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Sample records for affect gene transcription

  1. Land use type significantly affects microbial gene transcription in soil.

    PubMed

    Nacke, Heiko; Fischer, Christiane; Thürmer, Andrea; Meinicke, Peter; Daniel, Rolf

    2014-05-01

    Soil microorganisms play an essential role in sustaining biogeochemical processes and cycling of nutrients across different land use types. To gain insights into microbial gene transcription in forest and grassland soil, we isolated mRNA from 32 sampling sites. After sequencing of generated complementary DNA (cDNA), a total of 5,824,229 sequences could be further analyzed. We were able to assign nonribosomal cDNA sequences to all three domains of life. A dominance of bacterial sequences, which were affiliated to 25 different phyla, was found. Bacterial groups capable of aromatic compound degradation such as Phenylobacterium and Burkholderia were detected in significantly higher relative abundance in forest soil than in grassland soil. Accordingly, KEGG pathway categories related to degradation of aromatic ring-containing molecules (e.g., benzoate degradation) were identified in high abundance within forest soil-derived metatranscriptomic datasets. The impact of land use type forest on community composition and activity is evidently to a high degree caused by the presence of wood breakdown products. Correspondingly, bacterial groups known to be involved in lignin degradation and containing ligninolytic genes such as Burkholderia, Bradyrhizobium, and Azospirillum exhibited increased transcriptional activity in forest soil. Higher solar radiation in grassland presumably induced increased transcription of photosynthesis-related genes within this land use type. This is in accordance with high abundance of photosynthetic organisms and plant-infecting viruses in grassland. PMID:24553913

  2. Gene size differentially affects the binding of yeast transcription factor tau to two intragenic regions.

    PubMed Central

    Baker, R E; Camier, S; Sentenac, A; Hall, B D

    1987-01-01

    Yeast transcription factor tau (transcription factor IIIC) specifically interacts with tRNA genes, binding to both the A block and the B block elements of the internal promoter. To study the influence of A block-B block spacing, we analyzed the binding of purified tau protein to a series of internally deleted yeast tRNA(3Leu) genes with A and B blocks separated by 0 to 74 base pairs. Optimal binding occurred with genes having A block-B block distances of 30-60 base pairs; the relative helical orientation of the A and B blocks was unimportant. Results from DNase I "footprinting" and lambda exonuclease protection experiments were consistent with these findings and further revealed that changes in A block-B block distance primarily affect the ability of tau to interact with A block sequences; B block interactions are unaltered. When the A block-B block distance is 17 base pairs or less, tau interacts with a sequence located 15 base pairs upstream of the normal A block, and a new RNA initiation site is observed by in vitro transcription. We propose that the initial binding of tau to the B block activates transcription by enhancing its ability to bind at the A block, and that the A block interaction ultimately directs initiation by RNA polymerase III. Images PMID:2827154

  3. Analysis of Saccharomyces cerevisiae genome for the distributions of stress-response elements potentially affecting gene expression by transcriptional interference.

    PubMed

    Liu, Yunkai; Ye, Sujuan; Erkine, Alexandre M

    2009-01-01

    Cellular stress responses are characterized by coordinated transcriptional induction of genes encoding a group of conserved proteins known as molecular chaperones, most of which are also known as heat shock proteins (HSPs). In S. cerevisiae, transcriptional responses to stress are mediated via two trans-regulatory activators: heat shock transcription factors (HSFs) that bind to heat shock elements (HSEs), and the Msn2 and Msn4 transcription factors that bind to stress response elements (STREs). Recent studies in S. cerevisiae demonstrated that a significant portion of the non-coding region in the genome is transcribed and this intergenic transcription could regulate the transcription of adjacent genes by transcription interference. The goal of this study was to analyze the genomic distribution of HSF and Msn2/4 binding sites and to study the potential for transcription interference regulated by stress response systems. Our genome-wide analysis revealed that 297 genes have STREs in their promoter region, whereas 310 genes contained HSEs. Twenty-five genes had both HSEs and STREs in their promoters. The first set of genes is potentially regulated by the Msn2/Msn4/STRE interaction. For the second set of genes, regulation by heat shock could be mediated through HSF/HSE regulatory mechanisms. The overlap between these groups suggests a co-regulation by the two pathways. Our study yielded 239 candidate genes, whose regulation could potentially be affected by heat-shock via transcription interference directed both from upstream and downstream areas relative to the native promoters. In addition we have categorized 924 genes containing HSE and/or STRE elements within the Open Reading Frames (ORFs), which may also affect normal transcription. Our study revealed a widespread possibility for the regulation of genes via transcriptional interference initiated by stress response. We provided a categorization of genes potentially affected at the transcriptional level by known

  4. Advanced Glycation End-Products affect transcription factors regulating insulin gene expression

    SciTech Connect

    Puddu, A.; Storace, D.; Odetti, P.; Viviani, G.L.

    2010-04-23

    Advanced Glycation End-Products (AGEs) are generated by the covalent interaction of reducing sugars with proteins, lipids or nucleic acids. AGEs are implicated in diabetic complications and pancreatic {beta}-cell dysfunction. We previously demonstrated that exposure of the pancreatic islet cell line HIT-T15 to high concentrations of AGEs leads to a significant decrease of insulin secretion and content. Insulin gene transcription is positively regulated by the beta cell specific transcription factor PDX-1 (Pancreatic and Duodenal Homeobox-1). On the contrary, the forkhead transcription factor FoxO1 inhibits PDX-1 gene transcription. Activity of FoxO1 is regulated by post-translational modifications: phosphorylation deactivates FoxO1, and acetylation prevents FoxO1 ubiquitination. In this work we investigated whether AGEs affect expression and subcellular localization of PDX-1 and FoxO1. HIT-T15 cells were cultured for 5 days in presence of AGEs. Cells were then lysed and processed for subcellular fractionation. We determined intracellular insulin content, then we assessed the expression and subcellular localization of PDX-1, FoxO1, phosphoFoxO1 and acetylFoxO1. As expected intracellular insulin content was lower in HIT-T15 cells cultured with AGEs. The results showed that AGEs decreased expression and nuclear localization of PDX-1, reduced phosphorylation of FoxO1, and increased expression and acetylation of FoxO1. These results suggest that AGEs decrease insulin content unbalancing transcription factors regulating insulin gene expression.

  5. Genetic factors affecting gene transcription and catalytic activity of UDP-glucuronosyltransferases in human liver

    PubMed Central

    Liu, Wanqing; Ramírez, Jacqueline; Gamazon, Eric R.; Mirkov, Snezana; Chen, Peixian; Wu, Kehua; Sun, Chang; Cox, Nancy J.; Cook, Edwin; Das, Soma; Ratain, Mark J.

    2014-01-01

    The aim of this study was to discover cis- and trans-acting factors significantly affecting mRNA expression and catalytic activity of human hepatic UDP-glucuronosyltransferases (UGTs). Transcription levels of five major hepatic UGT1A (UGT1A1, UGT1A3, UGT1A4, UGT1A6 and UGT1A9) and five UGT2B (UGT2B4, UGT2B7, UGT2B10, UGT2B15 and UGT2B17) genes were quantified in human liver tissue samples (n = 125) using real-time PCR. Glucuronidation activities of 14 substrates were measured in 47 livers. We genotyped 167 tagSNPs (single-nucleotide polymorphisms) in UGT1A (n = 43) and UGT2B (n = 124), as well as the known functional UGT1A1*28 and UGT2B17 CNV (copy number variation) polymorphisms. Transcription levels of 15 transcription factors (TFs) known to regulate these UGTs were quantified. We found that UGT expression and activity were highly variable among the livers (median and range of coefficient of variations: 135%, 74–217% and 52%, 39–105%, respectively). CAR, PXR and ESR1 were found to be the most important trans-regulators of UGT transcription (median and range of correlation coefficients: 46%, 6–58%; 47%, 9–58%; and 52%, 24–75%, respectively). Hepatic UGT activities were mainly determined by UGT gene transcription levels. Twenty-one polymorphisms were significantly (FDR-adjusted P < 0.05) associated with mRNA expression and/or activities of UGT1A1, UGT1A3 and UGT2B17. We found novel SNPs in the UGT2B17 CNV region accounting for variability in UGT2B17 gene transcription and testosterone glucuronidation rate, in addition to that attributable to the UGT2B17 CNV. Our study discovered novel pharmacogenetic markers and provided detailed insight into the genetic network regulating hepatic UGTs. PMID:24879639

  6. Alteration of BRCA1 expression affects alcohol-induced transcription of RNA Pol III-dependent genes

    PubMed Central

    Zhong, Qian; Shi, Ganggang; Zhang, Yanmei; Lu, Lei; Levy, Daniel; Zhong, Shuping

    2014-01-01

    Emerging evidence has indicated that alcohol consumption is an established risk factor for breast cancer. Deregulation of RNA polymerase III (Pol III) transcription enhances cellular Pol III gene production, leading to an increase in translational capacity to promote cell transformation and tumor formation. We have reported that alcohol intake increases Pol III gene transcription to promote cell transformation and tumor formation in vitro and in vivo. Studies revealed that tumor suppressors, pRb, p53, PTEN and Maf1 repress the transcription of Pol III genes. BRCA1 is a tumor suppressor and its mutation is tightly related to breast cancer development. However, it is not clear whether BRCA1 expression affects alcohol-induced transcription of Pol III genes. At the present studies, we report that restoring BRCA1 in HCC 1937 cells, which is a BRCA1 deficient cell line, represses Pol III gene transcription. Expressing mutant or truncated BRCA1 in these cells does not affect the ability of repression on Pol III genes. Our analysis has demonstrated that alcohol induces Pol III gene transcription. More importantly, overexpression of BRCA1 in estrogen receptor positive (ER+) breast cancer cells (MCF-7) decreases the induction of tRNALeu and 5S rRNA genes by alcohol, whereas reduction of BRCA1 by its siRNA slightly increases the transcription of the class of genes. This suggests that BRCA1 is associated with alcohol-induced deregulation of Pol III genes. These studies for the first time demonstrate the role of BRCA1 in induction of Pol III genes by alcohol and uncover a novel mechanism of alcohol-associated breast cancer. PMID:25447904

  7. Transcription factor organic cation transporter 1 (OCT-1) affects the expression of porcine Klotho (KL) gene

    PubMed Central

    Zhou, Jiawei

    2016-01-01

    Klotho (KL), originally discovered as an aging suppressor, is a membrane protein that shares sequence similarity with the β-glucosidase enzymes. Recent reports showed Klotho might play a role in adipocyte maturation and systemic glucose metabolism. However, little is known about the transcription factors involved in regulating the expression of porcine KL gene. Deletion fragment analysis identified KL-D2 (−418 bp to −3 bp) as the porcine KL core promoter. MARC0022311SNP (A or G) in KL intron 1 was detected in Landrace × DIV pigs using the Porcine SNP60 BeadChip. The pGL-D2-A and pGL-D2-G were constructed with KL-D2 and the intron fragment of different alleles and relative luciferase activity of pGL3-D2-G was significantly higher than that of pGL3-D2-A in the PK cells and ST cells. This was possibly the result of a change in KL binding ability with transcription factor organic cation transporter 1 (OCT-1), which was confirmed using electrophoretic mobility shift assays (EMSA) and chromatin immune-precipitation (ChIP). Moreover, OCT-1 regulated endogenous KL expression by RNA interference experiments. Our study indicates SNP MARC0022311 affects porcine KL expression by regulating its promoter activity via OCT-1. PMID:27478698

  8. Transcription factor organic cation transporter 1 (OCT-1) affects the expression of porcine Klotho (KL) gene.

    PubMed

    Li, Yan; Wang, Lei; Zhou, Jiawei; Li, Fenge

    2016-01-01

    Klotho (KL), originally discovered as an aging suppressor, is a membrane protein that shares sequence similarity with the β-glucosidase enzymes. Recent reports showed Klotho might play a role in adipocyte maturation and systemic glucose metabolism. However, little is known about the transcription factors involved in regulating the expression of porcine KL gene. Deletion fragment analysis identified KL-D2 (-418 bp to -3 bp) as the porcine KL core promoter. MARC0022311SNP (A or G) in KL intron 1 was detected in Landrace × DIV pigs using the Porcine SNP60 BeadChip. The pGL-D2-A and pGL-D2-G were constructed with KL-D2 and the intron fragment of different alleles and relative luciferase activity of pGL3-D2-G was significantly higher than that of pGL3-D2-A in the PK cells and ST cells. This was possibly the result of a change in KL binding ability with transcription factor organic cation transporter 1 (OCT-1), which was confirmed using electrophoretic mobility shift assays (EMSA) and chromatin immune-precipitation (ChIP). Moreover, OCT-1 regulated endogenous KL expression by RNA interference experiments. Our study indicates SNP MARC0022311 affects porcine KL expression by regulating its promoter activity via OCT-1. PMID:27478698

  9. A compilation of composite regulatory elements affecting gene transcription in vertebrates.

    PubMed Central

    Kel, O V; Romaschenko, A G; Kel, A E; Wingender, E; Kolchanov, N A

    1995-01-01

    Over the past years, evidence has been accumulating for a fundamental role of protein-protein interactions between transcription factors in gene-specific transcription regulation. Many of these interactions run within composite elements containing binding sites for several factors. We have selected 101 composite regulatory elements identified experimentally in the regulatory regions of 64 genes of vertebrates and of their viruses and briefly described them in a compilation. Of these, 82 composite elements are of the synergistic type and 19 of the antagonistic type. Within the synergistic type composite elements, transcription factors bind to the corresponding sites simultaneously, thus cooperatively activating transcription. The factors, binding to their target sites within antagonistic type composite elements, produce opposing effects on transcription. The nucleotide sequence and localization in the genes, the names and brief description of transcription factors, are provided for each composite element, including a representation of experimental data on its functioning. Most of the composite elements (3/4) fall between -250 bp and the transcription start site. The distance between the binding sites within the composite elements described varies from complete overlapping to 80 bp. The compilation of composite elements is presented in the database COMPEL which is electronically accessible by anonymous ftp via internet. PMID:7479071

  10. Basal Body Structures Differentially Affect Transcription of RpoN- and FliA-Dependent Flagellar Genes in Helicobacter pylori

    PubMed Central

    Tsang, Jennifer

    2015-01-01

    ABSTRACT Flagellar biogenesis in Helicobacter pylori is regulated by a transcriptional hierarchy governed by three sigma factors, RpoD (σ80), RpoN (σ54), and FliA (σ28), that temporally coordinates gene expression with the assembly of the flagellum. Previous studies showed that loss of flagellar protein export apparatus components inhibits transcription of flagellar genes. The FlgS/FlgR two-component system activates transcription of RpoN-dependent genes though an unknown mechanism. To understand better the extent to which flagellar gene regulation is coupled to flagellar assembly, we disrupted flagellar biogenesis at various points and determined how these mutations affected transcription of RpoN-dependent (flaB and flgE) and FliA-dependent (flaA) genes. The MS ring (encoded by fliF) is one of the earliest flagellar structures assembled. Deletion of fliF resulted in the elimination of RpoN-dependent transcripts and an ∼4-fold decrease in flaA transcript levels. FliH is a cytoplasmic protein that functions with the C ring protein FliN to shuttle substrates to the export apparatus. Deletions of fliH and genes encoding C ring components (fliM and fliY) decreased transcript levels of flaB and flgE but had little or no effect on transcript levels of flaA. Transcript levels of flaB and flgE were elevated in mutants where genes encoding rod proteins (fliE and flgBC) were deleted, while transcript levels of flaA was reduced ∼2-fold in both mutants. We propose that FlgS responds to an assembly checkpoint associated with the export apparatus and that FliH and one or more C ring component assist FlgS in engaging this flagellar structure. IMPORTANCE The mechanisms used by bacteria to couple transcription of flagellar genes with assembly of the flagellum are poorly understood. The results from this study identified components of the H. pylori flagellar basal body that either positively or negatively affect expression of RpoN-dependent flagellar genes. Some of these

  11. Mod5 protein binds to tRNA gene complexes and affects local transcriptional silencing

    PubMed Central

    Pratt-Hyatt, Matthew; Pai, Dave A.; Haeusler, Rebecca A.; Wozniak, Glenn G.; Good, Paul D.; Miller, Erin L.; McLeod, Ian X.; Yates, John R.; Hopper, Anita K.; Engelke, David R.

    2013-01-01

    The tRNA gene-mediated (tgm) silencing of RNA polymerase II promoters is dependent on subnuclear clustering of the tRNA genes, but genetic analysis shows that the silencing requires additional mechanisms. We have identified proteins that bind tRNA gene transcription complexes and are required for tgm silencing but not required for gene clustering. One of the proteins, Mod5, is a tRNA modifying enzyme that adds an N6-isopentenyl adenosine modification at position 37 on a small number of tRNAs in the cytoplasm, although a subpopulation of Mod5 is also found in the nucleus. Recent publications have also shown that Mod5 has tumor suppressor characteristics in humans as well as confers drug resistance through prion-like misfolding in yeast. Here, we show that a subpopulation of Mod5 associates with tRNA gene complexes in the nucleolus. This association occurs and is required for tgm silencing regardless of whether the pre-tRNA transcripts are substrates for Mod5 modification. In addition, Mod5 is bound to nuclear pre-tRNA transcripts, although they are not substrates for the A37 modification. Lastly, we show that truncation of the tRNA transcript to remove the normal tRNA structure also alleviates silencing, suggesting that synthesis of intact pre-tRNAs is required for the silencing mechanism. These results are discussed in light of recent results showing that silencing near tRNA genes also requires chromatin modification. PMID:23898186

  12. Low-temperature affected LC-PUFA conversion and associated gene transcript level in Nannochloropsis oculata CS-179

    NASA Astrophysics Data System (ADS)

    Ma, Xiaolei; Zhang, Lin; Zhu, Baohua; Pan, Kehou; Li, Si; Yang, Guanpin

    2011-09-01

    Nannochloropsis oculata CS-179, a marine eukaryotic unicellular microalga, is rich in long-chain polyunsaturated fatty acids (LC-PUFAs). Culture temperature affected cell growth and the composition of LC-PUFAs. At an initial cell density of 1.5 × 106 cell mL-1, the highest growth was observed at 25°C and the cell density reached 3 × 107 cell mL-1 at the beginning of logarithmic phase. The content of LC-PUFAs varied with culture temperature. The highest content of LC-PUFAs (43.96%) and EPA (36.6%) was gained at 20°C. Real-time PCR showed that the abundance of Δ6-desaturase gene transcripts was significantly different among 5 culture temperatures and the highest transcript level (15°C) of Nanoc-D6D took off at cycle 21.45. The gene transcript of C20-elongase gene was higher at lower temperatures (10, 15, and 20°C), and the highest transcript level (20°C) of Nanoc-E took off at cycle 21.18. The highest conversion rate (39.3%) of Δ6-desaturase was also gained at 20°C. But the conversion rate of Nanoc-E was not detected. The higher content of LC-PUFAs was a result of higher gene transcript level and higher enzyme activity. Compared with C20-elongase gene, Δ6-desaturase gene transcript and enzyme activity varied significantly with temperature. It will be useful to study the mechanism of how the content of LC-PUFAs is affected by temperature.

  13. Power training and postmenopausal hormone therapy affect transcriptional control of specific co-regulated gene clusters in skeletal muscle

    PubMed Central

    Fey, Vidal; Törmäkangas, Timo; Ronkainen, Paula H. A.; Taaffe, Dennis R.; Takala, Timo; Koskinen, Satu; Cheng, Sulin; Puolakka, Jukka; Kujala, Urho M.; Suominen, Harri; Sipilä, Sarianna; Kovanen, Vuokko

    2010-01-01

    At the moment, there is no clear molecular explanation for the steeper decline in muscle performance after menopause or the mechanisms of counteractive treatments. The goal of this genome-wide study was to identify the genes and gene clusters through which power training (PT) comprising jumping activities or estrogen containing hormone replacement therapy (HRT) may affect skeletal muscle properties after menopause. We used musculus vastus lateralis samples from early stage postmenopausal (50–57 years old) women participating in a yearlong randomized double-blind placebo-controlled trial with PT and HRT interventions. Using microarray platform with over 24,000 probes, we identified 665 differentially expressed genes. The hierarchical clustering method was used to assort the genes. Additionally, enrichment analysis of gene ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways was carried out to clarify whether assorted gene clusters are enriched with particular functional categories. The analysis revealed transcriptional regulation of 49 GO/KEGG categories. PT upregulated transcription in “response to contraction”—category revealing novel candidate genes for contraction-related regulation of muscle function while HRT upregulated gene expression related to functionality of mitochondria. Moreover, several functional categories tightly related to muscle energy metabolism, development, and function were affected regardless of the treatment. Our results emphasize that during the early stages of the postmenopause, muscle properties are under transcriptional modulation, which both PT and HRT partially counteract leading to preservation of muscle power and potentially reducing the risk for aging-related muscle weakness. More specifically, PT and HRT may function through improving energy metabolism, response to contraction as well as by preserving functionality of the mitochondria. Electronic supplementary material The online version of this

  14. Copy number variants in patients with intellectual disability affect the regulation of ARX transcription factor gene.

    PubMed

    Ishibashi, Minaka; Manning, Elizabeth; Shoubridge, Cheryl; Krecsmarik, Monika; Hawkins, Thomas A; Giacomotto, Jean; Zhao, Ting; Mueller, Thomas; Bader, Patricia I; Cheung, Sau W; Stankiewicz, Pawel; Bain, Nicole L; Hackett, Anna; Reddy, Chilamakuri C S; Mechaly, Alejandro S; Peers, Bernard; Wilson, Stephen W; Lenhard, Boris; Bally-Cuif, Laure; Gecz, Jozef; Becker, Thomas S; Rinkwitz, Silke

    2015-11-01

    Protein-coding mutations in the transcription factor-encoding gene ARX cause various forms of intellectual disability (ID) and epilepsy. In contrast, variations in surrounding non-coding sequences are correlated with milder forms of non-syndromic ID and autism and had suggested the importance of ARX gene regulation in the etiology of these disorders. We compile data on several novel and some already identified patients with or without ID that carry duplications of ARX genomic region and consider likely genetic mechanisms underlying the neurodevelopmental defects. We establish the long-range regulatory domain of ARX and identify its brain region-specific autoregulation. We conclude that neurodevelopmental disturbances in the patients may not simply arise from increased dosage due to ARX duplication. This is further exemplified by a small duplication involving a non-functional ARX copy, but with duplicated enhancers. ARX enhancers are located within a 504-kb region and regulate expression specifically in the forebrain in developing and adult zebrafish. Transgenic enhancer-reporter lines were used as in vivo tools to delineate a brain region-specific negative and positive autoregulation of ARX. We find autorepression of ARX in the telencephalon and autoactivation in the ventral thalamus. Fluorescently labeled brain regions in the transgenic lines facilitated the identification of neuronal outgrowth and pathfinding disturbances in the ventral thalamus and telencephalon that occur when arxa dosage is diminished. In summary, we have established a model for how breakpoints in long-range gene regulation alter the expression levels of a target gene brain region-specifically, and how this can cause subtle neuronal phenotypes relating to the etiology of associated neuropsychiatric disease. PMID:26337422

  15. Parent-of-origin genetic background affects the transcriptional levels of circadian and neuronal plasticity genes following sleep loss

    PubMed Central

    Tinarelli, Federico; Garcia-Garcia, Celina; Nicassio, Francesco; Tucci, Valter

    2014-01-01

    Sleep homoeostasis refers to a process in which the propensity to sleep increases as wakefulness progresses and decreases as sleep progresses. Sleep is tightly organized around the circadian clock and is regulated by genetic and epigenetic mechanisms. The homoeostatic response of sleep, which is classically triggered by sleep deprivation, is generally measured as a rebound effect of electrophysiological measures, for example delta sleep. However, more recently, gene expression changes following sleep loss have been investigated as biomarkers of sleep homoeostasis. The genetic background of an individual may affect this sleep-dependent gene expression phenotype. In this study, we investigated whether parental genetic background differentially modulates the expression of genes following sleep loss. We tested the progeny of reciprocal crosses of AKR/J and DBA/2J mouse strains and we show a parent-of-origin effect on the expression of circadian, sleep and neuronal plasticity genes following sleep deprivation. Thus, we further explored, by in silico, specific functions or upstream mechanisms of regulation and we observed that several upstream mechanisms involving signalling pathways (i.e. DICER1, PKA), growth factors (CSF3 and BDNF) and transcriptional regulators (EGR2 and ELK4) may be differentially modulated by parental effects. This is the first report showing that a behavioural manipulation (e.g. sleep deprivation) in adult animals triggers specific gene expression responses according to parent-of-origin genomic mechanisms. Our study suggests that the same mechanism may be extended to other behavioural domains and that the investigation of gene expression following experimental manipulations should take seriously into account parent-of-origin effects. PMID:24446504

  16. Interaction between bisphenol A and dietary sugar affects global gene transcription in Drosophila melanogaster

    PubMed Central

    Branco, Alan T.; Lemos, Bernardo

    2014-01-01

    Human exposure to environmental toxins is a public health issue. The microarray data available in the Gene Expression Omnibus database under accession number GSE55655 and GSE55670GSE55655GSE55670 show the isolated and combined effects of dietary sugar and two organic compounds present in a variety of plastics [bisphenol A (BPA) and Bis(2-ethylhexyl) phthalate (DEHP)] on global gene expression in Drosophila melanogaster. The study was carried out with samples collected from flies exposed to these compounds for a limited period of time (48 h) in the adult stage, or throughout the entire development of the insect. The arrays were normalized using the limma/Bioconductor package. Differential expression was inferred using linear models in limma and BAGEL. The data show that each compound had its unique consequences to gene expression, and that the individual effect of each organic compound is maximized with the joint ingestion of dietary sugar. PMID:26484116

  17. Intragenic deletions affecting two alternative transcripts of the IMMP2L gene in patients with Tourette syndrome

    PubMed Central

    Bertelsen, Birgitte; Melchior, Linea; Jensen, Lars R; Groth, Camilla; Glenthøj, Birte; Rizzo, Renata; Debes, Nanette Mol; Skov, Liselotte; Brøndum-Nielsen, Karen; Paschou, Peristera; Silahtaroglu, Asli; Tümer, Zeynep

    2014-01-01

    Tourette syndrome is a neurodevelopmental disorder characterized by multiple motor and vocal tics, and the disorder is often accompanied by comorbidities such as attention-deficit hyperactivity-disorder and obsessive compulsive disorder. Tourette syndrome has a complex etiology, but the underlying environmental and genetic factors are largely unknown. IMMP2L (inner mitochondrial membrane peptidase, subunit 2) located on chromosome 7q31 is one of the genes suggested as a susceptibility factor in disease pathogenesis. Through screening of a Danish cohort comprising 188 unrelated Tourette syndrome patients for copy number variations, we identified seven patients with intragenic IMMP2L deletions (3.7%), and this frequency was significantly higher (P=0.0447) compared with a Danish control cohort (0.9%). Four of the seven deletions identified did not include any known exons of IMMP2L, but were within intron 3. These deletions were found to affect a shorter IMMP2L mRNA species with two alternative 5′-exons (one including the ATG start codon). We showed that both transcripts (long and short) were expressed in several brain regions, with a particularly high expression in cerebellum and hippocampus. The current findings give further evidence for the role of IMMP2L as a susceptibility factor in Tourette syndrome and suggest that intronic changes in disease susceptibility genes should be investigated further for presence of alternatively spliced exons. PMID:24549057

  18. DNA topology affects transcriptional regulation of the pertussis toxin gene of Bordetella pertussis in Escherichia coli and in vitro.

    PubMed Central

    Scarlato, V; Aricò, B; Rappuoli, R

    1993-01-01

    The bvg locus of Bordetella pertussis encodes an environmentally inducible operon essential for the expression of virulence genes. We show that in Escherichia coli, the PTOX promoter cloned in cis of the bvg locus is activated and environmentally regulated. Cotransformation of E. coli with the bvg locus cloned in a low-copy-number plasmid and with the PTOX promoter cloned in a high-copy-number plasmid can give rise to two different results. If the PTOX promoter is cloned in the pGem-3 vector, transcription is absent. If the PTOX promoter is cloned in the plasmid pKK232, containing the PTOX promoter between two ribosomal gene terminators of transcription, transcription occurs, although regulation of transcription is abolished. Under these conditions, the intracellular amount of RNA transcripts is increased by adding to the culture medium novobiocin, an inhibitor of bacterial gyrases. In vitro, the transcription of the PTOX promoter is activated on E. coli RNA polymerase supplemented with cell extracts from wild-type B. pertussis. Addition of DNA gyrase to the mixture dramatically reduces the amount of RNA synthesized. Our data show that the products of the bvg locus, BvgA and BvgS, are directly involved in the regulation of the PTOX promoter in E. coli and that DNA topology may play a role in the induction of transcription. Images PMID:8393006

  19. Quantitative gene expression profiling of mouse brain regions reveals differential transcripts conserved in human and affected in disease models.

    PubMed

    Brochier, Camille; Gaillard, Marie-Claude; Diguet, Elsa; Caudy, Nicolas; Dossat, Carole; Ségurens, Béatrice; Wincker, Patrick; Roze, Emmanuel; Caboche, Jocelyne; Hantraye, Philippe; Brouillet, Emmanuel; Elalouf, Jean-Marc; de Chaldée, Michel

    2008-04-22

    Using serial analysis of gene expression, we collected quantitative transcriptome data in 11 regions of the adult wild-type mouse brain: the orbital, prelimbic, cingulate, motor, somatosensory, and entorhinal cortices, the caudate-putamen, the nucleus accumbens, the thalamus, the substantia nigra, and the ventral tegmental area. With >1.2 million cDNA tags sequenced, this database is a powerful resource to explore brain functions and disorders. As an illustration, we performed interregional comparisons and found 315 differential transcripts. Most of them are poorly characterized and 20% lack functional annotation. For 78 differential transcripts, we provide independent expression level measurements in mouse brain regions by real-time quantitative RT-PCR. We also show examples where we used in situ hybridization to achieve infrastructural resolution. For 30 transcripts, we next demonstrated that regional enrichment is conserved in the human brain. We then quantified the expression levels of region-enriched transcripts in the R6/2 mouse model of Huntington disease and the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of Parkinson disease and observed significant alterations in the striatum, cerebral cortex, thalamus and substantia nigra of R6/2 mice and in the striatum of MPTP-treated mice. These results show that the gene expression data provided here for the mouse brain can be used to explore pathophysiological models and disclose transcripts differentially expressed in human brain regions. PMID:18252803

  20. The Zinc Finger Protein ZNF658 Regulates the Transcription of Genes Involved in Zinc Homeostasis and Affects Ribosome Biogenesis through the Zinc Transcriptional Regulatory Element

    PubMed Central

    Ogo, Ogo A.; Tyson, John; Cockell, Simon J.; Howard, Alison; Valentine, Ruth A.

    2015-01-01

    We previously identified the ZTRE (zinc transcriptional regulatory element) in genes involved in zinc homeostasis and showed that it mediates transcriptional repression in response to zinc. We now report that ZNF658 acts at the ZTRE. ZNF658 was identified by matrix-assisted laser desorption ionization–time of flight mass spectrometry of a band excised after electrophoretic mobility shift assay using a ZTRE probe. The protein contains a KRAB domain and 21 zinc fingers. It has similarity with ZAP1 from Saccharomyces cerevisiae, which regulates the response to zinc restriction, including a conserved DNA binding region we show to be functional also in ZNF658. Small interfering RNA (siRNA) targeted to ZNF658 abrogated the zinc-induced, ZTRE-dependent reduction in SLC30A5 (ZnT5 gene), SLC30A10 (ZnT10 gene), and CBWD transcripts in human Caco-2 cells and the ability of zinc to repress reporter gene expression from corresponding promoter-reporter constructs. Microarray analysis of the effect of reducing ZNF658 expression by siRNA uncovered a large decrease in rRNA. We find that ZTREs are clustered within the 45S rRNA precursor. We also saw effects on expression of multiple ribosomal proteins. ZNF658 thus links zinc homeostasis with ribosome biogenesis, the most active transcriptional, and hence zinc-demanding, process in the cell. ZNF658 is thus a novel transcriptional regulator that plays a fundamental role in the orchestrated cellular response to zinc availability. PMID:25582195

  1. Gene dosage of the transcription factor Fingerin (bHLHA9) affects digit development and links syndactyly to ectrodactyly.

    PubMed

    Schatz, Omri; Langer, Erez; Ben-Arie, Nissim

    2014-10-15

    Distal limb deformities are congenital malformations with phenotypic variability, genetic heterogeneity and complex inheritance. Among these, split-hand/foot malformation is an ectrodactyly with missing central fingers, yielding a lobster claw-like hand, which when combined with long-bone deficiency is defined as split-hand/foot malformation and long-bone deficiency (SHFLD) that is genetically heterogeneous. Copy number variation (CNV) consisting of 17p13.3 duplication was identified in unrelated pedigrees, underlying SHFLD3 (OMIM 612576). Although the transcription factor Fingerin (bHLHA9) is the only complete gene in the critical region, its biological role is not yet known and there are no data supporting its involvement in mammalian limb development. We have generated knockout mice in which only the entire coding region of Fingerin was deleted, and indeed found that most null mice display some limb defects. These include various levels of simple asymmetrical syndactyly, characterized by webbed fingers, generated by incomplete separation of soft, but not skeletal, tissues between forelimb digits 2 and 3. As expected, hand pads of Fingerin null embryos exhibited reduced apoptosis between digital rays 2 and 3. This defect was shown to cause syndactyly when the same limbs were grown ex vivo following the apoptosis assay. Extrapolating from mouse data, we suggest that Fingerin loss-of-function in humans may underlie MSSD syndactyly (OMIM 609432), which was mapped to the same locus. Taken together, Fingerin gene dosage links two different congenital limb malformations, syndactyly and ectrodactyly, which were previously postulated to share a common etiology. These results add limb disorders to the growing list of diseases resulting from CNV. PMID:24852374

  2. Mutations That Affect Transcription and Cyclic Amp-Crp Regulation of the Adenylate Cyclase Gene (Cya) of Salmonella Typhimurium

    PubMed Central

    Fandl, J. P.; Thorner, L. K.; Artz, S. W.

    1990-01-01

    We studied the expression of the cya promoter(s) in cya-lac fusion strains of Salmonella typhimurium and demonstrated cAMP receptor protein (CRP)-dependent repression by cAMP. Expression of cya was reduced about fourfold in cultures grown in acetate minimal medium as compared to cultures grown in glucose-6-phosphate minimal medium. Expression of cya was also reduced about fourfold by addition of 5 mM cAMP to cultures grown in glucose minimal medium. We constructed in vitro deletion and insertion mutations altering a major cya promoter (P2) and a putative CRP binding site overlapping P2. These mutations were recombined into the chromosome by allele replacement with M13mp::cya recombinant phages and the regulation of the mutant promoters was analyzed. A 4-bp deletion of the CRP binding site and a 4-bp insertion in this site nearly eliminated repression by cAMP. A mutant with the P2 promoter and the CRP binding site both deleted exhibited an 80% reduction in cya expression; the 20% residual expression was insensitive to cAMP repression. This mutant retained a Cya(+) phenotype. Taken together, the results establish that the cya gene is transcribed from multiple promoters one of which, P2, is negatively regulated by the cAMP-CRP complex. Correction for the contribution to transcription by the cAMP-CRP nonregulated cya promoters indicates that the P2 promoter is repressed at least eightfold by cAMP-CRP. PMID:2168849

  3. The Shwachman-Bodian-Diamond syndrome associated protein interacts with HsNip7 and its down-regulation affects gene expression at the transcriptional and translational levels

    SciTech Connect

    Hesling, Cedric; Oliveira, Carla C.; Castilho, Beatriz A.; Zanchin, Nilson I.T.

    2007-12-10

    The Shwachman-Bodian-Diamond syndrome (SDS) is an autosomal disorder with pleiotropic phenotypes including pancreatic, skeletal and bone marrow deficiencies and predisposition to hematological dysfunctions. SDS has been associated to mutations in the SBDS gene, encoding a highly conserved protein that was shown to function in ribosome biogenesis in yeast. In this work, we show that SBDS is found in complexes containing the human Nip7 ortholog. Analysis of pre-rRNA processing in a stable SBDS knock-down HEK293-derivative cell line revealed accumulation of a small RNA which is a further indication of SBDS involvement in rRNA biosynthesis. Global transcription and polysome-bound mRNA profiling revealed that SBDS knock-down affects expression of critical genes involved in brain development and function, bone morphogenesis, blood cell proliferation and differentiation, and cell adhesion. Expression of a group of growth and signal transduction factors and of DNA damage response genes is also affected. In SBDS knock-down cells, 34 mRNAs showed decreased and 55 mRNAs showed increased association to polysomes, among which is a group encoding proteins involved in alternative splicing and RNA modification. These results indicate that SBDS is required for accurate expression of genes important for proper brain, skeletal, and blood cell development.

  4. Polymorphisms at positions -22 and -348 in the promoter of the BAT1 gene affect transcription and the binding of nuclear factors.

    PubMed

    Price, Patricia; Wong, Agnes M-L; Williamson, David; Voon, Dominic; Baltic, Svetlana; Allcock, Richard J N; Boodhoo, Alvin; Christiansen, Frank T

    2004-05-01

    BAT1 (D6S81E, UAP56) lies in the central MHC between TNF and HLA-B, a region containing genes that affect susceptibility to immunopathologic disorders. BAT1 protein may be directly responsible for the genetic association, as antisense studies show it can down-regulate inflammatory cytokines. Here we investigate polymorphisms at positions -22 and -348 relative to the BAT1 transcription start site. DNA samples from healthy donors were used to confirm haplotypic associations with the type 1 diabetes-susceptible 8.1 ancestral haplotype (AH; HLA-A1,B8,BAT1-22*C,BAT1-348*C,DR3 ) and the diabetes-resistant 7.1 AH (HLA-A3,B7,BAT1-22*G,BAT1-348*T,DR15). Alleles carried at BAT1-22 and -348 were in linkage disequilibrium. Electrophoretic mobility shift assays using nuclear proteins from T-cells (Jurkat and HT2), monocytes (THP1, U937) and epithelial cells (HeLa and MDA468) demonstrated DNA : protein complexes binding oligonucleotides spanning positions -22 and -348 on the 7.1 AH only. Competition assays, supershifts and molecular weight determinations suggest the complexes include the transcription factors YY1 (at -348) and Oct1 (at -22). Promoter activity was demonstrated using 520 bp and 336 bp fragments cloned from immediately upstream of the transcription start site and carrying all combinations of -22 and -348 alleles, suggesting an unidentified non-polymorphic sequence within 336 bp of the start site drives transcription. The 520 bp fragment of the BAT1 promoter cloned from the 8.1 AH was slightly less efficient than the equivalent from the 7.1 AH, whilst the reverse was observed with 336 bp fragments. This suggests BAT1 transcription on the 7.1 AH is modified by interactions involving DNA flanking positions -22 and -348. PMID:15028669

  5. Transcriptional gene silencing in humans.

    PubMed

    Weinberg, Marc S; Morris, Kevin V

    2016-08-19

    It has been over a decade since the first observation that small non-coding RNAs can functionally modulate epigenetic states in human cells to achieve functional transcriptional gene silencing (TGS). TGS is mechanistically distinct from the RNA interference (RNAi) gene-silencing pathway. TGS can result in long-term stable epigenetic modifications to gene expression that can be passed on to daughter cells during cell division, whereas RNAi does not. Early studies of TGS have been largely overlooked, overshadowed by subsequent discoveries of small RNA-directed post-TGS and RNAi. A reappraisal of early work has been brought about by recent findings in human cells where endogenous long non-coding RNAs function to regulate the epigenome. There are distinct and common overlaps between the proteins involved in small and long non-coding RNA transcriptional regulatory mechanisms, suggesting that the early studies using small non-coding RNAs to modulate transcription were making use of a previously unrecognized endogenous mechanism of RNA-directed gene regulation. Here we review how non-coding RNA plays a role in regulation of transcription and epigenetic gene silencing in human cells by revisiting these earlier studies and the mechanistic insights gained to date. We also provide a list of mammalian genes that have been shown to be transcriptionally regulated by non-coding RNAs. Lastly, we explore how TGS may serve as the basis for development of future therapeutic agents. PMID:27060137

  6. Sequencing and Transcriptional Analysis of the Streptococcus thermophilus Histamine Biosynthesis Gene Cluster: Factors That Affect Differential hdcA Expression▿ †

    PubMed Central

    Calles-Enríquez, Marina; Eriksen, Benjamin Hjort; Andersen, Pia Skov; Rattray, Fergal P.; Johansen, Annette H.; Fernández, María; Ladero, Victor; Alvarez, Miguel A.

    2010-01-01

    Histamine, a toxic compound that is formed by the decarboxylation of histidine through the action of microbial decarboxylases, can accumulate in fermented food products. From a total of 69 Streptococcus thermophilus strains screened, two strains, CHCC1524 and CHCC6483, showed the capacity to produce histamine. The hdc clusters of S. thermophilus CHCC1524 and CHCC6483 were sequenced, and the factors that affect histamine biosynthesis and histidine-decarboxylating gene (hdcA) expression were studied. The hdc cluster began with the hdcA gene, was followed by a transporter (hdcP), and ended with the hdcB gene, which is of unknown function. The three genes were orientated in the same direction. The genetic organization of the hdc cluster showed a unique organization among the lactic acid bacterial group and resembled those of Staphylococcus and Clostridium species, thus indicating possible acquisition through a horizontal transfer mechanism. Transcriptional analysis of the hdc cluster revealed the existence of a polycistronic mRNA covering the three genes. The histidine-decarboxylating gene (hdcA) of S. thermophilus demonstrated maximum expression during the stationary growth phase, with high expression levels correlated with high histamine levels. Limited expression was evident during the lag and exponential growth phases. Low-temperature (4°C) incubation of milk inoculated with a histamine-producing strain showed lower levels of histamine than did inoculated milk kept at 42°C. This reduction was attributed to a reduction in the activity of the HdcA enzyme itself rather than a reduction in gene expression or the presence of a lower cell number. PMID:20656875

  7. Transcriptional regulation of tenascin genes

    PubMed Central

    Chiovaro, Francesca; Chiquet-Ehrismann, Ruth; Chiquet, Matthias

    2015-01-01

    Extracellular matrix proteins of the tenascin family resemble each other in their domain structure, and also share functions in modulating cell adhesion and cellular responses to growth factors. Despite these common features, the 4 vertebrate tenascins exhibit vastly different expression patterns. Tenascin-R is specific to the central nervous system. Tenascin-C is an “oncofetal” protein controlled by many stimuli (growth factors, cytokines, mechanical stress), but with restricted occurrence in space and time. In contrast, tenascin-X is a constituitive component of connective tissues, and its level is barely affected by external factors. Finally, the expression of tenascin-W is similar to that of tenascin-C but even more limited. In accordance with their highly regulated expression, the promoters of the tenascin-C and -W genes contain TATA boxes, whereas those of the other 2 tenascins do not. This article summarizes what is currently known about the complex transcriptional regulation of the 4 tenascin genes in development and disease. PMID:25793574

  8. Transcriptional regulation of tenascin genes.

    PubMed

    Chiovaro, Francesca; Chiquet-Ehrismann, Ruth; Chiquet, Matthias

    2015-01-01

    Extracellular matrix proteins of the tenascin family resemble each other in their domain structure, and also share functions in modulating cell adhesion and cellular responses to growth factors. Despite these common features, the 4 vertebrate tenascins exhibit vastly different expression patterns. Tenascin-R is specific to the central nervous system. Tenascin-C is an "oncofetal" protein controlled by many stimuli (growth factors, cytokines, mechanical stress), but with restricted occurrence in space and time. In contrast, tenascin-X is a constituitive component of connective tissues, and its level is barely affected by external factors. Finally, the expression of tenascin-W is similar to that of tenascin-C but even more limited. In accordance with their highly regulated expression, the promoters of the tenascin-C and -W genes contain TATA boxes, whereas those of the other 2 tenascins do not. This article summarizes what is currently known about the complex transcriptional regulation of the 4 tenascin genes in development and disease. PMID:25793574

  9. Histone deacetylase inhibitor abexinostat affects chromatin organization and gene transcription in normal B cells and in mantle cell lymphoma.

    PubMed

    Markozashvili, Diana; Pichugin, Andrei; Barat, Ana; Camara-Clayette, Valerie; Vasilyeva, Natalia V; Lelièvre, Hélène; Kraus-Berthier, Laurence; Depil, Stéphane; Ribrag, Vincent; Vassetzky, Yegor

    2016-04-15

    Mantle cell lymphoma (MCL) is a rare lymphoma caused by the t(11:14) juxtaposing the cyclin D1 (CCND1) locus on chromosome 11 and the immunoglobulin heavy chain (IgH) locus on chromosome 14. Several new treatments are proposed for MCL, including histone deacetylase inhibitors (HDACi). We have studied gene expression and chromatin organization in the translocated 11q13 locus in MCL cells as compared to lymphoblastoid cell lines as well as the effect of HDACi abexinostat on chromatin organization and gene expression in the 11q13 locus. We have identified a cluster of genes overexpressed in the translocation region on chromosome 11 in MCL cells. Abexinostat provokes a genome-wide disaggregation of heterochromatin. The genes upregulated after the t(11;14) translocation react to the HDACi treatment by increasing their expression, but their gene promoters do not show significant alterations in H3K9Ac and H3K9me2 levels in abexinostat-treated cells. PMID:26774800

  10. Transcriptional Silencing by Hairpin RNAs Complementary to a Gene Promoter

    PubMed Central

    Chu, Yongjun; Kalantari, Roya; Dodd, David W.

