States)

    Tao, Li; Gao, Na; Chen, Sanfeng; Yu, Jae-Hyuk

    2010-06-01

    Phosphatidylcholines (PCs) are a class of major cell membrane phospholipids that participate in many physiological processes. Three genes, choA, choB and choC, have been proposed to function in the endogenous biosynthesis of PC in Aspergillus nidulans. In this study, we characterize the choC gene encoding a putative highly conserved phospholipid methyltransferase. The previously reported choC3 mutant allele results from a mutation leading to the E177K amino acid substitution. The transcript of choC accumulates at high levels during vegetative growth and early asexual developmental phases. The deletion of choC causes severe impairment of vegetative growth, swelling of hyphal tips and the lack of both asexual and sexual development, suggesting the requirement of ChoC and PC in growth and development. Noticeably, supplementation of the mutant with the penultimate precursor of PC N, N-dimethylaminoethanol leads to full recovery of vegetative growth, but incomplete progression of asexual and sexual development, implying differential roles of PC and its intermediates in fungal growth and development. Importantly, while the choC deletion mutant shows reduced vegetative growth and precocious cell death until day 4, it regains hyphal proliferation and cell viability from day 5, indicating the presence of an alternative route for cellular membrane function in A. nidulans.

  7. Analysis of the expression of putative heat-stress related genes in relation to thermotolerance of cork oak.

    Science.gov (United States)

    Correia, Barbara; Rodriguez, José Luis; Valledor, Luis; Almeida, Tânia; Santos, Conceição; Cañal, Maria Jesús; Pinto, Glória

    2014-03-15

    Cork oak (Quercus suber L.) is a research priority in the Mediterranean area and because of cork oaks' distribution these stands are experiencing daily stress. Based on projections of intensifying climate change and considering the key role of exploring the recovery abilities, cork oak seedlings were subjected to a cumulative temperature increase from 25°C to 55°C and subsequent recovery. CO2 assimilation rate, chlorophyll fluorescence, anthocyanins, proline and lipid peroxidation were used to evaluate plant performance, while the relative abundance of seven genes encoding for proteins of cork oak with a putative role in thermal/stress regulation (POX1, POX2, HSP10.4, HSP17a.22, CHS, MTL and RBC) was analyzed by qPCR (quantitative Polymerase Chain Reaction). A temperature change to 35°C showed abundance alterations in the tested genes; at 45°C, the molecular changes were associated with an antioxidant response, possibly modulated by anthocyanins. At 55°C, HSP17a.22, MTL and proline accumulation were evident. After recovery, physiological balance was restored, whereas POX1, HSP10.4 and MTL abundances were suggested to be involved in increased thermotolerance. The data presented here are expected to pinpoint some pathways changes occurring during such stress and further recovery in this particular Mediterranean species.

  8. Identification of putative PPAR response elements in and around the murine UCP3 gene

    DEFF Research Database (Denmark)

    Siersbæk, Majken

    in thermogenesis in unerring, the physiological functions of UCP2 and UCP3 are at present not fully understood. Synthetic agonists for the peroxisome proliferator-activated receptors (PPARs) as well as fatty acids have been shown to increase murine UCP2 and UCP3 mRNA expression but response elements and mechanisms...... are not yet characterized. The aim of this study was to investigate the transcriptional regulation of UCP3 by the PPARs. Results: The PPAR agonists increase UCP2 and UCP3 mRNA expression in skeletal muscle cells (C2C12). In addition, UCP2 and UCP3 mRNA expression is upregulated during 3T3-L1 adipocyte...... differentiation. Results from sequencing of chromatin immunoprecipitated (ChIP) material from 3T3-L1 adipocytes revealed three PPAR and retinoid X receptor (RXR) binding sites in and around the murine UCP3 gene. The recruitment of PPAR and RXR to the three potential PPREs was determined by ChIP combined...

  9. Isolation and characterization of full-length putative alcohol dehydrogenase genes from polygonum minus

    Science.gov (United States)

    Hamid, Nur Athirah Abd; Ismail, Ismanizan

    2013-11-01

    Polygonum minus, locally named as Kesum is an aromatic herb which is high in secondary metabolite content. Alcohol dehydrogenase is an important enzyme that catalyzes the reversible oxidation of alcohol and aldehyde with the presence of NAD(P)(H) as co-factor. The main focus of this research is to identify the gene of ADH. The total RNA was extracted from leaves of P. minus which was treated with 150 μM Jasmonic acid. Full-length cDNA sequence of ADH was isolated via rapid amplification cDNA end (RACE). Subsequently, in silico analysis was conducted on the full-length cDNA sequence and PCR was done on genomic DNA to determine the exon and intron organization. Two sequences of ADH, designated as PmADH1 and PmADH2 were successfully isolated. Both sequences have ORF of 801 bp which encode 266 aa residues. Nucleotide sequence comparison of PmADH1 and PmADH2 indicated that both sequences are highly similar at the ORF region but divergent in the 3' untranslated regions (UTR). The amino acid is differ at the 107 residue; PmADH1 contains Gly (G) residue while PmADH2 contains Cys (C) residue. The intron-exon organization pattern of both sequences are also same, with 3 introns and 4 exons. Based on in silico analysis, both sequences contain "classical" short chain alcohol dehydrogenases/reductases ((c) SDRs) conserved domain. The results suggest that both sequences are the members of short chain alcohol dehydrogenase family.

  10. Phytohormone and Putative Defense Gene Expression Differentiates the Response of ‘Hayward’ Kiwifruit to Psa and Pfm Infections

    Directory of Open Access Journals (Sweden)

    Kirstin V. Wurms

    2017-08-01

    Full Text Available Pseudomonas syringae pv. actinidiae (Psa and Pseudomonas syringae pv. actinidifoliorum (Pfm are closely related pathovars infecting kiwifruit, but Psa is considered one of the most important global pathogens, whereas Pfm is not. In this study of Actinidia deliciosa ‘Hayward’ responses to the two pathovars, the objective was to test whether differences in plant defense responses mounted against the two pathovars correlated with the contrasting severity of the symptoms caused by them. Results showed that Psa infections were always more severe than Pfm infections, and were associated with highly localized, differential expression of phytohormones and putative defense gene transcripts in stem tissue closest to the inoculation site. Phytohormone concentrations of jasmonic acid (JA, jasmonate isoleucine (JA-Ile, salicylic acid (SA and abscisic acid were always greater in stem tissue than in leaves, and leaf phytohormones were not affected by pathogen inoculation. Pfm inoculation induced a threefold increase in SA in stems relative to Psa inoculation, and a smaller 1.6-fold induction of JA. Transcript expression showed no effect of inoculation in leaves, but Pfm inoculation resulted in the greatest elevation of the SA marker genes, PR1 and glucan endo-1,3-beta-glucosidase (β-1,3-glucosidase (32- and 25-fold increases, respectively in stem tissue surrounding the inoculation site. Pfm inoculation also produced a stronger response than Psa inoculation in localized stem tissue for the SA marker gene PR6, jasmonoyl-isoleucine-12-hydrolase (JIH1, which acts as a negative marker of the JA pathway, and APETALA2/Ethylene response factor 2 transcription factor (AP2 ERF2, which is involved in JA/SA crosstalk. WRKY40 transcription factor (a SA marker was induced equally in stems by wounding (mock inoculation and pathovar inoculation. Taken together, these results suggest that the host appears to mount a stronger, localized, SA-based defense response to Pfm

  11. The study of the transformer gene from Bactrocera dorsalis and B. correcta with putative core promoter regions.

    Science.gov (United States)

    Laohakieat, Kamoltip; Aketarawong, Nidchaya; Isasawin, Siriwan; Thitamadee, Siripong; Thanaphum, Sujinda

    2016-02-01

    The transformer (tra) is a sex determining switch in different orders of insects, including Diptera, as in the family Tephritidae. The lifelong autoregulatory loop of tra female-specific splicing can be reset by the intervention of male-specific primary signals (M factor). In early development, the functional female and truncated male TRA proteins relay the sexual fates to the alternative splicing of a bisexual switch gene, doublesex (dsx) cascading the sexual differentiation processes. Bactrocera dorsalis (Hendel) and Bactrocera correcta (Bezzi) are among the Bactrocera model worldwide key pests. Area-wide integrated pest management using the male-only Sterile Insect Technique (SIT) relying on genetic sexing systems is effective in control programs. We undertook the molecular characterization and comparative studies of the tra orthologues in the Bactrocera species, including the Salaya1 genetic sexing strain (GSS). RT-PCR revealed that B. dorsalis tra (Bdtra) and B. correcta tra (Bctra) transcripts contained conservation of both constitutive exons and male-specific exons as in other Bactrocera. However, new Bdtra male-specific exons were retained, diversifying the pattern of the male-specifically spliced transcripts. The coding sequences of tra were highly conserved in Bactrocera (86-95%) but less so among related genera (61-65%) within the same Tephritidae family. A conservation of deduced amino acid sequences (18 residues), called the TEP region, was identified to be distinctive among tephritids. The 5' regulatory sequence containing many structural characteristics of the putative core promoter was discovered in B. correcta. The expression patterns of Bdtra and Bctra were sex-specifically spliced and the signals relayed to the dsx genes in the adult wild-types. However, the coexistence of male- and female-specifically spliced transcripts (980 and 626 bp, respectively) of the B. dorsalis wild-type strain was found in the Salaya1 GSS adult males. The Bdtra RNA

  12. Polymorphism screening of four genes encoding advanced glycation end-product putative receptors. Association study with nephropathy in type 1 diabetic patients

    DEFF Research Database (Denmark)

    Poirier, Odette; Nicaud, Viviane; Vionnet, N

    2001-01-01

    Advanced glycation end-products (AGEs) may play an important role in the pathogenesis and progression of cardiovascular and renal complications of diabetes. Four putative AGE receptors (RAGEs), AGE-R1, AGE-R2, and AGE-R3 have been described. In this study, we scanned the sequence of the genes enc...

  13. Polymorphism screening of four genes encoding advanced glycation end-product putative receptors. Association study with nephropathy in type 1 diabetic patients

    DEFF Research Database (Denmark)

    Poirier, Odette; Nicaud, Viviane; Vionnet, N

    2001-01-01

    Advanced glycation end-products (AGEs) may play an important role in the pathogenesis and progression of cardiovascular and renal complications of diabetes. Four putative AGE receptors (RAGEs), AGE-R1, AGE-R2, and AGE-R3 have been described. In this study, we scanned the sequence of the genes....... The minor allele of a polymorphism located in the promoter region of the RAGE gene (C-1152A) conferred a weak protective effect (P

  14. Phylogenetic and comparative gene expression analysis of barley (Hordeum vulgare)WRKY transcription factor family reveals putatively retained functions betweenmonocots and dicots

    Energy Technology Data Exchange (ETDEWEB)

    Mangelsen, Elke; Kilian, Joachim; Berendzen, Kenneth W.; Kolukisaoglu, Uner; Harter, Klaus; Jansson, Christer; Wanke, Dierk

    2008-02-01

    WRKY proteins belong to the WRKY-GCM1 superfamily of zinc finger transcription factors that have been subject to a large plant-specific diversification. For the cereal crop barley (Hordeum vulgare), three different WRKY proteins have been characterized so far, as regulators in sucrose signaling, in pathogen defense, and in response to cold and drought, respectively. However, their phylogenetic relationship remained unresolved. In this study, we used the available sequence information to identify a minimum number of 45 barley WRKY transcription factor (HvWRKY) genes. According to their structural features the HvWRKY factors were classified into the previously defined polyphyletic WRKY subgroups 1 to 3. Furthermore, we could assign putative orthologs of the HvWRKY proteins in Arabidopsis and rice. While in most cases clades of orthologous proteins were formed within each group or subgroup, other clades were composed of paralogous proteins for the grasses and Arabidopsis only, which is indicative of specific gene radiation events. To gain insight into their putative functions, we examined expression profiles of WRKY genes from publicly available microarray data resources and found group specific expression patterns. While putative orthologs of the HvWRKY transcription factors have been inferred from phylogenetic sequence analysis, we performed a comparative expression analysis of WRKY genes in Arabidopsis and barley. Indeed, highly correlative expression profiles were found between some of the putative orthologs. HvWRKY genes have not only undergone radiation in monocot or dicot species, but exhibit evolutionary traits specific to grasses. HvWRKY proteins exhibited not only sequence similarities between orthologs with Arabidopsis, but also relatedness in their expression patterns. This correlative expression is indicative for a putative conserved function of related WRKY proteins in mono- and dicot species.

  15. Phylogenetic and comparative gene expression analysis of barley (Hordeum vulgare WRKY transcription factor family reveals putatively retained functions between monocots and dicots

    Directory of Open Access Journals (Sweden)

    Jansson Christer

    2008-04-01

    Full Text Available Abstract Background WRKY proteins belong to the WRKY-GCM1 superfamily of zinc finger transcription factors that have been subject to a large plant-specific diversification. For the cereal crop barley (Hordeum vulgare, three different WRKY proteins have been characterized so far as regulators in sucrose signaling, pathogen defense, and in response to cold and drought. However, their phylogenetic relationship remained unresolved. Results In this study, we used available sequence information to identify a minimum number of 45 barley WRKY transcription factor (HvWRKY genes. According to their structural features, the HvWRKY factors were classified into the previously defined polyphyletic WRKY subgroups 1 to 3. Furthermore, we could assign putative orthologs of the HvWRKY proteins in Arabidopsis and rice. While in most cases clades of orthologous proteins were formed within each group or subgroup, other clades were composed of paralogous proteins for the grasses and Arabidopsis only, which is indicative of specific gene radiation events. To gain insight into their putative functions, we examined expression profiles of WRKY genes from publicly available microarray data resources and found group specific expression patterns. While putative orthologs of the HvWRKY transcription factors have been inferred from phylogenetic sequence analysis, we performed a comparative expression analysis of WRKY genes in Arabidopsis and barley. Indeed, highly correlative expression profiles were found between some of the putative orthologs. Conclusion HvWRKY genes have not only undergone radiation in monocot or dicot species, but exhibit evolutionary traits specific to grasses. HvWRKY proteins exhibited not only sequence similarities between orthologs with Arabidopsis, but also relatedness in their expression patterns. This correlative expression is indicative for a putative conserved function of related WRKY proteins in monocot and dicot species.

  16. Identification of genes coding for putative wax ester synthase/diacylglycerol acyltransferase enzymes in terrestrial and marine environments.

    Science.gov (United States)

    Lanfranconi, Mariana P; Alvarez, Adrián F; Alvarez, Héctor M

    2015-12-01

    Synthesis of neutral lipids such as triacylglycerols (TAG) and wax esters (WE) is catalyzed in bacteria by wax ester synthase/diacylglycerol acyltransferase enzymes (WS/DGAT). We investigated the diversity of genes encoding this enzyme in contrasting natural environments from Patagonia (Argentina). The content of petroleum hydrocarbons in samples collected from oil-producing areas was measured. PCR-based analysis covered WS/DGAT occurrence in marine sediments and soil. No product was obtained in seawater samples. All clones retrieved from marine sediments affiliated with gammaproteobacterial sequences and within them, most phylotypes formed a unique cluster related to putative WS/DGAT belonging to marine OM60 clade. In contrast, soils samples contained phylotypes only related to actinomycetes. Among them, phylotypes affiliated with representatives largely or recently reported as oleaginous bacteria, as well as with others considered as possible lipid-accumulating bacteria based on the analysis of their annotated genomes. Our study shows for the first time that the environment could contain a higher variety of ws/dgat than that reported from bacterial isolates. The results of this study highlight the relevance of the environment in a natural process such as the synthesis and accumulation of neutral lipids. Particularly, both marine sediments and soil may serve as a useful source for novel WS/DGAT with biotechnological interest.

  17. RNA interference of a putative S-adenosyl-L-homocysteine hydrolase gene affects larval performance in Leptinotarsa decemlineata (Say).

    Science.gov (United States)

    Zhou, Li-Tao; Jia, Shuang; Wan, Pin-Jun; Kong, Ye; Guo, Wen-Chao; Ahmat, Tursun; Li, Guo-Qing

    2013-10-01

    In Leptinotarsa decemlineata, juvenile hormones (JHs) play primary roles in the regulation of metamorphosis, reproduction and diapause. In JH biosynthetic pathway in insect corpora allata, methylation of farnesoic acid or JH acid using S-adenosyl-L-methionine generates a potent feedback inhibitor S-adenosyl-L-homocysteine (AdoHcy). Rapid removal of AdoHcy is hypothesized to be essential for JH synthesis. AdoHcy hydrolase (SAHase) is the only eukaryotic enzyme catalyzing the removal. In the present paper, we firstly cloned a putative LdSAHase gene from L. decemlineata. The cDNA consists of 1806 bp and encodes a 525 amino acid protein. LdSAHase was expressed in all developmental stages. The gene had the highest and the lowest level of transcription respectively in the 3rd- and 4th-instars' heads that contain corpora allata, which was positively correlated with JH titer in the haemolymph and the mRNA level of a JH early-inducible gene, the Krüppel homolog 1 gene (Kr-h1). Secondly, dietary ingestion of bacterially-expressed LdSAHase-dsRNA significantly decreased LdSAHase and LdKr-h1 mRNA levels, reduced JH titer, and caused the death of the larvae, and the failure of pupation and adult emergence. After continuous exposure for 12 days, 42% of the larvae died, 65% of the prepupae failed to pupate and 100% of the pupae failed to emerge. Moreover, RNAi-mediated LdSAHase knockdown also reduced larval developing time, and decreased larval weight. Lastly, application of JH analogue pyriproxyfen to LdSAHase-dsRNA-exposed larvae did not greatly increase LdSAHase expression level and JH content, but up-regulated LdKr-h1 mRNA level. Expectedly, pyriproxyfen application could partially rescue the negative effects on the survival and the development. Thus, our results support the hypothesis that SAHase plays a critical role in JH biosynthesis in insects.

  18. Transcriptome analysis of Panax vietnamensis var. fuscidicus discovers putative ocotillol-type ginsenosides biosynthesis genes and genetic markers.

    Science.gov (United States)

    Zhang, Guang-Hui; Ma, Chun-Hua; Zhang, Jia-Jin; Chen, Jun-Wen; Tang, Qing-Yan; He, Mu-Han; Xu, Xiang-Zeng; Jiang, Ni-Hao; Yang, Sheng-Chao

    2015-03-08

    liquid chromatography (HPLC) and evaporative light scattering detector (ELSD). The genomic resources generated from P. vietnamensis var. fuscidiscus provide new insights into the identification of putative genes involved in triterpenoid saponins biosynthesis pathway. This will facilitate our understanding of the biosynthesis of triterpenoid saponins at molecular level. The SSR markers identified and developed in this study show genetic diversity for this important crop and will contribute to marker-assisted breeding for P. vietnamensis var. fuscidiscus.

  19. Global gene expression analysis reveals reduced abundance of putative microRNA targets in human prostate tumours

    Directory of Open Access Journals (Sweden)

    Xie Yi

    2009-02-01

    Full Text Available Abstract Background Recently, microRNAs (miRNAs have taken centre stage in the field of human molecular oncology. Several studies have shown that miRNA profiling analyses offer new possibilities in cancer classification, diagnosis and prognosis. However, the function of miRNAs that are dysregulated in tumours remains largely a mystery. Global analysis of miRNA-target gene expression has helped illuminate the role of miRNAs in developmental gene expression programs, but such an approach has not been reported in cancer transcriptomics. Results In this study, we globally analysed the expression patterns of miRNA target genes in prostate cancer by using several public microarray datasets. Intriguingly, we found that, in contrast to global mRNA transcript levels, putative miRNA targets showed a reduced abundance in prostate tumours relative to benign prostate tissue. Additionally, the down-regulation of these miRNA targets positively correlated with the number of types of miRNA target-sites in the 3' untranslated regions of these targets. Further investigation revealed that the globally low expression was mainly driven by the targets of 36 specific miRNAs that were reported to be up-regulated in prostate cancer by a miRNA expression profiling study. We also found that the transcript levels of miRNA targets were lower in androgen-independent prostate cancer than in androgen-dependent prostate cancer. Moreover, when the global analysis was extended to four other cancers, significant differences in transcript levels between miRNA targets and total mRNA backgrounds were found. Conclusion Global gene expression analysis, along with further investigation, suggests that miRNA targets have a significantly reduced transcript abundance in prostate cancer, when compared with the combined pool of all mRNAs. The abnormal expression pattern of miRNA targets in human cancer could be a common feature of the human cancer transcriptome. Our study may help to shed new

  20. SlDEAD31, a Putative DEAD-Box RNA Helicase Gene, Regulates Salt and Drought Tolerance and Stress-Related Genes in Tomato.

    Directory of Open Access Journals (Sweden)

    Mingku Zhu

    Full Text Available The DEAD-box RNA helicases are involved in almost every aspect of RNA metabolism, associated with diverse cellular functions including plant growth and development, and their importance in response to biotic and abiotic stresses is only beginning to emerge. However, none of DEAD-box genes was well characterized in tomato so far. In this study, we reported on the identification and characterization of two putative DEAD-box RNA helicase genes, SlDEAD30 and SlDEAD31 from tomato, which were classified into stress-related DEAD-box proteins by phylogenetic analysis. Expression analysis indicated that SlDEAD30 was highly expressed in roots and mature leaves, while SlDEAD31 was constantly expressed in various tissues. Furthermore, the expression of both genes was induced mainly in roots under NaCl stress, and SlDEAD31 mRNA was also increased by heat, cold, and dehydration. In stress assays, transgenic tomato plants overexpressing SlDEAD31 exhibited dramatically enhanced salt tolerance and slightly improved drought resistance, which were simultaneously demonstrated by significantly enhanced expression of multiple biotic and abiotic stress-related genes, higher survival rate, relative water content (RWC and chlorophyll content, and lower water loss rate and malondialdehyde (MDA production compared to wild-type plants. Collectively, these results provide a preliminary characterization of SlDEAD30 and SlDEAD31 genes in tomato, and suggest that stress-responsive SlDEAD31 is essential for salt and drought tolerance and stress-related gene regulation in plants.

  1. SlDEAD31, a Putative DEAD-Box RNA Helicase Gene, Regulates Salt and Drought Tolerance and Stress-Related Genes in Tomato.

    Science.gov (United States)

    Zhu, Mingku; Chen, Guoping; Dong, Tingting; Wang, Lingling; Zhang, Jianling; Zhao, Zhiping; Hu, Zongli

    2015-01-01

    The DEAD-box RNA helicases are involved in almost every aspect of RNA metabolism, associated with diverse cellular functions including plant growth and development, and their importance in response to biotic and abiotic stresses is only beginning to emerge. However, none of DEAD-box genes was well characterized in tomato so far. In this study, we reported on the identification and characterization of two putative DEAD-box RNA helicase genes, SlDEAD30 and SlDEAD31 from tomato, which were classified into stress-related DEAD-box proteins by phylogenetic analysis. Expression analysis indicated that SlDEAD30 was highly expressed in roots and mature leaves, while SlDEAD31 was constantly expressed in various tissues. Furthermore, the expression of both genes was induced mainly in roots under NaCl stress, and SlDEAD31 mRNA was also increased by heat, cold, and dehydration. In stress assays, transgenic tomato plants overexpressing SlDEAD31 exhibited dramatically enhanced salt tolerance and slightly improved drought resistance, which were simultaneously demonstrated by significantly enhanced expression of multiple biotic and abiotic stress-related genes, higher survival rate, relative water content (RWC) and chlorophyll content, and lower water loss rate and malondialdehyde (MDA) production compared to wild-type plants. Collectively, these results provide a preliminary characterization of SlDEAD30 and SlDEAD31 genes in tomato, and suggest that stress-responsive SlDEAD31 is essential for salt and drought tolerance and stress-related gene regulation in plants.

  2. Functional expression of SAV3818, a putative TetR-family transcriptional regulatory gene from Streptomyces avermitilis, stimulates antibiotic production in Streptomyces species.

    Science.gov (United States)

    Duong, Cac Thi Phung; Lee, Han-Na; Choi, Si-Sun; Lee, Sang Yup; Kim, Eung-Soo

    2009-02-01

    Avermectin and its analogs are major commercial antiparasitic agents in the fields of animal health, agriculture, and human infections. Previously, comparative transcriptome analysis between the low-producer S. avermitilis ATCC31267 and the high-producer S. avermitilis ATCC31780 using a S. avermitilis whole genome chip revealed that 50 genes were overexpressed at least two-fold higher in S. avermitilis ATCC31780. To verify the biological significance of some of the transcriptomics-guided targets, five putative regulatory genes were individually cloned under the strong-andconstitutive promoter of the Streptomyces expression vector pSE34, followed by the transformation into the lowproducer S. avermitilis ATCC31267. Among the putative genes tested, three regulatory genes including SAV213, SAV3818, and SAV4023 exhibited stimulatory effects on avermectin production in S. avermitilis ATCC31267. Moreover, overexpression of SAV3818 also stimulated actinorhodin production in both S. coelicolor M145 and S. lividans TK21, implying that the SAV3818, a putative TetR-family transcriptional regulator, could be a global upregulator acting in antibiotic production in Streptomyces species.

  3. An LTR Retrotransposon-Derived Gene Displays Lineage-Specific Structural and Putative Species-Specific Functional Variations in Eutherians

    Science.gov (United States)

    Irie, Masahito; Koga, Akihiko; Kaneko-Ishino, Tomoko; Ishino, Fumitoshi

    2016-01-01

    Amongst the 11 eutherian-specific genes acquired from a sushi-ichi retrotransposon is the CCHC type zinc-finger protein-encoding gene SIRH11/ZCCHC16. Its contribution to eutherian brain evolution is implied because of its involvement in cognitive function in mice, possibly via the noradrenergic system. Although, the possibility that Sirh11/Zcchc16 functions as a non-coding RNA still remains, dN/dS ratios in pairwise comparisons between its orthologs have provided supportive evidence that it acts as a protein. It became a pseudogene in armadillos (Cingulata) and sloths (Pilosa), the only two extant orders of xenarthra, which prompted us to examine the lineage-specific variations of SIRH11/ZCCHC16 in eutherians. We examined the predicted SIRH11/ZCCHC16 open reading frame (ORF) in 95 eutherian species based on the genomic DNA information in GenBank. A large variation in the SIRH11/ZCCHC16 ORF was detected in several lineages. These include a lack of a CCHC RNA-binding domain in its C-terminus, observed in gibbons (Hylobatidae: Primates) and megabats (Megachiroptera: Chiroptera). A lack of the N-terminal half, on the other hand, was observed in New World monkeys (Platyrrhini: Primates) and species belonging to New World and African Hystricognaths (Caviomorpha and Bathyergidae: Rodents) along with Cetacea and Ruminantia (Cetartiodactyla). Among the hominoids, interestingly, three out of four genera of gibbons have lost normal SIRH11/ZCCHC16 function by deletion or the lack of the CCHC RNA-binding domain. Our extensive dN/dS analysis suggests that such truncated SIRH11/ZCCHC16 ORFs are functionally diversified even within lineages. Combined, our results show that SIRH11/ZCCHC16 may contribute to the diversification of eutherians by lineage-specific structural changes after its domestication in the common eutherian ancestor, followed by putative species-specific functional changes that enhanced fitness and occurred as a consequence of complex natural selection events

  4. The putative-farnesoic acid O-methyl transferase (FAMeT) gene of Ceratitis capitata: characterization and pre-imaginal life expression.

    Science.gov (United States)

    Vannini, Laura; Ciolfi, Silvia; Spinsanti, Giacomo; Panti, Cristina; Frati, Francesco; Dallai, Romano

    2010-02-01

    Farnesoic acid O-methyl transferase (FAMeT) is the enzyme involved in the penultimate step of insect juvenile hormone (JH) biosynthesis and is thus a key regulator in insect development and reproduction. We report the characterization of the putative-FAMeT in the medfly or Mediterranean fruit fly, Ceratitis capitata. This gene was identified by suppressive subtractive hybridization and completely sequenced by the screening of a medfly cDNA library. The obtained sequence was analyzed for conserved protein domain identification and its expression profile was evaluated by quantitative Real-Time PCR in medfly pre-imaginal life. The tissue expression of the isolated gene was verified by in situ hybridization on third instar larvae sections. The characterization of the isolated gene pointed out several typical features of methyl transferase genes. The pre-imaginal putative-FAMeT expression levels were consistent with JH titer change in Diptera. As recognized in some crustaceans, this gene seems to be widely expressed in the medfly as well. Ceratitis capitata is one of the most relevant agricultural pests against which insecticides and the sterile insect technique (SIT) are extensively used in spite of the well-known limitations of these approaches. Although results are not conclusive for the physiological role of the isolated gene, they suggest the characterization of a new gene in the Mediterranean fruit fly potentially involved in JH biosynthesis and may, therefore, have implications for pest control. 2010 Wiley Periodicals, Inc.

  5. Screening for Genes Coding for Putative Antitumor Compounds, Antimicrobial and Enzymatic Activities from Haloalkalitolerant and Haloalkaliphilic Bacteria Strains of Algerian Sahara Soils

    Directory of Open Access Journals (Sweden)

    Okba Selama

    2014-01-01

    Full Text Available Extreme environments may often contain unusual bacterial groups whose physiology is distinct from those of normal environments. To satisfy the need for new bioactive pharmaceuticals compounds and enzymes, we report here the isolation of novel bacteria from an extreme environment. Thirteen selected haloalkalitolerant and haloalkaliphilic bacteria were isolated from Algerian Sahara Desert soils. These isolates were screened for the presence of genes coding for putative antitumor compounds using PCR based methods. Enzymatic, antibacterial, and antifungal activities were determined by using cultural dependant methods. Several of these isolates are typical of desert and alkaline saline soils, but, in addition, we report for the first time the presence of a potential new member of the genus Nocardia with particular activity against the yeast Saccharomyces cerevisiae. In addition to their haloalkali character, the presence of genes coding for putative antitumor compounds, combined with the antimicrobial activity against a broad range of indicator strains and their enzymatic potential, makes them suitable for biotechnology applications.

  6. Screening for genes coding for putative antitumor compounds, antimicrobial and enzymatic activities from haloalkalitolerant and haloalkaliphilic bacteria strains of Algerian Sahara Soils.

    Science.gov (United States)

    Selama, Okba; Amos, Gregory C A; Djenane, Zahia; Borsetto, Chiara; Laidi, Rabah Forar; Porter, David; Nateche, Farida; Wellington, Elizabeth M H; Hacène, Hocine

    2014-01-01

    Extreme environments may often contain unusual bacterial groups whose physiology is distinct from those of normal environments. To satisfy the need for new bioactive pharmaceuticals compounds and enzymes, we report here the isolation of novel bacteria from an extreme environment. Thirteen selected haloalkalitolerant and haloalkaliphilic bacteria were isolated from Algerian Sahara Desert soils. These isolates were screened for the presence of genes coding for putative antitumor compounds using PCR based methods. Enzymatic, antibacterial, and antifungal activities were determined by using cultural dependant methods. Several of these isolates are typical of desert and alkaline saline soils, but, in addition, we report for the first time the presence of a potential new member of the genus Nocardia with particular activity against the yeast Saccharomyces cerevisiae. In addition to their haloalkali character, the presence of genes coding for putative antitumor compounds, combined with the antimicrobial activity against a broad range of indicator strains and their enzymatic potential, makes them suitable for biotechnology applications.

  7. Comparison and Validation of Putative Pathogenicity-Related Genes Identified by T-DNA Insertional Mutagenesis and Microarray Expression Profiling in Magnaporthe oryzae

    Science.gov (United States)

    Wáng, Ying; Tan, Qi; Gao, Ying Nv; Li, Yan

    2017-01-01

    High-throughput technologies of functional genomics such as T-DNA insertional mutagenesis and microarray expression profiling have been employed to identify genes related to pathogenicity in Magnaporthe oryzae. However, validation of the functions of individual genes identified by these high-throughput approaches is laborious. In this study, we compared two published lists of genes putatively related to pathogenicity in M. oryzae identified by T-DNA insertional mutagenesis (comprising 1024 genes) and microarray expression profiling (comprising 236 genes), respectively, and then validated the functions of some overlapped genes between the two lists by knocking them out using the method of target gene replacement. Surprisingly, only 13 genes were overlapped between the two lists, and none of the four genes selected from the overlapped genes exhibited visible phenotypic changes on vegetative growth, asexual reproduction, and infection ability in their knockout mutants. Our results suggest that both of the lists might contain large proportions of unrelated genes to pathogenicity and therefore comparing the two gene lists is hardly helpful for the identification of genes that are more likely to be involved in pathogenicity as we initially expected.

  8. The tep1 gene of Sinorhizobium meliloti coding for a putative transmembrane efflux protein and N-acetyl glucosamine affect nod gene expression and nodulation of alfalfa plants

    Directory of Open Access Journals (Sweden)

    Soto María

    2009-01-01

    Full Text Available Abstract Background Soil bacteria collectively known as Rhizobium, characterized by their ability to establish beneficial symbiosis with legumes, share several common characteristics with pathogenic bacteria when infecting the host plant. Recently, it was demonstrated that a fadD mutant of Sinorhizobium meliloti is altered in the control of swarming, a type of co-ordinated movement previously associated with pathogenicity, and is also impaired in nodulation efficiency on alfalfa roots. In the phytopathogen Xanthomonas campestris, a fadD homolog (rpfB forms part of a cluster of genes involved in the regulation of pathogenicity factors. In this work, we have investigated the role in swarming and symbiosis of SMc02161, a S. meliloti fadD-linked gene. Results The SMc02161 locus in S. meliloti shows similarities with members of the Major Facilitator Superfamily (MFS of transporters. A S. meliloti null-mutant shows increased sensitivity to chloramphenicol. This indication led us to rename the locus tep1 for transmembrane efflux protein. The lack of tep1 does not affect the appearance of swarming motility. Interestingly, nodule formation efficiency on alfalfa plants is improved in the tep1 mutant during the first days of the interaction though nod gene expression is lower than in the wild type strain. Curiously, a nodC mutation or the addition of N-acetyl glucosamine to the wild type strain lead to similar reductions in nod gene expression as in the tep1 mutant. Moreover, aminosugar precursors of Nod factors inhibit nodulation. Conclusion tep1 putatively encodes a transmembrane protein which can confer chloramphenicol resistance in S. meliloti by expelling the antibiotic outside the bacteria. The improved nodulation of alfalfa but reduced nod gene expression observed in the tep1 mutant suggests that Tep1 transports compounds which influence nodulation. In contrast to Bradyrhizobium japonicum, we show that in S. meliloti there is no feedback regulation

  9. The tep1 gene of Sinorhizobium meliloti coding for a putative transmembrane efflux protein and N-acetyl glucosamine affect nod gene expression and nodulation of alfalfa plants.

    Science.gov (United States)

    van Dillewijn, Pieter; Sanjuán, Juan; Olivares, José; Soto, María José

    2009-01-27

    Soil bacteria collectively known as Rhizobium, characterized by their ability to establish beneficial symbiosis with legumes, share several common characteristics with pathogenic bacteria when infecting the host plant. Recently, it was demonstrated that a fadD mutant of Sinorhizobium meliloti is altered in the control of swarming, a type of co-ordinated movement previously associated with pathogenicity, and is also impaired in nodulation efficiency on alfalfa roots. In the phytopathogen Xanthomonas campestris, a fadD homolog (rpfB) forms part of a cluster of genes involved in the regulation of pathogenicity factors. In this work, we have investigated the role in swarming and symbiosis of SMc02161, a S. meliloti fadD-linked gene. The SMc02161 locus in S. meliloti shows similarities with members of the Major Facilitator Superfamily (MFS) of transporters. A S. meliloti null-mutant shows increased sensitivity to chloramphenicol. This indication led us to rename the locus tep1 for transmembrane efflux protein. The lack of tep1 does not affect the appearance of swarming motility. Interestingly, nodule formation efficiency on alfalfa plants is improved in the tep1 mutant during the first days of the interaction though nod gene expression is lower than in the wild type strain. Curiously, a nodC mutation or the addition of N-acetyl glucosamine to the wild type strain lead to similar reductions in nod gene expression as in the tep1 mutant. Moreover, aminosugar precursors of Nod factors inhibit nodulation. tep1 putatively encodes a transmembrane protein which can confer chloramphenicol resistance in S. meliloti by expelling the antibiotic outside the bacteria. The improved nodulation of alfalfa but reduced nod gene expression observed in the tep1 mutant suggests that Tep1 transports compounds which influence nodulation. In contrast to Bradyrhizobium japonicum, we show that in S. meliloti there is no feedback regulation of nodulation genes. Moreover, the Nod factor precursor

  10. Large number of putative chemoreception and pheromone biosynthesis genes revealed by analyzing transcriptome from ovipositor-pheromone glands of Chilo suppressalis.

    Science.gov (United States)

    Xia, Yi-Han; Zhang, Ya-Nan; Hou, Xiao-Qing; Li, Fei; Dong, Shuang-Lin

    2015-01-20

    The chemoreception role of moth ovipositor has long been suggested, but its molecular mechanism is mostly unknown. By transcriptomic analysis of the female ovipositor-pheromone glands (OV-PG) of Chilo suppressalis, we obtained 31 putative chemoreception genes (9 OBPs, 10 CSPs, 2 ORs, 1 SNMP, 8 CXEs and 1 AOX), in addition to 32 genes related to sex pheromone biosynthesis (1 FAS, 6 Dess, 10 FARs, 2 ACOs, 1 ACC, 4 FATPs, 3 ACBPs and 5 ELOs). Tissue expression profiles further revealed that CsupCSP2 and CsupCSP10 were OV-PG biased, while most chemoreception genes were highly and preferably expressed in antennae. This suggests that OV-PG employs mostly the same chemoreception proteins as in antennae, although the physiological roles of these proteins might be different in OV-PG. Of the 32 pheromone biosynthesis related genes, CsupDes4, CsupDes5 and CsupFAR2 are strongly OV-PG biased, and clustered with functionally validated genes from other moths, strongly indicating their involvement in specific step of the pheromone biosynthesis. Our study for the first time identified a large number of putative chemoreception genes, and provided an important basis for exploring the chemoreception mechanisms of OV-PG in C. suppressalis, as well as other moth species.

  11. Identification and Characterization of microRNA319a and Its Putative Target Gene, PvPCF5, in the Bioenergy Grass Switchgrass (Panicum virgatum)

    Science.gov (United States)

    Xie, Qi; Liu, Xue; Zhang, Yinbing; Tang, Jinfu; Yin, Dedong; Fan, Bo; Zhu, Lihuang; Han, Liebao; Song, Guilong; Li, Dayong

    2017-01-01

    Due to its high biomass yield, low environmental impact, and widespread adaptability to poor soils and harsh conditions, switchgrass (Panicum virgatum L.), a warm-region perennial herbaceous plant, has attracted much attention in recent years. However, little is known about microRNAs (miRNAs) and their functions in this bioenergy grass. Here, we identified and characterized a miRNA gene, Pvi-MIR319a, encoding microRNA319a in switchgrass. Transgenic rice lines generated by overexpressing the Pvi-MIR319a precursor gene exhibited broader leaves and delayed flowering compared with the control. Gene expression analysis indicated at least four putative target genes were downregulated. Additionally, we cloned a putative target gene (PvPCF5) of Pvi-MIR319a from switchgrass. PvPCF5, a TCP transcription factor, is a nuclear-localized protein with transactivation activity and control the development of leaf. Our results suggest that Pvi-MIR319a and its target genes may be used as potential genetic regulators for future switchgrass genetic improvement. PMID:28424710

  12. Gravistimulation changes expression of genes encoding putative carrier proteins of auxin polar transport in etiolated pea epicotyls

    Science.gov (United States)

    Hoshino, T.; Hitotsubashi, R.; Miyamoto, K.; Tanimoto, E.; Ueda, J.

    STS-95 space experiment has showed that auxin polar transport in etiolated epicotyls of pea (Pisum sativum L. cv. Alaska) seedlings is controlled by gravistimulation. In Arabidopsis thaliana auxin polar transport has considered to be regulated by efflux and influx carrier proteins in plasma membranes, AtPIN1 and AtAUX1, respectively. In order to know how gravistimuli control auxin polar transport in etiolated pea epicotyls at molecular levels, strenuous efforts have been made, resulting in successful isolation of full-length cDNAs of a putative auxin efflux and influx carriers, PsPIN2 and PsAUX1, respectively. Significantly high levels in homology were found on nucleotide and deduced amino acid sequences among PsPIN2, PsPIN1 (accession no. AY222857, Chawla and DeMason, 2003) and AtPINs, and also among PsAUX1, AtAUX1 and their related genes. Phylogenetic analyses based on the deduced amino acid sequences revealed that PsPIN2 belonged to a subclade including AtPIN3, AtPIN4 relating to lateral transport of auxin, while PsPIN1 belonged to the same clade as AtPIN1 relating to auxin polar transport. In the present study, we examined the effects of gravistimuli on the expression of PsPINs and PsAUX1 in etiolated pea seedlings by northern blot analysis. Expression of PsPIN1, PsPIN2 and PsAUX1 in hook region of 3.5-d-old etiolated pea seedlings grown under simulated microgravity conditions on a 3-D clinostat increased as compared with that of the seedlings grown under 1 g conditions. On the other hand, that of PsPIN1 and PsAUX1 in the 1st internode region under simulated microgravity conditions on a 3-D clinostat also increased, while that of PsPIN2 was affected little. These results suggest that expression of PsPIN1, PsPIN2 and PsAUX1 regulating polar/lateral transport of auxin is substantially under the control of gravity. A possible role of PsPINs and PsAUX1 of auxin polar transport in etiolated pea seedlings will also be discussed.

  13. Tumor-associated methylation of the putative tumor suppressor AJAP1 gene and association between decreased AJAP1 expression and shorter survival in patients with glioma

    Institute of Scientific and Technical Information of China (English)

    David Cogdell; Woonbok Chung; Yuexin Liu; Matthew McDonald; Kenneth Aldape; Jean-Pierre J. Issa; Gregory N. Fuller; Wei Zhang

    2011-01-01

    Allelic loss of the short arm of chromosome 1 has been observed frequently in a wide spectrum of cancers, most frequently in oligodendroglioma. In our previous studies, we evaluated 177 oligodendroglial tumor samples and identified the AJAP1 gene (formerly Shrew1) in the consensus region of deletion.AJAP1 is a transmembrane protein found in adheren junctions and functions to inhibit glioma cell adhesion and migration. Whereas a putative tumor suppressor gene, we did not detect AJAP1 gene mutations. In subsequent studies, we found that AJAP1 was underexpressed in oligodendrogliomas relative to normal brain tissues. Bioinformatic analysis revealed the presence of CpG islands in the promoter of AJAP1.Methylation analysis of the AJAP1 promoter identified hypermethylation in 21% of oligodendrogliomas (n = 27), and the degree of methylation correlated with Iow levels of AJAP1 expression (P = 0.045). The AJAP1 promoter was also highly methylated in a wide spectrum of cell lines (n = 22), including cell lines of glioblastoma. Analysis of the National Cancer Institute's REMBRANDT dataset, which contains 343 glioma samples, indicated that Iow AJAP1 gene expression was associated with decreased survival. Thus,both genetic (gene deletion) and epigenetic alterations (promoter methylation) are likely mechanisms that inactivate the putative tumor suppressor AJAP1 in gliomas, which contributes to poor prognosis.

  14. A KH Domain-Containing Putative RNA-Binding Protein Is Critical for Heat Stress-Responsive Gene Regulation and Thermotolerance in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Qingmei Guan; Changlong Wen; Haitao Zeng; Jianhua Zhu

    2013-01-01

    Heat stress is a severe environmental factor that significantly reduces plant growth and delays development.Heat stress factors (HSFs) are a class of transcription factors that are synthesized rapidly in response to elevations in temperature and are responsible for the transcription of many heat stress-responsive genes including those encoding heat shock proteins (HSPs).There are 21 HSFs in Arabidopsis,and recent studies have established that the HSFA1 family members are master regulators for the remaining HSFs.However,very little is known about upstream molecular factors that control the expression of HSFA1 genes and other HSF genes under heat stress.Through a forward genetic analysis,we identified RCF3,a K homology (KH) domain-containing nuclear-localized putative RNA-binding protein.RCF3 is a negative regulator of most HSFs,including HSFAla,HSFAlb,and HSFAld.In contrast,RCF3 positively controls the expression of HSFAle,HSFA3,HSFA9,HSFB3,and DREB2C.Consistently with the overall increased accumulation of heat-responsive genes,the rcf3 mutant plants are more tolerant than the wild-type to heat stress.Together,our results suggest that a KH domain-containing putative RNA-binding protein RCF3 is an important upstream regulator for heat stress-responsive gene expression and thermotolerance in Arabidopsis.

  15. Tumor-associated methylation of the putative tumor suppressor AJAP1 gene and association between decreased AJAP1 expression and shorter survival in patients with glioma.

    Science.gov (United States)

    Cogdell, David; Chung, Woonbok; Liu, Yuexin; McDonald, J Matthew; Aldape, Kenneth; Issa, Jean-Pierre J; Fuller, Gregory N; Zhang, Wei

    2011-04-01

    Allelic loss of the short arm of chromosome 1 has been observed frequently in a wide spectrum of cancers, most frequently in oligodendroglioma. In our previous studies, we evaluated 177 oligodendroglial tumor samples and identified the AJAP1 gene (formerly Shrew1) in the consensus region of deletion. AJAP1 is a transmembrane protein found in adheren junctions and functions to inhibit glioma cell adhesion and migration. Whereas a putative tumor suppressor gene, we did not detect AJAP1 gene mutations. In subsequent studies, we found that AJAP1 was underexpressed in oligodendrogliomas relative to normal brain tissues. Bioinformatic analysis revealed the presence of CpG islands in the promoter of AJAP1. Methylation analysis of the AJAP1 promoter identified hypermethylation in 21% of oligodendrogliomas (n =27), and the degree of methylation correlated with low levels of AJAP1 expression (P = 0.045). The AJAP1 promoter was also highly methylated in a wide spectrum of cell lines (n = 22), including cell lines of glioblastoma. Analysis of the National Cancer Institute's REMBRANDT dataset, which contains 343 glioma samples, indicated that low AJAP1 gene expression was associated with decreased survival. Thus, both genetic (gene deletion) and epigenetic alterations (promoter methylation) are likely mechanisms that inactivate the putative tumor suppressor AJAP1 in gliomas, which contributes to poor prognosis.

  16. Subgroup C avian metapneumovirus (MPV) and the recently isolated human MPV exhibit a common organization but have extensive sequence divergence in their putative SH and G genes.

    Science.gov (United States)

    Toquin, D; de Boisseson, C; Beven, V; Senne, D A; Eterradossi, N

    2003-08-01

    The genes encoding the putative small hydrophobic (SH), attachment (G) and polymerase (L) proteins of the Colorado isolate of subgroup C avian pneumovirus (APV) were entirely or partially sequenced. They all included metapneumovirus (MPV)-like gene start and gene end sequences. The deduced Colorado SH protein shared 26.9 and 21.7 % aa identity with its counterpart in human MPV (hMPV) and APV subgroup A, respectively, but its only significant aa similarities were to hMPV. Conserved features included a common hydrophobicity profile with an unique transmembrane domain and the conservation of most extracellular cysteine residues. The Colorado putative G gene encoded several ORFs, the longer of which encoded a 252 aa long type II glycoprotein with aa similarities to hMPV G only (20.6 % overall aa identity with seven conserved N-terminal residues). The putative Colorado G protein shared, at best, 21.0 % aa identity with its counterparts in the other APV subgroups and did not contain the extracellular cysteine residues and short aa stretch highly conserved in other APVs. The N-terminal end of the Colorado L protein exhibited 73.6 and 54.9 % aa identity with hMPV and APV subgroup A, respectively, with four aa blocks highly conserved among Pneumovirus: Phylogenetic analysis performed on the nt sequences confirmed that the L sequences from MPVs were genetically related, whereas analysis of the G sequences revealed that among MPVs, only APV subgroups A, B and D clustered together, independently of both the Colorado isolate and hMPV, which shared weak genetic relatedness at the G gene level.

  17. A temperature-sensitive allele of a putative mRNA splicing helicase down-regulates many cell wall genes and causes radial swelling in Arabidopsis thaliana.

    Science.gov (United States)

    Howles, Paul A; Gebbie, Leigh K; Collings, David A; Varsani, Arvind; Broad, Ronan C; Ohms, Stephen; Birch, Rosemary J; Cork, Ann H; Arioli, Tony; Williamson, Richard E

    2016-05-01

    The putative RNA helicase encoded by the Arabidopsis gene At1g32490 is a homolog of the yeast splicing RNA helicases Prp2 and Prp22. We isolated a temperature-sensitive allele (rsw12) of the gene in a screen for root radial swelling mutants. Plants containing this allele grown at the restrictive temperature showed weak radial swelling, were stunted with reduced root elongation, and contained reduced levels of cellulose. The role of the protein was further explored by microarray analysis. By using both fold change cutoffs and a weighted gene coexpression network analysis (WGCNA) to investigate coexpression of genes, we found that the radial swelling phenotype was not linked to genes usually associated with primary cell wall biosynthesis. Instead, the mutation has strong effects on expression of secondary cell wall related genes. Many genes potentially associated with secondary walls were present in the most significant WGCNA module, as were genes coding for arabinogalactans and proteins with GPI anchors. The proportion of up-regulated genes that possess introns in rsw12 was above that expected if splicing was unrelated to the activity of the RNA helicase, suggesting that the helicase does indeed play a role in splicing in Arabidopsis. The phenotype may be due to a change in the expression of one or more genes coding for cell wall proteins.

  18. Isolation and characterization of human patched 2 (PTCH2), a putative tumour suppressor gene inbasal cell carcinoma and medulloblastoma on chromosome 1p32.

    Science.gov (United States)

    Smyth, I; Narang, M A; Evans, T; Heimann, C; Nakamura, Y; Chenevix-Trench, G; Pietsch, T; Wicking, C; Wainwright, B J

    1999-02-01

    Mutations of the human Patched gene ( PTCH ) have been identified in individuals with the nevoid basal cell carcinoma syndrome (NBCCS) as well as in sporadic basal cell carcinomas and medulloblastomas. We have isolated a homologue of this tumour suppressor gene and localized it to the short arm of chromosome 1 (1p32.1-32.3). Patched 2 ( PTCH2 ) comprises 22 coding exons and spans approximately 15 kb of genomic DNA. The gene encodes a 1203 amino acid putative transmembrane protein which is highly homologous to the PTCH product. We have characterized the genomic structure of PTCH2 and have used single-stranded conformational polymorphism analysis to search for mutations in PTCH2 in NBCCS patients, basal cell carcinomas and in medulloblastomas. To date, we have identified one truncating mutation in a medulloblastoma and a change in a splice donor site in a basal cell carcinoma, suggesting that the gene plays a role in the development of some tumours.

  19. Validation of putative reference genes for qRT-PCR normalization in tissues and blood from pigs infected with Actinobacillus pleuropneumoniae

    DEFF Research Database (Denmark)

    Skovgaard, Kerstin; Mortensen, Shila; Poulsen, K.T.;

    2007-01-01

    The quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR) is a sensitive and very efficient technique for quantification of gene expression. However, qRT-PCR relies on accurate normalization of gene expression data, as RNA recovery and cDNA synthesis efficiency might vary...... from sample to sample. In the present study, six putative reference genes were validated for normalization of gene expression in three different tissues and in white blood cells from pigs experimentally infected with the common respiratory pathogen Actinobacillus pleuropneumoniae. Two dedicated...... (GAPDH). IL-6 expression was quantified in white blood cells, liver, lymph nodes and tonsils from 10 infected pigs and 5 control pigs. After normalization using either geNorm or Normfinder IL-6 was shown to be significantly up-regulated (P

  20. Characterization of BcMF23a and BcMF23b, two putative pectin methylesterase genes related to pollen development in Brassica campestris ssp. chinensis.

    Science.gov (United States)

    Lin, Sue; Huang, Li; Yu, Xiaolin; Xiong, Xingpeng; Yue, Xiaoyan; Liu, Tingting; Liang, Ying; Lv, Meiling; Cao, Jiashu

    2017-02-01

    Two homologous genes, Brassica campestris Male Fertility 23a (BcMF23a) and Brassica campestris Male Fertility 23b (BcMF23b), encoding putative pectin methylesterases (PMEs) were isolated from Brassica campestris ssp. chinensis (syn. Brassica rapa ssp. chinensis). These two genes sharing high sequence identity with each other were highly expressed in the fertile flower buds but silenced in the sterile ones of genic male sterile line system ('Bcajh97-01A/B'). Results of RT-PCR and in situ hybridization suggested that BcMF23a and BcMF23b were pollen-expressed genes, whose transcripts were first detected at the binucleate pollen and maintained throughout to the mature pollen grains. Western blot indicated that both of the putative BcMF23a and BcMF23b proteins are approximately 40 kDa, which exhibited extracellular localization revealed by transient expression analysis in the onion epidermal cells. The promoter of BcMF23a was active specifically in pollen during the late pollen developmental stages, while, in addition to the pollen, BcMF23b promoter drove an extra gene expression in the valve margins, abscission layer at the base of the first true leaves, taproot and lateral roots in seedlings.

  1. Genomic structure of the rat major AP endonuclease gene (Apex with an adjacent putative O-sialoglycoprotease gene (Prsmg1/Gcpl1 and a processed Apex pseudogene (Apexp1.

    Directory of Open Access Journals (Sweden)

    Yao,Ming

    1999-12-01

    Full Text Available Genomic sequencing and chromosomal assignment of the gene encoding rat APEX nuclease, a multifunctional DNA repair enzyme, were performed. An active Apex gene and a processed pseudogene were isolated from a rat genomic library. The active Apex gene consists of 5 exons and 4 introns spanning 2.1 kb. The putative promoter region of the Apex gene lacks the typical TATA box, but contains CAAT boxes and a CpG island having putative binding sites for several transcription factors, such as Sp1, AP-2, GATA-1 and ATF. A putative O-sialoglycoprotease (a homologue of Pasteurella haemolytica glycoprotease, gcp; abbreviated as Prsmg1/Gcpl1 gene consisting of 11 exons and 10 introns spanning 7.3 kb lies immediately adjacent to the Apex gene in a 5'-to-5' orientation. The Apex gene locus was mapped to rat chromosome 15p12 using in situ hybridization. The processed pseudogene (designated as rat Apexp1 has a nucleotide sequence 87.1% identical to that of the rat Apex cDNA, although several stop codons interrupting the coding sequences and multiple nucleotide deletions were observed. The Apexp1 is located in an inactive LINE sequence. Calculation of nucleotide substitution rates suggests that the immediate, active progenitor of Apexp1 arose 23 million years ago and that the non-functionalization occurred 15 million years ago.

  2. Differential expression patterns in chemosensory and non-chemosensory tissues of putative chemosensory genes identified by transcriptome analysis of insect pest the purple stem borer Sesamia inferens (Walker.

    Directory of Open Access Journals (Sweden)

    Ya-Nan Zhang

    Full Text Available BACKGROUND: A large number of insect chemosensory genes from different gene subfamilies have been identified and annotated, but their functional diversity and complexity are largely unknown. A systemic examination of expression patterns in chemosensory organs could provide important information. METHODOLOGY/PRINCIPAL FINDINGS: We identified 92 putative chemosensory genes by analysing the transcriptome of the antennae and female sex pheromone gland of the purple stem borer Sesamia inferens, among them 87 are novel in this species, including 24 transcripts encoding for odorant binding proteins (OBPs, 24 for chemosensory proteins (CSPs, 2 for sensory neuron membrane proteins (SNMPs, 39 for odorant receptors (ORs and 3 for ionotropic receptors (IRs. The transcriptome analyses were validated and quantified with a detailed global expression profiling by Reverse Transcription-PCR for all 92 transcripts and by Quantitative Real Time RT-PCR for selected 16 ones. Among the chemosensory gene subfamilies, CSP transcripts are most widely and evenly expressed in different tissues and stages, OBP transcripts showed a clear antenna bias and most of OR transcripts are only detected in adult antennae. Our results also revealed that some OR transcripts, such as the transcripts of SNMP2 and 2 IRs were expressed in non-chemosensory tissues, and some CSP transcripts were antenna-biased expression. Furthermore, no chemosensory transcript is specific to female sex pheromone gland and very few are found in the heads. CONCLUSION: Our study revealed that there are a large number of chemosensory genes expressed in S. inferens, and some of them displayed unusual expression profile in non-chemosensory tissues. The identification of a large set of putative chemosensory genes of each subfamily from a single insect species, together with their different expression profiles provide further information in understanding the functions of these chemosensory genes in S. inferens as

  3. Mutations in WSC genes for putative stress receptors result in sensitivity to multiple stress conditions and impairment of Rlm1-dependent gene expression in Saccharomyces cerevisiae.

    Science.gov (United States)

    Zu, T; Verna, J; Ballester, R

    2001-09-01

    Intracellular signaling by mitogen-activated protein (MAP) kinase cascades plays an essential role in the cellular response to environmental stress. In the yeast Saccharomyces cerevisiae, the PKC1-regulated, stress-activated MAP kinase pathway, the MPK1 cascade, is activated by heat and by a decrease in osmolarity. The genes WSC1, WSC2 and WSC3 encode putative receptors that maintain cell wall integrity under conditions of heat stress. Genetic studies place the function of the WSC genes upstream of the MPK1 kinase cascade. To further define the role of the WSC family in the stress response we determined whether: (1) the wscdelta mutants are sensitive to other environmental stress conditions, in addition to heat shock; (2) expression from four transcriptional control elements, known to be activated by stress, is impaired in wscdelta mutants; and (3) Wsc4, a Wsc homolog, has functions that overlap with those of the other Wsc family members. We report here that deletion of WSC and PKC1 causes hypersensitivity to ethanol, hydrogen peroxide and DNA-damaging drugs. In wscdelta mutants expression of beta-galactosidase from the AP-1 response element (ARE), the heat shock response element (HSE) or the stress response element (STRE) is not reduced. In contrast, expression of a reporter gene placed under the control of the Rlm1 (transcription factor)-dependent response element is significantly reduced in wscdelta mutants. This suggests that the lysis defect of wscdelta mutants is at least in part caused by a defect in transcriptional regulation by Rlm1. Phenotypic analysis of the effect of deleting WSC4 in a wsc1delta mutant show that, unlike WSC2 or WSC3, deletion of WSC4 does not exacerbate the lysis defect of a wsc1delta strain. In contrast, deletion of WSC4 enhances the sensitivity of the wsc1delta mutant to heat shock, ethanol, and a DNA-damaging drug, suggesting that WSC4 plays a role in the response to environmental stress but that its function may differ from those of

  4. Comparison of 454-ESTs from Huperzia serrata and Phlegmariurus carinatus reveals putative genes involved in lycopodium alkaloid biosynthesis and developmental regulation

    Directory of Open Access Journals (Sweden)

    Steinmetz André

    2010-09-01

    Full Text Available Abstract Background Plants of the Huperziaceae family, which comprise the two genera Huperzia and Phlegmariurus, produce various types of lycopodium alkaloids that are used to treat a number of human ailments, such as contusions, swellings and strains. Huperzine A, which belongs to the lycodine type of lycopodium alkaloids, has been used as an anti-Alzheimer's disease drug candidate. Despite their medical importance, little genomic or transcriptomic data are available for the members of this family. We used massive parallel pyrosequencing on the Roche 454-GS FLX Titanium platform to generate a substantial EST dataset for Huperzia serrata (H. serrata and Phlegmariurus carinatus (P. carinatus as representative members of the Huperzia and Phlegmariurus genera, respectively. H. serrata and P. carinatus are important plants for research on the biosynthesis of lycopodium alkaloids. We focused on gene discovery in the areas of bioactive compound biosynthesis and transcriptional regulation as well as genetic marker detection in these species. Results For H. serrata, 36,763 unique putative transcripts were generated from 140,930 reads totaling over 57,028,559 base pairs; for P. carinatus, 31,812 unique putative transcripts were generated from 79,920 reads totaling over 30,498,684 base pairs. Using BLASTX searches of public databases, 16,274 (44.3% unique putative transcripts from H. serrata and 14,070 (44.2% from P. carinatus were assigned to at least one protein. Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (KEGG orthology annotations revealed that the functions of the unique putative transcripts from these two species cover a similarly broad set of molecular functions, biological processes and biochemical pathways. In particular, a total of 20 H. serrata candidate cytochrome P450 genes, which are more abundant in leaves than in roots and might be involved in lycopodium alkaloid biosynthesis, were found based on the comparison of H

  5. A novel role for the immunophilin FKBP52 in motor coordination.

    Science.gov (United States)

    Young, Matthew J; Geiszler, Philippine C; Pardon, Marie-Christine

    2016-10-15

    FKBP52 is a ubiquitously distributed immunophilin that has been associated with wide-ranging functions in cell signalling as well as hormonal and stress responses. Amongst other pathways, it acts via complex-formation with corticosteroid receptors and has consequently been associated with stress- and age- related neurodegenerative disorders including Alzheimer's and Parkinson's diseases. Reduced levels of FKBP52 have been linked to tau dysfunction and amyloid beta toxicity in AD. However, FKBP52's role in cognition and neurodegenerative disorder-like phenotypes remain to be elucidated. The present study aimed therefore at investigating the cognitive and behavioural effects of reduced FKBP52 levels of genetically modified mice during ageing. Female and male FKBP52(+/+), FKBP52(+/-) and FKBP52(-/-) mice were compared at two-, ten-, twelve-, fifteen- and eighteen-months-of-age in a series of behavioural tests covering specie-specific behaviour, motor activity and coordination, fear-, spatial and recognition memory as well as curiosity and emotionality. Whilst cognitively unimpaired, FKBP52(+/-) mice performed worse on an accelerating rotating rod than FKBP52(+/+) littermates across all age-groups suggesting that FKBP52 is involved in processes controlling motor coordination. This deficit did not exacerbate with age but did worsen with repeated testing; pointing towards a role for FKBP52 in learning of tasks requiring motor coordination abilities. This study contributes to the knowledge base of FKBP52's implication in neurodegenerative diseases by demonstrating that FKBP52 by itself does not directly affect cognition and may therefore rather play an indirect, modulatory role in the functional pathology of AD, whereas it directly affects motor coordination, an early sign of neurodegenerative damages to the brain.

  6. Putative PPAR target genes express highly in skeletal muscle of insulin-resistant MetS model SHR/NDmc-cp rats.

    Science.gov (United States)

    Hariya, Natsuyo; Miyake, Kunio; Kubota, Takeo; Goda, Toshinao; Mochizuki, Kazuki

    2015-01-01

    It is known that insulin resistance in skeletal muscle induces subsequent metabolic diseases such as metabolic syndrome (MetS). However, which genes are altered in the skeletal muscle by development of insulin resistance in animal models has not been examined. In this study, we performed microarray and subsequent real-time RT-PCR analyses using total RNA extracted from the gastrocnemius muscle of the MetS model, spontaneously hypertensive corpulent congenic (SHR/NDmc-cp) rats, and control Wistar Kyoto (WKY) rats. SHR/NDmc-cp rats displayed overt insulin resistance relative to WKY rats. The expression of many genes related to fatty acid oxidation was higher in SHR/NDmc-cp rats than in WKY rats. Among 18 upregulated genes, putative peroxisome proliferator responsive elements were found in the upstream region of 15 genes. The protein expression of ACOX2, an upregulated gene, and peroxisome proliferator-activated receptor (PPAR) G1, but not of PPARG2, PPARA or PPARD, was higher in the gastrocnemius muscle of SHR/NDmc-cp rats than that in WKY rats. These results suggest that insulin resistance in the MetS model, SHR/NDmc-cp rats, is positively associated with the expression of fatty acid oxidation-related genes, which are presumably PPARs’ targets, in skeletal muscle.

  7. Structural basis of nucleic acid recognition by FK506-binding protein 25 (FKBP25), a nuclear immunophilin.

    Science.gov (United States)

    Prakash, Ajit; Shin, Joon; Rajan, Sreekanth; Yoon, Ho Sup

    2016-04-07

    The nuclear immunophilin FKBP25 interacts with chromatin-related proteins and transcription factors and is suggested to interact with nucleic acids. Currently the structural basis of nucleic acid binding by FKBP25 is unknown. Here we determined the nuclear magnetic resonance (NMR) solution structure of full-length human FKBP25 and studied its interaction with DNA. The FKBP25 structure revealed that the N-terminal helix-loop-helix (HLH) domain and C-terminal FK506-binding domain (FKBD) interact with each other and that both of the domains are involved in DNA binding. The HLH domain forms major-groove interactions and the basic FKBD loop cooperates to form interactions with an adjacent minor-groove of DNA. The FKBP25-DNA complex model, supported by NMR and mutational studies, provides structural and mechanistic insights into the nuclear immunophilin-mediated nucleic acid recognition. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  8. Control of Alzheimer's amyloid beta toxicity by the high molecular weight immunophilin FKBP52 and copper homeostasis in Drosophila.

    Directory of Open Access Journals (Sweden)

    Reiko Sanokawa-Akakura

    Full Text Available FK506 binding proteins (FKBPs, also called immunophilins, are prolyl-isomerases (PPIases that participate in a wide variety of cellular functions including hormone signaling and protein folding. Recent studies indicate that proteins that contain PPIase activity can also alter the processing of Alzheimer's Amyloid Precursor Protein (APP. Originally identified in hematopoietic cells, FKBP52 is much more abundantly expressed in neurons, including the hippocampus, frontal cortex, and basal ganglia. Given the fact that the high molecular weight immunophilin FKBP52 is highly expressed in CNS regions susceptible to Alzheimer's, we investigated its role in Abeta toxicity. Towards this goal, we generated Abeta transgenic Drosophila that harbor gain of function or loss of function mutations of FKBP52. FKBP52 overexpression reduced the toxicity of Abeta and increased lifespan in Abeta flies, whereas loss of function of FKBP52 exacerbated these Abeta phenotypes. Interestingly, the Abeta pathology was enhanced by mutations in the copper transporters Atox1, which interacts with FKBP52, and Ctr1A and was suppressed in FKBP52 mutant flies raised on a copper chelator diet. Using mammalian cultures, we show that FKBP52 (-/- cells have increased intracellular copper and higher levels of Abeta. This effect is reversed by reconstitution of FKBP52. Finally, we also found that FKBP52 formed stable complexes with APP through its FK506 interacting domain. Taken together, these studies identify a novel role for FKBP52 in modulating toxicity of Abeta peptides.

  9. Whole-transcriptome survey of the putative ATP-binding cassette (ABC) transporter family genes in the latex-producing laticifers of Hevea brasiliensis.

    Science.gov (United States)

    Zhiyi, Nie; Guijuan, Kang; Yu, Li; Longjun, Dai; Rizhong, Zeng

    2015-01-01

    The ATP-binding cassette (ABC) proteins or transporters constitute a large protein family in plants and are involved in many different cellular functions and processes, including solute transportation, channel regulation and molecular switches, etc. Through transcriptome sequencing, a transcriptome-wide survey and expression analysis of the ABC protein genes were carried out using the laticiferous latex from Hevea brasiliensis (rubber tree). A total of 46 putative ABC family proteins were identified in the H. brasiliensis latex. These consisted of 12 'full-size', 21 'half-size' and 13 other putative ABC proteins, and all of them showed strong conservation with their Arabidopsis thaliana counterparts. This study indicated that all eight plant ABC protein paralog subfamilies were identified in the H. brasiliensis latex, of which ABCB, ABCG and ABCI were the most abundant. Real-time quantitative reverse transcription-polymerase chain reaction assays demonstrated that gene expression of several latex ABC proteins was regulated by ethylene, jasmonic acid or bark tapping (a wound stress) stimulation, and that HbABCB15, HbABCB19, HbABCD1 and HbABCG21 responded most significantly of all to the abiotic stresses. The identification and expression analysis of the latex ABC family proteins could facilitate further investigation into their physiological involvement in latex metabolism and rubber biosynthesis by H. brasiliensis.

  10. Overexpression, crystallization and preliminary X-ray crystallographic analysis of a putative transposase from Thermoplasma acidophilum encoded by the Ta0474 gene

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Ji Yong; Lee, Hyung Ho; Kim, Do Jin; Han, Sang Hee; Kim, Olesya; Kim, Hyoun Sook; Lee, Sang Jae; Suh, Se Won, E-mail: sewonsuh@snu.ac.kr [Department of Chemistry, College of Natural Sciences, Seoul National University, Seoul 151-742 (Korea, Republic of)

    2006-11-01

    A putative transposase from T. acidophilum encoded by the Ta0474 gene was crystallized and X-ray diffraction data were collected to 1.78 Å. IS200 transposases, originally identified in Salmonella typhimurium LT2, are present in many bacteria and archaea and are distinct from other groups of transposases. To facilitate further structural comparisons among IS200-like transposases, structural analysis has been initiated of a putative transposase from Thermoplasma acidophilum encoded by the Ta0474 gene. Its 137-residue polypeptide shows high levels of sequence similarity to other members of the IS200 transposase family. The protein was overexpressed in intact form in Escherichia coli and crystallized at 297 K using a reservoir solution consisting of 100 mM Na HEPES pH 7.5 and 20%(v/v) ethanol. X-ray diffraction data were collected to 1.78 Å. The crystals belong to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 65.00, b = 34.07, c = 121.58 Å, α = 90, β = 100.20, γ = 90°. Four monomers, representing two copies of a dimeric molecule, are present in the asymmetric unit, giving a crystal volume per protein weight (V{sub M}) of 2.02 Å{sup 3} Da{sup −1} and a solvent content of 39.2%.

  11. Whole-transcriptome survey of the putative ATP-binding cassette (ABC transporter family genes in the latex-producing laticifers of Hevea brasiliensis.

    Directory of Open Access Journals (Sweden)

    Nie Zhiyi

    Full Text Available The ATP-binding cassette (ABC proteins or transporters constitute a large protein family in plants and are involved in many different cellular functions and processes, including solute transportation, channel regulation and molecular switches, etc. Through transcriptome sequencing, a transcriptome-wide survey and expression analysis of the ABC protein genes were carried out using the laticiferous latex from Hevea brasiliensis (rubber tree. A total of 46 putative ABC family proteins were identified in the H. brasiliensis latex. These consisted of 12 'full-size', 21 'half-size' and 13 other putative ABC proteins, and all of them showed strong conservation with their Arabidopsis thaliana counterparts. This study indicated that all eight plant ABC protein paralog subfamilies were identified in the H. brasiliensis latex, of which ABCB, ABCG and ABCI were the most abundant. Real-time quantitative reverse transcription-polymerase chain reaction assays demonstrated that gene expression of several latex ABC proteins was regulated by ethylene, jasmonic acid or bark tapping (a wound stress stimulation, and that HbABCB15, HbABCB19, HbABCD1 and HbABCG21 responded most significantly of all to the abiotic stresses. The identification and expression analysis of the latex ABC family proteins could facilitate further investigation into their physiological involvement in latex metabolism and rubber biosynthesis by H. brasiliensis.

  12. Glucocorticoids regulate the expression of the mouse urocortin II gene: a putative connection between the corticotropin-releasing factor receptor pathways.

    Science.gov (United States)

    Chen, Alon; Vaughan, Joan; Vale, Wylie W

    2003-08-01

    Peptides encoded by the urocortin II (Ucn II) gene were recently identified as new members of the corticotropin-releasing factor (CRF) family. Ucn II is a specific ligand for the type 2 CRF receptor. Using RT-PCR, DNA sequencing, and immunofluorescence staining, we report the expression of Ucn II mRNA in several human and mouse (m) neuronal cell lines. Using these neuronal cell lines, we provide evidence that exposure to glucocorticoid hormones increases mUcn II mRNA expression and promoter activation. The effect of glucocorticoids on mUcn II mRNA expression was tested in the Ucn II/glucocorticoid receptor-positive cell line NG108-15. The results demonstrate that mUcn II mRNA expression is up-regulated by dexamethasone in a dose- and time-dependent fashion. Computer analysis revealed the presence of 14 putative half-palindrome glucocorticoid response element sequences within 1.2 kb of the mUcn II 5' flanking region. Transfections with different fragments of the 5'-flanking region of the mUcn II gene fused to a luciferase reporter gene showed a promoter-dependent expression of the reporter gene and regulation by dexamethasone. Promoter deletion studies clarify the sufficient putative glucocorticoid response element site mediating this effect. The steroid hormone antagonist RU486 blocked the effect of dexamethasone on mUcn II mRNA expression and promoter activation, suggesting a direct glucocorticoid receptor-mediated effect of dexamethasone on mUcn II mRNA expression. Ucn II is expressed in vivo in the hypothalamus, brainstem, olfactory bulb, and pituitary. Low levels were also detected in the mouse cortex, hippocampus, and spinal cord. We demonstrated that mUcn II gene transcription was stimulated by glucocorticoid administration in vivo and inhibited by removal of glucocorticoids by adrenalectomy. Administration of dexamethasone to mice resulted in an increase of mUcn II levels in the hypothalamus and brainstem but not in the olfactory bulb region 12 h following

  13. Characterization and variation of a human inwardly-rectifying-K-channel gene (KCNJ6): a putative ATP-sensitive K-channel subunit.

    Science.gov (United States)

    Sakura, H; Bond, C; Warren-Perry, M; Horsley, S; Kearney, L; Tucker, S; Adelman, J; Turner, R; Ashcroft, F M

    1995-06-26

    The ATP-sensitive K-channel plays a central role in insulin release from pancreatic beta-cells. We report here the cloning of the gene (KCNJ6) encoding a putative subunit of a human ATP-sensitive K-channel expressed in brain and beta-cells, and characterisation of its exon-intron structure. Screening of a somatic cell mapping panel and fluorescent in situ hybridization place the gene on chromosome 21 (21q22.1-22.2). Analysis of single-stranded conformational polymorphisms revealed the presence of two silent polymorphisms (Pro-149: CCG-CCA and Asp-328: GAC-GAT) with similar frequencies in normal and non-insulin-dependent diabetic patients.

  14. Molecular identification of aiiA homologous gene from endophytic Enterobacter species and in silico analysis of putative tertiary structure of AHL-lactonase.

    Science.gov (United States)

    Rajesh, P S; Rai, V Ravishankar

    2014-01-03

    The aiiA homologous gene known to encode AHL- lactonase enzyme which hydrolyze the N-acylhomoserine lactone (AHL) quorum sensing signaling molecules produced by Gram negative bacteria. In this study, the degradation of AHL molecules was determined by cell-free lysate of endophytic Enterobacter species. The percentage of quorum quenching was confirmed and quantified by HPLC method (pEnterobacter asburiae VT65, Enterobacter aerogenes VT66 and Enterobacter ludwigii VT70 strains. Sequence alignment analysis revealed the presence of two zinc binding sites, "HXHXDH" motif as well as tyrosine residue at the position 194. Based on known template available at Swiss-Model, putative tertiary structure of AHL-lactonase was constructed. The result showed that novel endophytic strains of Enterobacter genera encode the novel aiiA homologous gene and its structural importance for future study.

  15. Putative recombination events and evolutionary history of five economically important viruses of fruit trees based on coat protein-encoding gene sequence analysis.

    Science.gov (United States)

    Boulila, Moncef

    2010-06-01

    To enhance the knowledge of recombination as an evolutionary process, 267 accessions retrieved from GenBank were investigated, all belonging to five economically important viruses infecting fruit crops (Plum pox, Apple chlorotic leaf spot, Apple mosaic, Prune dwarf, and Prunus necrotic ringspot viruses). Putative recombinational events were detected in the coat protein (CP)-encoding gene using RECCO and RDP version 3.31beta algorithms. Based on RECCO results, all five viruses were shown to contain potential recombination signals in the CP gene. Reconstructed trees with modified topologies were proposed. Furthermore, RECCO performed better than the RDP package in detecting recombination events and exhibiting their evolution rate along the sequences of the five viruses. RDP, however, provided the possible major and minor parents of the recombinants. Thus, the two methods should be considered complementary.

  16. Lactobacillus plantarum gene clusters encoding putative cell-surface protein complexes for carbohydrate utilization are conserved in specific gram-positive bacteria

    Directory of Open Access Journals (Sweden)

    Muscariello Lidia

    2006-05-01

    Full Text Available Abstract Background Genomes of gram-positive bacteria encode many putative cell-surface proteins, of which the majority has no known function. From the rapidly increasing number of available genome sequences it has become apparent that many cell-surface proteins are conserved, and frequently encoded in gene clusters or operons, suggesting common functions, and interactions of multiple components. Results A novel gene cluster encoding exclusively cell-surface proteins was identified, which is conserved in a subgroup of gram-positive bacteria. Each gene cluster generally has one copy of four new gene families called cscA, cscB, cscC and cscD. Clusters encoding these cell-surface proteins were found only in complete genomes of Lactobacillus plantarum, Lactobacillus sakei, Enterococcus faecalis, Listeria innocua, Listeria monocytogenes, Lactococcus lactis ssp lactis and Bacillus cereus and in incomplete genomes of L. lactis ssp cremoris, Lactobacillus casei, Enterococcus faecium, Pediococcus pentosaceus, Lactobacillius brevis, Oenococcus oeni, Leuconostoc mesenteroides, and Bacillus thuringiensis. These genes are neither present in the genomes of streptococci, staphylococci and clostridia, nor in the Lactobacillus acidophilus group, suggesting a niche-specific distribution, possibly relating to association with plants. All encoded proteins have a signal peptide for secretion by the Sec-dependent pathway, while some have cell-surface anchors, novel WxL domains, and putative domains for sugar binding and degradation. Transcriptome analysis in L. plantarum shows that the cscA-D genes are co-expressed, supporting their operon organization. Many gene clusters are significantly up-regulated in a glucose-grown, ccpA-mutant derivative of L. plantarum, suggesting catabolite control. This is supported by the presence of predicted CRE-sites upstream or inside the up-regulated cscA-D gene clusters. Conclusion We propose that the CscA, CscB, CscC and Csc

  17. Validation of putative reference genes for normalization of Q-RT-PCR data from paraffin-embedded lymphoid tissue

    DEFF Research Database (Denmark)

    Green, Tina Marie; de Stricker, Karin; Møller, Michael Boe

    2009-01-01

    , represented by non-neoplastic lymph nodes and diffuse large B-cell lymphomas, by using 2 statistical software applications, geNorm and NormFinder. In addition, we wanted to validate the usefulness of paraffin-embedded samples for Q-RT-PCR studies by investigating gene expressions of relevant target genes...

  18. Clustering of two genes putatively involved in cyanate detoxification evolved recently and independently in multiple fungal lineages

    Science.gov (United States)

    Fungi that have the enzymes cyanase and carbonic anhydrase show a limited capacity to detoxify cyanate, a fungicide employed by both plants and humans. Here, we describe a novel two-gene cluster that comprises duplicated cyanase and carbonic anhydrase copies, which we name the CCA gene cluster, trac...

  19. The wheat homolog of putative nucleotide-binding site-leucine-rich repeat resistance gene TaRGA contributes to resistance against powdery mildew.

    Science.gov (United States)

    Wang, Defu; Wang, Xiaobing; Mei, Yu; Dong, Hansong

    2016-03-01

    Powdery mildew, one of the most destructive wheat diseases worldwide, is caused by Blumeria graminis f. sp. tritici (Bgt), a fungal species with a consistently high mutation rate that makes individual resistance (R) genes ineffective. Therefore, effective resistance-related gene cloning is vital for breeding and studying the resistance mechanisms of the disease. In this study, a putative nucleotide-binding site-leucine-rich repeat (NBS-LRR) R gene (TaRGA) was cloned using a homology-based cloning strategy and analyzed for its effect on powdery mildew disease and wheat defense responses. Real-time reverse transcription-PCR (RT-PCR) analyses revealed that a Bgt isolate 15 and salicylic acid stimulation significantly induced TaRGA in the resistant variety. Furthermore, the silencing of TaRGA in powdery mildew-resistant plants increased susceptibility to Bgt15 and prompted conidia propagation at the infection site. However, the expression of TaRGA in leaf segments after single-cell transient expression assay highly increased the defense responses to Bgt15 by enhancing callose deposition and phenolic autofluorogen accumulation at the pathogen invading sites. Meanwhile, the expression of pathogenesis-related genes decreased in the TaRGA-silenced plants and increased in the TaRGA-transient-overexpressing leaf segments. These results implied that the TaRGA gene positively regulates the defense response to powdery mildew disease in wheat.

  20. Mating type gene homologues and putative sex pheromone-sensing pathway in arbuscular mycorrhizal fungi, a presumably asexual plant root symbiont.

    Science.gov (United States)

    Halary, Sébastien; Daubois, Laurence; Terrat, Yves; Ellenberger, Sabrina; Wöstemeyer, Johannes; Hijri, Mohamed

    2013-01-01

    The fungal kingdom displays a fascinating diversity of sex-determination systems. Recent advances in genomics provide insights into the molecular mechanisms of sex, mating type determination, and evolution of sexual reproduction in many fungal species in both ancient and modern phylogenetic lineages. All major fungal groups have evolved sexual differentiation and recombination pathways. However, sexuality is unknown in arbuscular mycorrhizal fungi (AMF) of the phylum Glomeromycota, an ecologically vital group of obligate plant root symbionts. AMF are commonly considered an ancient asexual lineage dating back to the Ordovician, approximately 460 M years ago. In this study, we used genomic and transcriptomic surveys of several AMF species to demonstrate the presence of conserved putative sex pheromone-sensing mitogen-activated protein (MAP) kinases, comparable to those described in Ascomycota and Basidiomycota. We also find genes for high mobility group (HMG) transcription factors, homologous to SexM and SexP genes in the Mucorales. The SexM genes show a remarkable sequence diversity among multiple copies in the genome, while only a single SexP sequence was detected in some isolates of Rhizophagus irregularis. In the Mucorales and Microsporidia, the sexM gene is flanked by genes for a triosephosphate transporter (TPT) and a RNA helicase, but we find no evidence for synteny in the vicinity of the Sex locus in AMF. Nonetheless, our results, together with previous observations on meiotic machinery, suggest that AMF could undergo a complete sexual reproduction cycle.

  1. Mating type gene homologues and putative sex pheromone-sensing pathway in arbuscular mycorrhizal fungi, a presumably asexual plant root symbiont.

    Directory of Open Access Journals (Sweden)

    Sébastien Halary

    Full Text Available The fungal kingdom displays a fascinating diversity of sex-determination systems. Recent advances in genomics provide insights into the molecular mechanisms of sex, mating type determination, and evolution of sexual reproduction in many fungal species in both ancient and modern phylogenetic lineages. All major fungal groups have evolved sexual differentiation and recombination pathways. However, sexuality is unknown in arbuscular mycorrhizal fungi (AMF of the phylum Glomeromycota, an ecologically vital group of obligate plant root symbionts. AMF are commonly considered an ancient asexual lineage dating back to the Ordovician, approximately 460 M years ago. In this study, we used genomic and transcriptomic surveys of several AMF species to demonstrate the presence of conserved putative sex pheromone-sensing mitogen-activated protein (MAP kinases, comparable to those described in Ascomycota and Basidiomycota. We also find genes for high mobility group (HMG transcription factors, homologous to SexM and SexP genes in the Mucorales. The SexM genes show a remarkable sequence diversity among multiple copies in the genome, while only a single SexP sequence was detected in some isolates of Rhizophagus irregularis. In the Mucorales and Microsporidia, the sexM gene is flanked by genes for a triosephosphate transporter (TPT and a RNA helicase, but we find no evidence for synteny in the vicinity of the Sex locus in AMF. Nonetheless, our results, together with previous observations on meiotic machinery, suggest that AMF could undergo a complete sexual reproduction cycle.

  2. Validation of candidate genes putatively associated with resistance to SCMV and MDMV in maize (Zea mays L.) by expression profiling

    DEFF Research Database (Denmark)

    Uzarowska, Anna; Dionisio, Giuseppe; Sarholz, Barbara

    2009-01-01

    Background The potyviruses sugarcane mosaic virus (SCMV) and maize dwarf mosaic virus (MDMV) are major pathogens of maize worldwide. Two loci, Scmv1 and Scmv2, have ealier been shown to confer complete resistance to SCMV. Custom-made microarrays containing previously identified SCMV resistance...... candidate genes and resistance gene analogs were utilised to investigate and validate gene expression and expression patterns of isogenic lines under pathogen infection in order to obtain information about the molecular mechanisms involved in maize-potyvirus interactions. Results By employing time course...... expressed genes in the SCMV experiment (75%) were identified one hour after virus inoculation, and about one quarter at multiple time points. Furthermore, most of the identified mapped genes were localised outside the Scmv QTL regions. Annotation revealed differential expression of promising pathogenesis...

  3. Chromosomal location and comparative genomics analysis of powdery mildew resistance gene Pm51 in a putative wheat-Thinopyrum ponticum introgression line.

    Science.gov (United States)

    Zhan, Haixian; Li, Guangrong; Zhang, Xiaojun; Li, Xin; Guo, Huijuan; Gong, Wenping; Jia, Juqing; Qiao, Linyi; Ren, Yongkang; Yang, Zujun; Chang, Zhijian

    2014-01-01

    Powdery mildew (PM) is a very destructive disease of wheat (Triticum aestivum L.). Wheat-Thinopyrum ponticum introgression line CH7086 was shown to possess powdery mildew resistance possibly originating from Th. ponticum. Genomic in situ hybridization and molecular characterization of the alien introgression failed to identify alien chromatin. To study the genetics of resistance, CH7086 was crossed with susceptible genotypes. Segregation in F2 populations and F2:3 lines tested with Chinese Bgt race E09 under controlled conditions indicated that CH7086 carries a single dominant gene for powdery mildew resistance. Fourteen SSR and EST-PCR markers linked with the locus were identified. The genetic distances between the locus and the two flanking markers were 1.5 and 3.2 cM, respectively. Based on the locations of the markers by nullisomic-tetrasomic and deletion lines of 'Chinese Spring', the resistance gene was located in deletion bin 2BL-0.89-1.00. Conserved orthologous marker analysis indicated that the genomic region flanking the resistance gene has a high level of collinearity to that of rice chromosome 4 and Brachypodium chromosome 5. Both resistance specificities and tests of allelism suggested the resistance gene in CH7086 was different from previously reported powdery mildew resistance genes on 2BL, and the gene was provisionally designated PmCH86. Molecular analysis of PmCH86 compared with other genes for resistance to Bgt in the 2BL-0.89-1.00 region suggested that PmCH86 may be a new PM resistance gene, and it was therefore designated as Pm51. The closely linked flanking markers could be useful in exploiting this putative wheat-Thinopyrum translocation line for rapid transfer of Pm51 to wheat breeding programs.

  4. Clustering of two genes putatively involved in cyanate detoxification evolved recently and independently in multiple fungal lineages.

    Science.gov (United States)

    Elmore, M Holly; McGary, Kriston L; Wisecaver, Jennifer H; Slot, Jason C; Geiser, David M; Sink, Stacy; O'Donnell, Kerry; Rokas, Antonis

    2015-03-01

    Fungi that have the enzymes cyanase and carbonic anhydrase show a limited capacity to detoxify cyanate, a fungicide employed by both plants and humans. Here, we describe a novel two-gene cluster that comprises duplicated cyanase and carbonic anhydrase copies, which we name the CCA gene cluster, trace its evolution across Ascomycetes, and examine the evolutionary dynamics of its spread among lineages of the Fusarium oxysporum species complex (hereafter referred to as the FOSC), a cosmopolitan clade of purportedly clonal vascular wilt plant pathogens. Phylogenetic analysis of fungal cyanase and carbonic anhydrase genes reveals that the CCA gene cluster arose independently at least twice and is now present in three lineages, namely Cochliobolus lunatus, Oidiodendron maius, and the FOSC. Genome-wide surveys within the FOSC indicate that the CCA gene cluster varies in copy number across isolates, is always located on accessory chromosomes, and is absent in FOSC's closest relatives. Phylogenetic reconstruction of the CCA gene cluster in 163 FOSC strains from a wide variety of hosts suggests a recent history of rampant transfers between isolates. We hypothesize that the independent formation of the CCA gene cluster in different fungal lineages and its spread across FOSC strains may be associated with resistance to plant-produced cyanates or to use of cyanate fungicides in agriculture.

  5. Genome sequencing and comparative genomics reveal a repertoire of putative pathogenicity genes in chilli anthracnose fungus Colletotrichum truncatum.

    Science.gov (United States)

    Rao, Soumya; Nandineni, Madhusudan R

    2017-01-01

    Colletotrichum truncatum, a major fungal phytopathogen, causes the anthracnose disease on an economically important spice crop chilli (Capsicum annuum), resulting in huge economic losses in tropical and sub-tropical countries. It follows a subcuticular intramural infection strategy on chilli with a short, asymptomatic, endophytic phase, which contrasts with the intracellular hemibiotrophic lifestyle adopted by most of the Colletotrichum species. However, little is known about the molecular determinants and the mechanism of pathogenicity in this fungus. A high quality whole genome sequence and gene annotation based on transcriptome data of an Indian isolate of C. truncatum from chilli has been obtained. Analysis of the genome sequence revealed a rich repertoire of pathogenicity genes in C. truncatum encoding secreted proteins, effectors, plant cell wall degrading enzymes, secondary metabolism associated proteins, with potential roles in the host-specific infection strategy, placing it next only to the Fusarium species. The size of genome assembly, number of predicted genes and some of the functional categories were similar to other sequenced Colletotrichum species. The comparative genomic analyses with other species and related fungi identified some unique genes and certain highly expanded gene families of CAZymes, proteases and secondary metabolism associated genes in the genome of C. truncatum. The draft genome assembly and functional annotation of potential pathogenicity genes of C. truncatum provide an important genomic resource for understanding the biology and lifestyle of this important phytopathogen and will pave the way for designing efficient disease control regimens.

  6. Transcriptome profiling of the eyestalk of precocious juvenile Chinese mitten crab reveals putative neuropeptides and differentially expressed genes.

    Science.gov (United States)

    Xu, Zhiqiang; Zhao, Muzi; Li, Xuguang; Lu, Quanping; Li, Yuehua; Ge, Jiachun; Pan, Jianlin

    2015-09-15

    Chinese mitten crabs that reach maturity 1 year earlier than normal crabs are known as precocious juvenile crabs. The molecular mechanisms underlying the precocity of the Chinese mitten crab are poorly understood. To identify the genes that may be involved in the control of precocity in Chinese mitten crab, we measured the expression profile of eyestalk genes in precocious and normally developed juvenile crabs using high-throughput sequencing on an Illumina HiSeq 2500 platform. We obtained 56,446,284 raw reads from the precocious crabs and 58,029,476 raw reads from the normally developed juvenile crabs. Reads from the two libraries were combined into a single data set. De novo assembly of the combined read set yielded 78,777 unigenes with an average length of 1563 bp. A total of 41,405 unigenes with predicted ORFs were selected for functional annotation. Among these genes, we identified three neuropeptide genes belonging to the crustacean hyperglycemic hormone family and two neuropeptide genes encoding the chromatophorotropic hormones. Transcriptome comparison between the two libraries revealed 42 genes that exhibited significant differential expression, of which 29 genes were up-regulated and 13 genes were down-regulated in the precocious crabs. To confirm the sequencing data, six differentially expressed genes with functional annotations were selected and validated by qRT-PCR. In conclusion, we obtained the comprehensive transcriptome of the eyestalk tissues of precocious juvenile crabs. The sequencing results may provide new insights into the biomolecular basis of precocity in the Chinese mitten crab.

  7. Nucleotide diversity and linkage disequilibrium of nine genes with putative effects on flowering time in perennial ryegrass (Lolium perenne L.)

    DEFF Research Database (Denmark)

    Fiil, Alice; Lenk, Ingo; Petersen, Klaus

    2011-01-01

    Optimization of flowering is an important breeding goal in forage and turf grasses, such as perennial ryegrass (Lolium perenne L.). Nine floral control genes including Lolium perenne CONSTANS (LpCO), SISTER OF FLOWERING LOCUS T (LpSFT), TERMINAL FLOWER1 (LpTFL1), VERNALIZATION1 (LpVRN1, identical......, one single nucleotide polymorphism (SNP) was present per 127 bp between two randomly sampled sequences for the nine genes (π = 0.00790). Two MADS-box genes, LpMADS1 and LpMADS10, involved in timing of flowering showed high nucleotide diversity and rapid LD decay, whereas MADS-box genes involved...

  8. Detection and distribution of putative virulence associated genes in Aeromonas species from freshwater and wastewater treatment plant.

    Science.gov (United States)

    Igbinosa, Isoken H; Okoh, Anthony I

    2013-11-01

    The detection of genes responsible for Aeromonas virulence is a vital tool in establishing the potential pathogenicity of the bacteria, as these virulence genes may act alone or in synergy in the establishment of infections. Freshwater and wastewater mixed liquor samples were collected from Kat river and Fort Beaufort wastewater treatment plant in the Eastern Cape Province of South Africa. Polymerase chain reaction was utilized for the amplification of the different genes coding for virulence. All virulence associated genes screened (alt, lip, fla, aer, ast, hlyA) were detected in at least one Aeromonas isolates. In fresh water sample, virulence genes were distributed as follows: lip (67%), aer (43%), alt (33%), fla (62%), ast (10%), and hlyA (86%), while in wastewater samples the occurrence were as follows: lip (92%), aer (21%), alt (54%), fla (83%), ast (29%), and hlyA (88%). The presence of these virulence genes in environmental Aeromonas isolates is of concern to public health as these organisms are potential pathogens in the environment and the virulence determinants could be transferred to aquatic organisms and humans by one mechanism or the other.

  9. Identification of Genes Putatively Involved in Chitin Metabolism and Insecticide Detoxification in the Rice Leaf Folder (Cnaphalocrocis medinalis Larvae through Transcriptomic Analysis

    Directory of Open Access Journals (Sweden)

    Hai-Zhong Yu

    2015-09-01

    Full Text Available The rice leaf roller (Cnaphalocrocis medinalis is one of the most destructive agricultural pests. Due to its migratory behavior, it is difficult to control worldwide. To date, little is known about major genes of C. medinalis involved in chitin metabolism and insecticide detoxification. In order to obtain a comprehensive genome dataset of C. medinalis, we conducted de novo transcriptome sequencing which focused on the major feeding stage of fourth-instar larvae, and our work revealed useful information on chitin metabolism and insecticide detoxification and target genes of C. medinalis. We acquired 29,367,797 Illumina reads and assembled these reads into 63,174 unigenes with an average length of 753 bp. Among these unigenes, 31,810 were annotated against the National Center for Biotechnology Information non-redundant (NCBI nr protein database, resulting in 24,246, 8669 and 18,176 assigned to Swiss-Prot, clusters of orthologous group (COG, and gene ontology (GO, respectively. We were able to map 10,043 unigenes into 285 pathways using the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG. Specifically, 16 genes, including five chitin deacetylases, two chitin synthases, five chitinases and four other related enzymes, were identified to be putatively involved in chitin biosynthesis and degradation, whereas 360 genes, including cytochrome P450s, glutathione S-transferases, esterases, and acetylcholinesterases, were found to be potentially involved in insecticide detoxification or as insecticide targets. The reliability of the transcriptome data was determined by reverse transcription quantitative PCR (RT-qPCR for the selected genes. Our data serves as a new and valuable sequence resource for genomic studies on C. medinalis. The findings should improve our understanding of C. medinalis genetics and contribute to management of this important agricultural pest.

  10. Identification of Genes Putatively Involved in Chitin Metabolism and Insecticide Detoxification in the Rice Leaf Folder (Cnaphalocrocis medinalis) Larvae through Transcriptomic Analysis.

    Science.gov (United States)

    Yu, Hai-Zhong; Wen, De-Fu; Wang, Wan-Lin; Geng, Lei; Zhang, Yan; Xu, Jia-Ping

    2015-09-10

    The rice leaf roller (Cnaphalocrocis medinalis) is one of the most destructive agricultural pests. Due to its migratory behavior, it is difficult to control worldwide. To date, little is known about major genes of C. medinalis involved in chitin metabolism and insecticide detoxification. In order to obtain a comprehensive genome dataset of C. medinalis, we conducted de novo transcriptome sequencing which focused on the major feeding stage of fourth-instar larvae, and our work revealed useful information on chitin metabolism and insecticide detoxification and target genes of C. medinalis. We acquired 29,367,797 Illumina reads and assembled these reads into 63,174 unigenes with an average length of 753 bp. Among these unigenes, 31,810 were annotated against the National Center for Biotechnology Information non-redundant (NCBI nr) protein database, resulting in 24,246, 8669 and 18,176 assigned to Swiss-Prot, clusters of orthologous group (COG), and gene ontology (GO), respectively. We were able to map 10,043 unigenes into 285 pathways using the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG). Specifically, 16 genes, including five chitin deacetylases, two chitin synthases, five chitinases and four other related enzymes, were identified to be putatively involved in chitin biosynthesis and degradation, whereas 360 genes, including cytochrome P450s, glutathione S-transferases, esterases, and acetylcholinesterases, were found to be potentially involved in insecticide detoxification or as insecticide targets. The reliability of the transcriptome data was determined by reverse transcription quantitative PCR (RT-qPCR) for the selected genes. Our data serves as a new and valuable sequence resource for genomic studies on C. medinalis. The findings should improve our understanding of C. medinalis genetics and contribute to management of this important agricultural pest.

  11. The Pratylenchus penetrans Transcriptome as a Source for the Development of Alternative Control Strategies: Mining for Putative Genes Involved in Parasitism and Evaluation of in planta RNAi.

    Science.gov (United States)

    Vieira, Paulo; Eves-van den Akker, Sebastian; Verma, Ruchi; Wantoch, Sarah; Eisenback, Jonathan D; Kamo, Kathryn

    2015-01-01

    The root lesion nematode Pratylenchus penetrans is considered one of the most economically important species within the genus. Host range studies have shown that nearly 400 plant species can be parasitized by this species. To obtain insight into the transcriptome of this migratory plant-parasitic nematode, we used Illumina mRNA sequencing analysis of a mixed population, as well as nematode reads detected in infected soybean roots 3 and 7 days after nematode infection. Over 140 million paired end reads were obtained for this species, and de novo assembly resulted in a total of 23,715 transcripts. Homology searches showed significant hit matches to 58% of the total number of transcripts using different protein and EST databases. In general, the transcriptome of P. penetrans follows common features reported for other root lesion nematode species. We also explored the efficacy of RNAi, delivered from the host, as a strategy to control P. penetrans, by targeted knock-down of selected nematode genes. Different comparisons were performed to identify putative nematode genes with a role in parasitism, resulting in the identification of transcripts with similarities to other nematode parasitism genes. Focusing on the predicted nematode secreted proteins found in this transcriptome, we observed specific members to be up-regulated at the early time points of infection. In the present study, we observed an enrichment of predicted secreted proteins along the early time points of parasitism by this species, with a significant number being pioneer candidate genes. A representative set of genes examined using RT-PCR confirms their expression during the host infection. The expression patterns of the different candidate genes raise the possibility that they might be involved in critical steps of P. penetrans parasitism. This analysis sheds light on the transcriptional changes that accompany plant infection by P. penetrans, and will aid in identifying potential gene targets for

  12. RT-qPCR analysis of putative beer-spoilage gene expression during growth of Lactobacillus brevis BSO 464 and Pediococcus claussenii ATCC BAA-344(T) in beer.

    Science.gov (United States)

    Bergsveinson, Jordyn; Pittet, Vanessa; Ziola, Barry

    2012-10-01

    Lactic acid bacteria (LAB) contamination of beer presents a continual economic threat to brewers. Interestingly, only certain isolates of LAB can grow in the hostile beer environment (e.g., as studied here, Lactobacillus brevis BSO 464 (Lb464) and a non-ropy isolate of Pediococcus claussenii ATCC BAA-344(T) (Pc344NR)), indicating that significant genetic specialization is required. The genes hitA, horA, horB, horC, and bsrA, which have been proposed to confer beer-spoiling ability to an organism, are suspected of counteracting the antimicrobial effects of hops. However, these genes are not present in the same combination (if at all) across beer-spoiling organisms. As such, we sought to investigate the extent to which these genes participate during Lb464 and Pc344NR mid-logarithmic growth in beer through reverse transcription quantitative PCR analysis. We first determined the optimal reference gene set needed for data normalization and, for each bacterium, established that two genes were needed for accurate assessment of gene expression. Following this, we found that horA expression was induced for Pc344NR, but not for Lb464, during growth in beer. Instead, horC expression was dramatically increased in Lb464 when growing in beer, whereas no change was detected for the other putative beer-spoilage-related genes. This indicates that HorC may be one of the principle mediators enabling growth of Lb464 in beer, whereas in Pc344NR, this may be attributable to HorA. These findings not only reveal that Lb464 and Pc344NR are unique in their beer-specific genetic expression profile but also indicate that a range of genetic specialization exists among beer-spoilage bacteria.

  13. Cloning and functional analysis of putative malonyl-CoA:acyl-carrier protein transacylase gene from the docosahexaenoic acid-producer Schizochytrium sp. TIO1101.

    Science.gov (United States)

    Cheng, Rubin; Ge, Yuqing; Yang, Bo; Zhong, Xiaoming; Lin, Xiangzhi; Huang, Zhen

    2013-06-01

    Malonyl-CoA:acyl-carrier protein transacylase (MCAT), which transfers the malonyl group from malonyl-CoA to holo-acyl carrier protein (ACP), is a key enzyme in fatty acid biosynthesis. Schizochytrium sp. TIO1101 is a marine protist with high levels of docosahexaenoic acid accumulation. In this study, the putative fabD gene coding MCAT was isolated from Schizochytrium sp. TIO1101. The Schizochytrium MCAT gene (ScTIOfabD) contained an 1176 bp open reading frame encoding a protein of 391 amino acids. The ScTIOfabD gene exhibited high novelty in nucleotide and amino acid sequence. The highest amino acid identity was only 35 % between ScTIOMCAT and the reported MCATs. Further studies demonstrated that ScTIOMCAT could bind malonyl-CoA directly and transfer malonyl group from malonyl-CoA to the ACP domain in vitro. Phylogenetic analysis suggested that ScTIOMCAT was relative close to MCATs of yeast strains. Overexpression of ScTIOMCAT in Saccharomyces cereviseae significantly increased the MCAT activity, without negative effects on the growth rate of the host strain. In addition, ScTIOMCAT generated 16.8 and 62 % increase in biomass and fatty acid accumulation, respectively, and did not alter the profile of fatty acid. Our results indicated that the novel MCAT gene from Schizochytrium sp. TIO1101 was crucial for fatty acid synthesis and had potential applications for genetic modifications of oil-producing species.

  14. Emerging putative associations between non-coding RNAs and protein-coding genes in Neuropathic Pain. Added value from re-using microarray data.

    Directory of Open Access Journals (Sweden)

    Enrico Capobianco

    2016-10-01

    Full Text Available Regeneration of injured nerves is likely occurring in the peripheral nervous system, but not in the central nervous system. Although protein-coding gene expression has been assessed during nerve regeneration, little is currently known about the role of non-coding RNAs (ncRNAs. This leaves open questions about the potential effects of ncRNAs at transcriptome level. Due to the limited availability of human neuropathic pain data, we have identified the most comprehensive time-course gene expression profile referred to sciatic nerve injury, and studied in a rat model, using two neuronal tissues, namely dorsal root ganglion (DRG and sciatic nerve (SN. We have developed a methodology to identify differentially expressed bioentities starting from microarray probes, and re-purposing them to annotate ncRNAs, while analyzing the expression profiles of protein-coding genes. The approach is designed to reuse microarray data and perform first profiling and then meta-analysis through three main steps. First, we used contextual analysis to identify what we considered putative or potential protein coding targets for selected ncRNAs. Relevance was therefore assigned to differential expression of neighbor protein-coding genes, with neighborhood defined by a fixed genomic distance from long or antisense ncRNA loci, and of parent genes associated with pseudogenes. Second, connectivity among putative targets was used to build networks, in turn useful to conduct inference at interactomic scale. Last, network paths were annotated to assess relevance to neuropathic pain. We found significant differential expression in long-intergenic ncRNAs (32 lincRNAs in SN, and 8 in DRG, antisense RNA (31 asRNA in SN, and 12 in DRG and pseudogenes (456 in SN, 56 in DRG. In particular, contextual analysis centered on pseudogenes revealed some targets with known association to neurodegeneration and/or neurogenesis processes. While modules of the olfactory receptors were clearly

  15. Putative resistance gene markers associated with quantitative trait loci for fire blight resistance in Malus ‘Robusta 5’ accessions

    Directory of Open Access Journals (Sweden)

    Gardiner Susan E

    2012-04-01

    Full Text Available Abstract Background Breeding of fire blight resistant scions and rootstocks is a goal of several international apple breeding programs, as options are limited for management of this destructive disease caused by the bacterial pathogen Erwinia amylovora. A broad, large-effect quantitative trait locus (QTL for fire blight resistance has been reported on linkage group 3 of Malus ‘Robusta 5’. In this study we identified markers derived from putative fire blight resistance genes associated with the QTL by integrating further genetic mapping studies with bioinformatics analysis of transcript profiling data and genome sequence databases. Results When several defined E.amylovora strains were used to inoculate three progenies from international breeding programs, all with ‘Robusta 5’ as a common parent, two distinct QTLs were detected on linkage group 3, where only one had previously been mapped. In the New Zealand ‘Malling 9’ X ‘Robusta 5’ population inoculated with E. amylovora ICMP11176, the proximal QTL co-located with SNP markers derived from a leucine-rich repeat, receptor-like protein ( MxdRLP1 and a closely linked class 3 peroxidase gene. While the QTL detected in the German ‘Idared’ X ‘Robusta 5’ population inoculated with E. amylovora strains Ea222_JKI or ICMP11176 was approximately 6 cM distal to this, directly below a SNP marker derived from a heat shock 90 family protein gene ( HSP90. In the US ‘Otawa3’ X ‘Robusta5’ population inoculated with E. amylovora strains Ea273 or E2002a, the position of the LOD score peak on linkage group 3 was dependent upon the pathogen strains used for inoculation. One of the five MxdRLP1 alleles identified in fire blight resistant and susceptible cultivars was genetically associated with resistance and used to develop a high resolution melting PCR marker. A resistance QTL detected on linkage group 7 of the US population co-located with another HSP90 gene-family member and a WRKY

  16. Emerging Putative Associations between Non-Coding RNAs and Protein-Coding Genes in Neuropathic Pain: Added Value from Reusing Microarray Data

    Science.gov (United States)

    Raju, Hemalatha B.; Tsinoremas, Nicholas F.; Capobianco, Enrico

    2016-01-01

    Regeneration of injured nerves is likely occurring in the peripheral nervous system, but not in the central nervous system. Although protein-coding gene expression has been assessed during nerve regeneration, little is currently known about the role of non-coding RNAs (ncRNAs). This leaves open questions about the potential effects of ncRNAs at transcriptome level. Due to the limited availability of human neuropathic pain (NP) data, we have identified the most comprehensive time-course gene expression profile referred to sciatic nerve (SN) injury and studied in a rat model using two neuronal tissues, namely dorsal root ganglion (DRG) and SN. We have developed a methodology to identify differentially expressed bioentities starting from microarray probes and repurposing them to annotate ncRNAs, while analyzing the expression profiles of protein-coding genes. The approach is designed to reuse microarray data and perform first profiling and then meta-analysis through three main steps. First, we used contextual analysis to identify what we considered putative or potential protein-coding targets for selected ncRNAs. Relevance was therefore assigned to differential expression of neighbor protein-coding genes, with neighborhood defined by a fixed genomic distance from long or antisense ncRNA loci, and of parental genes associated with pseudogenes. Second, connectivity among putative targets was used to build networks, in turn useful to conduct inference at interactomic scale. Last, network paths were annotated to assess relevance to NP. We found significant differential expression in long-intergenic ncRNAs (32 lincRNAs in SN and 8 in DRG), antisense RNA (31 asRNA in SN and 12 in DRG), and pseudogenes (456 in SN and 56 in DRG). In particular, contextual analysis centered on pseudogenes revealed some targets with known association to neurodegeneration and/or neurogenesis processes. While modules of the olfactory receptors were clearly identified in protein

  17. A comparative genomic analysis of putative pathogenicity genes in the host-specific sibling species Colletotrichum graminicola and Colletotrichum sublineola.