    2012-01-01

    Double-stranded RNAs can target gene promoters and inhibit transcription. To date, most research has focused on synthetic RNA duplexes. Transcriptional silencing by hairpin RNAs would facilitate a better understanding of endogenous RNA-mediated regulation of transcription within cells. Here we examine transcriptional silencing of progesterone receptor (PR) expression by hairpin RNAs. We identify the guide strand as the strand complementary to an antisense transcript at the PR promoter and that hairpin RNAs are active transcriptional silencing agents. The sequence of the hairpin loop affects activity, with the highest activity achieved when the loop has the potential for full complementarity to the antisense transcript target. Introduction of centrally mismatched bases relative to the target transcript does not prevent transcriptional silencing unless the mismatches are present on both the guide and passenger strands. These data demonstrate that hairpin RNAs can cause transcriptional silencing and offer insights into the mechanism of gene modulation by RNAs that target gene promoters. PMID:22703280

  11. Nascent transcription affected by RNA polymerase IV in Zea mays.

    PubMed

    Erhard, Karl F; Talbot, Joy-El R B; Deans, Natalie C; McClish, Allison E; Hollick, Jay B

    2015-04-01

    All eukaryotes use three DNA-dependent RNA polymerases (RNAPs) to create cellular RNAs from DNA templates. Plants have additional RNAPs related to Pol II, but their evolutionary role(s) remain largely unknown. Zea mays (maize) RNA polymerase D1 (RPD1), the largest subunit of RNA polymerase IV (Pol IV), is required for normal plant development, paramutation, transcriptional repression of certain transposable elements (TEs), and transcriptional regulation of specific alleles. Here, we define the nascent transcriptomes of rpd1 mutant and wild-type (WT) seedlings using global run-on sequencing (GRO-seq) to identify the broader targets of RPD1-based regulation. Comparisons of WT and rpd1 mutant GRO-seq profiles indicate that Pol IV globally affects transcription at both transcriptional start sites and immediately downstream of polyadenylation addition sites. We found no evidence of divergent transcription from gene promoters as seen in mammalian GRO-seq profiles. Statistical comparisons identify genes and TEs whose transcription is affected by RPD1. Most examples of significant increases in genic antisense transcription appear to be initiated by 3'-proximal long terminal repeat retrotransposons. These results indicate that maize Pol IV specifies Pol II-based transcriptional regulation for specific regions of the maize genome including genes having developmental significance. PMID:25653306

  12. Nascent Transcription Affected by RNA Polymerase IV in Zea mays

    PubMed Central

    Erhard, Karl F.; Talbot, Joy-El R. B.; Deans, Natalie C.; McClish, Allison E.; Hollick, Jay B.

    2015-01-01

    All eukaryotes use three DNA-dependent RNA polymerases (RNAPs) to create cellular RNAs from DNA templates. Plants have additional RNAPs related to Pol II, but their evolutionary role(s) remain largely unknown. Zea mays (maize) RNA polymerase D1 (RPD1), the largest subunit of RNA polymerase IV (Pol IV), is required for normal plant development, paramutation, transcriptional repression of certain transposable elements (TEs), and transcriptional regulation of specific alleles. Here, we define the nascent transcriptomes of rpd1 mutant and wild-type (WT) seedlings using global run-on sequencing (GRO-seq) to identify the broader targets of RPD1-based regulation. Comparisons of WT and rpd1 mutant GRO-seq profiles indicate that Pol IV globally affects transcription at both transcriptional start sites and immediately downstream of polyadenylation addition sites. We found no evidence of divergent transcription from gene promoters as seen in mammalian GRO-seq profiles. Statistical comparisons identify genes and TEs whose transcription is affected by RPD1. Most examples of significant increases in genic antisense transcription appear to be initiated by 3ʹ-proximal long terminal repeat retrotransposons. These results indicate that maize Pol IV specifies Pol II-based transcriptional regulation for specific regions of the maize genome including genes having developmental significance. PMID:25653306

  13. Transcriptional interference among the murine β-like globin genes

    PubMed Central

    Hu, Xiao; Eszterhas, Susan; Pallazzi, Nicolas; Bouhassira, Eric E.; Fields, Jennifer; Tanabe, Osamu; Gerber, Scott A.; Bulger, Michael; Engel, James Douglas; Groudine, Mark

    2007-01-01

    Mammalian β-globin loci contain multiple genes that are activated at different developmental stages. Studies have suggested that the transcription of one gene in a locus can influence the expression of the other locus genes. The prevalent model to explain this transcriptional interference is that all potentially active genes compete for locus control region (LCR) activity. To investigate the influence of transcription by the murine embryonic genes on transcription of the other β-like genes, we generated mice with deletions of the promoter regions of Ey and βh1 and measured transcription of the remaining genes. Deletion of the Ey and βh1 promoters increased transcription of βmajor and βminor 2-fold to 3-fold during primitive erythropoiesis. Deletion of Ey did not affect βh1 nor did deletion of βh1 affect Ey, but Ey deletion uniquely activated transcription from βh0, a β-like globin gene immediately downstream of Ey. Protein analysis showed that βh0 encodes a translatable β-like globin protein that can pair with alpha globin. The lack of transcriptional interference between Ey and βh1 and the gene-specific repression of βh0 did not support LCR competition among the embryonic genes and suggested that direct transcriptional interference from Ey suppressed βh0. PMID:17077320

  14. Ectopic expression of R3 MYB transcription factor gene OsTCL1 in Arabidopsis, but not rice, affects trichome and root hair formation.

    PubMed

    Zheng, Kaijie; Tian, Hainan; Hu, Qingnan; Guo, Hongyan; Yang, Li; Cai, Ling; Wang, Xutong; Liu, Bao; Wang, Shucai

    2016-01-01

    In Arabidopsis, a MYB-bHLH-WD40 (MBW) transcriptional activator complex activates the homeodomain protein gene GLABRA2 (GL2), leading to the promotion of trichome formation and inhibition of root hair formation. The same MBW complex also activates single-repeat R3 MYB genes. R3 MYBs in turn, play a negative feedback role by competing with R2R3 MYB proteins for binding bHLH proteins, thus blocking the formation of the MBW complex. By BLASTing the rice (Oryza sativa) protein database using the entire amino acid sequence of Arabidopsis R3 MYB transcription factor TRICHOMELESS1 (TCL1), we found that there are two genes in rice genome encoding R3 MYB transcription factors, namely Oryza sativa TRICHOMELESS1 (OsTCL1) and OsTCL2. Expressing OsTCL1 in Arabidopsis inhibited trichome formation and promoted root hair formation, and OsTCL1 interacted with GL3 when tested in Arabidopsis protoplasts. Consistent with these observations, expression levels of GL2, R2R3 MYB transcription factor gene GLABRA1 (GL1) and several R3 MYB genes were greatly reduced, indicating that OsTCL1 is functional R3 MYB. However, trichome and root hair formation in transgenic rice plants overexpressing OsTCL1 remained largely unchanged, and elevated expression of OsGL2 was observed in the transgenic rice plants, indicating that rice may use different mechanisms to regulate trichome formation. PMID:26758286

  15. Ectopic expression of R3 MYB transcription factor gene OsTCL1 in Arabidopsis, but not rice, affects trichome and root hair formation

    PubMed Central

    Zheng, Kaijie; Tian, Hainan; Hu, Qingnan; Guo, Hongyan; Yang, Li; Cai, Ling; Wang, Xutong; Liu, Bao; Wang, Shucai

    2016-01-01

    In Arabidopsis, a MYB-bHLH-WD40 (MBW) transcriptional activator complex activates the homeodomain protein gene GLABRA2 (GL2), leading to the promotion of trichome formation and inhibition of root hair formation. The same MBW complex also activates single-repeat R3 MYB genes. R3 MYBs in turn, play a negative feedback role by competing with R2R3 MYB proteins for binding bHLH proteins, thus blocking the formation of the MBW complex. By BLASTing the rice (Oryza sativa) protein database using the entire amino acid sequence of Arabidopsis R3 MYB transcription factor TRICHOMELESS1 (TCL1), we found that there are two genes in rice genome encoding R3 MYB transcription factors, namely Oryza sativa TRICHOMELESS1 (OsTCL1) and OsTCL2. Expressing OsTCL1 in Arabidopsis inhibited trichome formation and promoted root hair formation, and OsTCL1 interacted with GL3 when tested in Arabidopsis protoplasts. Consistent with these observations, expression levels of GL2, R2R3 MYB transcription factor gene GLABRA1 (GL1) and several R3 MYB genes were greatly reduced, indicating that OsTCL1 is functional R3 MYB. However, trichome and root hair formation in transgenic rice plants overexpressing OsTCL1 remained largely unchanged, and elevated expression of OsGL2 was observed in the transgenic rice plants, indicating that rice may use different mechanisms to regulate trichome formation. PMID:26758286

  16. Detection of ovine herpesvirus 2 major capsid gene transcripts as an indicator of virus replication in shedding sheep and clinically affected animals

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The aim of this study was to identify tissues where Ovine herpesvirus-2 (OvHV-2) replication occurs in vivo. A reverse-transcriptase PCR targeting the OvHV-2 major capsid protein gene (ORF 25) was developed and the presence of transcripts used as an indicator of virus replication in naturally infect...

  17. Thyroid active agents T3 and PTU differentially affect immune gene transcripts in the head kidney of rainbow trout (Oncorynchus mykiss).

    PubMed

    Quesada-García, Alba; Encinas, Paloma; Valdehita, Ana; Baumann, Lisa; Segner, Helmut; Coll, Julio M; Navas, José M

    2016-05-01

    In mammals, numerous reports describe an immunomodulating effect of thyroid-active compounds. In contrast, only few reports have been published on this subject in fish. We previously demonstrated that immune cells of rainbow trout (Oncorhynchus mykiss) possess thyroid hormone receptors (THRs) and that exposure of trout to the thyroid hormone 3,3',5-triiodo-l-thyronine (T3) or the antithyroid drug propylthiouracil (PTU) alters immune cell transcript levels of THR and several immune genes. The present study aims to further characterize the immunomodulating action of thyroid-active compounds in trout immune cells. We report here the use of a custom-designed 60-mer oligo immune-targeted microarray for rainbow trout to analyze the gene expression profiles induced in the head kidney by T3 and PTU. Morphometric analyses of the thyroid showed that PTU exposure increased the size of the epithelial cells, whereas T3 induced no significant effects. Both T3 and PTU had diverse and partly contrasting effects on immune transcript profiles. The strongest differential effects of T3 and PTU on gene expressions were those targeting the Mitogen Associated Protein Kinase (MAPK), NFkB, Natural Killer (NK) and Toll-Like Receptor (TLR) pathways, a number of multipath genes (MPG) such as those encoding pleiotropic transcription factors (atf1, junb, myc), as well as important pro-inflammatory genes (tnfa, tnf6, il1b) and interferon-related genes (ifng, irf10). With these results we show for the first time in a fish species that the in vivo thyroidal status modulates a diversity of immune genes and pathways. This knowledge provides the basis to investigate both mechanisms and consequences of thyroid hormone- and thyroid disruptor-mediated immunomodulation for the immunocompetence of fish. PMID:26963519

  18. Transcription factor CecR (YbiH) regulates a set of genes affecting the sensitivity of Escherichia coli against cefoperazone and chloramphenicol.

    PubMed

    Yamanaka, Yuki; Shimada, Tomohiro; Yamamoto, Kaneyoshi; Ishihama, Akira

    2016-07-01

    Genomic SELEX (systematic evolution of ligands by exponential enrichment) screening was performed for identification of the binding site of YbiH, an as yet uncharacterized TetR-family transcription factor, on the Escherichia coli genome. YbiH was found to be a unique single-target regulator that binds in vitro within the intergenic spacer located between the divergently transcribed ybiH-ybhGFSR and rhlE operons. YbhG is an inner membrane protein and YbhFSR forms a membrane-associated ATP-binding cassette (ABC) transporter while RhlE is a ribosome-associated RNA helicase. Gel shift assay and DNase footprinting analyses indicated one clear binding site of YbiH, including a complete palindromic sequence of AATTAGTT-AACTAATT. An in vivo reporter assay indicated repression of the ybiH operon and activation of the rhlE operon by YbiH. After phenotype microarray screening, YbiH was indicated to confer resistance to chloramphenicol and cefazoline (a first-generation cephalosporin). A systematic survey of the participation of each of the predicted YbiH-regulated genes in the antibiotic sensitivity indicated involvement of the YbhFSR ABC-type transporter in the sensitivity to cefoperazone (a third-generation cephalosporin) and of the membrane protein YbhG in the control of sensitivity to chloramphenicol. Taken together with the growth test in the presence of these two antibiotics and in vitro transcription assay, it was concluded that the hitherto uncharacterized YbiH regulates transcription of both the bidirectional transcription units, the ybiH-ybhGFSR operon and the rhlE gene, which altogether are involved in the control of sensitivity to cefoperazone and chloramphenicol. We thus propose to rename YbiH as CecR (regulator of cefoperazone and chloramphenicol sensitivity). PMID:27112147

  19. The poplar basic helix-loop-helix transcription factor BEE3 – Like gene affects biomass production by enhancing proliferation of xylem cells in poplar

    SciTech Connect

    Noh, Seol Ah Choi, Young-Im Cho, Jin-Seong Lee, Hyoshin

    2015-06-19

    Brassinosteroids (BRs) play important roles in many aspects of plant growth and development, including regulation of vascular cambium activities and cell elongation. BR-induced BEE3 (brassinosteroid enhanced expression 3) is required for a proper BR response. Here, we identified a poplar (Populus alba × Populus glandulosa) BEE3-like gene, PagBEE3L, encoding a putative basic helix-loop-helix (bHLH)-type transcription factor. Expression of PagBEE3L was induced by brassinolide (BL). Transcripts of PagBEE3L were mainly detected in stems, with the internode having a low level of transcription and the node having a relatively higher level. The function of the PagBEE3L gene was investigated through phenotypic analyses with PagBEE3L-overexpressing (ox) transgenic lines. This work particularly focused on a potential role of PagBEE3L in stem growth and development of polar. The PagBEE3L-ox poplar showed thicker and longer stems than wild-type plants. The xylem cells from the stems of PagBEE3L-ox plants revealed remarkably enhanced proliferation, resulting in an earlier thickening growth than wild-type plants. Therefore, this work suggests that xylem development of poplar is accelerated in PagBEE3L-ox plants and PagBEE3L plays a role in stem growth by increasing the proliferation of xylem cells to promote the initial thickening growth of poplar stems. - Highlights: • We identify the BEE3-like gene form hybrid poplar (Populus alba × Populus glandulosa). • We examine effects of overexpression of PagBEE3L on growth in poplar. • We found that 35S:BEE3L transgenic plants showed more rapid growth than wild-type plants. • BEE3L protein plays an important role in the development of plant stem.

  20. Novel TetR family transcriptional factor regulates expression of multiple transport-related genes and affects rifampicin resistance in Mycobacterium smegmatis

    PubMed Central

    Liu, Huicong; Yang, Min; He, Zheng-Guo

    2016-01-01

    Transport-related genes significantly affect bacterial antibiotic resistance. However, the effects of these genes and their regulation of bacterial drug resistance in several mycobacterial species, including the fast-growing Mycobacterium smegmatis, the pathogen M. tuberculosis and M. avium have not been clearly characterized. We identified Ms4022 (MSMEG_4022) as a novel TetR family regulator that activates the expression of seven transport-related genes and affects drug resistance in M. smegmatis. Overexpression of Ms4022 inhibited M. smegmatis growth and enhanced mycobacterial resistance to the anti-tuberculosis drug rifampicin (RIF). By contrast, the Ms4022-deleted mycobacterial strain has shown sensitive to RIF. Ms4022 recognized three 19 bp non-palindromic motifs containing a 9 bp conserved region at their 5′ end and it directly regulated seven transport-related genes, which affects mycobacterial resistance to RIF. Overexpression of three of seven transport-related genes (Ms1448, Ms1613, and Ms5278) inhibited the growth of M. smegmatis. This study improves our understanding of the function of mycobacterial transport-related genes and their regulation of bacterial drug resistance. PMID:27271013

  1. Novel TetR family transcriptional factor regulates expression of multiple transport-related genes and affects rifampicin resistance in Mycobacterium smegmatis.

    PubMed

    Liu, Huicong; Yang, Min; He, Zheng-Guo

    2016-01-01

    Transport-related genes significantly affect bacterial antibiotic resistance. However, the effects of these genes and their regulation of bacterial drug resistance in several mycobacterial species, including the fast-growing Mycobacterium smegmatis, the pathogen M. tuberculosis and M. avium have not been clearly characterized. We identified Ms4022 (MSMEG_4022) as a novel TetR family regulator that activates the expression of seven transport-related genes and affects drug resistance in M. smegmatis. Overexpression of Ms4022 inhibited M. smegmatis growth and enhanced mycobacterial resistance to the anti-tuberculosis drug rifampicin (RIF). By contrast, the Ms4022-deleted mycobacterial strain has shown sensitive to RIF. Ms4022 recognized three 19 bp non-palindromic motifs containing a 9 bp conserved region at their 5' end and it directly regulated seven transport-related genes, which affects mycobacterial resistance to RIF. Overexpression of three of seven transport-related genes (Ms1448, Ms1613, and Ms5278) inhibited the growth of M. smegmatis. This study improves our understanding of the function of mycobacterial transport-related genes and their regulation of bacterial drug resistance. PMID:27271013

  2. The peptide semax affects the expression of genes related to the immune and vascular systems in rat brain focal ischemia: genome-wide transcriptional analysis

    PubMed Central

    2014-01-01

    Background The nootropic neuroprotective peptide Semax (Met-Glu-His-Phe-Pro-Gly-Pro) has proved efficient in the therapy of brain stroke; however, the molecular mechanisms underlying its action remain obscure. Our genome-wide study was designed to investigate the response of the transcriptome of ischemized rat brain cortex tissues to the action of Semax in vivo. Results The gene-expression alteration caused by the action of the peptide Semax was compared with the gene expression of the “ischemia” group animals at 3 and 24 h after permanent middle cerebral artery occlusion (pMCAO). The peptide predominantly enhanced the expression of genes related to the immune system. Three hours after pMCAO, Semax influenced the expression of some genes that affect the activity of immune cells, and, 24 h after pMCAO, the action of Semax on the immune response increased considerably. The genes implicated in this response represented over 50% of the total number of genes that exhibited Semax-induced altered expression. Among the immune-response genes, the expression of which was modulated by Semax, genes that encode immunoglobulins and chemokines formed the most notable groups. In response to Semax administration, 24 genes related to the vascular system exhibited altered expression 3 h after pMCAO, whereas 12 genes were changed 24 h after pMCAO. These genes are associated with such processes as the development and migration of endothelial tissue, the migration of smooth muscle cells, hematopoiesis, and vasculogenesis. Conclusions Semax affects several biological processes involved in the function of various systems. The immune response is the process most markedly affected by the drug. Semax altered the expression of genes that modulate the amount and mobility of immune cells and enhanced the expression of genes that encode chemokines and immunoglobulins. In conditions of rat brain focal ischemia, Semax influenced the expression of genes that promote the formation and

  3. Gene transcription and electromagnetic fields

    SciTech Connect

    Henderson, A.S.

    1992-01-01

    Our overall aim is to obtain sufficient information to allow us to ultimately determine whether ELF EM field exposure is an initiating factor in neoplastic transformation and/or if exposure can mimic characteristics of the second-step counterpart in neoplastic disease. This aim is based on our previous findings that levels of some transcripts are increased in cells exposed to EM fields. While the research is basic in nature, the ramifications have bearing on the general safety of exposure to EM fields in industrial and everyday life. A large array of diverse biological effects are reported to occur as the result of exposure to elf EM fields, suggesting that the cell response to EM fields is at a basic level, presumably initiated by molecular and/or biophysical events at the cell membrane. The hypothesized route is a signal transduction pathway involving membrane calcium fluxes. Information flow resulting from signal transduction can mediate the induction of regulatory factors in the cell, and directly affect how transcription is regulated.

  4. The ts13 mutation in the TAF(II)250 subunit (CCG1) of TFIID directly affects transcription of D-type cyclin genes in cells arrested in G1 at the nonpermissive temperature.

    PubMed Central

    Suzuki-Yagawa, Y; Guermah, M; Roeder, R G

    1997-01-01

    The general transcription initiation factor TFIID contains the TATA-binding protein (TBP) and TBP-associated factors (TAFs) implicated in the function of gene-specific activators. Previous studies have indicated that a hamster cell line (ts13) with a point mutation in the TAF(II)250/CCG1 (TAF(II)250) gene shows temperature-sensitive expression of a subset of genes and arrests in late G1 at 39.5 degrees C. Here, we report the identification of cell cycle-specific (G1-specific) genes that appear to be regulated directly through TAF(II)250 both in vivo and in vitro. Transcription rates of several cell cycle-regulatory genes were determined by run-on assays in nuclei from ts13 cells grown at permissive (33 degrees C) and nonpermissive (39.5 degrees C) temperatures. Temperature-dependent differences in transcription rates were observed for cyclin A, D1, and D3 genes. In transient-transfection assays, the human cyclin D1 promoter fused to a luciferase reporter showed a temperature-dependent reduction in activity in ts13 cells but not in parental BHK cells. In in vitro assays, upstream sequence-dependent transcription from the human cyclin D1 promoter was significantly reduced in ts13 nuclear extracts preincubated at 30 degrees C but not in similarly treated BHK nuclear extracts, and transcription in the ts13 extract was restored by addition of an affinity-purified human TFIID. Preincubation of the ts13 nuclear extracts did not affect the function of several GAL4-activation domain fusion proteins (GAL4-VP16, GAL4-p65, and GAL4-p53) on either the adenovirus major late or cyclin D1 core promoter bearing GAL4 sites, further indicating that the effect of the TAF(II)250 mutation is both core promoter and activator specific. PMID:9154827

  5. A polymorphism of the GTP-cyclohydrolase I feedback regulator gene alters transcriptional activity and may affect response to SSRI antidepressants.

    PubMed

    McHugh, P C; Joyce, P R; Deng, X; Kennedy, M A

    2011-06-01

    Tetrahydrobiopterin (BH(4)) is an essential cofactor for synthesis of many neurotransmitters including serotonin. In serotonergic neurons, BH(4) is tightly regulated by GTP-cyclohydrolase I feedback regulator (GFRP). Given the pivotal role of the serotonergic system in mood disorders and selective serotonin reuptake inhibitors (SSRIs) antidepressant function, we tested the hypothesis that GFRP gene (GCHFR) variants would modify response to antidepressants in subjects with major depression. Two single nucleotide polymorphisms (rs7164342 and rs7163862) in the GCHFR promoter were identified and occurred as two haplotypes (GA or TT). A multiple regression analysis revealed that homozygous individuals for the TT haplotype were less likely to respond to the SSRI fluoxetine than to the tricyclic antidepressant nortriptyline (P = 0.037). Moreover, the TT haplotype showed a reduced transcription rate in luciferase reporter gene assays, which may impact on BH(4)-mediated neurotransmitter production, thus suggesting a biological process through which GCHFR promoter variants might influence antidepressant response. PMID:20351752

  6. Abscisic acid represses the transcription of chloroplast genes*

    PubMed Central

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

    2013-01-01

    Numerous studies have shown effects of abscisic acid (ABA) on nuclear genes encoding chloroplast-localized proteins. ABA effects on the transcription of chloroplast genes, however, have not been investigated yet thoroughly. This work, therefore, studied the effects of ABA (75 μM) on transcription and steady-state levels of transcripts in chloroplasts of basal and apical segments of primary leaves of barley (Hordeum vulgare L.). Basal segments consist of young cells with developing chloroplasts, while apical segments contain the oldest cells with mature chloroplasts. Exogenous ABA reduced the chlorophyll content and caused changes of the endogenous concentrations not only of ABA but also of cytokinins to different extents in the basal and apical segments. It repressed transcription by the chloroplast phage-type and bacteria-type RNA polymerases and lowered transcript levels of most investigated chloroplast genes drastically. ABA did not repress the transcription of psbD and a few other genes and even increased psbD mRNA levels under certain conditions. The ABA effects on chloroplast transcription were more pronounced in basal vs. apical leaf segments and enhanced by light. Simultaneous application of cytokinin (22 μM 6-benzyladenine) minimized the ABA effects on chloroplast gene expression. These data demonstrate that ABA affects the expression of chloroplast genes differentially and points to a role of ABA in the regulation and coordination of the activities of nuclear and chloroplast genes coding for proteins with functions in photosynthesis. PMID:24078671

  7. Ethylene negatively regulates transcript abundance of ROP-GAP rheostat-encoding genes and affects apoplastic reactive oxygen species homeostasis in epicarps of cold stored apple fruits.

    PubMed

    Zermiani, Monica; Zonin, Elisabetta; Nonis, Alberto; Begheldo, Maura; Ceccato, Luca; Vezzaro, Alice; Baldan, Barbara; Trentin, Annarita; Masi, Antonio; Pegoraro, Marco; Fadanelli, Livio; Teale, William; Palme, Klaus; Quintieri, Luigi; Ruperti, Benedetto

    2015-12-01

    Apple (Malus×domestica Borkh) fruits are stored for long periods of time at low temperatures (1 °C) leading to the occurrence of physiological disorders. 'Superficial scald' of Granny Smith apples, an economically important ethylene-dependent disorder, was used as a model to study relationships among ethylene action, the regulation of the ROP-GAP rheostat, and maintenance of H2O2 homeostasis in fruits during prolonged cold exposure. The ROP-GAP rheostat is a key module for adaptation to low oxygen in Arabidopsis through Respiratory Burst NADPH Oxidase Homologs (RBOH)-mediated and ROP GTPase-dependent regulation of reactive oxygen species (ROS) homeostasis. Here, it was shown that the transcriptional expression of several components of the apple ROP-GAP machinery, including genes encoding RBOHs, ROPs, and their ancillary proteins ROP-GEFs and ROP-GAPs, is coordinately and negatively regulated by ethylene in conjunction with the progressive impairment of apoplastic H2O2 homeostatic levels. RNA sequencing analyses showed that several components of the known ROP- and ROS-associated transcriptional networks are regulated along with the ROP-GAP rheostat in response to ethylene perception. These findings may extend the role of the ROP-GAP rheostat beyond hypoxic responses and suggest that it may be a functional regulatory node involved in the integration of ethylene and ROS signalling pathways in abiotic stress. PMID:26428066

  8. Ethylene negatively regulates transcript abundance of ROP-GAP rheostat-encoding genes and affects apoplastic reactive oxygen species homeostasis in epicarps of cold stored apple fruits

    PubMed Central

    Zermiani, Monica; Zonin, Elisabetta; Nonis, Alberto; Begheldo, Maura; Ceccato, Luca; Vezzaro, Alice; Baldan, Barbara; Trentin, Annarita; Masi, Antonio; Pegoraro, Marco; Fadanelli, Livio; Teale, William; Palme, Klaus; Quintieri, Luigi; Ruperti, Benedetto

    2015-01-01

    Apple (Malus×domestica Borkh) fruits are stored for long periods of time at low temperatures (1 °C) leading to the occurrence of physiological disorders. ‘Superficial scald’ of Granny Smith apples, an economically important ethylene-dependent disorder, was used as a model to study relationships among ethylene action, the regulation of the ROP-GAP rheostat, and maintenance of H2O2 homeostasis in fruits during prolonged cold exposure. The ROP-GAP rheostat is a key module for adaptation to low oxygen in Arabidopsis through Respiratory Burst NADPH Oxidase Homologs (RBOH)-mediated and ROP GTPase-dependent regulation of reactive oxygen species (ROS) homeostasis. Here, it was shown that the transcriptional expression of several components of the apple ROP-GAP machinery, including genes encoding RBOHs, ROPs, and their ancillary proteins ROP-GEFs and ROP-GAPs, is coordinately and negatively regulated by ethylene in conjunction with the progressive impairment of apoplastic H2O2 homeostatic levels. RNA sequencing analyses showed that several components of the known ROP- and ROS-associated transcriptional networks are regulated along with the ROP-GAP rheostat in response to ethylene perception. These findings may extend the role of the ROP-GAP rheostat beyond hypoxic responses and suggest that it may be a functional regulatory node involved in the integration of ethylene and ROS signalling pathways in abiotic stress. PMID:26428066

  9. Repetitive Sequence Variations in the Promoter Region of the Adhesin-Encoding Gene sabA of Helicobacter pylori Affect Transcription

    PubMed Central

    Harvey, Vivian C.; Acio, Catherine R.; Bredehoft, Amy K.; Zhu, Laurence; Hallinger, Daniel R.; Quinlivan-Repasi, Vanessa; Harvey, Samuel E.

    2014-01-01

    The pathogenesis of diseases elicited by the gastric pathogen Helicobacter pylori is partially determined by the effectiveness of adaptation to the variably acidic environment of the host stomach. Adaptation includes appropriate adherence to the gastric epithelium via outer membrane protein adhesins such as SabA. The expression of sabA is subject to regulation via phase variation in the promoter and coding regions as well as repression by the two-component system ArsRS. In this study, we investigated the role of a homopolymeric thymine [poly(T)] tract −50 to −33 relative to the sabA transcriptional start site in H. pylori strain J99. We quantified sabA expression in H. pylori J99 by quantitative reverse transcription-PCR (RT-PCR), demonstrating significant changes in sabA expression associated with experimental manipulations of poly(T) tract length. Mimicking the length increase of this tract by adding adenines instead of thymines had similar effects, while the addition of other nucleotides failed to affect sabA expression in the same manner. We hypothesize that modification of the poly(T) tract changes DNA topology, affecting regulatory protein interaction(s) or RNA polymerase binding efficiency. Additionally, we characterized the interaction between the sabA promoter region and ArsR, a response regulator affecting sabA expression. Using recombinant ArsR in electrophoretic mobility shift assays (EMSA), we localized binding to a sequence with partial dyad symmetry −20 and +38 relative to the sabA +1 site. The control of sabA expression by both ArsRS and phase variation at two distinct repeat regions suggests the control of sabA expression is both complex and vital to H. pylori infection. PMID:25022855

  10. The two-component system CpxR/A represses the expression of Salmonella virulence genes by affecting the stability of the transcriptional regulator HilD

    PubMed Central

    De la Cruz, Miguel A.; Pérez-Morales, Deyanira; Palacios, Irene J.; Fernández-Mora, Marcos; Calva, Edmundo; Bustamante, Víctor H.

    2015-01-01

    Salmonella enterica can cause intestinal or systemic infections in humans and animals mainly by the presence of pathogenicity islands SPI-1 and SPI-2, containing 39 and 44 genes, respectively. The AraC-like regulator HilD positively controls the expression of the SPI-1 genes, as well as many other Salmonella virulence genes including those located in SPI-2. A previous report indicates that the two-component system CpxR/A regulates the SPI-1 genes: the absence of the sensor kinase CpxA, but not the absence of its cognate response regulator CpxR, reduces their expression. The presence and absence of cell envelope stress activates kinase and phosphatase activities of CpxA, respectively, which in turn controls the level of phosphorylated CpxR (CpxR-P). In this work, we further define the mechanism for the CpxR/A-mediated regulation of SPI-1 genes. The negative effect exerted by the absence of CpxA on the expression of SPI-1 genes was counteracted by the absence of CpxR or by the absence of the two enzymes, AckA and Pta, which render acetyl-phosphate that phosphorylates CpxR. Furthermore, overexpression of the lipoprotein NlpE, which activates CpxA kinase activity on CpxR, or overexpression of CpxR, repressed the expression of SPI-1 genes. Thus, our results provide several lines of evidence strongly supporting that the absence of CpxA leads to the phosphorylation of CpxR via the AckA/Pta enzymes, which represses both the SPI-1 and SPI-2 genes. Additionally, we show that in the absence of the Lon protease, which degrades HilD, the CpxR-P-mediated repression of the SPI-1 genes is mostly lost; moreover, we demonstrate that CpxR-P negatively affects the stability of HilD and thus decreases the expression of HilD-target genes, such as hilD itself and hilA, located in SPI-1. Our data further expand the insight on the different regulatory pathways for gene expression involving CpxR/A and on the complex regulatory network governing virulence in Salmonella. PMID:26300871

  11. [Development genes encoding transcription factors and dysmorphology].

    PubMed

    Lacombe, Didier

    2009-04-01

    Studies of children with developmental abnormalities of genetic origin are necessary for accurate diagnosis, prognostication, patient management, and genetic counseling. Such studies can also help to identify genes involved in normal and abnormal morphogenesis, which often act as patterning genes and are also potential oncogenes. Many encode transcription factors that regulate other genes during embryonic development. PMID:20120282

  12. Transcription of denitrification genes and kinetics of NO, N2O and N2 by soil bacteria as affected by pH

    NASA Astrophysics Data System (ADS)

    Liu, B.; Bakken, L. R.; Frostegard, A.

    2010-12-01

    Nitrous oxide (N2O), which is to a large part derived from denitrification in soil, is a major greenhouse gas and was also recently shown to be the single most important ozone-depleting substance. Previous studies demonstrate that the N2O/N2 product ratio of denitrification is strongly dependent on pH, increasing with decreasing soil pH. The mechanisms involved are, however, poorly understood. We here present an investigation of soils from a long-term liming experiment. Since it is difficult to control which pH is actually experienced by bacterial cells in intact soils, we extracted cells on a Nycodenz gradient and exposed them to different pH levels. Bacteria extracted from soils of 3 different pHs (4.0, 6.1 and 8.0) were incubated in minimal medium supplemented with nitrate (2mM) and glutamic acid (5 mM), buffered at three pH levels (5.7, 6.1 and 7.6). Both the pH of the medium and original soil pH showed profound effect on the denitrification activity in terms of gas emission kinetics. N2O reductase (N2OR) activity was only present when cells from the high pH soils (pH 6.1 and 8.0) were exposed to high pH medium (pH 7.6). Functional genes (nirS, nirK and nosZ) and their transcripts were quantified in the extracts from pH6.1-soil. A 10-25 fold higher expression of nosZ vs nirS was found when incubated at pH 7.6 compared to pH 6.1 and 5.7. The low but significant transcription of nosZ at pH 6.1 and 5.7 did not result in detectable N2O reduction however. Cells that had been allowed to assemble their proteome while growing in pH7 medium showed N2OR activity which was practically unaffected by pH within the range 5-7. On the contrary, no N2OR activity was detected if the proteome had been formed at pH 6. The cells extracted from acid soils (pH 5.8 and 6.1) showed very low nosZ transcritption and no N2OR activity if exposed to pH 7 during the transition from oxic to anoxic conditions, suggesting an adaptation to low pH in the sense that they do not transcribe the gene

  13. OsARF12, a transcription activator on auxin response gene, regulates root elongation and affects iron accumulation in rice (Oryza sativa).

    PubMed

    Qi, YanHua; Wang, SuiKang; Shen, ChenJia; Zhang, SaiNa; Chen, Yue; Xu, YanXia; Liu, Yu; Wu, YunRong; Jiang, DeAn

    2012-01-01

    • Auxin has an important role in maintaining optimal root system architecture (RSA) that can cope with growth reductions of crops caused by water or nutrient shortages. However, the mechanism of controlling RSA remains largely unclear. Here, we found a limiting factor of RSA--OsARF12--an auxin response factor whose knockout led to decreased primary root length in rice (Oryza sativa). • OsARF12 as a transcription activator can facilitate the expression of the auxin response element DR5::GFP, and OsARF12 was inhibited by osa-miRNA167d by transient expression in tobacco and rice callus. • The root elongation zones of osarf12 and osarf12/25, which had lower auxin concentrations, were distinctly shorter than for the wild-type, possibly as a result of decreased expression of auxin synthesis genes OsYUCCAs and auxin efflux carriers OsPINs and OsPGPs. The knockout of OsARF12 also altered the abundance of mitochondrial iron-regulated (OsMIR), iron (Fe)-regulated transporter1 (OsIRT1) and short postembryonic root1 (OsSPR1) in roots of rice, and resulted in lower Fe content. • The data provide evidence for the biological function of OsARF12, which is implicated in regulating root elongation. Our investigation contributes a novel insight for uncovering regulation of RSA and the relationship between auxin response and Fe acquisition. PMID:21973088

  14. Does every transcript originate from a gene?

    PubMed

    Raabe, Carsten A; Brosius, Jürgen

    2015-04-01

    Outdated gene definitions favored regions corresponding to mature messenger RNAs, in particular, the open reading frame. In eukaryotes, the intergenic space was widely regarded nonfunctional and devoid of RNA transcription. Original concepts were based on the assumption that RNA expression was restricted to known protein-coding genes and a few so-called structural RNA genes, such as ribosomal RNAs or transfer RNAs. With the discovery of introns and, more recently, sensitive techniques for monitoring genome-wide transcription, this view had to be substantially modified. Tiling microarrays and RNA deep sequencing revealed myriads of transcripts, which cover almost entire genomes. The tremendous complexity of non-protein-coding RNA transcription has to be integrated into novel gene definitions. Despite an ever-growing list of functional RNAs, questions concerning the mass of identified transcripts are under dispute. Here, we examined genome-wide transcription from various angles, including evolutionary considerations, and suggest, in analogy to novel alternative splice variants that do not persist, that the vast majority of transcripts represent raw material for potential, albeit rare, exaptation events. PMID:25847549

  15. Methylation Affects Transposition and Splicing of a Large CACTA Transposon from a MYB Transcription Factor Regulating Anthocyanin Synthase Genes in Soybean Seed Coats

    PubMed Central

    Zabala, Gracia; Vodkin, Lila O.

    2014-01-01

    We determined the molecular basis of three soybean lines that vary in seed coat color at the R locus which is thought to encode a MYB transcription factor. RM55-rm is homozygous for a mutable allele (rm) that specifies black and brown striped seeds; RM30-R* is a stable black revertant isoline derived from the mutable line; and RM38-r has brown seed coats due to a recessive r allele shown to translate a truncated MYB protein. Using long range PCR, 454 sequencing of amplicons, and whole genome re-sequencing, we determined that the variegated RM55-rm line had a 13 kb CACTA subfamily transposon insertion (designated TgmR*) at a position 110 bp from the beginning of Intron2 of the R locus, Glyma09g36983. Although the MYB encoded by R was expressed at only very low levels in older seed coats of the black revertant RM30-R* line, it upregulated expression of anthocyanidin synthase genes (ANS2, ANS3) to promote the synthesis of anthocyanins. Surprisingly, the RM30-R* revertant also carried the 13 kb TgmR* insertion in Intron2. Using RNA-Seq, we showed that intron splicing was accurate, albeit at lower levels, despite the presence of the 13 kb TgmR* element. As determined by whole genome methylation sequencing, we demonstrate that the TgmR* sequence was relatively more methylated in RM30-R* than in the mutable RM55-rm progenitor line. The stabilized and more methylated RM30-R* revertant line apparently lacks effective binding of a transposae to its subterminal repeats, thus allowing intron splicing to proceed resulting in sufficient MYB protein to stimulate anthocyanin production and thus black seed coats. In this regard, the TgmR* element in soybean resembles McClintock's Spm-suppressible and change-of-state alleles of maize. This comparison explains the opposite effects of the TgmR* element on intron splicing of the MYB gene in which it resides depending on the methylation state of the element. PMID:25369033

  16. Transcriptional regulation of secretin gene expression.

    PubMed

    Nishitani, J; Rindi, G; Lopez, M J; Upchurch, B H; Leiter, A B

    1995-01-01

    Expression of the gene encoding the hormone secretin is restricted to a specific enteroendocrine cell type and to beta-cells in developing pancreatic islets. To characterize regulatory elements in the secretin gene responsible for its expression in secretin-producing cells, we used a series of reporter genes for transient expression assays in transfection studies carried out in secretin-producing islet cell lines. Analysis of the transcriptional activity of deletion mutants identified a positive cis regulatory domain between 174 and 53 base pairs upstream from the transcriptional initiation site which was required for secretin gene expression in secretin-producing HIT insulinoma cells. Within this enhancer were sequences resembling two binding sites for the transcription factor Sp1, as well as a consensus sequence for binding to helix-loop-helix proteins. Analysis of these three elements by site-directed mutagenesis suggests that each is important for full transcriptional activity. The role of proximal enhancer sequences in directing secretin gene expression to appropriate tissues is further supported by studies in transgenic mice revealing that 1.6 kilobases of the secretin gene 5' flanking sequence were sufficient to direct the expression of either human growth hormone or simian virus 40 large T-antigen reporter genes to all major secretin-producing tissues. PMID:8774991

  17. Transcriptional Control of the TNF Gene

    PubMed Central

    Falvo, James V.; Tsytsykova, Alla V.; Goldfeld, Anne E.

    2016-01-01

    The cytokine TNF is a critical mediator of immune and inflammatory responses. The TNF gene is an immediate early gene, rapidly transcribed in a variety of cell types following exposure to a broad range of pathogens and signals of inflammation and stress. Regulation of TNF gene expression at the transcriptional level is cell type- and stimulus-specific, involving the recruitment of distinct sets of transcription factors to a compact and modular promoter region. In this review, we describe our current understanding of the mechanisms through which TNF transcription is specifically activated by a variety of extracellular stimuli in multiple cell types, including T cells, B cells, macrophages, mast cells, dendritic cells, and fibroblasts. We discuss the role of nuclear factor of activated T cells and other transcription factors and coactivators in enhanceosome formation, as well as the contradictory evidence for a role for nuclear factor κB as a classical activator of the TNF gene. We describe the impact of evolutionarily conserved cis-regulatory DNA motifs in the TNF locus upon TNF gene transcription, in contrast to the neutral effect of single nucleotide polymorphisms. We also assess the regulatory role of chromatin organization, epigenetic modifications, and long-range chromosomal interactions at the TNF locus. PMID:20173386

  18. Topologies for perfect adaptation in gene transcription

    NASA Astrophysics Data System (ADS)

    Shi, Wenjia; Tang, Chao

    2014-03-01

    Adaptation is commonly used in sensory systems and signaling networks to allow the detection of further stimuli. Despite enzymatic network topologies for adaptation have been investigated systematically, the topology of transcriptional network that could perform adaptation still remains unclear, due to the complexity of transcriptional regulation. Here, we systematically investigated all three-node transcriptional networks, and found the topologies of transcriptional networks for adaptation are different from that of enzymatic ones. While both negative feedback loop (NFBL) and incoherent feed forward loop (IFFL) are capable of performing adaptation analytically, a positive self-regulation on buffer node is necessary for NFBL topology and more flexible structures emerge for IFFL than that of enzymatic networks. Most of the simulation results agree with analytical predictions. This study may explain the mechanism of adapted gene regulation behavior and supply a design table for gene regulatory adaptation.

  19. Aeromonas hydrophila Lateral Flagellar Gene Transcriptional Hierarchy

    PubMed Central

    Wilhelms, Markus; Gonzalez, Victor; Merino, Susana

    2013-01-01

    Aeromonas hydrophila AH-3 lateral flagella are not assembled when bacteria grow in liquid media; however, lateral flagellar genes are transcribed. Our results indicate that A. hydrophila lateral flagellar genes are transcribed at three levels (class I to III genes) and share some similarities with, but have many important differences from, genes of Vibrio parahaemolyticus. A. hydrophila lateral flagellum class I gene transcription is σ70 dependent, which is consistent with the fact that lateral flagellum is constitutively transcribed, in contrast to the characteristics of V. parahaemolyticus. The fact that multiple genes are included in class I highlights that lateral flagellar genes are less hierarchically transcribed than polar flagellum genes. The A. hydrophila lafK-fliEJL gene cluster (where the subscript L distinguishes genes for lateral flagella from those for polar flagella) is exclusively from class I and is in V. parahaemolyticus class I and II. Furthermore, the A. hydrophila flgAMNL cluster is not transcribed from the σ54/LafK-dependent promoter and does not contain class II genes. Here, we propose a gene transcriptional hierarchy for the A. hydrophila lateral flagella. PMID:23335410

  20. Synaptic, transcriptional, and chromatin genes disrupted in autism

    PubMed Central

    De Rubeis, Silvia; He, Xin; Goldberg, Arthur P.; Poultney, Christopher S.; Samocha, Kaitlin; Cicek, A Ercument; Kou, Yan; Liu, Li; Fromer, Menachem; Walker, Susan; Singh, Tarjinder; Klei, Lambertus; Kosmicki, Jack; Fu, Shih-Chen; Aleksic, Branko; Biscaldi, Monica; Bolton, Patrick F.; Brownfeld, Jessica M.; Cai, Jinlu; Campbell, Nicholas J.; Carracedo, Angel; Chahrour, Maria H.; Chiocchetti, Andreas G.; Coon, Hilary; Crawford, Emily L.; Crooks, Lucy; Curran, Sarah R.; Dawson, Geraldine; Duketis, Eftichia; Fernandez, Bridget A.; Gallagher, Louise; Geller, Evan; Guter, Stephen J.; Hill, R. Sean; Ionita-Laza, Iuliana; Gonzalez, Patricia Jimenez; Kilpinen, Helena; Klauck, Sabine M.; Kolevzon, Alexander; Lee, Irene; Lei, Jing; Lehtimäki, Terho; Lin, Chiao-Feng; Ma'ayan, Avi; Marshall, Christian R.; McInnes, Alison L.; Neale, Benjamin; Owen, Michael J.; Ozaki, Norio; Parellada, Mara; Parr, Jeremy R.; Purcell, Shaun; Puura, Kaija; Rajagopalan, Deepthi; Rehnström, Karola; Reichenberg, Abraham; Sabo, Aniko; Sachse, Michael; Sanders, Stephan J.; Schafer, Chad; Schulte-Rüther, Martin; Skuse, David; Stevens, Christine; Szatmari, Peter; Tammimies, Kristiina; Valladares, Otto; Voran, Annette; Wang, Li-San; Weiss, Lauren A.; Willsey, A. Jeremy; Yu, Timothy W.; Yuen, Ryan K.C.; Cook, Edwin H.; Freitag, Christine M.; Gill, Michael; Hultman, Christina M.; Lehner, Thomas; Palotie, Aarno; Schellenberg, Gerard D.; Sklar, Pamela; State, Matthew W.; Sutcliffe, James S.; Walsh, Christopher A.; Scherer, Stephen W.; Zwick, Michael E.; Barrett, Jeffrey C.; Cutler, David J.; Roeder, Kathryn; Devlin, Bernie; Daly, Mark J.; Buxbaum, Joseph D.