    Science.gov (United States)

    Buiate, E A S; Xavier, K V; Moore, N; Torres, M F; Farman, M L; Schardl, C L; Vaillancourt, L J

    2017-01-10

    Colletotrichum graminicola and C. sublineola cause anthracnose leaf and stalk diseases of maize and sorghum, respectively. In spite of their close evolutionary relationship, the two species are completely host-specific. Host specificity is often attributed to pathogen virulence factors, including specialized secondary metabolites (SSM), and small-secreted protein (SSP) effectors. Genes relevant to these categories were manually annotated in two co-occurring, contemporaneous strains of C. graminicola and C. sublineola. A comparative genomic and phylogenetic analysis was performed to address the evolutionary relationships among these and other divergent gene families in the two strains. Inoculation of maize with C. sublineola, or of sorghum with C. graminicola, resulted in rapid plant cell death at, or just after, the point of penetration. The two fungal genomes were very similar. More than 50% of the assemblies could be directly aligned, and more than 80% of the gene models were syntenous. More than 90% of the predicted proteins had orthologs in both species. Genes lacking orthologs in the other species (non-conserved genes) included many predicted to encode SSM-associated proteins and SSPs. Other common groups of non-conserved proteins included transporters, transcription factors, and CAZymes. Only 32 SSP genes appeared to be specific to C. graminicola, and 21 to C. sublineola. None of the SSM-associated genes were lineage-specific. Two different strains of C. graminicola, and three strains of C. sublineola, differed in no more than 1% percent of gene sequences from one another. Efficient non-host recognition of C. sublineola by maize, and of C. graminicola by sorghum, was observed in epidermal cells as a rapid deployment of visible resistance responses and plant cell death. Numerous non-conserved SSP and SSM-associated predicted proteins that could play a role in this non-host recognition were identified. Additional categories of genes that were also highly

  18. Transcriptome Analysis of Methyl Jasmonate-Elicited Panax ginseng Adventitious Roots to Discover Putative Ginsenoside Biosynthesis and Transport Genes

    Science.gov (United States)

    Cao, Hongzhe; Nuruzzaman, Mohammed; Xiu, Hao; Huang, Jingjia; Wu, Kunlu; Chen, Xianghui; Li, Jijia; Wang, Li; Jeong, Ji-Hak; Park, Sun-Jin; Yang, Fang; Luo, Junli; Luo, Zhiyong

    2015-01-01

    The Panax ginseng C.A. Meyer belonging to the Araliaceae has long been used as an herbal medicine. Although public databases are presently available for this family, no methyl jasmonate (MeJA) elicited transcriptomic information was previously reported on this species, with the exception of a few expressed sequence tags (ESTs) using the traditional Sanger method. Here, approximately 53 million clean reads of adventitious root transcriptome were separately filtered via Illumina HiSeq™2000 from two samples treated with MeJA (Pg-MeJA) and equal volumes of solvent, ethanol (Pg-Con). Jointly, a total of 71,095 all-unigenes from both samples were assembled and annotated, and based on sequence similarity search with known proteins, a total of 56,668 unigenes was obtained. Out of these annotated unigenes, 54,920 were assigned to the NCBI non-redundant protein (Nr) database, 35,448 to the Swiss-prot database, 43,051 to gene ontology (GO), and 19,986 to clusters of orthologous groups (COG). Searching in the Kyoto encyclopedia of genes and genomes (KEGG) pathway database indicated that 32,200 unigenes were mapped to 128 KEGG pathways. Moreover, we obtained several genes showing a wide range of expression levels. We also identified a total of 749 ginsenoside biosynthetic enzyme genes and 12 promising pleiotropic drug resistance (PDR) genes related to ginsenoside transport. PMID:25642758

  19. Transcriptome Analysis of Methyl Jasmonate-Elicited Panax ginseng Adventitious Roots to Discover Putative Ginsenoside Biosynthesis and Transport Genes

    Directory of Open Access Journals (Sweden)

    Hongzhe Cao

    2015-01-01

    Full Text Available The Panax ginseng C.A. Meyer belonging to the Araliaceae has long been used as an herbal medicine. Although public databases are presently available for this family, no methyl jasmonate (MeJA elicited transcriptomic information was previously reported on this species, with the exception of a few expressed sequence tags (ESTs using the traditional Sanger method. Here, approximately 53 million clean reads of adventitious root transcriptome were separately filtered via Illumina HiSeq™2000 from two samples treated with MeJA (Pg-MeJA and equal volumes of solvent, ethanol (Pg-Con. Jointly, a total of 71,095 all-unigenes from both samples were assembled and annotated, and based on sequence similarity search with known proteins, a total of 56,668 unigenes was obtained. Out of these annotated unigenes, 54,920 were assigned to the NCBI non-redundant protein (Nr database, 35,448 to the Swiss-prot database, 43,051 to gene ontology (GO, and 19,986 to clusters of orthologous groups (COG. Searching in the Kyoto encyclopedia of genes and genomes (KEGG pathway database indicated that 32,200 unigenes were mapped to 128 KEGG pathways. Moreover, we obtained several genes showing a wide range of expression levels. We also identified a total of 749 ginsenoside biosynthetic enzyme genes and 12 promising pleiotropic drug resistance (PDR genes related to ginsenoside transport.

  20. A cohort of balanced reciprocal translocations associated with dyslexia: identification of two putative candidate genes at DYX1

    DEFF Research Database (Denmark)

    Buonincontri, Roberta; Bache, Iben; Silahtaroglu, Asli

    2011-01-01

    Dyslexia is one of the most common neurodevelopmental disorders where likely many genes are involved in the pathogenesis. So far six candidate dyslexia genes have been proposed, and two of these were identified by rare chromosomal translocations in affected individuals. By systematic re......-examination of all translocation carriers in Denmark, we have identified 16 different translocations associated with dyslexia. In four families, where the translocation co-segregated with the phenotype, one of the breakpoints concurred (at the cytogenetic level) with either a known dyslexia linkage region--at 15q21...... (DYX1), 2p13 (DYX3) and 1p36 (DYX8)--or an unpublished linkage region at 19q13. As a first exploitation of this unique cohort, we identify three novel candidate dyslexia genes, ZNF280D and TCF12 at 15q21, and PDE7B at 6q23.3, by molecular mapping of the familial translocation with the 15q21 breakpoint....

  1. Sequence analysis of a 9873 bp fragment of the left arm of yeast chromosome XV that contains the ARG8 and CDC33 genes, a putative riboflavin synthase beta chain gene, and four new open reading frames.

    Science.gov (United States)

    Casas, C; Aldea, M; Casamayor, A; Lafuente, M J; Gamo, F J; Gancedo, C; Ariño, J; Herrero, E

    1995-09-15

    The DNA sequence of a 9873 bp fragment located near the left telomere of chromosome XV has been determined. Sequence analysis reveals seven open reading frames. One is the ARG8 gene coding for N-acetylornithine aminotransferase. Another corresponds to CDC33, which codes for the initiation factor 4E or cap binding protein. The open reading frame AOE169 can be considered as the putative gene for the Saccharomyces cerevisiae riboflavin synthase beta chain, since its translation product shows strong homology with four prokaryotic riboflavin synthase beta chains.

  2. Honeybee (Apis mellifera L.) mrjp gene family: computational analysis of putative promoters and genomic structure of mrjp1, the gene coding for the most abundant protein of larval food.

    Science.gov (United States)

    Malecová, Barbora; Ramser, Juliane; O'Brien, John K; Janitz, Michal; Júdová, Jana; Lehrach, Hans; Simúth, Jozef

    2003-01-16

    Mrjp1 gene belongs to the honeybee mrjp gene family encoding the major royal jelly proteins (MRJPs), secreted by nurse bees into the royal jelly. In this study, we have isolated the genomic clone containing the entire mrjp1 gene and determined its sequence. The mrjp1 gene sequence spans over 3038 bp and contains six exons separated by five introns. Seven mismatches between the mrjp1 gene sequence and two previously independently published cDNA sequences were found, but these differences do not lead to any change in the deduced amino acid sequence of MRJP1. With the aid of inverse polymerase chain reaction we obtained sequences flanking the 5' ends of other mrjp genes (mrjp2, mrjp3, mrjp4 and mrjp5). Putative promoters were predicted upstream of all mrjp genes (including mrjp1). The predicted promoters contain the TATA motif (TATATATT), highly conserved both in sequence and position. Ultraspiracle (USP) transcription factor (TF) binding sites in putative promoter regions and clusters of dead ringer TF binding sites upstream of these promoters were predicted computationally. We propose that USP, as a juvenile hormone (JH) binding TF, might possibly act as a mediator of mrjp expression in response to JH. Mrjp1's genomic locus is predicted to encode an antisense transcript, partially overlapping with five mrjp1 exons and entirely overlapping with the putative promoter and predicted transcriptional start point of mrjp1. This finding may shed light on the mechanisms of regulation of mrjps expression. Southern blot analysis of genomic DNA revealed that all so far known members of mrjp gene family (mrjp1, mrjp2, mrjp3, mrjp4 and mrjp5) are present as single-copy genes per haploid honeybee genome. Although MRJPs and the yellow protein of Drosophila melanogaster share a certain degree of similarity in aa sequence and although it has been shown that they share a common evolutionary origin, neither structural similarities in the gene organization, nor significant similarities

  3. A single base insertion in the putative transmembrane domain of the tyrosinase gene as a cause for tyrosinase-negative oculocutaneous albinism

    Energy Technology Data Exchange (ETDEWEB)

    Chintamaneni, C.D.; Kobayashi, Y.; Kwon, B.S. (Indiana Univ. School of Medicine, Indianapolis (United States)); Halaban, R. (Yale Univ. School of Medicine, New Haven, CT (United States)); Witkop, C.J. Jr. (Univ. of Minnesota, Minneapolis (United States))

    1991-06-15

    The authors have determined a molecular defect to be the likely basis for inactivity of the tyrosinase from a patient with tyrosinase-negative oculocutaneous albinism. A single base (thymine) was inserted in exon 5 of the tyrosinase gene following codon 471 in the putative transmembrane coding region. This insertion caused a shift in the reading frame of 19 amino acids at the 3{prime} end and introduced a premature termination signal that would be expected to truncate the protein by 21 amino acids at the carboxyl terminus. The albino tyrosinase was not recognized by antibodies directed to the carboxyl terminus of tyrosinase. Furthermore, as shown by gel electrophoresis of the immunoprecipitated protein, the tyrosinase was {approx} 3kDa smaller than normal. Similar immunoprecipitation data were obtained when cloned normal and mutant tyrosinases were expressed in COS-1 cells.

  4. Detecting population structure in a high gene-flow species, Atlantic herring (Clupea harengus): direct, simultaneous evaluation of neutral vs putatively selected loci

    DEFF Research Database (Denmark)

    André, C.; Larsson, L. C.; Laikre, L.;

    2010-01-01

    DNA, with one microsatellite locus, Cpa112, previously shown to be influenced by divergent selection associated with salinity, and one locus located in the major histocompatibility complex class IIA (MHC-IIA) gene, using the same individuals across analyses. Samples were collected in 2002 and 2003...... at two locations in the North Sea, one location in the Skagerrak and one location in the low-saline Baltic Sea. Levels of divergence for putatively neutral markers were generally low, with the exception of single outlier locus/sample combinations; microsatellites were the most statistically powerful...... to detect population structure in Atlantic herring (Clupea harengus), a migratory pelagic species with large effective population sizes. We compared the spatial and temporal patterns of divergence and statistical power of three traditional genetic marker types, microsatellites, allozymes and mitochondrial...

  5. Deletion of PREPL, a gene encoding a putative serine oligopeptidase, in patients with hypotonia-cystinuria syndrome

    NARCIS (Netherlands)

    Jaeken, J.; Martens, K.; Francois, I.; Eyskens, F.; Lecointre, C.; Derua, R.; Meulemans, S.; Slootstra, J.W.; Waelkens, E.; Zegher, de F.; Creemers, J.W.M.; Matthijs, G.

    2006-01-01

    In 11 patients with a recessive congenital disorder, which we refer to as ¿the hypotonia-cystinuria syndrome,¿ microdeletion of part of the SLC3A1 and PREPL genes on chromosome 2p21 was found. Patients present with generalized hypotonia at birth, nephrolithiasis, growth hormone deficiency, minor fac

  6. Isolation and characterization of the glnD gene of Gluconacetobacter diazotrophicus, encoding a putative uridylyltransferase/uridylyl-removing enzyme.

    Science.gov (United States)

    Perlova, Olena; Nawroth, Roman; Zellermann, Eva-Maria; Meletzus, Dietmar

    2002-09-04

    The glnD gene of Gluconacetobacter diazotrophicus was isolated by complementation of the Azotobacter vinelandii glnD (nfrX) mutant strain MV17 using a pLAFR3 cosmid library. The 5 kb chromosomal DNA region encoding the glnD gene on cosmid pAD401 was identified by introduction of deletions as well as subcloning of restriction fragments followed by subsequent DNA sequencing. Three open reading frames were identified with the deduced amino acid sequence of ORF1 showing significant homologies to known GlnD proteins of other proteobacteria such as Sinorhizobium meliloti, Rhizobium tropici, Escherichia coli and Azotobacter vinelandii.A mutagenesis of the chromosomal glnD gene was carried out by insertion of an interposon carrying the kanamycin resistance gene of Tn5. Mutants carrying the cassette inserted into a central region of glnD could not be isolated, while an interposon mutation at the 3' end of glnD was successful. The resulting strain showed a prolonged generation time in complex growth medium and was unable to utilize ammonium as sole nitrogen source. This phenotype appears to be pleiotropic, since the addition of single amino acids to the minimal medium was not sufficient to allow growth. Furthermore, the glnD mutant was able to express nitrogenase under diazotrophic as well as repressing growth conditions.

  7. Flavonoid Biosynthesis Genes Putatively Identified in the Aromatic Plant Polygonum minus via Expressed Sequences Tag (EST Analysis

    Directory of Open Access Journals (Sweden)

    Zamri Zainal

    2012-02-01

    Full Text Available P. minus is an aromatic plant, the leaf of which is widely used as a food additive and in the perfume industry. The leaf also accumulates secondary metabolites that act as active ingredients such as flavonoid. Due to limited genomic and transcriptomic data, the biosynthetic pathway of flavonoids is currently unclear. Identification of candidate genes involved in the flavonoid biosynthetic pathway will significantly contribute to understanding the biosynthesis of active compounds. We have constructed a standard cDNA library from P. minus leaves, and two normalized full-length enriched cDNA libraries were constructed from stem and root organs in order to create a gene resource for the biosynthesis of secondary metabolites, especially flavonoid biosynthesis. Thus, large‑scale sequencing of P. minus cDNA libraries identified 4196 expressed sequences tags (ESTs which were deposited in dbEST in the National Center of Biotechnology Information (NCBI. From the three constructed cDNA libraries, 11 ESTs encoding seven genes were mapped to the flavonoid biosynthetic pathway. Finally, three flavonoid biosynthetic pathway-related ESTs chalcone synthase, CHS (JG745304, flavonol synthase, FLS (JG705819 and leucoanthocyanidin dioxygenase, LDOX (JG745247 were selected for further examination by quantitative RT-PCR (qRT-PCR in different P. minus organs. Expression was detected in leaf, stem and root. Gene expression studies have been initiated in order to better understand the underlying physiological processes.

  8. Deletion of PREPL, a gene encoding a putative serine oligopeptidase, in patients with hypotonia-cystinuria syndrome

    NARCIS (Netherlands)

    Jaeken, J.; Martens, K.; Francois, I.; Eyskens, F.; Lecointre, C.; Derua, R.; Meulemans, S.; Slootstra, J.W.; Waelkens, E.; Zegher, de F.; Creemers, J.W.M.; Matthijs, G.

    2006-01-01

    In 11 patients with a recessive congenital disorder, which we refer to as ¿the hypotonia-cystinuria syndrome,¿ microdeletion of part of the SLC3A1 and PREPL genes on chromosome 2p21 was found. Patients present with generalized hypotonia at birth, nephrolithiasis, growth hormone deficiency, minor fac

  9. CPR1: a gene encoding a putative signal peptidase that functions in pathogenicity of Colletotrichum graminicola to maize.

    Science.gov (United States)

    Thon, M R; Nuckles, E M; Takach, J E; Vaillancourt, L J

    2002-02-01

    Colletotrichum graminicola causes anthracnose leaf blight and stalk rot of maize. We used restriction-enzyme mediated insertional (REMI) mutagenesis to identify a gene in this fungus that is required for pathogenicity to both stalks and leaves. The predicted polypeptide encoded by this gene, which we have named CPR1, is similar to a family of proteins that comprise one subunit of the eukaryotic microsomal signal peptidase. The nonpathogenic CPR1 REMI mutant contains a plasmid integration in the 3' untranslated region of the gene, 19 bp downstream from the stop codon. The result is a significant reduction in transcript levels in comparison to the wild type, perhaps as a result of increased transcript instability. We were unable to knock out the CPR1 gene, and it may be essential for viability. Microscopic examination of the REMI mutant on maize leaves revealed that it is fully capable of penetrating and colonizing host cells during the initial, biotrophic phases of the disease interaction but, unlike the wild type, it appears to be unable to switch to a necrotrophic mode of growth. We suggest that the CPR1 REMI mutant may be unable to secrete sufficient quantities of degradative enzymes to support that transition. The CPR1 REMI mutant provides us with a useful tool for future studies of the role of fungal protein transport in this important stalk rot disease of maize.

  10. Identification and cloning of putative water clarification genes of Moringa peregrina (Forssk. Fiori in E. coli Xl 1 blue cells

    Directory of Open Access Journals (Sweden)

    Reihaneh Ghodsi

    2014-01-01

    Results: We obtained 3 PCR products (approximately 200, 300, and 400 bps. Conclusion: After comparison of the sequences of 300bp band obtained from M. peregrina with Mo 1,2 gene in M. oleifera, it seems that 300bp band is a good candidate to investigate regarding its potential flocculent activity.

  11. Myosin IXB gene region and gluten intolerance : linkage to coeliac disease and a putative dermatitis herpetiformis association

    NARCIS (Netherlands)

    Koskinen, L. L. E.; Korponay-Szabo, I. R.; Viiri, K.; Juuti-Uusitalo, K.; Kaukinen, K.; Lindfors, K.; Mustalahti, K.; Kurppa, K.; Adany, R.; Pocsai, Z.; Szeles, G.; Einarsdottir, E.; Wijmenga, C.; Maeki, M.; Partanen, J.; Kere, J.; Saavalainen, P.

    2008-01-01

    Background: Coeliac disease is caused by dietary gluten, which triggers chronic inflammation of the small intestine in genetically predisposed individuals. In one quarter of the patients the disease manifests in the skin as dermatitis herpetiformis. Recently, a novel candidate gene, myosin IXB on ch

  12. A semidwarf phenotype of barley uzu results from a nucleotide substitution in the gene encoding a putative brassinosteroid receptor.

    Science.gov (United States)

    Chono, Makiko; Honda, Ichiro; Zeniya, Haruko; Yoneyama, Koichi; Saisho, Daisuke; Takeda, Kazuyoshi; Takatsuto, Suguru; Hoshino, Tsuguhiro; Watanabe, Yoshiaki

    2003-11-01

    Brassinosteroids (BRs) play important roles throughout plant growth and development. Despite the importance of clarifying the mechanism of BR-related growth regulation in cereal crops, BR-related cereal mutants have been identified only in rice (Oryza sativa). We previously found that semidwarf barley (Hordeum vulgare) accessions carrying the "uzu" gene, called "uzu" barley in Japan, are non-responding for brassinolide (BL). We then performed chemical and molecular analyses to clarify the mechanisms of uzu dwarfism using isogenic line pairs of uzu gene. The response of the uzu line to BL was significantly lower than that of its corresponding normal line. Measurement of BRs showed that the uzu line accumulates BRs, similar to known BR-insensitive mutants. The marker synteny of rice and barley chromosomes suggests that the uzu gene may be homologous to rice D61, a rice homolog of Arabidopsis BR-insensitive 1 (BRI1), encoding a BR-receptor protein. A barley homolog of BRI1, HvBRI1, was isolated by using degenerate primers. A comparison of HvBRI1 sequences in uzu and normal barley varieties showed that the uzu phenotype is correlated with a single nucleotide substitution. This substitution results in an amino acid change at a highly conserved residue in the kinase domain of the BR-receptor protein. These results may indicate that uzu dwarfism is caused by the missense mutation in HvBRI1. The uzu gene is being introduced into all hull-less barley cultivars in Japan as an effective dwarf gene for practical use, and this is the first report about an agronomically important mutation related to BRs.

  13. GhHyPRP4, a cotton gene encoding putative hybrid proline-rich protein, is preferentially expressed in leaves and involved in plant response to cold stress

    Institute of Scientific and Technical Information of China (English)

    Gengqing Huang; Siying Gong; Wenliang Xu; Peng Li; Dejing Zhang; Lixia Qin; Wen Li; Xuebao Li

    2011-01-01

    Plant hybrid proline-rich proteins (HyPRPs) usually consist of an N-terminal signal peptide, a central prolinerich domain, and a conserved eight-cysteine motif C-terminal domain. In this study, one gene (designated as GhHyPRP4) encoding putative HyPRP was isolated from cotton cDNA library. Northern blot and quantitative reverse transcriptase-polymerase chain reaction analyses revealed that GhHyPRP4 was preferentially expressed in leaves. Under cold stress, GhHyPRP4 expression was significantly up-regulated in leaves of cotton seedlings.Using the genome walking approach, a promoter fragment of GhHyPRP4 gene was isolated from cotton genome.GUS (β-glucuronidase) gene driven by GhHyPRP4 promoter was specifically expressed in leaves and cotyledons of the transgenic 4rabidopsis thaliana. Furthermore,GUS expression in leaves was remarkably induced by cold stress. Overexpression of GhHyPRP4 in yeast (Schizosaccharomyces pombe) significantly enhanced the cell survival rate upon treatment under -20℃ for 60 h.These data suggested that GhHyPRP4 may be involved in plant response to cold stress during seedling development of cotton.

  14. The promoter of the barley aleurone-specific gene encoding a putative 7 kDa lipid transfer protein confers aleurone cell-specific expression in transgenic rice.

    Science.gov (United States)

    Kalla, R; Shimamoto, K; Potter, R; Nielsen, P S; Linnestad, C; Olsen, O A

    1994-12-01

    This paper describes the aleurone-specific gene Ltp2 from barley, which encodes a putative 7 kDa non-specific lipid transfer protein. As shown by Northern and in situ hybridization analyses, the Ltp2 transcript is present in barley aleurone cells shortly after the initiation of aleurone cell differentiation. The expression of Ltp2 increases until grain mid-maturity, but the mRNA is absent from mature grains. The Ltp2 transcript is undetectable in the embryo and vegetative tissues, confirming the aleurone specificity of the Ltp2 gene. The ability of the isolated 801 bp Ltp2 promoter to direct aleurone-specific expression in immature barley grains is demonstrated by particle bombardment experiments. In these experiments, the activity of the Ltp2 promoter is 5% of the activity of the strong constitutive Actin1 promoter from rice, as quantified by GUS activity measurements. In stably transformed rice plants containing the Ltp2 promoter-Gus construct, the specificity of the Ltp2 promoter is confirmed in vivo by the presence of GUS activity exclusively in the aleurone layer. This study demonstrates the conserved nature of the regulatory signals involved in aleurone-specific gene transcription in cereal grains.

  15. Mapping of Powdery Mildew Resistance Gene pmCH89 in a Putative Wheat-Thinopyrum intermedium Introgression Line

    Directory of Open Access Journals (Sweden)

    Liyuan Hou

    2015-07-01

    Full Text Available Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt, is a globally serious disease adversely affecting wheat production. The Bgt-resistant wheat breeding line CH09W89 was derived after backcrossing a Bgt resistant wheat-Thinopyrum intermedium partial amphiploid TAI7045 with susceptible wheat cultivars. At the seedling stage, CH09W89 exhibited immunity or high resistance to Bgt pathotypes E09, E20, E21, E23, E26, Bg1, and Bg2, similar to its donor line TAI7045 and Th. intermedium. No Th. intermedium chromatin was detected based on genomic in situ hybridization of mitotic chromosomes. To determine the mode of inheritance of the Bgt resistance and the chromosomal location of the resistance gene, CH09W89 was crossed with two susceptible wheat cultivars. The results of the genetic analysis showed that the adult resistance to Bgt E09 in CH09W89 was controlled by a single recessive gene, which was tentatively designated as pmCH89. Two polymorphic SSR markers, Xwmc310 and Xwmc125, were linked to the resistance gene with genetic distances 3.1 and 2.7 cM, respectively. Using the Chinese Spring aneuploid and deletion lines, the resistance gene and its linked markers were assigned to chromosome arm 4BL in the bin 0.68–0.78. Due to its unique position on chromosome 4BL, pmCH89 appears to be a new locus for resistance to powdery mildew. These results will be of benefit for improving powdery mildew resistance in wheat breeding programs.

  16. Mapping of Powdery Mildew Resistance Gene pmCH89 in a Putative Wheat-Thinopyrum intermedium Introgression Line.

    Science.gov (United States)

    Hou, Liyuan; Zhang, Xiaojun; Li, Xin; Jia, Juqing; Yang, Huizhen; Zhan, Haixian; Qiao, Linyi; Guo, Huijuan; Chang, Zhijian

    2015-07-28

    Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is a globally serious disease adversely affecting wheat production. The Bgt-resistant wheat breeding line CH09W89 was derived after backcrossing a Bgt resistant wheat-Thinopyrum intermedium partial amphiploid TAI7045 with susceptible wheat cultivars. At the seedling stage, CH09W89 exhibited immunity or high resistance to Bgt pathotypes E09, E20, E21, E23, E26, Bg1, and Bg2, similar to its donor line TAI7045 and Th. intermedium. No Th. intermedium chromatin was detected based on genomic in situ hybridization of mitotic chromosomes. To determine the mode of inheritance of the Bgt resistance and the chromosomal location of the resistance gene, CH09W89 was crossed with two susceptible wheat cultivars. The results of the genetic analysis showed that the adult resistance to Bgt E09 in CH09W89 was controlled by a single recessive gene, which was tentatively designated as pmCH89. Two polymorphic SSR markers, Xwmc310 and Xwmc125, were linked to the resistance gene with genetic distances 3.1 and 2.7 cM, respectively. Using the Chinese Spring aneuploid and deletion lines, the resistance gene and its linked markers were assigned to chromosome arm 4BL in the bin 0.68-0.78. Due to its unique position on chromosome 4BL, pmCH89 appears to be a new locus for resistance to powdery mildew. These results will be of benefit for improving powdery mildew resistance in wheat breeding programs.

  17. Computational Analysis of Breast Cancer GWAS Loci Identifies the Putative Deleterious Effect of STXBP4 and ZNF404 Gene Variants.

    Science.gov (United States)

    Masoodi, Tariq Ahmad; Banaganapalli, Babajan; Vaidyanathan, Venkatesh; Talluri, Venkateswar R; Shaik, Noor A

    2017-04-19

    The genome-wide association studies (GWAS) have enabled us in identifying different breast cancer (BC) susceptibility loci. However, majority of these are non-coding variants with no annotated biological function. We investigated such 78 noncoding genome wide associated SNPs of BC and further expanded the list to 2,162 variants with strong linkage-disequilibrium (LD, r(2) ≥0.8). Using multiple publically available algorithms such as CADD, GWAVA, and FATHAMM, we classified all these variants into deleterious, damaging, or benign categories. Out of total 2,241 variants, 23 (1.02%) variants were extreme deleterious (rank 1), 70 (3.12%) variants were deleterious (rank 2), and 1,937 (86.43%) variants were benign (rank 3). The results show 14% of lead or associated variants are under strong negative selection (GERP++ RS ≥2), and ∼22% are under balancing selection (Tajima's D score >2) in CEU population of 1KGP-the regions being positively selected (GERP++ RS <0) in mammalian evolution. The expression quantitative trait loci of highest deleteriously ranked genes were tested on relevant adipose and breast tissues, the results of which were extended for protein expression on breast tissues. From the concordance analysis of ranking system of GWAVA, CADD, and FATHMM, eQTL and protein expression, we identified the deleterious SNPs localized in STXBP4 and ZNF404 genes which might play a role in BC development by dysregulating its gene expression. This simple approach will be easier to implement and to prioritize large scale GWAS data for variety of diseases and link to the potentially unrecognized functional roles of genes. J. Cell. Biochem. 9999: 1-12, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  18. Comprehensive analysis of cystatin family genes suggests their putative functions in sexual reproduction, embryogenesis, and seed formation.

    Science.gov (United States)

    Zhao, Peng; Zhou, Xue-mei; Zou, Jie; Wang, Wei; Wang, Lu; Peng, Xiong-bo; Sun, Meng-xiang

    2014-09-01

    Cystatins are tightly bound and reversible inhibitors of cysteine proteases in C1A and C13 peptidase families, which have been identified in several species and shown to function in vegetative development and response to biotic/abiotic stresses in plants. Recent work revealed their critical role in regulating programmed cell death during embryogenesis in tobacco and suggested their more comprehensive roles in the process of sexual plant reproduction, although little is known about cystatin family genes in the processes. Here, 10 cystatin family genes in Nicotiana tabacum were identified using an expressed sequence tag (EST)-based gene clone strategy. Analysis of their biochemical properties showed that nine of them have the potency to inhibit the activities of both commercial cathepsin L-like proteases and extracted cysteine proteases from seeds, but with different K i values depending on the types of proteases and the developmental stages of the seed tested. This suggests that cystatin-dependent cathepsin L-like proteolytic pathways are probably important for early seed development. Comprehensive expression profile analysis revealed that cystatin family genes showed manifold variations in their transcription levels in different plant cell types, including the sperm, egg, and zygote, especially in the embryo and seed at different developmental stages. More interestingly, intracellular localization analysis of each cystatin revealed that most members of cystatin families are recognized as secretory proteins with signal peptides that direct them to the endoplasmic reticulum. These results suggest their widespread roles in cell fate determination and cell-cell communication in the process of sexual reproduction, especially in gamete and embryo development, as well as in seed formation.

  19. Down-regulation of BdBRI1, a putative brassinosteroid receptor gene produces a dwarf phenotype with enhanced drought tolerance in Brachypodium distachyon.

    Science.gov (United States)

    Feng, Ying; Yin, Yanhai; Fei, Shuizhang

    2015-05-01

    Brassinosteroids (BRs) play important roles in plant growth, development and responses to a range of environmental cues. Although the mechanism of how BRs regulate growth and development is well-understood in Arabidopsis, the effect of BRs on stress tolerance, particularly drought tolerance remains unknown. We isolated a BRI1 (BRASSINOSTEROID INSENSITIVE 1) homologous gene, BdBRI1 from Brachypodium distachyon, a model for temperate grasses and cereals, created and characterized RNA interference (RNAi) knockdown mutants for BdBRI1 in Brachypodium. The loss-of-function BdBRI1-RNAi mutants exhibited reduced plant height, shortened internodes, narrow and short leaf, and reduced expression of BR signaling genes, BdBES1, BdBZR1, BdBLE2, and enhanced expression of BR biosynthesis genes BdD2, BdCPD and BdDWF4. More importantly, BdBRI1 RNAi mutants exhibited enhanced drought tolerance, accompanied by highly elevated expression of drought-responsive genes, BdP5CS, BdCOR47/BdRD17, together with BdERD1 and BdRD26, two putative targets of the transcription factors BES1 and BZR1 that are key components of the BR signaling pathway. Our results suggest that BR signaling and biosynthesis are largely conserved among Arabidopsis, rice and Brachypodium, and that BR signaling plays an important role in drought tolerance by directly regulating expression of key drought-responsive genes. The effect of BR biosynthesis or crosstalks between BR and other hormones or components of stress signaling pathways on drought tolerance is discussed.

  20. Identification of putative candidate genes for red rot resistance in sugarcane (Saccharum species hybrid) using LD-based association mapping.

    Science.gov (United States)

    Singh, Ram K; Banerjee, Nandita; Khan, M S; Yadav, Sonia; Kumar, Sanjeev; Duttamajumder, S K; Lal, Ram Ji; Patel, Jinesh D; Guo, H; Zhang, Dong; Paterson, Andrew H

    2016-06-01

    Red rot is a serious disease of sugarcane caused by the fungus Colletotrichum falcatum that has a colossal damage potential. The fungus, prevalent mainly in the Indian sub-continent, keeps on producing new pathogenic strains leading to breakdown of resistance in newly released varieties and hence the deployment of linked markers for marker-assisted selection for resistance to this disease can fine tune the breeding programme. This study based on a panel of 119 sugarcane genotypes fingerprinted for 944 SSR alleles was undertaken with an aim to identify marker-trait associations (MTAs) for resistance to red rot. Mixed linear model containing population structure and kinship as co-factor detected four MTAs that were able to explain 10-16 % of the trait variation, individually. Among the four MTAs, EST sequences diagnostic of three could be BLAST searched to the sorghum genome with significant sequence homology. Several genes encoding important plant defence related proteins, viz., cytochrome P450, Glycerol-3-phosphate transporter-1, MAP Kinase-4, Serine/threonine-protein kinase, Ring finger domain protein and others were localized to the vicinity of these MTAs. These positional candidate genes are worth of further investigation and possibly these could contribute directly to red rot resistance, and may find a potential application in marker-assisted sugarcane breeding.

  1. Identification of a reproductive-specific, putative lipid transport protein gene in a queenless ponerine ant Diacamma sp.

    Science.gov (United States)

    Okada, Yasukazu; Miyazaki, Satoshi; Koshikawa, Shigeyuki; Cornette, Richard; Maekawa, Kiyoto; Tsuji, Kazuki; Miura, Toru

    2010-11-01

    Of the various characteristics of social insects, communication for reproductive differentiation is one of the most important and basic social interactions among colony members. To elucidate the molecular basis underlying this process, genes responsible for reproductive differentiation in Diacamma were screened using fluorescent differential display. Differential display, together with real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), revealed that a gene belonging to the family of cellular retinaldehyde-binding proteins was specifically expressed in the epidermis of the head, legs, and thorax in reproductives. The deduced protein sequence in the coding region, obtained by rapid amplification of cDNA ends (RACE)-PCR, was found to include cellular retinaldehyde-binding domain (CRAL-TRIO domain), suggesting that DiaCRALDCP functions in transportation of lipids, such as cuticular hydrocarbons. DiaCRALDCP transcript levels immediately decreased 1 day after the gemma mutilation, suggesting that DiaCRALDCP is involved in the physiological changes provoked by the behavioral regulation. Considering these results, the social functions of DiaCRALDCP in Diacamma are discussed.

  2. Transcriptome Analysis to Identify the Putative Biosynthesis and Transport Genes Associated with the Medicinal Components of Achyranthes bidentata Bl.

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    Jinting Li

    2016-12-01

    Full Text Available Achyranthes bidentata is a popular perennial medicine herb used for thousands of years in China to treat various diseases. Although this herb has multiple pharmaceutical purposes in China, no transcriptomic information has been reported for this species. In addition, the understanding of several key pathways and enzymes involved in the biosynthesis of oleanolic acid and ecdysterone, two pharmacologically active classes of metabolites and major chemical constituents of A. bidentata root extracts, is limited. The aim of the present study was to characterize the transcriptome profile of the roots and leaves of A. bidentata to uncover the biosynthetic and transport mechanisms of the active components. In this study, we identified 100,987 transcripts, with an average length of 973.64 base pairs. A total of 31,634 (31.33% unigenes were annotated, and 12,762 unigenes were mapped to 303 pathways according to the Kyoto Encyclopedia of Genes and Genomes (KEGG pathway database. Moreover, we identified a total of 260 oleanolic acid and ecdysterone genes encoding biosynthetic enzymes. Furthermore, the key enzymes involved in the oleanolic acid and ecdysterone synthesis pathways were analyzed using quantitative real-time polymerase chain reaction (qRT-PCR, revealing that the roots expressed these enzymes to a greater extent than the leaves. In addition, we identified 85 ATP-binding cassette (ABC transporters, some of which might be involved in the translocation of secondary metabolites.

  3. Putative extremely high rate of proteome innovation in lancelets might be explained by high rate of gene prediction errors.

    Science.gov (United States)

    Bányai, László; Patthy, László

    2016-08-01

    A recent analysis of the genomes of Chinese and Florida lancelets has concluded that the rate of creation of novel protein domain combinations is orders of magnitude greater in lancelets than in other metazoa and it was suggested that continuous activity of transposable elements in lancelets is responsible for this increased rate of protein innovation. Since morphologically Chinese and Florida lancelets are highly conserved, this finding would contradict the observation that high rates of protein innovation are usually associated with major evolutionary innovations. Here we show that the conclusion that the rate of proteome innovation is exceptionally high in lancelets may be unjustified: the differences observed in domain architectures of orthologous proteins of different amphioxus species probably reflect high rates of gene prediction errors rather than true innovation.

  4. Variation analysis of the severe acute respiratory syndrome coronavirus putative non-structural protein 2 gene and construction of three-dimensional model

    Institute of Scientific and Technical Information of China (English)

    LU Jia-hai; CHEN Wei-qing; LING Wen-hua; YU Xin-bing; ZHONG Nan-shan; ZHANG Ding-mei; WANG Guo-ling; GUO Zhong-min; ZHANG Chuan-hai; TAN Bing-yan; OUYANG Li-ping; LIN Li; LIU Yi-min

    2005-01-01

    Background The rapid transmission and high mortality rate made severe acute respiratory syndrome (SARS) a global threat for which no efficacious therapy is available now. Without sufficient knowledge about the SARS coronavirus (SARS-CoV), it is impossible to define the candidate for the anti-SARS targets. The putative non-structural protein 2 (nsp2) (3CLpro, following the nomenclature by Gao et al, also known as nsp5 in Snidjer et al) of SARS-CoV plays an important role in viral transcription and replication, and is an attractive target for anti-SARS drug development, so we carried on this study to have an insight into putative polymerase nsp2 of SARS-CoV Guangdong (GD) strain.Methods The SARS-CoV strain was isolated from a SARS patient in Guangdong, China, and cultured in Vero E6 cells. The nsp2 gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and cloned into eukaryotic expression vector pCI-neo (pCI-neo/nsp2). Then the recombinant eukaryotic expression vector pCI-neo/nsp2 was transfected into COS-7 cells using lipofectin reagent to express the nsp2 protein. The expressive protein of SARS-CoV nsp2 was analyzed by 7% sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The nucleotide sequence and protein sequence of GD nsp2 were compared with that of other SARS-CoV strains by nucleotide-nucleotide basic local alignment search tool (BLASTN) and protein-protein basic local alignment search tool (BLASTP) to investigate its variance trend during the transmission. The secondary structure of GD strain and that of other strains were predicted by Garnier-Osguthorpe-Robson (GOR) Secondary Structure Prediction. Three-dimensional-PSSM Protein Fold Recognition (Threading) Server was employed to construct the three-dimensional model of the nsp2 protein.Results The putative polymerase nsp2 gene of GD strain was amplified by RT-PCR. The eukaryotic expression vector (pCI-neo/nsp2) was constructed and expressed the protein in COS-7

  5. Characterization of a new Vaccinia virus isolate reveals the C23L gene as a putative genetic marker for autochthonous Group 1 Brazilian Vaccinia virus.

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    Felipe L Assis

    Full Text Available Since 1999, several Vaccinia virus (VACV isolates, the etiological agents of bovine vaccinia (BV, have been frequently isolated and characterized with various biological and molecular methods. The results from these approaches have grouped these VACV isolates into two different clusters. This dichotomy has elicited debates surrounding the origin of the Brazilian VACV and its epidemiological significance. To ascertain vital information to settle these debates, we and other research groups have made efforts to identify molecular markers to discriminate VACV from other viruses of the genus Orthopoxvirus (OPV and other VACV-BR groups. In this way, some genes have been identified as useful markers to discriminate between the VACV-BR groups. However, new markers are needed to infer ancestry and to correlate each sample or group with its unique epidemiological and biological features. The aims of this work were to characterize a new VACV isolate (VACV DMTV-2005 molecularly and biologically using conserved and non-conserved gene analyses for phylogenetic inference and to search for new genes that would elucidate the VACV-BR dichotomy. The VACV DMTV-2005 isolate reported in this study is biologically and phylogenetically clustered with other strains of Group 1 VACV-BR, the most prevalent VACV group that was isolated during the bovine vaccinia outbreaks in Brazil. Sequence analysis of C23L, the gene that encodes for the CC-chemokine-binding protein, revealed a ten-nucleotide deletion, which is a new Group 1 Brazilian VACV genetic marker. This deletion in the C23L open reading frame produces a premature stop-codon that is shared by all Group 1 VACV-BR strains and may also reflect the VACV-BR dichotomy; the deletion can also be considered to be a putative genetic marker for non-virulent Brazilian VACV isolates and may be used for the detection and molecular characterization of new isolates.

  6. Genes encoding putative effector proteins of the type III secretion system of Salmonella pathogenicity island 2 are required for bacterial virulence and proliferation in macrophages.

    Science.gov (United States)

    Hensel, M; Shea, J E; Waterman, S R; Mundy, R; Nikolaus, T; Banks, G; Vazquez-Torres, A; Gleeson, C; Fang, F C; Holden, D W

    1998-10-01

    The type III secretion system of Salmonella pathogenicity island 2 (SPI-2) is required for systemic infection of this pathogen in mice. Cloning and sequencing of a central region of SPI-2 revealed the presence of genes encoding putative chaperones and effector proteins of the secretion system. The predicted products of the sseB, sseC and sseD genes display weak but significant similarity to amino acid sequences of EspA, EspD and EspB, which are secreted by the type III secretion system encoded by the locus of enterocyte effacement of enteropathogenic Escherichia coli. The transcriptional activity of an sseA::luc fusion gene was shown to be dependent on ssrA, which is required for the expression of genes encoding components of the secretion system apparatus. Strains carrying nonpolar mutations in sseA, sseB or sseC were severely attenuated in virulence, strains carrying mutations in sseF or sseG were weakly attenuated, and a strain with a mutation in sseE had no detectable virulence defect. These phenotypes were reflected in the ability of mutant strains to grow within a variety of macrophage cell types: strains carrying mutations in sseA, sseB or sseC failed to accumulate, whereas the growth rates of strains carrying mutations in sseE, sseF or sseG were only modestly reduced. These data suggest that, in vivo, one of the functions of the SPI-2 secretion system is to enable intracellular bacterial proliferation.

  7. De Novo Sequencing and Analysis of the Safflower Transcriptome to Discover Putative Genes Associated with Safflor Yellow in Carthamus tinctorius L.

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    Xiuming Liu

    2015-10-01

    Full Text Available Safflower (Carthamus tinctorius L., an important traditional Chinese medicine, is cultured widely for its pharmacological effects, but little is known regarding the genes related to the metabolic regulation of the safflower’s yellow pigment. To investigate genes related to safflor yellow biosynthesis, 454 pyrosequencing of flower RNA at different developmental stages was performed, generating large databases.In this study, we analyzed 454 sequencing data from different flowering stages in safflower. In total, 1,151,324 raw reads and 1,140,594 clean reads were produced, which were assembled into 51,591 unigenes with an average length of 679 bp and a maximum length of 5109 bp. Among the unigenes, 40,139 were in the early group, 39,768 were obtained from the full group and 28,316 were detected in both samples. With the threshold of “log2 ratio ≥ 1”, there were 34,464 differentially expressed genes, of which 18,043 were up-regulated and 16,421 were down-regulated in the early flower library. Based on the annotations of the unigenes, 281 pathways were predicted. We selected 12 putative genes and analyzed their expression levels using quantitative real time-PCR. The results were consistent with the 454 sequencing results. In addition, the expression of chalcone synthase, chalcone isomerase and anthocyanidin synthase, which are involved in safflor yellow biosynthesis and safflower yellow pigment (SYP content, were analyzed in different flowering periods, indicating that their expression levels were related to SYP synthesis. Moreover, to further confirm the results of the 454 pyrosequencing, full-length cDNA of chalcone isomerase (CHI and anthocyanidin synthase (ANS were cloned from safflower petal by RACE (Rapid-amplification of cDNA ends method according to fragment of the transcriptome.

  8. Analysis of the transcriptome of Erigeron breviscapus uncovers putative scutellarin and chlorogenic acids biosynthetic genes and genetic markers.

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    Ni-Hao Jiang

    Full Text Available Erigeron breviscapus (Vant. Hand-Mazz. is a famous medicinal plant. Scutellarin and chlorogenic acids are the primary active components in this herb. However, the mechanisms of biosynthesis and regulation for scutellarin and chlorogenic acids in E. breviscapus are considerably unknown. In addition, genomic information of this herb is also unavailable.Using Illumina sequencing on GAIIx platform, a total of 64,605,972 raw sequencing reads were generated and assembled into 73,092 non-redundant unigenes. Among them, 44,855 unigenes (61.37% were annotated in the public databases Nr, Swiss-Prot, KEGG, and COG. The transcripts encoding the known enzymes involved in flavonoids and in chlorogenic acids biosynthesis were discovered in the Illumina dataset. Three candidate cytochrome P450 genes were discovered which might encode flavone 6-hydroase converting apigenin to scutellarein. Furthermore, 4 unigenes encoding the homologues of maize P1 (R2R3-MYB transcription factors were defined, which might regulate the biosynthesis of scutellarin. Additionally, a total of 11,077 simple sequence repeat (SSR were identified from 9,255 unigenes. Of SSRs, tri-nucleotide motifs were the most abundant motif. Thirty-six primer pairs for SSRs were randomly selected for validation of the amplification and polymorphism. The result revealed that 34 (94.40% primer pairs were successfully amplified and 19 (52.78% primer pairs exhibited polymorphisms.Using next generation sequencing (NGS technology, this study firstly provides abundant genomic data for E. breviscapus. The candidate genes involved in the biosynthesis and transcriptional regulation of scutellarin and chlorogenic acids were obtained in this study. Additionally, a plenty of genetic makers were generated by identification of SSRs, which is a powerful tool for molecular breeding and genetics applications in this herb.

  9. Virus induced gene silencing of three putative prolyl 4-hydroxylases enhances plant growth in tomato (Solanum lycopersicum).

    Science.gov (United States)

    Fragkostefanakis, Sotirios; Sedeek, Khalid E M; Raad, Maya; Zaki, Marwa Samir; Kalaitzis, Panagiotis

    2014-07-01

    Proline hydroxylation is a major posttranslational modification of hydroxyproline-rich glycoproteins (HRGPs) that is catalyzed by prolyl 4-hydroxylases (P4Hs). HRGPs such as arabinogalactan proteins (AGPs) and extensios play significant roles on cell wall structure and function and their implication in cell division and expansion has been reported. We used tobacco rattle virus (TRV)-based virus induced gene silencing to investigate the role of three tomato P4Hs, out of ten present in the tomato genome, in growth and development. Eight-days old tomato seedlings were infected with the appropriate TRV vectors and plants were allowed to grow under standard conditions for 6 weeks. Lower P4H mRNA levels were associated with lower hydroxyproline content in root and shoot tissues indicating successful gene silencing. P4H-silenced plants had longer roots and shoots and larger leaves. The increased leaf area can be attributed to increased cell division as indicated by the higher leaf epidermal cell number in SlP4H1- and SlP4H9-silenced plants. In contrast, SlP4H7-silenced plants had larger leaves due to enhanced cell expansion. Western blot analysis revealed that silencing of SlP4H7 and SlP4H9 was associated with reduced levels of JIM8-bound AGP and JIM11-bound extensin epitopes, while silencing of SlP4H1 reduced only the levels of AGP proteins. Collectively these results show that P4Hs have significant and distinct roles in cell division and expansion of tomato leaves.

  10. Identification and analysis of unitary loss of long-established protein-coding genes in Poaceae shows evidences for biased gene loss and putatively functional transcription of relics.

    Science.gov (United States)

    Zhao, Yi; Tang, Liang; Li, Zhe; Jin, Jinpu; Luo, Jingchu; Gao, Ge

    2015-04-18

    Long-established protein-coding genes may lose their coding potential during evolution ("unitary gene loss"). Members of the Poaceae family are a major food source and represent an ideal model clade for plant evolution research. However, the global pattern of unitary gene loss in Poaceae genomes as well as the evolutionary fate of lost genes are still less-investigated and remain largely elusive. Using a locally developed pipeline, we identified 129 unitary gene loss events for long-established protein-coding genes from four representative species of Poaceae, i.e. brachypodium, rice, sorghum and maize. Functional annotation suggested that the lost genes in all or most of Poaceae species are enriched for genes involved in development and response to endogenous stimulus. We also found that 44 mutated genomic loci of lost genes, which we referred as relics, were still actively transcribed, and of which 84% (37 of 44) showed significantly differential expression across different tissues. More interestingly, we found that there were totally five expressed relics may function as competitive endogenous RNA in brachypodium, rice and sorghum genome. Based on comparative genomics and transcriptome data, we firstly compiled a comprehensive catalogue of unitary gene loss events in Poaceae species and characterized a statistically significant functional preference for these lost genes as well showed the potential of relics functioning as competitive endogenous RNAs in Poaceae genomes.

  11. Gene ercA, encoding a putative iron-containing alcohol dehydrogenase, is involved in regulation of ethanol utilization in Pseudomonas aeruginosa.

    Science.gov (United States)

    Hempel, Niels; Görisch, Helmut; Mern, Demissew S

    2013-09-01

    Several two-component regulatory systems are known to be involved in the signal transduction pathway of the ethanol oxidation system in Pseudomonas aeruginosa ATCC 17933. These sensor kinases and response regulators are organized in a hierarchical manner. In addition, a cytoplasmic putative iron-containing alcohol dehydrogenase (Fe-ADH) encoded by ercA (PA1991) has been identified to play an essential role in this regulatory network. The gene ercA (PA1991) is located next to ercS, which encodes a sensor kinase. Inactivation of ercA (PA1991) by insertion of a kanamycin resistance cassette created mutant NH1. NH1 showed poor growth on various alcohols. On ethanol, NH1 grew only with an extremely extended lag phase. During the induction period on ethanol, transcription of structural genes exa and pqqABCDEH, encoding components of initial ethanol oxidation in P. aeruginosa, was drastically reduced in NH1, which indicates the regulatory function of ercA (PA1991). However, transcription in the extremely delayed logarithmic growth phase was comparable to that in the wild type. To date, the involvement of an Fe-ADH in signal transduction processes has not been reported.

  12. ZDHHC8 as a candidate gene for schizophrenia: Analysis of a putative functional intronic marker in case-control and family-based association studies

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    Jabs Burkhard

    2005-10-01

    Full Text Available Abstract Background The chromosome 22q11 region is proposed as a major candidate locus for susceptibility genes to schizophrenia. Recently, the gene ZDHHC8 encoding a putative palmitoyltransferase at 22q11 was proposed to increase liability to schizophrenia based on both animal models and human association studies by significant over-transmission of allele rs175174A in female, but not male subjects with schizophrenia. Methods Given the genetic complexity of schizophrenia and the potential genetic heterogeneity in different populations, we examined rs175174 in 204 German proband-parent triads and in an independent case-control study (schizophrenic cases: n = 433; controls: n = 186. Results In the triads heterozygous parents transmitted allele G preferentially to females, and allele A to males (heterogeneity χ2 = 4.43; p = 0.035. The case-control sample provided no further evidence for overall or gender-specific effects regarding allele and genotype frequency distributions. Conclusion The findings on rs175174 at ZDHHC8 are still far from being conclusive, but evidence for sexual dimorphism is moderate, and our data do not support a significant genetic contribution of rs175174 to the aetiopathogenesis of schizophrenia.

  13. Phenotypes of gene disruptants in relation to a putative mitochondrial malate-citrate shuttle protein in citric acid-producing Aspergillus niger.

    Science.gov (United States)

    Kirimura, Kohtaro; Kobayashi, Keiichi; Ueda, Yuka; Hattori, Takasumi

    2016-09-01

    The mitochondrial citrate transport protein (CTP) functions as a malate-citrate shuttle catalyzing the exchange of citrate plus a proton for malate between mitochondria and cytosol across the inner mitochondrial membrane in higher eukaryotic organisms. In this study, for functional analysis, we cloned the gene encoding putative CTP (ctpA) of citric acid-producing Aspergillus niger WU-2223L. The gene ctpA encodes a polypeptide consisting 296 amino acids conserved active residues required for citrate transport function. Only in early-log phase, the ctpA disruptant DCTPA-1 showed growth delay, and the amount of citric acid produced by strain DCTPA-1 was smaller than that by parental strain WU-2223L. These results indicate that the CTPA affects growth and thereby citric acid metabolism of A. niger changes, especially in early-log phase, but not citric acid-producing period. This is the first report showing that disruption of ctpA causes changes of phenotypes in relation to citric acid production in A. niger.

  14. Identification of putative chemosensory receptor genes from yellow peach moth Conogethes punctiferalis (Guenée) antennae transcriptome

    Science.gov (United States)

    Ge, Xing; Zhang, Tiantao; Wang, Zhenying; He, Kanglai; Bai, Shuxiong

    2016-01-01

    The yellow peach moth, Conogethes punctiferalis, is an extremely important polyphagous insect in Asia. The chemosensory systems of moth play an important role in detecting food, oviposition sites and mate attraction. Several antennal chemosensory receptors are involved in odor detection. Our study aims to identify chemosensory receptor genes for potential applications in behavioral responses of yellow peach moth. By transcriptomic analysis of male and female antennae, 83 candidate chemosensory receptors, including 62 odorant receptors, 11 ionotropic receptors and 10 gustatory receptors were identified. Through Blast and sequence alignment, the highly conserved co-receptor Orco was annotated, eight unigenes clustered into pheromone receptors, and two clustered as sugar receptor. Among the IRs, one unigenes was similar with co-receptors IR25a. Expression levels of 50 odorant receptors were further evaluated by quantitative real-time PCR in antennae. All the ORs tested were detected in antennae and some of which were associated with sex-biased expression. The chemosensory receptors identified in C. punctiferalis provide a foundational resource for further analysis on olfaction for behavior. The expression profiles of ORs in antennae indicated variant functions in olfactory recognition, and our results provided the possibility for the potential application of semiochemical to control this pest moth. PMID:27659493

  15. Mycobacterium tuberculosis DosR regulon gene Rv0079 encodes a putative, 'dormancy associated translation inhibitor (DATIN'.

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    Ashutosh Kumar

    Full Text Available Mycobacterium tuberculosis is a major human pathogen that has evolved survival mechanisms to persist in an immune-competent host under a dormant condition. The regulation of M. tuberculosis metabolism during latent infection is not clearly known. The dormancy survival regulon (DosR regulon is chiefly responsible for encoding dormancy related functions of M. tuberculosis. We describe functional characterization of an important gene of DosR regulon, Rv0079, which appears to be involved in the regulation of translation through the interaction of its product with bacterial ribosomal subunits. The protein encoded by Rv0079, possibly, has an inhibitory role with respect to protein synthesis, as revealed by our experiments. We performed computational modelling and docking simulation studies involving the protein encoded by Rv0079 followed by in vitro translation and growth curve analysis experiments, involving recombinant E. coli and Bacille Calmette Guérin (BCG strains that overexpressed Rv0079. Our observations concerning the interaction of the protein with the ribosomes are supportive of its role in regulation/inhibition of translation. We propose that the protein encoded by locus Rv0079 is a 'dormancy associated translation inhibitor' or DATIN.

  16. Relative gene expression of bile salt hydrolase and surface proteins in two putative indigenous Lactobacillus plantarum strains under in vitro gut conditions.

    Science.gov (United States)

    Duary, Raj Kumar; Batish, Virender Kumar; Grover, Sunita

    2012-03-01

    Probiotic bacteria must overcome the toxicity of bile salts secreted in the gut and adhere to the epithelial cells to enable their better colonization with extended transit time. Expression of bile salt hydrolase and other proteins on the surface of probiotic bacteria can help in better survivability and optimal functionality in the gut. Two putative Lactobacillus plantarum isolates i.e., Lp9 and Lp91 along with standard strain CSCC5276 were used. A battery of six housekeeping genes viz. gapB, dnaG, gyrA, ldhD, rpoD and 16S rRNA were evaluated by using geNorm 3.4 excel based application for normalizing the expression of bile salt hydrolase (bsh), mucus-binding protein (mub), mucus adhesion promoting protein (mapA), and elongation factor thermo unstable (EF-Tu) in Lp9 and Lp91. The maximal level of relative bsh gene expression was recorded in Lp91 with 2.89 ± 0.14, 4.57 ± 0.37 and 6.38 ± 0.19 fold increase at 2% bile salt concentration after 1, 2 and 3 h, respectively. Similarly, mub and mapA genes were maximally expressed in Lp9 at the level of 20.07 ± 1.28 and 30.92 ± 1.51 fold, when MRS was supplemented with 0.05% mucin and 1% each of bile and pancreatin (pH 6.5). However, in case of EF-Tu, the maximal expression of 42.84 ± 5.64 fold was recorded in Lp91 in the presence of mucin alone (0.05%). Hence, the expression of bsh, mub, mapA and EF-Tu could be considered as prospective biomarkers for screening of novel probiotic lactobacillus strains for optimal functionality in the gut.

  17. A putative glucan synthase gene dps detected in exopolysaccharide-producing Pediococcus damnosus and Oenococcus oeni strains isolated from wine and cider.

    Science.gov (United States)

    Walling, Emilie; Gindreau, Emmanuel; Lonvaud-Funel, Aline

    2005-01-15

    Some lactic acid bacteria can induce viscosity in wine, beer and cider by production of exopolysaccharides (EPS). A polymerase chain reaction (PCR) assay was previously described for the detection of ropy Pediococcus damnosus strains in wine [J. Appl. Microbiol. 90 (2001) 535]. The primers used in that study, PF5 and PF6, are investigated in addition to new primers which broaden the range of spoiling agents detectable by PCR. Primers PF1 and PF8 allow the amplification of DNA from ropy P. damnosus strains isolated from wine, as was observed with PF5 and PF6. In addition, PF1 and PF8, unlike PF5 and PF6, are able to generate an amplicon using template DNA from a ropy P. damnosus strain isolated from cider and a ropy Oenococcus oeni strain isolated from champagne. Two different ropy Lactobacillus species were also isolated, but their DNA was not amplified using primers PF1 and PF8. The new primers PF1 and PF8 were chosen from the sequence of gene dps, a putative glucan synthase gene, found across all the ropy P. damnosus strains isolated, from both wine or cider, and also in a ropy O. oeni strain. To our knowledge, this is the first time that an EPS-producing O. oeni strain is described. Glucan biosynthesis was assessed by agglutination tests done with Streptococcus pneumoniae type 37-specific antibodies, which specifically detect glucan-producing cells. The results show that there is a direct correlation between glucan production and detection of gene dps. Therefore, Dps is considered a candidate for the glucan synthase enzyme responsible for EPS production by ropy strains of P. damnosus and O. oeni.

  18. Cloning and characterization of a putative human holocytochrome c-type synthetase gene (HCCS) isolated from the critical region for microphthalmia with linear skin defects (MLS)

    Energy Technology Data Exchange (ETDEWEB)

    Schaefer, L.; Ballabio, A.; Zoghbi, H.Y. [Baylor College of Medicine, Houston, TX (United States)

    1996-06-01

    Microphthalmia with linear skin defects syndrome (MLS) is an X-linked male-lethal disorder associated with X chromosomal rearrangements resulting in monosomy from Xpter to Xp22. Features include microphthalmia, sclerocornea, linear skin defects, and agenesis of the corpus callosum. Using a cross-species conservation strategy, an expressed sequence from the 450- to the 550-kb MLS critical region on Xp22 was identified by screening a human embryo cDNA library. Northern analysis revealed a transcript of {approx}2.6 kb in all tissues examined, with weaker expression of {approx}1.2- and {approx}5.2-kb transcripts. The strongest expression was observed in heart and skeletal muscle. Sequence analysis of a 3-kb cDNA contig revealed an 807-bp open reading frame encoding a putative 268-amino-acid-protein. Comparison of the sequence with sequences in the databases revealed homology with holocytochrome c-type synthetases, which catalyze the covalent addition of a heme group onto c-type cytochromes in the mitochondria. The c-type cytochromes are required for proper functioning of the electron transport pathway. The human gene (HGMW-approved symbol HCCS) and the corresponding murine gene characterized in this paper are the first mammalian holocytochrome c-type synthetases to be described in the literature. Because of the lack of a neuromuscular phenotype in MLS, it is uncertain whether the deletion of a mitochondrial holocytochrome synthetase would contribute to the phenotype seen in MLS. The expression pattern of this gene and knowledge about the function of holocytochrome synthetases, however, suggest that it is a good candidate for X-linked encephalomyopathies typically associated with mitochondrial dysfunction. 25 refs., 4 figs.

  19. De novo assembly of the Indo-Pacific humpback dolphin leucocyte transcriptome to identify putative genes involved in the aquatic adaptation and immune response.

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    Duan Gui

    Full Text Available BACKGROUND: The Indo-Pacific humpback dolphin (Sousa chinensis, a marine mammal species inhabited in the waters of Southeast Asia, South Africa and Australia, has attracted much attention because of the dramatic decline in population size in the past decades, which raises the concern of extinction. So far, this species is poorly characterized at molecular level due to little sequence information available in public databases. Recent advances in large-scale RNA sequencing provide an efficient approach to generate abundant sequences for functional genomic analyses in the species with un-sequenced genomes. PRINCIPAL FINDINGS: We performed a de novo assembly of the Indo-Pacific humpback dolphin leucocyte transcriptome by Illumina sequencing. 108,751 high quality sequences from 47,840,388 paired-end reads were generated, and 48,868 and 46,587 unigenes were functionally annotated by BLAST search against the NCBI non-redundant and Swiss-Prot protein databases (E-value<10(-5, respectively. In total, 16,467 unigenes were clustered into 25 functional categories by searching against the COG database, and BLAST2GO search assigned 37,976 unigenes to 61 GO terms. In addition, 36,345 unigenes were grouped into 258 KEGG pathways. We also identified 9,906 simple sequence repeats and 3,681 putative single nucleotide polymorphisms as potential molecular markers in our assembled sequences. A large number of unigenes were predicted to be involved in immune response, and many genes were predicted to be relevant to adaptive evolution and cetacean-specific traits. CONCLUSION: This study represented the first transcriptome analysis of the Indo-Pacific humpback dolphin, an endangered species. The de novo transcriptome analysis of the unique transcripts will provide valuable sequence information for discovery of new genes, characterization of gene expression, investigation of various pathways and adaptive evolution, as well as identification of genetic markers.

  20. Isolation and Expression Analysis of Two Cold-Inducible Genes Encoding Putative CBF Transcription Factors from Chinese Cabbage (Brassica pekinensis Rupr.)

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Two homologous genes of the Arabidopsis C-repeat/dehydration-responsive element binding factors (CBF/DREB1) transcriptional activator were isolated by RT-PCR from Chinese cabbage (Brassica pekinensis Rupr.cv. Qinbai 5) and were designated as BcCBF1 and BcCBF2. Each encodes a putative CBF/DREB1 protein with an AP2 (Apetal2) DNA-binding domain, a putative nuclear localization signal, and a possible acidic activation domain. Deduced amino acid sequences show that BcCBF1 is very similar to the Arabidopsis CBF1, whereas BcCBF2 is different in that it contains two extra regions of 24 and 20 amino acids in the acidic domain. The mRNA accumulation profiles indicated that the expression of BcCBF1 and BcCBF2 is strongly induced by cold treatment, but does not respond similarly to dehydration or abscisic acid (ABA) treatment. However,the cold-induced accumulation of BcCBF2 mRNA was rapid but short-lived compared with that of BcCBF1.The mRNA levels of both BcCBF1 and BcCBF2 were higher in leaves than in roots when plants were exposed to cold, whereas, salt stress caused higher accumulation of BcCBF2 mRNA in roots than in leaves,suggesting that the organ specificity of the gene expression of the BcCBFs is probably stress dependent.In addition, the accumulation of BcCBF1 and BcCBF2 mRNAs was greatly enhanced by light compared with darkness when seedlings were exposed to cold. It is concluded that the two BcCBF proteins may be involved in the process of plant response to cold stress through an ABA-independent pathway and that there is also a cross-talk between the light signaling conduction pathway and the cold response pathway in B. pekinensis as In Arabidopsis.

  1. Overexpression of a novel Arabidopsis gene related to putative zinc-transporter genes from animals can lead to enhanced zinc resistance and accumulation.

    Science.gov (United States)

    van der Zaal, B J; Neuteboom, L W; Pinas, J E; Chardonnens, A N; Schat, H; Verkleij, J A; Hooykaas, P J

    1999-03-01

    We describe the isolation of an Arabidopsis gene that is closely related to the animal ZnT genes (Zn transporter). The protein encoded by the ZAT (Zn transporter of Arabidopsis thaliana) gene has 398 amino acid residues and is predicted to have six membrane-spanning domains. To obtain evidence for the postulated function of the Arabidopsis gene, transgenic plants with the ZAT coding sequence under control of the cauliflower mosaic virus 35S promoter were analyzed. Plants obtained with ZAT in the sense orientation exhibited enhanced Zn resistance and strongly increased Zn content in the roots under high Zn exposure. Antisense mRNA-producing plants were viable, with a wild-type level of Zn resistance and content, like plants expressing a truncated coding sequence lacking the C-terminal cytoplasmic domain of the protein. The availability of ZAT can lead to a better understanding of the mechanism of Zn homeostasis and resistance in plants.

  2. Avocado cellulase: nucleotide sequence of a putative full-length cDNA clone and evidence for a small gene family.

    Science.gov (United States)

    Tucker, M L; Durbin, M L; Clegg, M T; Lewis, L N

    1987-05-01

    A cDNA library was prepared from ripe avocado fruit (Persea americana Mill. cv. Hass) and screened for clones hybridizing to a 600 bp cDNA clone (pAV5) coding for avocado fruit cellulase. This screening led to the isolation of a clone (pAV363) containing a 2021 nucleotide transcribed sequence and an approximately 150 nucleotide poly(A) tail. Hybridization of pAV363 to a northern blot shows that the length of the homologous message is approximately 2.2 kb. The nucleotide sequence of this putative full-length mRNA clone contains an open reading frame of 1482 nucleotides which codes for a polypeptide of 54.1 kD. The deduced amino acid composition compares favorably with the amino acid composition of native avocado cellulase determined by amino acid analysis. Southern blot analysis of Hind III and Eco RI endonuclease digested genomic DNA indicates a small family of cellulase genes.

  3. Distribution of genes encoding putative virulence factors and fragment length polymorphisms in the vrrA gene among Brazilian isolates of Bacillus cereus and Bacillus thuringiensis.

    Science.gov (United States)

    Zahner, Viviane; Cabral, Diana Aparecida; Régua-Mangia, Adriana Hamond; Rabinovitch, Leon; Moreau, Gaétan; McIntosh, Douglas

    2005-12-01

    One hundred twenty-one strains of the Bacillus cereus complex, of which 80 were isolated from a variety of sources in Brazil, were screened by PCR for the presence of sequences (bceT, hblA, nheBC, plc, sph, and vip3A) encoding putative virulence factors and for polymorphisms in variable-number tandem repeats (VNTR), using a variable region of the vrrA open reading frame as the target. Amplicons were generated from isolates of B. cereus and Bacillus thuringiensis for each of the sequences encoding factors suggested to play a role in infections of mammals. Intriguingly, the majority of these sequences were detected more frequently in Bacillus thuringiensis than in B. cereus. The vip3A sequence, which encodes an insecticidal toxin, was detected exclusively in B. thuringiensis. VNTR analysis demonstrated the presence of five different fragment length categories in both species, with two of these being widely distributed throughout both taxa. In common with data generated from previous studies examining European, Asian, or North American populations, our investigation of Brazilian isolates supports the notion that B. cereus and B. thuringiensis should be considered to represent a single species.

  4. Genomic diversity of Mycobacterium tuberculosis Beijing strains isolated in Tuscany, Italy, based on large sequence deletions, SNPs in putative DNA repair genes and MIRU-VNTR polymorphisms.

    Science.gov (United States)

    Garzelli, Carlo; Lari, Nicoletta; Rindi, Laura

    2016-03-01

    The Beijing genotype of Mycobacterium tuberculosis is cause of global concern as it is rapidly spreading worldwide, is considered hypervirulent, and is most often associated to massive spread of MDR/XDR TB, although these epidemiological or pathological properties have not been confirmed for all strains and in all geographic settings. In this paper, to gain new insights into the biogeographical heterogeneity of the Beijing family, we investigated a global sample of Beijing strains (22% from Italian-born, 78% from foreign-born patients) by determining large sequence polymorphism of regions RD105, RD181, RD150 and RD142, single nucleotide polymorphism of putative DNA repair genes mutT4 and mutT2 and MIRU-VNTR profiles based on 11 discriminative loci. We found that, although our sample of Beijing strains showed a considerable genomic heterogeneity, yielding both ancient and recent phylogenetic strains, the prevalent successful Beijing subsets were characterized by deletions of RD105 and RD181 and by one nucleotide substitution in one or both mutT genes. MIRU-VNTR analysis revealed 47 unique patterns and 9 clusters including a total of 33 isolates (41% of total isolates); the relatively high proportion of Italian-born Beijing TB patients, often occurring in mixed clusters, supports the possibility of an ongoing cross-transmission of the Beijing genotype to autochthonous population. High rates of extra-pulmonary localization and drug-resistance, particularly MDR, frequently reported for Beijing strains in other settings, were not observed in our survey.

  5. Discovery of precursor and mature microRNAs and their putative gene targets using high-throughput sequencing in pineapple (Ananas comosus var. comosus).

    Science.gov (United States)

    Yusuf, Noor Hydayaty Md; Ong, Wen Dee; Redwan, Raimi Mohamed; Latip, Mariam Abd; Kumar, S Vijay

    2015-10-15

    MicroRNAs (miRNAs) are a class of small, endogenous non-coding RNAs that negatively regulate gene expression, resulting in the silencing of target mRNA transcripts through mRNA cleavage or translational inhibition. MiRNAs play significant roles in various biological and physiological processes in plants. However, the miRNA-mediated gene regulatory network in pineapple, the model tropical non-climacteric fruit, remains largely unexplored. Here, we report a complete list of pineapple mature miRNAs obtained from high-throughput small RNA sequencing and precursor miRNAs (pre-miRNAs) obtained from ESTs. Two small RNA libraries were constructed from pineapple fruits and leaves, respectively, using Illumina's Solexa technology. Sequence similarity analysis using miRBase revealed 579,179 reads homologous to 153 miRNAs from 41 miRNA families. In addition, a pineapple fruit transcriptome library consisting of approximately 30,000 EST contigs constructed using Solexa sequencing was used for the discovery of pre-miRNAs. In all, four pre-miRNAs were identified (MIR156, MIR399, MIR444 and MIR2673). Furthermore, the same pineapple transcriptome was used to dissect the function of the miRNAs in pineapple by predicting their putative targets in conjunction with their regulatory networks. In total, 23 metabolic pathways were found to be regulated by miRNAs in pineapple. The use of high-throughput sequencing in pineapples to unveil the presence of miRNAs and their regulatory pathways provides insight into the repertoire of miRNA regulation used exclusively in this non-climacteric model plant.

  6. Cloning and characterisation of a putative pollen-specific polygalacturonase gene (CpPG1) differentially regulated during pollen development in zucchini (Cucurbita pepo L.).

    Science.gov (United States)

    Carvajal, F; Garrido, D; Jamilena, M; Rosales, R

    2014-03-01

    Studies in zucchini (Cucurbita pepo L. spp. pepo) pollen have been limited to the viability and morphology of the mature pollen grain. The enzyme polygalacturonase (PG) is involved in pollen development and pollination in many species. In this work, we study anther and pollen development of C. pepo and present the cloning and characterisation of a putative PG CpPG1 (Accession no. HQ232488) from pollen cDNA in C. pepo. The predicted protein for CpPG1 has 416 amino acids, with a high homology to other pollen PGs, such as P22 from Oenothera organensis (76%) and PGA3 from Arabidopsis thaliana (73%). CpPG1 belongs to clade C, which comprises PGs expressed in pollen, and presents a 34 amino acid signal peptide for secretion towards the cell wall. DNA-blot analysis revealed that there are at least another two genes that code for PGs in C. pepo. The spatial and temporal accumulation of CpPG1 was studied by semi-quantitative- and qRT-PCR. In addition, mRNA was detected only in anthers, pollen and the rudimentary anthers of bisexual flowers (only present in some zucchini cultivars under certain environmental conditions that trigger anther development in the third whorl of female flowers). However, no expression was detected in cotyledons, stem or fruit. Furthermore, CpPG1 mRNA was accumulated throughout anther development, with the highest expression found in mature pollen. Similarly, exo-PG activity increased from immature anther stages to mature anthers and mature pollen. Overall, these data support the pollen specificity of this gene and suggest an involvement of CpPG1 in pollen development in C. pepo.

  7. A putatively phase variable gene (dca) required for natural competence in Neisseria gonorrhoeae but not Neisseria meningitidis is located within the division cell wall (dcw) gene cluster.

    Science.gov (United States)

    Snyder, L A; Saunders, N J; Shafer, W M

    2001-02-01

    A cluster of 18 open reading frames (ORFs), 15 of which are homologous to genes involved in division and cell wall synthesis, has been identified in Neisseria gonorrhoeae and Neisseria meningitidis. The three additional ORFs, internal to the dcw cluster, are not homologous to dcw-related genes present in other bacterial species. Analysis of the N. meningitidis strain MC58 genome for foreign DNA suggests that these additional ORFs have not been acquired by recent horizontal exchange, indicating that they are a long-standing, integral part of the neisserial dcw gene cluster. Reverse transcription-PCR analysis of RNA extracted from N. gonorrhoeae strain FA19 confirmed that all three ORFs are transcribed in gonococci. One of these ORFs (dca, for division cluster competence associated), located between murE and murF, was studied in detail and found to be essential for competence in the gonococcal but not in the meningococcal strains tested. Computer analysis predicts that dca encodes an inner membrane protein similar to hypothetical proteins produced by other gram-negative bacteria. In some meningococcal strains dca is prematurely terminated following a homopolymeric tract of G's, the length of which differs between isolates of N. meningitidis, suggesting that dca is phase variable in this species. A deletion and insertional mutation was made in the dca gene of N. gonorrhoeae strain FA19 and N. meningitidis strain NMB. This mutation abrogated the ability of the gonococci to be transformed with chromosomal DNA. Thus, we conclude that the dca-encoded gene product is an essential competence factor for gonococci.

  8. Arabidopsis SOI33/AtENT8 Gene Encodes a Putative Equilibrative Nucleoside Transporter That Is Involved in Cytokinin Transport In Planta

    Institute of Scientific and Technical Information of China (English)

    Jiaqiang SUN; Naoya HIROSE; Xingchun WANG; Pei WEN; Li XUE; Hitoshi SAKAKIBARA; Jianru ZUO

    2005-01-01

    The plant phytohormone cytokinin plays an important role in many facets of plant growth and development by regulating cell division and differentiation. Recent studies have shed significant light into the mechanisms of cytokinin metabolism and signaling. However, little is known about how the hormone is transported in planta, although it has been proposed that the hormone is presumably transported in nucleoside-conjugated forms. Here, we report the identification and characterization of cytokinin transport ers in Arabidopsis. We previously reported that a gain-of-function mutation in the PGA22/AtIPT8 gene caused overproduction of cytokinins in planta. In an effort to screen for suppressor of pga22/atipt8 (soi) mutants, we identified a mutant soi33-1. Molecular and genetic analyses indicated that SOI33 encodes a putative equilibrative nucleoside transporter (ENT), previously designated as AtENT8. Members of this small gene family are presumed to be involved in the transport of nucleosides in eukaryotic cells. Under conditions of nitrogen starvation, loss-of-function mutations in SOI33/AtENT8 or in a related gene AtENT3 cause a reduced sensitivity to the nucleoside-type cytokinins isopentenyladenine riboside (iPR) and trans zeatin riboside (tZR), but display a normal response to the free base-type cytokinins isopentenyladenine (iP) and trans-zeatin (tZ). Conversely, overexpression of SOI33/AtENT8 renders transgenic plants hyper sensitive to iPR but not to iP. An in planta measurement experiment indicated that uptake efficiency of 3H labeled iPR was reduced more than 40% in soi33 and atent3 mutants. However, a mutation inAtENT1 had no substantial effect on the cytokinin response and iPR uptake efficiency. Our results suggest that SOI33/ AtENT8 and AtENT3 are involved in the transport of nucleoside-type cytokinins in Arabidopsis.

  9. Hypermethylation of BDNF and SST Genes in the Orbital Frontal Cortex of Older Individuals: A Putative Mechanism for Declining Gene Expression with Age.

    Science.gov (United States)

    McKinney, Brandon C; Lin, Chien-Wei; Oh, Hyunjung; Tseng, George C; Lewis, David A; Sibille, Etienne

    2015-10-01

    Expression of brain-derived neurotrophic factor (BDNF) and somatostatin (SST) mRNAs in the brain decreases progressively and robustly with age, and lower BDNF and SST expression in the brain has been observed in many brain disorders. BDNF is known to regulate SST expression; however, the mechanisms underlying decreased expression of both genes are not understood. DNA methylation (DNAm) is an attractive candidate mechanism. To investigate the contribution of DNAm to the age-related decline in BDNF and SST expression, the Illumina Infinium HumanMethylation450 Beadchip Array was used to quantify DNAm of BDNF (26 CpG loci) and SST (9 CpG loci) in the orbital frontal cortices of postmortem brains from 22 younger (age 60 years) with known age-dependent BDNF and SST expression differences. Relative to the younger individuals, 10 of the 26 CpG loci in BDNF and 8 of the 9 CpG loci in SST were significantly hypermethylated in the older individuals. DNAm in BDNF exons/promoters I, II, and IV negatively correlated with BDNF expression (r=-0.37, pgenes exhibited similar age-related changes in DNAm and correlation with gene expression. These results suggest that DNAm may be a proximal mechanism for decreased expression of BDNF, SST, and other BDNF- and GABA-related genes with brain aging and, by extension, for brain disorders in which their expression is decreased.

  10. Identification and characterization of potential NBS-encoding resistance genes and induction kinetics of a putative candidate gene associated with downy mildew resistance in Cucumis

    Directory of Open Access Journals (Sweden)

    Wan Hongjian

    2010-08-01

    Full Text Available Abstract Background Due to the variation and mutation of the races of Pseudoperonospora cubensis, downy mildew has in recent years become the most devastating leaf disease of cucumber worldwide. Novel resistance to downy mildew has been identified in the wild Cucumis species, C. hystrix Chakr. After the successful hybridization between C. hystrix and cultivated cucumber (C. sativus L., an introgression line (IL5211S was identified as highly resistant to downy mildew. Nucleotide-binding site and leucine-rich repeat (NBS-LRR genes are the largest class of disease resistance genes cloned from plant with highly conserved domains, which can be used to facilitate the isolation of candidate genes associated with downy mildew resistance in IL5211S. Results Degenerate primers that were designed based on the conserved motifs in the NBS domain of resistance (R proteins were used to isolate NBS-type sequences from IL5211S. A total of 28 sequences were identified and named as cucumber (C. sativus = CS resistance gene analogs as CSRGAs. Polygenetic analyses separated these sequences into four different classes. Quantitative real-time polymerase chain reaction (qRT-PCR analysis showed that these CSRGAs expressed at different levels in leaves, roots, and stems. In addition, introgression from C. hystrix induced expression of the partial CSRGAs in cultivated cucumber, especially CSRGA23, increased four-fold when compared to the backcross parent CC3. Furthermore, the expression of CSRGA23 under P. cubensis infection and abiotic stresses was also analyzed at different time points. Results showed that the P. cubensis treatment and four tested abiotic stimuli, MeJA, SA, ABA, and H2O2, triggered a significant induction of CSRGA23 within 72 h of inoculation. The results indicate that CSRGA23 may play a critical role in protecting cucumber against P. cubensis through a signaling the pathway triggered by these molecules. Conclusions Four classes of NBS-type RGAs were

  11. Overcoming qEEG abnormalities and reward gene deficits during protracted abstinence in male psychostimulant and polydrug abusers utilizing putative dopamine D₂ agonist therapy: part 2.

    Science.gov (United States)

    Blum, Kenneth; Chen, Thomas J H; Morse, Siobhan; Giordano, John; Chen, Amanda Lih Chaun; Thompson, James; Allen, Cameron; Smolen, Andrew; Lubar, Joel; Stice, Eric; Downs, B William; Waite, Roger L; Madigan, Margaret A; Kerner, Mallory; Fornari, Frank; Braverman, Eric R

    2010-11-01

    It is well established that in both food- and drug-addicted individuals there is "dopamine resistance" associated with the DRD2 gene A1 allele. Based on earlier studies, evidence is emerging wherein the potential of utilizing a natural, nonaddicting, safe, putative D2 agonist may play a significant role in the recovery of individuals with reward deficiency syndrome, including those addicted to psychoactive chemicals. Positive outcomes demonstrated by quantitative electroencephalographic (qEEG) imaging in a randomized, triple-blind, placebo-controlled, crossover study involving oral Synaptose Complex KB220Z™ showed an increase of alpha waves and low beta wave activity in the parietal brain region. Using t statistics, significant differences observed between placebo and Synaptose Complex KB220Z™ consistently occurred in the frontal regions after week 1 and then again after week 2 of analyses (P = 0.03). This is the first report to demonstrate involvement of the prefrontal cortex in the qEEG response to a natural putative D2 agonist (Synaptose Complex KB220Z™), especially evident in dopamine D2 A1 allele subjects. Independently, we have further supported this finding with an additional study of 3 serious polydrug abusers undergoing protracted abstinence who carried the DRD2 A1 allele. Significant qEEG differences were found between those who received 1 dose of placebo compared with those who were administered Synaptose Complex KB220Z™. Synaptose Complex KB220Z™ induced positive regulation of the dysregulated electrical activity of the brain in these addicts. The results are indicative of a phase change from low amplitude or low power in the brain to a more regulated state by increasing an average of 6.169 mV(2) across the prefrontal cortical region. In the first experiment we found that while 50% of the subjects carried the DRD2 A1 allele, 100% carried ≥ 1 risk allele. Specifically, based on the proposed addiction risk score for these 14 subjects, 72% had

  12. Proteomic identification of putative biomarkers for early detection of sudden cardiac death in a family with a LMNA gene mutation causing dilated cardiomyopathy.

    Science.gov (United States)

    Izquierdo, Irene; Rosa, Isaac; Bravo, Susana Belén; Guitián, Esteban; Pérez-Serra, Alexandra; Campuzano, Oscar; Brugada, Ramon; Mangas, Alipio; García, Ángel; Toro, Rocio

    2016-10-04

    Dilated cardiomyopathy (DCM) is a severe heart disease characterized by progressive ventricular dilation and impaired systolic function of the left ventricle. We recently identified a novel pathogenic mutation in the LMNA gene in a family affected by DCM showing sudden death background. We now aimed to identify potential biomarkers of disease status, as well as sudden death predictors, in members of this family. We analysed plasma samples from 14 family members carrying the mutation, four of which (with relevant clinical symptoms) were chosen for the proteomic analysis. Plasma samples from these four patients and from four sex- and age-matched healthy controls were processed for their enrichment in low- and medium-abundance proteins (ProteoMiner™) prior to proteomic analysis by 2D-DIGE and MS. 111 spots were found to be differentially regulated between mutation carriers and control groups, 83 of which were successfully identified by MS, corresponding to 41 different ORFs. Some proteins of interest were validated either by turbidimetry or western blot in family members and healthy controls. Actin, alpha-1-antytripsin, clusterin, vitamin-D binding protein and antithrombin-III showed increased levels in plasma from the diseased group. We suggest following these proteins as putative biomarkers for the evaluation of DCM status in LMNA mutation carriers. We developed a proteomic analysis of plasma samples from a family showing history of dilated cardiomyopathy caused by a LMNA mutation, which may lead to premature death or cardiac transplant. We identified a number of proteins augmented in mutation carriers that could be followed as potential biomarkers for dilated cardiomyopathy on these patients. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Differential regulation of hepatopancreatic vitellogenin (VTG) gene expression by two putative molt-inhibiting hormones (MIH1/2) in Pacific white shrimp (Litopenaeus vannamei).

    Science.gov (United States)

    Luo, Xing; Chen, Ting; Zhong, Ming; Jiang, Xiao; Zhang, Lvping; Ren, Chunhua; Hu, Chaoqun

    2015-06-01

    Molt-inhibiting hormone (MIH), a peptide member of the crustacean hyperglycemic hormone (CHH) family, is commonly considered as a negative regulator during the molt cycle in crustaceans. Phylogenetic analysis of CHH family peptides in penaeidae shrimps suggested that there is no significant differentiation between MIH and vitellogenesis-inhibiting hormone (VIH, another peptide member of CHH family), by far the most potent negative regulator of crustacean vitellogenesis known. Thus, MIH may also play a role in regulating vitellogenesis. In this study, two previously reported putative MIHs (LivMIH1 and LivMIH2) in the Pacific white shrimp (Litopenaeus vannamei) were expressed in Escherichia coli, purified by immobilized metal ion affinity chromatography (IMAC) and further confirmed by western blot. Regulation of vitellogenin (VTG) mRNA expression by recombinant LivMIH1 and LivMIH2 challenge was performed by both in vitro hepatopancreatic primary cells culture and in vivo injection approaches. In in vitro primary culture of shrimp hepatopancreatic cells, only LivMIH2 but not LivMIH1 administration could improve the mRNA expression of VTG. In in vivo injection experiments, similarly, only LivMIH2 but not LivMIH1 could stimulate hepatopancreatic VTG gene expression and induce ovary maturation. Our study may provide evidence for one isoform of MIH (MIH2 in L. vannamei) may serve as one of the mediators of the physiological progress of molting and vitellogenesis. Our study may also give new insight in CHH family peptides regulating reproduction in crustaceans, in particular penaeidae shrimps.

  14. A putative gene sbe3-rs for resistant starch mutated from SBE3 for starch branching enzyme in rice (Oryza sativa L..

    Directory of Open Access Journals (Sweden)

    Ruifang Yang

    Full Text Available Foods high in resistant starch (RS are beneficial to prevent various diseases including diabetes, colon cancers, diarrhea and chronic renal or hepatic diseases. Elevated RS in rice is important for public health since rice is a staple food for half of the world population. A japonica mutant 'Jiangtangdao 1' (RS = 11.67% was crossed with an indica cultivar 'Miyang 23' (RS = 0.41%. The mutant sbe3-rs that explained 60.4% of RS variation was mapped between RM6611 and RM13366 on chromosome 2 (LOD = 36 using 178 F(2 plants genotyped with 106 genome-wide polymorphic SSR markers. Using 656 plants from four F(3:4 families, sbe3-rs was fine mapped to a 573.3 Kb region between InDel 2 and InDel 6 using one STS, five SSRs and seven InDel markers. SBE3 which codes for starch branching enzyme was identified as a candidate gene within the putative region. Nine pairs of primers covering 22 exons were designed to sequence genomic DNA of the wild type for SBE3 and the mutant for sbe3-rs comparatively. Sequence analysis identified a missense mutation site where Leu-599 of the wild was changed to Pro-599 of the mutant in the SBE3 coding region. Because the point mutation resulted in the loss of a restriction enzyme site, sbe3-rs was not digested by a CAPS marker for SpeI site while SBE3 was. Co-segregation of the digestion pattern with RS content among 178 F(2 plants further supported sbe3-rs responsible for RS in rice. As a result, the CAPS marker could be used in marker-assisted breeding to develop rice cultivars with elevated RS which is otherwise difficult to accurately assess in crops. Transgenic technology should be employed for a definitive conclusion of the sbe3-rs.

  15. Functional validation of putative toxin-antitoxin genes from the Gram-positive pathogen Streptococcus pneumoniae: phd-doc is the fourth bona-fide operon

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    Wai Ting eChan

    2014-12-01

    Full Text Available Bacterial toxin-antitoxin (TAs loci usually consist of two genes organized as an operon, where their products are bound together and inert under normal conditions. However, under stressful circumstances the antitoxin, which is more labile, will be degraded more rapidly, thereby unleashing its cognate toxin to act on the cell. This, in turn, causes cell stasis or cell death, depending on the type of TAs and/or time of toxin exposure. Previously based on in silico analyses, we proposed that Streptococcus pneumoniae, a pathogenic Gram-positive bacterium, may harbor between four to ten putative TA loci depending on the strains. Here we have chosen the pneumococcal strain Hungary19A-6 which contains all possible ten TA loci. In addition to the three well-characterized operons, namely relBE2, yefM-yoeB, and pezAT, we show here the functionality of a fourth operon that encodes the pneumococcal equivalent of the phd-doc TA. Transcriptional fusions with gene encoding Green Fluorescent Protein showed that the promoter was slightly repressed by the Phd antitoxin, and exhibited almost background values when both Phd-Doc were expressed together. These findings demonstrate that phd-doc shows the negative self-regulatory features typical for an authentic TA. Further, we also show that the previously proposed TAs XreA-Ant and Bro-XreB, although they exhibit a genetic organization resembling those of typical TAs, did not appear to confer a functional behavior corresponding to bona fide TAs. In addition, we have also discovered new interesting bioinformatics results for the known pneumococcal TAs RelBE2 and PezAT. A global analysis of the four identified toxins-antitoxins in the pneumococcal genomes (PezAT, RelBE2, YefM-YoeB, and Phd-Doc showed that RelBE2 and Phd-Doc are the most conserved ones. Further, there was good correlation among TA types, clonal complexes and sequence types in the 48 pneumococcal strains analyzed.

  16. 3'-coterminal subgenomic RNAs and putative cis-acting elements of Grapevine leafroll-associated virus 3 reveals 'unique' features of gene expression strategy in the genus Ampelovirus

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    Dawson William O

    2010-08-01

    Full Text Available Abstract Background The family Closteroviridae comprises genera with monopartite genomes, Closterovirus and Ampelovirus, and with bipartite and tripartite genomes, Crinivirus. By contrast to closteroviruses in the genera Closterovirus and Crinivirus, much less is known about the molecular biology of viruses in the genus Ampelovirus, although they cause serious diseases in agriculturally important perennial crops like grapevines, pineapple, cherries and plums. Results The gene expression and cis-acting elements of Grapevine leafroll-associated virus 3 (GLRaV-3; genus Ampelovirus was examined and compared to that of other members of the family Closteroviridae. Six putative 3'-coterminal subgenomic (sg RNAs were abundantly present in grapevine (Vitis vinifera infected with GLRaV-3. The sgRNAs for coat protein (CP, p21, p20A and p20B were confirmed using gene-specific riboprobes in Northern blot analysis. The 5'-termini of sgRNAs specific to CP, p21, p20A and p20B were mapped in the 18,498 nucleotide (nt virus genome and their leader sequences determined to be 48, 23, 95 and 125 nt, respectively. No conserved motifs were found around the transcription start site or in the leader sequence of these sgRNAs. The predicted secondary structure analysis of sequences around the start site failed to reveal any conserved motifs among the four sgRNAs. The GLRaV-3 isolate from Washington had a 737 nt long 5' nontranslated region (NTR with a tandem repeat of 65 nt sequence and differed in sequence and predicted secondary structure with a South Africa isolate. Comparison of the dissimilar sequences of the 5'NTRs did not reveal any common predicted structures. The 3'NTR was shorter and more conserved. The lack of similarity among the cis-acting elements of the diverse viruses in the family Closteroviridae is another measure of the complexity of their evolution. Conclusions The results indicate that transcription regulation of GLRaV-3 sgRNAs appears to be different

  17. Identification and genetic mapping of the putative Thinopyrum intermedium-derived dominant powdery mildew resistance gene PmL962 on wheat chromosome arm 2BS

    Science.gov (United States)

    Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is a destructive disease affecting the production of wheat (Triticum aestivum). Powdery mildew resistance was putatively transferred from Thinopyrum intermedium to the common wheat line L962, which conferred resistance to multiple Ch...

  18. Transgenic cucumbers harboring the 54-kDa putative gene of Cucumber fruit mottle mosaic tobamovirus are highly resistant to viral infection and protect non-transgenic scions from soil infection.

    Science.gov (United States)

    Gal-On, Amit; Wolf, Dalia; Antignus, Yehezkel; Patlis, Larisa; Ryu, Ki Hyun; Min, Byoung Eun; Pearlsman, Malenia; Lachman, Oded; Gaba, Victor; Wang, Yongzeng; Shiboleth, Yoel Moshe; Yang, Jee; Zelcer, Aaron

    2005-02-01

    Cucumber fruit mottle mosaic tobamovirus (CFMMV) causes severe mosaic symptoms and yellow mottling on leaves and fruits and, occasionally, severe wilting of cucumber (Cucumis sativus L.) plants. No genetic source of resistance against this virus has been identified in cucumber. The gene coding for the putative 54-kDa replicase gene of CFMMV was cloned into an Agrobacterium tumefaciens binary vector, and transformation was performed on cotyledon explants of a parthenocarpic cucumber cultivar. R1 seedlings were screened for resistance to CFMMV by symptom expression, back inoculation on an alternative host and ELISA. From a total of 14 replicase-containing R1 lines, eight resistant lines were identified. Line 144--homozygous for the putative 54-kDa replicase gene--was immune to CFMMV infection by mechanical and graft inoculation, and to root infection following planting in CFMMV-infested soil. A substantial delay of symptom appearance was observed following infection by three additional cucurbit-infecting tobamoviruses. When used as a rootstock, line I44 protected susceptible cucumber scions from soil infection by CFMMV. This paper is the first report on protection of a susceptible cultivar against a soil-borne viral pathogen, by grafting onto a transgenic rootstock.

  19. Virus-Induced Gene Silencing-Based Functional Analyses Revealed the Involvement of Several Putative Trehalose-6-Phosphate Synthase/Phosphatase Genes in Disease Resistance against Botrytis cinerea and Pseudomonas syringae pv. tomato DC3000 in Tomato

    Science.gov (United States)

    Zhang, Huijuan; Hong, Yongbo; Huang, Lei; Liu, Shixia; Tian, Limei; Dai, Yi; Cao, Zhongye; Huang, Lihong; Li, Dayong; Song, Fengming

    2016-01-01

    Trehalose and its metabolism have been demonstrated to play important roles in control of plant growth, development, and stress responses. However, direct genetic evidence supporting the functions of trehalose and its metabolism in defense response against pathogens is lacking. In the present study, genome-wide characterization of putative trehalose-related genes identified 11 SlTPSs for trehalose-6-phosphate synthase, 8 SlTPPs for trehalose-6-phosphate phosphatase and one SlTRE1 for trehalase in tomato genome. Nine SlTPSs, 4 SlTPPs, and SlTRE1 were selected for functional analyses to explore their involvement in tomato disease resistance. Some selected SlTPSs, SlTPPs, and SlTRE1 responded with distinct expression induction patterns to Botrytis cinerea and Pseudomonas syringae pv. tomato (Pst) DC3000 as well as to defense signaling hormones (e.g., salicylic acid, jasmonic acid, and a precursor of ethylene). Virus-induced gene silencing-mediated silencing of SlTPS3, SlTPS4, or SlTPS7 led to deregulation of ROS accumulation and attenuated the expression of defense-related genes upon pathogen infection and thus deteriorated the resistance against B. cinerea or Pst DC3000. By contrast, silencing of SlTPS5 or SlTPP2 led to an increased expression of the defense-related genes upon pathogen infection and conferred an increased resistance against Pst DC3000. Silencing of SlTPS3, SlTPS4, SlTPS5, SlTPS7, or SlTPP2 affected trehalose level in tomato plants with or without infection of B. cinerea or Pst DC3000. These results demonstrate that SlTPS3, SlTPS4, SlTPS5, SlTPS7, and SlTPP2 play roles in resistance against B. cinerea and Pst DC3000, implying the importance of trehalose and tis metabolism in regulation of defense response against pathogens in tomato. PMID:27540389

  20. Molecular Characterization and Tissue-specific Expression of a Novel FKBP38 Gene in the Cashmere Goat (

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    X. Zheng

    2012-06-01

    Full Text Available As a member of a subclass of immunophilins, it is controversial that FKBP38 acts an upstream regulator of mTOR signaling pathway, which control the process of cell-growth, proliferation and differentiation. In order to explore the relationship between FKBP38 and mTOR in the Cashmere goat (Capra hircus cells, a full-length cDNA was cloned (GenBank accession number JF714970 and expression pattern was analyzed. The cloned FKBP38 gene is 1,248 bp in length, containing an open reading frame (ORF from nucleotide 13 to 1,248 which encodes 411 amino acids, and 12 nucleotides in front of the initiation codon. The full cDNA sequence shares 98% identity with cattle, 94% with horse and 90% with human. The putative amino acid sequence shows the higher homology which is 98%, 97% and 94%, correspondingly. The bioinformatics analysis showed that FKBP38 contained a FKBP_C domain, two TPR domains and a TM domain. Psite analysis suggested that the ORF encoding protein contained a leucine-zipper pattern and a Prenyl group binding site (CAAX box. Tissue-specific expression analysis was performed by semi-quantitative RT-PCR and showed that the FKBP38 expression was detected in all the tested tissues and the highest level of mRNA accumulation was detected in testis, suggesting that FKBP38 plays an important role in goat cells.

  1. Comparative genomic analysis of SET domain family reveals the origin, expansion, and putative function of the arthropod-specific SmydA genes as histone modifiers in insects

    Science.gov (United States)

    Jiang, Feng; Liu, Qing; Wang, Yanli; Zhang, Jie; Wang, Huimin; Song, Tianqi; Yang, Meiling

    2017-01-01

    Abstract The SET domain is an evolutionarily conserved motif present in histone lysine methyltransferases, which are important in the regulation of chromatin and gene expression in animals. In this study, we searched for SET domain–containing genes (SET genes) in all of the 147 arthropod genomes sequenced at the time of carrying out this experiment to understand the evolutionary history by which SET domains have evolved in insects. Phylogenetic and ancestral state reconstruction analysis revealed an arthropod-specific SET gene family, named SmydA, that is ancestral to arthropod animals and specifically diversified during insect evolution. Considering that pseudogenization is the most probable fate of the new emerging gene copies, we provided experimental and evolutionary evidence to demonstrate their essential functions. Fluorescence in situ hybridization analysis and in vitro methyltransferase activity assays showed that the SmydA-2 gene was transcriptionally active and retained the original histone methylation activity. Expression knockdown by RNA interference significantly increased mortality, implying that the SmydA genes may be essential for insect survival. We further showed predominantly strong purifying selection on the SmydA gene family and a potential association between the regulation of gene expression and insect phenotypic plasticity by transcriptome analysis. Overall, these data suggest that the SmydA gene family retains essential functions that may possibly define novel regulatory pathways in insects. This work provides insights into the roles of lineage-specific domain duplication in insect evolution. PMID:28444351

  2. The study of the E-class SEPALLATA3-like MADS-box genes in wild-type and mutant flowers of cultivated saffron crocus (Crocus sativus L.) and its putative progenitors.

    Science.gov (United States)

    Tsaftaris, Athanasios; Pasentsis, Konstantinos; Makris, Antonios; Darzentas, Nikos; Polidoros, Alexios; Kalivas, Apostolos; Argiriou, Anagnostis

    2011-09-15

    To further understand flowering and flower organ formation in the monocot crop saffron crocus (Crocus sativus L.), we cloned four MIKC(c) type II MADS-box cDNA sequences of the E-class SEPALLATA3 (SEP3) subfamily designated CsatSEP3a/b/c/c_as as well as the three respective genomic sequences. Sequence analysis showed that cDNA sequences of CsatSEP3 c and c_as are the products of alternative splicing of the CsatSEP3c gene. Bioinformatics analysis with putative orthologous sequences from various plant species suggested that all four cDNA sequences encode for SEP3-like proteins with characteristic motifs and amino acids, and highlighted intriguing sequence features. Phylogenetically, the isolated sequences were closest to the SEP3-like genes from monocots such as Asparagus virgatus, Oryza sativa, Zea mays, and the dicot Arabidopsis SEP3 gene. All four isolated C. sativus sequences were strongly expressed in flowers and in all flower organs: whorl1 tepals, whorl2 tepals, stamens and carpels, but not in leaves. Expression of CsatSEP3a/b/c/c_as cDNAs was compared in wild-type and mutant flowers. Expression of the isolatedCsatSEP3-like genes in whorl1 tepals together with E-class CsatAP1/FUL subfamily and B-class CsatAP3 and CsatPI subfamilies of genes, fits the ABCE "quartet model," an extended form of the original ABC model proposed to explain the homeotic transformation of whorl1 sepals into whorl1 tepals in Liliales and Asparagales plants such as C. sativus. This conclusion was also supported by the interaction of the CsatSEP3b protein with CsatAP1/FUL and CsatAP3 proteins. In contrast, expression of both B-class CsatAP3 and CsatPI genes and the C-class CsatAGAMOUS genes together with E-class CsatSEP3-like genes in carpels, without any phenotypic effects on carpels, raises questions about the role of these gene classes in carpel formation in this non-grass monocot and requires further experimentation. Finally, taking advantage of the size and sequence differences in

  3. [Changes in Cell Surface Properties and Biofilm Formation Efficiency in Azospirillum brasilense Sp245 Mutants in the Putative Genes of Lipid Metabolism mmsB1 and fabG1].

    Science.gov (United States)

    Shumilova, E; Shelud'ko, A V; Filip'echeva, Yu A; Evstigneeva, S S; Ponomareva, E G; Petrova, L P; Katsy, E I

    2016-01-01

    The previously obtained insertion mutants ofAzospirillum brasilense Sp245 in the genes mmsBl and fabG1 (strains SK039 and Sp245.1610, respectively) were characterized by impaired flagellation and motility. The putative products of expression of these genes are 3-hydroxyisobutyrate dehydrogenase and 3-oxoacyl-[acyl-carrier protein] reductase, respectively. In the present work, A. brasilense- Sp245 strains SK039 and Sp245.1610 were found to have differences in the content of 3-hydroxyhexadecanoic, hexadecanoic, 3-hydroxytetradecanoic, hexadecenoic, octadecenoic, and nonadecanoic acids in their lipopolysaccharide prepa- rations, as well as in cell hydrophobicity and hemagglutination activity and dynamics of cell aggregation, in biomass amount, and in the relative content of lipopolysaccharide antigens in mature biofilms formed on hydrophilic or hydrophobic surfaces.

  4. Identification of the gene encoding Brain Cell Membrane Protein 1 (BCMP1, a putative four-transmembrane protein distantly related to the Peripheral Myelin Protein 22 / Epithelial Membrane Proteins and the Claudins

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    Christophe Daniel

    2001-07-01

    Full Text Available Abstract Background A partial cDNA clone from dog thyroid presenting a very significant similarity with an uncharacterized mouse EST sequence was isolated fortuitously. We report here the identification of the complete mRNA and of the gene, the product of which was termed "brain cell membrane protein 1" (BCMP1. Results The 4 kb-long mRNA sequence exhibited an open-reading frame of only 543 b followed by a 3.2 kb-long 3' untranslated region containing several AUUUA instability motifs. Analysis of the encoded protein sequence identified the presence of four putative transmembrane domains. Similarity searches in protein domain databases identified partial sequence conservations with peripheral myelin protein 22 (PMP22/ epithelial membrane proteins (EMPs and Claudins, defining the encoded protein as representative of the existence of a novel subclass in this protein family. Northern-blot analysis of the expression of the corresponding mRNA in adult dog tissues revealed the presence of a huge amount of the 4 kb transcript in the brain. An EGFP-BCMP1 fusion protein expressed in transfected COS-7 cells exhibited a membranous localization as expected. The sequences encoding BCMP1 were assigned to chromosome X in dog, man and rat using radiation hybrid panels and were partly localized in the currently available human genome sequence. Conclusions We have identified the existence in several mammalian species of a gene encoding a putative four-transmembrane protein, BCMP1, wich defines a novel subclass in this family of proteins. In dog at least, the corresponding mRNA is highly present in brain cells. The chromosomal localization of the gene in man makes of it a likely candidate gene for X-linked mental retardation.

  5. Integrated network analysis identifies fight-club nodes as a class of hubs encompassing key putative switch genes that induce major transcriptome reprogramming during grapevine development.

    Science.gov (United States)

    Palumbo, Maria Concetta; Zenoni, Sara; Fasoli, Marianna; Massonnet, Mélanie; Farina, Lorenzo; Castiglione, Filippo; Pezzotti, Mario; Paci, Paola

    2014-12-01

    We developed an approach that integrates different network-based methods to analyze the correlation network arising from large-scale gene expression data. By studying grapevine (Vitis vinifera) and tomato (Solanum lycopersicum) gene expression atlases and a grapevine berry transcriptomic data set during the transition from immature to mature growth, we identified a category named "fight-club hubs" characterized by a marked negative correlation with the expression profiles of neighboring genes in the network. A special subset named "switch genes" was identified, with the additional property of many significant negative correlations outside their own group in the network. Switch genes are involved in multiple processes and include transcription factors that may be considered master regulators of the previously reported transcriptome remodeling that marks the developmental shift from immature to mature growth. All switch genes, expressed at low levels in vegetative/green tissues, showed a significant increase in mature/woody organs, suggesting a potential regulatory role during the developmental transition. Finally, our analysis of tomato gene expression data sets showed that wild-type switch genes are downregulated in ripening-deficient mutants. The identification of known master regulators of tomato fruit maturation suggests our method is suitable for the detection of key regulators of organ development in different fleshy fruit crops.

  6. Trichinella spiralis mtDNA: a nematode mitochondrial genome that encodes a putative ATP8 and normally structured tRNAS and has a gene arrangement relatable to those of coelomate metazoans.

    Science.gov (United States)

    Lavrov, D V; Brown, W M

    2001-01-01

    The complete mitochondrial DNA (mtDNA) of the nematode Trichinella spiralis has been amplified in four overlapping fragments and 16,656 bp of its sequence has been determined. This sequence contains the 37 genes typical of metazoan mtDNAs, including a putative atp8, which is absent from all other nematode mtDNAs examined. The genes are transcribed from both mtDNA strands and have an arrangement relatable to those of coelomate metazoans, but not to those of secernentean nematodes. All protein genes appear to initiate with ATN codons, typical for metazoans. Neither TTG nor GTT start codons, inferred for several genes of other nematodes, were found. The 22 T. spiralis tRNA genes fall into three categories: (i) those with the potential to form conventional "cloverleaf" secondary structures, (ii) those with TPsiC arm + variable arm replacement loops, and (iii) those with DHU-arm replacement loops. Mt-tRNA(R) has a 5'-UCG-3' anticodon, as in most other metazoans, instead of the very unusual 5'-ACG-3' present in the secernentean nematodes. The sequence also contains a large repeat region that is polymorphic in size at the population and/or individual level. PMID:11156984

  7. Abundance and distribution of archaeal acetyl-CoA/propionyl-CoA carboxylase genes indicative for putatively chemoautotrophic Archaea in the tropical Atlantic's interior.

    Science.gov (United States)

    Bergauer, Kristin; Sintes, Eva; van Bleijswijk, Judith; Witte, Harry; Herndl, Gerhard J

    2013-06-01

    Recently, evidence suggests that dark CO2 fixation in the pelagic realm of the ocean does not only occur in the suboxic and anoxic water bodies but also in the oxygenated meso- and bathypelagic waters of the North Atlantic. To elucidate the significance and phylogeny of the key organisms mediating dark CO2 fixation in the tropical Atlantic, we quantified functional genes indicative for CO2 fixation. We used a Q-PCR-based assay targeting the bifunctional acetyl-CoA/propionyl-CoA carboxylase (accA subunit), a key enzyme powering inter alia the 3-hydroxypropionate/4-hydroxybutyrate cycle (HP/HB) and the archaeal ammonia monooxygenase (amoA). Quantification of accA-like genes revealed a consistent depth profile in the upper mesopelagial with increasing gene abundances from subsurface layers towards the oxygen minimum zone (OMZ), coinciding with an increase in archaeal amoA gene abundance. Gene abundance profiles of metabolic marker genes (accA, amoA) were correlated with thaumarchaeal 16S rRNA gene abundances as well as CO2 fixation rates to link the genetic potential to actual rate measurements. AccA gene abundances correlated with archaeal amoA gene abundance throughout the water column (r(2)  = 0.309, P < 0.0001). Overall, a substantial genetic predisposition of CO2 fixation was present in the dark realm of the tropical Atlantic in both Archaea and Bacteria. Hence, dark ocean CO2 fixation might be more widespread among prokaryotes inhabiting the oxygenated water column of the ocean's interior than hitherto assumed. © 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  8. Cloning of a cDNA encoding a putative human very low density lipoprotein/Apolipoprotein E receptor and assignment of the gene to chromosome 9pter-p23[sup 6

    Energy Technology Data Exchange (ETDEWEB)

    Gafvels, M.E.; Strauss, J.F. III (Univ. of Pennyslvania, Philadelphia, PA (United States)); Caird, M.; Patterson, D. (Eleanor Roosevelt Institute, Denver, CO (United States)); Britt, D.; Jackson, C.L. (Brown Univ., Providence, RI (United States))

    1993-11-01

    The authors report the cloning of a 3656-bp cDNA encoding a putative human very low density lipoprotein (VLDL)/apolipoprotein E (ApoE) receptor. The gene encoding this protein was mapped to chromosome 9pter-p23. Northern analysis of human RNA identified cognate mRNAs of 6.0 and 3.8 kb with most abundant expression in heart and skeletal muscle, followed by kidney, placenta, pancreas, and brain. The pattern of expression generally paralleled that of lipoprotein lipase mRNA but differed from that of the low density lipoprotein (LDL) receptor and the low density lipoprotein receptor-related protein/[alpha][sub 2]-macroglobulin receptor (LRP), which are members of the same gene family. VLDL/ApoE receptor message was not detected in liver, whereas mRNAs for both LDL receptor and LRP were found in hepatic tissue. In mouse 3T3-L1 cells, VLDL/ApoE receptor mRNA was induced during the transformation of the cells into adipocytes. Expression was also detected in human choriocarcinoma cells, suggesting that at least part of the expression observed in placenta may be in trophoblasts, cells which would be exposed to maternal blood. Expression in brain may be related to high levels of ApoE expression in that organ, an observation of potential relevance to the recently hypothesized role for ApoE in late onset Alzheimer disease. The results suggest that the putative VLDL/ApoE receptor could play a role in the uptake of triglyceride-rich lipoprotein particles by specific organs including striated and cardiac muscle and adipose tissue and in the transport of maternal lipids across the placenta. The findings presented here, together with recent observations from other laboratories, bring up the possibility that a single gene, the VLDL/ApoE receptor, may play a role in the pathogenesis of certain forms of atherosclerosis, Alzheimer disease, and obesity.

  9. VMA11, a novel gene that encodes a putative proteolipid, is indispensable for expression of yeast vacuolar membrane H(+)-ATPase activity.

    Science.gov (United States)

    Umemoto, N; Ohya, Y; Anraku, Y

    1991-12-25

    A gene, VMA11, is indispensable for expression of the vacuolar membrane H(+)-ATPase activity in the yeast Saccharomyces cerevisiae (Ohya, Y., Umemoto, N., Tanida, I., Ohta, A., Iida, H., and Anraku, Y. (1991) J. Biol. Chem. 266, 13971-13977). The VMA11 gene was isolated from a yeast genomic DNA library by complementation of the vma11 mutation. The nucleotide sequence of the gene predicts a hydrophobic proteolipid of 164 amino acids with a calculated molecular mass of 17,037 daltons. The deduced amino acid sequence shows 56.7% identity, and significant coincidence in amino acid composition with the 16-kDa subunit c (a VMA3 gene product) of the yeast vacuolar membrane H(+)-ATPase. VMA11 and VMA3 on a multicopy plasmid did not suppress the vma3 and vma11 mutation, respectively, suggesting functional independence of the two gene products. Biochemical detection of the VMA11 gene product was unsuccessful, but vacuoles in the VMA11-disrupted cells were not assembled with either subunit c or subunits a and b of the H(+)-ATPase, resulting in defects of the activity and in vivo vacuolar acidification.