    2014-01-01

    Summary The genetic architecture of autism spectrum disorder involves the interplay of common and rare variation and their impact on hundreds of genes. Using exome sequencing, analysis of rare coding variation in 3,871 autism cases and 9,937 ancestry-matched or parental controls implicates 22 autosomal genes at a false discovery rate (FDR) < 0.05, and a set of 107 autosomal genes strongly enriched for those likely to affect risk (FDR < 0.30). These 107 genes, which show unusual evolutionary constraint against mutations, incur de novo loss-of-function mutations in over 5% of autistic subjects. Many of the genes implicated encode proteins for synaptic, transcriptional, and chromatin remodeling pathways. These include voltage-gated ion channels regulating propagation of action potentials, pacemaking, and excitability-transcription coupling, as well as histone-modifying enzymes and chromatin remodelers, prominently histone post-translational modifications involving lysine methylation/demethylation. PMID:25363760

  1. Poly(ADP-Ribosyl)ation Affects Histone Acetylation and Transcription

    PubMed Central

    Verdone, Loredana; La Fortezza, Marco; Ciccarone, Fabio; Caiafa, Paola; Zampieri, Michele; Caserta, Micaela

    2015-01-01

    Poly(ADP-ribosyl)ation (PARylation) is a posttranslational protein modification catalyzed by members of the poly(ADP-ribose) polymerase (PARP) enzyme family. PARylation regulates a wide variety of biological processes in most eukaryotic cells including energy metabolism and cell death, maintenance of genomic stability, chromatin structure and transcription. Inside the nucleus, cross-talk between PARylation and other epigenetic modifications, such as DNA and histone methylation, was already described. In the present work, using PJ34 or ABT888 to inhibit PARP activity or over-expressing poly(ADP-ribose) glycohydrolase (PARG), we show decrease of global histone H3 and H4 acetylation. This effect is accompanied by a reduction of the steady state mRNA level of p300, Pcaf, and Tnfα, but not of Dnmt1. Chromatin immunoprecipitation (ChIP) analyses, performed at the level of the Transcription Start Site (TSS) of these four genes, reveal that changes in histone acetylation are specific for each promoter. Finally, we demonstrate an increase of global deacetylase activity in nuclear extracts from cells treated with PJ34, whereas global acetyltransferase activity is not affected, suggesting a role for PARP in the inhibition of histone deacetylases. Taken together, these results show an important link between PARylation and histone acetylation regulated transcription. PMID:26636673

  2. Transcriptional enhancer from milk protein genes

    SciTech Connect

    Casperson, Gerald F.; Schmidhauser, Christian T.; Bissell, Mina J.

    1999-01-01

    The invention relates to novel enhancer nucleotide sequences which stimulate transcription of heterologous DNA in cells in culture. The enhancers are derived from major milk protein genes by the process of deletion mapping and functional analysis. The invention also relates to expression vectors containing the novel enhancers.

  3. Transcriptional enhancer from milk protein genes

    SciTech Connect

    Casperson, G.F.; Schmidhauser, C.T.; Bissell, M.J.

    1999-12-21

    The invention relates to novel enhancer nucleotide sequences which stimulate transcription of heterologous DNA in cells in culture. The enhancers are derived from major milk protein genes by the process of deletion mapping and functional analysis. The invention also relates to expression vectors containing the novel enhancers.

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

    SciTech Connect

    Tennyson, C.N.; Worton, R.G.

    1994-09-01

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

  5. Modular composition of gene transcription networks.

    PubMed

    Gyorgy, Andras; Del Vecchio, Domitilla

    2014-03-01

    Predicting the dynamic behavior of a large network from that of the composing modules is a central problem in systems and synthetic biology. Yet, this predictive ability is still largely missing because modules display context-dependent behavior. One cause of context-dependence is retroactivity, a phenomenon similar to loading that influences in non-trivial ways the dynamic performance of a module upon connection to other modules. Here, we establish an analysis framework for gene transcription networks that explicitly accounts for retroactivity. Specifically, a module's key properties are encoded by three retroactivity matrices: internal, scaling, and mixing retroactivity. All of them have a physical interpretation and can be computed from macroscopic parameters (dissociation constants and promoter concentrations) and from the modules' topology. The internal retroactivity quantifies the effect of intramodular connections on an isolated module's dynamics. The scaling and mixing retroactivity establish how intermodular connections change the dynamics of connected modules. Based on these matrices and on the dynamics of modules in isolation, we can accurately predict how loading will affect the behavior of an arbitrary interconnection of modules. We illustrate implications of internal, scaling, and mixing retroactivity on the performance of recurrent network motifs, including negative autoregulation, combinatorial regulation, two-gene clocks, the toggle switch, and the single-input motif. We further provide a quantitative metric that determines how robust the dynamic behavior of a module is to interconnection with other modules. This metric can be employed both to evaluate the extent of modularity of natural networks and to establish concrete design guidelines to minimize retroactivity between modules in synthetic systems. PMID:24626132

  6. Modular Composition of Gene Transcription Networks

    PubMed Central

    Gyorgy, Andras; Del Vecchio, Domitilla

    2014-01-01

    Predicting the dynamic behavior of a large network from that of the composing modules is a central problem in systems and synthetic biology. Yet, this predictive ability is still largely missing because modules display context-dependent behavior. One cause of context-dependence is retroactivity, a phenomenon similar to loading that influences in non-trivial ways the dynamic performance of a module upon connection to other modules. Here, we establish an analysis framework for gene transcription networks that explicitly accounts for retroactivity. Specifically, a module's key properties are encoded by three retroactivity matrices: internal, scaling, and mixing retroactivity. All of them have a physical interpretation and can be computed from macroscopic parameters (dissociation constants and promoter concentrations) and from the modules' topology. The internal retroactivity quantifies the effect of intramodular connections on an isolated module's dynamics. The scaling and mixing retroactivity establish how intermodular connections change the dynamics of connected modules. Based on these matrices and on the dynamics of modules in isolation, we can accurately predict how loading will affect the behavior of an arbitrary interconnection of modules. We illustrate implications of internal, scaling, and mixing retroactivity on the performance of recurrent network motifs, including negative autoregulation, combinatorial regulation, two-gene clocks, the toggle switch, and the single-input motif. We further provide a quantitative metric that determines how robust the dynamic behavior of a module is to interconnection with other modules. This metric can be employed both to evaluate the extent of modularity of natural networks and to establish concrete design guidelines to minimize retroactivity between modules in synthetic systems. PMID:24626132

  7. Asymmetric Regulation of Peripheral Genes by Two Transcriptional Regulatory Networks

    PubMed Central

    Li, Jing-Ru; Suzuki, Takahiro; Nishimura, Hajime; Kishima, Mami; Maeda, Shiori; Suzuki, Harukazu

    2016-01-01

    Transcriptional regulatory network (TRN) reconstitution and deconstruction occur simultaneously during reprogramming; however, it remains unclear how the starting and targeting TRNs regulate the induction and suppression of peripheral genes. Here we analyzed the regulation using direct cell reprogramming from human dermal fibroblasts to monocytes as the platform. We simultaneously deconstructed fibroblastic TRN and reconstituted monocytic TRN; monocytic and fibroblastic gene expression were analyzed in comparison with that of fibroblastic TRN deconstruction only or monocytic TRN reconstitution only. Global gene expression analysis showed cross-regulation of TRNs. Detailed analysis revealed that knocking down fibroblastic TRN positively affected half of the upregulated monocytic genes, indicating that intrinsic fibroblastic TRN interfered with the expression of induced genes. In contrast, reconstitution of monocytic TRN showed neutral effects on the majority of fibroblastic gene downregulation. This study provides an explicit example that demonstrates how two networks together regulate gene expression during cell reprogramming processes and contributes to the elaborate exploration of TRNs. PMID:27483142

  8. Gene duplication of type-B ARR transcription factors systematically extends transcriptional regulatory structures in Arabidopsis

    PubMed Central

    Choi, Seung Hee; Hyeon, Do Young; Lee, ll Hwan; Park, Su Jin; Han, Seungmin; Lee, In Chul; Hwang, Daehee; Nam, Hong Gil

    2014-01-01

    Many of duplicated genes are enriched in signaling pathways. Recently, gene duplication of kinases has been shown to provide genetic buffering and functional diversification in cellular signaling. Transcription factors (TFs) are also often duplicated. However, how duplication of TFs affects their regulatory structures and functions of target genes has not been explored at the systems level. Here, we examined regulatory and functional roles of duplication of three major ARR TFs (ARR1, 10, and 12) in Arabidopsis cytokinin signaling using wild-type and single, double, and triple deletion mutants of the TFs. Comparative analysis of gene expression profiles obtained from Arabidopsis roots in wild-type and these mutants showed that duplication of ARR TFs systematically extended their transcriptional regulatory structures, leading to enhanced robustness and diversification in functions of target genes, as well as in regulation of cellular networks of target genes. Therefore, our results suggest that duplication of TFs contributes to robustness and diversification in functions of target genes by extending transcriptional regulatory structures. PMID:25425016

  9. Transcriptional analysis of human survivin gene expression.

    PubMed Central

    Li, F; Altieri, D C

    1999-01-01

    The preservation of tissue and organ homoeostasis depends on the regulated expression of genes controlling apoptosis (programmed cell death). In this study, we have investigated the basal transcriptional requirements of the survivin gene, an IAP (inhibitor of apoptosis) prominently up-regulated in cancer. Analysis of the 5' flanking region of the human survivin gene revealed the presence of a TATA-less promoter containing a canonical CpG island of approximately 250 nt, three cell cycle dependent elements, one cell cycle homology region and numerous Sp1 sites. PCR-based analysis of human genomic DNA, digested with methylation-sensitive and -insensitive restriction enzymes, indicated that the CpG island was unmethylated in both normal and neoplastic tissues. Primer extension and S1 nuclease mapping of the human survivin gene identified two main transcription start sites at position -72 and within -57/-61 from the initiating ATG. Transfection of cervical carcinoma HeLa cells with truncated or nested survivin promoter-luciferase constructs revealed the presence of both enhancer and repressor sequences and identified a minimal promoter region within the proximal -230 nt of the human survivin gene. Unbiased mutagenesis analysis of the human survivin promoter revealed that targeting the Sp1 sequences at position -171 and -151 abolished basal transcriptional activity by approximately 63-82%. Electrophoretic mobility-shift assay with DNA oligonucleotides confirmed formation of a DNA-protein complex between the survivin Sp1 sequences and HeLa cell extracts in a reaction abolished by mutagenesis of the survivin Sp1 sites. These findings identify the basal transcriptional requirements of survivin gene expression. PMID:10567210

  10. Transcriptional regulation of the human biglycan gene.

    PubMed

    Ungefroren, H; Krull, N B

    1996-06-28

    The small leucine-rich proteoglycan biglycan is involved in several physiological and pathophysiological processes through the ability of its core protein to interact with other extracellular matrix molecules and transforming growth factor-beta (TGF-beta). To learn more about the regulation of biglycan core protein expression, we have cloned and sequenced 1218 base pairs from the 5'-flanking region of the human biglycan gene, demonstrated functional promoter activity, and investigated the molecular mechanisms through which various agents modulate its transcriptional activity. Sequencing revealed the presence of several cis-acting elements including multiple AP-2 sites and interleukin-6 response elements, a NF-kappaB site, a TGF-beta negative element, and an E-box. The TATA and CAAT box-lacking promoter possesses many features of a growth-related gene, e.g. a GC-rich immediate 5' region, many Sp1 sites, and the use of multiple transcriptional start sites. Transient transfections of the tumor cell lines MG-63, SK-UT-1, and T47D with various biglycan 5'-flanking region-luciferase reporter gene constructs showed that the proximal 78 base pairs are sufficient for full promoter activity. Several agents among them interleukin-6, and tumor necrosis factor-alpha. were capable of altering biglycan promoter activity. However, in MG-63 cells, TGF-beta1 failed to increase either activity of the biglycan promoter constructs or specific transcription from the endogenous biglycan gene. Since TGF-beta1 also did not alter the stability of cytoplasmic biglycan mRNA as determined from Northern analysis after inhibition of transcription with 5,6-dichloro-1beta-D-ribofuranosylbenzimidazole, an as yet unidentified nuclear post-transcriptional mechanism was considered responsible for the TGF-beta effect in this cell type. These results might help to elucidate the molecular pathways leading to pathological alterations of biglycan expression observed in atherosclerosis, glomerulonephritis

  11. Regulation of gene transcription by Polycomb proteins

    PubMed Central

    Aranda, Sergi; Mas, Gloria; Di Croce, Luciano

    2015-01-01

    The Polycomb group (PcG) of proteins defines a subset of factors that physically associate and function to maintain the positional identity of cells from the embryo to adult stages. PcG has long been considered a paradigmatic model for epigenetic maintenance of gene transcription programs. Despite intensive research efforts to unveil the molecular mechanisms of action of PcG proteins, several fundamental questions remain unresolved: How many different PcG complexes exist in mammalian cells? How are PcG complexes targeted to specific loci? How does PcG regulate transcription? In this review, we discuss the diversity of PcG complexes in mammalian cells, examine newly identified modes of recruitment to chromatin, and highlight the latest insights into the molecular mechanisms underlying the function of PcGs in transcription regulation and three-dimensional chromatin conformation. PMID:26665172

  12. Intersecting transcription networks constrain gene regulatory evolution.

    PubMed

    Sorrells, Trevor R; Booth, Lauren N; Tuch, Brian B; Johnson, Alexander D

    2015-07-16

    Epistasis-the non-additive interactions between different genetic loci-constrains evolutionary pathways, blocking some and permitting others. For biological networks such as transcription circuits, the nature of these constraints and their consequences are largely unknown. Here we describe the evolutionary pathways of a transcription network that controls the response to mating pheromone in yeast. A component of this network, the transcription regulator Ste12, has evolved two different modes of binding to a set of its target genes. In one group of species, Ste12 binds to specific DNA binding sites, while in another lineage it occupies DNA indirectly, relying on a second transcription regulator to recognize DNA. We show, through the construction of various possible evolutionary intermediates, that evolution of the direct mode of DNA binding was not directly accessible to the ancestor. Instead, it was contingent on a lineage-specific change to an overlapping transcription network with a different function, the specification of cell type. These results show that analysing and predicting the evolution of cis-regulatory regions requires an understanding of their positions in overlapping networks, as this placement constrains the available evolutionary pathways. PMID:26153861

  13. Intersecting transcription networks constrain gene regulatory evolution

    PubMed Central

    Sorrells, Trevor R; Booth, Lauren N; Tuch, Brian B; Johnson, Alexander D

    2015-01-01

    Epistasis—the non-additive interactions between different genetic loci—constrains evolutionary pathways, blocking some and permitting others1–8. For biological networks such as transcription circuits, the nature of these constraints and their consequences are largely unknown. Here we describe the evolutionary pathways of a transcription network that controls the response to mating pheromone in yeasts9. A component of this network, the transcription regulator Ste12, has evolved two different modes of binding to a set of its target genes. In one group of species, Ste12 binds to specific DNA binding sites, while in another lineage it occupies DNA indirectly, relying on a second transcription regulator to recognize DNA. We show, through the construction of various possible evolutionary intermediates, that evolution of the direct mode of DNA binding was not directly accessible to the ancestor. Instead, it was contingent on a lineage-specific change to an overlapping transcription network with a different function, the specification of cell type. These results show that analyzing and predicting the evolution of cis-regulatory regions requires an understanding of their positions in overlapping networks, as this placement constrains the available evolutionary pathways. PMID:26153861

  14. Involvement of GATA transcription factors in the regulation of endogenous bovine interferon-tau gene transcription.

    PubMed

    Bai, Hanako; Sakurai, Toshihiro; Kim, Min-Su; Muroi, Yoshikage; Ideta, Atsushi; Aoyagi, Yoshito; Nakajima, Hiromi; Takahashi, Masashi; Nagaoka, Kentaro; Imakawa, Kazuhiko

    2009-12-01

    Expression of interferon-tau (IFNT), necessary for pregnancy establishment in ruminant ungulates, is regulated in a temporal and spatial manner. However, molecular mechanisms by which IFNT gene transcription is regulated in this manner have not been firmly established. In this study, DNA microarray/RT-PCR analysis between bovine trophoblast CT-1 and Mardin-Darby bovine kidney (MDBK) cells was initially performed, finding that transcription factors GATA2, GATA3, and GATA6 mRNAs were specific to CT-1 cells. These mRNAs were also found in Days 17, 20, and 22 (Day 0 = day of estrus) bovine conceptuses. In examining other bovine cell lines, ovary cumulus granulosa (oCG) and ear fibroblast (EF) cells, GATA2 and GATA3, but not GATA6, were found specific to the bovine trophoblast cells. In transient transfection analyses using the upstream region (-631 to +59 bp) of bovine IFNT gene (bIFNT, IFN-tau-c1), over-expression of GATA2/GATA3 did not affect the transcription of bIFNT-reporter construct in human choriocarcinoma JEG3 cells. Transfection of GATA2, GATA3, ETS2, and/or CDX2, however, was effective in the up-regulation of the bIFNT construct transfected into bovine oCG and EF cells. One Point mutation studies revealed that among six potential GATA binding sites located on the upstream region of the bIFNT gene, the one next to ETS2 site exhibited reduced luciferase activity. In CT-1 cells, endogenous bIFNT gene transcription was up-regulated by over-expression of GATA2 or GATA3, but down-regulated by siRNA specific to GATA2 mRNA. These data suggest that GATA2/3 is involved in trophoblast-specific regulation of bIFNT gene transcription. PMID:19598245

  15. Investigation of factors affecting RNA-seq gene expression calls

    PubMed Central

    Harati, Sahar; Phan, John H.; Wang, May D.

    2016-01-01

    RNA-seq enables quantification of the human transcriptome. Estimation of gene expression is a fundamental issue in the analysis of RNA-seq data. However, there is an inherent ambiguity in distinguishing between genes with very low expression and experimental or transcriptional noise. We conducted an exploratory investigation of some factors that may affect gene expression calls. We observed that the distribution of reads that map to exonic, intronic, and intergenic regions are distinct. These distributions may provide useful insights into the behavior of gene expression noise. Moreover, we observed that these distributions are qualitatively similar between two sequence mapping algorithms. Finally, we examined the relationship between gene length and gene expression calls, and observed that they are correlated. This preliminary investigation is important for RNA-seq gene expression analysis because it may lead to more effective algorithms for distinguishing between true gene expression and experimental or transcriptional noise. PMID:25571173

  16. Circadian and feeding rhythms differentially affect rhythmic mRNA transcription and translation in mouse liver

    PubMed Central

    Atger, Florian; Gobet, Cédric; Marquis, Julien; Martin, Eva; Wang, Jingkui; Weger, Benjamin; Lefebvre, Grégory; Descombes, Patrick; Naef, Felix; Gachon, Frédéric

    2015-01-01

    Diurnal oscillations of gene expression are a hallmark of rhythmic physiology across most living organisms. Such oscillations are controlled by the interplay between the circadian clock and feeding rhythms. Although rhythmic mRNA accumulation has been extensively studied, comparatively less is known about their transcription and translation. Here, we quantified simultaneously temporal transcription, accumulation, and translation of mouse liver mRNAs under physiological light–dark conditions and ad libitum or night-restricted feeding in WT and brain and muscle Arnt-like 1 (Bmal1)-deficient animals. We found that rhythmic transcription predominantly drives rhythmic mRNA accumulation and translation for a majority of genes. Comparison of wild-type and Bmal1 KO mice shows that circadian clock and feeding rhythms have broad impact on rhythmic gene expression, Bmal1 deletion affecting surprisingly both transcriptional and posttranscriptional levels. Translation efficiency is differentially regulated during the diurnal cycle for genes with 5′-Terminal Oligo Pyrimidine tract (5′-TOP) sequences and for genes involved in mitochondrial activity, many harboring a Translation Initiator of Short 5′-UTR (TISU) motif. The increased translation efficiency of 5′-TOP and TISU genes is mainly driven by feeding rhythms but Bmal1 deletion also affects amplitude and phase of translation, including TISU genes. Together this study emphasizes the complex interconnections between circadian and feeding rhythms at several steps ultimately determining rhythmic gene expression and translation. PMID:26554015

  17. Imaging transcription dynamics at endogenous genes in living Drosophila tissues.

    PubMed

    Yao, Jie; Zobeck, Katie L; Lis, John T; Webb, Watt W

    2008-07-01

    How transcription of individual genes is regulated in a single, intact, three-dimensionally organized cell nucleus remains mysterious. Recently, live cell imaging has become an essential tool to dissect the in vivo mechanisms of gene transcription. It not only examines functions of transcription factors at their gene targets within the chromatin context, but it also provides a non-disruptive approach for observing the dynamics of a transcription cycle in real time. However, the identification of any endogenous gene loci and their associated transcription factors remains technically difficult. Here, we describe the method of imaging the transcriptional dynamics of heat shock genes in Drosophila polytene chromosomes in living salivary gland tissues by multiphoton microscopy (MPM). This method has provided the experimental capability to visualize the assembly and dynamics of individual transcription factors and regulators and to dissect their functions at their endogenous gene targets in living cells. PMID:18586105

  18. Transcriptional Characterization of Porcine Leptin and Leptin Receptor Genes.

    PubMed

    Pérez-Montarelo, Dafne; Fernández, Almudena; Barragán, Carmen; Noguera, Jose L; Folch, Josep M; Rodríguez, M Carmen; Ovilo, Cristina; Silió, Luis; Fernández, Ana I

    2013-01-01

    The leptin (LEP) and its receptor (LEPR) regulate food intake and energy balance through hypothalamic signaling. However, the LEP-LEPR axis seems to be more complex and its expression regulation has not been well described. In pigs, LEP and LEPR genes have been widely studied due to their relevance. Previous studies reported significant effects of SNPs located in both genes on growth and fatness traits. The aim of this study was to determine the expression profiles of LEP and LEPR across hypothalamic, adipose, hepatic and muscle tissues in Iberian x Landrace backcrossed pigs and to analyze the effects of gene variants on transcript abundance. To our knowledge, non porcine LEPR isoforms have been described rather than LEPRb. A short porcine LEPR isoform (LEPRa), that encodes a protein lacking the intracellular residues responsible of signal transduction, has been identified for the first time. The LEPRb isoform was only quantifiable in hypothalamus while LEPRa appeared widely expressed across tissues, but at higher levels in liver, suggesting that both isoforms would develop different roles. The unique LEP transcript showed expression in backfat and muscle. The effects of gene variants on transcript expression revealed interesting results. The LEPRc.1987C>T polymorphism showed opposite effects on LEPRb and LEPRa hypothalamic expression. In addition, one out of the 16 polymorphisms identified in the LEPR promoter region revealed high differential expression in hepatic LEPRa. These results suggest a LEPR isoform-specific regulation at tissue level. Conversely, non-differential expression of LEP conditional on the analyzed polymorphisms could be detected, indicating that its regulation is likely affected by other mechanisms rather than gene sequence variants. The present study has allowed a transcriptional characterization of LEP and LEPR isoforms on a range of tissues. Their expression patterns seem to indicate that both molecules develop peripheral roles apart from

  19. Post-transcriptional gene silencing, transcriptional gene silencing and human immunodeficiency virus

    PubMed Central

    Méndez, Catalina; Ahlenstiel, Chantelle L; Kelleher, Anthony D

    2015-01-01

    While human immunodeficiency virus 1 (HIV-1) infection is controlled through continuous, life-long use of a combination of drugs targeting different steps of the virus cycle, HIV-1 is never completely eradicated from the body. Despite decades of research there is still no effective vaccine to prevent HIV-1 infection. Therefore, the possibility of an RNA interference (RNAi)-based cure has become an increasingly explored approach. Endogenous gene expression is controlled at both, transcriptional and post-transcriptional levels by non-coding RNAs, which act through diverse molecular mechanisms including RNAi. RNAi has the potential to control the turning on/off of specific genes through transcriptional gene silencing (TGS), as well as fine-tuning their expression through post-transcriptional gene silencing (PTGS). In this review we will describe in detail the canonical RNAi pathways for PTGS and TGS, the relationship of TGS with other silencing mechanisms and will discuss a variety of approaches developed to suppress HIV-1 via manipulation of RNAi. We will briefly compare RNAi strategies against other approaches developed to target the virus, highlighting their potential to overcome the major obstacle to finding a cure, which is the specific targeting of the HIV-1 reservoir within latently infected cells. PMID:26279984

  20. Trans-Reactivation: A New Epigenetic Phenomenon Underlying Transcriptional Reactivation of Silenced Genes

    PubMed Central

    Cavalieri, Vincenzo; Ingrassia, Antonia M. R.; Pavesi, Giulio; Corona, Davide F. V.

    2015-01-01

    In order to study the role played by cellular RNA pools produced by homologous genomic loci in defining the transcriptional state of a silenced gene, we tested the effect of non-functional alleles of the white gene in the presence of a functional copy of white, silenced by heterochromatin. We found that non-functional alleles of white, unable to produce a coding transcript, could reactivate in trans the expression of a wild type copy of the same gene silenced by heterochromatin. This new epigenetic phenomenon of transcriptional trans-reactivation is heritable, relies on the presence of homologous RNA’s and is affected by mutations in genes involved in post-transcriptional gene silencing. Our data suggest a general new unexpected level of gene expression control mediated by homologous RNA molecules in the context of heterochromatic genes. PMID:26292210

  1. Trans-Reactivation: A New Epigenetic Phenomenon Underlying Transcriptional Reactivation of Silenced Genes.

    PubMed

    Onorati, Maria Cristina; Arancio, Walter; Cavalieri, Vincenzo; Ingrassia, Antonia M R; Pavesi, Giulio; Corona, Davide F V

    2015-08-01

    In order to study the role played by cellular RNA pools produced by homologous genomic loci in defining the transcriptional state of a silenced gene, we tested the effect of non-functional alleles of the white gene in the presence of a functional copy of white, silenced by heterochromatin. We found that non-functional alleles of white, unable to produce a coding transcript, could reactivate in trans the expression of a wild type copy of the same gene silenced by heterochromatin. This new epigenetic phenomenon of transcriptional trans-reactivation is heritable, relies on the presence of homologous RNA's and is affected by mutations in genes involved in post-transcriptional gene silencing. Our data suggest a general new unexpected level of gene expression control mediated by homologous RNA molecules in the context of heterochromatic genes. PMID:26292210

  2. Synaptic, transcriptional and chromatin genes disrupted in autism.

    PubMed

    De Rubeis, Silvia; He, Xin; Goldberg, Arthur P; Poultney, Christopher S; Samocha, Kaitlin; Cicek, A Erucment; Kou, Yan; Liu, Li; Fromer, Menachem; Walker, Susan; Singh, Tarinder; Klei, Lambertus; Kosmicki, Jack; Shih-Chen, Fu; Aleksic, Branko; Biscaldi, Monica; Bolton, Patrick F; Brownfeld, Jessica M; Cai, Jinlu; Campbell, Nicholas G; Carracedo, Angel; Chahrour, Maria H; Chiocchetti, Andreas G; Coon, Hilary; Crawford, Emily L; Curran, Sarah R; Dawson, Geraldine; Duketis, Eftichia; Fernandez, Bridget A; Gallagher, Louise; Geller, Evan; Guter, Stephen J; Hill, R Sean; Ionita-Laza, Juliana; Jimenz Gonzalez, Patricia; Kilpinen, Helena; Klauck, Sabine M; Kolevzon, Alexander; Lee, Irene; Lei, Irene; Lei, Jing; Lehtimäki, Terho; Lin, Chiao-Feng; Ma'ayan, Avi; Marshall, Christian R; McInnes, Alison L; Neale, Benjamin; Owen, Michael J; Ozaki, Noriio; Parellada, Mara; Parr, Jeremy R; Purcell, Shaun; Puura, Kaija; Rajagopalan, Deepthi; Rehnström, Karola; Reichenberg, Abraham; Sabo, Aniko; Sachse, Michael; Sanders, Stephan J; Schafer, Chad; Schulte-Rüther, Martin; Skuse, David; Stevens, Christine; Szatmari, Peter; Tammimies, Kristiina; Valladares, Otto; Voran, Annette; Li-San, Wang; Weiss, Lauren A; Willsey, A Jeremy; Yu, Timothy W; Yuen, Ryan K C; Cook, Edwin H; Freitag, Christine M; Gill, Michael; Hultman, Christina M; Lehner, Thomas; Palotie, Aaarno; Schellenberg, Gerard D; Sklar, Pamela; State, Matthew W; Sutcliffe, James S; Walsh, Christiopher A; Scherer, Stephen W; Zwick, Michael E; Barett, Jeffrey C; Cutler, David J; Roeder, Kathryn; Devlin, Bernie; Daly, Mark J; Buxbaum, Joseph D

    2014-11-13

    The genetic architecture of autism spectrum disorder involves the interplay of common and rare variants and their impact on hundreds of genes. Using exome sequencing, here we show that analysis of rare coding variation in 3,871 autism cases and 9,937 ancestry-matched or parental controls implicates 22 autosomal genes at a false discovery rate (FDR) < 0.05, plus a set of 107 autosomal genes strongly enriched for those likely to affect risk (FDR < 0.30). These 107 genes, which show unusual evolutionary constraint against mutations, incur de novo loss-of-function mutations in over 5% of autistic subjects. Many of the genes implicated encode proteins for synaptic formation, transcriptional regulation and chromatin-remodelling pathways. These include voltage-gated ion channels regulating the propagation of action potentials, pacemaking and excitability-transcription coupling, as well as histone-modifying enzymes and chromatin remodellers-most prominently those that mediate post-translational lysine methylation/demethylation modifications of histones. PMID:25363760

  3. Divergence of the Yeast Transcription Factor FZF1 Affects Sulfite Resistance

    PubMed Central

    Engle, Elizabeth K.; Fay, Justin C.

    2012-01-01

    Changes in gene expression are commonly observed during evolution. However, the phenotypic consequences of expression divergence are frequently unknown and difficult to measure. Transcriptional regulators provide a mechanism by which phenotypic divergence can occur through multiple, coordinated changes in gene expression during development or in response to environmental changes. Yet, some changes in transcriptional regulators may be constrained by their pleiotropic effects on gene expression. Here, we use a genome-wide screen for promoters that are likely to have diverged in function and identify a yeast transcription factor, FZF1, that has evolved substantial differences in its ability to confer resistance to sulfites. Chimeric alleles from four Saccharomyces species show that divergence in FZF1 activity is due to changes in both its coding and upstream noncoding sequence. Between the two closest species, noncoding changes affect the expression of FZF1, whereas coding changes affect the expression of SSU1, a sulfite efflux pump activated by FZF1. Both coding and noncoding changes also affect the expression of many other genes. Our results show how divergence in the coding and promoter region of a transcription factor alters the response to an environmental stress. PMID:22719269

  4. The transcriptional regulation of regucalcin gene expression.

    PubMed

    Yamaguchi, Masayoshi

    2011-01-01

    Regucalcin, which is discovered as a calcium-binding protein in 1978, has been shown to play a multifunctional role in many tissues and cell types; regucalcin has been proposed to play a pivotal role in keeping cell homeostasis and function for cell response. Regucalcin and its gene are identified in over 15 species consisting of regucalcin family. Comparison of the nucleotide sequences of regucalcin from vertebrate species is highly conserved in their coding region with throughout evolution. The regucalcin gene is localized on the chromosome X in rat and human. The organization of rat regucalcin gene consists of seven exons and six introns and several consensus regulatory elements exist upstream of the 5'-flanking region. AP-1, NF1-A1, RGPR-p117, β-catenin, and other factors have been found to be a transcription factor in the enhancement of regucalcin gene promoter activity. The transcription activity of regucalcin gene is enhanced through intracellular signaling factors that are mediated through the phosphorylation and dephosphorylation of nuclear protein in vitro. Regucalcin mRNA and its protein are markedly expressed in the liver and kidney cortex of rats. The expression of regucalcin mRNA in the liver and kidney cortex has been shown to stimulate by hormonal factors (including calcium, calcitonin, parathyroid hormone, insulin, estrogen, and dexamethasone) in vivo. Regucalcin mRNA expression is enhanced in the regenerating liver after partial hepatectomy of rats in vivo. The expression of regucalcin mRNA in the liver and kidney with pathophysiological state has been shown to suppress, suggesting an involvement of regucalcin in disease. Liver regucalcin expression is down-regulated in tumor cells, suggesting a suppressive role in the development of carcinogenesis. Liver regucalcin is markedly released into the serum of rats with chemically induced liver injury in vivo. Serum regucalcin has a potential sensitivity as a specific biochemical marker of chronic

  5. TEMPERATURE CONDITIONING ALTERS TRANSCRIPT ABUNDANCE OF GENES RELATED TO CHILLING STRESS IN GRAPEFRUIT.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Grapefruit (Citrus paradisi) are susceptible to chilling injury (CI) if held at temperatures below about 10C. Changes in transcript abundance for a number of genes have been correlated with chilling stress in citrus fruit. We tested the hypothesis that conditioning affects transcript abundance of ...

  6. Differential sensitivities of transcription factor target genes underlie cell type-specific gene expression profiles

    PubMed Central

    Johnson, Kirby D.; Kim, Shin-Il; Bresnick, Emery H.

    2006-01-01

    Changes in transcription factor levels and activities dictate developmental fate. Such a change might affect the full ensemble of target genes for a factor or only uniquely sensitive targets. We investigated the relationship among activity of the hematopoietic transcription factor GATA-1, chromatin occupancy, and target gene sensitivity. Graded activation of GATA-1 in GATA-1-null cells revealed high-, intermediate-, and low-sensitivity targets. GATA-1 activity requirements for occupancy and transcription often correlated. A GATA-1 amino-terminal deletion mutant severely deregulated the low-sensitivity gene Tac-2. Thus, cells expressing different levels of a cell type-specific activator can have qualitatively distinct target gene expression patterns, and factor mutations preferentially deregulate low-sensitivity genes. Unlike other target genes, GATA-1-mediated Tac-2 regulation was bimodal, with activation followed by repression, and the coregulator Friend of GATA-1 (FOG-1) selectively mediated repression. A GATA-1 mutant defective in FOG-1 binding occupied a Tac-2 regulatory region at levels higher than wild-type GATA-1, whereas FOG-1 facilitated chromatin occupancy at a distinct target site. These results indicate that FOG-1 is a determinant of GATA factor target gene sensitivity by either facilitating or opposing chromatin occupancy. PMID:17043224

  7. Transcriptional targets of the schizophrenia risk gene MIR137

    PubMed Central

    Collins, A L; Kim, Y; Bloom, R J; Kelada, S N; Sethupathy, P; Sullivan, P F

    2014-01-01

    Genome-wide association studies (GWAS) have strongly implicated MIR137 (the gene encoding the microRNA miR-137) in schizophrenia. A parsimonious hypothesis is that a pathway regulated by miR-137 is important in the etiology of schizophrenia. Full evaluation of this hypothesis requires more definitive knowledge about biological targets of miR-137, which is currently lacking. Our goals were to expand knowledge of the biology of miR-137 by identifying its empirical targets, and to test whether the resulting lists of direct and indirect targets were enriched for genes and pathways involved in risk for schizophrenia. We overexpressed miR-137 in a human neural stem cell line and analyzed gene expression changes at 24 and 48 h using RNA sequencing. Following miR-137 overexpression, 202 and 428 genes were differentially expressed after 24 and 48 h. Genes differentially expressed at 24 h were enriched for transcription factors and cell cycle genes, and differential expression at 48 h affected a wider variety of pathways. Pathways implicated in schizophrenia were upregulated in the 48 h findings (major histocompatibility complex, synapses, FMRP interacting RNAs and calcium channels). Critically, differentially expressed genes at 48 h were enriched for smaller association P-values in the largest published schizophrenia GWAS. This work provides empirical support for a role of miR-137 in the etiology of schizophrenia. PMID:24984191

  8. Gene expression in plant mitochondria: transcriptional and post-transcriptional control.

    PubMed Central

    Binder, Stefan; Brennicke, Axel

    2003-01-01

    The informational content of the mitochondrial genome in plants is, although small, essential for each cell. Gene expression in these organelles involves a number of distinct transcriptional and post-transcriptional steps. The complex post-transcriptional processes of plant mitochondria such as 5' and 3' RNA processing, intron splicing, RNA editing and controlled RNA stability extensively modify individual steady-state RNA levels and influence the mRNA quantities available for translation. In this overview of the processes in mitochondrial gene expression, we focus on confirmed and potential sites of regulatory interference and discuss the evolutionary origins of the transcriptional and post-transcriptional processes. PMID:12594926

  9. Quantitative characterization of gene regulation by Rho dependent transcription termination.

    PubMed

    Hussein, Razika; Lee, Tiffany Y; Lim, Han N

    2015-08-01

    Rho factor dependent transcription termination (RTT) is common within the coding sequences of bacterial genes and it acts to couple transcription and translation levels. Despite the importance of RTT for gene regulation, its effects on mRNA and protein concentrations have not been quantitatively characterized. Here we demonstrate that the exogenous cfp gene encoding the cyan fluorescent protein can serve as a model for gene regulation by RTT. This was confirmed by showing that Psu and bicyclomycin decrease RTT and increase full length cfp mRNAs (but remarkably they have little effect on protein production). We then use cfp to characterize the relationship between its protein and full length mRNA concentrations when the translation initiation rate is varied by sequence modifications of the translation initiation region (TIR). These experiments reveal that the fold change in protein concentration (RP) and the fold change in full length mRNA concentration (Rm) have the relationship RP≈Rm(b), where b is a constant. The average value of b was determined from three separate data sets to be ~3.6. We demonstrate that the above power law function can predict how altering the translation initiation rate of a gene in an operon will affect the mRNA concentrations of downstream genes and specify a lower bound for the associated changes in protein concentrations. In summary, this study defines a simple phenomenological model to help program expression from single genes and operons that are regulated by RTT, and to guide molecular models of RTT. PMID:25982507

  10. Mechanisms of specificity in neuronal activity-regulated gene transcription

    PubMed Central

    Lyons, Michelle R.; West, Anne E.

    2011-01-01

    The brain is a highly adaptable organ that is capable of converting sensory information into changes in neuronal function. This plasticity allows behavior to be accommodated to the environment, providing an important evolutionary advantage. Neurons convert environmental stimuli into long-lasting changes in their physiology in part through the synaptic activity-regulated transcription of new gene products. Since the neurotransmitter-dependent regulation of Fos transcription was first discovered nearly 25 years ago, a wealth of studies have enriched our understanding of the molecular pathways that mediate activity-regulated changes in gene transcription. These findings show that a broad range of signaling pathways and transcriptional regulators can be engaged by neuronal activity to sculpt complex programs of stimulus-regulated gene transcription. However, the shear scope of the transcriptional pathways engaged by neuronal activity raises the question of how specificity in the nature of the transcriptional response is achieved in order to encode physiologically relevant responses to divergent stimuli. Here we summarize the general paradigms by which neuronal activity regulates transcription while focusing on the molecular mechanisms that confer differential stimulus-, cell-type-, and developmental-specificity upon activity-regulated programs of neuronal gene transcription. In addition, we preview some of the new technologies that will advance our future understanding of the mechanisms and consequences of activity-regulated gene transcription in the brain. PMID:21620929

  11. Post-transcriptional RNA Regulons Affecting Cell Cycle and Proliferation

    PubMed Central

    Blackinton, Jeff G.

    2014-01-01

    The cellular growth cycle is initiated and maintained by punctual, yet agile, regulatory events involving modifications of cell cycle proteins as well as coordinated gene expression to support cyclic checkpoint decisions. Recent evidence indicates that post-transcriptional partitioning of messenger RNA subsets by RNA-binding proteins help physically localize, temporally coordinate, and efficiently translate cell cycle proteins. This dynamic organization of mRNAs encoding cell cycle components contributes to the overall economy of the cell cycle consistent with the post-transcriptional RNA regulon model of gene expression. This review examines several recent studies demonstrating the coordination of mRNA subsets encoding cell cycle proteins during nuclear export and subsequent coupling to protein synthesis, and discusses evidence for mRNA coordination of p53 targets and the DNA damage response pathway. We consider how these observations may connect to upstream and downstream post-transcriptional coordination and coupling of splicing, export, localization, and translation. Published examples from yeast, nematode, insect, and mammalian systems are discussed, and we consider genetic evidence supporting the conclusion that dysregulation of RNA regulons may promote pathogenic states of growth such as carcinogenesis. PMID:24882724

  12. GAD2 Alternative Transcripts in the Human Prefrontal Cortex, and in Schizophrenia and Affective Disorders

    PubMed Central

    Li, Chao; Gao, Yuan; Gondré-Lewis, Marjorie C.; Lipska, Barbara K.; Shin, Joo Heon; Xie, Bin; Ye, Tianzhang; Weinberger, Daniel R.; Kleinman, Joel E.; Hyde, Thomas M.

    2016-01-01

    Genetic variation and early adverse environmental events work together to increase risk for schizophrenia. γ-aminobutyric acid (GABA), the major inhibitory neurotransmitter in adult mammalian brain, plays a major role in normal brain development, and has been strongly implicated in the pathobiology of schizophrenia. GABA synthesis is controlled by two glutamic acid decarboxylase (GAD) genes, GAD1 and GAD2, both of which produce a number of alternative transcripts. Genetic variants in the GAD1 gene are associated with increased risk for schizophrenia, and reduced expression of its major transcript in the human dorsolateral prefrontal cortex (DLPFC). No consistent changes in GAD2 expression have been found in brains from patients with schizophrenia. In this work, with the use of RNA sequencing and PCR technologies, we confirmed and tracked the expression of an alternative truncated transcript of GAD2 (ENST00000428517) in human control DLPFC homogenates across lifespan besides the well-known full length transcript of GAD2. In addition, using quantitative RT-PCR, expression of GAD2 full length and truncated transcripts were measured in the DLPFC of patients with schizophrenia, bipolar disorder and major depression. The expression of GAD2 full length transcript is decreased in the DLPFC of schizophrenia and bipolar disorder patients, while GAD2 truncated transcript is increased in bipolar disorder patients but decreased in schizophrenia patients. Moreover, the patients with schizophrenia with completed suicide or positive nicotine exposure showed significantly higher expression of GAD2 full length transcript. Alternative transcripts of GAD2 may be important in the growth and development of GABA-synthesizing neurons as well as abnormal GABA signaling in the DLPFC of patients with schizophrenia and affective disorders. PMID:26848839

  13. 7SK small nuclear RNA directly affects HMGA1 function in transcription regulation

    PubMed Central

    Eilebrecht, Sebastian; Brysbaert, Guillaume; Wegert, Thomas; Urlaub, Henning; Benecke, Bernd-Joachim; Benecke, Arndt

    2011-01-01

    Non-coding (nc) RNAs are increasingly recognized to play important regulatory roles in eukaryotic gene expression. The highly abundant and essential 7SK ncRNA has been shown to negatively regulate RNA Polymerase II transcription by inactivating the positive transcription elongation factor b (P-TEFb) in cellular and Tat-dependent HIV transcription. Here, we identify a more general, P-TEFb-independent role of 7SK RNA in directly affecting the function of the architectural transcription factor and chromatin regulator HMGA1. An important regulatory role of 7SK RNA in HMGA1-dependent cell differentiation and proliferation regulation is uncovered with the identification of over 1500 7SK-responsive HMGA1 target genes. Elevated HMGA1 expression is observed in nearly every type of cancer making the use of a 7SK substructure in the inhibition of HMGA1 activity, as pioneered here, potentially useful in therapy. The 7SK-HMGA1 interaction not only adds an essential facet to the comprehension of transcriptional plasticity at the coupling of initiation and elongation, but also might provide a molecular link between HIV reprogramming of cellular gene expression-associated oncogenesis. PMID:21087998

  14. Evolution of gene regulation during transcription and translation.

    PubMed

    Wang, Zhe; Sun, Xuepeng; Zhao, Yi; Guo, Xiaoxian; Jiang, Huifeng; Li, Hongye; Gu, Zhenglong

    2015-04-01

    Understanding how gene regulation evolves is a key area in the current evolutionary field. Gene regulation occurs at various levels. Previous work on the evolution of gene regulation has largely focused on gene transcription. In this study, we used a recently developed ribosomal footprint profiling method to investigate how gene regulation evolves at both the transcription (mRNA abundance) and translation (ribosomal density) levels. By constructing a hybrid between Saccharomyces cerevisiae (Scer) and Saccharomyces bayanus (Sbay), which diverged ∼20 Ma, and quantifying transcriptome and translatome in both parental strains and their hybrid, we showed that translation is much more conserved than transcription, mostly due to the buffering effect of translational regulation for the transcriptional divergence. More conservation in translation than transcription is also confirmed by the inheritance mode of transcription and translation between two species. Furthermore, cis and trans effects are widely involved in changes at both transcription and translation levels. Finally, our results showed that genes with certain functions and sequence features might employ specific modes for evolution at these two critical levels of gene regulation. Our results demonstrated that it is essential to investigate the evolution of gene regulation at various levels from different genetic backgrounds to obtain a complete picture of its evolutionary modes in nature. PMID:25877616

  15. Transcriptional Regulation of Gene Expression in C. elegans

    PubMed Central

    Reinke, Valerie; Krause, Michael; Okkema, Peter

    2013-01-01

    Protein coding gene sequences are converted to mRNA by the highly regulated process of transcription. The precise temporal and spatial control of transcription for many genes is an essential part of development in metazoans. Thus, understanding the molecular mechanisms underlying transcriptional control is essential to understanding cell fate determination during embryogenesis, post-embryonic development, many environmental interactions, and disease-related processes. Studies of transcriptional regulation in C. elegans exploit its genomic simplicity and physical characteristics to define regulatory events with single cell and minute time scale resolution. When combined with the genetics of the system, C. elegans offers a unique and powerful vantage point from which to study how chromatin-associated protein and their modifications interact with transcription factors and their binding sites to yield precise control of gene expression through transcriptional regulation. PMID:23801596

  16. The relationship between gene transcription and combinations of histone modifications

    NASA Astrophysics Data System (ADS)

    Cui, Xiangjun; Li, Hong; Luo, Liaofu

    2012-09-01

    Histone modification is an important subject of epigenetics which plays an intrinsic role in transcriptional regulation. It is known that multiple histone modifications act in a combinatorial fashion. In this study, we demonstrated that the pathways within constructed Bayesian networks can give an indication for the combinations among 12 histone modifications which have been studied in the TSS+1kb region in S. cerevisiae. After Bayesian networks for the genes with high transcript levels (H-network) and low transcript levels (L-network) were constructed, the combinations of modifications within the two networks were analyzed from the view of transcript level. The results showed that different combinations played dissimilar roles in the regulation of gene transcription when there exist differences for gene expression at transcription level.