  10. Characterization of a gene coding for a putative adenosine deaminase-related growth factor by RNA interference in the basidiomycete Flammulina velutipes.

    Science.gov (United States)

    Sekiya, Shuichi; Yamada, Masato; Shibata, Kou; Okuhara, Toru; Yoshida, Masumi; Inatomi, Satoshi; Taguchi, Goro; Shimosaka, Makoto

    2013-04-01

    A full-length cDNA coding for a putative adenosine deaminase (Fv-ada) was isolated from the basidiomycete Flammulina velutipes. Fv-ada encodes a polypeptide consisting of 537 amino acid residues, which has a consensus sequence conserved among adenosine deaminase-related growth factors (ADGF) found in several metazoa, including chordates and insects. Fv-ada transcript was detected at all stages of growth in dikaryotic F. velutipes cells, with a peak at the primordial stage. Heterologous expression of Fv-ada in the yeast Pichia pastoris produced recombinant Fv-ADA that catalyzed the conversion of adenosine to inosine. Dikaryotic mycelia from F. velutipes were transformed with the binary plasmid pFungiway-Fv-ada, which was designed to suppress the expression of Fv-ada through RNA interference. The growth rates of the resulting transformants were retarded in response to the degree of suppression, indicating that Fv-ada plays an important role in the mycelial growth of F. velutipes. These results suggested that ADGF could function as growth factors in fungi, as is seen in other eukaryotes.

  11. Cloning of cellobiose phosphoenolpyruvate-dependent phosphotransferase genes: Functional expression in recombinant Escherichia coli and identification of a putative binding region for disaccharides

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Xiaokuang; Davis, F.C.; Ingram, L.O. [Univ. of Florida, Gainesville, FL (United States); Hespell, R.B. [USDA Agricultural Research Service, Peoria, IL (United States)

    1997-02-01

    Genomic libraries from nine cellobiose-metabolizing bacteria were screened for cellobiose utilization. Positive clones were recovered from six libraries, all of which encode phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) proteins. Clones from Bacillus subtilis, Butyrivibrio fibrisolvens, and Klebsiella oxytoca allowed the growth of recombinant Escherichia coli in cellobiose-M9 minimal medium. The K. oxytoca clone, pLOI1906, exhibited an unusually broad substrate range (cellobiose, arbutin, salicin, and methylumbelliferyl derivatives of glucose, cellobiose, mannose, and xylose) and was sequenced. The insert in this plasmid encoded the carboxy-terminal region of a putative regulatory protein, cellobiose permease (single polypeptide), and phospho-{beta}-glucosidase, which appear to form an operon (casRAB). Subclones allowed both casA and casB to be expressed independently, as evidenced by in vitro complementation. An analysis of the translated sequences from the EIIC domains of cellobiose, aryl-{beta}-glucoside, and other disaccharide permeases allowed the identification of a 50-amino-acid conserved region. A disaccharide consensus sequence is proposed for the most conserved segment (13 amino acids), which may represent part of the EIIC active site for binding and phosphorylation. 63 refs., 4 figs., 4 tabs.

  12. A universal algorithm for genome-wide in silicio identification of biologically significant gene promoter putative cis-regulatory-elements; identification of new elements for reactive oxygen species and sucrose signaling in Arabidopsis.

    Science.gov (United States)

    Geisler, Matt; Kleczkowski, Leszek A; Karpinski, Stanislaw

    2006-02-01

    Short motifs of many cis-regulatory elements (CREs) can be found in the promoters of most Arabidopsis genes, and this raises the question of how their presence can confer specific regulation. We developed a universal algorithm to test the biological significance of CREs by first identifying every Arabidopsis gene with a CRE and then statistically correlating the presence or absence of the element with the gene expression profile on multiple DNA microarrays. This algorithm was successfully verified for previously characterized abscisic acid, ethylene, sucrose and drought responsive CREs in Arabidopsis, showing that the presence of these elements indeed correlates with treatment-specific gene induction. Later, we used standard motif sampling methods to identify 128 putative motifs induced by excess light, reactive oxygen species and sucrose. Our algorithm was able to filter 20 out of 128 novel CREs which significantly correlated with gene induction by either heat, reactive oxygen species and/or sucrose. The position, orientation and sequence specificity of CREs was tested in silicio by analyzing the expression of genes with naturally occurring sequence variations. In three novel CREs the forward orientation correlated with sucrose induction and the reverse orientation with sucrose suppression. The functionality of the predicted novel CREs was experimentally confirmed using Arabidopsis cell-suspension cultures transformed with short promoter fragments or artificial promoters fused with the GUS reporter gene. Our genome-wide analysis opens up new possibilities for in silicio verification of the biological significance of newly discovered CREs, and allows for subsequent selection of such CREs for experimental studies.

  13. Transcriptome profiling of Pinus radiata juvenile wood with contrasting stiffness identifies putative candidate genes involved in microfibril orientation and cell wall mechanics

    Directory of Open Access Journals (Sweden)

    Wu Harry X

    2011-10-01

    Full Text Available Abstract Background The mechanical properties of wood are largely determined by the orientation of cellulose microfibrils in secondary cell walls. Several genes and their allelic variants have previously been found to affect microfibril angle (MFA and wood stiffness; however, the molecular mechanisms controlling microfibril orientation and mechanical strength are largely uncharacterised. In the present study, cDNA microarrays were used to compare gene expression in developing xylem with contrasting stiffness and MFA in juvenile Pinus radiata trees in order to gain further insights into the molecular mechanisms underlying microfibril orientation and cell wall mechanics. Results Juvenile radiata pine trees with higher stiffness (HS had lower MFA in the earlywood and latewood of each ring compared to low stiffness (LS trees. Approximately 3.4 to 14.5% out of 3, 320 xylem unigenes on cDNA microarrays were differentially regulated in juvenile wood with contrasting stiffness and MFA. Greater variation in MFA and stiffness was observed in earlywood compared to latewood, suggesting earlywood contributes most to differences in stiffness; however, 3-4 times more genes were differentially regulated in latewood than in earlywood. A total of 108 xylem unigenes were differentially regulated in juvenile wood with HS and LS in at least two seasons, including 43 unigenes with unknown functions. Many genes involved in cytoskeleton development and secondary wall formation (cellulose and lignin biosynthesis were preferentially transcribed in wood with HS and low MFA. In contrast, several genes involved in cell division and primary wall synthesis were more abundantly transcribed in LS wood with high MFA. Conclusions Microarray expression profiles in Pinus radiata juvenile wood with contrasting stiffness has shed more light on the transcriptional control of microfibril orientation and the mechanical properties of wood. The identified candidate genes provide an

  14. A non-sense mutation in the putative anti-mutator gene ada/alkA of Mycobacterium tuberculosis and M. bovis isolates suggests convergent evolution

    Directory of Open Access Journals (Sweden)

    Gicquel Brigitte

    2007-05-01

    Full Text Available Abstract Background Previous studies have suggested that variations in DNA repair genes of W-Beijing strains may have led to transient mutator phenotypes which in turn may have contributed to host adaptation of this strain family. Single nucleotide polymorphism (SNP in the DNA repair gene mutT1 was identified in MDR-prone strains from the Central African Republic. A Mycobacteriumtuberculosis H37Rv mutant inactivated in two DNA repair genes, namely ada/alkA and ogt, was shown to display a hypermutator phenotype. We then looked for polymorphisms in these genes in Central African Republic strains (CAR. Results In this study, 55 MDR and 194 non-MDR strains were analyzed. Variations in DNA repair genes ada/alkA and ogt were identified. Among them, by comparison to M. tuberculosis published sequences, we found a non-sense variation in ada/alkA gene which was also observed in M. bovis AF2122 strain. SNPs that are present in the adjacent regions to the amber variation are different in M. bovis and in M. tuberculosis strain. Conclusion An Amber codon was found in the ada/alkA locus of clustered M. tuberculosis isolates and in M. bovis strain AF2122. This is likely due to convergent evolution because SNP differences between strains are incompatible with horizontal transfer of an entire gene. This suggests that such a variation may confer a selective advantage and be implicated in hypermutator phenotype expression, which in turn contributes to adaptation to environmental changes.

  15. Evaluation of RNA extraction methods and identification of putative reference genes for real-time quantitative polymerase chain reaction expression studies on olive (Olea europaea L.) fruits.

    Science.gov (United States)

    Nonis, Alberto; Vezzaro, Alice; Ruperti, Benedetto

    2012-07-11

    Genome wide transcriptomic surveys together with targeted molecular studies are uncovering an ever increasing number of differentially expressed genes in relation to agriculturally relevant processes in olive (Olea europaea L). These data need to be supported by quantitative approaches enabling the precise estimation of transcript abundance. qPCR being the most widely adopted technique for mRNA quantification, preliminary work needs to be done to set up robust methods for extraction of fully functional RNA and for the identification of the best reference genes to obtain reliable quantification of transcripts. In this work, we have assessed different methods for their suitability for RNA extraction from olive fruits and leaves and we have evaluated thirteen potential candidate reference genes on 21 RNA samples belonging to fruit developmental/ripening series and to leaves subjected to wounding. By using two different algorithms, GAPDH2 and PP2A1 were identified as the best reference genes for olive fruit development and ripening, and their effectiveness for normalization of expression of two ripening marker genes was demonstrated.

  16. Lack of ligand-selective binding of the aryl hydrocarbon receptor to putative DNA binding sites regulating expression of Bax and paraoxonase 1 genes.

    Science.gov (United States)

    DeGroot, Danica E; Hayashi, Ai; Denison, Michael S

    2014-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that mediates the biological and toxicological effects of structurally diverse chemicals through its ability to bind specific DNA recognition sites (dioxin responsive elements (DREs)), and activate transcription of adjacent genes. While the DRE has a highly conserved consensus sequence, it has been suggested that the nucleotide specificity of AhR DNA binding may be ligand-dependent. The upstream regulatory regions of the murine Bax and human paraoxonase 1 (PON1) genes reportedly contain unique DRE-like sequences that respond to AhRs activated by some ligands but not others. Given the significant implications of this observation to understanding the diversity in AhR responses and that of other ligand-dependent nuclear receptors, a combination of DNA binding, nuclear translocation and gene expression analysis was used to investigate the molecular mechanisms underlying these ligand-selective responses. Although known AhR agonists stimulated AhR nuclear translocation, DRE binding and gene expression, the ligand-selective DRE-like DNA elements identified in the Bax and PON1 upstream regulatory regions failed to bind ligand-activated AhR or confer AhR-responsiveness upon a reporter gene. These results argue against the reported ligand-selectivity of AhR DNA binding and suggest DNA binding by ligand activated AhR involves DRE-containing DNA.

  17. A low-pungency S3212 genotype of Capsicum frutescens caused by a mutation in the putative aminotransferase (p-AMT) gene.

    Science.gov (United States)

    Park, Young-Jun; Nishikawa, Tomotaro; Minami, Mineo; Nemoto, Kazuhiro; Iwasaki, Tomohiro; Matsushima, Kenichi

    2015-12-01

    The purpose of this study was to identify the genetic mechanism underlying capsinoid biosynthesis in S3212, a low-pungency genotype of Capsicum frutescens. Screening of C. frutescens accessions for capsaicinoid and capsiate contents by high-performance liquid chromatography revealed that low-pungency S3212 contained high levels of capsiate but no capsaicin. Comparison of DNA coding sequences of pungent (T1 and Bird Eye) and low-pungency (S3212) genotypes uncovered a significant 12-bp deletion mutation in exon 7 of the p-AMT gene of S3212. In addition, p-AMT gene transcript levels in placental tissue were positively correlated with the degree of pungency. S3212, the low-pungency genotype, exhibited no significant p-AMT transcript levels, whereas T1, one of the pungent genotypes, displayed high transcript levels of this gene. We therefore conclude that the deletion mutation in the p-AMT gene is related to the loss of pungency in placental tissue and has given rise to the low-pungency S3212 C. frutescens genotype. C. frutescens S3212 represents a good natural source of capsinoids. Finally, our basic characterization of the uncovered p-AMT gene mutation should contribute to future studies of capsinoid biosynthesis in Capsicum.

  18. Detection of a putative virulence cadF gene of Campylobacter jejuni obtained from different sources using a microfabricated PCR chip

    DEFF Research Database (Denmark)

    Poulsen, Claus Riber; El-Ali, Jamil; Perch-Nielsen, Ivan R.

    2005-01-01

    A microfabricated polymerase chain reaction (PCR) chip made of epoxy-based photoresist (SU-8) was recently designed and developed. In this study, we tested whether the PCR chip could be used for rapid detection of a potential virulence determinant, the cadF gene of Campylobacter jejuni. PCR...... was performed using published PCR conditions and primers for the C. jejuni cadF gene. DNA isolated from a C. jejuni reference strain CCUG 11284, C. jejuni isolates obtained from different sources (chicken and human), and Campylobacter whole cells were used as templates in the PCR tests. Conventional PCR in tube...... was used as the control. After optimization of the PCR chip, PCR positives on the chip were obtained from 91.0% (10/11) of the tested chips. A fast transition time was achieved with the PCR chip, and therefore a faster cycling time and a shorter PCR program were obtained. Using the PCR chip, the cadF gene...

  19. Seasonal Differences in Relative Gene Expression of Putative Central Appetite Regulators in Arctic Charr (Salvelinus alpinus Do Not Reflect Its Annual Feeding Cycle.

    Directory of Open Access Journals (Sweden)

    Anja Striberny

    Full Text Available The highly seasonal anadromous Arctic charr (Salvelinus alpinus was used to investigate the possible involvement of altered gene expression of brain neuropeptides in seasonal appetite regulation. Pro-opiomelanocortin (POMCA1, POMCA2, Cocaine and amphetamine regulated transcript (CART, Agouti related Peptide (AgRP, Neuropeptide Y (NPY and Melanocortin Receptor 4 (MC4-R genes were examined. The function of centrally expressed Leptin (Lep in fish remains unclear, so Lep (LepA1, LepA2 and Leptin Receptor (LepR genes were included in the investigation. In a ten months study gene expression was analysed in hypothalamus, mesencephalon and telencephalon of immature charr held under natural photoperiod (69°38'N and ambient temperature and given excess feed. From April to the beginning of June the charr did not feed and lost weight, during July and August they were feeding and had a marked increase in weight and condition factor, and from November until the end of the study the charr lost appetite and decreased in weight and condition factor. Brain compartments were sampled from non-feeding charr (May, feeding charr (July, and non-feeding charr (January. Reverse transcription real-time quantitative PCR revealed temporal patterns of gene expression that differed across brain compartments. The non-feeding charr (May, January had a lower expression of the anorexigenic LepA1, MC4-R and LepR in hypothalamus and a higher expression of the orexigenic NPY and AgRP in mesencephalon, than the feeding charr (July. In the telencephalon, LepR was more highly expressed in January and May than in July. These results do not indicate that changes in central gene expression of the neuropeptides investigated here directly induce seasonal changes in feeding in Arctic charr.

  20. TCDD and a putative endogenous AhR ligand, ITE, elicit the same immediate changes in gene expression in mouse lung fibroblasts.

    Science.gov (United States)

    Henry, Ellen C; Welle, Stephen L; Gasiewicz, Thomas A

    2010-03-01

    The aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, mediates toxicity of several classes of xenobiotics and also has important physiological roles in differentiation, reproduction, and immunity, although the endogenous ligand(s) mediating these functions is/are as yet unidentified. One candidate endogenous ligand, 2-(1'H-indolo-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE), is a potent AhR agonist in vitro, activates the murine AhR in vivo, but does not induce toxicity. We hypothesized that ITE and the toxic ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), may modify transcription of different sets of genes to account for their different toxicity. To test this hypothesis, primary mouse lung fibroblasts were exposed to 0.5muM ITE, 0.2nM TCDD, or vehicle for 4 h, and total gene expression was evaluated using microarrays. After this short-term and low-dose treatment, several hundred genes were changed significantly, and the response to ITE and TCDD was remarkably similar, both qualitatively and quantitatively. Induced gene sets included the expected battery of AhR-dependent xenobiotic-metabolizing enzymes, as well as several sets that reflect the inflammatory role of lung fibroblasts. Real time quantitative RT-qPCR assay of several selected genes confirmed these microarray data and further suggested that there may be kinetic differences in expression between ligands. These data suggest that ITE and TCDD elicit an analogous change in AhR conformation such that the initial transcription response is the same. Furthermore, if the difference in toxicity between TCDD and ITE is mediated by differences in gene expression, then it is likely that secondary changes enabled by the persistent TCDD, but not by the shorter lived ITE, are responsible.

  1. Integrated genome-wide genotyping and gene expression profiling reveals BCL11B as a putative oncogene in acute myeloid leukemia with 14q32 aberrations.

    Science.gov (United States)

    Abbas, Saman; Sanders, Mathijs A; Zeilemaker, Annelieke; Geertsma-Kleinekoort, Wendy M C; Koenders, Jasper E; Kavelaars, Francois G; Abbas, Zabiollah G; Mahamoud, Souad; Chu, Isabel W T; Hoogenboezem, Remco; Peeters, Justine K; van Drunen, Ellen; van Galen, Janneke; Beverloo, H Berna; Löwenberg, Bob; Valk, Peter J M

    2014-05-01

    Acute myeloid leukemia is a neoplasm characterized by recurrent molecular aberrations traditionally demonstrated by cytogenetic analyses. We used high density genome-wide genotyping and gene expression profiling to reveal acquired cryptic abnormalities in acute myeloid leukemia. By genome-wide genotyping of 137 cases of primary acute myeloid leukemia, we disclosed a recurrent focal amplification on chromosome 14q32, which included the genes BCL11B, CCNK, C14orf177 and SETD3, in two cases. In the affected cases, the BCL11B gene showed consistently high mRNA expression, whereas the expression of the other genes was unperturbed. Fluorescence in situ hybridization on 40 cases of acute myeloid leukemia with high BCL11B mRNA expression [2.5-fold above median; 40 out of 530 cases (7.5%)] revealed 14q32 abnormalities in two additional cases. In the four BCL11B-rearranged cases the 14q32 locus was fused to different partner chromosomes. In fact, in two cases, we demonstrated that the focal 14q32 amplifications were integrated into transcriptionally active loci. The translocations involving BCL11B result in increased expression of full-length BCL11B protein. The BCL11B-rearranged acute myeloid leukemias expressed both myeloid and T-cell markers. These biphenotypic acute leukemias all carried FLT3 internal tandem duplications, a characteristic marker of acute myeloid leukemia. BCL11B mRNA expression in acute myeloid leukemia appeared to be strongly associated with expression of other T-cell-specific genes. Myeloid 32D(GCSF-R) cells ectopically expressing Bcl11b showed decreased proliferation rate and less maturation. In conclusion, by an integrated approach involving high-throughput genome-wide genotyping and gene expression profiling we identified BCL11B as a candidate oncogene in acute myeloid leukemia.

  2. Determination of microbial diversity of Aeromonas strains on the basis of multilocus sequence typing, phenotype, and presence of putative virulence genes.

    Science.gov (United States)

    Martino, Maria Elena; Fasolato, Luca; Montemurro, Filomena; Rosteghin, Marina; Manfrin, Amedeo; Patarnello, Tomaso; Novelli, Enrico; Cardazzo, Barbara

    2011-07-01

    The genus Aeromonas has been described as comprising several species associated with the aquatic environment, which represents their principal reservoir. Aeromonas spp. are commonly isolated from diseased and healthy fish, but the involvement of such bacteria in human infection and gastroenteritis has frequently been reported. The primary challenge in establishing an unequivocal link between the Aeromonas genus and pathogenesis in humans is the extremely complicated taxonomy. With the aim of clarifying taxonomic relationships among the strains and phenotypes, a multilocus sequencing approach was developed and applied to characterize 23 type and reference strains of Aeromonas spp. and a collection of 77 field strains isolated from fish, crustaceans, and mollusks. All strains were also screened for putative determinants of virulence by PCR (ast, ahh1, act, asa1, eno, ascV, and aexT) and the production of acylated homoserine lactones (AHLs). In addition, the phenotypic fingerprinting obtained from 29 biochemical tests was submitted to the nonparametric combination (NPC) test methodology to define the statistical differences among the identified genetic clusters. Multilocus sequence typing (MLST) achieved precise strain genotyping, and the phylogenetic analysis of concatenated sequences delineated the relationship among the taxa belonging to the genus Aeromonas, providing a powerful tool for outbreak traceability, host range diffusion, and ecological studies. The NPC test showed the feasibility of phenotypic differentiation among the majority of the MLST clusters by using a selection of tests or the entire biochemical fingerprinting. A Web-based MLST sequence database (http://pubmlst.org/aeromonas) specific for the Aeromonas genus was developed and implemented with all the results.

  3. Roles of Prolyl Isomerases in RNA-Mediated Gene Expression

    Directory of Open Access Journals (Sweden)

    Roopa Thapar

    2015-05-01

    Full Text Available The peptidyl-prolyl cis-trans isomerases (PPIases that include immunophilins (cyclophilins and FKBPs and parvulins (Pin1, Par14, Par17 participate in cell signaling, transcription, pre-mRNA processing and mRNA decay. The human genome encodes 19 cyclophilins, 18 FKBPs and three parvulins. Immunophilins are receptors for the immunosuppressive drugs cyclosporin A, FK506, and rapamycin that are used in organ transplantation. Pin1 has also been targeted in the treatment of Alzheimer’s disease, asthma, and a number of cancers. While these PPIases are characterized as molecular chaperones, they also act in a nonchaperone manner to promote protein-protein interactions using surfaces outside their active sites. The immunosuppressive drugs act by a gain-of-function mechanism by promoting protein-protein interactions in vivo. Several immunophilins have been identified as components of the spliceosome and are essential for alternative splicing. Pin1 plays roles in transcription and RNA processing by catalyzing conformational changes in the RNA Pol II C-terminal domain. Pin1 also binds several RNA binding proteins such as AUF1, KSRP, HuR, and SLBP that regulate mRNA decay by remodeling mRNP complexes. The functions of ribonucleoprotein associated PPIases are largely unknown. This review highlights PPIases that play roles in RNA-mediated gene expression, providing insight into their structures, functions and mechanisms of action in mRNP remodeling in vivo.

  4. Functional characterization of the gene FoOCH1 encoding a putative α-1,6-mannosyltransferase in Fusarium oxysporum f. sp. cubense.

    Science.gov (United States)

    Li, Min-Hui; Xie, Xiao-Ling; Lin, Xian-Feng; Shi, Jin-Xiu; Ding, Zhao-Jian; Ling, Jin-Feng; Xi, Ping-Gen; Zhou, Jia-Nuan; Leng, Yueqiang; Zhong, Shaobin; Jiang, Zi-De

    2014-04-01

    Fusarium oxysporum f. sp. cubense (FOC) is the causal agent of banana Fusarium wilt and has become one of the most destructive pathogens threatening the banana production worldwide. However, few genes related to morphogenesis and pathogenicity of this fungal pathogen have been functionally characterized. In this study, we identified and characterized the disrupted gene in a T-DNA insertional mutant (L953) of FOC with significantly reduced virulence on banana plants. The gene disrupted by T-DNA insertion in L953 harbors an open reading frame, which encodes a protein with homology to α-1,6-mannosyltransferase (OCH1) in fungi. The deletion mutants (ΔFoOCH1) of the OCH1 orthologue (FoOCH1) in FOC were impaired in fungal growth, exhibited brighter staining with fluorescein isothiocyanate (FITC)-Concanavalin A, had less cell wall proteins and secreted more proteins into liquid media than the wild type. Furthermore, the mutation or deletion of FoOCH1 led to loss of ability to penetrate cellophane membrane and decline in hyphal attachment and colonization as well as virulence to the banana host. The mutant phenotypes were fully restored by complementation with the wild type FoOCH1 gene. Our data provide a first evidence for the critical role of FoOCH1 in maintenance of cell wall integrity and virulence of F. oxysporum f. sp. cubense. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Detection of a Putative TetR-Like Gene Related to Mycobacterium bovis BCG Growth in Cholesterol Using a gfp-Transposon Mutagenesis System

    Science.gov (United States)

    Otal, Isabel; Pérez-Herrán, Esther; Garcia-Morales, Lazaro; Menéndez, María C.; Gonzalez-y-Merchand, Jorge A.; Martín, Carlos; García, María J.

    2017-01-01

    In vitro transposition is a powerful genetic tool for identifying mycobacterial virulence genes and studying virulence factors in relation to the host. Transposon shuttle mutagenesis is a method for constructing stable insertions in the genome of different microorganisms including mycobacteria. Using an IS1096 derivative, we have constructed the Tngfp, a transposon containing a promoterless green fluorescent protein (gfp) gene. This transposon was able to transpose randomly in Mycobacterium bovis BCG. Bacteria with a single copy of the gfp gene per chromosome from an M. bovis BCG::Tngfp library were analyzed and cells exhibiting high levels of fluorescence were detected by flow cytometry. Application of this approach allowed for the selection of a mutant, BCG_2177c::Tngfp (BCG-Tn), on the basis of high level of long-standing fluorescence at stationary phase. This BCG-Tn mutant showed some particular phenotypic features compared to the wild type strain, mainly during stationary phase, when cholesterol was used as a sole carbon source, thus supporting the relationships of the targeted gene with the regulation of cholesterol metabolism in this bacteria. This approach showed that Tngfp is a potentially useful tool for studying the involvement of the targeted loci in metabolic pathways of mycobacteria. PMID:28321208

  6. PtSRR1, a putative Pisolithus tinctorius symbiosis related receptor gene is expressed during the first hours of mycorrhizal interaction with Castanea sativa roots.

    Science.gov (United States)

    Acioli-Santos, B; Malosso, E; Calzavara-Silva, C E; Lima, C E P; Figueiredo, A; Sebastiana, M; Pais, M S

    2009-04-01

    PtSRR1 EST was previously identified in the first hours of Pisolithus tinctorius and Castanea sativa interaction. QRT-PCR confirmed PtSRR1 early expression and in silico preliminary translated peptide analysis indicated a strong probability that PtSRR1 be a transmembrane protein. These data stimulate the PtSRR1 gene research during ectomycorrhiza formation.

  7. Potentially harmful advantage to athletes: a putative connection between UGT2B17 gene deletion polymorphism and renal disorders with prolonged use of anabolic androgenic steroids

    Directory of Open Access Journals (Sweden)

    Barker James

    2010-04-01

    Full Text Available Abstract Background and objective With prolonged use of anabolic androgenic steroids (AAS, occasional incidents of renal disorders have been observed. Independently, it has also been established that there are considerable inter-individual and inter-ethnic differences, in particular with reference to the uridine diphosphate-glucuronosyltransferase 2B17 (UGT2B17 gene, in metabolising these compounds. This report postulates the association of deletion polymorphism in the UGT2B17 gene with the occurrence of renal disorders on chronic exposure to AAS. Presentation of the hypothesis The major deactivation and elimination pathway of AASs is through glucuronide conjugation, chiefly catalyzed by the UGT2B17 enzyme, followed by excretion in urine. Excretion of steroids is affected in individuals with a deletion mutation in the UGT2B17 gene. We hypothesize that UGT2B17 deficient individuals are more vulnerable to developing renal disorders with prolonged use of AAS owing to increases in body mass index and possible direct toxic effects of steroids on the kidneys. Elevated serum levels of biologically active steroids due to inadequate elimination can lead to prolonged muscle build up. An increase in body mass index may cause renal injuries due to sustained elevated glomerular pressure and flow rate. Testing the hypothesis In the absence of controlled clinical trials in humans, observational studies can be carried out. Real time PCR with allelic discrimination should be employed to examine the prevalence of different UGT2B17 genotypes in patients with impaired renal function and AAS abuse. In individuals with the UGT2B17 deletion polymorphism, blood tests, biofluid analyses, urinalysis, and hair analyses following the administration of an anabolic steroid can be used to determine the fate of the substance once in the body. Implications of the hypothesis If the hypothesis is upheld, anabolic steroid users with a deletion mutation in the UGT2B17 gene may be

  8. Elimination of manganese(II,III) oxidation in Pseudomonas putida GB-1 by a double knockout of two putative multicopper oxidase genes.

    Science.gov (United States)

    Geszvain, Kati; McCarthy, James K; Tebo, Bradley M

    2013-01-01

    Bacterial manganese(II) oxidation impacts the redox cycling of Mn, other elements, and compounds in the environment; therefore, it is important to understand the mechanisms of and enzymes responsible for Mn(II) oxidation. In several Mn(II)-oxidizing organisms, the identified Mn(II) oxidase belongs to either the multicopper oxidase (MCO) or the heme peroxidase family of proteins. However, the identity of the oxidase in Pseudomonas putida GB-1 has long remained unknown. To identify the P. putida GB-1 oxidase, we searched its genome and found several homologues of known or suspected Mn(II) oxidase-encoding genes (mnxG, mofA, moxA, and mopA). To narrow this list, we assumed that the Mn(II) oxidase gene would be conserved among Mn(II)-oxidizing pseudomonads but not in nonoxidizers and performed a genome comparison to 11 Pseudomonas species. We further assumed that the oxidase gene would be regulated by MnxR, a transcription factor required for Mn(II) oxidation. Two loci met all these criteria: PputGB1_2447, which encodes an MCO homologous to MnxG, and PputGB1_2665, which encodes an MCO with very low homology to MofA. In-frame deletions of each locus resulted in strains that retained some ability to oxidize Mn(II) or Mn(III); loss of oxidation was attained only upon deletion of both genes. These results suggest that PputGB1_2447 and PputGB1_2665 encode two MCOs that are independently capable of oxidizing both Mn(II) and Mn(III). The purpose of this redundancy is unclear; however, differences in oxidation phenotype for the single mutants suggest specialization in function for the two enzymes.

  9. Identification of Genes Putatively Involved in Chitin Metabolism and Insecticide Detoxification in the Rice Leaf Folder (Cnaphalocrocis medinalis) Larvae through Transcriptomic Analysis

    OpenAIRE

    2015-01-01

    The rice leaf roller (Cnaphalocrocis medinalis) is one of the most destructive agricultural pests. Due to its migratory behavior, it is difficult to control worldwide. To date, little is known about major genes of C. medinalis involved in chitin metabolism and insecticide detoxification. In order to obtain a comprehensive genome dataset of C. medinalis, we conducted de novo transcriptome sequencing which focused on the major feeding stage of fourth-instar larvae, and our work revealed usef...

  10. Ribosomal genes and heat shock proteins as putative markers for chronic, sublethal heat stress in Arctic charr: applications for aquaculture and wild fish.

    Science.gov (United States)

    Quinn, Nicole L; McGowan, Colin R; Cooper, Glenn A; Koop, Ben F; Davidson, William S

    2011-09-22

    Arctic charr thrive at high densities and can live in freshwater year round, making this species especially suitable for inland, closed containment aquaculture. However, it is a cold-water salmonid, which both limits where the species can be farmed and places wild populations at particular risk to climate change. Previously, we identified genes associated with tolerance and intolerance to acute, lethal temperature stress in Arctic charr. However, there remained a need to examine the genes involved in the stress response to more realistic temperatures that could be experienced during a summer heat wave in grow-out tanks that are not artificially cooled, or under natural conditions. Here, we exposed Arctic charr to sublethal heat stress of 15-18°C for 72 h, and gill tissues extracted before, during (i.e., at 72 h), immediately after cooling and after 72 h of recovery at ambient temperature (6°C) were used for gene expression profiling by microarray and qPCR analyses. The results revealed an expected pattern for heat shock protein expression, which was highest during heat exposure, with significantly reduced expression (approaching control levels) quickly thereafter. We also found that the expression of numerous ribosomal proteins was significantly elevated immediately and 72 h after cooling, suggesting that the gill tissues were undergoing ribosome biogenesis while recovering from damage caused by heat stress. We suggest that these are candidate gene targets for the future development of genetic markers for broodstock development or for monitoring temperature stress and recovery in wild or cultured conditions.

  11. Molecular Modeling Approaches for Determining Gene Function: application to a Putative Poly-A Binding Protein from Leishmania amazonensis (LaPABP

    Directory of Open Access Journals (Sweden)

    Silva-Jr FP

    2002-01-01

    Full Text Available The great expansion in the number of genome sequencing projects has revealed the importance of computational methods to speed up the characterization of unknown genes. These studies have been improved by the use of three dimensional information from the predicted proteins generated by molecular modeling techniques. In this work, we disclose the structure-function relationship of a gene product from Leishmania amazonensis by applying molecular modeling and bioinformatics techniques. The analyzed sequence encodes a 159 aminoacids polypeptide (estimated 18 kDa and was denoted LaPABP for its high homology with poly-A binding proteins from trypanosomatids. The domain structure, clustering analysis and a three dimensional model of LaPABP, basically obtained by homology modeling on the structure of the human poly-A binding protein, are described. Based on the analysis of the electrostatic potential mapped on the model's surface and conservation of intramolecular contacts responsible for folding stabilization we hypothesize that this protein may have less avidity to RNA than it's L. major counterpart but still account for a significant functional activity in the parasite. The model obtained will help in the design of mutagenesis experiments aimed to elucidate the mechanism of gene expression in trypanosomatids and serve as a starting point for its exploration as a potential source of targets for a rational chemotherapy.

  12. HACE1 is a putative tumor suppressor gene in B-cell lymphomagenesis and is down-regulated by both deletion and epigenetic alterations.

    Science.gov (United States)

    Bouzelfen, Abdelilah; Alcantara, Marion; Kora, Hafid; Picquenot, Jean-Michel; Bertrand, Philippe; Cornic, Marie; Mareschal, Sylvain; Bohers, Elodie; Maingonnat, Catherine; Ruminy, Philippe; Adriouch, Sahil; Boyer, Olivier; Dubois, Sydney; Bastard, Christian; Tilly, Hervé; Latouche, Jean-Baptiste; Jardin, Fabrice

    2016-06-01

    HECT domain and ankyrin repeat containing E3 ubiquitin protein ligase 1, HACE1, located on chromosome 6q, encodes an E3 ubiquitin ligase and is downregulated in many human tumors. Here, we report HACE1 as a candidate tumor suppressor gene down-regulated by a combination of deletion and epigenetic mechanisms. HACE1 deletions were observed in 40% of B-cell lymphoma tumors. Hypermethylation of the HACE1 promoter CpG177 island was found in 60% (68/111) of cases and in all tested B-cell lymphoma lines. Using HDAC inhibitors, we observed predominantly inactive chromatin conformation (methylated H3 histones H3K9me2) in HACE1 gene promoter region. We demonstrated in Ramos and Raji cells that down-regulation of HACE1 expression was associated with a significant decrease in apoptosis and an accumulation of cells in the S and G2/M phases. Our experiments indicate that HACE1 can act as a haploinsufficient tumor suppressor gene in most B-cell lymphomas and can be downregulated by deacetylation of its promoter region chromatin, which makes HACE1 a potential target for HDAC inhibitors.

  13. cDNA cloning, tissue distribution, and chromosomal localization of Ocp2, a gene encoding a putative transcription-associated factor predominantly expressed in the auditory organs

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Hong; Thalmann, I.; Thalmann, R. [Washington Univ., St. Louis, MO (United States)] [and others

    1995-06-10

    We report the cloning of the Ocp2 gene encoding OCP-II from a guinea pig organ-of-Corti cDNA library. The predicted open reading frame encodes a protein of 163 amino acids with an estimated molecular mass of 18.6 kDa. A homology search revealed that Ocp2 shares significant sequence similarity with p15, a sub-unit of transcription factor SIII that regulates the activity of the RNA polymerase II elongation complex. The Ocp2 messenger RNA is expressed abundantly in the cochlea while not significantly in any other tissues examined, including brain, eye, heart, intestine, kidney, liver, lung, thigh muscle, and testis, demonstrating that the expression of this gene may be restricted to auditory organs. A polyclonal antiserum was raised against the N-terminal region of OCP-II. Immunohistochemical staining of paraffin-embedded sections of the cochlea showed that OCP-II is localized abundantly in nonsensory cells in the organ of Corti; in addition, it was also detected, at a lower concentration, in vestibular sensory organs, as well as auditory and vestibular brain stem nuclei. The Ocp2 gene was mapped to mouse chromosome 4 as well as 11. Our results suggest that OCP-II may be involved in transcription regulation for the development or maintenance of specialized functions of the inner ear. 40 refs., 5 figs.

  14. Identification and characterization of genes, encoding the 3-hydroxybutyrate dehydrogenase and a putative lipase, in an avirulent spontaneous Legionella pneumophila serogroup 6 mutant.

    Science.gov (United States)

    Scaturro, Maria; Barello, Cristina; Giusti, Melania De; Fontana, Stefano; Pinci, Federica; Giuffrida, Maria Gabriella; Ricci, Maria Luisa

    2015-04-01

    Legionella pneumophila is a pathogen widespread in aquatic environment, able to multiply both within amoebae and human macrophages. The aim of this study was to identify genes differently expressed in a spontaneous avirulent Legionella pneumophila serogroup 6 mutant, named Vir-, respect the parental strain (Vir+), and to determine their role in the loss of virulence. Protein profiles revealed some differences in Vir- proteomic maps, and among the identified proteins the undetectable 3-hydroxybutyrate dehydrogenase (BdhA) and a down-produced lipase. Both Legionella enzymes were studied before and were here further characterized at genetic level. A significant down-regulation of both genes was observed in Vir- at the transcriptional level, but the use of defined mutants demonstrated that they did not affect the intracellular multiplication. A mutant (MS1) showed an accumulation of poly-3-hydroxybutyrate (PHB) granules suggesting a role of bdhA gene in its degradation process. The lipase deduced amino acid sequence revealed a catalytic triad, typical of the 'lipase box' characteristic of PHB de-polymerase enzymes, that let us suppose a possible involvement of lipase in the PHB granule degradation process. Our results revealed unexpected alterations in secondary metabolic pathways possibly linking the loss of virulence to Legionella lack of energy sources.

  15. Genetic susceptibility on CagA-interacting molecules and gene-environment interaction with phytoestrogens: a putative risk factor for gastric cancer.

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    Jae Jeong Yang

    Full Text Available OBJECTIVES: To evaluate whether genes that encode CagA-interacting molecules (SRC, PTPN11, CRK, CRKL, CSK, c-MET and GRB2 are associated with gastric cancer risk and whether an interaction between these genes and phytoestrogens modify gastric cancer risk. METHODS: In the discovery phase, 137 candidate SNPs in seven genes were analyzed in 76 incident gastric cancer cases and 322 matched controls from the Korean Multi-Center Cancer Cohort. Five significant SNPs in three genes (SRC, c-MET and CRK were re-evaluated in 386 cases and 348 controls in the extension phase. Odds ratios (ORs for gastric cancer risk were estimated adjusted for age, smoking, H. pylori seropositivity and CagA strain positivity. Summarized ORs in the total study population (462 cases and 670 controls were presented using pooled- and meta-analysis. Plasma concentrations of phytoestrogens (genistein, daidzein, equol and enterolactone were measured using the time-resolved fluoroimmunoassay. RESULTS: SRC rs6122566, rs6124914, c-MET rs41739, and CRK rs7208768 showed significant genetic effects for gastric cancer in both the pooled and meta-analysis without heterogeneity (pooled OR = 3.96 [95% CI 2.05-7.65], 1.24 [95% CI = 1.01-1.53], 1.19 [95% CI = 1.01-1.41], and 1.37 [95% CI = 1.15-1.62], respectively; meta OR = 4.59 [95% CI 2.74-7.70], 1.36 [95% CI = 1.09-1.70], 1.20 [95% CI = 1.00-1.44], and 1.32 [95% CI = 1.10-1.57], respectively. Risk allele of CRK rs7208768 had a significantly increased risk for gastric cancer at low phytoestrogen levels (p interaction<0.05. CONCLUSIONS: Our findings suggest that SRC, c-MET and CRK play a key role in gastric carcinogenesis by modulating CagA signal transductions and interaction between CRK gene and phytoestrogens modify gastric cancer risk.

  16. The dtd gene from Bacillus amyloliquefaciens encodes a putative D-tyrosyl-tRNATyr deacylase and is a selectable marker for Bacillus subtilis.

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    Geraskina, Natalia V; Butov, Ivan A; Yomantas, Yurgis A V; Stoynova, Nataliya V

    2015-02-01

    Genetically engineered microbes are of high practical importance due to their cost-effective production of valuable metabolites and enzymes, and the search for new selectable markers for genetic manipulation is of particular interest. Here, we revealed that the soil bacterium Bacillus amyloliquefaciens A50 is tolerant to the non-canonical amino acid D-tyrosine (D-Tyr), in contrast to the closely related Bacillus strain B. subtilis 168, which is a widely used "domesticated" laboratory strain. The gene responsible for resistance to D-Tyr was identified. The resistance was associated with the activity of a potential D-tyrosyl-tRNA(Tyr) deacylase. Orthologs of this enzyme are capable of hydrolyzing the ester bond and recycling misacetylated D-aminoacyl-tRNA molecules into free tRNAs and D-amino acids. This gene, yrvI (dtd), is applicable as a convenient, small selectable marker for non-antibiotic resistance selection in experiments aimed at genome editing of D-Tyr-sensitive microorganisms.

  17. An unusual gene arrangement for the putative chromosome replication origin and circadian expression of dnaN in Synechococcus sp. strain PCC 7942.

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    Liu, Y; Tsinoremas, N F

    1996-06-12

    In eubacteria, the clustering of DnaA boxes around the dnaN (beta subunit of DNA polymerase III) and dnaA genes usually defines the chromosome replication origin (oriC). In this study, the dnaN locus from the cyanobacterium Synechococcus sp. strain PCC 7942 was sequenced. The gene order in this region is cbbZp-dnaN-orf288-purL-purF which contrasts with other eubacteria. A cluster of eleven DnaA boxes (consensus sequence: TTTTCCACA) was found in the intergenic region between dnaN and cbbZp. We also found a 41-bp sequence within this region that is 80% identical to the proposed oriC of Streptomyces coelicolor. Therefore, we propose that this intergenic region may serve as an oriC in Synechococcus. Using bacterial luciferase as a reporter, we also showed that dnaN is rhythmically expressed, suggesting that DNA replication could be under circadian control in this organism.

  18. The maize glossy13 gene, cloned via BSR-Seq and Seq-walking encodes a putative ABC transporter required for the normal accumulation of epicuticular waxes.

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    Li Li

    Full Text Available Aerial plant surfaces are covered by epicuticular waxes that among other purposes serve to control water loss. Maize glossy mutants originally identified by their "glossy" phenotypes exhibit alterations in the accumulation of epicuticular waxes. By combining data from a BSR-Seq experiment and the newly developed Seq-Walking technology, GRMZM2G118243 was identified as a strong candidate for being the glossy13 gene. The finding that multiple EMS-induced alleles contain premature stop codons in GRMZM2G118243, and the one knockout allele of gl13, validates the hypothesis that gene GRMZM2G118243 is gl13. Consistent with this, GRMZM2G118243 is an ortholog of AtABCG32 (Arabidopsis thaliana, HvABCG31 (barley and OsABCG31 (rice, which encode ABCG subfamily transporters involved in the trans-membrane transport of various secondary metabolites. We therefore hypothesize that gl13 is involved in the transport of epicuticular waxes onto the surfaces of seedling leaves.

  19. Association of elevated rotavirus-specific antibody titers with HBGA secretor status in Swedish individuals: The FUT2 gene as a putative susceptibility determinant for infection.

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    Günaydın, Gökçe; Nordgren, Johan; Sharma, Sumit; Hammarström, Lennart

    2016-01-04

    The histo-blood group antigens (HBGAs) have recently been suggested to serve as attachment factors for rotavirus VP8* (P-genotype) in vitro and associated with susceptibility in vivo. We thus investigated whether rotavirus antibody titers and genotype specific neutralization titers correlate with HBGA status in Swedish individuals. We investigated the effect of inactivating mutations in the secretor FUT2 (rs601338) and Lewis FUT3 genes (rs28362459, rs3894326, rs812936 and rs778986) on serum IgG antibody titers and neutralizing antibody titers to rotavirus strains of the P[8] and P[6] genotypes in Swedish healthy blood donors and patients with IgA deficiency using genotyping, enzyme linked immunosorbent assay and a neutralization assay. Rotavirus-specific serum IgG and neutralizing antibody titers to the Wa strain (G1P[8]), but not to the ST3 (G4P[6]) strain, were significantly higher in secretors (with at least one functional FUT2 gene) than in non-secretors (Protavirus specific serum antibodies, suggesting a higher susceptibility to rotavirus infections, as compared to non-secretors in Sweden.

  20. De novo assembly, functional annotation and comparative analysis of Withania somnifera leaf and root transcriptomes to identify putative genes involved in the withanolides biosynthesis.

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    Gupta, Parul; Goel, Ridhi; Pathak, Sumya; Srivastava, Apeksha; Singh, Surya Pratap; Sangwan, Rajender Singh; Asif, Mehar Hasan; Trivedi, Prabodh Kumar

    2013-01-01

    Withania somnifera is one of the most valuable medicinal plants used in Ayurvedic and other indigenous medicine systems due to bioactive molecules known as withanolides. As genomic information regarding this plant is very limited, little information is available about biosynthesis of withanolides. To facilitate the basic understanding about the withanolide biosynthesis pathways, we performed transcriptome sequencing for Withania leaf (101L) and root (101R) which specifically synthesize withaferin A and withanolide A, respectively. Pyrosequencing yielded 8,34,068 and 7,21,755 reads which got assembled into 89,548 and 1,14,814 unique sequences from 101L and 101R, respectively. A total of 47,885 (101L) and 54,123 (101R) could be annotated using TAIR10, NR, tomato and potato databases. Gene Ontology and KEGG analyses provided a detailed view of all the enzymes involved in withanolide backbone synthesis. Our analysis identified members of cytochrome P450, glycosyltransferase and methyltransferase gene families with unique presence or differential expression in leaf and root and might be involved in synthesis of tissue-specific withanolides. We also detected simple sequence repeats (SSRs) in transcriptome data for use in future genetic studies. Comprehensive sequence resource developed for Withania, in this study, will help to elucidate biosynthetic pathway for tissue-specific synthesis of secondary plant products in non-model plant organisms as well as will be helpful in developing strategies for enhanced biosynthesis of withanolides through biotechnological approaches.

  1. De novo assembly, functional annotation and comparative analysis of Withania somnifera leaf and root transcriptomes to identify putative genes involved in the withanolides biosynthesis.

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    Parul Gupta

    Full Text Available Withania somnifera is one of the most valuable medicinal plants used in Ayurvedic and other indigenous medicine systems due to bioactive molecules known as withanolides. As genomic information regarding this plant is very limited, little information is available about biosynthesis of withanolides. To facilitate the basic understanding about the withanolide biosynthesis pathways, we performed transcriptome sequencing for Withania leaf (101L and root (101R which specifically synthesize withaferin A and withanolide A, respectively. Pyrosequencing yielded 8,34,068 and 7,21,755 reads which got assembled into 89,548 and 1,14,814 unique sequences from 101L and 101R, respectively. A total of 47,885 (101L and 54,123 (101R could be annotated using TAIR10, NR, tomato and potato databases. Gene Ontology and KEGG analyses provided a detailed view of all the enzymes involved in withanolide backbone synthesis. Our analysis identified members of cytochrome P450, glycosyltransferase and methyltransferase gene families with unique presence or differential expression in leaf and root and might be involved in synthesis of tissue-specific withanolides. We also detected simple sequence repeats (SSRs in transcriptome data for use in future genetic studies. Comprehensive sequence resource developed for Withania, in this study, will help to elucidate biosynthetic pathway for tissue-specific synthesis of secondary plant products in non-model plant organisms as well as will be helpful in developing strategies for enhanced biosynthesis of withanolides through biotechnological approaches.

  2. A gene-rich linkage map in the dioecious species Actinidia chinensis (kiwifruit reveals putative X/Y sex-determining chromosomes

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    Gill Geoffrey P

    2009-03-01

    Full Text Available Abstract Background The genus Actinidia (kiwifruit consists of woody, scrambling vines, native to China, and only recently propagated as a commercial crop. All species described are dioecious, but the genetic mechanism for sex-determination is unknown, as is the genetic basis for many of the cluster of characteristics making up the unique fruit. It is, however, an important crop in the New Zealand economy, and a classical breeding program would benefit greatly by knowledge of the trait alleles carried by both female and male parents. The application of marker assisted selection (MAS in seedling populations would also aid the accurate and efficient development of novel fruit types for the market. Results Gene-rich female, male and consensus linkage maps of the diploid species A. chinensis have been constructed with 644 microsatellite markers. The maps consist of twenty-nine linkage groups corresponding to the haploid number n = 29. We found that sex-linked sequence characterized amplified region (SCAR markers and the 'Flower-sex' phenotype consistently mapped to a single linkage group, in a subtelomeric region, in a section of inconsistent marker order. The region also contained markers of expressed genes, some of unknown function. Recombination, assessed by allelic distribution and marker order stability, was, in the remainder of the linkage group, in accordance with other linkage groups. Fully informative markers to other genes in this linkage group identified the comparative linkage group in the female map, where recombination ratios determining marker order were similar to the autosomes. Conclusion We have created genetic linkage maps that define the 29 linkage groups of the haploid genome, and have revealed the position and extent of the sex-determining locus in A. chinensis. As all Actinidia species are dioecious, we suggest that the sex-determining loci of other Actinidia species will be similar to that region defined in our maps. As the

  3. Characterization and expression analysis of mcoln1.1 and mcoln1.2, the putative zebrafish co-orthologs of the gene responsible for human mucolipidosis type IV.

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    Benini, Anna; Bozzato, Andrea; Mantovanelli, Silvia; Calvarini, Laura; Giacopuzzi, Edoardo; Bresciani, Roberto; Moleri, Silvia; Zizioli, Daniela; Beltrame, Monica; Borsani, Giuseppe

    2013-01-01

    Mucolipidosis type IV (MLIV) is an autosomal recessive lysosomal storage disorder caused by mutations in the MCOLN1 gene coding for mucolipin-1 (TRPML1). TRPML1 belongs to a transient receptor potential channels (TRP) subfamily, which in mammals includes two other members: mucolipin-2 (TRPML2) and mucolipin-3 (TRPML3). Bioinformatic analysis of the Danio rerio (zebrafish) genome and trascriptome revealed the presence of five different genes related to human mucolipins: mcoln1.1, mcoln1.2, mcoln2, mcoln3.1 and mcoln3.2. We focused our efforts on the characterization of the two putative zebrafish MCOLN1 co-orthologs. Transient-expression experiments in human HeLa cells demonstrated that fish Mcoln1.1 and Mcoln1.2, similarly to TRPML1, localize to late endosomal/lysosomal compartments. Real-Time PCR (RT-PCR) experiments showed that both genes are maternally expressed and transcribed at different levels during embryogenesis. RT-PCR analysis in different zebrafish tissues displayed ubiquitary expression for mcoln1.1 and a more tissue-specific pattern for mcoln1.2. Spatial and temporal expression studies using whole-mount in situ hybridization confirmed that both genes are maternally expressed and ubiquitously transcribed during gastrulation and early somitogenesis. Notably, in the next developmental stages they are more expressed in neural regions and in retina layers, tissues affected in MLIV. Interestingly, mcoln1.1 is detected, from 10 somite-stage until to 36 hpf, in the yolk syncytial layer (YSL) and in the intermediate cell mass (ICM), the earliest site of hematopoiesis. Overall, the redundancy of mucolipins together with their expression profile support the biological relevance of this class of proteins in zebrafish. The data herein presented indicate that Danio rerio could be a suitable vertebrate model for the study of some aspects of MLIV pathogenesis.

  4. Thermostable and site-specific DNA binding of the gene product ORF56 from the Sulfolobus islandicus plasmid pRN1, a putative archael plasmid copy control protein

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    Lipps, Georg; Stegert, Mario; Krauss, Gerhard

    2001-01-01

    There is still a lack of information on the specific characteristics of DNA-binding proteins from hyperthermophiles. Here we report on the product of the gene orf56 from plasmid pRN1 of the acidophilic and thermophilic archaeon Sulfolobus islandicus. orf56 has not been characterised yet but low sequence similarily to several eubacterial plasmid-encoded genes suggests that this 6.5 kDa protein is a sequence-specific DNA-binding protein. The DNA-binding properties of ORF56, expressed in Escherichia coli, have been investigated by EMSA experiments and by fluorescence anisotropy measurements. Recombinant ORF56 binds to double-stranded DNA, specifically to an inverted repeat located within the promoter of orf56. Binding to this site could down-regulate transcription of the orf56 gene and also of the overlapping orf904 gene, encoding the putative initiator protein of plasmid replication. By gel filtration and chemical crosslinking we have shown that ORF56 is a dimeric protein. Stoichiometric fluorescence anisotropy titrations further indicate that ORF56 binds as a tetramer to the inverted repeat of its target binding site. CD spectroscopy points to a significant increase in ordered secondary structure of ORF56 upon binding DNA. ORF56 binds without apparent cooperativity to its target DNA with a dissociation constant in the nanomolar range. Quantitative analysis of binding isotherms performed at various salt concentrations and at different temperatures indicates that approximately seven ions are released upon complex formation and that complex formation is accompanied by a change in heat capacity of –6.2 kJ/mol. Furthermore, recombinant ORF56 proved to be highly thermostable and is able to bind DNA up to 85°C. PMID:11160922

  5. Cloning of a putative monogalactosyldiacylglycerol synthase gene from rice (Oryza sativa L.) plants and its expression in response to submergence and other stresses.

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    Qi, Yanhua; Yamauchi, Yasuo; Ling, Jianqun; Kawano, Naoyoshi; Li, Debao; Tanaka, Kiyoshi

    2004-07-01

    Suppression subtractive hybridization was used to construct a subtractive cDNA library from plants of non-submerged and 7-day-submerged rice (Oryza sativa L., FR13A, a submergence-tolerant cultivar). One clone of the subtractive cDNA library, S23, was expressed abundantly during submergence. The full length of S23 was amplified using 5'- and 3'-rapid amplification of cDNA ends, and found to consist of 1,671 bp with an open reading frame of 1,077 bp (181-1257) encoding 358 amino acids. Its deduced amino acid sequence showed a high homology with monogalactosyldiacylglycerol synthase (UDPgalactose: 1,2-diacylglycerol 3-beta-D-galactosyl transferase; EC 2.4.1.46, MGDG synthase) from Arabidopsis thaliana; therefore, we named the gene OsMGD. Time-course studies showed that the expression of OsMGD in the rice cultivars FR13A and IR42 (submergence-susceptive cultivar) during submergence was gradually increased and that expression in FR13A was higher than in IR42. The expression of OsMGD in FR13A was influenced by benzyladenine and illumination. The accumulation of OsMGD mRNA in both FR13A and IR42 was also increased by ethephon, gibberellin, drought and salt treatment, but cold stress had no effect on the expression of the gene. These results suggest that the expression of OsMGD mRNA requires benzyladenine or illumination, and that the process is also mediated by ethephon and gibberellin. Salt and drought stress have an effect similar to that of submergence. Furthermore, the enhanced expression of OsMGD may relate to photosynthesis, and play an important role during submergence.