  17. Does inbreeding affect gene expression in birds?

    PubMed Central

    Hansson, Bengt; Naurin, Sara; Hasselquist, Dennis

    2014-01-01

    Inbreeding increases homozygosity, exposes genome-wide recessive deleterious alleles and often reduces fitness. The physiological and reproductive consequences of inbreeding may be manifested already during gene regulation, but the degree to which inbreeding influences gene expression is unknown in most organisms, including in birds. To evaluate the pattern of inbreeding-affected gene expression over the genome and in relation to sex, we performed a transcriptome-wide gene expression (10 695 genes) study of brain tissue of 10-day-old inbred and outbred, male and female zebra finches. We found significantly lower gene expression in females compared with males at Z-linked genes, confirming that dosage compensation is incomplete in female birds. However, inbreeding did not affect gene expression at autosomal or sex-linked genes, neither in males nor in females. Analyses of single genes again found a clear sex-biased expression at Z-linked genes, whereas only a single gene was significantly affected by inbreeding. The weak effect of inbreeding on gene expression in zebra finches contrasts to the situation, for example, in Drosophila where inbreeding has been found to influence gene expression more generally and at stress-related genes in particular. PMID:25232028

  18. Large-scale analysis of antisense transcription in wheat using the Affymetrix GeneChip Wheat Genome Array

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Natural antisense transcripts (NATs) are transcripts of the opposite DNA strand to the sense-strand either at the same locus (cis-encoded) or a different locus (trans-encoded). They can affect gene expression at multiple stages including transcription, RNA processing and transport, and translation....

  19. Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products

    SciTech Connect

    Kuchka, M.R.

    1992-01-01

    Many individual chloroplast genes require the products of a collection of nuclear genes for their successful expression. These nuclear gene products apparently work with great specificity, each committed to the expression of a single chloroplast gene. We have chosen as a model nuclear mutants of Chlamydomonas affected in different stages in the expression of the chloroplast encoded Photosystem II polypeptide, D2. We have made the progress in understanding how nuclear gene products affect the translation of the D2 encoding MRNA. Two nuclear genes are required for this process which have been mapped genetically. In contrast to other examples of nuclear control of translation in the chloroplast, these nuclear gene products appear to be required either for specific stages in translation elongation or for the post-translational stabilization of the nascent D2 protein. Pseudoreversion analysis has led us to a locus which may be directly involved in D2 expression. We have made considerable progress in pursuing the molecular basis of psbd MRNA stabilization. psbD 5' UTR specific transcripts have been synthesized in vitro and used in gel mobility shift assays. UV-crosslinking studies are underway to identify the transacting factors which bind to these sequences. The continued examination of these mutants will help us to understand how nuclear gene products work in this specific case of chloroplast gene expression, and will elucidate how two distinct genomes can interact generally.

  20. Neurotoxocarosis alters myelin protein gene transcription and expression.

    PubMed

    Heuer, Lea; Beyerbach, Martin; Lühder, Fred; Beineke, Andreas; Strube, Christina

    2015-06-01

    Neurotoxocarosis is an infection of the central nervous system caused by migrating larvae of the common dog and cat roundworms (Toxocara canis and Toxocara cati), which are zoonotic agents. As these parasites are prevalent worldwide and neuropathological and molecular investigations on neurotoxocarosis are scare, this study aims to characterise nerve fibre demyelination associated with neurotoxocarosis on a molecular level. Transcription of eight myelin-associated genes (Cnp, Mag, Mbp, Mog, Mrf-1, Nogo-A, Plp1, Olig2) was determined in the mouse model during six time points of the chronic phase of infection using qRT-PCR. Expression of selected proteins was analysed by Western blotting or immunohistochemistry. Additionally, demyelination and neuronal damage were investigated histologically. Significant differences (p ≤ 0.05) between transcription rates of T. canis-infected and uninfected control mice were detected for all analysed genes while T. cati affected five of eight investigated genes. Interestingly, 2', 3 ´-cyclic nucleotide 3'-phosphodiesterase (Cnp) and myelin oligodendrocyte glycoprotein (Mog) were upregulated in both T. canis- and T. cati-infected mice preceding demyelination. Later, CNPase expression was additionally enhanced. As expected, myelin basic protein (Mbp) was downregulated in cerebra and cerebella of T. canis-infected mice when severe demyelination was present 120 days post infectionem (dpi). The transcriptional pattern observed in the present study appears to reflect direct traumatic and hypoxic effects of larval migration as well as secondary processes including host immune reactions, demyelination and attempts to remyelinate damaged areas. PMID:25773181

  1. Reactivation of apolipoprotein II gene transcription by cycloheximide reveals two steps in the deactivation of estrogen receptor-mediated transcription.

    PubMed

    Sensel, M G; Binder, R; Lazier, C B; Williams, D L

    1994-03-01

    In this report, we describe apolipoprotein II (apoII) gene expression in cell lines derived by stable expression of the chicken estrogen receptor in LMH chicken hepatoma cells. In cell lines expressing high levels of receptor (LMH/2A), apoII gene expression is increased by estrogen 300-fold compared with levels in the receptor-deficient parent LMH line. LMH/2A cells show apoII mRNA induction and turnover kinetics similar to those in chicken liver. Inhibition of protein synthesis with cycloheximide (CHX) or puromycin following estrogen withdrawal superinduces apoII mRNA without affecting apoII mRNA stability. Superinduction is due to an estrogen-independent reactivation of apoII gene transcription. The apoII gene can be reactivated by CHX for up to 24 h following hormone withdrawal, suggesting that the gene is in a repressed yet transcriptionally competent state. These results reveal two distinct events necessary for termination of estrogen receptor-mediated transcription. The first event, removal of hormone, is sufficient to stop transcription when translation is ongoing. The second event is revealed by the CHX-induced superinduction of apoII mRNA following hormone withdrawal. This superinduction suggests that deactivation of estrogen receptor-mediated transcription requires a labile protein. Furthermore, reactivation of apoII gene expression by CHX and estrogen is additive, suggesting that estrogen is unable to overcome repression completely. Thus, a labile protein may act to repress estrogen receptor-mediated transcription of the apoII gene. PMID:8114707

  2. Structural basis of eukaryotic gene transcription.

    PubMed

    Boeger, Hinrich; Bushnell, David A; Davis, Ralph; Griesenbeck, Joachim; Lorch, Yahli; Strattan, J Seth; Westover, Kenneth D; Kornberg, Roger D

    2005-02-01

    An RNA polymerase II promoter has been isolated in transcriptionally activated and repressed states. Topological and nuclease digestion analyses have revealed a dynamic equilibrium between nucleosome removal and reassembly upon transcriptional activation, and have further shown that nucleosomes are removed by eviction of histone octamers rather than by sliding. The promoter, once exposed, assembles with RNA polymerase II, general transcription factors, and Mediator in a approximately 3 MDa transcription initiation complex. X-ray crystallography has revealed the structure of RNA polymerase II, in the act of transcription, at atomic resolution. Extension of this analysis has shown how nucleotides undergo selection, polymerization, and eventual release from the transcribing complex. X-ray and electron crystallography have led to a picture of the entire transcription initiation complex, elucidating the mechanisms of promoter recognition, DNA unwinding, abortive initiation, and promoter escape. PMID:15680971

  3. Precisely modulated pathogenicity island interference with late phage gene transcription.

    PubMed

    Ram, Geeta; Chen, John; Ross, Hope F; Novick, Richard P

    2014-10-01

    Having gone to great evolutionary lengths to develop resistance to bacteriophages, bacteria have come up with resistance mechanisms directed at every aspect of the bacteriophage life cycle. Most genes involved in phage resistance are carried by plasmids and other mobile genetic elements, including bacteriophages and their relatives. A very special case of phage resistance is exhibited by the highly mobile phage satellites, staphylococcal pathogenicity islands (SaPIs), which carry and disseminate superantigen and other virulence genes. Unlike the usual phage-resistance mechanisms, the SaPI-encoded interference mechanisms are carefully crafted to ensure that a phage-infected, SaPI-containing cell will lyse, releasing the requisite crop of SaPI particles as well as a greatly diminished crop of phage particles. Previously described SaPI interference genes target phage functions that are not required for SaPI particle production and release. Here we describe a SaPI-mediated interference system that affects expression of late phage gene transcription and consequently is required for SaPI and phage. Although when cloned separately, a single SaPI gene totally blocks phage production, its activity in situ is modulated accurately by a second gene, achieving the required level of interference. The advantage for the host bacteria is that the SaPIs curb excessive phage growth while enhancing their gene transfer activity. This activity is in contrast to that of the clustered regularly interspaced short palindromic repeats (CRISPRs), which totally block phage growth at the cost of phage-mediated gene transfer. In staphylococci the SaPI strategy seems to have prevailed during evolution: The great majority of Staphylococcus aureus strains carry one or more SaPIs, whereas CRISPRs are extremely rare. PMID:25246539

  4. Oxytocin Regulates Stress-Induced Crf Gene Transcription through CREB-Regulated Transcription Coactivator 3

    PubMed Central

    Jurek, Benjamin; Slattery, David A.; Hiraoka, Yuichi; Liu, Ying; Nishimori, Katsuhiko; Aguilera, Greti; van den Burg, Erwin H.

    2015-01-01

    The major regulator of the neuroendocrine stress response in the brain is corticotropin releasing factor (CRF), whose transcription is controlled by CREB and its cofactors CRTC2/3 (TORC2/3). Phosphorylated CRTCs are sequestered in the cytoplasm, but rapidly dephosphorylated and translocated into the nucleus following a stressful stimulus. As the stress response is attenuated by oxytocin (OT), we tested whether OT interferes with CRTC translocation and, thereby, Crf expression. OT (1 nmol, i.c.v.) delayed the stress-induced increase of nuclear CRTC3 and Crf hnRNA levels in the paraventricular nucleus of male rats and mice, but did not affect either parameter in the absence of the stressor. The increase in Crf hnRNA levels at later time points was parallel to elevated nuclear CRTC2/3 levels. A direct effect of Thr4 Gly7-OT (TGOT) on CRTC3 translocation and Crf expression was found in rat primary hypothalamic neurons, amygdaloid (Ar-5), hypothalamic (H32), and human neuroblastoma (Be(2)M17) cell lines. CRTC3, but not CRCT2, knockdown using siRNA in Be(2)M17 cells prevented the effect of TGOT on Crf hnRNA levels. Chromatin-immunoprecipitation demonstrated that TGOT reduced CRTC3, but not CRTC2, binding to the Crf promoter after 10 min of forskolin stimulation. Together, the results indicate that OT modulates CRTC3 translocation, the binding of CRTC3 to the Crf promoter and, ultimately, transcription of the Crf gene. SIGNIFICANCE STATEMENT The neuropeptide oxytocin has been proposed to reduce hypothalamic-pituitary-adrenal (HPA) axis activation during stress. The underlying mechanisms are, however, elusive. In this study we show that activation of the oxytocin receptor in the paraventricular nucleus delays transcription of the gene encoding corticotropin releasing factor (Crf), the main regulator of the stress response. It does so by sequestering the coactivator of the transcription factor CREB, CRTC3, in the cytosol, resulting in reduced binding of CRTC3 to the Crf

  5. Negative elongation factor NELF controls transcription of immediate early genes in a stimulus-specific manner

    SciTech Connect

    Fujita, Toshitsugu; Piuz, Isabelle; Schlegel, Werner

    2009-01-15

    The transcription rate of immediate early genes (IEGs) is controlled directly by transcription elongation factors at the transcription elongation step. Negative elongation factor (NELF) and 5,6-dichloro-1-{beta}-D-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor (DSIF) stall RNA polymerase II (pol II) soon after transcription initiation. Upon induction of IEG transcription, DSIF is converted into an accelerator for pol II elongation. To address whether and how NELF as well as DSIF controls overall IEG transcription, its expression was reduced using stable RNA interference in GH4C1 cells. NELF knock-down reduced thyrotropin-releasing hormone (TRH)-induced transcription of the IEGs c-fos, MKP-1, and junB. In contrast, epidermal growth factor (EGF)-induced transcription of these IEGs was unaltered or even slightly increased by NELF knock-down. Thus, stable knock-down of NELF affects IEG transcription stimulation-specifically. Conversely, DSIF knock-down reduced both TRH- and EGF-induced transcription of the three IEGs. Interestingly, TRH-induced activation of the MAP kinase pathway, a pathway essential for transcription of the three IEGs, was down-regulated by NELF knock-down. Thus, stable knock-down of NELF, by modulating intracellular signaling pathways, caused stimulation-specific loss of IEG transcription. These observations indicate that NELF controls overall IEG transcription via multiple mechanisms both directly and indirectly.

  6. Transcriptional and Post-Transcriptional Regulation of Nucleotide Excision Repair Genes in Human Cells

    PubMed Central

    Lefkofsky, Hailey B.; Veloso, Artur; Ljungman, Mats

    2014-01-01

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death. PMID:26255935

  7. Transcriptional and post-transcriptional regulation of nucleotide excision repair genes in human cells.

    PubMed

    Lefkofsky, Hailey B; Veloso, Artur; Ljungman, Mats

    2015-06-01

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death. PMID:26255935

  8. Transcriptional regulation differs in affected facioscapulohumeral muscular dystrophy patients compared to asymptomatic related carriers

    PubMed Central

    Arashiro, Patricia; Eisenberg, Iris; Kho, Alvin T.; Cerqueira, Antonia M. P.; Canovas, Marta; Silva, Helga C. A.; Pavanello, Rita C. M.; Verjovski-Almeida, Sergio; Kunkel, Louis M.; Zatz, Mayana

    2009-01-01

    Facioscapulohumeral muscular dystrophy (FSHD) is a progressive muscle disorder that has been associated with a contraction of 3.3-kb repeats on chromosome 4q35. FSHD is characterized by a wide clinical inter- and intrafamilial variability, ranging from wheelchair-bound patients to asymptomatic carriers. Our study is unique in comparing the gene expression profiles from related affected, asymptomatic carrier, and control individuals. Our results suggest that the expression of genes on chromosome 4q is altered in affected and asymptomatic individuals. Remarkably, the changes seen in asymptomatic samples are largely in products of genes encoding several chemokines, whereas the changes seen in affected samples are largely in genes governing the synthesis of GPI-linked proteins and histone acetylation. Besides this, the affected patient and related asymptomatic carrier share the 4qA161 haplotype. Thus, these polymorphisms by themselves do not explain the pathogenicity of the contracted allele. Interestingly, our results also suggest that the miRNAs might mediate the regulatory network in FSHD. Together, our results support the previous evidence that FSHD may be caused by transcriptional dysregulation of multiple genes, in cis and in trans, and suggest some factors potentially important for FSHD pathogenesis. The study of the gene expression profiles from asymptomatic carriers and related affected patients is a unique approach to try to enhance our understanding of the missing link between the contraction in D4Z4 repeats and muscle disease, while minimizing the effects of differences resulting from genetic background. PMID:19339494

  9. Gene transcription and chromosome replication in Escherichia coli.

    PubMed Central

    Zhou, P; Bogan, J A; Welch, K; Pickett, S R; Wang, H J; Zaritsky, A; Helmstetter, C E

    1997-01-01

    Transcript levels of several Escherichia coli genes involved in chromosome replication and cell division were measured in dnaC2(Ts) mutants synchronized for chromosome replication by temperature shifts. Levels of transcripts from four of the genes, dam, nrdA, mukB, and seqA, were reduced at a certain stage during chromosome replication. The magnitudes of the decreases were similar to those reported previously ftsQ and ftsZ (P. Zhou and C. E. Helmstetter, J. Bacteriol. 176:6100-6106, 1994) but considerably less than those seen with dnaA, gidA, and mioC (P. W. Theisen, J. E. Grimwade, A. C. Leonard, J. A. Bogan, and C. E. Helmstetter, Mol. Microbiol. 10:575-584, 1993). The decreases in transcripts appeared to correlate with the estimated time at which the genes replicated. This same conclusion was reached in studies with synchronous cultures obtained with the baby machine in those instances in which periodicities in transcript levels were clearly evident. The transcriptional levels for two genes, minE and tus, did not fluctuate significantly, whereas the transcripts for one gene, iciA, appeared to increase transiently. The results support the idea that cell cycle timing in E. coli is not governed by timed bursts of gene expression, since the overall findings summarized in this report are generally consistent with cell cycle-dependent transient inhibitions of transcription rather than stimulations. PMID:8981994

  10. Transcription mediated insulation and interference direct gene cluster expression switches

    PubMed Central

    Nguyen, Tania; Brown, David; Murray, Struan C; Haenni, Simon; Halstead, James M; O'Connor, Leigh; Shipkovenska, Gergana; Steinmetz, Lars M; Mellor, Jane

    2014-01-01

    In yeast, many tandemly arranged genes show peak expression in different phases of the metabolic cycle (YMC) or in different carbon sources, indicative of regulation by a bi-modal switch, but it is not clear how these switches are controlled. Using native elongating transcript analysis (NET-seq), we show that transcription itself is a component of bi-modal switches, facilitating reciprocal expression in gene clusters. HMS2, encoding a growth-regulated transcription factor, switches between sense- or antisense-dominant states that also coordinate up- and down-regulation of transcription at neighbouring genes. Engineering HMS2 reveals alternative mono-, di- or tri-cistronic and antisense transcription units (TUs), using different promoter and terminator combinations, that underlie state-switching. Promoters or terminators are excluded from functional TUs by read-through transcriptional interference, while antisense TUs insulate downstream genes from interference. We propose that the balance of transcriptional insulation and interference at gene clusters facilitates gene expression switches during intracellular and extracellular environmental change. DOI: http://dx.doi.org/10.7554/eLife.03635.001 PMID:25407679

  11. LEF-1 Regulates Tyrosinase Gene Transcription In Vitro

    PubMed Central

    Wang, Xueping; Liu, Yalan; Chen, Hongsheng; Mei, Lingyun; He, Chufeng; Jiang, Lu; Niu, Zhijie; Sun, Jie; Luo, Hunjin; Li, Jiada; Feng, Yong

    2015-01-01

    TYR, DCT and MITF are three important genes involved in maintaining the mature phenotype and producing melanin; they therefore participate in neural crest cell development into melanocytes. Previous studies have revealed that the Wnt signaling factor lymphoid enhancer-binding factor (LEF-1) can enhance DCT and MITF gene expression. However, whether LEF-1 also affects TYR gene expression remains unclear. In the present study, we found that LEF-1 regulated TYR transcription in vitro. LEF-1 overexpression increased TYR gene promoter activity, whereas LEF-1 knockdown by RNA interference significantly decreased TYR expression. Moreover, the core GTTTGAT sequence (-56 to -50) within the TYR promoter is essential for the effect of LEF-1 on TYR expression, and chromatin immunoprecipitation (ChIP) assay indicated that endogenous LEF-1 interacts with the TYR promoter. In addition, we observed a synergistic transactivation of the TYR promoter by LEF-1 and MITF. These data suggest that Wnt signaling plays an important role in regulating melanocyte development and differentiation. PMID:26580798

  12. Genome-Wide Analysis of Factors Affecting Transcription Elongation and DNA Repair: A New Role for PAF and Ccr4-Not in Transcription-Coupled Repair

    PubMed Central

    Gaillard, Hélène; Tous, Cristina; Botet, Javier; González-Aguilera, Cristina; Quintero, Maria José; Viladevall, Laia; García-Rubio, María L.; Rodríguez-Gil, Alfonso; Marín, Antonio; Ariño, Joaquín; Revuelta, José Luis; Chávez, Sebastián; Aguilera, Andrés

    2009-01-01

    RNA polymerases frequently deal with a number of obstacles during transcription elongation that need to be removed for transcription resumption. One important type of hindrance consists of DNA lesions, which are removed by transcription-coupled repair (TC-NER), a specific sub-pathway of nucleotide excision repair. To improve our knowledge of transcription elongation and its coupling to TC-NER, we used the yeast library of non-essential knock-out mutations to screen for genes conferring resistance to the transcription-elongation inhibitor mycophenolic acid and the DNA-damaging agent 4-nitroquinoline-N-oxide. Our data provide evidence that subunits of the SAGA and Ccr4-Not complexes, Mediator, Bre1, Bur2, and Fun12 affect transcription elongation to different extents. Given the dependency of TC-NER on RNA Polymerase II transcription and the fact that the few proteins known to be involved in TC-NER are related to transcription, we performed an in-depth TC-NER analysis of a selection of mutants. We found that mutants of the PAF and Ccr4-Not complexes are impaired in TC-NER. This study provides evidence that PAF and Ccr4-Not are required for efficient TC-NER in yeast, unraveling a novel function for these transcription complexes and opening new perspectives for the understanding of TC-NER and its functional interconnection with transcription elongation. PMID:19197357

  13. Glucose repression of yeast mitochondrial transcription: kinetics of derepression and role of nuclear genes.

    PubMed Central

    Ulery, T L; Jang, S H; Jaehning, J A

    1994-01-01

    Yeast mitochondrial transcript and gene product abundance has been observed to increase upon release from glucose repression, but the mechanism of regulation of this process has not been determined. We report a kinetic analysis of this phenomenon, which demonstrates that the abundance of all classes of mitochondrial RNA changes slowly relative to changes observed for glucose-repressed nuclear genes. Several cell doublings are required to achieve the 2- to 20-fold-higher steady-state levels observed after a shift to a nonrepressing carbon source. Although we observed that in some yeast strains the mitochondrial DNA copy number also increases upon derepression, this does not seem to play the major role in increased RNA abundance. Instead we found that three- to sevenfold increases in RNA synthesis rates, measured by in vivo pulse-labelling experiments, do correlate with increased transcript abundance. We found that mutations in the SNF1 and REG1 genes, which are known to affect the expression of many nuclear genes subject to glucose repression, affect derepression of mitochondrial transcript abundance. These genes do not appear to regulate mitochondrial transcript levels via regulation of the nuclear genes RPO41 and MTF1, which encode the subunits of the mitochondrial RNA polymerase. We conclude that a nuclear gene-controlled factor(s) in addition to the two RNA polymerase subunits must be involved in glucose repression of mitochondrial transcript abundance. Images PMID:8289797

  14. Characterizing transcriptional heterogeneity through pathway and gene set overdispersion analysis

    PubMed Central

    Fan, Jean; Salathia, Neeraj; Liu, Rui; Kaeser, Gwendolyn E.; Yung, Yun C.; Herman, Joseph L.; Kaper, Fiona; Fan, Jian-Bing; Zhang, Kun; Chun, Jerold; Kharchenko, Peter V.

    2016-01-01

    The transcriptional state of a cell reflects a variety of biological factors, from persistent cell-type specific features to transient processes such as cell cycle. Depending on biological context, all such aspects of transcriptional heterogeneity may be of interest, but detecting them from noisy single-cell RNA-seq data remains challenging. We developed PAGODA to resolve multiple, potentially overlapping aspects of transcriptional heterogeneity by testing gene sets for coordinated variability amongst measured cells. PMID:26780092

  15. Gene Transcription Profile of the Detached Retina (An AOS Thesis)

    PubMed Central

    Zacks, David N.

    2009-01-01

    Purpose: Separation of the neurosensory retina from the retinal pigment epithelium (RPE) yields many morphologic and functional consequences, including death of the photoreceptor cells, Müller cell hypertrophy, and inner retinal rewiring. Many of these changes are due to the separation-induced activation of specific genes. In this work, we define the gene transcription profile within the retina as a function of time after detachment. We also define the early activation of kinases that might be responsible for the detachment-induced changes in gene transcription. Methods: Separation of the retina from the RPE was induced in Brown-Norway rats by the injection of 1% hyaluronic acid into the subretinal space. Retinas were harvested at 1, 7, and 28 days after separation. Gene transcription profiles for each time point were determined using the Affymetrix Rat 230A gene microarray chip. Transcription levels in detached retinas were compared to those of nondetached retinas with the BRB-ArrayTools Version 3.6.0 using a random variance analysis of variance (ANOVA) model. Confirmation of the significant transcriptional changes for a subset of the genes was performed using microfluidic quantitative real-time polymerase chain reaction (qRT-PCR) assays. Kinase activation was explored using Western blot analysis to look for early phosphorylation of any of the 3 main families of mitogen-activated protein kinases (MAPK): the p38 family, the Janus kinase family, and the p42/p44 family. Results: Retinas separated from the RPE showed extensive alterations in their gene transcription profile. Many of these changes were initiated as early as 1 day after separation, with significant increases by 7 days. ANOVA analysis defined 144 genes that had significantly altered transcription levels as a function of time after separation when setting a false discovery rate at ≤0.1. Confirmatory RT-PCR was performed on 51 of these 144 genes. Differential transcription detected on the microarray

  16. Stochastic models of gene expression and post-transcriptional regulation

    NASA Astrophysics Data System (ADS)

    Pendar, Hodjat; Kulkarni, Rahul; Jia, Tao

    2011-10-01

    The intrinsic stochasticity of gene expression can give rise to phenotypic heterogeneity in a population of genetically identical cells. Correspondingly, there is considerable interest in understanding how different molecular mechanisms impact the 'noise' in gene expression. Of particular interest are post-transcriptional regulatory mechanisms involving genes called small RNAs, which control important processes such as development and cancer. We propose and analyze general stochastic models of gene expression and derive exact analytical expressions quantifying the noise in protein distributions [1]. Focusing on specific regulatory mechanisms, we analyze a general model for post-transcriptional regulation of stochastic gene expression [2]. The results obtained provide new insights into the role of post-transcriptional regulation in controlling the noise in gene expression. [4pt] [1] T. Jia and R. V. Kulkarni, Phys. Rev. Lett.,106, 058102 (2011) [0pt] [2] T. Jia and R. V. Kulkarni, Phys. Rev. Lett., 105, 018101 (2010)

  17. Transcription factor trapping by RNA in gene regulatory elements

    PubMed Central

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

    2016-01-01

    Transcription factors (TFs) bind specific sequences in promoter-proximal and distal DNA elements in order to regulate gene transcription. RNA is transcribed from both of these DNA elements, and some DNA-binding TFs bind RNA. Hence, RNA transcribed from regulatory elements may contribute to stable TF occupancy at these sites. We show that the ubiquitously expressed TF YY1 binds to both gene regulatory elements and also to their associated RNA species genome-wide. Reduced transcription of regulatory elements diminishes YY1 occupancy whereas artificial tethering of RNA enhances YY1 occupancy at these elements. We propose that RNA makes a modest but important contribution to the maintenance of certain TFs at gene regulatory elements and suggest that transcription of regulatory elements produces a positive feedback loop that contributes to the stability of gene expression programs. PMID:26516199

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

    PubMed

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

    2015-11-20

    Transcription factors (TFs) bind specific sequences in promoter-proximal and -distal DNA elements to regulate gene transcription. RNA is transcribed from both of these DNA elements, and some DNA binding TFs bind RNA. Hence, RNA transcribed from regulatory elements may contribute to stable TF occupancy at these sites. We show that the ubiquitously expressed TF Yin-Yang 1 (YY1) binds to both gene regulatory elements and their associated RNA species across the entire genome. Reduced transcription of regulatory elements diminishes YY1 occupancy, whereas artificial tethering of RNA enhances YY1 occupancy at these elements. We propose that RNA makes a modest but important contribution to the maintenance of certain TFs at gene regulatory elements and suggest that transcription of regulatory elements produces a positive-feedback loop that contributes to the stability of gene expression programs. PMID:26516199

  19. Higher plant mitochondrial DNA: Genomes, genes, mutants, transcription, translation

    SciTech Connect

    Not Available

    1986-01-01

    This volume contains brief summaries of 63 presentations given at the International Workshop on Higher Plant Mitochondrial DNA. The presentations are organized into topical discussions addressing plant genomes, mitochondrial genes, cytoplasmic male sterility, transcription, translation, plasmids and tissue culture. (DT)

  20. Two control regions for eukaryotic tRNA gene transcription.

    PubMed Central

    DeFranco, D; Schmidt, O; Söll, D

    1980-01-01

    Two Drosophila tRNALys genes with identical coding sequences were shown to transcribe with very different efficiences in nuclear extracts from Xenopus oocytes. The use of recombinant plasmids in which the 5'-flanking sequences of these genes were either "switched" or replaced by defined pBR322 sequences revealed two control regions for tRNA gene transcription. An internal control region comprising the mature tRNA coding sequence (and possibly its 3'-flanking sequences) is sufficient for transcription initiation, and an external control region comprising the 5'-flanking sequences represses this transcription. All transcripts have short leader sequences. Altered precursor tRNAs transcribed from truncated tRNALys genes (missing a single base pair in the acceptor stem) are not processed well in vitro. Images PMID:6774336

  1. Transcription Interference and ORF Nature Strongly Affect Promoter Strength in a Reconstituted Metabolic Pathway

    PubMed Central

    Carquet, Marie; Pompon, Denis; Truan, Gilles

    2015-01-01

    Fine tuning of individual enzyme expression level is necessary to alleviate metabolic imbalances in synthetic heterologous pathways. A known approach consists of choosing a suitable combination of promoters, based on their characterized strengths in model conditions. We questioned whether each step of a multiple-gene synthetic pathway could be independently tunable at the transcription level. Three open reading frames, coding for enzymes involved in a synthetic pathway, were combinatorially associated to different promoters on an episomal plasmid in Saccharomyces cerevisiae. We quantified the mRNA levels of the three genes in each strain of our generated combinatorial metabolic library. Our results evidenced that the ORF nature, position, and orientation induce strong discrepancies between the previously reported promoters’ strengths and the observed ones. We conclude that, in the context of metabolic reconstruction, the strength of usual promoters can be dramatically affected by many factors. Among them, transcriptional interference and ORF nature seem to be predominant. PMID:25767795

  2. Age and Diet Affect Gene Expression Profile in Canine Skeletal Muscle

    PubMed Central

    Middelbos, Ingmar S.; Vester, Brittany M.; Karr-Lilienthal, Lisa K.; Schook, Lawrence B.; Swanson, Kelly S.

    2009-01-01

    We evaluated gene transcription in canine skeletal muscle (biceps femoris) using microarray analysis to identify effects of age and diet on gene expression. Twelve female beagles were used (six 1-year olds and six 12-year olds) and they were fed one of two experimental diets for 12 months. One diet contained primarily plant-based protein sources (PPB), whereas the second diet contained primarily animal-based protein sources (APB). Affymetrix GeneChip Canine Genome Arrays were used to hybridize extracted RNA. Age had the greatest effect on gene transcription (262 differentially expressed genes), whereas the effect of diet was relatively small (22 differentially expressed genes). Effects of age (regardless of diet) were most notable on genes related to metabolism, cell cycle and cell development, and transcription function. All these genes were predominantly down-regulated in geriatric dogs. Age-affected genes that were differentially expressed on only one of two diets were primarily noted in the PPB diet group (144/165 genes). Again, genes related to cell cycle (22/35) and metabolism (15/19) had predominantly decreased transcription in geriatric dogs, but 6/8 genes related to muscle development had increased expression. Effects of diet on muscle gene expression were mostly noted in geriatric dogs, but no consistent patterns in transcription were observed. The insight these data provide into gene expression profiles of canine skeletal muscle as affected by age, could serve as a foundation for future research pertaining to age-related muscle diseases. PMID:19221602

  3. CrBPF1 overexpression alters transcript levels of terpenoid indole alkaloid biosynthetic and regulatory genes.

    PubMed

    Li, Chun Yao; Leopold, Alex L; Sander, Guy W; Shanks, Jacqueline V; Zhao, Le; Gibson, Susan I

    2015-01-01

    Terpenoid indole alkaloid (TIA) biosynthesis in Catharanthus roseus is a complex and highly regulated process. Understanding the biochemistry and regulation of the TIA pathway is of particular interest as it may allow the engineering of plants to accumulate higher levels of pharmaceutically important alkaloids. Toward this end, we generated a transgenic C. roseus hairy root line that overexpresses the CrBPF1 transcriptional activator under the control of a β-estradiol inducible promoter. CrBPF1 is a MYB-like protein that was previously postulated to help regulate the expression of the TIA biosynthetic gene STR. However, the role of CrBPF1 in regulation of the TIA and related pathways had not been previously characterized. In this study, transcriptional profiling revealed that overexpression of CrBPF1 results in increased transcript levels for genes from both the indole and terpenoid biosynthetic pathways that provide precursors for TIA biosynthesis, as well as for genes in the TIA biosynthetic pathway. In addition, overexpression of CrBPF1 causes increases in the transcript levels for 11 out of 13 genes postulated to act as transcriptional regulators of genes from the TIA and TIA feeder pathways. Interestingly, overexpression of CrBPF1 causes increased transcript levels for both TIA transcriptional activators and repressors. Despite the fact that CrBPF1 overexpression affects transcript levels of a large percentage of TIA biosynthetic and regulatory genes, CrBPF1 overexpression has only very modest effects on the levels of the TIA metabolites analyzed. This finding may be due, at least in part, to the up-regulation of both transcriptional activators and repressors in response to CrBPF1 overexpression, suggesting that CrBPF1 may serve as a "fine-tune" regulator for TIA biosynthesis, acting to help regulate the timing and amplitude of TIA gene expression. PMID:26483828

  4. A mutation in the C31 subunit of Saccharomyces cerevisiae RNA polymerase III affects transcription initiation.

    PubMed Central

    Thuillier, V; Stettler, S; Sentenac, A; Thuriaux, P; Werner, M

    1995-01-01

    The C31 subunit belongs to a complex of three subunits (C31, C34 and C82) specific to RNA polymerase (pol) III that have no counterparts in other RNA polymerases. This complex is thought to play a role in transcription initiation since it interacts with the general initiation factor TFIIIB via subunit C34. We have obtained a conditional mutation of pol III by partially deleting the acidic C-terminus of the C31 subunit. A Saccharomyces cerevisiae strain carrying this truncated C31 subunit is impaired in in vivo transcription of tRNAs and failed to grow at 37 degrees C. This conditional growth phenotype was suppressed by overexpression of the gene coding for the largest subunit of pol III (C160), suggesting an interaction between C160 and C31. The mutant pol III enzyme transcribed non-specific templates at wild-type rates in vitro, but was impaired in its capacity to transcribe tRNA genes in the presence of general initiation factors. Transcription initiation, but not termination or recycling of the enzyme, was affected in the mutant, suggesting that it could be altered on interaction with initiation factors or on the formation of the open complex. Interestingly, the C-terminal deletion was also suppressed by a high gene dosage of the DED1 gene encoding a putative helicase. Images PMID:7835345

  5. Genes on a Wire: The Nucleoid-Associated Protein HU Insulates Transcription Units in Escherichia coli.

    PubMed

    Berger, Michael; Gerganova, Veneta; Berger, Petya; Rapiteanu, Radu; Lisicovas, Viktoras; Dobrindt, Ulrich

    2016-01-01

    The extent to which chromosomal gene position in prokaryotes affects local gene expression remains an open question. Several studies have shown that chromosomal re-positioning of bacterial transcription units does not alter their expression pattern, except for a general decrease in gene expression levels from chromosomal origin to terminus proximal positions, which is believed to result from gene dosage effects. Surprisingly, the question as to whether this chromosomal context independence is a cis encoded property of a bacterial transcription unit, or if position independence is a property conferred by factors acting in trans, has not been addressed so far. For this purpose, we established a genetic test system assessing the chromosomal positioning effects by means of identical promoter-fluorescent reporter gene fusions inserted equidistantly from OriC into both chromosomal replichores of Escherichia coli K-12. Our investigations of the reporter activities in mutant cells lacking the conserved nucleoid associated protein HU uncovered various drastic chromosomal positional effects on gene transcription. In addition we present evidence that these positional effects are caused by transcriptional activity nearby the insertion site of our reporter modules. We therefore suggest that the nucleoid-associated protein HU is functionally insulating transcription units, most likely by constraining transcription induced DNA supercoiling. PMID:27545593

  6. Genes on a Wire: The Nucleoid-Associated Protein HU Insulates Transcription Units in Escherichia coli

    PubMed Central

    Berger, Michael; Gerganova, Veneta; Berger, Petya; Rapiteanu, Radu; Lisicovas, Viktoras; Dobrindt, Ulrich

    2016-01-01

    The extent to which chromosomal gene position in prokaryotes affects local gene expression remains an open question. Several studies have shown that chromosomal re-positioning of bacterial transcription units does not alter their expression pattern, except for a general decrease in gene expression levels from chromosomal origin to terminus proximal positions, which is believed to result from gene dosage effects. Surprisingly, the question as to whether this chromosomal context independence is a cis encoded property of a bacterial transcription unit, or if position independence is a property conferred by factors acting in trans, has not been addressed so far. For this purpose, we established a genetic test system assessing the chromosomal positioning effects by means of identical promoter-fluorescent reporter gene fusions inserted equidistantly from OriC into both chromosomal replichores of Escherichia coli K-12. Our investigations of the reporter activities in mutant cells lacking the conserved nucleoid associated protein HU uncovered various drastic chromosomal positional effects on gene transcription. In addition we present evidence that these positional effects are caused by transcriptional activity nearby the insertion site of our reporter modules. We therefore suggest that the nucleoid-associated protein HU is functionally insulating transcription units, most likely by constraining transcription induced DNA supercoiling. PMID:27545593

  7. Genomewide Identification of Genes Under Directional Selection: Gene Transcription QST Scan in Diverging Atlantic Salmon Subpopulations

    PubMed Central

    Roberge, C.; Guderley, H.; Bernatchez, L.

    2007-01-01

    Evolutionary genomics has benefited from methods that allow identifying evolutionarily important genomic regions on a genomewide scale, including genome scans and QTL mapping. Recently, genomewide scanning by means of microarrays has permitted assessing gene transcription differences among species or populations. However, the identification of differentially transcribed genes does not in itself suffice to measure the role of selection in driving evolutionary changes in gene transcription. Here, we propose and apply a “transcriptome scan” approach to investigating the role of selection in shaping differential profiles of gene transcription among populations. We compared the genomewide transcription levels between two Atlantic salmon subpopulations that have been diverging for only six generations. Following assessment of normality and unimodality on a gene-per-gene basis, the additive genetic basis of gene transcription was estimated using the animal model. Gene transcription h2 estimates were significant for 1044 (16%) of all detected cDNA clones. In an approach analogous to that of genome scans, we used the distribution of the QST values estimated from intra- and intersubpopulation additive genetic components of the transcription profiles to identify 16 outlier genes (average QST estimate = 0.11) whose transcription levels are likely to have evolved under the influence of directional selection within six generations only. Overall, this study contributes both empirically and methodologically to the quantitative genetic exploration of gene transcription data. PMID:17720934

  8. Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products

    SciTech Connect

    Kuchka, M.R.

    1992-01-01

    The following is a review of research accomplished in the first two years of funding for the above mentioned project. The work performed is a molecular characterization of nuclear mutants of Chlamydomonas reinhardtii which are deficient in different stages in the post-transcriptional expression of a single chloroplast encoded polypeptide, the D2 protein of Photosystem II. Our long-term goals are to understand the molecular mechanisms by which nuclear gene products affect the expression of chloroplast genes. Specifically, we which to understand how specific nuclear gene products affect the turnover rate of the D2 encoding mRNA (psbD), how other nuclear encoded factors work to promote the translation of psbD mRNA and/or stabilize the D2 protein, and what the role of the D2 protein itself is in Photosystem II assembly and in the control of expression of other chloroplast genes. This progress report will be organized into four major sections concerning (I) The characterization of nuclear mutants affected in D2 translation/turnover, (II) The study of trans-acting factors which associate with the 5{prime} end of the psbD mRNA, (III) In vitro mutagenesis of the psbD gene, and (IV) Additional studies.

  9. Topics in Transcriptional Control of Lipid Metabolism: from Transcription Factors to Gene-Promoter Polymorphisms

    PubMed Central

    Bergen, Werner G.; Burnett, Derris D.

    2013-01-01

    The central dogma of biology (DNA>>RNA>>Protein) has remained as an extremely useful scaffold to guide the study of molecular regulation of cellular metabolism. Molecular regulation of cellular metabolism has been pursued from an individual enzyme to a global assessment of protein function at the genomic (DNA), transcriptomic (RNA) and translation (Protein) levels. Details of a key role by inhibitory small RNAs and post-translational processing of cellular proteins on a whole cell/global basis are now just emerging. Below we emphasize the role of transcription factors (TF) in regulation of adipogenesis and lipogenesis. Additionally we have also focused on emerging additional TF that may also have hitherto unrecognized roles in adipogenesis and lipogenesis as compared to our present understanding. It is generally recognized that SNPs in structural genes can affect the final structure/function of a given protein. The implications of SNPs located in the non-transcribed promoter region on transcription have not been examined as extensively at this time. Here we have also summarized some emerging results on promoter SNPs for lipid metabolism and related cellular processes. PMID:25031651

  10. Dynamic enhancer–gene body contacts during transcription elongation

    PubMed Central

    Lee, Kiwon; Hsiung, Chris C.-S.; Huang, Peng; Raj, Arjun; Blobel, Gerd A.

    2015-01-01

    Enhancers govern transcription through multiple mechanisms, including the regulation of elongation by RNA polymerase II (RNAPII). We characterized the dynamics of looped enhancer contacts during synchronous transcription elongation. We found that many distal enhancers form stable contacts with their target promoters during the entire interval of elongation. Notably, we detected additional dynamic enhancer contacts throughout the gene bodies that track with elongating RNAPII and the leading edge of RNA synthesis. These results support a model in which the gene body changes its position relative to a stable enhancer–promoter complex, which has broad ramifications for enhancer function and architectural models of transcriptional elongation. PMID:26443845

  11. TransFind—predicting transcriptional regulators for gene sets

    PubMed Central

    Kiełbasa, Szymon M.; Klein, Holger; Roider, Helge G.; Vingron, Martin; Blüthgen, Nils

    2010-01-01

    The analysis of putative transcription factor binding sites in promoter regions of coregulated genes allows to infer the transcription factors that underlie observed changes in gene expression. While such analyses constitute a central component of the in-silico characterization of transcriptional regulatory networks, there is still a lack of simple-to-use web servers able to combine state-of-the-art prediction methods with phylogenetic analysis and appropriate multiple testing corrected statistics, which returns the results within a short time. Having these aims in mind we developed TransFind, which is freely available at http://transfind.sys-bio.net/. PMID:20511592

  12. Identification of putative target genes of the transcription factor RUNX2.

    PubMed

    Kuhlwilm, Martin; Davierwala, Armaity; Pääbo, Svante

    2013-01-01

    Comparisons of the genomes of Neandertals and Denisovans with present-day human genomes have suggested that the gene RUNX2, which encodes a transcription factor, may have been positively selected during early human evolution. Here, we overexpress RUNX2 in ten human cell lines and identify genes that are directly or indirectly affected by RUNX2 expression. We find a number of genes not previously known to be affected by RUNX2 expression, in particular BIRC3, genes encoded on the mitochondrial genome, and several genes involved in bone and tooth formation. These genes are likely to provide inroads into pathways affected by RUNX2 and potentially by the evolutionary changes that affected RUNX2 in modern humans. PMID:24349465

  13. Transcriptional landscape and essential genes of Neisseria gonorrhoeae

    PubMed Central

    Remmele, Christian W.; Xian, Yibo; Albrecht, Marco; Faulstich, Michaela; Fraunholz, Martin; Heinrichs, Elisabeth; Dittrich, Marcus T.; Müller, Tobias; Reinhardt, Richard; Rudel, Thomas

    2014-01-01

    The WHO has recently classified Neisseria gonorrhoeae as a super-bacterium due to the rapid spread of antibiotic resistant derivatives and an overall dramatic increase in infection incidences. Genome sequencing has identified potential genes, however, little is known about the transcriptional organization and the presence of non-coding RNAs in gonococci. We performed RNA sequencing to define the transcriptome and the transcriptional start sites of all gonococcal genes and operons. Numerous new transcripts including 253 potentially non-coding RNAs transcribed from intergenic regions or antisense to coding genes were identified. Strikingly, strong antisense transcription was detected for the phase-variable opa genes coding for a family of adhesins and invasins in pathogenic Neisseria, that may have regulatory functions. Based on the defined transcriptional start sites, promoter motifs were identified. We further generated and sequenced a high density Tn5 transposon library to predict a core of 827 gonococcal essential genes, 133 of which have no known function. Our combined RNA-Seq and Tn-Seq approach establishes a detailed map of gonococcal genes and defines the first core set of essential gonococcal genes. PMID:25143534

  14. Transcriptional modulator ZBED6 affects cell cycle and growth of human colorectal cancer cells

    PubMed Central

    Akhtar Ali, Muhammad; Younis, Shady; Wallerman, Ola; Gupta, Rajesh; Andersson, Leif; Sjöblom, Tobias

    2015-01-01

    The transcription factor ZBED6 (zinc finger, BED-type containing 6) is a repressor of IGF2 whose action impacts development, cell proliferation, and growth in placental mammals. In human colorectal cancers, IGF2 overexpression is mutually exclusive with somatic mutations in PI3K signaling components, providing genetic evidence for a role in the PI3K pathway. To understand the role of ZBED6 in tumorigenesis, we engineered and validated somatic cell ZBED6 knock-outs in the human colorectal cancer cell lines RKO and HCT116. Ablation of ZBED6 affected the cell cycle and led to increased growth rate in RKO cells but reduced growth in HCT116 cells. This striking difference was reflected in the transcriptome analyses, which revealed enrichment of cell-cycle–related processes among differentially expressed genes in both cell lines, but the direction of change often differed between the cell lines. ChIP sequencing analyses displayed enrichment of ZBED6 binding at genes up-regulated in ZBED6-knockout clones, consistent with the view that ZBED6 modulates gene expression primarily by repressing transcription. Ten differentially expressed genes were identified as putative direct gene targets, and their down-regulation by ZBED6 was validated experimentally. Eight of these genes were linked to the Wnt, Hippo, TGF-β, EGF receptor, or PI3K pathways, all involved in colorectal cancer development. The results of this study show that the effect of ZBED6 on tumor development depends on the genetic background and the transcriptional state of its target genes. PMID:26056301

  15. Transcriptional analysis of Penaeus stylirostris densovirus genes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Penaeus stylirostris densovirus (PstDNV) genome contains three open reading frames (ORFs), left, middle, and right, which encode a non-structural (NS) protein, an unknown protein, and a capsid protein (CP), respectively. Transcription mapping revealed that P2, P11 and P61 promoters transcribe the le...

  16. Transcriptional unit of the murine Thy-1 gene: different distribution of transcription initiation sites in brain.

    PubMed Central

    Spanopoulou, E; Giguere, V; Grosveld, F

    1988-01-01

    Structural analysis of the mouse Thy-1.2 gene has shown that the major promoter of the gene is characterized by a tissue-specific DNase I-hypersensitive site and is located within a methylation-free island. The gene is regulated at the transcriptional level, and steady-state mRNA analysis reveals that the previously reported exon Ib contributes at most 5% of the total mRNA. The major promoter uses several transcription initiation sites within a region of 100 base pairs. The frequency of usage of these sites in brain is markedly different from that in other tissues. Images PMID:2906111

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

    PubMed Central

    Miyazato, Paola; Matsuo, Misaki; Katsuya, Hiroo; Satou, Yorifumi

    2016-01-01

    Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus associated with human diseases, such as adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/Tropic spastic paraparesis (HAM/TSP). As a retrovirus, its life cycle includes a step where HTLV-1 is integrated into the host genomic DNA and forms proviral DNA. In the chronic phase of the infection, HTLV‑1 is known to proliferate as a provirus via the mitotic division of the infected host cells. There are generally tens of thousands of infected clones within an infected individual. They exist not only in peripheral blood, but also in various lymphoid organs. Viral proteins encoded in HTLV-1 genome play a role in the proliferation and survival of the infected cells. As is the case with other chronic viral infections, HTLV-1 gene expression induces the activation of the host immunity against the virus. Thus, the transcription from HTLV-1 provirus needs to be controlled in order to evade the host immune surveillance. There should be a dynamic and complex regulation in vivo, where an equilibrium between viral antigen expression and host immune surveillance is achieved. The mechanisms regulating viral gene expression from the provirus are a key to understanding the persistent/latent infection with HTLV-1 and its pathogenesis. In this article, we would like to review our current understanding on this topic. PMID:27322309

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

    PubMed

    Miyazato, Paola; Matsuo, Misaki; Katsuya, Hiroo; Satou, Yorifumi

    2016-01-01

    Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus associated with human diseases, such as adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/Tropic spastic paraparesis (HAM/TSP). As a retrovirus, its life cycle includes a step where HTLV-1 is integrated into the host genomic DNA and forms proviral DNA. In the chronic phase of the infection, HTLV‑1 is known to proliferate as a provirus via the mitotic division of the infected host cells. There are generally tens of thousands of infected clones within an infected individual. They exist not only in peripheral blood, but also in various lymphoid organs. Viral proteins encoded in HTLV-1 genome play a role in the proliferation and survival of the infected cells. As is the case with other chronic viral infections, HTLV-1 gene expression induces the activation of the host immunity against the virus. Thus, the transcription from HTLV-1 provirus needs to be controlled in order to evade the host immune surveillance. There should be a dynamic and complex regulation in vivo, where an equilibrium between viral antigen expression and host immune surveillance is achieved. The mechanisms regulating viral gene expression from the provirus are a key to understanding the persistent/latent infection with HTLV-1 and its pathogenesis. In this article, we would like to review our current understanding on this topic. PMID:27322309

  19. Alpha-amylase gene transcription in tissues of normal dog.

    PubMed

    Mocharla, H; Mocharla, R; Hodes, M E

    1990-02-25

    We studied the distribution of alpha-amylase mRNA in normal dog tissues by northern blotting (NB) and reverse transcription-polymerase chain reaction (RT-PCR) with human pancreatic (AMY2) and salivary (AMY1) alpha-amylase cDNA-specific primers. Analysis of poly(A+) RNA from various normal tissues by NB indicated the presence of detectable levels of alpha-amylase mRNA transcripts only in pancreas. Dot-blot analysis of DNA amplified with primers common to both (human) isoamylase mRNAs showed presence of alpha-amylase gene transcripts not only in pancreas but also in liver, small intestine, large intestine and fallopian tube. Traces of amylase gene transcripts were also observed in ovary, uterus and lung. Interestingly, amylase transcripts were not detectable in the parotid gland by NB or RT-PCR. We have also localized alpha-amylase mRNA transcripts to dog pancreas by in situ transcription and in situ hybridization. Our results suggest that there is high degree of homology between the alpha-amylase mRNA sequences in dog and human at least in the exon 3-4 regions of the human gene. PMID:2315015

  20. The Role of Multiple Transcription Factors In Archaeal Gene Expression

    SciTech Connect

    Charles J. Daniels

    2008-09-23

    Since the inception of this research program, the project has focused on two central questions: What is the relationship between the 'eukaryal-like' transcription machinery of archaeal cells and its counterparts in eukaryal cells? And, how does the archaeal cell control gene expression using its mosaic of eukaryal core transcription machinery and its bacterial-like transcription regulatory proteins? During the grant period we have addressed these questions using a variety of in vivo approaches and have sought to specifically define the roles of the multiple TATA binding protein (TBP) and TFIIB-like (TFB) proteins in controlling gene expression in Haloferax volcanii. H. volcanii was initially chosen as a model for the Archaea based on the availability of suitable genetic tools; however, later studies showed that all haloarchaea possessed multiple tbp and tfb genes, which led to the proposal that multiple TBP and TFB proteins may function in a manner similar to alternative sigma factors in bacterial cells. In vivo transcription and promoter analysis established a clear relationship between the promoter requirements of haloarchaeal genes and those of the eukaryal RNA polymerase II promoter. Studies on heat shock gene promoters, and the demonstration that specific tfb genes were induced by heat shock, provided the first indication that TFB proteins may direct expression of specific gene families. The construction of strains lacking tbp or tfb genes, coupled with the finding that many of these genes are differentially expressed under varying growth conditions, provided further support for this model. Genetic tools were also developed that led to the construction of insertion and deletion mutants, and a novel gene expression scheme was designed that allowed the controlled expression of these genes in vivo. More recent studies have used a whole genome array to examine the expression of these genes and we have established a linkage between the expression of specific tfb

  1. Alkane Biosynthesis Genes in Cyanobacteria and Their Transcriptional Organization

    PubMed Central

    Klähn, Stephan; Baumgartner, Desirée; Pfreundt, Ulrike; Voigt, Karsten; Schön, Verena; Steglich, Claudia; Hess, Wolfgang R.

    2014-01-01

    In cyanobacteria, alkanes are synthesized from a fatty acyl-ACP by two enzymes, acyl–acyl carrier protein reductase and aldehyde deformylating oxygenase. Despite the great interest in the exploitation for biofuel production, nothing is known about the transcriptional organization of their genes or the physiological function of alkane synthesis. The comparison of 115 microarray datasets indicates the relatively constitutive expression of aar and ado genes. The analysis of 181 available genomes showed that in 90% of the genomes both genes are present, likely indicating their physiological relevance. In 61% of them they cluster together with genes encoding acetyl-CoA carboxyl transferase and a short-chain dehydrogenase, strengthening the link to fatty acid metabolism and in 76% of the genomes they are located in tandem, suggesting constraints on the gene arrangement. However, contrary to the expectations for an operon, we found in Synechocystis sp. PCC 6803 specific promoters for the two genes, sll0208 (ado) and sll0209 (aar), which give rise to monocistronic transcripts. Moreover, the upstream located ado gene is driven by a proximal as well as a second, distal, promoter, from which a third transcript, the ~160 nt sRNA SyR9 is transcribed. Thus, the transcriptional organization of the alkane biosynthesis genes in Synechocystis sp. PCC 6803 is of substantial complexity. We verified all three promoters to function independently from each other and show a similar promoter arrangement also in the more distant Nodularia spumigena, Trichodesmium erythraeum, Anabaena sp. PCC 7120, Prochlorococcus MIT9313, and MED4. The presence of separate regulatory elements and the dominance of monocistronic mRNAs suggest the possible autonomous regulation of ado and aar. The complex transcriptional organization of the alkane synthesis gene cluster has possible metabolic implications and should be considered when manipulating the expression of these genes in cyanobacteria. PMID

  2. Dual requirement for the yeast MMS19 gene in DNA repair and RNA polymerase II transcription.

    PubMed Central

    Lauder, S; Bankmann, M; Guzder, S N; Sung, P; Prakash, L; Prakash, S

    1996-01-01

    Genetic and biochemical studies of Saccharomyces cerevisiae have indicated the involvement of a large number of protein factors in nucleotide excision repair (NER) of UV-damaged DNA. However, how MMS19 affects this process has remained unclear. Here, we report on the isolation of the MMS19 gene and the determination of its role in NER and other cellular processes. Genetic and biochemical evidence indicates that besides its function in NER, MMS19 also affects RNA polymerase II (Pol II) transcription. mms19delta cells do not grow at 37 degrees C, and mutant extract exhibits a thermolabile defect in Pol II transcription. Thus, Mms19 protein resembles TFIIH in that it is required for both transcription and DNA repair. However, addition of purified Mms19 protein does not alleviate the transcriptional defect of the mms19delta extract, nor does it stimulate the incision of UV-damaged DNA reconstituted from purified proteins. Interestingly, addition of purified TFIIH corrects the transcriptional defect of the mms19delta extract. Mms19 is, however, not a component of TFIIH or of Pol II holoenzyme. These and other results suggest that Mms19 affects NER and transcription by influencing the activity of TFIIH as an upstream regulatory element. It is proposed that mutations in the human MMS19 counterpart could result in syndromes in which both NER and transcription are affected. PMID:8943333

  3. Gene looping facilitates TFIIH kinase-mediated termination of transcription

    PubMed Central

    Medler, Scott; Ansari, Athar

    2015-01-01

    TFIIH is a general transcription factor with kinase and helicase activities. The kinase activity resides in the Kin28 subunit of TFIIH. The role of Kin28 kinase in the early steps of transcription is well established. Here we report a novel role of Kin28 in the termination of transcription. We show that RNAPII reads through a termination signal upon kinase inhibition. Furthermore, the recruitment of termination factors towards the 3′ end of a gene was compromised in the kinase mutant, thus confirming the termination defect. A concomitant decrease in crosslinking of termination factors near the 5′ end of genes was also observed in the kinase-defective mutant. Simultaneous presence of termination factors towards both the ends of a gene is indicative of gene looping; while the loss of termination factor occupancy from the distal ends suggest the abolition of a looped gene conformation. Accordingly, CCC analysis revealed that the looped architecture of genes was severely compromised in the Kin28 kinase mutant. In a looping defective sua7-1 mutant, even the enzymatically active Kin28 kinase could not rescue the termination defect. These results strongly suggest a crucial role of Kin28 kinase-dependent gene looping in the termination of transcription in budding yeast. PMID:26286112

  4. Spatially coordinated dynamic gene transcription in living pituitary tissue

    PubMed Central

    Featherstone, Karen; Hey, Kirsty; Momiji, Hiroshi; McNamara, Anne V; Patist, Amanda L; Woodburn, Joanna; Spiller, David G; Christian, Helen C; McNeilly, Alan S; Mullins, John J; Finkenstädt, Bärbel F; Rand, David A; White, Michael RH; Davis, Julian RE

    2016-01-01

    Transcription at individual genes in single cells is often pulsatile and stochastic. A key question emerges regarding how this behaviour contributes to tissue phenotype, but it has been a challenge to quantitatively analyse this in living cells over time, as opposed to studying snap-shots of gene expression state. We have used imaging of reporter gene expression to track transcription in living pituitary tissue. We integrated live-cell imaging data with statistical modelling for quantitative real-time estimation of the timing of switching between transcriptional states across a whole tissue. Multiple levels of transcription rate were identified, indicating that gene expression is not a simple binary ‘on-off’ process. Immature tissue displayed shorter durations of high-expressing states than the adult. In adult pituitary tissue, direct cell contacts involving gap junctions allowed local spatial coordination of prolactin gene expression. Our findings identify how heterogeneous transcriptional dynamics of single cells may contribute to overall tissue behaviour. DOI: http://dx.doi.org/10.7554/eLife.08494.001 PMID:26828110

  5. Expression of a Mutant kcnj2 Gene Transcript in Zebrafish

    PubMed Central

    Leong, Ivone U. S.; Skinner, Jonathan R.; Shelling, Andrew N.; Love, Donald R.

    2013-01-01

    Long QT 7 syndrome (LQT7, also known as Andersen-Tawil syndrome) is a rare autosomal-dominant disorder that causes cardiac arrhythmias, periodic paralysis, and dysmorphic features. Mutations in the human KCNJ2 gene, which encodes for the subunit of the potassium inwardly-rectifying channel (IK1), have been associated with the disorder. The majority of mutations are considered to be dominant-negative as mutant proteins interact to limit the function of wild type KCNJ2 proteins. Several LQT7 syndrome mouse models have been created that vary in the physiological similarity to the human disease. To complement the LQT7 mouse models, we investigated the usefulness of the zebrafish as an alternative model via a transient approach. Initial bioinformatic analysis identified the zebrafish orthologue of the human KCNJ2 gene, together with a spatial expression profile that was similar to that of human. The expression of a kcnj2-12 transcript carrying an in-frame deletion of critical amino acids identified in human studies resulted in embryos that exhibited defects in muscle development, thereby affecting movement, a decrease in jaw size, pupil-pupil distance, and signs of scoliosis. These defects correspond to some phenotypes expressed by human LQT7 patients.

  6. Quantification of Yeast and Bacterial Gene Transcripts in Retail Cheeses by Reverse Transcription-Quantitative PCR

    PubMed Central

    Straub, Cécile; Castellote, Jessie; Onesime, Djamila; Bonnarme, Pascal; Irlinger, Françoise

    2013-01-01

    The cheese microbiota contributes to a large extent to the development of the typical color, flavor, and texture of the final product. Its composition is not well defined in most cases and varies from one cheese to another. The aim of the present study was to establish procedures for gene transcript quantification in cheeses by reverse transcription-quantitative PCR. Total RNA was extracted from five smear-ripened cheeses purchased on the retail market, using a method that does not involve prior separation of microbial cells. 16S rRNA and malate:quinone oxidoreductase gene transcripts of Corynebacterium casei, Brevibacterium aurantiacum, and Arthrobacter arilaitensis and 26S rRNA and beta tubulin gene transcripts of Geotrichum candidum and Debaryomyces hansenii could be detected and quantified in most of the samples. Three types of normalization were applied: against total RNA, against the amount of cheese, and against a reference gene. For the first two types of normalization, differences of reverse transcription efficiencies from one sample to another were taken into account by analysis of exogenous control mRNA. No good correlation was found between the abundances of target mRNA or rRNA transcripts and the viable cell concentration of the corresponding species. However, in most cases, no mRNA transcripts were detected for species that did not belong to the dominant species. The applications of gene expression measurement in cheeses containing an undefined microbiota, as well as issues concerning the strategy of normalization and the assessment of amplification specificity, are discussed. PMID:23124230

  7. Circadian transcriptional regulation by the posttranslational oscillator without de novo clock gene expression in Synechococcus

    PubMed Central

    Hosokawa, Norimune; Hatakeyama, Tetsuhiro S.; Kojima, Takashi; Kikuchi, Yoshiyuki; Ito, Hiroshi; Iwasaki, Hideo

    2011-01-01

    Circadian rhythms are a fundamental property of most organisms, from cyanobacteria to humans. In the unicellular obligately photoautotrophic cyanobacterium Synechococcus elongatus PCC 7942, essentially all promoter activities are controlled by the KaiABC-based clock under continuous light conditions. When Synechococcus cells are transferred from the light to continuous dark (DD) conditions, the expression of most genes, including the clock genes kaiA and kaiBC, is rapidly down-regulated, whereas the KaiC phosphorylation cycle persists. Therefore, we speculated that the posttranslational oscillator might not drive the transcriptional circadian output without de novo expression of the kai genes. Here we show that the cyanobacterial clock regulates the transcriptional output even in the dark. The expression of a subset of genes in the genomes of cells grown in the dark was dramatically affected by kaiABC nullification, and the magnitude of dark induction was dependent on the time at which the cells were transferred from the light to the dark. Moreover, under DD conditions, the expression of some dark-induced gene transcripts exhibited temperature-compensated damped oscillations, which were nullified in kaiABC-null strains and were affected by a kaiC period mutation. These results indicate that the Kai protein-based posttranslational oscillator can drive the circadian transcriptional output even without the de novo expression of the clock genes. PMID:21896749

  8. A transcription map of a yeast centromere plasmid: unexpected transcripts and altered gene expression.

    PubMed Central

    Marczynski, G T; Jaehning, J A

    1985-01-01

    YCp19 is a yeast centromere plasmid capable of autonomous replication in both yeast and E. coli (J. Mol. Biol., 158: 157-179, 1982). It is stably maintained as a single copy in the yeast cell and is therefore a model yeast "minichromosome" and cloning vector. We have located the positions and measured the abundance of the in vivo yeast transcripts from YCp19. Transcripts from the selectable marker genes TRP1 and URA3 were present at increased levels relative to chromosomal copies of the genes. Unanticipated transcripts from the yeast CEN4 and E. coli pBR322 sequences were also found. Although much of the plasmid vector is actively transcribed in vivo, the regions around the most useful cloning sites (BamHI, EcoRI, SalI) are free of transcripts. We have analyzed transcription of BamHI inserts containing promoter variants of the HIS3 gene and determined that although initiation events are accurate, plasmid context may alter levels of gene expression. Images PMID:3909105

  9. Molecular weight abnormalities of the CTCF transcription factor: CTCF migrates aberrantly in SDS-PAGE and the size of the expressed protein is affected by the UTRs and sequences within the coding region of the CTCF gene.

    PubMed Central

    Klenova, E M; Nicolas, R H; U, S; Carne, A F; Lee, R E; Lobanenkov, V V; Goodwin, G H

    1997-01-01

    CTCF belongs to the Zn finger transcription factors family and binds to the promoter region of c-myc. CTCF is highly conserved between species, ubiquitous and localised in nuclei. The endogenous CTCF migrates as a 130 kDa (CTCF-130) protein on SDS-PAGE, however, the open reading frame (ORF) of the CTCF cDNA encodes only a 82 kDa protein (CTCF-82). In the present study we investigate this phenomenon and show with mass-spectra analysis that this occurs due to aberrant mobility of the CTCF protein. Another paradox is that our original cDNA, composed of the ORF and 3'-untranslated region (3'-UTR), produces a protein with the apparent molecular weight of 70 kDa (CTCF-70). This paradox has been found to be an effect of the UTRs and sequences within the coding region of the CTCF gene resulting in C-terminal truncation of CTCF-130. The potential attenuator has been identified and point-mutated. This restored the electrophoretic mobility of the CTCF protein to 130 kDa. CTCF-70, the aberrantly migrating CTCF N-terminus per se, is also detected in some cell types and therefore may have some biological implications. In particular, CTCF-70 interferes with CTCF-130 normal function, enhancing transactivation induced by CTCF-130 in COS6 cells. The mechanism of CTCF-70 action and other possible functions of CTCF-70 are discussed. PMID:9016583

  10. Transcription of Inflammatory Genes: Long Noncoding RNA and Beyond

    PubMed Central

    Carpenter, Susan

    2015-01-01

    The innate immune system must coordinate elaborate signaling pathways to turn on expression of hundreds of genes to provide protection against pathogens and resolve acute inflammation. Multiple genes within distinct functional categories are coordinately and temporally regulated by transcriptional on and off switches in response to distinct external stimuli. Three classes of transcription factors act together with transcriptional coregulators and chromatin-modifying complexes to control these programs. In addition, newer studies implicate long noncoding RNA (lncRNA) as additional regulators of these responses. LncRNAs promote, fine-tune, and restrain the inflammatory program. In this study, we provide an overview of gene regulation and the emerging importance of lncRNAs in the immune system. PMID:25250698

  11. Transcriptional targeting of tumor endothelial cells for gene therapy

    PubMed Central

    Dong, Zhihong; Nör, Jacques E.

    2009-01-01

    It is well known that angiogenesis plays a critical role in the pathobiology of tumors. Recent clinical trials have shown that inhibition of angiogenesis can be an effective therapeutic strategy for patients with cancer. However, one of the outstanding issues in anti-angiogenic treatment for cancer is the development of toxicities related to off-target effects of drugs. Transcriptional targeting of tumor endothelial cells involves the use of specific promoters for selective expression of therapeutic genes in the endothelial cells lining the blood vessels of tumors. Recently, several genes that are expressed specifically in tumor-associated endothelial cells have been identified and characterized. These discoveries have enhanced the prospectus of transcriptionaly targeting tumor endothelial cells for cancer gene therapy. In this manuscript, we review the promoters, vectors, and therapeutic genes that have been used for transcriptional targeting of tumor endothelial cells, and discuss the prospects of such approaches for cancer gene therapy. PMID:19393703

  12. Single nucleotide variants in transcription factors associate more tightly with phenotype than with gene expression.

    PubMed

    Sudarsanam, Priya; Cohen, Barak A

    2014-05-01

    Mapping the polymorphisms responsible for variation in gene expression, known as Expression Quantitative Trait Loci (eQTL), is a common strategy for investigating the molecular basis of disease. Despite numerous eQTL studies, the relationship between the explanatory power of variants on gene expression versus their power to explain ultimate phenotypes remains to be clarified. We addressed this question using four naturally occurring Quantitative Trait Nucleotides (QTN) in three transcription factors that affect sporulation efficiency in wild strains of the yeast, Saccharomyces cerevisiae. We compared the ability of these QTN to explain the variation in both gene expression and sporulation efficiency. We find that the amount of gene expression variation explained by the sporulation QTN is not predictive of the amount of phenotypic variation explained. The QTN are responsible for 98% of the phenotypic variation in our strains but the median gene expression variation explained is only 49%. The alleles that are responsible for most of the variation in sporulation efficiency do not explain most of the variation in gene expression. The balance between the main effects and gene-gene interactions on gene expression variation is not the same as on sporulation efficiency. Finally, we show that nucleotide variants in the same transcription factor explain the expression variation of different sets of target genes depending on whether the variant alters the level or activity of the transcription factor. Our results suggest that a subset of gene expression changes may be more predictive of ultimate phenotypes than the number of genes affected or the total fraction of variation in gene expression variation explained by causative variants, and that the downstream phenotype is buffered against variation in the gene expression network. PMID:24784239

  13. Impact of ACTH Signaling on Transcriptional Regulation of Steroidogenic Genes

    PubMed Central

    Ruggiero, Carmen; Lalli, Enzo

    2016-01-01

    The trophic peptide hormone adrenocorticotropic (ACTH) stimulates steroid hormone biosynthesis evoking both a rapid, acute response and a long-term, chronic response, via the activation of cAMP/protein kinase A (PKA) signaling. The acute response is initiated by the mobilization of cholesterol from lipid stores and its delivery to the inner mitochondrial membrane, a process that is mediated by the steroidogenic acute regulatory protein. The chronic response results in the increased coordinated transcription of genes encoding steroidogenic enzymes. ACTH binding to its cognate receptor, melanocortin 2 receptor (MC2R), stimulates adenylyl cyclase, thus inducing cAMP production, PKA activation, and phosphorylation of specific nuclear factors, which bind to target promoters and facilitate coactivator protein recruitment to direct steroidogenic gene transcription. This review provides a general view of the transcriptional control exerted by the ACTH/cAMP system on the expression of genes encoding for steroidogenic enzymes in the adrenal cortex. Special emphasis will be given to the transcription factors required to mediate ACTH-dependent transcription of steroidogenic genes. PMID:27065945

  14. Regulation of neural gene transcription by optogenetic inhibition of the RE1-silencing transcription factor.

    PubMed

    Paonessa, Francesco; Criscuolo, Stefania; Sacchetti, Silvio; Amoroso, Davide; Scarongella, Helena; Pecoraro Bisogni, Federico; Carminati, Emanuele; Pruzzo, Giacomo; Maragliano, Luca; Cesca, Fabrizia; Benfenati, Fabio

    2016-01-01

    Optogenetics provides new ways to activate gene transcription; however, no attempts have been made as yet to modulate mammalian transcription factors. We report the light-mediated regulation of the repressor element 1 (RE1)-silencing transcription factor (REST), a master regulator of neural genes. To tune REST activity, we selected two protein domains that impair REST-DNA binding or recruitment of the cofactor mSin3a. Computational modeling guided the fusion of the inhibitory domains to the light-sensitive Avena sativa light-oxygen-voltage-sensing (LOV) 2-phototrophin 1 (AsLOV2). By expressing AsLOV2 chimeras in Neuro2a cells, we achieved light-dependent modulation of REST target genes that was associated with an improved neural differentiation. In primary neurons, light-mediated REST inhibition increased Na(+)-channel 1.2 and brain-derived neurotrophic factor transcription and boosted Na(+) currents and neuronal firing. This optogenetic approach allows the coordinated expression of a cluster of genes impinging on neuronal activity, providing a tool for studying neuronal physiology and correcting gene expression changes taking place in brain diseases. PMID:26699507

  15. Regulation of neural gene transcription by optogenetic inhibition of the RE1-silencing transcription factor

    PubMed Central

    Paonessa, Francesco; Criscuolo, Stefania; Sacchetti, Silvio; Amoroso, Davide; Scarongella, Helena; Pecoraro Bisogni, Federico; Carminati, Emanuele; Pruzzo, Giacomo; Maragliano, Luca; Cesca, Fabrizia; Benfenati, Fabio

    2016-01-01

    Optogenetics provides new ways to activate gene transcription; however, no attempts have been made as yet to modulate mammalian transcription factors. We report the light-mediated regulation of the repressor element 1 (RE1)-silencing transcription factor (REST), a master regulator of neural genes. To tune REST activity, we selected two protein domains that impair REST-DNA binding or recruitment of the cofactor mSin3a. Computational modeling guided the fusion of the inhibitory domains to the light-sensitive Avena sativa light–oxygen–voltage-sensing (LOV) 2-phototrophin 1 (AsLOV2). By expressing AsLOV2 chimeras in Neuro2a cells, we achieved light-dependent modulation of REST target genes that was associated with an improved neural differentiation. In primary neurons, light-mediated REST inhibition increased Na+-channel 1.2 and brain-derived neurotrophic factor transcription and boosted Na+ currents and neuronal firing. This optogenetic approach allows the coordinated expression of a cluster of genes impinging on neuronal activity, providing a tool for studying neuronal physiology and correcting gene expression changes taking place in brain diseases. PMID:26699507

  16. Simultaneous live imaging of the transcription and nuclear position of specific genes

    PubMed Central

    Ochiai, Hiroshi; Sugawara, Takeshi; Yamamoto, Takashi

    2015-01-01

    The relationship between genome organization and gene expression has recently been established. However, the relationships between spatial organization, dynamics, and transcriptional regulation of the genome remain unknown. In this study, we developed a live-imaging method for simultaneous measurements of the transcriptional activity and nuclear position of endogenous genes, which we termed the ‘Real-time Observation of Localization and EXpression (ROLEX)’ system. We demonstrated that ROLEX is highly specific and does not affect the expression level of the target gene. ROLEX enabled detection of sub-genome-wide mobility changes that depended on the state of Nanog transactivation in embryonic stem cells. We believe that the ROLEX system will become a powerful tool for exploring the relationship between transcription and nuclear dynamics in living cells. PMID:26092696

  17. The CREB Transcription Factor Controls Transcriptional Activity of the Human RIC8B Gene.

    PubMed

    Maureira, Alejandro; Sánchez, Rodolfo; Valenzuela, Nicole; Torrejón, Marcela; Hinrichs, María V; Olate, Juan; Gutiérrez, José L

    2016-08-01

    Proper regulation of gene expression is essential for normal development, cellular growth, and differentiation. Differential expression profiles of mRNA coding for vertebrate Ric-8B during embryo and adult stages have been observed. In addition, Ric-8B is expressed in few cerebral nuclei subareas. These facts point to a dynamic control of RIC8B gene expression. In order to understand the transcriptional regulation of this gene, we searched for cis-elements in the sequence of the human RIC8B promoter region, identifying binding sites for the basic/leucine zipper (bZip) CREB transcription factor family (CRE sites) and C/EBP transcription factor family (C/EBP sites). CRE sites were found clustered near the transcription start site, while the C/EBP sites were found clustered at around 300 bp upstream the CRE sites. Here, we demonstrate the ability of CREB1 and C/EBPβ to bind their respective elements identified in the RIC8B promoter. Comparative protein-DNA interaction analyses revealed only the proximal elements as high affinity sites for CREB1 and only the distal elements as high affinity sites for C/EBPβ. Chromatin immunoprecipitation analyses, carried out using a human neuroblastoma cell line, confirmed the preferential association of CREB to the proximal region of the RIC8B promoter. By performing luciferase reporter assays, we found the CRE sites as the most relevant elements for its transcriptional activity. Taken together, these data show the existence of functional CREB and C/EBP binding sites in the human RIC8B gene promoter, a particular distribution of these sites and demonstrate a relevant role of CREB in stimulating transcriptional activity of this gene. J. Cell. Biochem. 117: 1797-1805, 2016. © 2016 Wiley Periodicals, Inc. PMID:26729411

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

    PubMed

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

    2016-01-01

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

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

    PubMed Central

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

    2016-01-01

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

  20. Transcriptional promiscuity of the human /alpha/-globin gene

    SciTech Connect

    Whitelaw, E.; Hogben, P.; Hanscombe, O.; Proudfoot, N.J.

    1989-01-01

    The human /alpha/-globin gene displays the unusual property of transcriptional promiscuity: that is, it functions in the absence of an enhancer when transfected into nonerythroid cell lines. It is also unusual in that its promoter region lies in a hypomethylated HpaII tiny fragment (HTF) island containing multiple copies of the consensus sequence for the SP1-binding site. The authors have investigated whether there is a relationship between these two observations. First, they investigated the mouse /alpha/-globin gene since it does not lie in an HTF island. They have demonstrated that it was not transcriptionally promiscuous. Second, they studied the transcriptional activity of the human /alpha/-globin gene in the absence of the GC-rich region containing putative SP1-binding sites and found a small (two- to threefold) but consistent positive effect of this region on transcriptional activity in both nonerythroid and erythroid cell lines. However, this effect did not account for the promiscuous nature of the human /alpha/-globin gene. They found that in a nonreplicating system, the human //a/-globin gene, like that of the mouse, required a simian virus 40 enhancer in order to be transcriptionally active in nonerythroid and erythroid cell lines. Since they only observed enhancer independence of the human /alpha/-globin gene in a high-copy-number replicating system, they suggest that competition for trans-acting factors could explain these results. Finally, the authors' experiments with the erythroid cell line Putko suggest that there are no tissue-specific enhancers within 1 kilobase 5' of the human /alpha/-globin cap site or within the gene itself.

  1. Epigenetic regulation of intragenic transposable elements impacts gene transcription in Arabidopsis thaliana

    PubMed Central

    Le, Tu N.; Miyazaki, Yuji; Takuno, Shohei; Saze, Hidetoshi

    2015-01-01

    Genomes of higher eukaryotes, including plants, contain numerous transposable elements (TEs), that are often silenced by epigenetic mechanisms, such as histone modifications and DNA methylation. Although TE silencing adversely affects expression of nearby genes, recent studies reveal the presence of intragenic TEs marked by repressive heterochromatic epigenetic marks within transcribed genes. However, even for the well-studied plant model Arabidopsis thaliana, the abundance of intragenic TEs, how they are epigenetically regulated, and their potential impacts on host gene expression, remain unexplored. In this study, we comprehensively analyzed genome-wide distribution and epigenetic regulation of intragenic TEs in A. thaliana. Our analysis revealed that about 3% of TEs are located within gene bodies, dominantly at intronic regions. Most of them are shorter and less methylated than intergenic TEs, but they are still targeted by RNA-directed DNA methylation-dependent and independent pathways. Surprisingly, the heterochromatic epigenetic marks at TEs are maintained within actively transcribed genes. Moreover, the heterochromatic state of intronic TEs is critical for proper transcription of associated genes. Our study provides the first insight into how intragenic TEs affect the transcriptional landscape of the A. thaliana genome, and suggests the importance of epigenetic mechanisms for regulation of TEs within transcriptional gene units. PMID:25813042

  2. Transcriptional regulation of human small nuclear RNA genes

    PubMed Central

    Jawdekar, Gauri W.; Henry, R. William

    2009-01-01

    The products of human snRNA genes have been frequently described as performing housekeeping functions and their synthesis refractory to regulation. However, recent studies have emphasized that snRNA and other related non-coding RNA molecules control multiple facets of the central dogma, and their regulated expression is critical to cellular homeostasis during normal growth and in response to stress. Human snRNA genes contain compact and yet powerful promoters that are recognized by increasingly well-characterized transcription factors, thus providing a premier model system to study gene regulation. This review summarizes many recent advances deciphering the mechanism by which the transcription of human snRNA and related genes are regulated. PMID:18442490

  3. Association of a transcription factor 21 gene polymorphism with hypertension

    PubMed Central

    FUJIMAKI, TETSUO; OGURI, MITSUTOSHI; HORIBE, HIDEKI; KATO, KIMIHIKO; MATSUOKA, REIKO; ABE, SHINTARO; TOKORO, FUMITAKA; ARAI, MASAZUMI; NODA, TOSHIYUKI; WATANABE, SACHIRO; YAMADA, YOSHIJI

    2015-01-01

    Various loci and genes that confer susceptibility to coronary artery disease (CAD) have been identified mainly in Caucasian populations by genome-wide association studies (GWASs). As hypertension is a major risk factor for CAD, certain polymorphisms may contribute to the genetic susceptibility to CAD through affecting the predisposition to hypertension. The aim of the present study was to examine a possible association of hypertension with 29 single-nucleotide polymorphisms (SNPs) previously identified by meta-analyses of GWASs as susceptibility loci for CAD. Study subjects comprised of 5,460 individuals (3,348 subjects with hypertension and 2,112 controls). The genotypes of SNPs were determined by the multiplex bead-based Luminex assay. The χ2 test revealed that genotype distributions and allele frequencies for rs12190287 of the transcription factor 21 gene (TCF21) and rs1122608 of the SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a, member 4 gene (SMARCA4) were significantly (P<0.05) associated with hypertension. Allele frequencies for rs9369640 of the phosphatase and actin regulator 1 gene (PHACTR1) and genotype distributions for rs599839 of the proline/serine-rich coiled-coil 1 gene (PSRC1) were also significantly associated with hypertension. Multivariable logistic regression analysis with adjustment for age, gender, body mass index and smoking status revealed that rs12190287 of TCF21 (P=0.0014; recessive model; odds ratio, 1.21) was significantly associated with hypertension, and the C allele represented a risk factor for this condition. Similar analyses revealed that rs1122608 of SMARCA4 (P=0.0305; dominant model; odds ratio, 0.86), rs9369640 of PHACTR1 (P=0.0119; dominant model; odds ratio, 0.82) and rs599839 of PSRC1 (P=0.0248; dominant model; odds ratio, 0.84) were also related to hypertension, with the minor T, C and G alleles, respectively, being protective against this condition. Thus, the present results

  4. Association of a transcription factor 21 gene polymorphism with hypertension.

    PubMed

    Fujimaki, Tetsuo; Oguri, Mitsutoshi; Horibe, Hideki; Kato, Kimihiko; Matsuoka, Reiko; Abe, Shintaro; Tokoro, Fumitaka; Arai, Masazumi; Noda, Toshiyuki; Watanabe, Sachiro; Yamada, Yoshiji

    2015-01-01

    Various loci and genes that confer susceptibility to coronary artery disease (CAD) have been identified mainly in Caucasian populations by genome-wide association studies (GWASs). As hypertension is a major risk factor for CAD, certain polymorphisms may contribute to the genetic susceptibility to CAD through affecting the predisposition to hypertension. The aim of the present study was to examine a possible association of hypertension with 29 single-nucleotide polymorphisms (SNPs) previously identified by meta-analyses of GWASs as susceptibility loci for CAD. Study subjects comprised of 5,460 individuals (3,348 subjects with hypertension and 2,112 controls). The genotypes of SNPs were determined by the multiplex bead-based Luminex assay. The χ(2) test revealed that genotype distributions and allele frequencies for rs12190287 of the transcription factor 21 gene (TCF21) and rs1122608 of the SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a, member 4 gene (SMARCA4) were significantly (P<0.05) associated with hypertension. Allele frequencies for rs9369640 of the phosphatase and actin regulator 1 gene (PHACTR1) and genotype distributions for rs599839 of the proline/serine-rich coiled-coil 1 gene (PSRC1) were also significantly associated with hypertension. Multivariable logistic regression analysis with adjustment for age, gender, body mass index and smoking status revealed that rs12190287 of TCF21 (P=0.0014; recessive model; odds ratio, 1.21) was significantly associated with hypertension, and the C allele represented a risk factor for this condition. Similar analyses revealed that rs1122608 of SMARCA4 (P=0.0305; dominant model; odds ratio, 0.86), rs9369640 of PHACTR1 (P=0.0119; dominant model; odds ratio, 0.82) and rs599839 of PSRC1 (P=0.0248; dominant model; odds ratio, 0.84) were also related to hypertension, with the minor T, C and G alleles, respectively, being protective against this condition. Thus, the present results

  5. The 26S Proteasome and Initiation of Gene Transcription

    PubMed Central

    Durairaj, Geetha; Kaiser, Peter

    2014-01-01

    Transcription activation is the foremost step of gene expression and is modulated by various factors that act in synergy. Misregulation of this process and its associated factors has severe effects and hence requires strong regulatory control. In recent years, growing evidence has highlighted the 26S proteasome as an important contributor to the regulation of transcription initiation. Well known for its role in protein destruction, its contribution to protein synthesis was initially viewed with skepticism. However, studies over the past several years have established the proteasome as an important component of transcription initiation through proteolytic and non-proteolytic activities. In this review, we discuss findings made so far in understanding the connections between transcription initiation and the 26S proteasome complex. PMID:25211636

  6. The Transcription Factor Titration Effect Dictates Level of Gene Expression

    PubMed Central

    Brewster, Robert C.; Weinert, Franz M.; Garcia, Hernan G.; Song, Dan; Rydenfelt, Mattias; Phillips, Rob

    2014-01-01

    Models of transcription are often built around a picture of RNA polymerase and transcription factors (TFs) acting on a single copy of a promoter. However, most TFs are shared between multiple genes with varying binding affinities. Beyond that, genes often exist at high copy number; in multiple, identical copies on the chromosome or on plasmids or viral vectors with copy numbers in the hundreds. Using a thermodynamic model, we characterize the interplay between TF copy number and the demand for that TF. We demonstrate the parameter-free predictive power of this model as a function of the copy number of the TF and the number and affinities of the available specific binding sites; such predictive control is important for the understanding of transcription and the desire to quantitatively design the output of genetic circuits. Finally we use these experiments to dynamically measure plasmid copy number through the cell cycle. PMID:24612990

  7. Synthetic Transcription Amplifier System for Orthogonal Control of Gene Expression in Saccharomyces cerevisiae

    PubMed Central

    Rantasalo, Anssi; Czeizler, Elena; Virtanen, Riitta; Rousu, Juho; Lähdesmäki, Harri; Penttilä, Merja

    2016-01-01

    This work describes the development and characterization of a modular synthetic expression system that provides a broad range of adjustable and predictable expression levels in S. cerevisiae. The system works as a fixed-gain transcription amplifier, where the input signal is transferred via a synthetic transcription factor (sTF) onto a synthetic promoter, containing a defined core promoter, generating a transcription output signal. The system activation is based on the bacterial LexA-DNA-binding domain, a set of modified, modular LexA-binding sites and a selection of transcription activation domains. We show both experimentally and computationally that the tuning of the system is achieved through the selection of three separate modules, each of which enables an adjustable output signal: 1) the transcription-activation domain of the sTF, 2) the binding-site modules in the output promoter, and 3) the core promoter modules which define the transcription initiation site in the output promoter. The system has a novel bidirectional architecture that enables generation of compact, yet versatile expression modules for multiple genes with highly diversified expression levels ranging from negligible to very strong using one synthetic transcription factor. In contrast to most existing modular gene expression regulation systems, the present system is independent from externally added compounds. Furthermore, the established system was minimally affected by the several tested growth conditions. These features suggest that it can be highly useful in large scale biotechnology applications. PMID:26901642

  8. Synthetic Transcription Amplifier System for Orthogonal Control of Gene Expression in Saccharomyces cerevisiae.

    PubMed

    Rantasalo, Anssi; Czeizler, Elena; Virtanen, Riitta; Rousu, Juho; Lähdesmäki, Harri; Penttilä, Merja; Jäntti, Jussi; Mojzita, Dominik

    2016-01-01

    This work describes the development and characterization of a modular synthetic expression system that provides a broad range of adjustable and predictable expression levels in S. cerevisiae. The system works as a fixed-gain transcription amplifier, where the input signal is transferred via a synthetic transcription factor (sTF) onto a synthetic promoter, containing a defined core promoter, generating a transcription output signal. The system activation is based on the bacterial LexA-DNA-binding domain, a set of modified, modular LexA-binding sites and a selection of transcription activation domains. We show both experimentally and computationally that the tuning of the system is achieved through the selection of three separate modules, each of which enables an adjustable output signal: 1) the transcription-activation domain of the sTF, 2) the binding-site modules in the output promoter, and 3) the core promoter modules which define the transcription initiation site in the output promoter. The system has a novel bidirectional architecture that enables generation of compact, yet versatile expression modules for multiple genes with highly diversified expression levels ranging from negligible to very strong using one synthetic transcription factor. In contrast to most existing modular gene expression regulation systems, the present system is independent from externally added compounds. Furthermore, the established system was minimally affected by the several tested growth conditions. These features suggest that it can be highly useful in large scale biotechnology applications. PMID:26901642

  9. Transcription control of ribulose bisphosphate carboxylase/oxygenase activase and adjacent genes in Anabaena species.

    PubMed Central

    Li, L A; Tabita, F R

    1994-01-01

    The gene encoding ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) activase (rca) was uniformly localized downstream from the genes encoding the large and small subunits of RubisCO (rbcL and rbcS) in three strains of Anabaena species. However, two open reading frames (ORF1 and ORF2), situated between rbcS and rca in Anabaena sp. strain CA, were not found in the intergenic region of Anabaena variabilis and Anabaena sp. strain PCC 7120. During autotrophic growth of Anabaena cells, rca and rbc transcripts accumulated in the light and diminished in the dark; light-dependent expression of these genes was not affected by the nitrogen source and the concentration of exogenous CO2 supplied to the cells. When grown on fructose, rca- and rbc-specific transcripts accumulated in A. variabilis regardless of whether the cells were illuminated. Transcript levels, however, were much lower in dark-grown heterotrophic cultures than in photoheterotrophic cultures. In photoheterotrophic cultures, the expression of the rca and rbc genes was similar to that in cultures grown with CO2 as the sole source of carbon. Although the rbcL-rbcS and rca genes are linked and are in the same transcriptional orientation in Anabaena strains, hybridization of rbc and rca to distinct transcripts suggested that these genes are not cotranscribed, consistent with the results of primer extension and secondary structure analysis of the nucleotide sequence. Transcription from ORF1 and ORF2 was not detected under the conditions examined, and the function of these putative genes remains unknown. Images PMID:7961423

  10. Dietary control of late trypsin gene transcription in Aedes aegypti.

    PubMed

    Noriega, F G; Barillas-Mury, C; Wells, M A

    1994-06-01

    In Aedes aegypti the levels of midgut trypsin activity after feeding are directly proportional to the protein concentration in the meal. The mechanisms of this up-regulatory event were investigated by analyzing the expression of the late trypsin gene under different dietary conditions. Transcription of the gene was dependent on both the quality and quantity of protein in the meal. As measured by Northern blot analysis, the levels of late trypsin gene expression increased up to 100-fold 24 h after feeding on gamma-globulin, hemoglobin or albumin (100 mg/ml). In contrast, gelatin, histone, amino acids, saline or agarose were very poor inducers of transcription. The rates of late trypsin transcription induced during the first 24 h were directly proportional to the concentration of protein in the meal. These data further support the suggestion that the primary mechanism that regulates the synthesis of trypsin in the mosquito midgut is transcriptional regulation of the gene. This regulatory mechanism enables the midgut to maintain the appropriate balance between protease synthesis and the protein content of the meal. PMID:7519098

  11. Transcription Profiling and Mutation Detection of Soybean Homoeologous Genes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The soybean genome maintains numerous gene duplications, many of which are derived from ancient large-scale duplication. We are interested in exploring the evolutionary fate of duplicated genes and the extent to which gene duplication affects selectable trait variation. We are applying quantitative ...

  12. Transcription of the putative tumor suppressor gene HCCS1 requires binding of ETS-2 to its consensus near the transcription start site.