  6. Ectopic Expression in Arabidopsis thaliana of an NB-ARC Encoding Putative Disease Resistance Gene from Wild Chinese Vitis pseudoreticulata Enhances Resistance to Phytopathogenic Fungi and Bacteria

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    Zhifeng eWen

    2015-12-01

    Full Text Available Plant resistance proteins mediate pathogen recognition and activate innate immune responses to restrict pathogen proliferation. One common feature of these proteins is an NB-ARC domain. In this study, we characterized a gene encoding a protein with an NB-ARC domain from wild Chinese grapevine Vitis pseudoreticulata accession Baihe-35-1, which was identified in a transcriptome analysis of the leaves following inoculation with Erysiphe necator (Schw., a causal agent of powdery mildew. Transcript levels of this gene, designated VpCN (GenBank accession number KT265084, increased strongly after challenge of grapevine leaves with E. necator. The deduced amino acid sequence was predicted to contain an NB-ARC domain in the C-terminus and an RxCC-like domain similar to CC domain of Rx protein in the N-terminus. Ectopic expression of VpCN in Arabidopsis thaliana resulted in either a wild-type phenotype or a dwarf phenotype. The phenotypically normal transgenic A. thaliana showed enhance resistance to A. thaliana powdery mildew Golovinomyces cichoracearum, as well as to a virulent bacterial pathogen Pseudomonas syringae pv. tomato DC3000. Moreover, promoter::GUS (β-glucuronidase analysis revealed that powdery mildew infection induced the promoter activity of VpCN in grapevine leaves. Finally, a promoter deletion analysis showed that TC rich repeat elements likely play an important role in the response to E. necator infection. Taken together, our results suggest that VpCN contribute to powdery mildew disease resistant in grapevine.

  7. Characterization of Three New Glutaredoxin Genes in the Arbuscular Mycorrhizal Fungus Rhizophagus irregularis: Putative Role of RiGRX4 and RiGRX5 in Iron Homeostasis

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    Tamayo, Elisabeth; Benabdellah, Karim; Ferrol, Nuria

    2016-01-01

    Glutaredoxins (GRXs) are small ubiquitous oxidoreductases involved in the regulation of the redox state in living cells. In an attempt to identify the full complement of GRXs in the arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis, three additional GRX homologs, besides the formerly characterized GintGRX1 (renamed here as RiGRX1), were identified. The three new GRXs (RiGRX4, RiGRX5 and RiGRX6) contain the CXXS domain of monothiol GRXs, but whereas RiGRX4 and RiGRX5 belong to class II GRXs, RiGRX6 belongs to class I together with RiGRX1. By using a yeast expression system, we observed that the newly identified homologs partially reverted sensitivity of the GRX deletion yeast strains to external oxidants. Furthermore, our results indicated that RiGRX4 and RiGRX5 play a role in iron homeostasis in yeast. Gene expression analyses revealed that RiGRX1 and RiGRX6 were more highly expressed in the intraradical (IRM) than in the extraradical mycelium (ERM). Exposure of the ERM to hydrogen peroxide induced up-regulation of RiGRX1, RiGRX4 and RiGRX5 gene expression. RiGRX4 expression was also up-regulated in the ERM when the fungus was grown in media supplemented with a high iron concentration. These data indicate the two monothiol class II GRXs, RiGRX4 and RiGRX5, might be involved in oxidative stress protection and in the regulation of fungal iron homeostasis. Increased expression of RiGRX1 and RiGRX6 in the IRM suggests that these GRXs should play a key role in oxidative stress protection of R. irregularis during its in planta phase. PMID:26900849

  8. Characterization of Three New Glutaredoxin Genes in the Arbuscular Mycorrhizal Fungus Rhizophagus irregularis: Putative Role of RiGRX4 and RiGRX5 in Iron Homeostasis.

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    Elisabeth Tamayo

    Full Text Available Glutaredoxins (GRXs are small ubiquitous oxidoreductases involved in the regulation of the redox state in living cells. In an attempt to identify the full complement of GRXs in the arbuscular mycorrhizal (AM fungus Rhizophagus irregularis, three additional GRX homologs, besides the formerly characterized GintGRX1 (renamed here as RiGRX1, were identified. The three new GRXs (RiGRX4, RiGRX5 and RiGRX6 contain the CXXS domain of monothiol GRXs, but whereas RiGRX4 and RiGRX5 belong to class II GRXs, RiGRX6 belongs to class I together with RiGRX1. By using a yeast expression system, we observed that the newly identified homologs partially reverted sensitivity of the GRX deletion yeast strains to external oxidants. Furthermore, our results indicated that RiGRX4 and RiGRX5 play a role in iron homeostasis in yeast. Gene expression analyses revealed that RiGRX1 and RiGRX6 were more highly expressed in the intraradical (IRM than in the extraradical mycelium (ERM. Exposure of the ERM to hydrogen peroxide induced up-regulation of RiGRX1, RiGRX4 and RiGRX5 gene expression. RiGRX4 expression was also up-regulated in the ERM when the fungus was grown in media supplemented with a high iron concentration. These data indicate the two monothiol class II GRXs, RiGRX4 and RiGRX5, might be involved in oxidative stress protection and in the regulation of fungal iron homeostasis. Increased expression of RiGRX1 and RiGRX6 in the IRM suggests that these GRXs should play a key role in oxidative stress protection of R. irregularis during its in planta phase.

  9. A putatively functional polymorphism in the HTR2C gene is associated with depressive symptoms in white females reporting significant life stress.

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    Beverly H Brummett

    Full Text Available Psychosocial stress is well known to be positively associated with subsequent depressive symptoms. Cortisol response to stress may be one of a number of biological mechanisms that links psychological stress to depressive symptoms, although the precise causal pathway remains unclear. Activity of the x-linked serotonin 5-HTR2C receptor has also been shown to be associated with depression and with clinical response to antidepressant medications. We recently demonstrated that variation in a single nucleotide polymorphism on the HTR2C gene, rs6318 (Ser23Cys, is associated with different cortisol release and short-term changes in affect in response to a series of stress tasks in the laboratory. Based on this observation, we decided to examine whether rs6318 might moderate the association between psychosocial stress and subsequent depressive symptoms. In the present study we use cross-sectional data from a large population-based sample of young adult White men (N = 2,366 and White women (N = 2,712 in the United States to test this moderation hypothesis. Specifically, we hypothesized that the association between self-reported stressful life events and depressive symptoms would be stronger among homozygous Ser23 C females and hemizygous Ser23 C males than among Cys23 G carriers. In separate within-sex analyses a genotype-by-life stress interaction was observed for women (p = .022 but not for men (p = .471. Homozygous Ser23 C women who reported high levels of life stress had depressive symptom scores that were about 0.3 standard deviations higher than female Cys23 G carriers with similarly high stress levels. In contrast, no appreciable difference in depressive symptoms was observed between genotypes at lower levels of stress. Our findings support prior work that suggests a functional SNP on the HTR2C gene may confer an increased risk for depressive symptoms in White women with a history of significant life stress.

  10. De novo sequencing and analysis of the American ginseng root transcriptome using a GS FLX Titanium platform to discover putative genes involved in ginsenoside biosynthesis

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    Lui Edmund MK

    2010-04-01

    Full Text Available Abstract Background American ginseng (Panax quinquefolius L. is one of the most widely used herbal remedies in the world. Its major bioactive constituents are the triterpene saponins known as ginsenosides. However, little is known about ginsenoside biosynthesis in American ginseng, especially the late steps of the pathway. Results In this study, a one-quarter 454 sequencing run produced 209,747 high-quality reads with an average sequence length of 427 bases. De novo assembly generated 31,088 unique sequences containing 16,592 contigs and 14,496 singletons. About 93.1% of the high-quality reads were assembled into contigs with an average 8-fold coverage. A total of 21,684 (69.8% unique sequences were annotated by a BLAST similarity search against four public sequence databases, and 4,097 of the unique sequences were assigned to specific metabolic pathways by the Kyoto Encyclopedia of Genes and Genomes. Based on the bioinformatic analysis described above, we found all of the known enzymes involved in ginsenoside backbone synthesis, starting from acetyl-CoA via the isoprenoid pathway. Additionally, a total of 150 cytochrome P450 (CYP450 and 235 glycosyltransferase unique sequences were found in the 454 cDNA library, some of which encode enzymes responsible for the conversion of the ginsenoside backbone into the various ginsenosides. Finally, one CYP450 and four UDP-glycosyltransferases were selected as the candidates most likely to be involved in ginsenoside biosynthesis through a methyl jasmonate (MeJA inducibility experiment and tissue-specific expression pattern analysis based on a real-time PCR assay. Conclusions We demonstrated, with the assistance of the MeJA inducibility experiment and tissue-specific expression pattern analysis, that transcriptome analysis based on 454 pyrosequencing is a powerful tool for determining the genes encoding enzymes responsible for the biosynthesis of secondary metabolites in non-model plants. Additionally, the

  11. rtfA, a putative RNA-Pol II transcription elongation factor gene, is necessary for normal morphological and chemical development in Aspergillus flavus.

    Science.gov (United States)

    Lohmar, Jessica M; Harris-Coward, Pamela Y; Cary, Jeffrey W; Dhingra, Sourabh; Calvo, Ana M

    2016-06-01

    The filamentous fungus Aspergillus flavus is an agriculturally important opportunistic plant pathogen that produces potent carcinogenic compounds called aflatoxins. We identified the A. flavus rtfA gene, the ortholog of rtf1 in Saccharomyces cerevisiae and rtfA in Aspergillus nidulans. Interestingly, rtfA has multiple cellular roles in this mycotoxin-producing fungus. In this study, we show that rtfA regulates conidiation. The rtfA deletion mutant presented smaller conidiophores with significantly reduced conidial production compared to the wild-type strain. The absence of rtfA also resulted in a significant decrease or lack of sclerotial production under conditions that allowed abundant production of these resistance structures in the wild type. Importantly, the deletion of rtfA notably reduced the production of aflatoxin B1, indicating that rtfA is a regulator of mycotoxin biosynthesis in A. flavus. In addition, the deletion rtfA also altered the production of several unknown secondary metabolites indicating a broader regulatory scope. Furthermore, our study revealed that rtfA controls the expression of the global regulators veA and laeA, which further influence morphogenesis and secondary metabolism in A. flavus.

  12. Molecular cloning of the cDNA and chromosomal localization of the gene for a putative seven-transmembrane segment (7-TMS) receptor isolated from human spleen

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    Federsppiel, B.; Melhado, I.G.; Delaney, A.; Clark-Lewis, I. (Univ. of British Columbia, Vancouver (Canada)); Duncan, A.M.V. (Queens Univ., Kinston, Ontario (Canada)); Jirik, F.R. (Hospital for Sick Children, Toronto, Ontario (Canada))

    1993-06-01

    A family of proinflammatory cytokines sharing several structural features has been described and includes, for example, interleukin-8, monocyte chemoattractant protein-1, and melanocyte growth stimulatory activity. Recently, the receptors for interleukin-8 have been isolated and found to belong to the seven-transmembrane domain class of G protein-coupled receptors. As other members of this cytokine family likely interact with similar receptors, the polymerase chain reaction was employed to isolate related receptors from human peripheral blood adherent cells. Degenerate oligonucleotide primers based on the rabbit interleukin-8 receptor sequence were used. The corresponding full-length cDNA was isolated from a human spleen cDNA library. The predicted protein sequence of this clone, designated pBE1.3, was 93% identical to that of a cDNA isolated from bovine locus coeruleus, which apparently encodes a neuropeptide Y receptor, and also shows similarity with the interleukin-8 receptor and the human cytomegalovirus US28 sequences. The gene, designated D2S201E, was localized to human chromosome 2q21. By Northern blotting, transcripts hybridizing to this cDNA were present in a variety of tissues and cells, including those of hemopoietic origin. 32 refs., 5 figs.

  13. Molecular characterization of the Jatropha curcas JcR1MYB1 gene encoding a putative R1-MYB transcription factor

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    Hui-Liang Li

    2014-09-01

    Full Text Available The cDNA encoding the R1-MYB transcription factor, designated as JcR1MYB1, was isolated from Jatropha curcas using rapid amplification of cDNA ends. JcR1MYB1 contains a 951 bp open reading frame that encodes 316 amino acids. The deduced JcR1MYB1 protein was predicted to possess the conserved, 56-amino acid-long DNA-binding domain, which consists of a single helix-turn-helix module and usually occurs in R1-MYBs. JcR1MYB1 is a member of the R1-MYB transcription factor subfamily. A subcellular localization study confirmed the nuclear localization of JcR1MYB1. Expression analysis showed that JcR1MYB1 transcripts accumulated in various examined tissues, with high expression levels in the root and low levels in the stem. JcR1MYB1 transcription was up-regulated by polyethylene glycol, NaCl, and cold treatments, as well as by abscisic acid, jasmonic acid, and ethylene treatment. Analysis of transgenic tobacco plants over-expressing JcR1MYB1 indicates an inportant function for this gene in salt stress.

  14. Characterization of the putative amino acid transporter genes AtCAT2, 3 &4: the tonoplast localized AtCAT2 regulates soluble leaf amino acids.

    Science.gov (United States)

    Yang, Huaiyu; Krebs, Melanie; Stierhof, York-Dieter; Ludewig, Uwe

    2014-05-01

    The plant vacuole constitutes a large transient storage compartment for nutrients, proteins and metabolites, and is a major cellular sink for toxic waste compounds. Amino acids can cross the vacuolar membrane via specific transport proteins, which are molecularly not well characterized. Two members of a small subfamily of the cationic amino acid transporters, AtCAT2 and AtCAT4, were primarily localized at the tonoplast when tagged with GFP. The closely related AtCAT3, by contrast, was detected in the endoplasmic reticulum membrane. The exchange of a di-acidic motif at the carboxy-tail affected their sub-cellular localization, with larger effects visible in transiently transformed protoplasts compared to stably expressing plant lines. The genes have broad, partially overlapping tissue expression, with CAT2 dominating in most tissues. Loss-of-function mutants of individual CATs showed no visible phenotype under various conditions, but the overall tissue concentration of amino acids was increased in soil-grown cat2 mutants. The data suggest that CAT2 is a critical target of leaf amino acid concentrations and manipulation of this tonoplast transporter can significantly alter total tissue amino acid concentrations.

  15. Osteo-chondroprogenitor-specific deletion of the selenocysteine tRNA gene, Trsp, leads to chondronecrosis and abnormal skeletal development: a putative model for Kashin-Beck disease.

    Directory of Open Access Journals (Sweden)

    Charlene M Downey

    2009-08-01

    Full Text Available Kashin-Beck disease, a syndrome characterized by short stature, skeletal deformities, and arthropathy of multiple joints, is highly prevalent in specific regions of Asia. The disease has been postulated to result from a combination of different environmental factors, including contamination of barley by mold mycotoxins, iodine deficiency, presence of humic substances in drinking water, and, importantly, deficiency of selenium. This multifunctional trace element, in the form of selenocysteine, is essential for normal selenoprotein function, including attenuation of excessive oxidative stress, and for the control of redox-sensitive molecules involved in cell growth and differentiation. To investigate the effects of skeletal selenoprotein deficiency, a Cre recombinase transgenic mouse line was used to trigger Trsp gene deletions in osteo-chondroprogenitors. Trsp encodes selenocysteine tRNA([Ser]Sec, required for the incorporation of selenocysteine residues into selenoproteins. The mutant mice exhibited growth retardation, epiphyseal growth plate abnormalities, and delayed skeletal ossification, as well as marked chondronecrosis of articular, auricular, and tracheal cartilages. Phenotypically, the mice thus replicated a number of the pathological features of Kashin-Beck disease, supporting the notion that selenium deficiency is important to the development of this syndrome.

  16. A putative multi-replicon plasmid co-harboring beta-lactamase genes blaKPC-2, blaCTX-M-14 and blaTEM-1 and trimethoprim resistance gene dfrA25 from a Klebsiella pneumoniae sequence type (ST) 11 strain in China

    Science.gov (United States)

    Tang, Yu; Shen, Pinghua; Liang, Wei; Jin, Jialin; Jiang, Xiaofei

    2017-01-01

    The global emergence of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae poses a major public health threat requiring immediate and aggressive action. Some older generation antibiotics, such as trimethoprim, serve as alternatives for treatment of infections. Here, we determined the complete nucleotide sequence of plasmid pHS091147, which co-harbored the carbapenemase (blaKPC-2) and trimethoprim resistance genes (dfrA25) from a Klebsiella pneumoniae sequence type (ST) 11 clone recovered in Shanghai, China. pHS091147 had three replication genes, several plasmid-stability genes and an intact type IV secretion system gene cluster. Besides blaKPC-2 and dfrA25, pHS091147 carried several other resistance genes, including β-lactamase genes blaTEM-1 and blaCTX-M-14, sulphonamide resistance gene sul1, a quinolone resistance gene remnant (ΔqnrB2), and virulence associated gene iroN. Notably, the multidrug-resistance region was a chimeric structure composed of three subregions, which shared strong sequence homology with several plasmids previously assigned in Genbank. To our knowledge, this is the first report of the co-localization of blaKPC-2 and dfrA25 on a novel putative multi-replicon plasmid in a Klebsiella pneumoniae ST11 clone. PMID:28152085

  17. In vitro and in vivo silencing of plasmodial dhs and eIf-5a genes in a putative, non-canonical RNAi-related pathway

    Directory of Open Access Journals (Sweden)

    Schwentke Andreas

    2012-06-01

    days post infection before animals succumbed to hyperparasitemia similar to infections with the related but non-lethal phenotype P. berghei strain NK65. RT-PCR and Western blot experiments performed with blood from the transfected erythrocytic stages showed that both genes are important for the proliferation of the parasite. Moreover, these experiments clearly demonstrate that the hypusine pathway in Plasmodium is linked to human iNos induction.

  18. Putative archaeal viruses from the mesopelagic ocean.

    Science.gov (United States)

    Vik, Dean R; Roux, Simon; Brum, Jennifer R; Bolduc, Ben; Emerson, Joanne B; Padilla, Cory C; Stewart, Frank J; Sullivan, Matthew B

    2017-01-01

    Oceanic viruses that infect bacteria, or phages, are known to modulate host diversity, metabolisms, and biogeochemical cycling, while the viruses that infect marine Archaea remain understudied despite the critical ecosystem roles played by their hosts. Here we introduce "MArVD", for Metagenomic Archaeal Virus Detector, an annotation tool designed to identify putative archaeal virus contigs in metagenomic datasets. MArVD is made publicly available through the online iVirus analytical platform. Benchmarking analysis of MArVD showed it to be >99% accurate and 100% sensitive in identifying the 127 known archaeal viruses among the 12,499 viruses in the VirSorter curated dataset. Application of MArVD to 10 viral metagenomes from two depth profiles in the Eastern Tropical North Pacific (ETNP) oxygen minimum zone revealed 43 new putative archaeal virus genomes and large genome fragments ranging in size from 10 to 31 kb. Network-based classifications, which were consistent with marker gene phylogenies where available, suggested that these putative archaeal virus contigs represented six novel candidate genera. Ecological analyses, via fragment recruitment and ordination, revealed that the diversity and relative abundances of these putative archaeal viruses were correlated with oxygen concentration and temperature along two OMZ-spanning depth profiles, presumably due to structuring of the host Archaea community. Peak viral diversity and abundances were found in surface waters, where Thermoplasmata 16S rRNA genes are prevalent, suggesting these archaea as hosts in the surface habitats. Together these findings provide a baseline for identifying archaeal viruses in sequence datasets, and an initial picture of the ecology of such viruses in non-extreme environments.

  19. Construction and analysis of antennal cDNA library from rice striped stem borer, Chilo suppressalis (Walker) (Lepidoptera: Pyralidae), and expression profiles of putative odorant-binding protein and chemosensory protein genes.

    Science.gov (United States)

    Gong, Zhong-Jun; Liu, Su; Jiang, Yan-Dong; Zhou, Wen-Wu; Liang, Qing-Mei; Cheng, Jiaan; Zhang, Chuan-Xi; Zhu, Zeng-Rong; Gurr, Geoff M

    2015-05-01

    In this study, we constructed a high-quality cDNA library from the antennae of the Chilo suppressalis (Walker) (Lepidoptera: Pyralidae). A total of 1,235 colonies with inserts greater than 0.7 kb were sequenced and analyzed. Homology searching coupled with bioinformatics analysis identified 15 and 7 cDNA sequences, respectively, encoding putative odorant-binding proteins (OBPs) and chemosensory proteins (CSPs). A phylogenetic tree of CsupCSPs showed that each CsupCSP has orthologs in Manduca sexta and Bombyx mori with strong bootstrapping support. One CSP was either very specific or more related to the CSPs of another species than to conspecific CSP. The expression profiles of the OBPs and CSPs in different tissues were measured by real-time quantitative PCR. The results revealed that of the 11 OBP genes, the transcript levels of CsupOBP1, CsupOBP5, and CsupOBP7 were higher in both male and female antennae than those in other tissues. And CsupCSP7 was highly expressed in both male and female antennae. Based on these results, the possible physiological functions of CsupOBPs and CsupCSPs were discussed.

  20. The SUD1 Gene Encodes a Putative E3 Ubiquitin Ligase and Is a Positive Regulator of 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase Activity in Arabidopsis[C][W

    Science.gov (United States)

    Doblas, Verónica G.; Amorim-Silva, Vítor; Posé, David; Rosado, Abel; Esteban, Alicia; Arró, Montserrat; Azevedo, Herlander; Bombarely, Aureliano; Borsani, Omar; Valpuesta, Victoriano; Ferrer, Albert; Tavares, Rui M.; Botella, Miguel A.

    2013-01-01

    The 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) enzyme catalyzes the major rate-limiting step of the mevalonic acid (MVA) pathway from which sterols and other isoprenoids are synthesized. In contrast with our extensive knowledge of the regulation of HMGR in yeast and animals, little is known about this process in plants. To identify regulatory components of the MVA pathway in plants, we performed a genetic screen for second-site suppressor mutations of the Arabidopsis thaliana highly drought-sensitive drought hypersensitive2 (dry2) mutant that shows decreased squalene epoxidase activity. We show that mutations in SUPPRESSOR OF DRY2 DEFECTS1 (SUD1) gene recover most developmental defects in dry2 through changes in HMGR activity. SUD1 encodes a putative E3 ubiquitin ligase that shows sequence and structural similarity to yeast Degradation of α factor (Doα10) and human TEB4, components of the endoplasmic reticulum–associated degradation C (ERAD-C) pathway. While in yeast and animals, the alternative ERAD-L/ERAD-M pathway regulates HMGR activity by controlling protein stability, SUD1 regulates HMGR activity without apparent changes in protein content. These results highlight similarities, as well as important mechanistic differences, among the components involved in HMGR regulation in plants, yeast, and animals. PMID:23404890

  1. Comparison of ex vivo harvested and in vitro cultured materials from Echinococcus granulosus by measuring expression levels of five genes putatively involved in the development and maturation of adult worms.

    Science.gov (United States)

    Dezaki, Ebrahim Saedi; Yaghoubi, Mohammad Mehdi; Spiliotis, Markus; Boubaker, Ghalia; Taheri, Elham; Almani, Pooya Ghaseminejad; Tohidi, Farideh; Harandi, Majid Fasihi; Gottstein, Bruno

    2016-11-01

    Parts of the natural life cycle of Echinococcus granulosus can be retraced in vitro such as the development of protoscoleces into semiadult worms with three or more proglottids, or the redifferentiation of in vitro cultured protoscoleces into metacestode-like cystic structures. Most in vitro generated samples share-at the microscopical level-high similarities with those naturally grown, but developmental differences have also been documented, such as missing egg production in in vitro grown adults or unusual bladder/vesicle formation in protoscoleces cultured into the metacestode direction. The aim of the present study was to explore how far different in vitro generated stage-specific materials/structures match the natural situation on the transcriptome level, based on testing five exemplarily chosen different genes: the frizzled receptor eg-fz4 (posterior marker), the FGF receptor-like factor eg-fgfrl (anterior association), the cell differentiation protein eg-rcd1 (part of the CCR4-NOT complex, a key regulator of eukaryotic gene expression), the rapidly accelerated fibrosarcoma serin/threonin kinase eg-braf (part of the MAPK pathway involved, e.g., in EGF signaling) and the co-smad eg-smadD (downstream factor of TGFβ/BMP2/activin signaling). These genes-tested via qPCR-were selected such as to allow a discussion on their potential role in the development of E. granulosus into the adult stage. Thus, testing took place with three ex vivo isolated samples, namely (i) egg-containing adult worms, (ii) invaginated protoscoleces, and (iii) protoscolex-free germinal layer tissue. Respective data were compared (a) with in vitro generated metacestode-like microcysts developed from protoscolices, and (b) different development stages of protoscoleces in vitro cultured toward adult maturation. As a finding, only eg-smadD and partially eg-fz4 showed high expression similarities between ex vivo harvested and in vitro cultured E. granulosus, thus suggesting a putative role in

  2. Immunophilins, Refsum disease, and lupus nephritis: the peroxisomal enzyme phytanoyl-COA alpha-hydroxylase is a new FKBP-associated protein.

    Science.gov (United States)

    Chambraud, B; Radanyi, C; Camonis, J H; Rajkowski, K; Schumacher, M; Baulieu, E E

    1999-03-01

    FKBP52 (FKBP59, FKBP4) is a "macro" immunophilin that, although sharing high structural and functional homologies in its amino-terminal domain with FKBP12 (FKBP1), does not have immunosuppressant activity when complexed with FK506, unlike FKBP12. To investigate the physiological function of FKBP52, we used the yeast two-hybrid system as an approach to find its potential protein partners and, from that, its cellular role. This methodology, which already has allowed us to find the FK506-binding protein (FKBP)-associated protein FAP48, also led to the detection of another FKBP-associated protein. Determination of the sequence of this protein permitted its identification as phytanoyl-CoA alpha-hydroxylase (PAHX), a peroxisomal enzyme that so far was unknown as an FKBP-associated protein. Inactivation of this enzyme is responsible for Refsum disease in humans. The protein also corresponds to the mouse protein LN1, which could be involved in the progress of lupus nephritis. We show here that PAHX has the physical capacity to interact with the FKBP12-like domain of FKBP52, but not with FKBP12, suggesting that it is a particular and specific target of FKBP52. Whereas the binding of calcineurin to FKBP12 is potentiated by FK506, the specific association of PAHX and FKBP52 is maintained in the presence of FK506. This observation suggests that PAHX is a serious candidate for studying the cellular signaling pathway(s) involving FKBP52 in the presence of immunosuppressant drugs.

  3. Crystal structure of Arabidopsis cyclophilin38 reveals a previously uncharacterized immunophilin fold and a possible autoinhibitory mechanism.

    Science.gov (United States)

    Vasudevan, Dileep; Fu, Aigen; Luan, Sheng; Swaminathan, Kunchithapadam

    2012-06-01

    Cyclophilin38 (CYP38) is one of the highly divergent cyclophilins from Arabidopsis thaliana. Here, we report the crystal structure of the At-CYP38 protein (residues 83 to 437 of 437 amino acids) at 2.39-Å resolution. The structure reveals two distinct domains: an N-terminal helical bundle and a C-terminal cyclophilin β-barrel, connected by an acidic loop. Two N-terminal β-strands become part of the C-terminal cyclophilin β-barrel, thereby making a previously undiscovered domain organization. This study shows that CYP38 does not possess peptidyl-prolyl cis/trans isomerase activity and identifies a possible interaction of CYP38 with the E-loop of chlorophyll protein47 (CP47), a component of photosystem II. The interaction of CYP38 with the E-loop of CP47 is mediated through its cyclophilin domain. The N-terminal helical domain is closely packed together with the putative C-terminal cyclophilin domain and establishes a strong intramolecular interaction, thereby preventing the access of the cyclophilin domain to other proteins. This was further verified by protein-protein interaction assays using the yeast two-hybrid system. Furthermore, the non-Leucine zipper N-terminal helical bundle contains several new elements for protein-protein interaction that may be of functional significance. Together, this study provides the structure of a plant cyclophilin and explains a possible mechanism for autoinhibition of its function through an intramolecular interaction.

  4. Congenital insensitivity to pain with anhidrosis (CIPA): Novel mutations of the TRKA (NTRK1) gene, a putative uniparental disomy, and a linkage of the mutant TRKA and PKLR genes in a family with CIPA and pyruvate kinase deficiency

    NARCIS (Netherlands)

    Y. Indo (Yasuhiro); S. Mardy (Sek); Y. Miura (Yuichi); A. Moosa (Allie); E.A.R. Ismail (Essam A.); E. Toscano (Ennio); G. Andria (Generoso); V. Pavone (Vito); D.L. Brown (Deborah); A.S. Brooks (Alice); F. Endo (Fumio); I. Matsuda (Ichiro)

    2001-01-01

    textabstractCongenital insensitivity to pain with anhidrosis is an autosomal recessive hereditary disorder characterized by recurrent episodic fever, anhidrosis (inability to sweat), absence of reaction to noxious stimuli, self-mutilating behavior, and mental retardation. The human TRKA gene (NTRK1)

  5. Genome-Wide Analysis of Secondary Metabolite Gene Clusters in Ophiostoma ulmi and Ophiostoma novo-ulmi Reveals a Fujikurin-Like Gene Cluster with a Putative Role in Infection

    Directory of Open Access Journals (Sweden)

    Nicolau Sbaraini

    2017-06-01

    Full Text Available The emergence of new microbial pathogens can result in destructive outbreaks, since their hosts have limited resistance and pathogens may be excessively aggressive. Described as the major ecological incident of the twentieth century, Dutch elm disease, caused by ascomycete fungi from the Ophiostoma genus, has caused a significant decline in elm tree populations (Ulmus sp. in North America and Europe. Genome sequencing of the two main causative agents of Dutch elm disease (Ophiostoma ulmi and Ophiostoma novo-ulmi, along with closely related species with different lifestyles, allows for unique comparisons to be made to identify how pathogens and virulence determinants have emerged. Among several established virulence determinants, secondary metabolites (SMs have been suggested to play significant roles during phytopathogen infection. Interestingly, the secondary metabolism of Dutch elm pathogens remains almost unexplored, and little is known about how SM biosynthetic genes are organized in these species. To better understand the metabolic potential of O. ulmi and O. novo-ulmi, we performed a deep survey and description of SM biosynthetic gene clusters (BGCs in these species and assessed their conservation among eight species from the Ophiostomataceae family. Among 19 identified BGCs, a fujikurin-like gene cluster (OpPKS8 was unique to Dutch elm pathogens. Phylogenetic analysis revealed that orthologs for this gene cluster are widespread among phytopathogens and plant-associated fungi, suggesting that OpPKS8 may have been horizontally acquired by the Ophiostoma genus. Moreover, the detailed identification of several BGCs paves the way for future in-depth research and supports the potential impact of secondary metabolism on Ophiostoma genus’ lifestyle.

  6. Sequence analysis of a 13.4 kbp fragment from the left arm of chromosome XV reveals a malate dehydrogenase gene, a putative Ser/Thr protein kinase, the ribosomal L25 gene and four new open reading frames.

    Science.gov (United States)

    Casamayor, A; Khalid, H; Balcells, L; Aldea, M; Casas, C; Herrero, E; Ariño, J

    1996-09-01

    A 13421 bp fragment located near the left telomere of chromosome XV (cosmid pEOA461) has been sequenced. Seven non-overlapping open reading frames (ORFs) encoding polypeptides longer than 100 residues have been found (AOB859, AOC184, AOE375, AOX142i, AOE423, AOA476 and AOE433). An additional ORF (AOE131) is found within AOA476. Three of them (AOC184, AOA476 and AOE433) show no remarkable identity with proteins deposited in the data banks. ORF AOB859 is quite similar to a hypothetical yeast protein of similar size located in chromosome VI, particularly within the C-terminal half. AOE375 encodes a new member of the glycogen synthase kinase-3 subfamily of Ser/Thr protein kinases. AOX142i is the gene encoding the previously described ribosomal protein L25. AOE423 codes for a protein virtually identical to the MDH2 malate dehydrogenase isozyme. However, our DNA sequence shows a single one-base insertion upstream of the reported initiating codon. This would produce a larger ORF by extending 46 residues the N-terminus of the protein. The existence of this insertion has been confirmed in three different yeast strains, including FY1679.

  7. Trypanosoma brucei: a putative RNA polymerase II promoter.

    Science.gov (United States)

    Bayele, Henry K

    2009-12-01

    RNA polymerase II (pol II) promoters are rare in the African trypanosome Trypanosoma brucei because gene regulation in the parasite is complex and polycistronic. Here, we describe a putative pol II promoter and its structure-function relationship. The promoter has features of an archetypal eukaryotic pol II promoter including putative canonical CCAAT and TATA boxes, and an initiator element. However, the spatial arrangement of these elements is only similar to yeast pol II promoters. Deletion mapping and transcription assays enabled delineation of a minimal promoter that could drive orientation-independent reporter gene expression suggesting that it may be a bidirectional promoter. In vitro transcription in a heterologous nuclear extract revealed that the promoter can be recognized by the basal eukaryotic transcription complex. This suggests that the transcription machinery in the parasite may be very similar to those of other eukaryotes.

  8. Molecular cloning of a putative crustacean hyperglycemic hormone (CHH) isoform from extra-eyestalk tissue of the blue crab (Callinectes sapidus), and determination of temporal and spatial patterns of CHH gene expression.

    Science.gov (United States)

    Zheng, Junying; Chen, Hsiang-Yin; Choi, Cheol Young; Roer, Robert D; Watson, R Douglas

    2010-11-01

    Crustacean hyperglycemic hormone (CHH) is a polypeptide neurohormone involved in regulation of multiple physiological processes. We report here the cloning from thoracic ganglia of the blue crab (Callinectes sapidus) a cDNA (CsCHH-2) encoding a putative CHH isoform (CsCHH-2). CsCHH-2 is structurally similar to a putative preproCHH (CsCHH-1) previously cloned from eyestalk ganglia of C. sapidus. The two preprohormones possess an identical signal peptide and CHH precursor related peptide, but differ in the mature CHH polypeptide. An analysis by RT-PCR of the tissue distribution of CsCHH-1 and CsCHH-2 revealed the former is restricted to eyestalk neural ganglia, while the latter is widely distributed among tissues. The type of CHH transcript present in eyestalk and thoracic ganglia did not vary as a function of the molt cycle. An assessment of transcript abundance in tissues of intermolt crabs showed the abundance of the CsCHH-1 transcript in eyestalk ganglia far exceeds the abundance of the CsCHH-2 transcript in extra-eyestalk tissue. An assessment of transcript abundance during a molt cycle showed CsCHH-1 transcript abundance in eyestalk ganglia was low during intermolt, rose during premolt, reaching a peak in D(3), then fell prior to molting, and remained low during postmolt. By contrast, CsCHH-2 transcript abundance in thoracic ganglia was low during intermolt, rose sharply during D(2), then dropped in D(3) and remained low during postmolt. The results are consistent with the hypothesis that CsCHH-1 and CsCHH-2 differ with respect to physiological function. Copyright © 2010 Elsevier Inc. All rights reserved.

  9. The Biogeography of Putative Microbial Antibiotic Production.

    Directory of Open Access Journals (Sweden)

    Hélène Morlon

    Full Text Available Understanding patterns in the distribution and abundance of functional traits across a landscape is of fundamental importance to ecology. Mapping these distributions is particularly challenging for species-rich groups with sparse trait measurement coverage, such as flowering plants, insects, and microorganisms. Here, we use likelihood-based character reconstruction to infer and analyze the spatial distribution of unmeasured traits. We apply this framework to a microbial dataset comprised of 11,732 ketosynthase alpha gene sequences extracted from 144 soil samples from three continents to document the spatial distribution of putative microbial polyketide antibiotic production. Antibiotic production is a key competitive strategy for soil microbial survival and performance. Additionally, novel antibiotic discovery is highly relevant to human health, making natural antibiotic production by soil microorganisms a major target for bioprospecting. Our comparison of trait-based biogeographical patterns to patterns based on taxonomy and phylogeny is relevant to our basic understanding of microbial biogeography as well as the pressing need for new antibiotics.

  10. 转录调控蛋白PrfA对两组新近发现的单核细胞增生李斯特菌基因的体外转录作用的研究%In vitro Transcription of Two Groups of Newly Identified and Putatively in vivo PrfA-Regulated Genes of Listeria monocytogenes

    Institute of Scientific and Technical Information of China (English)

    罗勤; 周青春; 邓灵福; 高强; 刘德立; GOEBEL Werner

    2006-01-01

    目的 研究转录调控蛋白PffA对两组新近发现的单核细胞增生李斯特菌基因的体外转录作用.方法 利用本室近年来建立的体外转录系统,对两组基于转录基因组体内研究发现的5个可能的受PffA不同调节的单核细胞增生李斯特菌基因进行了体外转录活性的研究.结果 第一组中的hpt基因的体外转录活性受PffA正调节,而其它4个基因既不被PffA正调节也不被负调节.结论 除hpt基因外,其它4个基因体外转录结果与体内实验不相一致,说明PrfA在体内可能通过复杂多样的非直接方式、或者还需要一些目前未知的因子来调控这些新近发现的基因的表达.%Objective The aim of study was to investigate PrfA-dependent transcription activities of two groups of newly identified and putatively in vivo PrfA-regulated genes of listeria monocytogenes. Methods Transcription initiation at promoters of five genes selected from the two groups based on the transcriptome analysis was studied by means of in vitro transcription system. Results The data showed that, among these new putative" PrfA-regulated" promoters, only the promoter of hpt gene belonging to group I was significantly activated by PrfA. In addition, this promoter was the only one exhibiting all essential features of a typical PrfA-dependent promoter. Neither positively nor negatively, was in vitro transcription starting at most of other promoters affected by PrfA. Conclusion These in vitro transcription results indicate that the effect of PrfA in vivo on the expression of these new genes might be committed in an indirect manner or via a complex process, and that the PrfA-mediated transcription of these genes, in contrast to the PrfA-dependent transcription of the known virulence genes ( including hpt), probably requires additional factors lacked in the in vitro transcription system.

  11. Putative regulatory factors associated with intramuscular fat content.

    Directory of Open Access Journals (Sweden)

    Aline S M Cesar

    Full Text Available Intramuscular fat (IMF content is related to insulin resistance, which is an important prediction factor for disorders, such as cardiovascular disease, obesity and type 2 diabetes in human. At the same time, it is an economically important trait, which influences the sensorial and nutritional value of meat. The deposition of IMF is influenced by many factors such as sex, age, nutrition, and genetics. In this study Nellore steers (Bos taurus indicus subspecies were used to better understand the molecular mechanisms involved in IMF content. This was accomplished by identifying differentially expressed genes (DEG, biological pathways and putative regulatory factors. Animals included in this study had extreme genomic estimated breeding value (GEBV for IMF. RNA-seq analysis, gene set enrichment analysis (GSEA and co-expression network methods, such as partial correlation coefficient with information theory (PCIT, regulatory impact factor (RIF and phenotypic impact factor (PIF were utilized to better understand intramuscular adipogenesis. A total of 16,101 genes were analyzed in both groups (high (H and low (L GEBV and 77 DEG (FDR 10% were identified between the two groups. Pathway Studio software identified 13 significantly over-represented pathways, functional classes and small molecule signaling pathways within the DEG list. PCIT analyses identified genes with a difference in the number of gene-gene correlations between H and L group and detected putative regulatory factors involved in IMF content. Candidate genes identified by PCIT include: ANKRD26, HOXC5 and PPAPDC2. RIF and PIF analyses identified several candidate genes: GLI2 and IGF2 (RIF1, MPC1 and UBL5 (RIF2 and a host of small RNAs, including miR-1281 (PIF. These findings contribute to a better understanding of the molecular mechanisms that underlie fat content and energy balance in muscle and provide important information for the production of healthier beef for human consumption.

  12. Orally administered lactoperoxidase increases expression of the FK506 binding protein 5 gene in epithelial cells of the small intestine of mice: a DNA microarray study.

    Science.gov (United States)

    Wakabayashi, Hiroyuki; Miyauchi, Hirofumi; Shin, Kouichirou; Yamauchi, Koji; Matsumoto, Ichiro; Abe, Keiko; Takase, Mitsunori

    2007-09-01

    Lactoperoxidase (LPO) is a component of milk and other external secretions. To study the influence of ingested LPO on the digestive tract, we performed DNA microarray analysis of the small intestine of mice administered LPO. LPO administration upregulated 78 genes, including genes involved in metabolism, immunity, apoptosis, and the cell cycle, and downregulated nine genes, including immunity-related genes. The most upregulated gene was FK506 binding protein 5 (FKBP5), a glucocorticoid regulating immunophilin. The upregulation of this gene was confirmed by quantitative RT-PCR in other samples. In situ hybridization revealed that expression of the FKBP5 gene in the crypt epithelial cells of the small intestine was enhanced by LPO. These results suggest that ingested LPO modulates gene expression in the small intestine and especially increases FKBP5 gene expression in the epithelial cells of the intestine.

  13. The sequence of an 11.1 kb fragment on the left arm of Saccharomyces cerevisiae chromosome VII reveals six open reading frames including NSP49, KEM1 and four putative new genes.

    Science.gov (United States)

    Bertani, I; Coglievina, M; Zaccaria, P; Klima, R; Bruschi, C V

    1995-09-30

    We report the sequence of an 11.1 kb fragment located on the left arm of chromosome VII of Saccharomyces cerevisiae. By sequence analysis we have detected six open reading frames (ORFs) longer that 300 bp, which cover 87% of the entire sequence. ORF G1645 is 100% identical to the KEM1 gene, also identified as DST2, XRN1, SEP1 and RAR5, while G1648 is 100% identical to the NSP49 or NUP49 gene. ORF G1642 shares some identity with a hypothetical protein of Caenorhabditis elegans, while the other four ORFs show no significant homology to known proteins.

  14. Fluoroquinolone-Resistant Haemophilus parasuis Isolates Exhibit More Putative Virulence Factors than Their Susceptible Counterparts

    OpenAIRE

    Zhang, Qiang; Liu, Jiantao; Yan, Shuxian; Yang, Yujie; Zhang, Anding; Jin, Meilin

    2013-01-01

    The prevalence of 23 putative virulence factors among fluoroquinolone-susceptible and -resistant Haemophilus parasuis isolates was analyzed. Putative hemolysin precursor, fimbrial assembly chaperone, and type I site-specific restriction modification system R subunit genes were more prevalent among fluoroquinolone-resistant H. parasuis isolates than among fluoroquinolone-susceptible H. parasuis isolates. Fluoroquinolone resistance may be associated with an increase in the presence of some viru...

  15. A putative helicase, the SUA5, PMR1, tRNALys1 genes and four open reading frames have been detected in the DNA sequence of an 8.8 kb fragment of the left arm of chromosome VII of Saccharomyces cerevisiae.

    Science.gov (United States)

    Klima, R; Coglievina, M; Zaccaria, P; Bertani, I; Bruschi, C V

    1996-09-01

    We report the sequence of an 8.8 kb segment of DNA from the left arm of chromosome VII of Saccharomyces cerevisiae. The sequence reveals seven open reading frames (ORFs) G1651, G1654, G1660, G1663, G1666, G1667 and G1669 greater than 100 amino acids in length and the tRNALys1 gene. ORF G1651 shows 100% identity with the ROK1 protein which is a putative RNA helicase of the 'DEAD box' protein family. ORF G1654 exhibits a motif highly conserved in ATP/GTP binding proteins generally referred to as 'P-loop'. From FastA analysis, G1660 and G1666 were found to be previously sequenced genes, respectively SUA5 and PMR1. The three other ORFs identified are partially (G1663) or completely (G1667 and G1669) overlapping with the PMR1 sequence on the complementary strand. This feature, together with their low codon adaptation indexes and the absence of significant homology with known proteins suggest that they do not correspond to real genes.

  16. CLONING, SEQUENCE ANALYSIS, AND CHARACTERIZATION OF PUTATIVE BETA-LACTAMASE OF STENOTROPHOMONAS MALTOPHILIA

    Directory of Open Access Journals (Sweden)

    Chong Seng Shueh

    2012-10-01

    Full Text Available The main objective of current study was to explore the function of chromosomal putative beta-lactamase gene (smlt 0115 in clinical Stenotrophomonas maltophilia. Antibiotic susceptibility test (AST screening for current antimicrobial drugs was done and Minimum Inhibitory Concentration (MIC level towards beta-lactams was determined by E-test. Putative beta-lactamase gene of S. maltophilia was amplified via PCR, with specific primers, then cloned into pET-15 expression plasmid and transformed into Escherichia coli BL21. The gene was sequenced and analyzed. The expressed protein was purified by affinity chromatography and the kinetic assay was performed. S. maltophilia ATCC 13637 was included in this experiment. Besides, a hospital strain which exhibited resistant to a series of beta-lactams including cefepime was identified via AST and MIC, hence it was named as S2 strain and was considered in this study. Sequencing result showed that putative beta-lactamase gene obtained from ATCC 13637 and S2 strains were predicted to have cephalosporinase activity by National Center for Biotechnology Information (NCBI blast program. Differences in the sequences of both ATCC 13637 and S2 strains were found via ClustalW alignment software. Kinetic assay proved a cephalosporinase characteristic produced by E. coli BL21 clone that overexpressed the putative beta-lactamase gene cloned under the control of an external promoter. Yet, expressed protein purified from S2 strain had high catalytic activity against beta-lactam antibiotics which was 14-fold higher than expressed protein purified from ATCC 13637 strain. This study represents the characterization analysis of putative beta-lactamase gene (smlt 0115 of S. maltophilia. The presence of the respective gene in the chromosome of S. maltophilia suggested that putative beta-lactamase gene (smlt 0115 of S. maltophilia plays a role in beta-lactamase resistance.

  17. 牦牛 HOXA1基因的时空表达及其对机体调控机制的研究%Expression Pattern of HOXA1 Gene and Its Putative Regulating Mechanism of Yak

    Institute of Scientific and Technical Information of China (English)

    张亚南; 石仙; 熊显荣; 兰道亮; 李键

    2016-01-01

    The aim of study was to obtain the yak HOXA1 gene sequence and related bioinformat-ics information,and analyze its expression spectrum and temporal expression profiles for providing related information of the formation of multiple vertebrae and its mechanism.RT-PCR technology was applied to clone the yak HOXA1 gene,semi-quantitative RT-PCR technique was used to de-tect the gene expression level in multiple organisms,and Real-time quantitative PCR technique was used to detect the gene expression level in the organisms in different development periods. The results showed that the obtained HOXA1 gene was 885 bp including the ORF of 870 bp,en-coding 290 amino acids.The yak HOXA1 gene shared a homology of 75.4% to 98.1% with ordi-nary cow,wild yak,human,wild boar,horse,mice,chimpanzees,jungle fowl and wild boar.There were different expression level of HOXA1 gene in heart,liver,spleen,lung,kidney,large intes-tine,small intestine,muscle,stomach,ovaries,uterus,fallopian tubes,breast and testicular tissue. With the growth of the yak,HOXA1 gene expression level in tissue also increased.%试验旨在获得牦牛 HOXA1基因序列并进行生物信息学分析,同时分析其组织表达谱和时序表达谱,为进一步研究多脊椎骨形成的原因及其机制奠定基础。通过 RT-PCR 技术克隆多脊椎骨牦牛 HOXA1基因,利用半定量 RT-PCR 技术检测该基因在多脊椎骨牦牛组织中的表达情况,利用实时荧光定量 PCR 技术检测该基因在多脊椎骨牦牛不同发育时期各组织中的表达情况。RT-PCR 结果表明,多脊椎骨牦牛 HOXA1基因序列全长为885 bp,其中开放阅读框(ORF)为870 bp,编码290个氨基酸。同源性分析表明,牦牛与普通牛、野牦牛、人、野猪、马、家鼠、黑猩猩、原鸡和野猪的同源性为75.4%~98.1%。组织表达分析表明,HOXA1基因在心脏、肝脏、脾脏、肺脏、肾脏、大肠、小肠、肌肉、胃、卵巢、子宫、输卵管、乳腺、睾丸中均有

  18. Major Quantitative Trait Loci and Putative Candidate Genes for Powdery Mildew Resistance and Fruit-Related Traits Revealed by an Intraspecific Genetic Map for Watermelon (Citrullus lanatus var. lanatus).

    Science.gov (United States)

    Kim, Kwang-Hwan; Hwang, Ji-Hyun; Han, Dong-Yeup; Park, Minkyu; Kim, Seungill; Choi, Doil; Kim, Yongjae; Lee, Gung Pyo; Kim, Sun-Tae; Park, Young-Hoon

    2015-01-01

    An intraspecific genetic map for watermelon was constructed using an F2 population derived from 'Arka Manik' × 'TS34' and transcript sequence variants and quantitative trait loci (QTL) for resistance to powdery mildew (PMR), seed size (SS), and fruit shape (FS) were analyzed. The map consists of 14 linkage groups (LGs) defined by 174 cleaved amplified polymorphic sequences (CAPS), 2 derived-cleaved amplified polymorphic sequence markers, 20 sequence-characterized amplified regions, and 8 expressed sequence tag-simple sequence repeat markers spanning 1,404.3 cM, with a mean marker interval of 6.9 cM and an average of 14.6 markers per LG. Genetic inheritance and QTL analyses indicated that each of the PMR, SS, and FS traits is controlled by an incompletely dominant effect of major QTLs designated as pmr2.1, ss2.1, and fsi3.1, respectively. The pmr2.1, detected on chromosome 2 (Chr02), explained 80.0% of the phenotypic variation (LOD = 30.76). This QTL was flanked by two CAPS markers, wsb2-24 (4.00 cM) and wsb2-39 (13.97 cM). The ss2.1, located close to pmr2.1 and CAPS marker wsb2-13 (1.00 cM) on Chr02, explained 92.3% of the phenotypic variation (LOD = 68.78). The fsi3.1, detected on Chr03, explained 79.7% of the phenotypic variation (LOD = 31.37) and was flanked by two CAPS, wsb3-24 (1.91 cM) and wsb3-9 (7.00 cM). Candidate gene-based CAPS markers were developed from the disease resistance and fruit shape gene homologs located on Chr.02 and Chr03 and were mapped on the intraspecific map. Colocalization of these markers with the major QTLs indicated that watermelon orthologs of a nucleotide-binding site-leucine-rich repeat class gene containing an RPW8 domain and a member of SUN containing the IQ67 domain are candidate genes for pmr2.1 and fsi3.1, respectively. The results presented herein provide useful information for marker-assisted breeding and gene cloning for PMR and fruit-related traits.

  19. The structure at 2.4 Å resolution of the protein from gene locus At3g21360, a putative Fe{sup II}/2-oxoglutarate-dependent enzyme from Arabidopsis thaliana

    Energy Technology Data Exchange (ETDEWEB)

    Bitto, Eduard; Bingman, Craig A.; Allard, Simon T. M.; Wesenberg, Gary E.; Aceti, David J.; Wrobel, Russell L.; Frederick, Ronnie O.; Sreenath, Hassan; Vojtik, Frank C.; Jeon, Won Bae; Newman, Craig S.; Primm, John; Sussman, Michael R.; Fox, Brian G.; Markley, John L.; Phillips, George N. Jr, E-mail: phillips@biochem.wisc.edu [Center for Eukaryotic Structural Genomics, Department of Biochemistry, University of Wisconsin-Madison (United States)

    2005-05-01

    The crystal structure of the 37.2 kDa At3g21360 gene product from A. thaliana was determined at 2.4 Å resolution. The structure establishes that this protein binds a metal ion and is a member of a clavaminate synthase-like superfamily in A. thaliana. The crystal structure of the gene product of At3g21360 from Arabidopsis thaliana was determined by the single-wavelength anomalous dispersion method and refined to an R factor of 19.3% (R{sub free} = 24.1%) at 2.4 Å resolution. The crystal structure includes two monomers in the asymmetric unit that differ in the conformation of a flexible domain that spans residues 178–230. The crystal structure confirmed that At3g21360 encodes a protein belonging to the clavaminate synthase-like superfamily of iron(II) and 2-oxoglutarate-dependent enzymes. The metal-binding site was defined and is similar to the iron(II) binding sites found in other members of the superfamily.

  20. Molecular diagnosis of putative Stargardt disease by capture next generation sequencing.

    Science.gov (United States)

    Zhang, Xiao; Ge, Xianglian; Shi, Wei; Huang, Ping; Min, Qingjie; Li, Minghan; Yu, Xinping; Wu, Yaming; Zhao, Guangyu; Tong, Yi; Jin, Zi-Bing; Qu, Jia; Gu, Feng

    2014-01-01

    Stargardt Disease (STGD) is the commonest genetic form of juvenile or early adult onset macular degeneration, which is a genetically heterogeneous disease. Molecular diagnosis of STGD remains a challenge in a significant proportion of cases. To address this, seven patients from five putative STGD families were recruited. We performed capture next generation sequencing (CNGS) of the probands and searched for potentially disease-causing genetic variants in previously identified retinal or macular dystrophy genes. Seven disease-causing mutations in ABCA4 and two in PROM1 were identified by CNGS, which provides a confident genetic diagnosis in these five families. We also provided a genetic basis to explain the differences among putative STGD due to various mutations in different genes. Meanwhile, we show for the first time that compound heterozygous mutations in PROM1 gene could cause cone-rod dystrophy. Our findings support the enormous potential of CNGS in putative STGD molecular diagnosis.

  1. The structure at 2.4 Å resolution of the protein from gene locus At3g21360, a putative FeII/2-oxo­glutarate-dependent enzyme from Arabidopsis thaliana

    Science.gov (United States)

    Bitto, Eduard; Bingman, Craig A.; Allard, Simon T. M.; Wesenberg, Gary E.; Aceti, David J.; Wrobel, Russell L.; Frederick, Ronnie O.; Sreenath, Hassan; Vojtik, Frank C.; Jeon, Won Bae; Newman, Craig S.; Primm, John; Sussman, Michael R.; Fox, Brian G.; Markley, John L.; Phillips, George N.

    2005-01-01

    The crystal structure of the gene product of At3g21360 from Arabidopsis thaliana was determined by the single-wavelength anomalous dispersion method and refined to an R factor of 19.3% (R free = 24.1%) at 2.4 Å resolution. The crystal structure includes two monomers in the asymmetric unit that differ in the conformation of a flexible domain that spans residues 178–230. The crystal structure confirmed that At3g21360 encodes a protein belonging to the clavaminate synthase-like superfamily of iron(II) and 2-oxoglutarate-dependent enzymes. The metal-binding site was defined and is similar to the iron(II) binding sites found in other members of the superfamily. PMID:16511070

  2. Characterization of putative effectors from the cereal cyst nematode Heterodera avenae.

    Science.gov (United States)

    Cui, Jiangkuan; Peng, Huan; Qiao, Fen; Wang, Gaofeng; Huang, Wenkun; Wu, Duqign; Peng, Deliang

    2017-09-20

    Few molecular details of effectors of Heterodera avenae parasitism are known. We performed a high-throughput sequencing analysis of the H. avenae transcriptome at five developmental stages. A total of 82,549 unigenes were ultimately obtained, and 747 transcripts showed best hits to genes putatively encoding carbohydrate-active enzymes in plant parasitic nematodes that play an important role in the invasion process. A total of 1480 unigenes were homologous to known phytonematode effectors, and 63 putative novel effectors were identified in the H. avenae transcriptomes. Twenty-three unigenes were analyzed by qRT-PCR and confirmed to be highly expressed during at least one developmental stage. For in situ hybridization, 17 of the 22 tested putative effectors were specifically expressed and located in the subventral gland cells, and five putative novel effectors were specifically expressed in the dorsal gland. Furthermore, 115 transcripts were found to have putative lethal RNA interference (RNAi) phenotypes. Three target genes with lethal RNAi phenotypes and two of the four tested putative effectors were associated with a decrease in the number of cysts through in vitro RNAi technology. These transcriptomic data lay a foundation for further studies of interactions of H. avenae with cereal and H. avenae parasitic control.