    PubMed

    Zhu, Jing De; Fei, Qi; Wang, Peng; Lan, Fei; Mao, Da Qin; Zhang, Hong Yu; Yao, Xue Biao

    2006-09-01

    The hepatocellular carcinoma suppressor 1 (HCCS1) gene was identified by both positional cloning from a predominant region of loss of heterozygosity (17p13.3) in liver cancer and by functional screening for genes affecting cell proliferation in large-scale transfection assays. Its overexpression results in inhibition of cell proliferation in cell culture and tumor growth in nude mice. To understand its transcription regulation, the promoter architecture has been dissected in detail. The major start of transcription was mapped by primer extension to a C residue, 177 nucleotides upstream of the ATG codon. By assessing the promoter activity of a set of linker-scanning mutants of the minimal promoter (-60 to +148 region) in a transient transfection assay, we found that the +1 to + 40 region is critical to HCCS1 gene transcription, containing binding sites for transcription factors NF-kappaB (-21 to +7 and +40 to +26), p53 (+29 to +9) and ETS (+4 to +20 and +23 to +39). Biochemical and molecular analyses revealed that the ETS transcription factors ETS-2 and Elf-1 bind to the two ETS sites in situ and contribute significantly to the transcriptionally active state of the HCCS1 gene, while NF-kappaB, p53 and two other members of the ETS family (ETS-1 and NERF2) appear to play little role. Our observations provide insight into the mechanistic aspects of HCCS1 transcription regulation. PMID:16953216

  13. Transcriptional regulation of Bacillus subtilis citrate synthase genes.

    PubMed Central

    Jin, S; Sonenshein, A L

    1994-01-01

    The Bacillus subtilis citrate synthase genes citA and citZ were repressed during early exponential growth phase in nutrient broth medium and were induced as cells reached the end of exponential phase. Both genes were also induced by treatment of cells with the drug decoyinine. After induction, the steady-state level of citZ mRNA was about five times higher than that of citA mRNA. At least some of the citZ transcripts read through into the isocitrate dehydrogenase (citC) gene. Transcription from an apparent promoter site located near the 3' end of the citZ gene also contributed to expression of citC. In minimal medium, citA transcription was about 6-fold lower when glucose was the sole carbon source than it was when succinate was the carbon source. Expression of the citZ gene was repressed 2-fold by glucose and 10-fold when glucose and glutamate were present simultaneously. This latter synergistic repression is similar to the effect of glucose and glutamate on steady-state citrate synthase enzyme activity. CitR, a protein of the LysR family, appeared to be a repressor of citA but not of citZ. Images PMID:8045899

  14. Cohesin: genomic insights into controlling gene transcription and development

    PubMed Central

    Dorsett, Dale

    2011-01-01

    Over the past decade it has emerged that the cohesin protein complex, which functions in sister chromatid cohesion, chromosome segregation and DNA repair, also regulates gene expression and development. Even minor changes in cohesin activity alter several aspects of development. Genome-wide analysis indicates that cohesin directly regulates transcription of genes involved in cell proliferation, pluripotency, and differentiation through multiple mechanisms. These mechanisms are poorly understood, but involve both partial gene repression in concert with Polycomb group proteins, and facilitating long-range looping, both between enhancers and promoters, and between CTCF protein binding sites. PMID:21324671

  15. Transcriptional control of human p53-regulated genes.

    PubMed

    Riley, Todd; Sontag, Eduardo; Chen, Patricia; Levine, Arnold

    2008-05-01

    The p53 protein regulates the transcription of many different genes in response to a wide variety of stress signals. Following DNA damage, p53 regulates key processes, including DNA repair, cell-cycle arrest, senescence and apoptosis, in order to suppress cancer. This Analysis article provides an overview of the current knowledge of p53-regulated genes in these pathways and others, and the mechanisms of their regulation. In addition, we present the most comprehensive list so far of human p53-regulated genes and their experimentally validated, functional binding sites that confer p53 regulation. PMID:18431400

  16. Gene transcription in polar bears (Ursus maritimus) from disparate populations

    USGS Publications Warehouse

    Bowen, Lizabeth; Miles, A. Keith; Waters, Shannon C.; Meyerson, Randi; Rode, Karyn D.; Atwood, Todd C.

    2015-01-01

    Polar bears in the Beaufort (SB) and Chukchi (CS) Seas experience different environments due primarily to a longer history of sea ice loss in the Beaufort Sea. Ecological differences have been identified as a possible reason for the generally poorer body condition and reproduction of Beaufort polar bears compared to those from the Chukchi, but the influence of exposure to other stressors remains unknown. We use molecular technology, quantitative PCR, to identify gene transcription differences among polar bears from the Beaufort and Chukchi Seas as well as captive healthy polar bears. We identified significant transcriptional differences among a priori groups (i.e., captive bears, SB 2012, SB 2013, CS 2013) for ten of the 14 genes of interest (i.e., CaM, HSP70, CCR3, TGFβ, COX2, THRα, T-bet, Gata3, CD69, and IL17); transcription levels of DRβ, IL1β, AHR, and Mx1 did not differ among groups. Multivariate analysis also demonstrated separation among the groups of polar bears. Specifically, we detected transcript profiles consistent with immune function impairment in polar bears from the Beaufort Sea, when compared with Chukchi and captive polar bears. Although there is no strong indication of differential exposure to contaminants or pathogens between CS and SB bears, there are clearly differences in important transcriptional responses between populations. Further investigation is warranted to refine interpretation of potential effects of described stress-related conditions for the SB population.

  17. A combination of independent transcriptional regulators shapes bacterial virulence gene expression during infection.

    PubMed

    Shelburne, Samuel A; Olsen, Randall J; Suber, Bryce; Sahasrabhojane, Pranoti; Sumby, Paul; Brennan, Richard G; Musser, James M

    2010-03-01

    Transcriptional regulatory networks are fundamental to how microbes alter gene expression in response to environmental stimuli, thereby playing a critical role in bacterial pathogenesis. However, understanding how bacterial transcriptional regulatory networks function during host-pathogen interaction is limited. Recent studies in group A Streptococcus (GAS) suggested that the transcriptional regulator catabolite control protein A (CcpA) influences many of the same genes as the control of virulence (CovRS) two-component gene regulatory system. To provide new information about the CcpA and CovRS networks, we compared the CcpA and CovR transcriptomes in a serotype M1 GAS strain. The transcript levels of several of the same genes encoding virulence factors and proteins involved in basic metabolic processes were affected in both DeltaccpA and DeltacovR isogenic mutant strains. Recombinant CcpA and CovR bound with high-affinity to the promoter regions of several co-regulated genes, including those encoding proteins involved in carbohydrate and amino acid metabolism. Compared to the wild-type parental strain, DeltaccpA and DeltacovRDeltaccpA isogenic mutant strains were significantly less virulent in a mouse myositis model. Inactivation of CcpA and CovR alone and in combination led to significant alterations in the transcript levels of several key GAS virulence factor encoding genes during infection. Importantly, the transcript level alterations in the DeltaccpA and DeltacovRDeltaccpA isogenic mutant strains observed during infection were distinct from those occurring during growth in laboratory medium. These data provide new knowledge regarding the molecular mechanisms by which pathogenic bacteria respond to environmental signals to regulate virulence factor production and basic metabolic processes during infection. PMID:20333240

  18. Transcriptional control in the segmentation gene network of Drosophila.

    PubMed

    Schroeder, Mark D; Pearce, Michael; Fak, John; Fan, HongQing; Unnerstall, Ulrich; Emberly, Eldon; Rajewsky, Nikolaus; Siggia, Eric D; Gaul, Ulrike

    2004-09-01

    The segmentation gene network of Drosophila consists of maternal and zygotic factors that generate, by transcriptional (cross-) regulation, expression patterns of increasing complexity along the anterior-posterior axis of the embryo. Using known binding site information for maternal and zygotic gap transcription factors, the computer algorithm Ahab recovers known segmentation control elements (modules) with excellent success and predicts many novel modules within the network and genome-wide. We show that novel module predictions are highly enriched in the network and typically clustered proximal to the promoter, not only upstream, but also in intronic space and downstream. When placed upstream of a reporter gene, they consistently drive patterned blastoderm expression, in most cases faithfully producing one or more pattern elements of the endogenous gene. Moreover, we demonstrate for the entire set of known and newly validated modules that Ahab's prediction of binding sites correlates well with the expression patterns produced by the modules, revealing basic rules governing their composition. Specifically, we show that maternal factors consistently act as activators and that gap factors act as repressors, except for the bimodal factor Hunchback. Our data suggest a simple context-dependent rule for its switch from repressive to activating function. Overall, the composition of modules appears well fitted to the spatiotemporal distribution of their positive and negative input factors. Finally, by comparing Ahab predictions with different categories of transcription factor input, we confirm the global regulatory structure of the segmentation gene network, but find odd skipped behaving like a primary pair-rule gene. The study expands our knowledge of the segmentation gene network by increasing the number of experimentally tested modules by 50%. For the first time, the entire set of validated modules is analyzed for binding site composition under a uniform set of

  19. Transcriptional Control in the Segmentation Gene Network of Drosophila

    PubMed Central

    Fan, HongQing; Unnerstall, Ulrich; Emberly, Eldon; Rajewsky, Nikolaus; Siggia, Eric D

    2004-01-01

    The segmentation gene network of Drosophila consists of maternal and zygotic factors that generate, by transcriptional (cross-) regulation, expression patterns of increasing complexity along the anterior-posterior axis of the embryo. Using known binding site information for maternal and zygotic gap transcription factors, the computer algorithm Ahab recovers known segmentation control elements (modules) with excellent success and predicts many novel modules within the network and genome-wide. We show that novel module predictions are highly enriched in the network and typically clustered proximal to the promoter, not only upstream, but also in intronic space and downstream. When placed upstream of a reporter gene, they consistently drive patterned blastoderm expression, in most cases faithfully producing one or more pattern elements of the endogenous gene. Moreover, we demonstrate for the entire set of known and newly validated modules that Ahab's prediction of binding sites correlates well with the expression patterns produced by the modules, revealing basic rules governing their composition. Specifically, we show that maternal factors consistently act as activators and that gap factors act as repressors, except for the bimodal factor Hunchback. Our data suggest a simple context-dependent rule for its switch from repressive to activating function. Overall, the composition of modules appears well fitted to the spatiotemporal distribution of their positive and negative input factors. Finally, by comparing Ahab predictions with different categories of transcription factor input, we confirm the global regulatory structure of the segmentation gene network, but find odd skipped behaving like a primary pair-rule gene. The study expands our knowledge of the segmentation gene network by increasing the number of experimentally tested modules by 50%. For the first time, the entire set of validated modules is analyzed for binding site composition under a uniform set of

  20. From immunogenetics to immunomics: functional prospecting of genes and transcripts.

    PubMed

    Schönbach, Christian

    2003-01-01

    Human and mouse genome and transcriptome projects have expanded the field of 'immunogenetics' beyond the traditional study of the genetics and evolution of MHC, TCR and Ig loci into the new interdisciplinary area of 'immunomics'. Immunomics is the study of the molecular functions associated with all immune-related coding and non-coding mRNA transcripts. To unravel the function, regulation and diversity of the immunome requires that we identify and correctly categorize all immune-related transcripts. The importance of intercalated genes, antisense transcripts and non-coding RNAs and their potential role in regulation of immune development and function are only just starting to be appreciated. To better understand immune function and regulation, transcriptome projects (e.g. Functional Annotation of the Mouse, FANTOM), that focus on sequencing full-length transcripts from multiple tissue sources, ideally should include specific immune cells (e.g. T cell, B cells, macrophages, dendritic cells) at various states of development, in activated and unactivated states and in different disease contexts. Progress in deciphering immune regulatory networks will require the cooperative efforts of immunologists, immunogeneticists, molecular biologists and bioinformaticians. Although primary sequence analysis remains useful for annotation of new transcripts it is less useful for identifying novel functions of known transcripts in a new context (protein interaction network or pathway). The most efficient approach to mine useful information from the vast a priori knowledge contained in biological databases and the scientific literature, is to use a combination of computational and expert-driven knowledge discovery strategies. This paper will illustrate the challenges posed in attempts to functionally infer transcriptional regulation and interaction of immune-related genes from text and sequence-based data sources. PMID:14712938

  1. Mechanisms of post-transcriptional gene regulation in bacterial biofilms

    PubMed Central

    Martínez, Luary C.; Vadyvaloo, Viveka

    2014-01-01

    Biofilms are characterized by a dense multicellular community of microorganisms that can be formed by the attachment of bacteria to an inert surface and to each other. The development of biofilm involves the initial attachment of planktonic bacteria to a surface, followed by replication, cell-to-cell adhesion to form microcolonies, maturation, and detachment. Mature biofilms are embedded in a self-produced extracellular polymeric matrix composed primarily of bacterial-derived exopolysaccharides, specialized proteins, adhesins, and occasionally DNA. Because the synthesis and assembly of biofilm matrix components is an exceptionally complex process, the transition between its different phases requires the coordinate expression and simultaneous regulation of many genes by complex genetic networks involving all levels of gene regulation. The finely controlled intracellular level of the chemical second messenger molecule, cyclic-di-GMP is central to the post-transcriptional mechanisms governing the switch between the motile planktonic lifestyle and the sessile biofilm forming state in many bacteria. Several other post-transcriptional regulatory mechanisms are known to dictate biofilm development and assembly and these include RNA-binding proteins, small non-coding RNAs, toxin-antitoxin systems, riboswitches, and RNases. Post-transcriptional regulation is therefore a powerful molecular mechanism employed by bacteria to rapidly adjust to the changing environment and to fine tune gene expression to the developmental needs of the cell. In this review, we discuss post-transcriptional mechanisms that influence the biofilm developmental cycle in a variety of pathogenic bacteria. PMID:24724055

  2. Sperm is epigenetically programmed to regulate gene transcription in embryos.

    PubMed

    Teperek, Marta; Simeone, Angela; Gaggioli, Vincent; Miyamoto, Kei; Allen, George E; Erkek, Serap; Kwon, Taejoon; Marcotte, Edward M; Zegerman, Philip; Bradshaw, Charles R; Peters, Antoine H F M; Gurdon, John B; Jullien, Jerome

    2016-08-01

    For a long time, it has been assumed that the only role of sperm at fertilization is to introduce the male genome into the egg. Recently, ideas have emerged that the epigenetic state of the sperm nucleus could influence transcription in the embryo. However, conflicting reports have challenged the existence of epigenetic marks on sperm genes, and there are no functional tests supporting the role of sperm epigenetic marking on embryonic gene expression. Here, we show that sperm is epigenetically programmed to regulate embryonic gene expression. By comparing the development of sperm- and spermatid-derived frog embryos, we show that the programming of sperm for successful development relates to its ability to regulate transcription of a set of developmentally important genes. During spermatid maturation into sperm, these genes lose H3K4me2/3 and retain H3K27me3 marks. Experimental removal of these epigenetic marks at fertilization de-regulates gene expression in the resulting embryos in a paternal chromatin-dependent manner. This demonstrates that epigenetic instructions delivered by the sperm at fertilization are required for correct regulation of gene expression in the future embryos. The epigenetic mechanisms of developmental programming revealed here are likely to relate to the mechanisms involved in transgenerational transmission of acquired traits. Understanding how parental experience can influence development of the progeny has broad potential for improving human health. PMID:27034506

  3. Sperm is epigenetically programmed to regulate gene transcription in embryos

    PubMed Central

    Teperek, Marta; Simeone, Angela; Gaggioli, Vincent; Miyamoto, Kei; Allen, George E.; Erkek, Serap; Kwon, Taejoon; Marcotte, Edward M.; Zegerman, Philip; Bradshaw, Charles R.; Peters, Antoine H.F.M.; Gurdon, John B.; Jullien, Jerome

    2016-01-01

    For a long time, it has been assumed that the only role of sperm at fertilization is to introduce the male genome into the egg. Recently, ideas have emerged that the epigenetic state of the sperm nucleus could influence transcription in the embryo. However, conflicting reports have challenged the existence of epigenetic marks on sperm genes, and there are no functional tests supporting the role of sperm epigenetic marking on embryonic gene expression. Here, we show that sperm is epigenetically programmed to regulate embryonic gene expression. By comparing the development of sperm- and spermatid-derived frog embryos, we show that the programming of sperm for successful development relates to its ability to regulate transcription of a set of developmentally important genes. During spermatid maturation into sperm, these genes lose H3K4me2/3 and retain H3K27me3 marks. Experimental removal of these epigenetic marks at fertilization de-regulates gene expression in the resulting embryos in a paternal chromatin-dependent manner. This demonstrates that epigenetic instructions delivered by the sperm at fertilization are required for correct regulation of gene expression in the future embryos. The epigenetic mechanisms of developmental programming revealed here are likely to relate to the mechanisms involved in transgenerational transmission of acquired traits. Understanding how parental experience can influence development of the progeny has broad potential for improving human health. PMID:27034506

  4. Modular construction of mammalian gene circuits using TALE transcriptional repressors.

    PubMed

    Li, Yinqing; Jiang, Yun; Chen, He; Liao, Weixi; Li, Zhihua; Weiss, Ron; Xie, Zhen

    2015-03-01

    An important goal of synthetic biology is the rational design and predictable implementation of synthetic gene circuits using standardized and interchangeable parts. However, engineering of complex circuits in mammalian cells is currently limited by the availability of well-characterized and orthogonal transcriptional repressors. Here, we introduce a library of 26 reversible transcription activator-like effector repressors (TALERs) that bind newly designed hybrid promoters and exert transcriptional repression through steric hindrance of key transcriptional initiation elements. We demonstrate that using the input-output transfer curves of our TALERs enables accurate prediction of the behavior of modularly assembled TALER cascade and switch circuits. We also show that TALER switches using feedback regulation exhibit improved accuracy for microRNA-based HeLa cancer cell classification versus HEK293 cells. Our TALER library is a valuable toolkit for modular engineering of synthetic circuits, enabling programmable manipulation of mammalian cells and helping elucidate design principles of coupled transcriptional and microRNA-mediated post-transcriptional regulation. PMID:25643171

  5. Transcriptional Regulation of the p16 Tumor Suppressor Gene.

    PubMed

    Kotake, Yojiro; Naemura, Madoka; Murasaki, Chihiro; Inoue, Yasutoshi; Okamoto, Haruna

    2015-08-01

    The p16 tumor suppressor gene encodes a specific inhibitor of cyclin-dependent kinase (CDK) 4 and 6 and is found altered in a wide range of human cancers. p16 plays a pivotal role in tumor suppressor networks through inducing cellular senescence that acts as a barrier to cellular transformation by oncogenic signals. p16 protein is relatively stable and its expression is primary regulated by transcriptional control. Polycomb group (PcG) proteins associate with the p16 locus in a long non-coding RNA, ANRIL-dependent manner, leading to repression of p16 transcription. YB1, a transcription factor, also represses the p16 transcription through direct association with its promoter region. Conversely, the transcription factors Ets1/2 and histone H3K4 methyltransferase MLL1 directly bind to the p16 locus and mediate p16 induction during replicative and premature senescence. In the present review, we discuss the molecular mechanisms by which these factors regulate p16 transcription. PMID:26168478

  6. ULTRAPETALA trxG genes interact with KANADI transcription factor genes to regulate Aradopsis Gynoecium patterning

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Organ formation relies upon precise patterns of gene expression that are under tight spatial and temporal regulation. Transcription patterns are specified by several cellular processes during development, including chromatin remodeling, but little is known about how chromatin remodeling factors cont...

  7. Demonstration of transcriptional regulation of specific genes by phytochrome action

    PubMed Central

    Silverthorne, Jane; Tobin, Elaine M.

    1984-01-01

    We have developed an in vitro transcription system that uses nuclei isolated from Lemna gibba G-3. The in vitro transcripts include sequences homologous to hybridization probes for the small subunit of ribulose-1,5-bisphosphate carboxylase [3-phospho-D-glycerate carboxy-lyase (dimerizing), EC 4.1.1.39], the light-harvesting chlorophyll a/b-protein, and rRNA. Light-harvesting chlorophyll a/b-protein sequences are transcribed to a greater extent in nuclei isolated from plants grown in darkness with 2 min of red light every 8 hr than in nuclei isolated from dark-treated plants. Furthermore, the amount of these transcripts measured in plants given a single minute of red light after dark treatment is increased over the amount measured in dark-treated plants. The effect of red light is at least partially reversible by 10 min of far-red light given immediately after the red light pulse. Transcription of both rRNA and small subunit sequences is also stimulated by a single minute of red light as compared to dark-treated tissue. However, the relative magnitudes of the increases compared to the dark levels are smaller than the increase seen for the chlorophyll a/b-protein, possibly because of the higher level of transcription of these sequences in the dark. The effect of red light on the transcription of small subunit and rRNA sequences is also reversible by immediate treatment with 10 min of far-red light. Pulse chase studies of dark-treated nuclei for up to 110 min do not show substantial turnover of in vitro labeled small subunit and chlorophyll a/b-protein transcripts. We therefore conclude that phytochrome action has induced specific changes in transcription of these genes. Images PMID:16593420

  8. Transcriptional regulatory elements downstream of the JunB gene.

    PubMed Central

    Perez-Albuerne, E D; Schatteman, G; Sanders, L K; Nathans, D

    1993-01-01

    JunB is an immediate early transcription factor that is induced by a variety of extracellular signaling agents, including growth factors, phorbol esters, and agents that elevate cyclic AMP. The mechanism of activation of the gene encoding JunB by these agents is not well understood. By using the JunB gene together with flanking DNA in transfection experiments, we show that a serum response element (SRE) and/or a cAMP response element (CRE) downstream of the gene mediate the response of the gene in mouse NIH 3T3 cells to serum, platelet-derived growth factor, basic fibroblast growth factor, phorbol ester, and forskolin. In addition, a segment of DNA just upstream of the TATA box is required for optimal activation of the gene. Images Fig. 1 Fig. 5 PMID:8265655

  9. A hormone-encoding gene identifies a pathway for cardiac but not skeletal muscle gene transcription.

    PubMed Central

    Grépin, C; Dagnino, L; Robitaille, L; Haberstroh, L; Antakly, T; Nemer, M

    1994-01-01

    In contrast to skeletal muscle, the mechanisms responsible for activation and maintenance of tissue-specific transcription in cardiac muscle remain poorly understood. A family of hormone-encoding genes is expressed in a highly specific manner in cardiac but not skeletal myocytes. This includes the A- and B-type natriuretic peptide (ANP and BNP) genes, which encode peptide hormones with crucial roles in the regulation of blood volume and pressure. Since these genes are markers of cardiac cells, we have used them to probe the mechanisms for cardiac muscle-specific transcription. Cloning and functional analysis of the rat BNP upstream sequences revealed unexpected structural resemblance to erythroid but not to muscle-specific promoters and enhancers, including a requirement for regulatory elements containing GATA motifs. A cDNA clone corresponding to a member of the GATA family of transcription factors was isolated from a cardiomyocyte cDNA library. Transcription of this GATA gene is restricted mostly to the heart and is undetectable in skeletal muscle. Within the heart, GATA transcripts are localized in ANP- and BNP-expressing myocytes, and forced expression of the GATA protein in heterologous cells markedly activates transcription from the natural cardiac muscle-specific ANP and BNP promoters. This GATA-dependent pathway defines the first mechanism for cardiac muscle-specific transcription. Moreover, the present findings reveal striking similarities between the mechanisms controlling gene expression in hematopoietic and cardiac cells and may have important implications for studies of cardiogenesis. Images PMID:8164667

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

    PubMed

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

    2015-07-01

    Predictive modelling of gene expression provides a powerful framework for exploring the regulatory logic underpinning transcriptional regulation. Recent studies have demonstrated the utility of such models in identifying dysregulation of gene and miRNA expression associated with abnormal patterns of transcription factor (TF) binding or nucleosomal histone modifications (HMs). Despite the growing popularity of such approaches, a comparative review of the various modelling algorithms and feature extraction methods is lacking. We define and compare three methods of quantifying pairwise gene-TF/HM interactions and discuss their suitability for integrating the heterogeneous chromatin immunoprecipitation (ChIP)-seq binding patterns exhibited by TFs and HMs. We then construct log-linear and ϵ-support vector regression models from various mouse embryonic stem cell (mESC) and human lymphoblastoid (GM12878) data sets, considering both ChIP-seq- and position weight matrix- (PWM)-derived in silico TF-binding. The two algorithms are evaluated both in terms of their modelling prediction accuracy and ability to identify the established regulatory roles of individual TFs and HMs. Our results demonstrate that TF-binding and HMs are highly predictive of gene expression as measured by mRNA transcript abundance, irrespective of algorithm or cell type selection and considering both ChIP-seq and PWM-derived TF-binding. As we encourage other researchers to explore and develop these results, our framework is implemented using open-source software and made available as a preconfigured bootable virtual environment. PMID:25231769

  11. Functional divergence and convergence between the transcript network and gene network in lung adenocarcinoma

    PubMed Central

    Hsu, Min-Kung; Pan, Chia-Lin; Chen, Feng-Chi

    2016-01-01

    Introduction Alternative RNA splicing is a critical regulatory mechanism during tumorigenesis. However, previous oncological studies mainly focused on the splicing of individual genes. Whether and how transcript isoforms are coordinated to affect cellular functions remain underexplored. Also of great interest is how the splicing regulome cooperates with the transcription regulome to facilitate tumorigenesis. The answers to these questions are of fundamental importance to cancer biology. Results Here, we report a comparative study between the transcript-based network (TN) and the gene-based network (GN) derived from the transcriptomes of paired tumor–normal tissues from 77 lung adenocarcinoma patients. We demonstrate that the two networks differ significantly from each other in terms of patient clustering and the number and functions of network modules. Interestingly, the majority (89.5%) of multi-transcript genes have their transcript isoforms distributed in at least two TN modules, suggesting regulatory and functional divergences between transcript isoforms. Furthermore, TN and GN modules share onlŷ50%–60% of their biological functions. TN thus appears to constitute a regulatory layer separate from GN. Nevertheless, our results indicate that functional convergence and divergence both occur between TN and GN, implying complex interactions between the two regulatory layers. Finally, we report that the expression profiles of module members in both TN and GN shift dramatically yet concordantly during tumorigenesis. The mechanisms underlying this coordinated shifting remain unclear yet are worth further explorations. Conclusion We show that in lung adenocarcinoma, transcript isoforms per se are coordinately regulated to conduct biological functions not conveyed by the network of genes. However, the two networks may interact closely with each other by sharing the same or related biological functions. Unraveling the effects and mechanisms of such interactions will

  12. A Novel POK Family Transcription Factor, ZBTB5, Represses Transcription of p21CIP1 Gene*

    PubMed Central

    Koh, Dong-In; Choi, Won-Il; Jeon, Bu-Nam; Lee, Choong-Eun; Yun, Chae-Ok; Hur, Man-Wook

    2009-01-01

    Transcriptional repression through chromatin remodeling and histone deacetylation has been postulated as a driving force for tumorigenesis. We isolated and characterized a novel POZ domain Krüppel-like zinc finger transcription repressor, ZBTB5 (zinc finger and BTB domain-containing 5). Serial analysis of gene expression (SAGE) analysis showed that ZBTB5 expression is higher in retinoblastoma and muscle cancer tissues. Immunocytochemistry showed that ZBTB5 was localized to the nucleus, particularly nuclear speckles. ZBTB5 directly repressed transcription of cell cycle arrest gene p21 by binding to the proximal GC-box 5/6 elements and the two distal p53-responsive elements (bp −2323 ∼ −2299; bp −1416 ∼ −1392). Chromatin immunoprecipitation assays showed that ZBTB5 and p53 competed with each other in occupying the p53 binding elements. ZBTB5 interacted with co-repressor-histone deacetylase complexes such as BCoR (BCL-6-interacting corepressor), NCoR (nuclear receptor corepressor), and SMRT (silencing mediator for retinoid and thyroid receptors) via its POZ domain. These interactions resulted in deacetylation of histones Ac-H3 and Ac-H4 at the proximal promoter, which is important in the transcriptional repression of p21. MTT (3-(4,5-di meth yl thi azol-2-yl)-2,5-diphenyltetrazolium bromide) assays and fluorescent-activated cell sorter analysis revealed that ZBTB5 stimulated both cell proliferation and cell cycle progression, significantly increasing the number of cells in S-phase. Overall, our data suggest that ZBTB5 is a potent transcription repressor of cell cycle arrest gene p21 and a potential proto-oncogene stimulating cell proliferation. PMID:19491398

  13. NFAT5 regulates transcription of the mouse telomerase reverse transcriptase gene

    SciTech Connect

    Fujiki, Tsukasa; Udono, Miyako; Kotake, Yojiro; Yamashita, Makiko; Shirahata, Sanetaka; Katakura, Yoshinori

    2010-12-10

    We aimed to clarify the transcription-regulation mechanisms of the mouse telomerase reverse transcriptase gene (mTERT). First, we searched for the promoter region required for transcriptional activation of mTERT and identified an enhancer cis-element (named mTERT-EE) located between - 200 and - 179 bp of the mouse TERT gene (mTERT). EMSA results suggested that nuclear factor of activated T cells (NFAT) member proteins bind to mTERT-EE. We then identified NFAT5 as the factor binding to mTERT-EE and found that it activates the transcription of the mTERT core promoter. The results that siRNA directed against NFAT5 significantly reduced mTERT expression and mTERT core promoter activity and that the expressions of NFAT5 and mTERT were well correlated in various mouse tissues except liver suggest that NFAT5 dominantly and directly regulates mTERT expression. To clarify their functionality further, we investigated the effect of hypertonic stress, a known stimulus affecting the expression and transcriptional activity of NFAT5, on mTERT expression. The result indicated that hypertonic stress activates mTERT transcription via the activation and recruitment of NFAT5 to the mTERT promoter. These results provide useful information about the transcription-regulation mechanisms of mTERT.

  14. Post-Transcriptional Control of Chloroplast Gene Expression

    PubMed Central

    del Campo, Eva M.

    2009-01-01

    Chloroplasts contain their own genome, organized as operons, which are generally transcribed as polycistronic transcriptional units. These primary transcripts are processed into smaller RNAs, which are further modified to produce functional RNAs. The RNA processing mechanisms remain largely unknown and represent an important step in the control of chloroplast gene expression. Such mechanisms include RNA cleavage of pre-existing RNAs, RNA stabilization, intron splicing, and RNA editing. Recently, several nuclear-encoded proteins that participate in diverse plastid RNA processing events have been characterised. Many of them seem to belong to the pentatricopeptide repeat (PPR) protein family that is implicated in many crucial functions including organelle biogenesis and plant development. This review will provide an overview of current knowledge of the post-transcriptional processing in chloroplasts. PMID:19838333

  15. Effect of replication on epigenetic memory and consequences on gene transcription.

    PubMed

    Zerihun, Mehari B; Vaillant, Cédric; Jost, Daniel

    2015-04-01

    Gene activity in eukaryotes is in part regulated at the level of chromatin through the assembly of local chromatin states that are more or less permissive to transcription. How do these chromatin states achieve their functions and whether or not they contribute to the epigenetic inheritance of the transcriptional program remain to be elucidated. In cycling cells, stability is indeed strongly challenged by the periodic occurrence of replication and cell division. To address this question, we perform simulations of the stochastic dynamics of chromatin states when driven out-of-equilibrium by periodic perturbations. We show how epigenetic memory is significantly affected by the cell cycle length. In addition, we develop a simple model to connect the epigenetic state to the transcriptional state and gene activity. In particular, it suggests that replication may induce transcriptional bursting at repressive loci. Finally, we discuss how our findings-effect of replication and link to gene transcription-have original and deep implications to various biological contexts of epigenetic memory. PMID:25884278

  16. Widespread transcriptional gene inactivation initiated by a repair intermediate of 8-oxoguanine

    PubMed Central

    Allgayer, Julia; Kitsera, Nataliya; Bartelt, Solveig; Epe, Bernd; Khobta, Andriy

    2016-01-01

    DNA damage can significantly modulate expression of the affected genes either by direct structural interference with transcription components or as a collateral outcome of cellular repair attempts. Thus, DNA glycosylases of the base excision repair (BER) pathway have been implicated in negative transcriptional response to several spontaneously generated DNA base modifications, including a common oxidative DNA base modification 8-oxoguanine (8-oxoG). Here, we report that single 8-oxoG situated in the non-transcribed DNA strand of a reporter gene has a pronounced negative effect on transcription, driven by promoters of various strength and with different structural properties, including viral, human, and artificial promoters. We further show that the magnitude of the negative effect on the gene expression correlates with excision of the modified base by OGG1 in all promoter constructs tested. Moreover, by using expression vectors with nuclease resistant backbone modifications, we demonstrate that OGG1 does not catalyse DNA strand cleavage in vivo. Rather, cleavage of the phosphate bond 5′ to 8-oxodG (catalysed by APE1) is essential and universally required for the onset of transcriptional silencing, regardless of the promoter structure. Hence, induction of transcriptional silencing emerges as a ubiquitous mode of biological response to 8-oxoG in DNA. PMID:27220469

  17. Interaction of nucleolin with ribosomal RNA genes and its role in RNA polymerase I transcription

    PubMed Central

    Cong, Rong; Das, Sadhan; Ugrinova, Iva; Kumar, Sanjeev; Mongelard, Fabien; Wong, Jiemin; Bouvet, Philippe

    2012-01-01

    Nucleolin is a multi-functional nucleolar protein that is required for ribosomal RNA gene (rRNA) transcription in vivo, but the mechanism by which nucleolin modulates RNA polymerase I (RNAPI) transcription is not well understood. Nucleolin depletion results in an increase in the heterochromatin mark H3K9me2 and a decrease in H4K12Ac and H3K4me3 euchromatin histone marks in rRNA genes. ChIP-seq experiments identified an enrichment of nucleolin in the ribosomal DNA (rDNA) coding and promoter region. Nucleolin is preferentially associated with unmethylated rRNA genes and its depletion leads to the accumulation of RNAPI at the beginning of the transcription unit and a decrease in UBF along the coding and promoter regions. Nucleolin is able to affect the binding of transcription termination factor-1 on the promoter-proximal terminator T0, thus inhibiting the recruitment of TIP5 and HDAC1 and the establishment of a repressive heterochromatin state. These results reveal the importance of nucleolin for the maintenance of the euchromatin state and transcription elongation of rDNA. PMID:22859736

  18. Noninvasive Tracking of Gene Transcript and Neuroprotection after Gene Therapy

    PubMed Central

    Ren, Jiaqian; Chen, Y. Iris; Liu, Christina H.; Chen, Po-Chih; Prentice, Howard; Wu, Jang-Yen; Liu, Philip K.

    2015-01-01

    Gene therapy holds exceptional potential for translational medicine by improving the products of defective genes in diseases and/or providing necessary biologics from endogenous sources during recovery processes. However, validating methods for the delivery, distribution and expression of the exogenous genes from such therapy can generally not be applicable to monitor effects over the long term because they are invasive. We report here that human granulocyte colony-stimulating factor (hG-CSF) cDNA encoded in scAAV-type 2 adeno-associated virus, as delivered through eye drops at multiple time points after cerebral ischemia using bilateral carotid occlusion for 60 min (BCAO-60) led to significant reduction in mortality rates, cerebral atrophy, and neurological deficits in C57black6 mice. Most importantly, we validated hG-CSF cDNA expression using translatable magnetic resonance imaging (MRI) in living brains. This noninvasive approach for monitoring exogenous gene expression in the brains has potential for great impact in the area of experimental gene therapy in animal models of heart attack, stroke, Alzheimer’s dementia, Parkinson’s disorder and amyotrophic lateral sclerosis, and the translation of such techniques to emergency medicine. PMID:26207935

  19. Noninvasive tracking of gene transcript and neuroprotection after gene therapy.

    PubMed

    Ren, J; Chen, Y I; Liu, C H; Chen, P-C; Prentice, H; Wu, J-Y; Liu, P K

    2016-01-01

    Gene therapy holds exceptional potential for translational medicine by improving the products of defective genes in diseases and/or providing necessary biologics from endogenous sources during recovery processes. However, validating methods for the delivery, distribution and expression of the exogenous genes from such therapy can generally not be applicable to monitor effects over the long term because they are invasive. We report here that human granulocyte colony-stimulating factor (hG-CSF) complimentary DNA (cDNA) encoded in self-complementary adeno-associated virus-type 2 adeno-associated virus, as delivered through eye drops at multiple time points after cerebral ischemia using bilateral carotid occlusion for 60 min (BCAO-60) led to significant reduction in mortality rates, cerebral atrophy and neurological deficits in C57black6 mice. Most importantly, we validated hG-CSF cDNA expression using translatable magnetic resonance imaging (MRI) in living brains. This noninvasive approach for monitoring exogenous gene expression in the brains has potential for great impact in the area of experimental gene therapy in animal models of heart attack, stroke, Alzheimer's dementia, Parkinson's disorder and amyotrophic lateral sclerosis, and the translation of such techniques to emergency medicine. PMID:26207935

  20. Multiple controls affect arsenite oxidase gene expression in Herminiimonas arsenicoxydans

    PubMed Central

    2010-01-01

    Background Both the speciation and toxicity of arsenic are affected by bacterial transformations, i.e. oxidation, reduction or methylation. These transformations have a major impact on environmental contamination and more particularly on arsenic contamination of drinking water. Herminiimonas arsenicoxydans has been isolated from an arsenic- contaminated environment and has developed various mechanisms for coping with arsenic, including the oxidation of As(III) to As(V) as a detoxification mechanism. Results In the present study, a differential transcriptome analysis was used to identify genes, including arsenite oxidase encoding genes, involved in the response of H. arsenicoxydans to As(III). To get insight into the molecular mechanisms of this enzyme activity, a Tn5 transposon mutagenesis was performed. Transposon insertions resulting in a lack of arsenite oxidase activity disrupted aoxR and aoxS genes, showing that the aox operon transcription is regulated by the AoxRS two-component system. Remarkably, transposon insertions were also identified in rpoN coding for the alternative N sigma factor (σ54) of RNA polymerase and in dnaJ coding for the Hsp70 co-chaperone. Western blotting with anti-AoxB antibodies and quantitative RT-PCR experiments allowed us to demonstrate that the rpoN and dnaJ gene products are involved in the control of arsenite oxidase gene expression. Finally, the transcriptional start site of the aoxAB operon was determined using rapid amplification of cDNA ends (RACE) and a putative -12/-24 σ54-dependent promoter motif was identified upstream of aoxAB coding sequences. Conclusion These results reveal the existence of novel molecular regulatory processes governing arsenite oxidase expression in H. arsenicoxydans. These data are summarized in a model that functionally integrates arsenite oxidation in the adaptive response to As(III) in this microorganism. PMID:20167112

  1. Transcriptional regulation of the phosphoenolpyruvate carboxykinase gene by cooperation between hepatic nuclear factors.

    PubMed Central

    Yanuka-Kashles, O; Cohen, H; Trus, M; Aran, A; Benvenisty, N; Reshef, L

    1994-01-01

    To study the transcriptional regulation of the liver gluconeogenic phenotype, the underdifferentiated mouse Hepa-1c1c7 (Hepa) hepatoma cell line was used. These cells mimicked the fetal liver by appreciably expressing the alpha-fetoprotein and albumin genes but not the phosphoenolpyruvate carboxykinase (PEPCK) gene. Unlike the fetal liver, however, Hepa cells failed to express the early-expressed factors hepatocyte nuclear factor 1 alpha (HNF-1 alpha) and HNF-4 and the late-expressed factor C/EBP alpha, thereby providing a suitable system for examining possible cooperation between these factors in the transcriptional regulation of the PEPCK gene. Transient transfection assays of a chimeric PEPCK-chloramphenicol acetyltransferase construct showed a residual PEPCK promoter activity in the Hepa cell line, which was slightly stimulated by cotransfection with a single transcription factor from either the C/EBP family or HNF-1 alpha but not at all affected by cotransfection of HNF-4. In contrast, cotransfection of the PEPCK construct with members from the C/EBP family plus HNF-1 alpha resulted in a synergistic stimulation of the PEPCK promoter activity. This synergistic effect depended on the presence in the PEPCK promoter region of the HNF-1 recognition sequence and on the presence of two C/EBP recognition sequences. The results demonstrate a requirement for coexistence and cooperation between early and late liver-enriched transcription factors in the transcriptional regulation of the PEPCK gene. In addition, the results suggest redundancy between members of the C/EBP family of transcription factors in the regulation of PEPCK gene expression. Images PMID:7935427

  2. Glucocorticoid modulation of casein gene transcription in mouse mammary gland.

    PubMed Central

    Ganguly, R; Mehta, N M; Ganguly, N; Banerjee, M R

    1979-01-01

    The influence of cortisol and prolactin on casein gene expression in the mammary gland of lactating BALB/c mice was measured by using a specific cDNA probe to 15S casein mRNA (cDNAcsn). Casein mRNA (mRNAcsn) level in the mammary gland was decreased by 85% 5 days after adrenal ablation, but then was increased 4.4-fold 12 hr after a single injection of hydrocortisone-21-acetate. An 80% decrease in serum prolactin level, induced by the prolactin inhibitor 2-bromo-alpha-ergocryptin (CB-154), did not alter the level of mRNAcsn in the gland. Specific transcription of the casein gene in nuclei isolated from lactating mammary glands was measured by cDNAcsn hybridization to the in vitro synthesized Hg-CTP-containing RNA (Hg-RNA), which was purified by SH-agarose chromatography. The level of the mRNAcsn in Hg-RNA synthesized in the isolated nuclei was 0.09% and this was decreased 85% by alpha-amanitin, indicating that the mRNAcsn sequences in the Hg-RNA were the products of RNA polymerase II-directed DNA-dependent RNA synthesis. Transcription of the mRNAcsn in isolated nuclei was decreased by 70% 5 days after adrenalectomy and a single injection of the glucocorticoid then increased the transcription level 2-fold at 6 hr. Essentially no alteration of the level of transcription was detectable in mammary nuclei isolated from lactating mice with 80% decreased serum prolactin level, induced by CB-154 treatment. The results thus demonstrate a glucocorticoid involvement on the modulation of casein gene expression at the transcriptional level of control. PMID:293734

  3. Transcriptional Activation of the Cyclin A Gene by the Architectural Transcription Factor HMGA2

    PubMed Central

    Tessari, Michela A.; Gostissa, Monica; Altamura, Sandro; Sgarra, Riccardo; Rustighi, Alessandra; Salvagno, Clio; Caretti, Giuseppina; Imbriano, Carol; Mantovani, Roberto; Del Sal, Giannino; Giancotti, Vincenzo; Manfioletti, Guidalberto

    2003-01-01

    The HMGA2 protein belongs to the HMGA family of architectural transcription factors, which play an important role in chromatin organization. HMGA proteins are overexpressed in several experimental and human tumors and have been implicated in the process of neoplastic transformation. Hmga2 knockout results in the pygmy phenotype in mice and in a decreased growth rate of embryonic fibroblasts, thus indicating a role for HMGA2 in cell proliferation. Here we show that HMGA2 associates with the E1A-regulated transcriptional repressor p120E4F, interfering with p120E4F binding to the cyclin A promoter. Ectopic expression of HMGA2 results in the activation of the cyclin A promoter and induction of the endogenous cyclin A gene. In addition, chromatin immunoprecipitation experiments show that HMGA2 associates with the cyclin A promoter only when the gene is transcriptionally activated. These data identify the cyclin A gene as a cellular target for HMGA2 and, for the first time, suggest a mechanism for HMGA2-dependent cell cycle regulation. PMID:14645522

  4. Stochastic model for gene transcription on Drosophila melanogaster embryos.

    PubMed

    Prata, Guilherme N; Hornos, José Eduardo M; Ramos, Alexandre F

    2016-02-01

    We examine immunostaining experimental data for the formation of stripe 2 of even-skipped (eve) transcripts on D. melanogaster embryos. An estimate of the factor converting immunofluorescence intensity units into molecular numbers is given. The analysis of the eve dynamics at the region of stripe 2 suggests that the promoter site of the gene has two distinct regimes: an earlier phase when it is predominantly activated until a critical time when it becomes mainly repressed. That suggests proposing a stochastic binary model for gene transcription on D. melanogaster embryos. Our model has two random variables: the transcripts number and the state of the source of mRNAs given as active or repressed. We are able to reproduce available experimental data for the average number of transcripts. An analysis of the random fluctuations on the number of eves and their consequences on the spatial precision of stripe 2 is presented. We show that the position of the anterior or posterior borders fluctuate around their average position by ∼1% of the embryo length, which is similar to what is found experimentally. The fitting of data by such a simple model suggests that it can be useful to understand the functions of randomness during developmental processes. PMID:26986358

  5. Stochastic model for gene transcription on Drosophila melanogaster embryos

    NASA Astrophysics Data System (ADS)

    Prata, Guilherme N.; Hornos, José Eduardo M.; Ramos, Alexandre F.

    2016-02-01

    We examine immunostaining experimental data for the formation of stripe 2 of even-skipped (eve) transcripts on D. melanogaster embryos. An estimate of the factor converting immunofluorescence intensity units into molecular numbers is given. The analysis of the eve dynamics at the region of stripe 2 suggests that the promoter site of the gene has two distinct regimes: an earlier phase when it is predominantly activated until a critical time when it becomes mainly repressed. That suggests proposing a stochastic binary model for gene transcription on D. melanogaster embryos. Our model has two random variables: the transcripts number and the state of the source of mRNAs given as active or repressed. We are able to reproduce available experimental data for the average number of transcripts. An analysis of the random fluctuations on the number of eves and their consequences on the spatial precision of stripe 2 is presented. We show that the position of the anterior or posterior borders fluctuate around their average position by ˜1 % of the embryo length, which is similar to what is found experimentally. The fitting of data by such a simple model suggests that it can be useful to understand the functions of randomness during developmental processes.

  6. A mutation that uncouples flagellum assembly from transcription alters the temporal pattern of flagellar gene expression in Caulobacter crescentus.

    PubMed Central

    Mangan, E K; Bartamian, M; Gober, J W

    1995-01-01

    The transcription of flagellar genes in Caulobacter crescentus is regulated by cell cycle events that culminate in the synthesis of a new flagellum once every cell division. Early flagellar gene products regulate the expression of late flagellar genes at two distinct stages of the flagellar trans-acting hierarchy. Here we investigate the coupling of early flagellar biogenesis with middle and late flagellar gene expression. We have isolated mutants (bfa) that do not require early class II flagellar gene products for the transcription of middle or late flagellar genes. bfa mutant strains are apparently defective in a negative regulatory pathway that couples early flagellar biogenesis to late flagellar gene expression. The bfa regulatory pathway functions solely at the level of transcription. Although flagellin promoters are transcribed in class II/bfa double mutants, there is no detectable flagellin protein on immunoblots prepared from mutant cell extracts. This finding suggests that early flagellar biogenesis is coupled to gene expression by two distinct mechanisms: one that negatively regulates transcription, mediated by bfa, and another that functions posttranscriptionally. To determine whether bfa affects the temporal pattern of late flagellar gene expression, cell cycle experiments were performed in bfa mutant strains. In a bfa mutant strain, flagellin expression fails to shut off at its normal time in the cell division cycle. This experimental result indicates that bfa may function as a regulator of flagellar gene transcription late in the cell cycle, after early flagellar structures have been assembled. PMID:7768816

  7. Rapid parallel evolutionary changes of gene transcription profiles in farmed Atlantic salmon.

    PubMed

    Roberge, Christian; Einum, Sigurd; Guderley, Helga; Bernatchez, Louis

    2006-01-01

    Farmed salmon strains have been selected to improve growth rates as well as other traits of commercial interest but the 2 million farmed salmon escaping annually may enhance the risk of extinction of wild populations through genetic and ecological interactions. Here, we compare the transcription profiles of 3557 genes in the progeny of farmed and wild Atlantic salmon from Norway and Canada grown in controlled conditions, and demonstrate that five to seven generations of artificial selection led to heritable changes in gene transcription profiles, the average magnitude of the differences being 25% and 18% for at least 1.4% and 1.7% of the expressed genes in juvenile salmon from Norway and Canada, respectively. Moreover, genes showing significant transcription profile differences in both farmed strains (16%) all exhibited parallel changes. These findings, along with the identification of several genes whose expression profiles were modified through artificial selection, provide new insights into the molecular basis of parallel evolution, and suggest how gene flow from farmed escapees may affect the genetic integrity of wild populations. PMID:16367826

  8. Transcriptional Regulation of Tlr11 Gene Expression in Epithelial Cells*

    PubMed Central

    Cai, Zhenyu; Shi, Zhongcheng; Sanchez, Amir; Zhang, Tingting; Liu, Mingyao; Yang, Jianghua; Wang, Fen; Zhang, Dekai

    2009-01-01

    As sensors of invading microorganisms, Toll-like receptors (TLRs) are expressed not only on macrophages and dendritic cells (DCs) but also on epithelial cells. In the TLR family, Tlr11 appears to have the unique feature in that it is expressed primarily on epithelial cells, although it is also expressed on DCs and macrophages. Here, we demonstrate that transcription of the Tlr11 gene is regulated through two cis-acting elements, one Ets-binding site and one interferon regulatory factor (IRF)-binding site. The Ets element interacts with the epithelium-specific transcription factors, ESE-1 and ESE-3, and the IRF motif interacts with IRF-8. Thus, Tlr11 expression on epithelial cells is regulated by the transcription factors that are presumably distinct from transcription factors that regulate the expression of TLRs in innate immune cells such as macrophages and DCs. Our results imply that the distinctive transcription regulatory machinery for TLRs on epithelium may represent a promising new avenue for the development of epithelia-specific therapeutic interventions. PMID:19801549

  9. Transcript degradation and noise of small RNA-controlled genes in a switch activated network in Escherichia coli.

    PubMed

    Arbel-Goren, Rinat; Tal, Asaf; Parasar, Bibudha; Dym, Alvah; Costantino, Nina; Muñoz-García, Javier; Court, Donald L; Stavans, Joel

    2016-08-19

    Post-transcriptional regulatory processes may change transcript levels and affect cell-to-cell variability or noise. We study small-RNA downregulation to elucidate its effects on noise in the iron homeostasis network of Escherichia coli In this network, the small-RNA RyhB undergoes stoichiometric degradation with the transcripts of target genes in response to iron stress. Using single-molecule fluorescence in situ hybridization, we measured transcript numbers of the RyhB-regulated genes sodB and fumA in individual cells as a function of iron deprivation. We observed a monotonic increase of noise with iron stress but no evidence of theoretically predicted, enhanced stoichiometric fluctuations in transcript numbers, nor of bistable behavior in transcript distributions. Direct detection of RyhB in individual cells shows that its noise is much smaller than that of these two targets, when RyhB production is significant. A generalized two-state model of bursty transcription that neglects RyhB fluctuations describes quantitatively the dependence of noise and transcript distributions on iron deprivation, enabling extraction of in vivo RyhB-mediated transcript degradation rates. The transcripts' threshold-linear behavior indicates that the effective in vivo interaction strength between RyhB and its two target transcripts is comparable. Strikingly, the bacterial cell response exhibits Fur-dependent, switch-like activation instead of a graded response to iron deprivation. PMID:27085802

  10. Manganese peroxidase gene transcription in Phanerochaete chrysosporium: Activation by manganese

    SciTech Connect

    Brown, J.A.; Alic, M. Gold, M.H. )

    1991-07-01

    The expression of manganese peroxidase in nitrogen-limited cultures of Phanerochaete chrysosporium is dependent on Mn, and initial work suggested that Mn regulates transcription of the mnp gene. In this study, using Northern (RNA) blot analysis of kinetic, dose-response, and inhibitor experiments, the authors demonstrate unequivocally that Mn regulates mnp gene transcription. The amount of mnp mRNA in cells of 4-day-old nitrogen-limited cultures is a direct function of the concentration of Mn in the culture medium up to a maximum of 180 {mu}M. Addition of Mn to nitrogen-limited Mn-deficient secondary metabolic (4-, 5-, and 6-day-old) cultures results in the appearance of mnp mRNA within 40 min. The appearance of this message is completely inhibited by the RNA synthesis inhibitor dactinomycin but not by the protein synthesis inhibitor cycloheximide. Furthermore, the amount of mnp mRNA produced is a direct function of the concentration of added Mn. In contrast, addition of Mn to low-nitrogen Mn-deficient 2- or 3-day-old cultures does not result in the appearance of mnp mRNA. Manganese peroxidase protein is detected by specific immunoprecipitation of the in vitro translation products of poly(A) RNA isolated from Mn-supplemented (but nor from Mn-deficient) cells. All of these results demonstrate that Mn, the substrate for the enzyme, regulates mnp gene transcription via a growth-stage-specific and concentration-dependent mechanism.

  11. Transcriptional regulation of cathelicidin genes in chicken bone marrow cells.

    PubMed

    Lee, Sang In; Jang, Hyun June; Jeon, Mi-hyang; Lee, Mi Ock; Kim, Jeom Sun; Jeon, Ik-Soo; Byun, Sung June

    2016-04-01

    Cathelicidins form a family of vertebrate-specific immune molecules with an evolutionarily conserved gene structure. We analyzed the expression patterns of cathelicidin genes (CAMP, CATH3, and CATHB1) in chicken bone marrow cells (BMCs) and chicken embryonic fibroblasts (CEFs). We found that CAMP and CATHB1 were significantly up-regulated in BMCs, whereas the expression of CATH3 did not differ significantly between BMCs and CEFs. To study the mechanism underlying the up-regulation of cathelicidin genes in BMCs, we predicted the transcription factors (TFs) that bind to the 5'-flanking regions of cathelicidin genes. CEBPA, EBF1, HES1, MSX1, and ZIC3 were up-regulated in BMCs compared to CEFs. Subsequently, when a siRNA-mediated knockdown assay was performed for MSX1, the expression of CAMP and CATHB1 was decreased in BMCs. We also showed that the transcriptional activity of the CAMP promoter was decreased by mutation of the MSX1-binding sites present within the 5'-flanking region of CAMP. These results increase our understanding of the regulatory mechanisms controlling cathelicidin genes in BMCs. PMID:26908883

  12. Housekeeping gene transcript abundance in bovine fertilized and cloned embryos.

    PubMed

    Ross, Pablo J; Wang, Kai; Kocabas, Arif; Cibelli, Jose B

    2010-12-01

    The objective of this study was to compare housekeeping gene expression levels, relative to total mRNA, across different stages of bovine preimplantation development in embryos generated by IVF and somatic cell nuclear transfer (SCNT). We first analyzed the levels of total RNA recovered from different stages of preimplantation development. A similar RNA level was observed from oocytes to 16-cell stage embryos with a significant increase at morula and blastocyst stages. Then we used an absolute mRNA determination method that accounts for the RNA level in the embryo by quantifying copies of transcripts normalized to loaded cDNA amount. The number of housekeeping genes mRNA copies per nanogram of cDNA was compared among samples obtained from different stages of preimplantation IVF-derived embryos. None of the genes analyzed (GAPDH, PPIA, ACTB, RPL15, GUSB, and Histone H2A.2) maintained constant levels throughout preimplantation development, indicating that they are not suitable for normalizing gene expression across developmental stages. We then compared expression of housekeeping genes between IVF and SCNT embryos at different embryonic stages. We found different levels of transcript abundance between IVF and SCNT embryos for GAPDH, RPL15, GUSB, and ACTB. On the other hand, Histone H2A.2 and PPIA were similar between IVF and SCNT embryos at each stage analyzed, although they varied across stages as previously mentioned. PMID:20973679

  13. Non-Equilibrium Thermodynamics of Gene Expression and Transcriptional Regulation

    NASA Astrophysics Data System (ADS)

    Lemus, Enrique Hernández

    2009-12-01

    In recent times whole-genome gene expression analysis has turned out to be a highly important tool to study the coordinated function of a very large number of genes within their corresponding cellular environment, especially in relation to phenotypic diversity and disease. A wide variety of methods of quantitative analysis has been developed to cope with high throughput data sets generated by gene expression profiling experiments. Due to the complexity associated with transcriptomics, especially in the case of gene regulation phenomena, most of these methods are of a probabilistic or statistical nature. Even if these methods have reached a central status in the development of an integrative, systematic understanding of the associated biological processes, they very rarely constitute a concrete guide to the actual physicochemical mechanisms behind biological function, and the role of these methods is more on a hypotheses generating line. An important improvement could lie in the development of a thermodynamic theory for gene expression and transcriptional regulation that will build the foundations for a proper integration of the vast amount of molecular biophysical data and could lead, in the future, to a systemic view of genetic transcription and regulation.

  14. New insights on the reorganization of gene transcription in Pseudomonas putida KT2440 at elevated pressure

    PubMed Central

    2013-01-01

    Background Elevated pressure, elevated oxygen tension (DOT) and elevated carbon dioxide tension (DCT) are readily encountered at the bottom of large industrial bioreactors and during bioprocesses where pressure is applied for enhancing the oxygen transfer. Yet information about their effect on bacteria and on the gene expression thereof is scarce. To shed light on the cellular functions affected by these specific environmental conditions, the transcriptome of Pseudomonas putida KT2440, a bacterium of great relevance for the production of medium-chain-length polyhydroxyalkanoates, was thoroughly investigated using DNA microarrays. Results Very well defined chemostat cultivations were carried out with P. putida to produce high quality RNA samples and ensure that differential gene expression was caused exclusively by changes of pressure, DOT and/or DCT. Cellular stress was detected at 7 bar and elevated DCT in the form of heat shock and oxidative stress-like responses, and indicators of cell envelope perturbations were identified as well. Globally, gene transcription was not considerably altered when DOT was increased from 40 ± 5 to 235 ± 20% at 7 bar and elevated DCT. Nevertheless, differential transcription was observed for a few genes linked to iron-sulfur cluster assembly, terminal oxidases, glutamate metabolism and arginine deiminase pathway, which shows their particular sensitivity to variations of DOT. Conclusions This study provides a comprehensive overview on the changes occurring in the transcriptome of P. putida upon mild variations of pressure, DOT and DCT. Interestingly, whereas the changes of gene transcription were widespread, the cell physiology was hardly affected, which illustrates how efficient reorganization of the gene transcription is for dealing with environmental changes that may otherwise be harmful. Several particularly sensitive cellular functions were identified, which will certainly contribute to the understanding of the

  15. Cloning and transcriptional analysis of Crepis alpina fatty acid desaturases affecting the biosynthesis of crepenynic acid.

    PubMed

    Nam, Jeong-Won; Kappock, T Joseph

    2007-01-01

    Crepis alpina acetylenase is a variant FAD2 desaturase that catalyses the insertion of a triple bond at the Delta12 position of linoleic acid, forming crepenynic acid in developing seeds. Seeds contain a high level of crepenynic acid but other tissues contain none. Using reverse transcriptase-coupled PCR (RT-PCR), acetylenase transcripts were identified in non-seed C. alpina tissues, which were highest in flower heads. To understand why functional expression of the acetylenase is limited to seeds, genes that affect acetylenase activity by providing substrate (FAD2) or electrons (cytochrome b5), or that compete for substrate (FAD3), were cloned. RT-PCR analysis indicated that the availability of a preferred cytochrome b5 isoform is not a limiting factor. Developing seeds co-express acetylenase and FAD2 isoform 2 (FAD2-2) at high levels. Flower heads co-express FAD2-3 and FAD3 at high levels, and FAD2-2 and acetylenase at moderate levels. FAD2-3 was not expressed in developing seed. Real-time RT-PCR absolute transcript quantitation showed 10(4)-fold higher acetylenase expression in developing seeds than in flower heads. Collectively, the results show that both the acetylenase expression level and the co-expression of other desaturases may contribute to the tissue specificity of crepenynate production. Helianthus annuus contains a Delta12 acetylenase in a polyacetylene biosynthetic pathway, so does not accumulate crepenynate. Real-time RT-PCR analysis showed relatively strong acetylenase expression in young sunflowers. Acetylenase transcription is observed in both species without accumulation of the enzymatic product, crepenynate. Functional expression of acetylenase appears to be affected by competition and collaboration with other enzymes. PMID:17329262

  16. Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products. Progress report, June 1, 1990--June 30, 1992

    SciTech Connect

    Kuchka, M.R.

    1992-08-01

    Many individual chloroplast genes require the products of a collection of nuclear genes for their successful expression. These nuclear gene products apparently work with great specificity, each committed to the expression of a single chloroplast gene. We have chosen as a model nuclear mutants of Chlamydomonas affected in different stages in the expression of the chloroplast encoded Photosystem II polypeptide, D2. We have made the progress in understanding how nuclear gene products affect the translation of the D2 encoding MRNA. Two nuclear genes are required for this process which have been mapped genetically. In contrast to other examples of nuclear control of translation in the chloroplast, these nuclear gene products appear to be required either for specific stages in translation elongation or for the post-translational stabilization of the nascent D2 protein. Pseudoreversion analysis has led us to a locus which may be directly involved in D2 expression. We have made considerable progress in pursuing the molecular basis of psbd MRNA stabilization. psbD 5` UTR specific transcripts have been synthesized in vitro and used in gel mobility shift assays. UV-crosslinking studies are underway to identify the transacting factors which bind to these sequences. The continued examination of these mutants will help us to understand how nuclear gene products work in this specific case of chloroplast gene expression, and will elucidate how two distinct genomes can interact generally.

  17. Transcriptional Truncation of the Long Coding Imprinted Gene Usp29

    PubMed Central

    He, Hongzhi; Ye, An; Kim, Joomyeong

    2016-01-01

    Usp29 (Ubiquitin-specific protease 29) is a paternally expressed gene located upstream of another imprinted gene Peg3. In the current study, the transcription of this long coding gene spanning a 250-kb genomic distance was truncated using a knockin allele. According to the results, paternal transmission of the mutant allele resulted in reduced body and litter sizes whereas the maternal transmission caused no obvious effects. In the paternal mutant, the expression levels of Usp29 were reduced to 14–18% level of the wild-type littermates due to the Poly-A signal included in the knockin cassette. Expression analyses further revealed an unusual female-specific up-regulation of the adjacent imprinted gene Zfp264 in the mutant. Consistent with this, the promoter of Zfp264 was hypomethylated only in the female mutant. Interestingly, this female-specific hypomethylation by the knockin allele was not detected in the offspring of an interspecific crossing, indicating its sensitivity to genetic background. Overall, the results suggest that the transcription of Usp29 may be involved in DNA methylation setting of Zfp264 promoter in a sex-specific manner. PMID:27327533

  18. WRKY transcription factor genes in wild rice Oryza nivara.

    PubMed

    Xu, Hengjian; Watanabe, Kenneth A; Zhang, Liyuan; Shen, Qingxi J

    2016-08-01

    The WRKY transcription factor family is one of the largest gene families involved in plant development and stress response. Although many WRKY genes have been studied in cultivated rice (Oryza sativa), the WRKY genes in the wild rice species Oryza nivara, the direct progenitor of O. sativa, have not been studied. O. nivara shows abundant genetic diversity and elite drought and disease resistance features. Herein, a total of 97 O. nivara WRKY (OnWRKY) genes were identified. RNA-sequencing demonstrates that OnWRKY genes were generally expressed at higher levels in the roots of 30-day-old plants. Bioinformatic analyses suggest that most of OnWRKY genes could be induced by salicylic acid, abscisic acid, and drought. Abundant potential MAPK phosphorylation sites in OnWRKYs suggest that activities of most OnWRKYs can be regulated by phosphorylation. Phylogenetic analyses of OnWRKYs support a novel hypothesis that ancient group IIc OnWRKYs were the original ancestors of only some group IIc and group III WRKYs. The analyses also offer strong support that group IIc OnWRKYs containing the HVE sequence in their zinc finger motifs were derived from group Ia WRKYs. This study provides a solid foundation for the study of the evolution and functions of WRKY genes in O. nivara. PMID:27345721

  19. WRKY transcription factor genes in wild rice Oryza nivara

    PubMed Central

    Xu, Hengjian; Watanabe, Kenneth A.; Zhang, Liyuan; Shen, Qingxi J.

    2016-01-01

    The WRKY transcription factor family is one of the largest gene families involved in plant development and stress response. Although many WRKY genes have been studied in cultivated rice (Oryza sativa), the WRKY genes in the wild rice species Oryza nivara, the direct progenitor of O. sativa, have not been studied. O. nivara shows abundant genetic diversity and elite drought and disease resistance features. Herein, a total of 97 O. nivara WRKY (OnWRKY) genes were identified. RNA-sequencing demonstrates that OnWRKY genes were generally expressed at higher levels in the roots of 30-day-old plants. Bioinformatic analyses suggest that most of OnWRKY genes could be induced by salicylic acid, abscisic acid, and drought. Abundant potential MAPK phosphorylation sites in OnWRKYs suggest that activities of most OnWRKYs can be regulated by phosphorylation. Phylogenetic analyses of OnWRKYs support a novel hypothesis that ancient group IIc OnWRKYs were the original ancestors of only some group IIc and group III WRKYs. The analyses also offer strong support that group IIc OnWRKYs containing the HVE sequence in their zinc finger motifs were derived from group Ia WRKYs. This study provides a solid foundation for the study of the evolution and functions of WRKY genes in O. nivara. PMID:27345721

  20. Zinc triggers a complex transcriptional and post-transcriptional regulation of the metal homeostasis gene FRD3 in Arabidopsis relatives

    PubMed Central

    Charlier, Jean-Benoit; Polese, Catherine; Nouet, Cécile; Carnol, Monique; Bosman, Bernard; Krämer, Ute; Motte, Patrick; Hanikenne, Marc

    2015-01-01

    In Arabidopsis thaliana, FRD3 (FERRIC CHELATE REDUCTASE DEFECTIVE 3) plays a central role in metal homeostasis. FRD3 is among a set of metal homeostasis genes that are constitutively highly expressed in roots and shoots of Arabidopsis halleri, a zinc hyperaccumulating and hypertolerant species. Here, we examined the regulation of FRD3 by zinc in both species to shed light on the evolutionary processes underlying the evolution of hyperaccumulation in A. halleri. We combined gene expression studies with the use of β-glucuronidase and green fluorescent protein reporter constructs to compare the expression profile and transcriptional and post-transcriptional regulation of FRD3 in both species. The AtFRD3 and AhFRD3 genes displayed a conserved expression profile. In A. thaliana, alternative transcription initiation sites from two promoters determined transcript variants that were differentially regulated by zinc supply in roots and shoots to favour the most highly translated variant under zinc-excess conditions. In A. halleri, a single transcript variant with higher transcript stability and enhanced translation has been maintained. The FRD3 gene thus undergoes complex transcriptional and post-transcriptional regulation in Arabidopsis relatives. Our study reveals that a diverse set of mechanisms underlie increased gene dosage in the A. halleri lineage and illustrates how an environmental challenge can alter gene regulation. PMID:25900619

  1. Transcriptional regulation of the presenilin-1 gene controls gamma-secretase activity.

    PubMed

    Lee, Sebum; Das, Hriday K

    2010-01-01

    Inhibition of basal JNK activity by JNK inhibitor SP600125 or JNK1siRNA repressed presenilin-1 (PS1) expression in SK-N-SH cells by augmenting the level of p53, a repressor of the PS1 gene (1). We now showed that repression of PS1 transcription by JNK inhibitor SP600125 inhibited gamma-secretase mediated processing of amyloid precursor protein (APP) resulting in the accumulation of C99 fragment and the reduction of secreted Abeta40 level without altering the expression of nicastrin (NCT). Co-treatment of cells with SP600125 and p53 inhibitor, pifithrin-alpha, partially nullified the suppressive effects of SP610025 on PS1 expression and secreted Abeta40 level. Suppression of JNK1 by JNK1siRNA also decreased Abeta40 level. Furthermore, overexpression of the repressors p53, ZNF237 and CHD3 of the PS1 gene also suppressed the processing of APP through repression of PS1 transcription by deacetylation of histone at the PS1 promoter. Transcriptional activator Ets2 increased PS1 protein and secreted Abeta40 levels without affecting the expression of NCT by activating PS1 transcription via hyper-acetylation of histone at the PS1 promoter. Therefore, regulation of PS1 transcription modulates gamma-secretase activity. PMID:20036849

  2. Stability of mRNA/DNA and DNA/DNA Duplexes Affects mRNA Transcription

    PubMed Central

    Kraeva, Rayna I.; Krastev, Dragomir B.; Roguev, Assen; Ivanova, Anna; Nedelcheva-Veleva, Marina N.; Stoynov, Stoyno S.

    2007-01-01

    Nucleic acids, due to their structural and chemical properties, can form double-stranded secondary structures that assist the transfer of genetic information and can modulate gene expression. However, the nucleotide sequence alone is insufficient in explaining phenomena like intron-exon recognition during RNA processing. This raises the question whether nucleic acids are endowed with other attributes that can contribute to their biological functions. In this work, we present a calculation of thermodynamic stability of DNA/DNA and mRNA/DNA duplexes across the genomes of four species in the genus Saccharomyces by nearest-neighbor method. The results show that coding regions are more thermodynamically stable than introns, 3′-untranslated regions and intergenic sequences. Furthermore, open reading frames have more stable sense mRNA/DNA duplexes than the potential antisense duplexes, a property that can aid gene discovery. The lower stability of the DNA/DNA and mRNA/DNA duplexes of 3′-untranslated regions and the higher stability of genes correlates with increased mRNA level. These results suggest that the thermodynamic stability of DNA/DNA and mRNA/DNA duplexes affects mRNA transcription. PMID:17356699

  3. Regulation of HDL genes: transcriptional, posttranscriptional, and posttranslational.

    PubMed

    Kardassis, Dimitris; Gafencu, Anca; Zannis, Vassilis I; Davalos, Alberto

    2015-01-01

    HDL regulation is exerted at multiple levels including regulation at the level of transcription initiation by transcription factors and signal transduction cascades; regulation at the posttranscriptional level by microRNAs and other noncoding RNAs which bind to the coding or noncoding regions of HDL genes regulating mRNA stability and translation; as well as regulation at the posttranslational level by protein modifications, intracellular trafficking, and degradation. The above mechanisms have drastic effects on several HDL-mediated processes including HDL biogenesis, remodeling, cholesterol efflux and uptake, as well as atheroprotective functions on the cells of the arterial wall. The emphasis is on mechanisms that operate in physiologically relevant tissues such as the liver (which accounts for 80% of the total HDL-C levels in the plasma), the macrophages, the adrenals, and the endothelium. Transcription factors that have a significant impact on HDL regulation such as hormone nuclear receptors and hepatocyte nuclear factors are extensively discussed both in terms of gene promoter recognition and regulation but also in terms of their impact on plasma HDL levels as was revealed by knockout studies. Understanding the different modes of regulation of this complex lipoprotein may provide useful insights for the development of novel HDL-raising therapies that could be used to fight against atherosclerosis which is the underlying cause of coronary heart disease. PMID:25522987

  4. Genetic Variants in the STMN1 Transcriptional Regulatory Region Affect Promoter Activity and Fear Behavior in English Springer Spaniels

    PubMed Central

    Zhang, Hanying; Xu, Yinxue

    2016-01-01

    Stathmin 1 (STMN1) is a neuronal growth-associated protein that is involved in microtubule dynamics and plays an important role in synaptic outgrowth and plasticity. Given that STMN1 affects fear behavior, we hypothesized that genetic variations in the STMN1 transcriptional regulatory region affect gene transcription activity and control fear behavior. In this study, two single nucleotide polymorphisms (SNPs), g. -327 A>G and g. -125 C>T, were identified in 317 English Springer Spaniels. A bioinformatics analysis revealed that both were loci located in the canine STMN1 putative promoter region and affected transcription factor binding. A statistical analysis revealed that the TT genotype at g.-125 C>T produced a significantly greater fear level than that of the CC genotype (P < 0.05). Furthermore, the H4H4 (GTGT) haplotype combination was significantly associated with canine fear behavior (P < 0.01). Using serially truncated constructs of the STMN1 promoters and the luciferase reporter, we found that a 395 bp (−312 nt to +83 nt) fragment constituted the core promoter region. The luciferase assay also revealed that the H4 (GT) haplotype promoter had higher activity than that of other haplotypes. Overall, our results suggest that the two SNPs in the canine STMN1 promoter region could affect canine fear behavior by altering STMN1 transcriptional activity. PMID:27390866

  5. Novel Transcriptional Regulons for Autotrophic Cycle Genes in Crenarchaeota

    PubMed Central

    Leyn, Semen A.; Rodionova, Irina A.; Li, Xiaoqing

    2015-01-01

    ABSTRACT Autotrophic microorganisms are able to utilize carbon dioxide as their only carbon source, or, alternatively, many of them can grow heterotrophically on organics. Different variants of autotrophic pathways have been identified in various lineages of the phylum Crenarchaeota. Aerobic members of the order Sulfolobales utilize the hydroxypropionate-hydroxybutyrate cycle (HHC) to fix inorganic carbon, whereas anaerobic Thermoproteales use the dicarboxylate-hydroxybutyrate cycle (DHC). Knowledge of transcriptional regulation of autotrophic pathways in Archaea is limited. We applied a comparative genomics approach to predict novel autotrophic regulons in the Crenarchaeota. We report identification of two novel DNA motifs associated with the autotrophic pathway genes in the Sulfolobales (HHC box) and Thermoproteales (DHC box). Based on genome context evidence, the HHC box regulon was attributed to a novel transcription factor from the TrmB family named HhcR. Orthologs of HhcR are present in all Sulfolobales genomes but were not found in other lineages. A predicted HHC box regulatory motif was confirmed by in vitro binding assays with the recombinant HhcR protein from Metallosphaera yellowstonensis. For the DHC box regulon, we assigned a different potential regulator, named DhcR, which is restricted to the order Thermoproteales. DhcR in Thermoproteus neutrophilus (Tneu_0751) was previously identified as a DNA-binding protein with high affinity for the promoter regions of two autotrophic operons. The global HhcR and DhcR regulons reconstructed by comparative genomics were reconciled with available omics data in Metallosphaera and Thermoproteus spp. The identified regulons constitute two novel mechanisms for transcriptional control of autotrophic pathways in the Crenarchaeota. IMPORTANCE Little is known about transcriptional regulation of carbon dioxide fixation pathways in Archaea. We previously applied the comparative genomics approach for reconstruction of Dtx

  6. The transcriptional terminator sequences downstream of the covR gene terminate covR/S operon transcription to generate covR monocistronic transcripts in Streptococcus pyogenes.

    PubMed

    Chiang-Ni, Chuan; Tsou, Chih-Cheng; Lin, Yee-Shin; Chuang, Woei-Jer; Lin, Ming-T; Liu, Ching-Chuan; Wu, Jiunn-Jong

    2008-12-31

    CovR/S is an important two component regulatory system, which regulates about 15% of the gene expression in Streptococcus pyogenes. The covR/S locus was identified as an operon generating an RNA transcript around 2.5-kb in size. In this study, we found the covR/S operon produced three RNA transcripts (around 2.5-, 1.0-, and 0.8-kb in size). Using RNA transcriptional terminator sequence prediction and transcriptional terminator analysis, we identified two atypical rho-independent terminator sequences downstream of the covR gene and showed these terminator sequences terminate RNA transcription efficiently. These results indicate that covR/S operon generates covR/S transcript and monocistronic covR transcripts. PMID:18824088

  7. Effect of replication on epigenetic memory and consequences on gene transcription

    NASA Astrophysics Data System (ADS)

    Zerihun, Mehari B.; Vaillant, Cédric; Jost, Daniel

    2015-04-01

    Gene activity in eukaryotes is in part regulated at the level of chromatin through the assembly of local chromatin states that are more or less permissive to transcription. How do these chromatin states achieve their functions and whether or not they contribute to the epigenetic inheritance of the transcriptional program remain to be elucidated. In cycling cells, stability is indeed strongly challenged by the periodic occurrence of replication and cell division. To address this question, we perform simulations of the stochastic dynamics of chromatin states when driven out-of-equilibrium by periodic perturbations. We show how epigenetic memory is significantly affected by the cell cycle length. In addition, we develop a simple model to connect the epigenetic state to the transcriptional state and gene activity. In particular, it suggests that replication may induce transcriptional bursting at repressive loci. Finally, we discuss how our findings—effect of replication and link to gene transcription—have original and deep implications to various biological contexts of epigenetic memory.

  8. Effect of nonylphenol on response of physiology and photosynthesis-related gene transcription of Chlorella vulgaris.

    PubMed

    Qian, Haifeng; Pan, Xiangjie; Shi, Shutian; Yu, Shuqiong; Jiang, Haiyan; Lin, Zhifan; Fu, Zhengwei

    2011-11-01

    Nonylphenol (NP) is regarded as a kind of persistent organic pollutant which exists ubiquitously in the environment. The objective of this study was to evaluate the effects of NP on Chlorella vulgaris physiological indices and gene transcription. The results showed that NP stress inhibited algal growth in short-term bioassay. NP also decreased chlorophyll content, including chl a, chl b, and total chlorophyll. NP caused oxidant hurt by overproducing reactive oxygen species (ROS), which might destroy the overall membrane system to cause malondialdehyde content increase. NP inhibited photosynthesis-related gene transcription in C. vulgaris after 24 to 48 h exposure. The lowest transcript levels of psaB, psbA, and rbcL in C. vulgaris decreased to only 18.5%, 7%, and 4% of the control, respectively. Taken together, our results demonstrate that NP is toxic to fresh algae growth by affecting the photosynthesis-related genes transcription and overproducing ROS to disrupt cell structure in a short period. PMID:21207133

  9. Transcriptional regulation of FoxO3 gene by glucocorticoids in murine myotubes.

    PubMed

    Kuo, Taiyi; Liu, Patty H; Chen, Tzu-Chieh; Lee, Rebecca A; New, Jenny; Zhang, Danyun; Lei, Cassandra; Chau, Andy; Tang, Yicheng; Cheung, Edna; Wang, Jen-Chywan

    2016-04-01

    Glucocorticoids and FoxO3 exert similar metabolic effects in skeletal muscle. FoxO3 gene expression was increased by dexamethasone (Dex), a synthetic glucocorticoid, both in vitro and in vivo. In C2C12 myotubes the increased expression is due to, at least in part, the elevated rate of FoxO3 gene transcription. In the mouse FoxO3 gene, we identified three glucocorticoid receptor (GR) binding regions (GBRs): one being upstream of the transcription start site, -17kbGBR; and two in introns, +45kbGBR and +71kbGBR. Together, these three GBRs contain four 15-bp glucocorticoid response elements (GREs). Micrococcal nuclease (MNase) assay revealed that Dex treatment increased the sensitivity to MNase in the GRE of +45kbGBR and +71kbGBR upon 30- and 60-min Dex treatment, respectively. Conversely, Dex treatment did not affect the chromatin structure near the -17kbGBR, in which the GRE is located in the linker region. Dex treatment also increased histone H3 and/or H4 acetylation in genomic regions near all three GBRs. Moreover, using chromatin conformation capture (3C) assay, we showed that Dex treatment increased the interaction between the -17kbGBR and two genomic regions: one located around +500 bp and the other around +73 kb. Finally, the transcriptional coregulator p300 was recruited to all three GBRs upon Dex treatment. The reduction of p300 expression decreased FoxO3 gene expression and Dex-stimulated interaction between distinct genomic regions of FoxO3 gene identified by 3C. Overall, our results demonstrate that glucocorticoids activated FoxO3 gene transcription through multiple GREs by chromatin structural change and DNA looping. PMID:26758684

  10. Controlling for Gene Expression Changes in Transcription Factor Protein Networks*

    PubMed Central

    Banks, Charles A. S.; Lee, Zachary T.; Boanca, Gina; Lakshminarasimhan, Mahadevan; Groppe, Brad D.; Wen, Zhihui; Hattem, Gaye L.; Seidel, Chris W.; Florens, Laurence; Washburn, Michael P.

    2014-01-01

    The development of affinity purification technologies combined with mass spectrometric analysis of purified protein mixtures has been used both to identify new protein–protein interactions and to define the subunit composition of protein complexes. Transcription factor protein interactions, however, have not been systematically analyzed using these approaches. Here, we investigated whether ectopic expression of an affinity tagged transcription factor as bait in affinity purification mass spectrometry experiments perturbs gene expression in cells, resulting in the false positive identification of bait-associated proteins when typical experimental controls are used. Using quantitative proteomics and RNA sequencing, we determined that the increase in the abundance of a set of proteins caused by overexpression of the transcription factor RelA is not sufficient for these proteins to then co-purify non-specifically and be misidentified as bait-associated proteins. Therefore, typical controls should be sufficient, and a number of different baits can be compared with a common set of controls. This is of practical interest when identifying bait interactors from a large number of different baits. As expected, we found several known RelA interactors enriched in our RelA purifications (NFκB1, NFκB2, Rel, RelB, IκBα, IκBβ, and IκBε). We also found several proteins not previously described in association with RelA, including the small mitochondrial chaperone Tim13. Using a variety of biochemical approaches, we further investigated the nature of the association between Tim13 and NFκB family transcription factors. This work therefore provides a conceptual and experimental framework for analyzing transcription factor protein interactions. PMID:24722732

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

    PubMed Central

    Smita, Shuchi; Katiyar, Amit; Chinnusamy, Viswanathan; Pandey, Dev M.; Bansal, Kailash C.

    2015-01-01

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

  12. Transcriptional analysis of the Pseudomonas aeruginosa exoenzyme S structural gene.

    PubMed Central

    Yahr, T L; Hovey, A K; Kulich, S M; Frank, D W

    1995-01-01

    The transcriptional regulation of the Pseudomonas aeruginosa exoS gene was investigated. Expression of exoS in P. aeruginosa PA103 was dependent upon growth in a low-cation environment and the presence of a functional exsA gene. Promoter fusion analysis indicated that a 285-bp PstI-NsiI fragment, located 5' of the exoS coding region, contained a functional promoter for exoS. Expression of the reporter gene was inducible in a low-cation growth environment and required a functional copy of exsA. Divergent promoters, coordinately regulated with exoS transcription, were identified within the PstI-NsiI fragment. A fusion derivative of ExsA, MALA3A2, was shown to bind directly to the PstI-NsiI probe. DNase I protection analysis demonstrated that MALA3A2 bound to the intergenic region between the postulated -35 boxes of each promoter region. Northern (RNA) blot analysis with probes internal to and upstream of exoS demonstrated that separate, coordinately regulated mRNAs were expressed in P. aeruginosa. These data suggested that a locus, coregulated with exoS transcription, was located upstream of exoS. DNA sequence analysis of the exoS upstream region revealed three open reading frames, ORF 1, ORF 2, and ORF 3. ORF 1 demonstrated significant homology to the SycE/YerA protein of Yersinia sp. SycE/YerA is postulated to function as a chaperone for the YopE cytotoxin. The loci encoding YopE and ExoS show similarities in genetic organization, protein composition, and regulation. PMID:7868588

  13. Estrogen Signaling Multiple Pathways to Impact Gene Transcription

    PubMed Central

    Marino, Maria; Galluzzo, Paola; Ascenzi, Paolo

    2006-01-01

    Steroid hormones exert profound effects on cell growth, development, differentiation, and homeostasis. Their effects are mediated through specific intracellular steroid receptors that act via multiple mechanisms. Among others, the action mechanism starting upon 17β-estradiol (E2) binds to its receptors (ER) is considered a paradigmatic example of how steroid hormones function. Ligand-activated ER dimerizes and translocates in the nucleus where it recognizes specific hormone response elements located in or near promoter DNA regions of target genes. Behind the classical genomic mechanism shared with other steroid hormones, E2 also modulates gene expression by a second indirect mechanism that involves the interaction of ER with other transcription factors which, in turn, bind their cognate DNA elements. In this case, ER modulates the activities of transcription factors such as the activator protein (AP)-1, nuclear factor-κB (NF-κB) and stimulating protein-1 (Sp-1), by stabilizing DNA-protein complexes and/or recruiting co-activators. In addition, E2 binding to ER may also exert rapid actions that start with the activation of a variety of signal transduction pathways (e.g. ERK/MAPK, p38/MAPK, PI3K/AKT, PLC/PKC). The debate about the contribution of different ER-mediated signaling pathways to coordinate the expression of specific sets of genes is still open. This review will focus on the recent knowledge about the mechanism by which ERs regulate the expression of target genes and the emerging field of integration of membrane and nuclear receptor signaling, giving examples of the ways by which the genomic and non-genomic actions of ERs on target genes converge. PMID:18369406

  14. Transcriptional profiling reveals regulated genes in the hippocampus during memory formation

    NASA Technical Reports Server (NTRS)

    Donahue, Christine P.; Jensen, Roderick V.; Ochiishi, Tomoyo; Eisenstein, Ingrid; Zhao, Mingrui; Shors, Tracey; Kosik, Kenneth S.

    2002-01-01

    Transcriptional profiling (TP) offers a powerful approach to identify genes activated during memory formation and, by inference, the molecular pathways involved. Trace eyeblink conditioning is well suited for the study of regional gene expression because it requires the hippocampus, whereas the highly parallel task, delay conditioning, does not. First, we determined when gene expression was most regulated during trace conditioning. Rats were exposed to 200 trials per day of paired and unpaired stimuli each day for 4 days. Changes in gene expression were most apparent 24 h after exposure to 200 trials. Therefore, we profiled gene expression in the hippocampus 24 h after 200 trials of trace eyeblink conditioning, on multiple arrays using additional animals. Of 1,186 genes on the filter array, seven genes met the statistical criteria and were also validated by real-time polymerase chain reaction. These genes were growth hormone (GH), c-kit receptor tyrosine kinase (c-kit), glutamate receptor, metabotropic 5 (mGluR5), nerve growth factor-beta (NGF-beta), Jun oncogene (c-Jun), transmembrane receptor Unc5H1 (UNC5H1), and transmembrane receptor Unc5H2 (UNC5H2). All these genes, except for GH, were downregulated in response to trace conditioning. GH was upregulated; therefore, we also validated the downregulation of the GH inhibitor, somatostatin (SST), even though it just failed to meet criteria on the arrays. By during situ hybridization, GH was expressed throughout the cell layers of the hippocampus in response to trace conditioning. None of the genes regulated in trace eyeblink conditioning were similarly affected by delay conditioning, a task that does not require the hippocampus. These findings demonstrate that transcriptional profiling can exhibit a repertoire of genes sensitive to the formation of hippocampal-dependent associative memories.

  15. Variability of Gene Expression Identifies Transcriptional Regulators of Early Human Embryonic Development

    PubMed Central

    Hasegawa, Yu; Taylor, Deanne; Ovchinnikov, Dmitry A.; Wolvetang, Ernst J.; de Torrenté, Laurence; Mar, Jessica C.

    2015-01-01

    An analysis of gene expression variability can provide an insightful window into how regulatory control is distributed across the transcriptome. In a single cell analysis, the inter-cellular variability of gene expression measures the consistency of transcript copy numbers observed between cells in the same population. Application of these ideas to the study of early human embryonic development may reveal important insights into the transcriptional programs controlling this process, based on which components are most tightly regulated. Using a published single cell RNA-seq data set of human embryos collected at four-cell, eight-cell, morula and blastocyst stages, we identified genes with the most stable, invariant expression across all four developmental stages. Stably-expressed genes were found to be enriched for those sharing indispensable features, including essentiality, haploinsufficiency, and ubiquitous expression. The stable genes were less likely to be associated with loss-of-function variant genes or human recessive disease genes affected by a DNA copy number variant deletion, suggesting that stable genes have a functional impact on the regulation of some of the basic cellular processes. Genes with low expression variability at early stages of development are involved in regulation of DNA methylation, responses to hypoxia and telomerase activity, whereas by the blastocyst stage, low-variability genes are enriched for metabolic processes as well as telomerase signaling. Based on changes in expression variability, we identified a putative set of gene expression markers of morulae and blastocyst stages. Experimental validation of a blastocyst-expressed variability marker demonstrated that HDDC2 plays a role in the maintenance of pluripotency in human ES and iPS cells. Collectively our analyses identified new regulators involved in human embryonic development that would have otherwise been missed using methods that focus on assessment of the average expression

  16. Transcriptional profiling reveals regulated genes in the hippocampus during memory formation.

    PubMed

    Donahue, Christine P; Jensen, Roderick V; Ochiishi, Tomoyo; Eisenstein, Ingrid; Zhao, Mingrui; Shors, Tracey; Kosik, Kenneth S

    2002-01-01

    Transcriptional profiling (TP) offers a powerful approach to identify genes activated during memory formation and, by inference, the molecular pathways involved. Trace eyeblink conditioning is well suited for the study of regional gene expression because it requires the hippocampus, whereas the highly parallel task, delay conditioning, does not. First, we determined when gene expression was most regulated during trace conditioning. Rats were exposed to 200 trials per day of paired and unpaired stimuli each day for 4 days. Changes in gene expression were most apparent 24 h after exposure to 200 trials. Therefore, we profiled gene expression in the hippocampus 24 h after 200 trials of trace eyeblink conditioning, on multiple arrays using additional animals. Of 1,186 genes on the filter array, seven genes met the statistical criteria and were also validated by real-time polymerase chain reaction. These genes were growth hormone (GH), c-kit receptor tyrosine kinase (c-kit), glutamate receptor, metabotropic 5 (mGluR5), nerve growth factor-beta (NGF-beta), Jun oncogene (c-Jun), transmembrane receptor Unc5H1 (UNC5H1), and transmembrane receptor Unc5H2 (UNC5H2). All these genes, except for GH, were downregulated in response to trace conditioning. GH was upregulated; therefore, we also validated the downregulation of the GH inhibitor, somatostatin (SST), even though it just failed to meet criteria on the arrays. By during situ hybridization, GH was expressed throughout the cell layers of the hippocampus in response to trace conditioning. None of the genes regulated in trace eyeblink conditioning were similarly affected by delay conditioning, a task that does not require the hippocampus. These findings demonstrate that transcriptional profiling can exhibit a repertoire of genes sensitive to the formation of hippocampal-dependent associative memories. PMID:12542233

  17. Identification of a novel transcript of human MD2 gene.

    PubMed

    Shen, Chen; Shen, A-Dong

    2016-09-15

    Myeloid differentiation protein 2 (MD2) regulates bacterial lipopolysaccharide (LPS) triggered anti-bacterial immune response as a broker between LPS and Toll-like receptor 4 (TLR4). In this study, we identified a novel naturally occurring spliceosome of human MD2, termed as MD2-T3. This transcript lacked two exons of MD2 gene. By protein structure analysis and literature review, we predicted that MD2-T3 isoform might execute regulatory biological effects such as limiting LPS-triggered TLR4 signaling. PMID:27317890

  18. Structure and transcription of the actin gene of Trypanosoma brucei

    SciTech Connect

    Ben Amar, M.F.; Pays, A.; Tebabi, P.; Dero, B.; Seebeck, T.; Steinert, M.; Pays, E.

    1988-05-01

    In Trypanosoma brucei, the actin gene is present in a cluster of two, three, or four tandemly linked copies, depending on the strain. Each cluster seems to exist in two allelic versions, as suggested by the polymorphism of both gene number and restriction fragment length in the DNA from cloned trypanosomes. The amplification of the gene copy number probably occurs through unequal sister chromatic exchange. The chromosomes harboring the actin genes belong to the large size class. The coding sequence was 1,128 nucleotides long and showed 60 to 70% homology to other eucaryotic actin genes. Surprisingly, this homology seemed weaker with Trypanosoma congolense, Trypanosoma cruzi, Trypanosoma vivax, Trypanosoma mega, or Leishmania acting-specific sequences. The mRNA was around 1.6 kilobases long and was synthesized at the same level in bloodstream and procyclic forms of the parasite. Large RNA precursors, up to 7.7 kilobases, were found in a pattern identical in strains containing either two or three gene copies. Probing of the flanking regions of the gene with either steady-state or in vitro transcripts, as well as S1 nuclease protection and primer extension experiments, allowed mapping of the 3' splice site of the actin mRNA, 38 nucleotides upstream from the translation initiation codon. A variably sized poly(dT) tract was found about 30 base pairs ahead of the splice site. The largest detected actin mRNA precursor seemed to give rise to at least two additional stable mRNAs. The RNA polymerase transcribing the actin gene exhibited the same sensitivity to inhibition by ..cap alpha..-amanitin as that transcribing both the spliced leader and the bulk of polyadenylated mRNAs.

  19. The Trypanosoma brucei protein phosphatase gene: polycistronic transcription with the RNA polymerase II largest subunit gene.

    PubMed Central

    Evers, R; Cornelissen, A W

    1990-01-01

    We have previously described the trypanosomal gene encoding the largest subunit of RNA polymerase II (RNAP II) and found that two almost identical genes are encoded within the Trypanosoma brucei genome. Here we show by Southern analyses that the 5' breakpoint between both loci is located approximately 7.5 kb upstream of the RNAP II genes. Northern analyses revealed that the 5' duplicated segment contains at least four other genes, which are transcribed in both bloodstream and procyclic trypanosomes. The gene located immediately upstream of the RNAP II gene in both loci was characterized by sequence analyses. The deduced amino acid sequences show a high degree of similarity to the catalytic subunit of protein phosphatase class 1 (PP1) genes. S1 mapping provided strong evidence in support of the fact that the PP1 and RNAP II genes belong to a single transcription unit. Images PMID:2169604

  20. Transcription factor IIIB generates extended DNA interactions in RNA polymerase III transcription complexes on tRNA genes.