  3. Sint1, a common integration site in SL3-3-induced T-cell lymphomas, harbors a putative proto-oncogene with homology to the septin gene family

    DEFF Research Database (Denmark)

    Sørensen, A B; Lund, Anders Henrik; Ethelberg, S;

    2000-01-01

    The murine retrovirus SL3-3 is a potent inducer of T-cell lymphomas when inoculated into susceptible newborn mice. Previously, DNAs from twenty SL3-3-induced tumors were screened by PCR for provirus integration sites. Two out of 20 tumors demonstrated clonal provirus insertion into a common region....... This region has now been isolated and characterized. The region, named SL3-3 integration site 1 (Sint1), maps to the distal end of mouse chromosome 11, corresponding to human chromosome 17q25, and may be identical to a mouse mammary tumor virus integration site in a T-cell lymphoma, Pad3. Two overlapping...... genomic lambda clones spanning about 35 kb were isolated and used as a starting point for a search for genes in the neighborhood of the virus integration sites. A genomic fragment was used as a hybridization probe to isolate a 3-kb cDNA clone, the expression of which was upregulated in one of two tumors...

  4. Amino acid transport in taxonomically diverse cyanobacteria and identification of two genes encoding elements of a neutral amino acid permease putatively involved in recapture of leaked hydrophobic amino acids.

    Science.gov (United States)

    Montesinos, M L; Herrero, A; Flores, E

    1997-02-01

    The activities of uptake of thirteen 14C-labeled amino acids were determined in nine cyanobacteria, including the unicellular strains Synechococcus sp. strain PCC 7942 and Synechocystis sp. strain PCC 6803; the filamentous strain Pseudanabaena sp. strain PCC 6903, and the filamentous, heterocyst-forming strains Anabaena sp. strains PCC 7120 and PCC 7937; Nostoc sp. strains PCC 7413 and PCC 7107; Calothrix sp. strain PCC 7601 (which is a mutant unable to develop heterocysts); and Fischerella muscicola UTEX 1829. Amino acid transport mutants, selected as mutants resistant to some amino acid analogs, were isolated from the Anabaena, Nostoc, Calothrix, and Pseudanabaena strains. All of the tested cyanobacteria bear at least a neutral amino acid transport system, and some strains also bear transport systems specific for basic or acidic amino acids. Two genes, natA and natB, encoding elements (conserved component, NatA, and periplasmic binding protein, NatB) of an ABC-type permease for neutral amino acids were identified by insertional mutagenesis of strain PCC 6803 open reading frames from the recently published genomic DNA sequence of this cyanobacterium. DNA sequences homologous to natA and natB from strain PCC 6803 were detected by hybridization in eight cyanobacterial strains tested. Mutants unable to transport neutral amino acids, including natA and natB insertional mutants, accumulated in the extracellular medium a set of amino acids that always included Ala, Val, Phe, Ile, and Leu. A general role for a cyanobacterial neutral amino acid permease in recapture of hydrophobic amino acids leaked from the cells is suggested.

  5. Identification of putative candidate gene markers for grain zinc ...

    African Journals Online (AJOL)

    Gande

    2014-01-29

    Jan 29, 2014 ... with 96 rice genotypes showed three markers (OsZIP8a, OsNAC and OsZIP4b) with phenotypic variation of 11.0, 5.8 .... tent in wheat mapping population, ranging from 19.9 to .... Genotype×environment interaction for iron con-.

  6. Transcript accumulation of putative drought responsive genes in ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-09-15

    Sep 15, 2009 ... technique is a suitable, low-cost technique to identify differentially .... sequence Ct value and ∆∆Ct involves subtraction of ∆Ct of the irrigated from ∆Ct of the ..... In addition, stress conditions reduce rates of photosyn- thetic CO2 ...

  7. Differential expressions of putative genes in various floral organs of ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-06-03

    Jun 3, 2009 ... 1Department of Biology, Faculty Science, University Putra Malaysia. ... investigation, further molecular expression investigations are required and essential to develop .... Semi-quantitative RT-PCR and Southern hybridization.

  8. Expression of putative virulence factors in the potato pathogen Clavibacter michiganensis subsp. sepedonicus during infection.

    Science.gov (United States)

    Holtsmark, Ingrid; Takle, Gunnhild W; Brurberg, May Bente

    2008-02-01

    The Gram-positive bacterium Clavibacter michiganensis subsp. sepedonicus is the causal agent of bacterial wilt and ring rot of potato. So far, only two proteins have been shown to be essential for virulence, namely a plasmid-encoded cellulase CelA and a hypersensitive response-inducing protein. We have examined the relative expression of CelA and eight putative virulence factors during infection of potato and in liquid culture, using quantitative real-time PCR. The examined putative virulence genes were celB, a cellulase-encoding gene and genes encoding a pectate lyase, a xylanase and five homologues of the Clavibacter michiganensis subsp. michiganensis pathogenicity factor Pat-1 thought to encode a serine protease. Six of the nine assayed genes were up-regulated during infection of potato, including celA, celB, the xylanase gene, and two of the pat genes. The pectate lyase gene showed only slightly elevated expression, whereas three of the five examined pat genes were down-regulated during infection in potato. Interestingly, the two up-regulated pat genes showed a noticeable sequence difference compared to the three down-regulated pat genes. These results reveal several new proteins that are likely to be involved in Clavibacter michiganensis subsp. sepedonicus pathogenicity.

  9. A new putative sigma factor of Myxococcus xanthus.

    Science.gov (United States)

    Apelian, D; Inouye, S

    1993-06-01

    A third putative sigma factor gene, sigC, has been isolated from Myxococcus xanthus by using the sigA gene (formerly rpoD of M. xanthus) as a probe. The nucleotide sequence of sigC has been determined, and an open reading frame of 295 residues (M(r) = 33,430) has been identified. The deduced amino acid sequence of sigC exhibits the features which are characteristic of other bacterial sigma factors. The characterization of a sigC-lacZ strain has demonstrated that sigC expression is induced immediately after cells enter into the developmental cycle and is dramatically reduced at the onset of sporulation. A deletion mutant of sigC grows normally in vegetative culture and is able to develop normally. However, in contrast to the wild-type cells, the sigC deletion mutant cells became capable of forming fruiting bodies and myxospores on semirich agar plates. This suggests that sigC may play a role in expression of genes involved in negatively regulating the initiation of fruiting body formation.

  10. In vitro culture and characterization of putative porcine embryonic germ cells derived from domestic breeds and yucatan mini pig embryos at days 20-24 of gestation

    DEFF Research Database (Denmark)

    Petkov, Stoyan Gueorguiev; Marks, Hendrik; Klein, Tino

    2011-01-01

    annotation clustering of the gene expression pattern of the putative EGC suggests partial differentiation toward endo/mesodermal lineages. The putative EGC were able to form embryoid bodies in suspension culture and to differentiate into epithelial-like, mesenchymal-like, and neuronal-like cells. However...

  11. Ten Putative Contributors to the Obesity Epidemic

    Science.gov (United States)

    McAllister, Emily J.; Dhurandhar, Nikhil V.; Keith, Scott W.; Aronne, Louis J.; Barger, Jamie; Baskin, Monica; Benca, Ruth M.; Biggio, Joseph; Boggiano, Mary M.; Eisenmann, Joe C.; Elobeid, Mai; Fontaine, Kevin R.; Gluckman, Peter; Hanlon, Erin C.; Katzmarzyk, Peter; Pietrobelli, Angelo; Redden, David T.; Ruden, Douglas M.; Wang, Chenxi; Waterland, Robert A.; Wright, Suzanne M.; Allison, David B.

    2010-01-01

    The obesity epidemic is a global issue and shows no signs of abating, while the cause of this epidemic remains unclear. Marketing practices of energy-dense foods and institutionally-driven declines in physical activity are the alleged perpetrators for the epidemic, despite a lack of solid evidence to demonstrate their causal role. While both may contribute to obesity, we call attention to their unquestioned dominance in program funding and public efforts to reduce obesity, and propose several alternative putative contributors that would benefit from equal consideration and attention. Evidence for microorganisms, epigenetics, increasing maternal age, greater fecundity among people with higher adiposity, assortative mating, sleep debt, endocrine disruptors, pharmaceutical iatrogenesis, reduction in variability of ambient temperatures, and intrauterine and intergenerational effects, as contributing factors to the obesity epidemic are reviewed herein. While the evidence is strong for some contributors such as pharmaceutical-induced weight gain, it is still emerging for other reviewed factors. Considering the role of such putative etiological factors of obesity may lead to comprehensive, cause specific, and effective strategies for prevention and treatment of this global epidemic. PMID:19960394

  12. 脂肪细胞分化过程中影响PAI-1基因转录表达的Dex和C/EBPs顺式调控元件的分析%Analysis of a Novel Dexamethasone Response Element and a Putative C/EBPs cis-Motif: Controlling PAI-1 Gene Expression During Adipocyte Differentiation

    Institute of Scientific and Technical Information of China (English)

    陈可洋; 马春姑; 汤其群; 宋后燕

    2002-01-01

    在研究胰岛素(Ins)、地塞米松(Dex)和甲基异丁基黄嘌呤(Mix)对脂肪细胞分化过程中PAI-1基因表达的影响基础上,为进一步探讨Ins、Dex调控PAI-1基因转录表达的调控机制,应用DNA重组技术,构建含萤光素酶(luciferase)报告基因和PAI-1启动子不同长度片段的嵌合质粒,转染3T3-L1前脂肪细胞并测定报告基因荧光素酶的活性.结果表明,小鼠PAI-1基因起动子-690至-850碱基序列之间有一个Dex的正调控元件.用计算机软件进行分析发现:Dex顺式元件位于PAI-1启动子的-750至-770碱基序列.其组成为:5′ GGTAACCTCTGTTCTCAT 3′.同时还发现在PAI-1启动子的-720至-740碱基序列中,存在一个C/EBPs的结合元件5′CCAAT3′并用凝胶电泳迁移实验对这些元件进行了鉴定.表明Dex正是通过激活转录因子(糖皮质激素受体,GR)和C/EBPα一起与各自的顺式元件结合来促进PAI-1基因的表达.%It has been reported that there is a significant increase in PAI-1 expression level in obese subjects. To explore the linkage between PAI-1 gene expression and obesity, the restriction enzymes and DNA recombination technologies were used to construct the chimeric plasmids with luciferase and different lengths of PAI-1 promoter. After transfection of the chimeric plasmids into 3T3-L1 preadipocyte and detection of luciferase activity, the results indicated that a positive dexamethasone cis-acting element (bases -690 to -850) may be present in mouse PAI-1 promoter. In addition, computer analysis using Match-Search Software found that a new motif of DexRE (dexamethasone response element) 5′ GGTAACCTCTGTTCTCAT 3′ and a putative C/EBPs binding site (cis-motif) exist respectively in the fragment (nucleotides -751 to -770) of, and a sequence (bases -720 to -740) of, mouse PAI-1 promoter,and GMSA and competition assays identified that the trans-acting factors induced by dexamethasone can specifically bind to those cis-motifs. Meanwhile

  13. Putative Nitrogen Sensing Systems in Higher Plants

    Institute of Scientific and Technical Information of China (English)

    Hon-Ming Lam; Ying Ann Chiao; Man-Wah Li; Yuk-Kwong Yung; Sang Ji

    2006-01-01

    Nitrogen (N) metabolism is essential for the biosynthesis of vital biomolecules. N status thus exerts profound effects on plant growth and development, and must be closely monitored. In bacteria and fungi, a few sophisticated N sensing systems have been extensively studied. In animals, the ability to receive amino acid signals has evolved to become an integral part of the nervous coordination system. In this review, we will summarize recent developments in the search for putative N sensing systems in higher plants based on homologous systems in bacteria, fungi, and animals. Apparently, although plants have separated and diversified from other organisms during the evolution process, striking similarities can be found in their N sensing systems compared with those of their counterparts; however, our understanding of these systems is still incomplete. Significant modifications of the N sensing systems (including cross-talk with other signal transduction pathways) in higher plants may be a strategy of adaptation to their unique mode of life.

  14. Putative respiratory chain of Porphyromonas gingivalis.

    Science.gov (United States)

    Meuric, Vincent; Rouillon, Astrid; Chandad, Fatiha; Bonnaure-Mallet, Martine

    2010-05-01

    The electron transfer chain in Porphyromonas gingivalis, or periodontopathogens, has not yet been characterized. P. gingivalis, a strict anaerobic bacteria and the second colonizer of the oral cavity, is considered to be a major causal agent involved in periodontal diseases. Primary colonizers create a favorable environment for P. gingivalis growth by decreasing oxygen pressure. Oxygen does not appear to be the final electron acceptor of the respiratory chain. Fumarate and cytochrome b have been implicated as major components of the respiratory activity. However, the P. gingivalis genome shows many other enzymes that could be implicated in aerobic or nitrite respiration. Using bioinformatic tools and literature studies of respiratory pathways, the ATP synthesis mechanism from the sodium cycle and nutrients metabolism, the putative respirasome of P. gingivalis has been proposed.

  15. Molecular diagnosis of putative Stargardt disease probands by exome sequencing

    Directory of Open Access Journals (Sweden)

    Strom Samuel P

    2012-08-01

    Full Text Available Abstract Background The commonest genetic form of juvenile or early adult onset macular degeneration is Stargardt Disease (STGD caused by recessive mutations in the gene ABCA4. However, high phenotypic and allelic heterogeneity and a small but non-trivial amount of locus heterogeneity currently impede conclusive molecular diagnosis in a significant proportion of cases. Methods We performed whole exome sequencing (WES of nine putative Stargardt Disease probands and searched for potentially disease-causing genetic variants in previously identified retinal or macular dystrophy genes. Follow-up dideoxy sequencing was performed for confirmation and to screen for mutations in an additional set of affected individuals lacking a definitive molecular diagnosis. Results Whole exome sequencing revealed seven likely disease-causing variants across four genes, providing a confident genetic diagnosis in six previously uncharacterized participants. We identified four previously missed mutations in ABCA4 across three individuals. Likely disease-causing mutations in RDS/PRPH2, ELOVL, and CRB1 were also identified. Conclusions Our findings highlight the enormous potential of whole exome sequencing in Stargardt Disease molecular diagnosis and research. WES adequately assayed all coding sequences and canonical splice sites of ABCA4 in this study. Additionally, WES enables the identification of disease-related alleles in other genes. This work highlights the importance of collecting parental genetic material for WES testing as the current knowledge of human genome variation limits the determination of causality between identified variants and disease. While larger sample sizes are required to establish the precision and accuracy of this type of testing, this study supports WES for inherited early onset macular degeneration disorders as an alternative to standard mutation screening techniques.

  16. Cryptic species in putative ancient asexual darwinulids (Crustacea, Ostracoda.

    Directory of Open Access Journals (Sweden)

    Isa Schön

    Full Text Available BACKGROUND: Fully asexually reproducing taxa lack outcrossing. Hence, the classic Biological Species Concept cannot be applied. METHODOLOGY/PRINCIPAL FINDINGS: We used DNA sequences from the mitochondrial COI gene and the nuclear ITS2 region to check species boundaries according to the evolutionary genetic (EG species concept in five morphospecies in the putative ancient asexual ostracod genera, Penthesilenula and Darwinula, from different continents. We applied two methods for detecting cryptic species, namely the K/θ method and the General Mixed Yule Coalescent model (GMYC. We could confirm the existence of species in all five darwinulid morphospecies and additional cryptic diversity in three morphospecies, namely in Penthesilenula brasiliensis, Darwinula stevensoni and in P. aotearoa. The number of cryptic species within one morphospecies varied between seven (P. brasiliensis, five to six (D. stevensoni and two (P. aotearoa, respectively, depending on the method used. Cryptic species mainly followed continental distributions. We also found evidence for coexistence at the local scale for Brazilian cryptic species of P. brasiliensis and P. aotearoa. Our ITS2 data confirmed that species exist in darwinulids but detected far less EG species, namely two to three cryptic species in P. brasiliensis and no cryptic species at all in the other darwinulid morphospecies. CONCLUSIONS/SIGNIFICANCE: Our results clearly demonstrate that both species and cryptic diversity can be recognized in putative ancient asexual ostracods using the EG species concept, and that COI data are more suitable than ITS2 for this purpose. The discovery of up to eight cryptic species within a single morphospecies will significantly increase estimates of biodiversity in this asexual ostracod group. Which factors, other than long-term geographic isolation, are important for speciation processes in these ancient asexuals remains to be investigated.

  17. Small intestinal mucosa expression of putative chaperone fls485

    Directory of Open Access Journals (Sweden)

    Raupach Kerstin

    2010-03-01

    Full Text Available Abstract Background Maturation of enterocytes along the small intestinal crypt-villus axis is associated with significant changes in gene expression profiles. fls485 coding a putative chaperone protein has been recently suggested as a gene involved in this process. The aim of the present study was to analyze fls485 expression in human small intestinal mucosa. Methods fls485 expression in purified normal or intestinal mucosa affected with celiac disease was investigated with a molecular approach including qRT-PCR, Western blotting, and expression strategies. Molecular data were corroborated with several in situ techniques and usage of newly synthesized mouse monoclonal antibodies. Results fls485 mRNA expression was preferentially found in enterocytes and chromaffine cells of human intestinal mucosa as well as in several cell lines including Rko, Lovo, and CaCo2 cells. Western blot analysis with our new anti-fls485 antibodies revealed at least two fls485 proteins. In a functional CaCo2 model, an increase in fls485 expression was paralleled by cellular maturation stage. Immunohistochemistry demonstrated fls485 as a cytosolic protein with a slightly increasing expression gradient along the crypt-villus axis which was impaired in celiac disease Marsh IIIa-c. Conclusions Expression and synthesis of fls485 are found in surface lining epithelia of normal human intestinal mucosa and deriving epithelial cell lines. An interdependence of enterocyte differentiation along the crypt-villus axis and fls485 chaperone activity might be possible.

  18. Small intestinal mucosa expression of putative chaperone fls485.

    Science.gov (United States)

    Reinartz, Andrea; Ehling, Josef; Franz, Susanne; Simon, Verena; Bravo, Ignacio G; Tessmer, Claudia; Zentgraf, Hanswalter; Lyer, Stefan; Schneider, Ursula; Köster, Jan; Raupach, Kerstin; Kämmerer, Elke; Klaus, Christina; Tischendorf, Jens J W; Kopitz, Jürgen; Alonso, Angel; Gassler, Nikolaus

    2010-03-07

    Maturation of enterocytes along the small intestinal crypt-villus axis is associated with significant changes in gene expression profiles. fls485 coding a putative chaperone protein has been recently suggested as a gene involved in this process. The aim of the present study was to analyze fls485 expression in human small intestinal mucosa. fls485 expression in purified normal or intestinal mucosa affected with celiac disease was investigated with a molecular approach including qRT-PCR, Western blotting, and expression strategies. Molecular data were corroborated with several in situ techniques and usage of newly synthesized mouse monoclonal antibodies. fls485 mRNA expression was preferentially found in enterocytes and chromaffine cells of human intestinal mucosa as well as in several cell lines including Rko, Lovo, and CaCo2 cells. Western blot analysis with our new anti-fls485 antibodies revealed at least two fls485 proteins. In a functional CaCo2 model, an increase in fls485 expression was paralleled by cellular maturation stage. Immunohistochemistry demonstrated fls485 as a cytosolic protein with a slightly increasing expression gradient along the crypt-villus axis which was impaired in celiac disease Marsh IIIa-c. Expression and synthesis of fls485 are found in surface lining epithelia of normal human intestinal mucosa and deriving epithelial cell lines. An interdependence of enterocyte differentiation along the crypt-villus axis and fls485 chaperone activity might be possible.

  19. Putative impact of RNA editing on drug discovery.

    Science.gov (United States)

    Decher, Niels; Netter, Michael F; Streit, Anne K

    2013-01-01

    Virtually all organisms use RNA editing as a powerful post-transcriptional mechanism to recode genomic information and to increase functional protein diversity. The enzymatic editing of pre-mRNA by ADARs and CDARs is known to change the functional properties of neuronal receptors and ion channels regulating cellular excitability. However, RNA editing is also an important mechanism for genes expressed outside the brain. The fact that RNA editing breaks the 'one gene encodes one protein' hypothesis is daunting for scientists and a probable drawback for drug development, as scientists might search for drugs targeting the 'wrong' protein. This possible difficulty for drug discovery and development became more evident from recent publications, describing that RNA editing events have profound impact on the pharmacology of some common drug targets. These recent studies highlight that RNA editing can cause massive discrepancies between the in vitro and in vivo pharmacology. Here, we review the putative impact of RNA editing on drug discovery, as RNA editing has to be considered before using high-throughput screens, rational drug design or choosing the right model organism for target validation.

  20. Structure, regulation, and putative function of the arginine deiminase system of Streptococcus suis.

    Science.gov (United States)

    Gruening, Petra; Fulde, Marcus; Valentin-Weigand, Peter; Goethe, Ralph

    2006-01-01

    Streptococcus suis is an important cause of infectious diseases in young pigs. Little is known about the virulence factors or protective antigens of S. suis. Recently, we have identified two proteins of the arginine deiminase system (ADS) of S. suis, which were temperature induced and expressed on the streptococcal surface (N. Winterhoff, R. Goethe, P. Gruening, M. Rohde, H. Kalisz, H. E. Smith, and P. Valentin-Weigand, J. Bacteriol. 184:6768-6776, 2002). In the present study, we analyzed the complete ADS of S. suis. Due to their homologies to the recently published S. gordonii ADS genes, the genes for arginine deiminase, ornithine carbamoyl-transferase, and carbamate kinase, which were previously designated adiS, octS, and ckS, respectively, were renamed arcA, arcB, and arcC, respectively. Our data revealed that arcA, arcB, and arcC of the S. suis ADS are transcribed from an operon (arcABC operon). Additionally, putative ADS-associated genes were cloned and sequenced which, however, did not belong to the arcABC operon. These were the flpS gene upstream of the arcABC operon with homology to the flp transcription regulator of S. gordonii and the arcD, arcT, arcH, and argR genes downstream of the arcABC operon with high homologies to a putative arginine-ornithine antiporter, a putative dipeptidase of S. gordonii, a putative beta-N-acetylhexosaminidase of S. pneumoniae, and a putative arginine repressor of S. gordonii, respectively. The transcriptional start point of the arcABC operon was determined, and promoter analysis provided evidence that multiple factors contribute to the regulation of the ADS. Thus, a putative binding site for a transcription regulator of the Crp/Fnr family, an ArgR-binding site, and two cis-acting catabolite response elements were identified in the promoter-operator region of the operon. Consistent with this, we could demonstrate that the ADS of S. suis is inducible by arginine and reduced O2 tension and subject to carbon catabolite

  1. A putative ABC transporter is involved in negative regulation of biofilm formation by Listeria monocytogenes

    DEFF Research Database (Denmark)

    Zhu, Xinna; Long, Fei; Chen, Yonghui

    2008-01-01

    Listeria monocytogenes may persist for long periods in food processing environments. In some instances, this may be due to aggregation or biofilm formation. To investigate the mechanism controlling biofilm formation in the food-borne pathogen L. monocytogenes, we characterized LM-49, a mutant...... with enhanced ability of biofilm-formation generated via transposon Tn917 mutagenesis of L. monocytogenes 4b G. In this mutant, a Tn917 insertion has disrupted the coding region of the gene encoding a putative ATP binding cassette (ABC) transporter permease identical to Lmof2365_1771 (a putative ABC......-transporter permease) presented in the sequenced strain L. monocytogenes str. 4b F2365. This disrupted gene, denoted lm.G_1771, encoded a protein with 10 transmembrane helixes. The revertant, LM-49RE, was obtained by replacing lm.G_1771::Tn917 with lm.G_1771 via homologous recombination. We found that LM-49RE formed...

  2. Putative Enzymes of UV Photoproduct Repair

    Directory of Open Access Journals (Sweden)

    Cynthia J. Sakofsky

    2011-01-01

    Full Text Available In order to determine the biological relevance of two S. acidocaldarius proteins to the repair of UV photoproducts, the corresponding genes (Saci_1227 and Saci_1096 were disrupted, and the phenotypes of the resulting mutants were examined by various genetic assays. The disruption used integration by homologous recombination of a functional but heterologous pyrE gene, promoted by short sequences attached to both ends via PCR. The phenotypic analyses of the disruptants confirmed that ORF Saci_1227 encodes a DNA photolyase which functions in vivo, but they could not implicate ORF Saci_1096 in repair of UV- or other externally induced DNA damage despite its similarity to genes encoding UV damage endonucleases. The success of the gene-disruption strategy, which used 5′ extensions of PCR primers to target cassette integration, suggests potential advantages for routine construction of Sulfolobus strains.

  3. Putative bronchopulmonary flagellated protozoa in immunosuppressed patients.

    Science.gov (United States)

    Kilimcioglu, Ali Ahmet; Havlucu, Yavuz; Girginkardesler, Nogay; Celik, Pınar; Yereli, Kor; Özbilgin, Ahmet

    2014-01-01

    Flagellated protozoa that cause bronchopulmonary symptoms in humans are commonly neglected. These protozoal forms which were presumed to be "flagellated protozoa" have been previously identified in immunosuppressed patients in a number of studies, but have not been certainly classified so far. Since no human cases of bronchopulmonary flagellated protozoa were reported from Turkey, we aimed to investigate these putative protozoa in immunosuppressed patients who are particularly at risk of infectious diseases. Bronchoalveolar lavage fluid samples of 110 immunosuppressed adult patients who were admitted to the Department of Chest Diseases, Hafsa Sultan Hospital of Celal Bayar University, Manisa, Turkey, were examined in terms of parasites by light microscopy. Flagellated protozoal forms were detected in nine (8.2%) of 110 cases. Metronidazole (500 mg b.i.d. for 30 days) was given to all positive cases and a second bronchoscopy was performed at the end of the treatment, which revealed no parasites. In conclusion, immunosuppressed patients with bronchopulmonary symptoms should attentively be examined with regard to flagellated protozoa which can easily be misidentified as epithelial cells.

  4. Mechanosensory neurons, cutaneous mechanoreceptors, and putative mechanoproteins.

    Science.gov (United States)

    Del Valle, M E; Cobo, T; Cobo, J L; Vega, J A

    2012-08-01

    The mammalian skin has developed sensory structures (mechanoreceptors) that are responsible for different modalities of mechanosensitivity like touch, vibration, and pressure sensation. These specialized sensory organs are anatomically and functionally connected to a special subset of sensory neurons called mechanosensory neurons, which electrophysiologically correspond with Aβ fibers. Although mechanosensory neurons and cutaneous mechanoreceptors are rather well known, the biology of the sense of touch still remains poorly understood. Basically, the process of mechanosensitivity requires the conversion of a mechanical stimulus into an electrical signal through the activation of ion channels that gate in response to mechanical stimuli. These ion channels belong primarily to the family of the degenerin/epithelium sodium channels, especially the subfamily acid-sensing ion channels, and to the family of transient receptor potential channels. This review compiles the current knowledge on the occurrence of putative mechanoproteins in mechanosensory neurons and mechanoreceptors, as well as the involvement of these proteins on the biology of touch. Furthermore, we include a section about what the knock-out mice for mechanoproteins are teaching us. Finally, the possibilities for mechanotransduction in mechanoreceptors, and the common involvement of the ion channels, extracellular membrane, and cytoskeleton, are revisited.

  5. Phylogeny of algal sequences encoding carbohydrate sulfotransferases, formylglycine-dependent sulfatases and putative sulfatase modifying factors

    Directory of Open Access Journals (Sweden)

    Chai-Ling eHo

    2015-11-01

    Full Text Available Many algae are rich sources of sulfated polysaccharides with biological activities. The physicochemical/rheological properties and biological activities of sulfated polysaccharides are affected by the pattern and number of sulfate moieties. Sulfation of carbohydrates is catalyzed by carbohydrate sulfotransferases (CHSTs while modification of sulfate moieties on sulfated polysaccharides was presumably catalyzed by sulfatases including formylglycine-dependent sulfatases (FGly-SULFs. Post-translationally modification of Cys to FGly in FGly-SULFs by sulfatase modifiying factors (SUMFs is necessary for the activity of this enzyme. The aims of this study are to mine for sequences encoding algal CHSTs, FGly-SULFs and putative SUMFs from the fully sequenced algal genomes and to infer their phylogenetic relationships to their well characterized counterparts from other organisms. Algal sequences encoding CHSTs, FGly-SULFs, SUMFs and SUMF-like proteins were successfully identified from green and brown algae. However, red algal FGly-SULFs and SUMFs were not identified. In addition, a group of SUMF-like sequences with different gene structure and possibly different functions were identified for green, brown and red algae. The phylogeny of these putative genes contributes to the corpus of knowledge of an unexplored area. The analyses of these putative genes contribute towards future production of existing and new sulfated carbohydrate polymers through enzymatic synthesis and metabolic engineering.

  6. Localization of putative carbonic anhydrases in two marine diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana.

    Science.gov (United States)

    Tachibana, Masaaki; Allen, Andrew E; Kikutani, Sae; Endo, Yuri; Bowler, Chris; Matsuda, Yusuke

    2011-09-01

    It is believed that intracellular carbonic anhydrases (CAs) are essential components of carbon concentrating mechanisms in microalgae. In this study, putative CA-encoding genes were identified in the genome sequences of the marine diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana. Subsequently, the subcellular localizations of the encoded proteins were determined. Nine and thirteen CA sequences were found in the genomes of P. tricornutum and T. pseudonana, respectively. Two of the β-CA genes in P. tricornutum corresponded to ptca1 and ptca2 identified previously. Immunostaining transmission electron microscopy of a PtCA1:YFP fusion expressed in the cells of P. tricornutum clearly showed the localization of PtCA1 within the central part of the pyrenoid structure in the chloroplast. Besides these two β-CA genes, P. tricornutum likely contains five α- and two γ-CA genes, whereas T. pseudonana has three α-, five γ-, four δ-, and one ζ-CA genes. Semi-quantitative reverse transcription PCR performed on mRNA from the two diatoms grown in changing light and CO(2) conditions revealed that levels of six putative α- and γ-CA mRNAs in P. tricornutum did not change between cells grown in air-level CO(2) and 5% CO(2). However, mRNA levels of one putative α-CA gene, CA-VII in P. tricornutum, were reduced in the dark compared to that in the light. In T. pseudonana, mRNA accumulation levels of putative α-CA (CA-1), ζ-CA (CA-3) and δ-CA (CA-7) were analyzed and all levels found to be significantly reduced when cells were grown in 0.16% CO(2). Intercellular localizations of eight putative CAs were analyzed by expressing GFP fusion in P. tricornutum and T. pseudonana. In P. tricornutum, CA-I and II localized in the periplastidial compartment, CA-III, VI, VII were found in the chloroplast endoplasmic reticulum, and CA-VIII was localized in the mitochondria. On the other hand, T. pseudonana CA-1 localized in the stroma and CA-3 was found in the periplasm

  7. Transcriptome of Aphanomyces euteiches: new oomycete putative pathogenicity factors and metabolic pathways.

    Directory of Open Access Journals (Sweden)

    Elodie Gaulin

    Full Text Available Aphanomyces euteiches is an oomycete pathogen that causes seedling blight and root rot of legumes, such as alfalfa and pea. The genus Aphanomyces is phylogenically distinct from well-studied oomycetes such as Phytophthora sp., and contains species pathogenic on plants and aquatic animals. To provide the first foray into gene diversity of A. euteiches, two cDNA libraries were constructed using mRNA extracted from mycelium grown in an artificial liquid medium or in contact to plant roots. A unigene set of 7,977 sequences was obtained from 18,864 high-quality expressed sequenced tags (ESTs and characterized for potential functions. Comparisons with oomycete proteomes revealed major differences between the gene content of A. euteiches and those of Phytophthora species, leading to the identification of biosynthetic pathways absent in Phytophthora, of new putative pathogenicity genes and of expansion of gene families encoding extracellular proteins, notably different classes of proteases. Among the genes specific of A. euteiches are members of a new family of extracellular proteins putatively involved in adhesion, containing up to four protein domains similar to fungal cellulose binding domains. Comparison of A. euteiches sequences with proteomes of fully sequenced eukaryotic pathogens, including fungi, apicomplexa and trypanosomatids, allowed the identification of A. euteiches genes with close orthologs in these microorganisms but absent in other oomycetes sequenced so far, notably transporters and non-ribosomal peptide synthetases, and suggests the presence of a defense mechanism against oxidative stress which was initially characterized in the pathogenic trypanosomatids.

  8. Gene

    Data.gov (United States)

    U.S. Department of Health & Human Services — Gene integrates information from a wide range of species. A record may include nomenclature, Reference Sequences (RefSeqs), maps, pathways, variations, phenotypes,...

  9. Phytophthora infestans specific phosphorylation patterns and new putative control targets.

    Science.gov (United States)

    Frades, Itziar; Andreasson, Erik

    2016-04-01

    In this study we applied biomathematical searches of gene regulatory mechanisms to learn more about oomycete biology and to identify new putative targets for pesticides or biological control against Phytophthora infestans. First, oomycete phylum-specific phosphorylation motifs were found by discriminative n-gram analysis. We found 11.600 P. infestans specific n-grams, mapping 642 phosphoproteins. The most abundant group among these related to phosphatidylinositol metabolism. Due to the large number of possible targets found and our hypothesis that multi-level control is a sign of usefulness as targets for intervention, we identified overlapping targets with a second screen. This was performed to identify proteins dually regulated by small RNA and phosphorylation. We found 164 proteins to be regulated by both sRNA and phosphorylation and the dominating functions where phosphatidylinositol signalling/metabolism, endocytosis, and autophagy. Furthermore we performed a similar regulatory study and discriminative n-gram analysis of proteins with no clear orthologs in other species and proteins that are known to be unique to P. infestans such as the RxLR effectors, Crinkler (CRN) proteins and elicitins. We identified CRN proteins with specific phospho-motifs present in all life stages. PITG_12626, PITG_14042 and PITG_23175 are CRN proteins that have species-specific phosphorylation motifs and are subject to dual regulation.

  10. Rapid Discrimination Among Putative Mechanistic Models of Biochemical Systems.

    Science.gov (United States)

    Lomnitz, Jason G; Savageau, Michael A

    2016-08-31

    An overarching goal in molecular biology is to gain an understanding of the mechanistic basis underlying biochemical systems. Success is critical if we are to predict effectively the outcome of drug treatments and the development of abnormal phenotypes. However, data from most experimental studies is typically noisy and sparse. This allows multiple potential mechanisms to account for experimental observations, and often devising experiments to test each is not feasible. Here, we introduce a novel strategy that discriminates among putative models based on their repertoire of qualitatively distinct phenotypes, without relying on knowledge of specific values for rate constants and binding constants. As an illustration, we apply this strategy to two synthetic gene circuits exhibiting anomalous behaviors. Our results show that the conventional models, based on their well-characterized components, cannot account for the experimental observations. We examine a total of 40 alternative hypotheses and show that only 5 have the potential to reproduce the experimental data, and one can do so with biologically relevant parameter values.

  11. Epigenetic regulation of putative tumor suppressor TGFBI in human leukemias

    Institute of Scientific and Technical Information of China (English)

    Fang Hongbo; Liu Jing; Guo Dan; Liu Peixiang; Zhao Yongliang

    2014-01-01

    Background Both in vitro and in vivo data have demonstrated the TGFBI gene functions as a putative tumor suppressor and is frequently downregulated in human tumors of different histological types.The hypermethylation of the TGFBI promoter,as one of the main regulatory mechanisms,is associated with TGFBI silencing.In this study,we used a methylation-specific PCR (MSP) method to evaluate the methylation status of the TGFBI promoter in human leukemias.Methods Real-time RT-PCR and methylation-specific PCR approaches were performed to define the TGFBI expression and promoter methylation in human leukemia call lines and clinical samples.Genomic DNA was isolated from peripheral blood mononuclear cells from leukemia patients,bisulfite-converted,and analyzed by the MSP method.Results Hypermethylation of the TGFBI promoter occurred in leukemia cell lines and demethylation treatment reexpressed TGFBI at a substantially increased level in most of leukemia cell lines tested.Furthermore,a much higher level of CpG island methylation and a significantly lower TGFBI expression were also identified in clinical leukemia samples.Conclusion The results suggest an important role of promoter methylation in regulating TGFBI expression in leukemia,which provides a useful diagnostic marker for clinical management of human leukemias.

  12. The DNA sequence of a 7941 bp fragment of the left arm of chromosome VII of Saccharomyces cerevisiae contains four open reading frames including the multicopy suppressor gene of the pop2 mutation and a putative serine/threonine protein kinase gene.

    Science.gov (United States)

    Coglievina, M; Bertani, I; Klima, R; Zaccaria, P; Bruschi, C V

    1995-06-30

    We report the sequence of a 7941 bp DNA fragment from the left arm of chromosome VII of Saccharomyces cerevisiae which contains four open reading frames (ORFs) of greater than 100 amino acid residues. ORF biC834 shows 100% bp identity with the recently identified multicopy suppressor gene of the pop2 mutation (MPT5); its deduced protein product carries an eight-repeat domain region, homologous to that found in the hypothetical regulatory YGL023 protein of S. cerevisiae and the Pumilio protein of Drosophila. ORF biE560 protein exhibits patterns typical of serine/threonine protein kinases, with which it shares high degrees of homology.

  13. Functional characterization of PaLAX1, a putative auxin permease, in heterologous plant systems.

    Science.gov (United States)

    Hoyerová, Klára; Perry, Lucie; Hand, Paul; Lanková, Martina; Kocábek, Tomás; May, Sean; Kottová, Jana; Paces, Jan; Napier, Richard; Zazímalová, Eva

    2008-03-01

    We have isolated the cDNA of the gene PaLAX1 from a wild cherry tree (Prunus avium). The gene and its product are highly similar in sequences to both the cDNAs and the corresponding protein products of AUX/LAX-type genes, coding for putative auxin influx carriers. We have prepared and characterized transformed Nicotiana tabacum and Arabidopsis thaliana plants carrying the gene PaLAX1. We have proved that constitutive overexpression of PaLAX1 is accompanied by changes in the content and distribution of free indole-3-acetic acid, the major endogenous auxin. The increase in free indole-3-acetic acid content in transgenic plants resulted in various phenotype changes, typical for the auxin-overproducing plants. The uptake of synthetic auxin, 2,4-dichlorophenoxyacetic acid, was 3 times higher in transgenic lines compared to the wild-type lines and the treatment with the auxin uptake inhibitor 1-naphthoxyacetic acid reverted the changes caused by the expression of PaLAX1. Moreover, the agravitropic response could be restored by expression of PaLAX1 in the mutant aux1 plants, which are deficient in auxin influx carrier activity. Based on our data, we have concluded that the product of the gene PaLAX1 promotes the uptake of auxin into cells, and, as a putative auxin influx carrier, it affects the content and distribution of free endogenous auxin in transgenic plants.

  14. Identification and Characterization of Putative Integron-Like Elements of the Heavy-Metal-Hypertolerant Strains of Pseudomonas spp.

    Science.gov (United States)

    Ciok, Anna; Adamczuk, Marcin; Bartosik, Dariusz; Dziewit, Lukasz

    2016-11-28

    Pseudomonas strains isolated from the heavily contaminated Lubin copper mine and Zelazny Most post-flotation waste reservoir in Poland were screened for the presence of integrons. This analysis revealed that two strains carried homologous DNA regions composed of a gene encoding a DNA_BRE_C domain-containing tyrosine recombinase (with no significant sequence similarity to other integrases of integrons) plus a three-component array of putative integron gene cassettes. The predicted gene cassettes encode three putative polypeptides with homology to (i) transmembrane proteins, (ii) GCN5 family acetyltransferases, and (iii) hypothetical proteins of unknown function (homologous proteins are encoded by the gene cassettes of several class 1 integrons). Comparative sequence analyses identified three structural variants of these novel integron-like elements within the sequenced bacterial genomes. Analysis of their distribution revealed that they are found exclusively in strains of the genus Pseudomonas.

  15. Hippocampal and thalamic neuronal metabolism in a putative rat model of schizophrenia○

    Institute of Scientific and Technical Information of China (English)

    Guolin Ma; Tianbin Song; Min Chen; Yuan Fu; Yong Xu; Ensen Ma; Wu Wang; Jiang Du; Mingxiong Huang

    2013-01-01

    The transcription factor early growth response protein 3 (EGR3) is involved in schizophrenia. We developed a putative rat model of schizophrenia by transfecting lentiviral particles carrying the Egr3 gene into bilateral hippocampal dentate gyrus. We assessed spatial working memory using the Morris water maze test, and neuronal metabolite levels in bilateral hippocampus and thalamus were determined by 3.0 T proton magnetic resonance spectroscopy. Choline content was significantly greater in the hippocampus after transfection, while N-acetylaspartate and the ratio of N-acetylaspartate to creatine/phosphocreatine in the thalamus were lower than in controls. This study is the first to report evaluation of brain metabolites using 3.0 T proton magnetic resonance spectroscopy in rats transfected with Egr3, and reveals metabolic abnormalities in the hippocampus and thalamus in this putative model of schizophrenia.

  16. Characterization of three putative xylulose 5-phosphate/fructose 6-phosphate phosphoketolases in the cyanobacterium Anabaena sp. PCC 7120.

    Science.gov (United States)

    Moriyama, Takashi; Tajima, Naoyuki; Sekine, Kohsuke; Sato, Naoki

    2015-01-01

    Xylulose 5-phosphate/fructose 6-phosphate phosphoketolase (Xfp) is a key enzyme in the central carbohydrate metabolism in heterofermentative bacteria, in which enzymatic property of Xfps is well characterized. This is not the case in other microbes. The cyanobacterium Anabaena sp. PCC 7120 possesses three putative genes encoding Xfp, all1483, all2567, and alr1850. We purified three putative Xfps as recombinant proteins. The results of gel filtration indicated that these proteins form homomultimer complex. All1483 and All2567 showed phosphoketolase activity, whereas Alr1850 did not show the activity. Kinetic analyses demonstrated that substrates, fructose 6-phosphate and inorganic phosphate, are cooperatively bound to enzymes positively and negatively, respectively.

  17. Regulation of Arabidopsis Early Anther Development by Putative Cell-Cell Signaling Molecules and Transcriptional Regulators

    Institute of Scientific and Technical Information of China (English)

    Yu-Jin Sun; Carey LH Hord; Chang-Bin Chen; Hong Ma

    2007-01-01

    Anther development in flowering plants involves the formation of several cell types, including the tapetal and pollen mother cells. The use of genetic and molecular tools has led to the identification and characterization of genes that are critical for normal cell division and differentiation in Arabidopsis early anther development. We review here several recent studies on these genes, including the demonstration that the putative receptor protein kinases BAM1 and BAM2 together play essential roles in the control of early cell division and differentiation. In addition, we discuss the hypothesis that BAM1/2 may form a positive-negative feedback regulatory loop with a previously identified key regulator, SPOROCYTELESS (also called NOZZLE),to control the balance between sporogenous and somatic cell types in the anther. Furthermore, we summarize the isolation and functional analysis of the DYSFUNCTIONAL TAPETUM1 (DYT1) gene in promoting proper tapetal cell differentiation. Our finding that DYT1 encodes a putative transcription factor of the bHLH family, as well as relevant expression analyses, strongly supports a model that DYT1 serves as a critical link between upstream factors and downstream target genes that are critical for normal tapetum development and function. These studies, together with other recently published works, indicate that cell-cell communication and transcriptional control are key processes essential for cell fate specification in anther development.

  18. Unprecedented diversity of catalytic domains in the first four modules of the putative pederin polyketide synthase.

    Science.gov (United States)

    Piel, Jörn; Wen, Gaiping; Platzer, Matthias; Hui, Dequan

    2004-01-03

    Polyketides of the pederin group are highly potent antitumor compounds found in terrestrial beetles and marine sponges. Pederin is used by beetles of the genera Paederus and Paederidus as a chemical defense. We have recently identified a group of putative pederin biosynthesis genes and localized them to the genome of an as yet unculturable Pseudomonas sp. symbiont, the likely true pederin producer. However, this polyketide synthase cluster lacks several genes expected for pederin production. Here we report an additional polyketide synthase encoded on a separate region of the symbiont genome. It contains at least three novel catalytic domains that are predicted to be involved in pederin chain initiation and the formation of an unusual exomethylene bond. The region is bordered by mobility pseudogenes; this suggests that gene transposition led to the disjointed cluster organization. With this work, all putative pederin genes have been identified. Their heterologous expression in a culturable bacterium will provide important insights into how sustainable sources of invertebrate-derived drug candidates can be created.

  19. A fruit quality gene map of Prunus

    National Research Council Canada - National Science Library

    Ogundiwin, Ebenezer A; Peace, Cameron P; Gradziel, Thomas M; Parfitt, Dan E; Bliss, Fredrick A; Crisosto, Carlos H

    2009-01-01

    ... to marketing and consumption. Here we present an integrated fruit quality gene map of Prunus containing 133 genes putatively involved in the determination of fruit texture, pigmentation, flavor, and chilling injury resistance...

  20. Putative Corneal Neuralgia Responding to Vitamin D Supplementation

    Directory of Open Access Journals (Sweden)

    Eric L. Singman

    2013-09-01

    Full Text Available A patient with putative corneal neuralgia was incidentally discovered to have hypovitaminosis D. Supplementation of vitamin D appears to have led to a resolution of the patient's pain, whereas other efforts to treat the patient were unsuccessful.

  1. Evaluating the function of putative hormone transporters.

    Science.gov (United States)

    Frommer, Wolf B; Schulz, Burkhard; Murphy, Angus S

    2009-02-01

    Hormones typically serve as long distance signaling molecules. To reach their site of action, hormones need to be transported from the sites of synthesis. Many plant hormones are mobile, thus requiring specific transport systems for the export from their source cells as well as subsequent import into target cells. Hormone transport in general is still poorly understood. Auxin is probably the most intensively studied plant hormone concerning transport in the moment. To advance our understanding of hormone transport we need two principal data sets: information on the properties of the transport systems including substrate specificity and kinetics, and we need to identify candidate genes for the respective transporters. Physiological transport data can provide an important basis for identifying and characterizing candidate transporters and to define their in vivo role. A recent publication in Plant Physiology highlights how kinetic and specificity studies may help to identify cytokinin transporters.

  2. Putative ancient microorganisms from amber nuggets.

    Science.gov (United States)

    Veiga-Crespo, Patricia; Blasco, Lucía; Poza, Margarita; Villa, Tomás G

    2007-06-01

    Evolutionary microbiology studies based on the isolation of ancient DNA and/or microbial samples are scarce due to the difficulty of finding well preserved biological specimens. However, amber is a fossil resin with natural preserving properties for microbial cells and DNA. The visualization by transmission electron microscopy of different microorganism-like specimens found in amber nuggets from both the Miocene and the Cretaceous periods was accompanied by studies of ancient DNA obtained from the nuggets. After the design of specific primers based on the present sequences of both genes in Saccharomyces cerevisiae, the ancestral AGP2 sequence from the Miocene, as well as the 18S rRNA from the Cretaceous, were amplified.

  3. The expression of superoxide dismutase (SOD) and a putative ABC transporter permease is inversely correlated during biofilm formation in Listeria monocytogenes 4b G

    Science.gov (United States)

    Little is known about the molecular basis of biofilm formation in Listeria monocytogenes. The superoxide dismutase (SOD) of the deletion mutant of lm.G_1771 gene, which encodes for a putative ABC_transporter permease, is highly expressed in biofilm. In this study, the sod gene deletion mutant delta ...

  4. Identification of putative cis-regulatory elements in Cryptosporidium parvum by de novo pattern finding

    Directory of Open Access Journals (Sweden)

    Kissinger Jessica C

    2007-01-01

    Full Text Available Abstract Background Cryptosporidium parvum is a unicellular eukaryote in the phylum Apicomplexa. It is an obligate intracellular parasite that causes diarrhea and is a significant AIDS-related pathogen. Cryptosporidium parvum is not amenable to long-term laboratory cultivation or classical molecular genetic analysis. The parasite exhibits a complex life cycle, a broad host range, and fundamental mechanisms of gene regulation remain unknown. We have used data from the recently sequenced genome of this organism to uncover clues about gene regulation in C. parvum. We have applied two pattern finding algorithms MEME and AlignACE to identify conserved, over-represented motifs in the 5' upstream regions of genes in C. parvum. To support our findings, we have established comparative real-time -PCR expression profiles for the groups of genes examined computationally. Results We find that groups of genes that share a function or belong to a common pathway share upstream motifs. Different motifs are conserved upstream of different groups of genes. Comparative real-time PCR studies show co-expression of genes within each group (in sub-sets during the life cycle of the parasite, suggesting co-regulation of these genes may be driven by the use of conserved upstream motifs. Conclusion This is one of the first attempts to characterize cis-regulatory elements in the absence of any previously characterized elements and with very limited expression data (seven genes only. Using de novo pattern finding algorithms, we have identified specific DNA motifs that are conserved upstream of genes belonging to the same metabolic pathway or gene family. We have demonstrated the co-expression of these genes (often in subsets using comparative real-time-PCR experiments thus establishing evidence for these conserved motifs as putative cis-regulatory elements. Given the lack of prior information concerning expression patterns and organization of promoters in C. parvum we

  5. Genetic diversity of bovine papillomavirus types, including two putative new types, in teat warts from dairy cattle herds.

    Science.gov (United States)

    Lunardi, Michele; de Camargo Tozato, Claudia; Alfieri, Alice Fernandes; de Alcântara, Brígida Kussumoto; Vilas-Boas, Laurival Antonio; Otonel, Rodrigo Alejandro Arellano; Headley, Selwyn Arlington; Alfieri, Amauri Alcindo

    2016-06-01

    Teat papillomatosis affects dairy cows worldwide. Milking can become difficult due to teat warts, and maintaining affected cows in the herds may diminish economic profit in the dairy industry. Currently, 13 bovine papillomavirus (BPV) types have been fully characterized, and numerous putative BPV types have been identified through partial L1 gene PCR. In order to identify the viral types present in warts on the udders of dairy cows, 40 teat lesions from 24 cows from 13 cattle farms in three States of Brazil were evaluated by PV L1 gene PCR. The warts that were evaluated contained sequences from BPVs 6-10, the putative BPV types BAPV9 and BAPV4, and two unreported putative papillomavirus (PV) types, named BPV/BR-UEL6 and BPV/BR-UEL7. In addition, mixed infections and coinfections were identified, since more than one lesion was observed on the udders of 13 cows. Phylogenetic analysis showed that BPV/BR-UEL6 is closely related to BPVs belonging to the genus Xipapillomavirus, while BPV/BR-UEL7 clustered with the previously reported strains Cervus timorensis and Pudu puda PVs, which represent a putative new PV type, and it was only distantly related to xi-, epsilon-, delta- and dyoxi-PVs. These results provide information that will assist in the understanding of the association of BPVs 6, 7, 8, 9, and 10, as well as putative BPV types BAPV4 and BAPV9, with mammary papillomatosis. This is the first characterization of putative novel PV types BPV/BR-UEL6 and BPV/BR-UEL7 in teat warts of dairy cows, highlighting the high genetic diversity of BPVs associated with teat papillomatosis.

  6. Salivary PYY: a putative bypass to satiety.

    Directory of Open Access Journals (Sweden)

    Andres Acosta

    Full Text Available Peptide YY(3-36 is a satiation hormone released postprandially into the bloodstream from L-endocrine cells in the gut epithelia. In the current report, we demonstrate PYY(3-36 is also present in murine as well as in human saliva. In mice, salivary PYY(3-36 derives from plasma and is also synthesized in the taste cells in taste buds of the tongue. Moreover, the cognate receptor Y2R is abundantly expressed in the basal layer of the progenitor cells of the tongue epithelia and von Ebner's gland. The acute augmentation of salivary PYY(3-36 induced stronger satiation as demonstrated in feeding behavioral studies. The effect is mediated through the activation of the specific Y2 receptor expressed in the lingual epithelial cells. In a long-term study involving diet-induced obese (DIO mice, a sustained increase in PYY(3-36 was achieved using viral vector-mediated gene delivery targeting salivary glands. The chronic increase in salivary PYY(3-36 resulted in a significant long-term reduction in food intake (FI and body weight (BW. Thus this study provides evidence for new functions of the previously characterized gut peptide PYY(3-36 suggesting a potential simple and efficient alternative therapeutic approach for the treatment of obesity.

  7. The molecular dimension of microbial species: 1. Ecological distinctions among, and homogeneity within, putative ecotypes of Synechococcus inhabiting the cyanobacterial mat of Mushroom Spring, Yellowstone National Park

    Directory of Open Access Journals (Sweden)

    Eric Daniel Becraft

    2015-06-01

    Full Text Available Based on the Stable Ecotype Model, evolution leads to the divergence of ecologically distinct populations (e.g., with different niches and/or behaviors of ecologically interchangeable membership. In this study, pyrosequencing was used to provide deep sequence coverage of Synechococcus psaA genes and transcripts over a large number of habitat types in the Mushroom Spring microbial mat. Putative ecological species (putative ecotypes, which were predicted by an evolutionary simulation based on the Stable Ecotype Model (Ecotype Simulation, exhibited distinct distributions relative to temperature-defined positions in the effluent channel and vertical position in the upper 1 mm-thick mat layer. Importantly, in most cases variants predicted to belong to the same putative ecotype formed unique clusters relative to temperature and depth in the mat in canonical correspondence analysis, supporting the hypothesis that while the putative ecotypes are ecologically distinct, the members of each ecotype are ecologically homogeneous. Putative ecotypes responded differently to experimental perturbations of temperature and light, but the genetic variation within each putative ecotype was maintained as the relative abundances of putative ecotypes changed, further indicating that each population responded as a set of ecologically interchangeable individuals. Compared to putative ecotypes that predominate deeper within the mat photic zone, the timing of transcript abundances for selected genes differed for putative ecotypes that predominate in microenvironments closer to upper surface of the mat with spatiotemporal differences in light and O2 concentration. All of these findings are consistent with the hypotheses that Synechococcus species in hot spring mats are sets of ecologically interchangeable individuals that are differently adapted, that these adaptations control their distributions, and that the resulting distributions constrain the activities of the species

  8. Identification of Putative Coffee Rust Mycoparasites via Single-Molecule DNA Sequencing of Infected Pustules.

    Science.gov (United States)

    James, Timothy Y; Marino, John A; Perfecto, Ivette; Vandermeer, John

    2015-11-13

    The interaction of crop pests with their natural enemies is a fundament to their control. Natural enemies of fungal pathogens of crops are poorly known relative to those of insect pests, despite the diversity of fungal pathogens and their economic importance. Currently, many regions across Latin America are experiencing unprecedented epidemics of coffee rust (Hemileia vastatrix). Identification of natural enemies of coffee rust could aid in developing management strategies or in pinpointing species that could be used for biocontrol. In the present study, we characterized fungal communities associated with coffee rust lesions by single-molecule DNA sequencing of fungal rRNA gene bar codes from leaf discs (≈28 mm(2)) containing rust lesions and control discs with no rust lesions. The leaf disc communities were hyperdiverse in terms of fungi, with up to 69 operational taxonomic units (putative species) per control disc, and the diversity was only slightly reduced in rust-infected discs, with up to 63 putative species. However, geography had a greater influence on the fungal community than whether the disc was infected by coffee rust. Through comparisons between control and rust-infected leaf discs, as well as taxonomic criteria, we identified 15 putative mycoparasitic fungi. These fungi are concentrated in the fungal family Cordycipitaceae and the order Tremellales. These data emphasize the complexity of diverse fungi of unknown ecological function within a leaf that might influence plant disease epidemics or lead to the development of species for biocontrol of fungal disease.

  9. Genomic identification of a putative circadian system in the cladoceran crustacean Daphnia pulex

    Science.gov (United States)

    Tilden, Andrea R.; McCoole, Matthew D.; Harmon, Sarah M.; Baer, Kevin N.; Christie, Andrew E.

    2011-01-01

    Essentially nothing is known about the molecular underpinnings of crustacean circadian clocks. The genome of Daphnia pulex, the only crustacean genome available for public use, provides a unique resource for identifying putative circadian proteins in this species. Here, the Daphnia genome was mined for putative circadian protein genes using Drosophila melanogaster queries. The sequences of core clock (e.g. CLOCK, CYCLE, PERIOD, TIMELESS and CRYPTOCHROME 2), clock input (CRYPTOCHROME 1) and clock output (PIGMENT DISPERSING HORMONE RECEPTOR) proteins were deduced. Structural analyses and alignment of the Daphnia proteins with their Drosophila counterparts revealed extensive sequence conservation, particularly in functional domains. Comparisons of the Daphnia proteins with other sequences showed that they are, in most cases, more similar to homologs from other species, including vertebrates, than they are to those of Drosophila. The presence of both CRYPTOCHROME 1 and 2 in Daphnia suggests the organization of its clock may be more similar to that of the butterfly Danaus plexippus than to that of Drosophila (which possesses CRYPTOCHROME 1 but not CRYPTOCHROME 2). These data represent the first description of a putative circadian system from any crustacean, and provide a foundation for future molecular, anatomical and physiological investigations of circadian signaling in Daphnia. PMID:21798832

  10. Identification and site of action of the remaining four putative pseudouridine synthases in Escherichia coli.

    Science.gov (United States)

    Del Campo, M; Kaya, Y; Ofengand, J

    2001-11-01

    There are 10 known putative pseudouridine synthase genes in Escherichia coli. The products of six have been previously assigned, one to formation of the single pseudouridine in 16S RNA, three to the formation of seven pseudouridines in 23S RNA, and three to the formation of three pseudouridines in tRNA (one synthase makes pseudouridine in 23S RNA and tRNA). Here we show that the remaining four putative synthase genes make bona fide pseudouridine synthases and identify which pseudouridines they make. RluB (formerly YciL) and RluE (formerly YmfC) make pseudouridine2605 and pseudouridine2457, respectively, in 23S RNA. RluF (formerly YjbC) makes the newly discovered pseudouridine2604 in 23S RNA, and TruC (formerly YqcB) makes pseudouridine65 in tRNA(Ile1) and tRNA(Asp). Deletion of each of these synthase genes individually had no effect on exponential growth in rich media at 25 degrees C, 37 degrees C, or 42 degrees C. A strain lacking RluB and RluF also showed no growth defect under these conditions. Mutation of a conserved aspartate in a common sequence motif, previously shown to be essential for the other six E. coli pseudouridine synthases and several yeast pseudouridine synthases, also caused a loss of in vivo activity in all four of the synthases studied in this work.

  11. Biochemical Characterization of Putative Adenylate Dimethylallyltransferase and Cytokinin Dehydrogenase from Nostoc sp. PCC 7120.

    Science.gov (United States)

    Frébortová, Jitka; Greplová, Marta; Seidl, Michael F; Heyl, Alexander; Frébort, Ivo

    2015-01-01

    Cytokinins, a class of phytohormones, are adenine derivatives common to many different organisms. In plants, these play a crucial role as regulators of plant development and the reaction to abiotic and biotic stress. Key enzymes in the cytokinin synthesis and degradation in modern land plants are the isopentyl transferases and the cytokinin dehydrogenases, respectively. Their encoding genes have been probably introduced into the plant lineage during the primary endosymbiosis. To shed light on the evolution of these proteins, the genes homologous to plant adenylate isopentenyl transferase and cytokinin dehydrogenase were amplified from the genomic DNA of cyanobacterium Nostoc sp. PCC 7120 and expressed in Escherichia coli. The putative isopentenyl transferase was shown to be functional in a biochemical assay. In contrast, no enzymatic activity was detected for the putative cytokinin dehydrogenase, even though the principal domains necessary for its function are present. Several mutant variants, in which conserved amino acids in land plant cytokinin dehydrogenases had been restored, were inactive. A combination of experimental data with phylogenetic analysis indicates that adenylate-type isopentenyl transferases might have evolved several times independently. While the Nostoc genome contains a gene coding for protein with characteristics of cytokinin dehydrogenase, the organism is not able to break down cytokinins in the way shown for land plants.

  12. Evaluation of two putative susceptibility loci for oral clefts in the Danish population

    DEFF Research Database (Denmark)

    Mitchell, L E; Murray, J C; O'Brien, S;

    2001-01-01

    Previous studies suggest that the risk of nonsyndromic cleft lip with or without cleft palate (CL+/-P) and isolated cleft palate (CP) is influenced by genetic variation at several loci and that the relation between specific genetic variants and disease risk may be modified by environmental factors....... The present study evaluated potential associations between CL+/-P and CP and two putative clefting susceptibility loci, MSX1 and TGFB3, using data from a nationwide case-control study conducted in Denmark from 1991 to 1994. The potential effects of interactions between these genes and two common environmental...

  13. A putative transglycosylase encoded by SCO4132 influences morphological differentiation and actinorhodin production in Streptomyces coelicolor

    Institute of Scientific and Technical Information of China (English)

    Pengfei Xie; Ana Zeng; Xiaoting Lv; Qiuxiang Cheng; Zhongjun Qin

    2013-01-01

    Here we report that tgdA,a novel gene encoding a putative transglycosylase,affects both the morphological differentiation and the yield of blue-pigmented compound actinorhodin in Streptomyces coelicolor.The tgdA null mutant displays sparse aerial hyphae and irregular spore chains frequently lacking chromosomal DNA.Elevated actinorhodin production coincides with the overexpression of actⅡ-orf4 in mutant.tgdA expression is temporally and developmentally regulated.The tgdA orthologs in Streptomyces avermilitis and Streptomyces lividans also affect differentiation.

  14. The putative oncogene Pim-1 in the mouse: its linkage and variation among t haplotypes.

    Science.gov (United States)

    Nadeau, J H; Phillips, S J

    1987-11-01

    Pim-1, a putative oncogene involved in T-cell lymphomagenesis, was mapped between the pseudo-alpha globin gene Hba-4ps and the alpha-crystallin gene Crya-1 on mouse chromosome 17 and therefore within the t complex. Pim-1 restriction fragment variants were identified among t haplotypes. Analysis of restriction fragment sizes obtained with 12 endonucleases demonstrated that the Pim-1 genes in some t haplotypes were indistinguishable from the sizes for the Pim-1b allele in BALB/c inbred mice. There are now three genes, Pim-1, Crya-1 and H-2 I-E, that vary among independently derived t haplotypes and that have indistinguishable alleles in t haplotypes and inbred strains. These genes are closely linked within the distal inversion of the t complex. Because it is unlikely that these variants arose independently in t haplotypes and their wild-type homologues, we propose that an exchange of chromosomal segments, probably through double crossingover, was responsible for indistinguishable Pim-1 genes shared by certain t haplotypes and their wild-type homologues. There was, however, no apparent association between variant alleles of these three genes among t haplotypes as would be expected if a single exchange introduced these alleles into t haplotypes. If these variant alleles can be shown to be identical to the wild-type allele, then lack of association suggests that multiple exchanges have occurred during the evolution of the t complex.

  15. Genome-wide analysis of putative peroxiredoxin in unicellular and filamentous cyanobacteria

    Directory of Open Access Journals (Sweden)

    Cui Hongli

    2012-11-01

    Full Text Available Abstract Background Cyanobacteria are photoautotrophic prokaryotes with wide variations in genome sizes and ecological habitats. Peroxiredoxin (PRX is an important protein that plays essential roles in protecting own cells against reactive oxygen species (ROS. PRXs have been identified from mammals, fungi and higher plants. However, knowledge on cyanobacterial PRXs still remains obscure. With the availability of 37 sequenced cyanobacterial genomes, we performed a comprehensive comparative analysis of PRXs and explored their diversity, distribution, domain structure and evolution. Results Overall 244 putative prx genes were identified, which were abundant in filamentous diazotrophic cyanobacteria, Acaryochloris marina MBIC 11017, and unicellular cyanobacteria inhabiting freshwater and hot-springs, while poor in all Prochlorococcus and marine Synechococcus strains. Among these putative genes, 25 open reading frames (ORFs encoding hypothetical proteins were identified as prx gene family members and the others were already annotated as prx genes. All 244 putative PRXs were classified into five major subfamilies (1-Cys, 2-Cys, BCP, PRX5_like, and PRX-like according to their domain structures. The catalytic motifs of the cyanobacterial PRXs were similar to those of eukaryotic PRXs and highly conserved in all but the PRX-like subfamily. Classical motif (CXXC of thioredoxin was detected in protein sequences from the PRX-like subfamily. Phylogenetic tree constructed of catalytic domains coincided well with the domain structures of PRXs and the phylogenies based on 16s rRNA. Conclusions The distribution of genes encoding PRXs in different unicellular and filamentous cyanobacteria especially those sub-families like PRX-like or 1-Cys PRX correlate with the genome size, eco-physiology, and physiological properties of the organisms. Cyanobacterial and eukaryotic PRXs share similar conserved motifs, indicating that cyanobacteria adopt similar catalytic

  16. Genome-wide analysis of putative peroxiredoxin in unicellular and filamentous cyanobacteria

    Science.gov (United States)

    2012-01-01

    Background Cyanobacteria are photoautotrophic prokaryotes with wide variations in genome sizes and ecological habitats. Peroxiredoxin (PRX) is an important protein that plays essential roles in protecting own cells against reactive oxygen species (ROS). PRXs have been identified from mammals, fungi and higher plants. However, knowledge on cyanobacterial PRXs still remains obscure. With the availability of 37 sequenced cyanobacterial genomes, we performed a comprehensive comparative analysis of PRXs and explored their diversity, distribution, domain structure and evolution. Results Overall 244 putative prx genes were identified, which were abundant in filamentous diazotrophic cyanobacteria, Acaryochloris marina MBIC 11017, and unicellular cyanobacteria inhabiting freshwater and hot-springs, while poor in all Prochlorococcus and marine Synechococcus strains. Among these putative genes, 25 open reading frames (ORFs) encoding hypothetical proteins were identified as prx gene family members and the others were already annotated as prx genes. All 244 putative PRXs were classified into five major subfamilies (1-Cys, 2-Cys, BCP, PRX5_like, and PRX-like) according to their domain structures. The catalytic motifs of the cyanobacterial PRXs were similar to those of eukaryotic PRXs and highly conserved in all but the PRX-like subfamily. Classical motif (CXXC) of thioredoxin was detected in protein sequences from the PRX-like subfamily. Phylogenetic tree constructed of catalytic domains coincided well with the domain structures of PRXs and the phylogenies based on 16s rRNA. Conclusions The distribution of genes encoding PRXs in different unicellular and filamentous cyanobacteria especially those sub-families like PRX-like or 1-Cys PRX correlate with the genome size, eco-physiology, and physiological properties of the organisms. Cyanobacterial and eukaryotic PRXs share similar conserved motifs, indicating that cyanobacteria adopt similar catalytic mechanisms as eukaryotes. All

  17. Regulators of gene expression in Enteric Neural Crest Cells are putative Hirschsprung disease genes

    NARCIS (Netherlands)

    Schriemer, Duco; Sribudiani, Yunia; IJpma, Arne; Natarajan, Dipa; MacKenzie, Katherine C.; Metzger, Marco; Binder, Ellen; Burns, Alan J.; Thapar, Nikhil; Hofstra, Robert M. W.; Eggen, Bart J. L.

    2016-01-01

    The enteric nervous system (ENS) is required for peristalsis of the gut and is derived from Enteric Neural Crest Cells (ENCCs). During ENS development, the RET receptor tyrosine kinase plays a critical role in the proliferation and survival of ENCCs, their migration along the developing gut, and dif

  18. Regulators of gene expression in Enteric Neural Crest Cells are putative Hirschsprung disease genes

    NARCIS (Netherlands)

    Schriemer, Duco; Sribudiani, Yunia; IJpma, Arne; Natarajan, Dipa; MacKenzie, Katherine C; Metzger, Marco; Binder, Ellen; Burns, Alan J; Thapar, Nikhil; Hofstra, Robert M W; Eggen, Bart J L

    2016-01-01

    The enteric nervous system (ENS) is required for peristalsis of the gut and is derived from enteric neural crest cells (ENCCs). During ENS development, the RET receptor tyrosine kinase plays a critical role in the proliferation and survival of ENCCs, their migration along the developing gut, and dif

  19. A comparative study on efficiency of adult fibroblast, putative embryonic stem cell and lymphocyte as donor cells for production of handmade cloned embryos in goat and characterization of putative ntES cells obtained from these embryos.

    Science.gov (United States)

    Dutta, Rahul; Malakar, Dhruba; Khate, Keviletsu; Sahu, Shailendra; Akshey, Yogesh; Mukesh, Manishi

    2011-09-15

    The main purpose of the experiment was to compare the efficiency of three cell types, namely adult fibroblast, putative embryonic stem (ES) cell, and lymphocyte, as donor cells for somatic cell nuclear transfer by handmade cloning in goats. The outcome clearly shows that putative embryonic stem cells, with a cleavage and blastocyst production rate of 74.69% ± 3.92 and 39.75% ± 3.86, respectively, performs better in comparison to adult fibroblast cell and lymphocyte. Between adult fibroblast cell and lymphocyte no statistically significant difference exists at P II DRB genes of cloned embryos and three donor cells were performed to verify the cloned embryos. The amplified PCR products were subjected to SSCP to confirm their genetic identity. The karyotyping of the cloned embryos showed normal chromosomal status as expected in goat. Significantly, in the second stage of the experiment, the produced cloned embryos were successfully used to derive ntES-like cells. The rate of primary colony formation rate was 62.50% ± 4.62 for fibroblast donor cell derived embryos. The same was 60.60% ± 4.62 for putative ES donor cell derived embryos and 66.66% ± 4.62 for lymphocyte donor cell derived embryos, respectively. The putative ntES colonies were positively characterized for alkaline phosphatase, Oct-4, TRA-1-60, TRA-1-81, Sox-2, and Nanog by Immunocytochemistry and Reverse Transcription PCR. To further validate the stem ness, the produced putative ntES colonies were differentiated to embryoid bodies. Immunocytochemistry revealed that embryoid bodies expressed NESTIN specific for ectodermal lineage; GATA-4 for endodermal lineage and smooth muscle actin-I, and troponin-I specific for mesodermal lineage. The study has established an efficient protocol for putative ntES cell derivation from HMC embryos. It could be of substantial significance as patient specific ntES cells have proven therapeutic significance.

  20. Putative functions of extracellular matrix glycoproteins in secondary palate morphogenesis

    Science.gov (United States)

    d'Amaro, Rocca; Scheidegger, Rolf; Blumer, Susan; Pazera, Pawel; Katsaros, Christos; Graf, Daniel; Chiquet, Matthias

    2012-01-01

    Cleft palate is a common birth defect in humans. Elevation and fusion of paired palatal shelves are coordinated by growth and transcription factors, and mutations in these can cause malformations. Among the effector genes for growth factor signaling are extracellular matrix (ECM) glycoproteins. These provide substrates for cell adhesion (e.g., fibronectin, tenascins), but also regulate growth factor availability (e.g., fibrillins). Cleft palate in Bmp7 null mouse embryos is caused by a delay in palatal shelf elevation. In contrast, palatal shelves of Tgf-β3 knockout mice elevate normally, but a cleft develops due to their failure to fuse. However, nothing is known about a possible functional interaction between specific ECM proteins and Tgf-β/Bmp family members in palatogenesis. To start addressing this question, we studied the mRNA and protein distribution of relevant ECM components during secondary palate development, and compared it to growth factor expression in wildtypewild type and mutant mice. We found that fibrillin-2 (but not fibrillin-1) mRNA appeared in the mesenchyme of elevated palatal shelves adjacent to the midline epithelial cells, which were positive for Tgf-β3 mRNA. Moreover, midline epithelial cells started expressing fibronectin upon contact of the two palatal shelves. These findings support the hypothesis that fibrillin-2 and fibronectin are involved in regulating the activity of Tgf-β3 at the fusing midline. In addition, we observed that tenascin-W (but not tenascin-C) was misexpressed in palatal shelves of Bmp7-deficient mouse embryos. In contrast to tenascin-C, tenascin-W secretion was strongly induced by Bmp7 in embryonic cranial fibroblasts in vitro. These results are consistent with a putative function for tenascin-W as a target of Bmp7 signaling during palate elevation. Our results indicate that distinct ECM proteins are important for morphogenesis of the secondary palate, both as downstream effectors and as regulators of Tgf

  1. A Combined Computational and Experimental Study on the Structure-Regulation Relationships of Putative Mammalian DNA Replication Initiator GINS

    Institute of Scientific and Technical Information of China (English)

    Reiko Hayashi; Takako Arauchi; Moe Tategu; Yuya Goto; Kenichi Yoshida

    2006-01-01

    GINS, a heterotetramer of SLD5, PSF1, PSF2, and PSF3 proteins, is an emerging chromatin factor recognized to be involved in the initiation and elongation step of DNA replication. Although the yeast and Xenopus GINS genes are well documented, their orthologous genes in higher eukaryotes are not fully characterized.In this study, we report the genomic structure and transcriptional regulation of mammalian GINS genes. Serum stimulation increased the GINS Mrna levels in human cells. Reporter gene assay using putative GINS promoter sequences revealed that the expression of mammalian GINS is regulated by 17β-Estradiolstimulated estrogen receptor α, and human PSF3 acts as a gene responsive to transcription factor E2F1. The goal of this study is to present the current data so as to encourage further work in the field of GINS gene regulation and functions in mammalian cells.

  2. Identification of putative rhamnogalacturonan-II specific glycosyltransferases in Arabidopsis using a combination of bioinformatics approaches.

    Science.gov (United States)

    Voxeur, Aline; André, Aurélie; Breton, Christelle; Lerouge, Patrice

    2012-01-01

    Rhamnogalacturonan-II (RG-II) is a complex plant cell wall polysaccharide that is composed of an α(1,4)-linked homogalacturonan backbone substituted with four side chains. It exists in the cell wall in the form of a dimer that is cross-linked by a borate di-ester. Despite its highly complex structure, RG-II is evolutionarily conserved in the plant kingdom suggesting that this polymer has fundamental functions in the primary wall organisation. In this study, we have set up a bioinformatics strategy aimed at identifying putative glycosyltransferases (GTs) involved in RG-II biosynthesis. This strategy is based on the selection of candidate genes encoding type II membrane proteins that are tightly coexpressed in both rice and Arabidopsis with previously characterised genes encoding enzymes involved in the synthesis of RG-II and exhibiting an up-regulation upon isoxaben treatment. This study results in the final selection of 26 putative Arabidopsis GTs, including 10 sequences already classified in the CAZy database. Among these CAZy sequences, the screening protocol allowed the selection of α-galacturonosyltransferases involved in the synthesis of α4-GalA oligogalacturonides present in both homogalacturonans and RG-II, and two sialyltransferase-like sequences previously proposed to be involved in the transfer of Kdo and/or Dha on the pectic backbone of RG-II. In addition, 16 non-CAZy GT sequences were retrieved in the present study. Four of them exhibited a GT-A fold. The remaining sequences harbored a GT-B like fold and a fucosyltransferase signature. Based on homologies with glycosyltransferases of known functions, putative roles in the RG-II biosynthesis are proposed for some GT candidates.

  3. Dengue Virus Infection of Aedes aegypti Requires a Putative Cysteine Rich Venom Protein.

    Directory of Open Access Journals (Sweden)

    Berlin Londono-Renteria

    2015-10-01

    Full Text Available Dengue virus (DENV is a mosquito-borne flavivirus that causes serious human disease and mortality worldwide. There is no specific antiviral therapy or vaccine for DENV infection. Alterations in gene expression during DENV infection of the mosquito and the impact of these changes on virus infection are important events to investigate in hopes of creating new treatments and vaccines. We previously identified 203 genes that were ≥5-fold differentially upregulated during flavivirus infection of the mosquito. Here, we examined the impact of silencing 100 of the most highly upregulated gene targets on DENV infection in its mosquito vector. We identified 20 genes that reduced DENV infection by at least 60% when silenced. We focused on one gene, a putative cysteine rich venom protein (SeqID AAEL000379; CRVP379, whose silencing significantly reduced DENV infection in Aedes aegypti cells. Here, we examine the requirement for CRVP379 during DENV infection of the mosquito and investigate the mechanisms surrounding this phenomenon. We also show that blocking CRVP379 protein with either RNAi or specific antisera inhibits DENV infection in Aedes aegypti. This work identifies a novel mosquito gene target for controlling DENV infection in mosquitoes that may also be used to develop broad preventative and therapeutic measures for multiple flaviviruses.

  4. Dengue Virus Infection of Aedes aegypti Requires a Putative Cysteine Rich Venom Protein.

    Science.gov (United States)

    Londono-Renteria, Berlin; Troupin, Andrea; Conway, Michael J; Vesely, Diana; Ledizet, Michael; Roundy, Christopher M; Cloherty, Erin; Jameson, Samuel; Vanlandingham, Dana; Higgs, Stephen; Fikrig, Erol; Colpitts, Tonya M

    2015-10-01

    Dengue virus (DENV) is a mosquito-borne flavivirus that causes serious human disease and mortality worldwide. There is no specific antiviral therapy or vaccine for DENV infection. Alterations in gene expression during DENV infection of the mosquito and the impact of these changes on virus infection are important events to investigate in hopes of creating new treatments and vaccines. We previously identified 203 genes that were ≥5-fold differentially upregulated during flavivirus infection of the mosquito. Here, we examined the impact of silencing 100 of the most highly upregulated gene targets on DENV infection in its mosquito vector. We identified 20 genes that reduced DENV infection by at least 60% when silenced. We focused on one gene, a putative cysteine rich venom protein (SeqID AAEL000379; CRVP379), whose silencing significantly reduced DENV infection in Aedes aegypti cells. Here, we examine the requirement for CRVP379 during DENV infection of the mosquito and investigate the mechanisms surrounding this phenomenon. We also show that blocking CRVP379 protein with either RNAi or specific antisera inhibits DENV infection in Aedes aegypti. This work identifies a novel mosquito gene target for controlling DENV infection in mosquitoes that may also be used to develop broad preventative and therapeutic measures for multiple flaviviruses.

  5. Glycosyltransferases and oligosaccharyltransferases in Archaea: putative components of the N-glycosylation pathway in the third domain of life.

    Science.gov (United States)

    Magidovich, Hilla; Eichler, Jerry

    2009-11-01

    The ability of Eukarya, Bacteria and Archaea to perform N-glycosylation underlies the importance and possible antiquity of this post-translational protein modification. However, in contrast to the relatively well-studied eukaryal and bacterial pathways, the archaeal N-glycosylation process is less understood. To remedy this disparity, the following study has examined 56 available archaeal genomes with the aim of identifying glycosyltransferases and oligosaccharyltransferases, including those putatively catalyzing this post-translational processing event. This analysis reveals that while oligosaccharyltransferases, central components of the N-glycosylation pathway, are found across the range of archaeal phenotypes, the N-glycosylation machinery of hyperthermophilic Archaea may well rely on fewer components than do the parallel systems of nonhyperthermophilic Archaea. Moreover, genes encoding predicted glycosyltransferases of hyperthermophilic Archaea tend to be far more scattered within the genome than is the case with nonhyperthermophilic species, where putative glycosyltransferase genes are often clustered around identified oligosaccharyltransferase-encoding sequences.

  6. Transcriptome-Based Examination of Putative Pollen Allergens of Rice(Oryza sativa ssp.japonica)

    Institute of Scientific and Technical Information of China (English)

    Scott D.Russell; Prem L Bhalla; Mohan B.Singh

    2008-01-01

    Pollen allergens are among the most abundantly transcribed and translated products in the Iife history of plants,and particularly grasses.To identify different pollen allergens in rice,putative allergens were identified in the rice genome and their expression characterized using the Affymetrix 57K rice GeneChip microarray.Among the most abundant pollen-specific candidate transcripts were Ory s 1 beta-expansin.Ory s 2,Ory s 7 EFhand,Ory s 11,Ory s 12 profilin A,Ory s 23,glycosyl hydrolase family 28(polygalacturonase).and FAD binding proteins.Highly expressed pollen proteins are frequently present in multiple copy numbers,sometimes with mirror images Iocated on nearby regions of the opposite DNA strand.Many of these are intronless and inserted as copies that retain nearly exact copies of their regulatory elements.Ory s 23 reflects low variability and high copy number,suggesting recent gene amplification.Some copies contain pseudogenes,which may reflect their origin through activity of retrotransposition;some putative allergenic sequences bear fusion products with repeat sequences of transposable elements(LTRs).The abundance of nearby repetitive sequences,activation of transposable elements.and high production of mRNA transcripts appear to coincide in pollen and may contribute to a syndrome in which highly transcribed proteins may be copied and inserted with streamlined features for translation,including grouping and removaI of introns.

  7. Identification of calcium-transporting ATPases of Entamoeba histolytica and cellular localization of the putative SERCA.

    Science.gov (United States)

    Martinez-Higuera, Aarón; Salas-Casas, Andrés; Calixto-Gálvez, Mercedes; Chávez-Munguía, Bibiana; Pérez-Ishiwara, D Guillermo; Ximénez, Cecilia; Rodríguez, Mario A

    2013-09-01

    Calcium has an important role on signaling of different cellular processes in the protozoa parasite Entamoeba histolytica, including development and pathogenesis. However, the systems that control calcium responses in this parasite are incompletely understood. Calcium-ATPases (Ca(2+)-ATPases) are proteins that play an important role in calcium homeostasis by catalyzing the active efflux of this ion from cytoplasm and are essential to the correct functioning of the cell machinery. Here, we reported the identification of five E. histolytica genes encoding putative Ca(2+)-ATPases, three related to PMCA, and two related to organellar ATPases. RT-PCR assays showed that all those genes are expressed in trophozoites and specific antibodies against the SERCA-like member located this protein in a continuous cytoplasmic network, supporting the hypothesis that it corresponds to the Ca(2+)-ATPase responsible to sequester calcium in the endoplasmic reticulum of this parasite.

  8. A putative viral defence mechanism in archaeal cells

    DEFF Research Database (Denmark)

    Lillestøl, Reidun K; Redder, Peter; Garrett, Roger Antony

    2006-01-01

    in cells, and that both the mode of inhibition of viral propagation and the mechanism of adding spacer-repeat units to clusters, are dependent on RNAs transcribed from the clusters. Moreover, the putative inhibitory apparatus (piRNA-based) may be evolutionarily related to the interference RNA systems (si...

  9. Putative Lineage of Novel African Usutu Virus, Central Europe

    Centers for Disease Control (CDC) Podcasts

    2015-10-15

    Sarah Gregory reads an abridged version of "Putative Lineage of Novel African Usutu Virus, Central Europe.".  Created: 10/15/2015 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 10/15/2015.

  10. Putative golden proportions as predictors of facial esthetics in adolescents.

    NARCIS (Netherlands)

    Kiekens, R.M.A.; Kuijpers-Jagtman, A.M.; Hof, M.A. van 't; Hof, BE van 't; Maltha, J.C.

    2008-01-01

    INTRODUCTION: In orthodontics, facial esthetics is assumed to be related to golden proportions apparent in the ideal human face. The aim of the study was to analyze the putative relationship between facial esthetics and golden proportions in white adolescents. METHODS: Seventy-six adult laypeople

  11. KLONING GEN PUTATIVE CLEAVAGE PROTEIN 1 (PCP-1 PADA UDANG VANAME (Litopenaeus vannamei YANG TERSERANG INFECTIOUS MYONECROSIS VIRUS

    Directory of Open Access Journals (Sweden)

    Hessy Novita

    2016-12-01

    Full Text Available Penanggulangan penyakit ikan dapat dilakukan dengan cara meningkatkan kekebalan tubuh ikan melalui program vaksinasi. Namun vaksinasi tidak tepat untuk udang, karena udang tidak mempunyai immunological memory seperti ikan. Oleh karena itu, perlindungan udang terhadap serangan penyakit viral dengan menggunakan RNA interference (RNAi. Teknologi RNAi digunakan untuk menghalangi (interfere proses replikasi infectious myonecrosis virus (IMNV pada udang vaname dengan cara menon-aktifkan gen putative cleavage protein 1 (PCP-1, yang berfungsi dalam pembentukan capsid dan proses transkripsi RNA IMNV. Penelitian ini bertujuan untuk melakukan kloning gen putative cleavage protein 1 dalam rangka perakitan teknologi RNAi untuk pengendalian penyakit IMNV pada udang vaname. Tahapan penelitian meliputi koleksi sampel, isolasi RNA, sintesis cDNA, amplifikasi PCR, purifikasi DNA, transformasi, isolasi plasmid, serta sekuensing dan analisis data. Hasil isolasi plasmid cDNA PCP-1 memperlihatkan semua koloni bakteri terseleksi ternyata membawa plasmid hasil insersi DNA gen PCP–1, hasil sekuen dengan nilai homologinya mencapai 100% dan 99% yang dibandingkan dengan sekuen di Genebank. Hasil penelitian menunjukkan bahwa kloning gen putative cleavage protein 1 (PCP-1 dari udang vaname yang terserang Infectious Myonecrosis Virus berhasil dikloning yang nantinya digunakan untuk perakitan RNAi. The prevention of fish diseases can be done by increasing of the fish immune through vaccination programs. However, the vaccination can not be done for the shrimp,due to the absence of  immunological memory. Therefore, the protection of shrimp against viral diseases was done by using of RNA interference (RNAi. RNAi technology is used to interfere infectious myonecrosis virus (IMNV replication process on white shrimp by disabling of putative cleavage protein 1 (PCP-1gene, which functions in capsid formation and RNA transcription process. The study was conducted to perform putative

  12. Identification and temporal expression of putative circadian clock transcripts in the amphipod crustacean Talitrus saltator.

    Science.gov (United States)

    O'Grady, Joseph F; Hoelters, Laura S; Swain, Martin T; Wilcockson, David C

    2016-01-01

    Talitrus saltator is an amphipod crustacean that inhabits the supralittoral zone on sandy beaches in the Northeast Atlantic and Mediterranean. T. saltator exhibits endogenous locomotor activity rhythms and time-compensated sun and moon orientation, both of which necessitate at least one chronometric mechanism. Whilst their behaviour is well studied, currently there are no descriptions of the underlying molecular components of a biological clock in this animal, and very few in other crustacean species. We harvested brain tissue from animals expressing robust circadian activity rhythms and used homology cloning and Illumina RNAseq approaches to sequence and identify the core circadian clock and clock-related genes in these samples. We assessed the temporal expression of these genes in time-course samples from rhythmic animals using RNAseq. We identified a comprehensive suite of circadian clock gene homologues in T. saltator including the 'core' clock genes period (Talper), cryptochrome 2 (Talcry2), timeless (Taltim), clock (Talclk), and bmal1 (Talbmal1). In addition we describe the sequence and putative structures of 23 clock-associated genes including two unusual, extended isoforms of pigment dispersing hormone (Talpdh). We examined time-course RNAseq expression data, derived from tissues harvested from behaviourally rhythmic animals, to reveal rhythmic expression of these genes with approximately circadian period in Talper and Talbmal1. Of the clock-related genes, casein kinase IIβ (TalckIIβ), ebony (Talebony), jetlag (Taljetlag), pigment dispensing hormone (Talpdh), protein phosphatase 1 (Talpp1), shaggy (Talshaggy), sirt1 (Talsirt1), sirt7 (Talsirt7) and supernumerary limbs (Talslimb) show temporal changes in expression. We report the sequences of principle genes that comprise the circadian clock of T. saltator and highlight the conserved structural and functional domains of their deduced cognate proteins. Our sequencing data contribute to the growing inventory

  13. Evidence for positive selection in putative virulence factors within the Paracoccidioides brasiliensis species complex.

    Directory of Open Access Journals (Sweden)

    Daniel R Matute

    Full Text Available Paracoccidioides brasiliensis is a dimorphic fungus that is the causative agent of paracoccidioidomycosis, the most important prevalent systemic mycosis in Latin America. Recently, the existence of three genetically isolated groups in P. brasiliensis was demonstrated, enabling comparative studies of molecular evolution among P. brasiliensis lineages. Thirty-two gene sequences coding for putative virulence factors were analyzed to determine whether they were under positive selection. Our maximum likelihood-based approach yielded evidence for selection in 12 genes that are involved in different cellular processes. An in-depth analysis of four of these genes showed them to be either antigenic or involved in pathogenesis. Here, we present evidence indicating that several replacement mutations in gp43 are under positive balancing selection. The other three genes (fks, cdc42 and p27 show very little variation among the P. brasiliensis lineages and appear to be under positive directional selection. Our results are consistent with the more general observations that selective constraints are variable across the genome, and that even in the genes under positive selection, only a few sites are altered. We present our results within an evolutionary framework that may be applicable for studying adaptation and pathogenesis in P. brasiliensis and other pathogenic fungi.

  14. Coral bleaching under thermal stress: putative involvement of host/symbiont recognition mechanisms

    Directory of Open Access Journals (Sweden)

    Tambutte Sylvie

    2009-08-01

    Full Text Available Abstract Background Coral bleaching can be defined as the loss of symbiotic zooxanthellae and/or their photosynthetic pigments from their cnidarian host. This major disturbance of reef ecosystems is principally induced by increases in water temperature. Since the beginning of the 1980s and the onset of global climate change, this phenomenon has been occurring at increasing rates and scales, and with increasing severity. Several studies have been undertaken in the last few years to better understand the cellular and molecular mechanisms of coral bleaching but the jigsaw puzzle is far from being complete, especially concerning the early events leading to symbiosis breakdown. The aim of the present study was to find molecular actors involved early in the mechanism leading to symbiosis collapse. Results In our experimental procedure, one set of Pocillopora damicornis nubbins was subjected to a gradual increase of water temperature from 28°C to 32°C over 15 days. A second control set kept at constant temperature (28°C. The differentially expressed mRNA between the stressed states (sampled just before the onset of bleaching and the non stressed states (control were isolated by Suppression Subtractive Hybridization. Transcription rates of the most interesting genes (considering their putative function were quantified by Q-RT-PCR, which revealed a significant decrease in transcription of two candidates six days before bleaching. RACE-PCR experiments showed that one of them (PdC-Lectin contained a C-Type-Lectin domain specific for mannose. Immunolocalisation demonstrated that this host gene mediates molecular interactions between the host and the symbionts suggesting a putative role in zooxanthellae acquisition and/or sequestration. The second gene corresponds to a gene putatively involved in calcification processes (Pdcyst-rich. Its down-regulation could reflect a trade-off mechanism leading to the arrest of the mineralization process under stress

  15. Two Genes Encoding Uracil Phosphoribosyltransferase Are Present in Bacillus subtilis

    DEFF Research Database (Denmark)

    Martinussen, Jan; Glaser, Philippe; Andersen, Paal S.

    1995-01-01

    Uracil phosphoribosyltransferase (UPRTase) catalyzes the key reaction in the salvage of uracil in many microorganisms. Surprisingly, two genes encoding UPRTase activity were cloned from Bacillus subtilis by complementation of an Escherichia coli mutant. The genes were sequenced, and the putative...

  16. Characterization of ERAS, a putative novel human oncogene, in skin and breast

    Energy Technology Data Exchange (ETDEWEB)

    Peña Avalos, B.L. de la

    2014-07-01

    Most human tumors have mutations in genes of the RAS small GTPase protein family. RAS works as a molecular switch for signaling pathways that modulate many aspects of cell behavior, including proliferation, differentiation, motility and death. Oncogenic mutations in RAS prevent GTP hydrolysis, locking RAS in a permanently active state, being the most common mutations in HRAS, KRAS and NRAS. The human RAS family consists of at least 36 different genes, many of which have been scarcely studied. One of these relatively unknown genes is ERAS (ES cell-expressed RAS), which is a constitutively active RAS protein, localized in chromosome X and expressed only in embryonic cells, being undetectable in adult tissues. New high throughput technologies have made it possible to screen complete cancer genomes for identification of mutations associated to cancer. Using the Sleeping Beauty (SB) transposon system, ERAS was identified as a putative novel oncogene in non-melanoma skin and breast cancers. The major aim of this project is to determine the general characteristics of ERAS as a putative novel human oncogene in skin and breast cells. Forced expression of ERAS results in drastic changes in cell shape, proliferation and motility. When ERAS is overexpressed in skin and breast human cells it is mainly localized in the cytoplasmic membrane. ERAS activates the phosphatidylinositol-3-OH kinase (PI3K) pathway but not the mitogen-activated protein kinase (MAPK) pathway. ERAS-expressing cells suffer spontaneous morphologic and phenotypic EMT-like changes, including cytoskeleton reorganization, vimentin and N-cadherin up-regulation and down-regulation of E-cadherin, which can be associated with increased malignancy, and invasive and metastatic potential. Our results suggest that inappropriate expression of ERAS lead to transformation of human cells. (Author)

  17. Putative melatonin receptors in a human biological clock

    Energy Technology Data Exchange (ETDEWEB)

    Reppert, S.M.; Weaver, D.R.; Rivkees, S.A.; Stopa, E.G.

    1988-10-07

    In vitro autoradiography with /sup 125/I-labeled melatonin was used to examine melatonin binding sites in human hypothalamus. Specific /sup 125/I-labeled melatonin binding was localized to the suprachiasmatic nuclei, the site of a putative biological clock, and was not apparent in other hypothalamic regions. Specific /sup 125/I-labeled melatonin binding was consistently found in the suprachiasmatic nuclei of hypothalami from adults and fetuses. Densitometric analysis of competition experiments with varying concentrations of melatonin showed monophasic competition curves, with comparable half-maximal inhibition values for the suprachiasmatic nuclei of adults (150 picomolar) and fetuses (110 picomolar). Micromolar concentrations of the melatonin agonist 6-chloromelatonin completely inhibited specific /sup 125/I-labeled melatonin binding, whereas the same concentrations of serotonin and norepinephrine caused only a partial reduction in specific binding. The results suggest that putative melatonin receptors are located in a human biological clock.

  18. Cloning of partial putative gonadotropin hormone receptor sequence from fish

    Indian Academy of Sciences (India)

    G Kumaresan; T Venugopal; A Vikas; T J Pandian; S M Athavan

    2000-03-01

    A search for the presence of mariner-like elements in the Labeo rohita genome by polymerase chain reaction led to the amplification of a partial DNA sequence coding for a putative transmembrane domain of gonadotropin hormone receptor. The amplified DNA sequence shows a high degree of homology to the available turkey and human luteinizing and follicle stimulating hormone receptor coding sequences. This is the first report on cloning such sequences of piscine origin.

  19. A putative role for apelin in the etiology of obesity.

    Science.gov (United States)

    Rayalam, Srujana; Della-Fera, Mary Anne; Krieg, Paul A; Cox, Christopher M; Robins, Allan; Baile, Clifton A

    2008-04-11

    Apelin, the endogenous ligand of the G protein-coupled APJ receptor has been shown to promote tumor angiogenesis. However, the effect of apelin on inducing angiogenesis in adipose tissue has not been investigated. In this review, we propose a putative role for apelin in promoting angiogenesis in adipose tissue. We further propose that targeting adipose tissue vasculature by blocking apelin signaling with anti-apelin antibodies will lead not only to inhibition of angiogenesis in adipose tissue but also to decreased adiposity.

  20. Functional Characterization of PaLAX1, a Putative Auxin Permease, in Heterologous Plant Systems1[W][OA

    Science.gov (United States)

    Hoyerová, Klára; Perry, Lucie; Hand, Paul; Laňková, Martina; Kocábek, Tomáš; May, Sean; Kottová, Jana; Pačes, Jan; Napier, Richard; Zažímalová, Eva

    2008-01-01

    We have isolated the cDNA of the gene PaLAX1 from a wild cherry tree (Prunus avium). The gene and its product are highly similar in sequences to both the cDNAs and the corresponding protein products of AUX/LAX-type genes, coding for putative auxin influx carriers. We have prepared and characterized transformed Nicotiana tabacum and Arabidopsis thaliana plants carrying the gene PaLAX1. We have proved that constitutive overexpression of PaLAX1 is accompanied by changes in the content and distribution of free indole-3-acetic acid, the major endogenous auxin. The increase in free indole-3-acetic acid content in transgenic plants resulted in various phenotype changes, typical for the auxin-overproducing plants. The uptake of synthetic auxin, 2,4-dichlorophenoxyacetic acid, was 3 times higher in transgenic lines compared to the wild-type lines and the treatment with the auxin uptake inhibitor 1-naphthoxyacetic acid reverted the changes caused by the expression of PaLAX1. Moreover, the agravitropic response could be restored by expression of PaLAX1 in the mutant aux1 plants, which are deficient in auxin influx carrier activity. Based on our data, we have concluded that the product of the gene PaLAX1 promotes the uptake of auxin into cells, and, as a putative auxin influx carrier, it affects the content and distribution of free endogenous auxin in transgenic plants. PMID:18184737

  1. Identification of putative insulin-like peptides and components of insulin signaling pathways in parasitic platyhelminths by the use of genome-wide screening.

    Science.gov (United States)

    Wang, Shuai; Luo, Xuenong; Zhang, Shaohua; Yin, Cai; Dou, Yongxi; Cai, Xuepeng

    2014-02-01

    No endogenous insulin-like peptides in parasitic flatworms have been reported. Insulin receptors from flukes and tapeworms have been shown to interact directly with the host-derived insulin molecule, which suggests the exploitation of host-derived insulin. In this study, a strategy of genome-wide searches followed by comprehensive analyses of strictly conserved features of the insulin family was used to demonstrate the presence of putative insulin-like peptides in the genomes of six tapeworms and two flukes. In addition, whole insulin signaling pathways were annotated on a genome-wide scale. Two putative insulin-like peptide genes in each genome of tapeworms and one insulin-like peptide gene in each genome of flukes were identified. The comprehensive analyses revealed that all of these peptides showed the common features shared by other members of the insulin family, and the phylogenetic analysis implied a putative gene duplication event in the Cestoda during the evolution of insulin-like peptide genes. The quantitative expression analysis and immunolocalization results suggested a putative role of these peptides in reproduction. Entire sets of major components of the classic insulin signaling pathway were successfully identified, suggesting that this pathway in parasitic flatworms might also regulate many other important biological activities. We believe that the identification of the insulin-like peptides gives us a better understanding of the insulin signaling pathway in these parasites, as well as host-parasite interactions.

  2. The genome of tolypocladium inflatum: evolution, organization, and expression of the cyclosporin biosynthetic gene cluster.

    Science.gov (United States)

    Bushley, Kathryn E; Raja, Rajani; Jaiswal, Pankaj; Cumbie, Jason S; Nonogaki, Mariko; Boyd, Alexander E; Owensby, C Alisha; Knaus, Brian J; Elser, Justin; Miller, Daniel; Di, Yanming; McPhail, Kerry L; Spatafora, Joseph W

    2013-06-01

    The ascomycete fungus Tolypocladium inflatum, a pathogen of beetle larvae, is best known as the producer of the immunosuppressant drug cyclosporin. The draft genome of T. inflatum strain NRRL 8044 (ATCC 34921), the isolate from which cyclosporin was first isolated, is presented along with comparative analyses of the biosynthesis of cyclosporin and other secondary metabolites in T. inflatum and related taxa. Phylogenomic analyses reveal previously undetected and complex patterns of homology between the nonribosomal peptide synthetase (NRPS) that encodes for cyclosporin synthetase (simA) and those of other secondary metabolites with activities against insects (e.g., beauvericin, destruxins, etc.), and demonstrate the roles of module duplication and gene fusion in diversification of NRPSs. The secondary metabolite gene cluster responsible for cyclosporin biosynthesis is described. In addition to genes necessary for cyclosporin biosynthesis, it harbors a gene for a cyclophilin, which is a member of a family of immunophilins known to bind cyclosporin. Comparative analyses support a lineage specific origin of the cyclosporin gene cluster rather than horizontal gene transfer from bacteria or other fungi. RNA-Seq transcriptome analyses in a cyclosporin-inducing medium delineate the boundaries of the cyclosporin cluster and reveal high levels of expression of the gene cluster cyclophilin. In medium containing insect hemolymph, weaker but significant upregulation of several genes within the cyclosporin cluster, including the highly expressed cyclophilin gene, was observed. T. inflatum also represents the first reference draft genome of Ophiocordycipitaceae, a third family of insect pathogenic fungi within the fungal order Hypocreales, and supports parallel and qualitatively distinct radiations of insect pathogens. The T. inflatum genome provides additional insight into the evolution and biosynthesis of cyclosporin and lays a foundation for further investigations of the role

  3. KLIKK proteases of Tannerella forsythia: putative virulence factors with a unique domain structure

    Science.gov (United States)

    Ksiazek, Miroslaw; Mizgalska, Danuta; Eick, Sigrum; Thøgersen, Ida B.; Enghild, Jan J.; Potempa, Jan

    2015-01-01

    Comparative genomics of virulent Tannerella forsythia ATCC 43037 and a close health-associated relative, Tannerella BU063, revealed, in the latter, the absence of an entire array of genes encoding putative secretory proteases that possess a nearly identical C-terminal domain (CTD) that ends with a -Lys-Leu-Ile-Lys-Lys motif. This observation suggests that these proteins, referred to as KLIKK proteases, may function as virulence factors. Re-sequencing of the loci of the KLIKK proteases found only six genes grouped in two clusters. All six genes were expressed by T. forsythia in routine culture conditions, although at different levels. More importantly, a transcript of each gene was detected in gingival crevicular fluid (GCF) from periodontitis sites infected with T. forsythia indicating that the proteases are expressed in vivo. In each protein, a protease domain was flanked by a unique N-terminal profragment and a C-terminal extension ending with the CTD. Partially purified recombinant proteases showed variable levels of proteolytic activity in zymography gels and toward protein substrates, including collagen, gelatin, elastin, and casein. Taken together, these results indicate that the pathogenic strain of T. forsythia secretes active proteases capable of degrading an array of host proteins, which likely represents an important pathogenic feature of this bacterium. PMID:25954253

  4. KLIKK proteases of Tannerella forsythia: putative virulence factors with a unique domain structure

    Directory of Open Access Journals (Sweden)

    Miroslaw eKsiazek

    2015-04-01

    Full Text Available Comparative genomics of virulent Tannerella forsythia ATCC 43037 and a close health-associated relative, Tannerella BU063, revealed, in the latter, the absence of an entire array of genes encoding putative secretory proteases that possess a nearly identical C-terminal domain (CTD that ends with a -Lys-Leu-Ile-Lys-Lys motif. This observation suggests that these proteins, referred to as KLIKK proteases, may function as virulence factors. Re-sequencing of the loci of the KLIKK proteases found only six genes grouped in two clusters. All six genes were expressed by T. forsythia in routine culture conditions, although at different levels. More importantly, a transcript of each gene was detected in gingival crevicular fluid (GCF from periodontitis sites infected with T. forsythia indicating that the proteases are expressed in vivo. In each protein, a protease domain was flanked by a unique N-terminal profragment and a C-terminal extension ending with the CTD. Partially purified recombinant proteases showed variable levels of proteolytic activity in zymography gels and toward protein substrates, including collagen, gelatin, elastin, and casein. Taken together, these results indicate that the pathogenic strain of T. forsythia secretes active proteases capable of degrading an array of host proteins, which likely represents an important pathogenic feature of this bacterium.

  5. KLIKK proteases of Tannerella forsythia: putative virulence factors with a unique domain structure.

    Science.gov (United States)

    Ksiazek, Miroslaw; Mizgalska, Danuta; Eick, Sigrum; Thøgersen, Ida B; Enghild, Jan J; Potempa, Jan

    2015-01-01

    Comparative genomics of virulent Tannerella forsythia ATCC 43037 and a close health-associated relative, Tannerella BU063, revealed, in the latter, the absence of an entire array of genes encoding putative secretory proteases that possess a nearly identical C-terminal domain (CTD) that ends with a -Lys-Leu-Ile-Lys-Lys motif. This observation suggests that these proteins, referred to as KLIKK proteases, may function as virulence factors. Re-sequencing of the loci of the KLIKK proteases found only six genes grouped in two clusters. All six genes were expressed by T. forsythia in routine culture conditions, although at different levels. More importantly, a transcript of each gene was detected in gingival crevicular fluid (GCF) from periodontitis sites infected with T. forsythia indicating that the proteases are expressed in vivo. In each protein, a protease domain was flanked by a unique N-terminal profragment and a C-terminal extension ending with the CTD. Partially purified recombinant proteases showed variable levels of proteolytic activity in zymography gels and toward protein substrates, including collagen, gelatin, elastin, and casein. Taken together, these results indicate that the pathogenic strain of T. forsythia secretes active proteases capable of degrading an array of host proteins, which likely represents an important pathogenic feature of this bacterium.

  6. Evolution and structural diversification of PILS putative auxin carriers in plants

    Directory of Open Access Journals (Sweden)

    Elena eFeraru

    2012-10-01

    Full Text Available The phytohormone auxin contributes to virtually every aspect of the plant development. The spatiotemporal distribution of auxin depends on a complex interplay between auxin metabolism and intercellular auxin transport. Intracellular auxin compartmentalization provides another link between auxin transport processes and auxin metabolism. The PIN-LIKES (PILS putative auxin carriers localize to the endoplasmic reticulum (ER and contribute to cellular auxin homeostasis. PILS proteins regulate intracellular auxin accumulation, the rate of auxin conjugation and subsequently, affect nuclear auxin signalling. Here, we investigate sequence diversification of the PILS family in Arabidopsis thaliana and provide insights into the evolution of these novel putative auxin carriers in plants. Our data suggest that PILS proteins are conserved throughout the plant lineage and expanded during higher plant evolution. PILS proteins diversified early during plant evolution into three clades. Besides the ancient Clade I encompassing non-land plant species, PILS proteins evolved into two clades. The diversification of Clade II and Clade III occurred already at the level of non-vascular plant evolution and, hence, both clades contain vascular and non-vascular plant species. Nevertheless, Clade III contains fewer non- and increased numbers of vascular plants, indicating higher importance of Clade III for vascular plant evolution. Notably, PILS proteins are distinct and appear evolutionarily older than the prominent PIN-FORMED auxin carriers. Moreover, we revealed particular PILS sequence divergence in Arabidopsis and assume that these alterations could contribute to distinct gene regulations and protein functions.

  7. Characterization of putative multidrug resistance transporters of the major facilitator-superfamily expressed in Salmonella Typhi.

    Science.gov (United States)

    Shaheen, Aqsa; Ismat, Fouzia; Iqbal, Mazhar; Haque, Abdul; De Zorzi, Rita; Mirza, Osman; Walz, Thomas; Rahman, Moazur

    2015-05-01

    Multidrug resistance mediated by efflux pumps is a well-known phenomenon in infectious bacteria. Although much work has been carried out to characterize multidrug efflux pumps in Gram-negative and Gram-positive bacteria, such information is still lacking for many deadly pathogens. The aim of this study was to gain insight into the substrate specificity of previously uncharacterized transporters of Salmonella Typhi to identify their role in the development of multidrug resistance. S. Typhi genes encoding putative members of the major facilitator superfamily were cloned and expressed in the drug-hypersensitive Escherichia coli strain KAM42, and tested for transport of 25 antibacterial compounds, including representative antibiotics of various classes, antiseptics, dyes and detergents. Of the 15 tested putative transporters, STY0901, STY2458 and STY4874 exhibited a drug-resistance phenotype. Among these, STY4874 conferred resistance to at least ten of the tested antimicrobials: ciprofloxacin, norfloxacin, levofloxacin, kanamycin, streptomycin, gentamycin, nalidixic acid, chloramphenicol, ethidium bromide, and acriflavine, including fluoroquinolone antibiotics, which were drugs of choice to treat S. Typhi infections. Cell-based functional studies using ethidium bromide and acriflavine showed that STY4874 functions as a H(+)-dependent exporter. These results suggest that STY4874 may be an important drug target, which can now be tested by studying the susceptibility of a STY4874-deficient S. Typhi strain to antimicrobials.

  8. Isolation and Identification of Putative Oral Cancer Stem Cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Min; ZHAO Yan-Hua; TANG Xiao-Fei

    2011-01-01

    Objective: To isolate and characterize putative cancer stem cells in Tea8113 oral squmous cell carcinoma cell line. Methods: Putative cancer stem cells were isolated by limited dilution assay in Tea8113 cell line. Biological features of putative cancer stem cells were detected by MTT assay, flow cytometry, immunofluorescence, Colony Forming Efficiency assays, cell motility assay and in vivo tumor formation experiment. Results: Compared with untreated Tea8113 cells, the putative cancer stem cells proliferated more quickly and showed heteroploid cell cycle,higher G0/G1-arrested cells, higher CFE and higher expression levels of ABCG2 belonged to tumor stem cell phenotypes. The putative cancer stem cells had stronger capacity to generate tumors in vivo. Conclusion: The holoclone cells have higher proliferation and self-renewal abilities, which may be cancer stem cells existed in Tea8113 oral squmous cell carcinoma cell line.%目的:分离鉴定口腔鳞癌细胞系Tca8113中的肿瘤干细胞.方法:利用有限稀释的方法分离Tca8113细胞系中的肿瘤干细胞.通过MTT法、流式细胞技术、细胞免疫荧光、克隆形成率分析、细胞迁移能力检测和裸鼠皮下成瘤实验确定分离得到的肿瘤干细胞的生物学特点.结果:分离得到的紧密型克隆肿瘤细胞表现为异倍体样细胞周期,大部分细胞处于G0/G1期,增殖能力、克隆形成率和体外迁移能力都明显高于未分离的肿瘤细胞.紧密型克隆肿瘤细胞肿瘤干细胞标记物ABCG2表达也高于未分离的肿瘤细胞,并且具有更强的裸鼠皮下成瘤能力.结论:我们分离得到的紧密型克隆细胞具有较强的细胞增殖和自我更新能力,可能就是口腔鳞癌细胞系Tca8113中的肿瘤干细胞.