    PubMed Central

    Kassavetis, G A; Riggs, D L; Negri, R; Nguyen, L H; Geiduschek, E P

    1989-01-01

    Transcription complexes that assemble on tRNA genes in a crude Saccharomyces cerevisiae cell extract extend over the entire transcription unit and approximately 40 base pairs of contiguous 5'-flanking DNA. We show here that the interaction with 5'-flanking DNA is due to a protein that copurifies with transcription factor TFIIIB through several steps of purification and shares characteristic properties that are normally ascribed to TFIIIB: dependence on prior binding of TFIIIC and great stability once the TFIIIC-TFIIIB-DNA complex is formed. SUP4 gene (tRNATyr) DNA that was cut within the 5'-flanking sequence (either 31 or 28 base pairs upstream of the transcriptional start site) was no longer able to stably incorporate TFIIIB into a transcription complex. The TFIIIB-dependent 5'-flanking DNA protein interaction was predominantly not sequence specific. The extension of the transcription complex into this DNA segment does suggest two possible explanations for highly diverse effects of flanking-sequence substitutions on tRNA gene transcription: either (i) proteins that are capable of binding to these upstream DNA segments are also potentially capable of stimulating or interfering with the incorporation of TFIIIB into transcription complexes or (ii) 5'-flanking sequence influences the rate of assembly of TFIIIB into stable transcription complexes. Images PMID:2668737

  1. Separation of mutational and transcriptional enhancers in immunoglobulin genes

    PubMed Central

    Kothapalli, Naga Rama; Collura, Kaitlin M.; Norton, Darrell D.; Fugmann, Sebastian D.

    2011-01-01

    Secondary immunoglobulin (Ig) gene diversification relies on activation-induced cytidine deaminase (AID) to create U:G mismatches that are subsequently fixed by mutagenic repair pathways. AID activity is focused to Ig loci by cis-regulatory DNA sequences named targeting elements. Here we show that in contrast to prevailing thought in the field, the targeting elements in the chicken IGL locus are distinct from classical transcriptional enhancers. These mutational enhancer elements (MEEs) are required over and above transcription to recruit AID-mediated mutagenesis to Ig loci. We identified a small 222 bp fragment in the chicken IGL locus that enhances mutagenesis without boosting transcription, and this sequence represents a key component of a MEE. Lastly, MEEs are evolutionarily conserved amongst birds, both in sequence and function, and contain several highly conserved sequence modules that are likely involved in recruiting trans-acting targeting factors. We propose that MEEs represent a novel class of cis-regulatory elements whose function is to control genomic integrity. PMID:21844395

  2. Separation of mutational and transcriptional enhancers in Ig genes.

    PubMed

    Kothapalli, Naga Rama; Collura, Kaitlin M; Norton, Darrell D; Fugmann, Sebastian D

    2011-09-15

    Secondary Ig gene diversification relies on activation-induced cytidine deaminase (AID) to create U:G mismatches that are subsequently fixed by mutagenic repair pathways. AID activity is focused to Ig loci by cis-regulatory DNA sequences named targeting elements. In this study, we show that in contrast to prevailing thought in the field, the targeting elements in the chicken IGL locus are distinct from classical transcriptional enhancers. These mutational enhancer elements (MEEs) are required over and above transcription to recruit AID-mediated mutagenesis to Ig loci. We identified a small 222-bp fragment in the chicken IGL locus that enhances mutagenesis without boosting transcription, and this sequence represents a key component of an MEE. Lastly, MEEs are evolutionarily conserved among birds, both in sequence and function, and contain several highly conserved sequence modules that are likely involved in recruiting trans-acting targeting factors. We propose that MEEs represent a novel class of cis-regulatory elements for which the function is to control genomic integrity. PMID:21844395

  3. Analysis of the Highly Diverse Gene Borders in Ebola Virus Reveals a Distinct Mechanism of Transcriptional Regulation

    PubMed Central

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

    2014-01-01

    ABSTRACT Ebola virus (EBOV) belongs to the group of nonsegmented negative-sense RNA viruses. The seven EBOV genes are separated by variable gene borders, including short (4- or 5-nucleotide) intergenic regions (IRs), a single long (144-nucleotide) IR, and gene overlaps, where the neighboring gene end and start signals share five conserved nucleotides. The unique structure of the gene overlaps and the presence of a single long IR are conserved among all filoviruses. Here, we sought to determine the impact of the EBOV gene borders during viral transcription. We show that readthrough mRNA synthesis occurs in EBOV-infected cells irrespective of the structure of the gene border, indicating that the gene overlaps do not promote recognition of the gene end signal. However, two consecutive gene end signals at the VP24 gene might improve termination at the VP24-L gene border, ensuring efficient L gene expression. We further demonstrate that the long IR is not essential for but regulates transcription reinitiation in a length-dependent but sequence-independent manner. Mutational analysis of bicistronic minigenomes and recombinant EBOVs showed no direct correlation between IR length and reinitiation rates but demonstrated that specific IR lengths not found naturally in filoviruses profoundly inhibit downstream gene expression. Intriguingly, although truncation of the 144-nucleotide-long IR to 5 nucleotides did not substantially affect EBOV transcription, it led to a significant reduction of viral growth. IMPORTANCE Our current understanding of EBOV transcription regulation is limited due to the requirement for high-containment conditions to study this highly pathogenic virus. EBOV is thought to share many mechanistic features with well-analyzed prototype nonsegmented negative-sense RNA viruses. A single polymerase entry site at the 3′ end of the genome determines that transcription of the genes is mainly controlled by gene order and cis-acting signals found at the gene

  4. Light affects gene expression during leafhopper diapause

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The condition of diapause in the glassy-winged sharpshooter, GWSS, Homalodisca vitripennis, (Hemiptera: Cicadellidae) is poorly understood. Diapause is better known from other, non hemipteran insects. We used oligonucleotide microarrays to address the specificities of transcriptional responses of ad...

  5. The transcriptional repressor DREAM is involved in thyroid gene expression

    SciTech Connect

    D'Andrea, Barbara; Di Palma, Tina; Mascia, Anna; Motti, Maria Letizia; Viglietto, Giuseppe; Nitsch, Lucio; Zannini, Mariastella . E-mail: stella@szn.it

    2005-04-15

    Downstream regulatory element antagonistic modulator (DREAM) was originally identified in neuroendocrine cells as a calcium-binding protein that specifically binds to downstream regulatory elements (DRE) on DNA, and represses transcription of its target genes. To explore the possibility that DREAM may regulate the endocrine activity of the thyroid gland, we analyzed its mRNA expression in undifferentiated and differentiated thyroid cells. We demonstrated that DREAM is expressed in the normal thyroid tissue as well as in differentiated thyroid cells in culture while it is absent in FRT poorly differentiated cells. In the present work, we also show that DREAM specifically binds to DRE sites identified in the 5' untranslated region (UTR) of the thyroid-specific transcription factors Pax8 and TTF-2/FoxE1 in a calcium-dependent manner. By gel retardation assays we demonstrated that thapsigargin treatment increases the binding of DREAM to the DRE sequences present in Pax8 and TTF-2/Foxe1 5' UTRs, and this correlates with a significant reduction of the expression of these genes. Interestingly, in poorly differentiated thyroid cells overexpression of exogenous DREAM strongly inhibits Pax8 expression. Moreover, we provide evidence that a mutated form of DREAM unable to bind Ca{sup 2+} interferes with thyroid cell proliferation. Therefore, we propose that in thyroid cells DREAM is a mediator of the calcium-signaling pathway and it is involved in the regulation of thyroid cell function.

  6. Genomewide identification of genes under directional selection: gene transcription Q(ST) scan in diverging Atlantic salmon subpopulations.

    PubMed

    Roberge, C; Guderley, H; Bernatchez, L

    2007-10-01

    Evolutionary genomics has benefited from methods that allow identifying evolutionarily important genomic regions on a genomewide scale, including genome scans and QTL mapping. Recently, genomewide scanning by means of microarrays has permitted assessing gene transcription differences among species or populations. However, the identification of differentially transcribed genes does not in itself suffice to measure the role of selection in driving evolutionary changes in gene transcription. Here, we propose and apply a "transcriptome scan" approach to investigating the role of selection in shaping differential profiles of gene transcription among populations. We compared the genomewide transcription levels between two Atlantic salmon subpopulations that have been diverging for only six generations. Following assessment of normality and unimodality on a gene-per-gene basis, the additive genetic basis of gene transcription was estimated using the animal model. Gene transcription h(2) estimates were significant for 1044 (16%) of all detected cDNA clones. In an approach analogous to that of genome scans, we used the distribution of the Q(ST) values estimated from intra- and intersubpopulation additive genetic components of the transcription profiles to identify 16 outlier genes (average Q(ST) estimate = 0.11) whose transcription levels are likely to have evolved under the influence of directional selection within six generations only. Overall, this study contributes both empirically and methodologically to the quantitative genetic exploration of gene transcription data. PMID:17720934

  7. Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products. Progress report, June 1, 1991--May 31, 1992

    SciTech Connect

    Kuchka, M.R.

    1992-05-01

    The following is a review of research accomplished in the first two years of funding for the above mentioned project. The work performed is a molecular characterization of nuclear mutants of Chlamydomonas reinhardtii which are deficient in different stages in the post-transcriptional expression of a single chloroplast encoded polypeptide, the D2 protein of Photosystem II. Our long-term goals are to understand the molecular mechanisms by which nuclear gene products affect the expression of chloroplast genes. Specifically, we which to understand how specific nuclear gene products affect the turnover rate of the D2 encoding mRNA (psbD), how other nuclear encoded factors work to promote the translation of psbD mRNA and/or stabilize the D2 protein, and what the role of the D2 protein itself is in Photosystem II assembly and in the control of expression of other chloroplast genes. This progress report will be organized into four major sections concerning (I) The characterization of nuclear mutants affected in D2 translation/turnover, (II) The study of trans-acting factors which associate with the 5{prime} end of the psbD mRNA, (III) In vitro mutagenesis of the psbD gene, and (IV) Additional studies.

  8. Alternative promoters are used for genes within maize chloroplast polycistronic transcription units.

    PubMed

    Haley, J; Bogorad, L

    1990-04-01

    Many chloroplast genes are co-transcribed in polycistronic transcription units that give rise to numerous overlapping RNAs, but the significance of this pattern of transcript accumulation is not understood. An analysis of the transcripts of the adjacent and divergent maize psbE-psbF-psbL-ORF40 and ORF31-petE-ORF42 gene clusters indicates that transcription initiation at alternative promoters contributes to the generation of overlapping RNAs for both clusters. Furthermore, developmentally varying transcript ratios for the ORF31-petE-ORF42 gene cluster are determined at least in part by selective promoter usage. During light-induced plastid maturation, increased levels of primarily monocistronic petE transcripts accumulate from a promoter upstream of the internal petE gene. Dark-predominant and non-light-responsive bi- and tricistronic transcripts result from transcription initiation upstream of ORF31, the proximal gene of the cluster. In addition to the transcriptional overlap within gene clusters, divergent transcription units for the two gene clusters overlap and reciprocal antisense RNAs accumulate. The organization of the transcription units in this region raises the possibility of promoter interdependence or other functional interaction between transcription units. PMID:2152119

  9. Saxitoxin Modulates Immunological Parameters and Gene Transcription in Mytilus chilensis Hemocytes.

    PubMed

    Astuya, Allisson; Carrera, Crisleri; Ulloa, Viviana; Aballay, Ambbar; Núñez-Acuña, Gustavo; Hégaret, Hélène; Gallardo-Escárate, Cristian

    2015-01-01

    Saxitoxin (STX) is a neurotoxin produced by dinoflagellates in diverse species, such as Alexandrium spp., and it causes paralytic shellfish poisoning (PSP) in humans after the ingestion of contaminated shellfish. Recent studies have suggested that the immune functions of bivalves could be affected by harmful algae and/or by their toxins. Herein, hemocytes are the main effector cells of the immune cellular response. In this study, we evaluated the response of hemocytes from the mussel Mytilus chilensis to STX exposure in a primary culture. Cell cultures were characterized according to size and complexity, while reactive oxygen species (ROS) production was evaluated using a dichlorofluorescein diacetate (DCFH-DA) assay. Finally, phagocytic activity was measured using both flow cytometry and fluorescence microscopy assays. Additionally, gene transcription of candidate genes was evaluated by qPCR assays. The results evidenced that exposures to different concentrations of STX (1-100 nM) for 24 h did not affect cell viability, as determined by an MTT assay. However, when hemocytes were exposed for 4 or 16 h to STX (1-100 nM), there was a modulation of phagocytic activity and ROS production. Moreover, hemocytes exposed to 100 nM of STX for 4 or 16 h showed a significant increase in transcript levels of genes encoding for antioxidant enzymes (SOD, CAT), mitochondrial enzymes (COI, COIII, CYTB, ATP6, ND1) and ion channels (K+, Ca2+). Meanwhile, C-type lectin and toll-like receptor genes revealed a bi-phase transcriptional response after 16 and 24-48 h of exposure to STX. These results suggest that STX can negatively affect the immunocompetence of M. chilensis hemocytes, which were capable of responding to STX exposure in vitro by increasing the mRNA levels of antioxidant enzymes. PMID:26154765

  10. Saxitoxin Modulates Immunological Parameters and Gene Transcription in Mytilus chilensis Hemocytes

    PubMed Central

    Astuya, Allisson; Carrera, Crisleri; Ulloa, Viviana; Aballay, Ambbar; Núñez-Acuña, Gustavo; Hégaret, Hélène; Gallardo-Escárate, Cristian

    2015-01-01

    Saxitoxin (STX) is a neurotoxin produced by dinoflagellates in diverse species, such as Alexandrium spp., and it causes paralytic shellfish poisoning (PSP) in humans after the ingestion of contaminated shellfish. Recent studies have suggested that the immune functions of bivalves could be affected by harmful algae and/or by their toxins. Herein, hemocytes are the main effector cells of the immune cellular response. In this study, we evaluated the response of hemocytes from the mussel Mytilus chilensis to STX exposure in a primary culture. Cell cultures were characterized according to size and complexity, while reactive oxygen species (ROS) production was evaluated using a dichlorofluorescein diacetate (DCFH-DA) assay. Finally, phagocytic activity was measured using both flow cytometry and fluorescence microscopy assays. Additionally, gene transcription of candidate genes was evaluated by qPCR assays. The results evidenced that exposures to different concentrations of STX (1–100 nM) for 24 h did not affect cell viability, as determined by an MTT assay. However, when hemocytes were exposed for 4 or 16 h to STX (1–100 nM), there was a modulation of phagocytic activity and ROS production. Moreover, hemocytes exposed to 100 nM of STX for 4 or 16 h showed a significant increase in transcript levels of genes encoding for antioxidant enzymes (SOD, CAT), mitochondrial enzymes (COI, COIII, CYTB, ATP6, ND1) and ion channels (K+, Ca2+). Meanwhile, C-type lectin and toll-like receptor genes revealed a bi-phase transcriptional response after 16 and 24–48 h of exposure to STX. These results suggest that STX can negatively affect the immunocompetence of M. chilensis hemocytes, which were capable of responding to STX exposure in vitro by increasing the mRNA levels of antioxidant enzymes. PMID:26154765

  11. VITELLOGENIN GENE TRANSCRIPTION: A RELATIVE QUANTITATIVE EXPOSURE INDICATOR OF ENVIRONMENTAL ESTROGENS

    EPA Science Inventory

    We report the development of a quantifiable exposure indicator for measuring the presence of environmental estrogens in aquatic systems. Synthetic oligonucleotides, designed specifically for the vitellogenin gene (Vg) transcription product, were used in a Reverse Transcription Po...

  12. Inferring transcriptional gene regulation network of starch metabolism in Arabidopsis thaliana leaves using graphical Gaussian model

    PubMed Central

    2012-01-01

    Background Starch serves as a temporal storage of carbohydrates in plant leaves during day/night cycles. To study transcriptional regulatory modules of this dynamic metabolic process, we conducted gene regulation network analysis based on small-sample inference of graphical Gaussian model (GGM). Results Time-series significant analysis was applied for Arabidopsis leaf transcriptome data to obtain a set of genes that are highly regulated under a diurnal cycle. A total of 1,480 diurnally regulated genes included 21 starch metabolic enzymes, 6 clock-associated genes, and 106 transcription factors (TF). A starch-clock-TF gene regulation network comprising 117 nodes and 266 edges was constructed by GGM from these 133 significant genes that are potentially related to the diurnal control of starch metabolism. From this network, we found that β-amylase 3 (b-amy3: At4g17090), which participates in starch degradation in chloroplast, is the most frequently connected gene (a hub gene). The robustness of gene-to-gene regulatory network was further analyzed by TF binding site prediction and by evaluating global co-expression of TFs and target starch metabolic enzymes. As a result, two TFs, indeterminate domain 5 (AtIDD5: At2g02070) and constans-like (COL: At2g21320), were identified as positive regulators of starch synthase 4 (SS4: At4g18240). The inference model of AtIDD5-dependent positive regulation of SS4 gene expression was experimentally supported by decreased SS4 mRNA accumulation in Atidd5 mutant plants during the light period of both short and long day conditions. COL was also shown to positively control SS4 mRNA accumulation. Furthermore, the knockout of AtIDD5 and COL led to deformation of chloroplast and its contained starch granules. This deformity also affected the number of starch granules per chloroplast, which increased significantly in both knockout mutant lines. Conclusions In this study, we utilized a systematic approach of microarray analysis to discover

  13. Transcriptional analysis of the recA gene of Streptococcus thermophilus

    PubMed Central

    Giliberti, Gabriele; Baccigalupi, Loredana; Cordone, Angelina; Ricca, Ezio; De Felice, Maurilio

    2006-01-01

    Background RecA is a highly conserved prokaryotic protein that not only plays several important roles connected to DNA metabolism but also affects the cell response to various stress conditions. While RecA is highly conserved, the mechanism of transcriptional regulation of its structural gene is less conserved. In Escherichia coli the LexA protein acts as a recA repressor and is able, in response to DNA damage, of RecA-promoted self-cleavage, thus allowing recA transcription. The LexA paradigm, although confirmed in a wide number of cases, is not universally valid. In some cases LexA does not control recA transcription while in other RecA-containing bacteria a LexA homologue is not present. Results We have studied the recA transcriptional regulation in S. thermophilus, a bacterium that does not contain a LexA homologue. We have characterized the promoter region of the gene and observed that its expression is strongly induced by DNA damage. The analysis of deletion mutants and of translational gene fusions showed that a DNA region of 83 base pairs, containg the recA promoter and the transcriptional start site, is sufficient to ensure normal expression of the gene. Unlike LexA of E. coli, the factor controlling recA expression in S. thermophilus acts in a RecA-independent way since recA induction was observed in a strain carrying a recA null mutation. Conclusion In S. thermophilus, as in many other bacteria,recA expression is strongly induced by DNA damage, however, in this organism expression of the gene is controlled by a factor different from those well characterized in other bacteria. A small DNA region extending from 62 base pairs upstream of the recA transcriptional start site to 21 base pairs downstream of it carries all the information needed for normal regulation of the S. thermophilus recA gene. PMID:16972988

  14. Herpes simplex virus type 1 protein IE63 affects the nuclear export of virus intron-containing transcripts.

    PubMed Central

    Phelan, A; Dunlop, J; Clements, J B

    1996-01-01

    Using in situ hybridization labelling methods, we have determined that the herpes simplex virus type 1 immediate-early protein IE63 (ICP27) affects the cellular localization of virus transcripts. Intronless transcripts from the IE63, UL38, and UL44 genes are rapidly exported to and accumulate in the cytoplasm throughout infection, in either the presence or absence of IE63 expression. The intron-containing transcripts from the IE110 and UL15 genes, while initially cytoplasmic, are increasingly retained in the nucleus in distinct clumps as infection proceeds, and the clumps colocalize with the redistributed small nuclear ribonucleoprotein particles. Infections with the IE63 mutant virus 27-lacZ demonstrated that in the absence of IE63 expression, nuclear retention of intron-containing transcripts was lost. The nuclear retention of UL15 transcripts, which demonstrated both nuclear and cytoplasmic label, was not as pronounced as that of the IE110 transcripts, and we propose that this is due to the late expression of UL15. Infections with the mutant virus 110C1, in which both introns of IE110 have been precisely removed (R.D. Everett, J. Gen. Virol. 72:651-659, 1991), demonstrated IE110 transcripts in both the nucleus and the cytoplasm; thus, exon definition sequences which regulate viral RNA transport are present in the IE110 transcript. By in situ hybridization a stable population of polyadenylated RNAs was found to accumulate in the nucleus in spots, most of which were separate from the small nuclear ribonucleoprotein particle clumps. The IE63 protein has an involvement, either direct or indirect, in the regulation of nucleocytoplasmic transport of viral transcripts, a function which contrasts with the recently proposed role of herpes simplex virus type 1 Us11 in promoting the nuclear export of partially spliced or unspliced transcripts (J.-J. Diaz, M. Duc Dodon, N. Schaerer-Uthurraly, D. Simonin, K. Kindbeiter, L. Gazzolo, and J.-J. Madjar, Nature [London] 379

  15. GOLDEN 2-LIKE transcription factors for chloroplast development affect ozone tolerance through the regulation of stomatal movement.

    PubMed

    Nagatoshi, Yukari; Mitsuda, Nobutaka; Hayashi, Maki; Inoue, Shin-Ichiro; Okuma, Eiji; Kubo, Akihiro; Murata, Yoshiyuki; Seo, Mitsunori; Saji, Hikaru; Kinoshita, Toshinori; Ohme-Takagi, Masaru

    2016-04-12

    Stomatal movements regulate gas exchange, thus directly affecting the efficiency of photosynthesis and the sensitivity of plants to air pollutants such as ozone. The GARP family transcription factors GOLDEN 2-LIKE1 (GLK1) and GLK2 have known functions in chloroplast development. Here, we show that Arabidopsis thaliana (A. thaliana) plants expressing the chimeric repressors for GLK1 and -2 (GLK1/2-SRDX) exhibited a closed-stomata phenotype and strong tolerance to ozone. By contrast, plants that overexpress GLK1/2 exhibited an open-stomata phenotype and higher sensitivity to ozone. The plants expressing GLK1-SRDX had reduced expression of the genes for inwardly rectifying K(+) (K(+) in) channels and reduced K(+) in channel activity. Abscisic acid treatment did not affect the stomatal phenotype of 35S:GLK1/2-SRDX plants or the transcriptional activity for K(+) in channel gene, indicating that GLK1/2 act independently of abscisic acid signaling. Our results indicate that GLK1/2 positively regulate the expression of genes for K(+) in channels and promote stomatal opening. Because the chimeric GLK1-SRDX repressor driven by a guard cell-specific promoter induced a closed-stomata phenotype without affecting chloroplast development in mesophyll cells, modulating GLK1/2 activity may provide an effective tool to control stomatal movements and thus to confer resistance to air pollutants. PMID:27035938

  16. The transcript elongation factor SPT4/SPT5 is involved in auxin-related gene expression in Arabidopsis

    PubMed Central

    Dürr, Julius; Lolas, Ihab B.; Sørensen, Brian B.; Schubert, Veit; Houben, Andreas; Melzer, Michael; Deutzmann, Rainer; Grasser, Marion; Grasser, Klaus D.

    2014-01-01

    The heterodimeric complex SPT4/SPT5 is a transcript elongation factor (TEF) that directly interacts with RNA polymerase II (RNAPII) to regulate messenger RNA synthesis in the chromatin context. We provide biochemical evidence that in Arabidopsis, SPT4 occurs in a complex with SPT5, demonstrating that the SPT4/SPT5 complex is conserved in plants. Each subunit is encoded by two genes SPT4-1/2 and SPT5-1/2. A mutant affected in the tissue-specifically expressed SPT5-1 is viable, whereas inactivation of the generally expressed SPT5-2 is homozygous lethal. RNAi-mediated downregulation of SPT4 decreases cell proliferation and causes growth reduction and developmental defects. These plants display especially auxin signalling phenotypes. Consistently, auxin-related genes, most strikingly AUX/IAA genes, are downregulated in SPT4–RNAi plants that exhibit an enhanced auxin response. In Arabidopsis nuclei, SPT5 clearly localizes to the transcriptionally active euchromatin, and essentially co-localizes with transcribing RNAPII. Typical for TEFs, SPT5 is found over the entire transcription unit of RNAPII-transcribed genes. In SPT4–RNAi plants, elevated levels of RNAPII and SPT5 are detected within transcribed regions (including those of downregulated genes), indicating transcript elongation defects in these plants. Therefore, SPT4/SPT5 acts as a TEF in Arabidopsis, regulating transcription during the elongation stage with particular impact on the expression of certain auxin-related genes. PMID:24497194

  17. IKK{epsilon} modulates RSV-induced NF-{kappa}B-dependent gene transcription

    SciTech Connect

    Bao Xiaoyong; Indukuri, Hemalatha; Liu Tianshuang; Liao Suiling; Tian, Bing; Brasier, Allan R.; Garofalo, Roberto P.; Casola, Antonella

    2010-12-20

    Respiratory syncytial virus (RSV), a negative-strand RNA virus, is the most common cause of epidemic respiratory disease in infants and young children. RSV infection of airway epithelial cells induces the expression of immune/inflammatory genes through the activation of a subset of transcription factors, including Nuclear Factor-{kappa}B (NF-{kappa}B). In this study we have investigated the role of the non canonical I{kappa}B kinase (IKK){epsilon} in modulating RSV-induced NF-{kappa}B activation. Our results show that inhibition of IKK{epsilon} activation results in significant impairment of viral-induced NF-{kappa}B-dependent gene expression, through a reduction in NF-{kappa}B transcriptional activity, without changes in nuclear translocation or DNA-binding activity. Absence of IKK{epsilon} results in a significant decrease of RSV-induced NF-{kappa}B phosphorylation on serine 536, a post-translational modification important for RSV-induced NF-{kappa}B-dependent gene expression, known to regulate NF-{kappa}B transcriptional activity without affecting nuclear translocation. This study identifies a novel mechanism by which IKK{epsilon} regulates viral-induced cellular signaling.

  18. EGR1 regulates hepatic clock gene amplitude by activating Per1 transcription

    PubMed Central

    Tao, Weiwei; Wu, Jing; Zhang, Qian; Lai, Shan-Shan; Jiang, Shan; Jiang, Chen; Xu, Ying; Xue, Bin; Du, Jie; Li, Chao-Jun

    2015-01-01

    The mammalian clock system is composed of a master clock and peripheral clocks. At the molecular level, the rhythm-generating mechanism is controlled by a molecular clock composed of positive and negative feedback loops. However, the underlying mechanisms for molecular clock regulation that affect circadian clock function remain unclear. Here, we show that Egr1 (early growth response 1), an early growth response gene, is expressed in mouse liver in a circadian manner. Consistently, Egr1 is transactivated by the CLOCK/BMAL1 heterodimer through a conserved E-box response element. In hepatocytes, EGR1 regulates the transcription of several core clock genes, including Bmal1, Per1, Per2, Rev-erbα and Rev-erbβ, and the rhythm amplitude of their expression is dependent on EGR1’s transcriptional function. Further mechanistic studies indicated that EGR1 binds to the proximal region of the Per1 promoter to activate its transcription directly. When the peripheral clock is altered by light or feeding behavior transposition in Egr1-deficient mice, the expression phase of hepatic clock genes shifts normally, but the amplitude is also altered. Our data reveal a critical role for EGR1 in the regulation of hepatic clock circuitry, which may contribute to the rhythm stability of peripheral clock oscillators. PMID:26471974

  19. Sucrose regulation of ADP-glucose pyrophosphorylase subunit genes transcript levels in leaves and fruits

    NASA Technical Reports Server (NTRS)

    Li, Xiangyang; Xing, Jinpeng; Gianfagna, Thomas J.; Janes, Harry W.

    2002-01-01

    ADP-glucose pyrophosphorylase (AGPase, EC2.7.7.27) is a key regulatory enzyme in starch biosynthesis. The enzyme is a heterotetramer with two S and two B subunits. In tomato, there are three multiple forms of the S subunit gene. Agp S1, S2 and B are highly expressed in fruit from 10 to 25 days after anthesis. Agp S3 is only weakly expressed in fruit. Sucrose significantly elevates expression of Agp S1, S2 and B in both leaves and fruits. Agp S1 exhibits the highest degree of regulation by sucrose. In fact, sucrose may be required for Agp S1 expression. For excised leaves incubated in water, no transcripts for Agp S1 could be detected in the absence of sucrose, whereas it took up to 16 h in water before transcripts were no longer detectable for Agp S2 and B. Neither Agp S3 nor the tubulin gene is affected by sucrose, demonstrating that this response is specifically regulated by a carbohydrate metabolic signal, and is not due to a general increase in metabolism caused by sucrose treatment. Truncated versions of the promoter for Agp S1 indicate that a specific region 1.3-3.0 kb upstream from the transcription site is responsible for sucrose sensitivity. This region of the S1 promoter contains several cis-acting elements present in the promoters of other genes that are also regulated by sucrose. c2002 Elsevier Science Ireland Ltd. All rights reserved.

  20. IKKε MODULATES RSV-INDUCED NF-κB-DEPENDENT GENE TRANSCRIPTION

    PubMed Central

    Bao, Xiaoyong; Indukuri, Hemalatha; Liu, Tianshuang; Liao, Sui-Ling; Tian, Bing; Brasier, Allan R.; Garofalo, Roberto P.; Casola, Antonella

    2010-01-01

    Respiratory syncytial virus (RSV), a negative-strand RNA virus, is the most common cause of epidemic respiratory disease in infants and young children. RSV infection of airway epithelial cells induces the expression of immune/inflammatory genes through the activation of a subset of transcription factors, including Nuclear Factor-κB (NF-κB). In this study we have investigated the role of the non canonical IκB kinase (IKK)ε in modulating RSV-induced NF-κB activation. Our results show that inhibition of IKKε activation results in significant impairment of viral-induced NF-κB-dependent gene expression, through a reduction in NF-κB transcriptional activity, without changes in nuclear translocation or DNA-binding activity. Absence of IKKε results in a significant decrease of RSV-induced NF-κB phosphorylation on serine 536, a post-translational modification important for RSV-induced NF-κB-dependent gene expression, known to regulate NF-κB transcriptional activity without affecting nuclear translocation. This study identifies a novel mechanism by which IKKε regulates viral-induced cellular signaling. PMID:20961594

  1. Kinetics of transcription of infectious laryngotracheitis virus genes.

    PubMed

    Mahmoudian, Alireza; Markham, Philip F; Noormohammadi, Amir H; Browning, Glenn F

    2012-03-01

    The kinetics of expression of only a few genes of infectious laryngotracheitis virus (ILTV) have been determined, using northern blot analysis. We used quantitative reverse transcriptase PCR to examine the kinetics of expression of 74 ILTV genes in LMH cells. ICP4 was the only gene fully expressed in the presence of cycloheximide, and thus classified as immediate-early. The genes most highly expressed early in infection, and thus classified as early, included UL1 (gL), UL2, UL3, UL4, UL5, UL6, UL7, UL8, UL13, UL14, UL19, UL20, UL23 (TK), UL25, UL28, UL29, UL31, UL33, UL34, UL38, UL39, UL40, UL42, UL43, UL44 (gC), UL47, UL48 (α-TIF), UL49, UL54 (ICP27), US3 and US10. ORF A, ORF B, ORF C, ORF E, sORF 4/3, UL[-1], UL0, UL3.5, UL9, UL10 (gM), UL11, UL15a, UL15b, UL18, UL22 (gH), UL24, UL26, UL30, UL32, UL36, UL45, UL49.5 (gN), UL52, US2, US4 (gG), US5 (gJ) and US9 were most highly expressed late in infection and were thus considered late genes. Several genes, including ORF D, UL12, UL17, UL21, UL27 (gB), UL35, UL37, UL41, UL46, UL50, UL51, UL53 (gK), US8 (gE), US6 (gD) and US7 (gI), had features of both early and late genes and were classified as early/late. Our findings suggest transcription from most of ILTV genes is leaky or subject to more complex patterns of regulation than those classically described for herpesviruses. This is the first study examining global expression of ILTV genes and the data provide a basis for future investigations of the pathogenesis of infection with ILTV. PMID:22195977

  2. Single chromosome transcriptional profiling reveals chromosome-level regulation of gene expression

    PubMed Central

    Levesque, Marshall J.; Raj, Arjun

    2013-01-01

    Here we report iceFISH, a multiplex imaging method for measuring gene expression and chromosome structure simultaneously on single chromosomes. We demonstrate that chromosomal translocations can alter transcription chromosome-wide, finding substantial differences in transcriptional frequency between genes located on a translocated chromosome in comparison to the normal chromosome in the same cell. Examination of correlations between genes on a single chromosome revealed a cis chromosome-level transcriptional interaction spanning 14.3 megabases. PMID:23416756

  3. Nucleosome-driven transcription factor binding and gene regulation.

    PubMed

    Ballaré, Cecilia; Castellano, Giancarlo; Gaveglia, Laura; Althammer, Sonja; González-Vallinas, Juan; Eyras, Eduardo; Le Dily, Francois; Zaurin, Roser; Soronellas, Daniel; Vicent, Guillermo P; Beato, Miguel

    2013-01-10

    Elucidating the global function of a transcription factor implies the identification of its target genes and genomic binding sites. The role of chromatin in this context is unclear, but the dominant view is that factors bind preferentially to nucleosome-depleted regions identified as DNaseI-hypersensitive sites (DHS). Here we show by ChIP, MNase, and DNaseI assays followed by deep sequencing that the progesterone receptor (PR) requires nucleosomes for optimal binding and function. In breast cancer cells treated with progestins, we identified 25,000 PR binding sites (PRbs). The majority of these sites encompassed several copies of the hexanucleotide TGTYCY, which is highly abundant in the genome. We found that functional PRbs accumulate around progesterone-induced genes, mainly in enhancers. Most of these sites overlap with DHS but exhibit high nucleosome occupancy. Progestin stimulation results in remodeling of these nucleosomes with displacement of histones H1 and H2A/H2B dimers. Our results strongly suggest that nucleosomes are crucial for PR binding and hormonal gene regulation. PMID:23177737

  4. Transcriptional Targeting in the Airway Using Novel Gene Regulatory Elements

    PubMed Central

    Burnight, Erin R.; Wang, Guoshun; McCray, Paul B.

    2012-01-01

    The delivery of cystic fibrosis transmembrane conductance regulator (CFTR) to airway epithelia is a goal of many gene therapy strategies to treat cystic fibrosis. Because the native regulatory elements of the CFTR are not well characterized, the development of vectors with heterologous promoters of varying strengths and specificity would aid in our selection of optimal reagents for the appropriate expression of the vector-delivered CFTR gene. Here we contrasted the performance of several novel gene-regulatory elements. Based on airway expression analysis, we selected putative regulatory elements from BPIFA1 and WDR65 to investigate. In addition, we selected a human CFTR promoter region (∼ 2 kb upstream of the human CFTR transcription start site) to study. Using feline immunodeficiency virus vectors containing the candidate elements driving firefly luciferase, we transduced murine nasal epithelia in vivo. Luciferase expression persisted for 30 weeks, which was the duration of the experiment. Furthermore, when the nasal epithelium was ablated using the detergent polidocanol, the mice showed a transient loss of luciferase expression that returned 2 weeks after administration, suggesting that our vectors transduced a progenitor cell population. Importantly, the hWDR65 element drove sufficient CFTR expression to correct the anion transport defect in CFTR-null epithelia. These results will guide the development of optimal vectors for sufficient, sustained CFTR expression in airway epithelia. PMID:22447971

  5. Transcriptional regulation of the Arabidopsis thaliana chalcone synthase gene

    SciTech Connect

    Feinbaum, R.L.; Ausubel, F.M.

    1988-05-01

    The authors cloned an Arabiodpsis thaliana chalcone synthase (CHS) gene on the basis of cross-hybridization with a Petroselinum hortense CHS cDNA clone. The protein sequence deduced from the A. thaliana CHS DNA sequence is at least 85% homologous to the CHS sequences from P. hortense, Antirrhinum majus, and Petunia hybrida. Southern blot analysis indicated that CHS is a single-copy gene in A. thaliana. High-intensity light treatment of A. thaliana plants for 24 h caused a 50-fold increase in CHS enzyme activity and an accumulation of visibly detectable levels of anthocyanin pigments in the vegetative structures of these plants. A corresponding increase in the steady-state level of CHS mRNA was detected after high-intensity light treatment for the same period of time. The accumulation of CHS mRNA in response to high-intensity light was due, at least in part, to an increased rate of transcription of the CHS gene as demonstrated by nuclear runoff experiment.

  6. Specific regulation of transcription of the discoidin gene family in Dictyostelium discoideum.

    PubMed Central

    Alexander, S; Shinnick, T M

    1985-01-01

    Dictyostelium discoideum strains that carry the dis mutations fail to express the family of developmentally regulated discoidin lectin genes during morphogenesis. We show here that this absence of discoidin lectin expression is due to the failure to transcribe the discoidin genes. Furthermore, the dis mutations appear to affect only discoidin expression and not the expression of other proteins during development, as assessed by a two-dimensional gel analysis of pulse-labeled proteins and by the accumulation of developmentally regulated enzymes. The dis mutations appear to define trans-acting regulatory loci, the products of which act at the transcriptional level to control specifically the developmental expression of the discoidin gene family. Images PMID:4000124

  7. Pairwise comparisons of ten porcine tissues identify differential transcriptional regulation at the gene, isoform, promoter and transcription start site level

    SciTech Connect

    Farajzadeh, Leila; Hornshøj, Henrik; Momeni, Jamal; Thomsen, Bo; Larsen, Knud; Hedegaard, Jakob; Bendixen, Christian; Madsen, Lone Bruhn

    2013-08-23

    Highlights: •Transcriptome sequencing yielded 223 mill porcine RNA-seq reads, and 59,000 transcribed locations. •Establishment of unique transcription profiles for ten porcine tissues including four brain tissues. •Comparison of transcription profiles at gene, isoform, promoter and transcription start site level. •Highlights a high level of regulation of neuro-related genes at both gene, isoform, and TSS level. •Our results emphasize the pig as a valuable animal model with respect to human biological issues. -- Abstract: The transcriptome is the absolute set of transcripts in a tissue or cell at the time of sampling. In this study RNA-Seq is employed to enable the differential analysis of the transcriptome profile for ten porcine tissues in order to evaluate differences between the tissues at the gene and isoform expression level, together with an analysis of variation in transcription start sites, promoter usage, and splicing. Totally, 223 million RNA fragments were sequenced leading to the identification of 59,930 transcribed gene locations and 290,936 transcript variants using Cufflinks with similarity to approximately 13,899 annotated human genes. Pairwise analysis of tissues for differential expression at the gene level showed that the smallest differences were between tissues originating from the porcine brain. Interestingly, the relative level of differential expression at the isoform level did generally not vary between tissue contrasts. Furthermore, analysis of differential promoter usage between tissues, revealed a proportionally higher variation between cerebellum (CBE) versus frontal cortex and cerebellum versus hypothalamus (HYP) than in the remaining comparisons. In addition, the comparison of differential transcription start sites showed that the number of these sites is generally increased in comparisons including hypothalamus in contrast to other pairwise assessments. A comprehensive analysis of one of the tissue contrasts, i

  8. Transcriptional regulation of the Bacillus subtilis glucitol dehydrogenase gene.

    PubMed Central

    Ye, R; Wong, S L

    1994-01-01

    The regulatory region of the Bacillus subtilis glucitol dehydrogenase (gutB) gene was divided into three subregions: a promoter, an upstream positive regulatory region, and a downstream negative regulatory region. Data from primer extension, deletion, and site-directed mutagenesis analyses were consistent with two possible models for the gutB promoter. It is either a sigma A-type promoter with an unusually short spacer region (15 bp) or a special sigma A promoter which requires only the hexameric -10 sequence for its function. Sequence carrying just the promoter region (from -48 to +6) failed to direct transcription in vivo. An upstream regulatory sequence was essential for glucitol induction. When this sequence was inserted in a high-copy-number plasmid, an effect characteristic of titration of a transcriptional activator was seen. Downstream from the promoter, there is an imperfect, AT-rich inverted repeat sequence. Deletion of this element did not lead to constitutive expression of gutB. However, the induced gutB expression level was enhanced three- to fourfold. Images PMID:8195086

  9. RNAi related mechanisms affect both transcriptional and posttranscriptional transgene silencing in Drosophila.

    PubMed

    Pal-Bhadra, Manika; Bhadra, Utpal; Birchler, James A

    2002-02-01

    Two types of transgene silencing were found for the Alcohol dehydrogenase (Adh) transcription unit. Transcriptional gene silencing (TGS) is Polycomb dependent and occurs when Adh is driven by the white eye color gene promoter. Full-length Adh transgenes are silenced posttranscriptionally at high copy number or by a pulsed increase over a threshold. The posttranscriptional gene silencing (PTGS) exhibits molecular hallmarks typical of RNA interference (RNAi), including the production of 21--25 bp length sense and antisense RNAs homologous to the silenced RNA. Mutations in piwi, which belongs to a gene family with members required for RNAi, block PTGS and one aspect of TGS, indicating a connection between the two types of silencing. PMID:11864605

  10. Dynamic Encounters of Genes and Transcripts with the Nuclear Pore.

    PubMed

    Ben-Yishay, Rakefet; Ashkenazy, Asaf J; Shav-Tal, Yaron

    2016-07-01

    Transcribed mRNA molecules must reach the cytoplasm to undergo translation. Technological developments in imaging have placed mRNAs under the spotlight, allowing the quantitative study of the spatial and temporal dynamics of the nucleocytoplasmic mRNA export process. Here, we discuss studies that have used such experimental approaches to demonstrate that gene tethering at the nuclear pore complex (NPC) regulates mRNA expression, and to characterize mRNA dynamics during transport in real time. The paths taken by mRNAs as they move from their sites of transcription and travel through the nucleoplasm, in between chromatin domains, and finally through the NPC, can now be observed in detail. PMID:27185238

  11. Post-transcriptional gene silencing activity of human GIGYF2.

    PubMed

    Kryszke, Marie-Hélène; Adjeriou, Badia; Liang, Feifei; Chen, Hong; Dautry, François

    2016-07-01

    In mammalian post-transcriptional gene silencing, the Argonaute protein AGO2 indirectly recruits translation inhibitors, deadenylase complexes, and decapping factors to microRNA-targeted mRNAs, thereby repressing mRNA translation and accelerating mRNA decay. However, the exact composition and assembly pathway of the microRNA-induced silencing complex are not completely elucidated. As the GYF domain of human GIGYF2 was shown to bind AGO2 in pulldown experiments, we wondered whether GIGYF2 could be a novel protein component of the microRNA-induced silencing complex. Here we show that full-length GIGYF2 coimmunoprecipitates with AGO2 in human cells, and demonstrate that, upon tethering to a reporter mRNA, GIGYF2 exhibits strong, dose-dependent silencing activity, involving both mRNA destabilization and translational repression. PMID:27157137

  12. Environmental regulation of virulence in group A streptococci: transcription of the gene encoding M protein is stimulated by carbon dioxide.

    PubMed Central

    Caparon, M G; Geist, R T; Perez-Casal, J; Scott, J R

    1992-01-01

    We have found that different atmospheres can have significant effects on the transcription of emm, the gene that encodes M protein, the major virulence factor of the group A streptococcus (Streptococcus pyogenes). Expression of emm was monitored by constructing a transcriptional fusion of the promoter for emm6.1 from S. pyogenes JRS4 to a promoterless chloramphenicol acetyltransferase gene. Transcription, as measured by determining chloramphenicol acetyltransferase specific activity, was stimulated by as much as 25-fold by increased carbon dioxide tension. Expression was greater in the latter stages of growth and was not affected by growth at 30 instead of 37 degrees C. Insertional inactivation of mry, a gene encoding a positive regulator of emm6.1, reduced chloramphenicol acetyltransferase activity below the detectable level. We conclude that expression of emm is influenced by environmental factors and that the level of carbon dioxide is one signal that may influence expression of M protein during infection. Images PMID:1512202

  13. Transcriptional and Posttranscriptional Regulations of the HLA-G Gene

    PubMed Central

    Castelli, Erick C.; Veiga-Castelli, Luciana C.; Yaghi, Layale; Donadi, Eduardo A.

    2014-01-01

    HLA-G has a relevant role in immune response regulation. The overall structure of the HLA-G coding region has been maintained during the evolution process, in which most of its variable sites are synonymous mutations or coincide with introns, preserving major functional HLA-G properties. The HLA-G promoter region is different from the classical class I promoters, mainly because (i) it lacks regulatory responsive elements for IFN-γ and NF-κB, (ii) the proximal promoter region (within 200 bases from the first translated ATG) does not mediate transactivation by the principal HLA class I transactivation mechanisms, and (iii) the presence of identified alternative regulatory elements (heat shock, progesterone and hypoxia-responsive elements) and unidentified responsive elements for IL-10, glucocorticoids, and other transcription factors is evident. At least three variable sites in the 3′ untranslated region have been studied that may influence HLA-G expression by modifying mRNA stability or microRNA binding sites, including the 14-base pair insertion/deletion, +3142C/G and +3187A/G polymorphisms. Other polymorphic sites have been described, but there are no functional studies on them. The HLA-G coding region polymorphisms might influence isoform production and at least two null alleles with premature stop codons have been described. We reviewed the structure of the HLA-G promoter region and its implication in transcriptional gene control, the structure of the HLA-G 3′UTR and the major actors of the posttranscriptional gene control, and, finally, the presence of regulatory elements in the coding region. PMID:24741620

  14. Transcriptional regulation of human UDP-glucuronosyltransferase genes.

    PubMed

    Hu, Dong Gui; Meech, Robyn; McKinnon, Ross A; Mackenzie, Peter I

    2014-11-01

    Glucuronidation is an important metabolic pathway for many small endogenous and exogenous lipophilic compounds, including bilirubin, steroid hormones, bile acids, carcinogens and therapeutic drugs. Glucuronidation is primarily catalyzed by the UDP-glucuronosyltransferase (UGT) 1A and two subfamilies, including nine functional UGT1A enzymes (1A1, 1A3-1A10) and 10 functional UGT2 enzymes (2A1, 2A2, 2A3, 2B4, 2B7, 2B10, 2B11, 2B15, 2B17 and 2B28). Most UGTs are expressed in the liver and this expression relates to the major role of hepatic glucuronidation in systemic clearance of toxic lipophilic compounds. Hepatic glucuronidation activity protects the body from chemical insults and governs the therapeutic efficacy of drugs that are inactivated by UGTs. UGT mRNAs have also been detected in over 20 extrahepatic tissues with a unique complement of UGT mRNAs seen in almost every tissue. This extrahepatic glucuronidation activity helps to maintain homeostasis and hence regulates biological activity of endogenous molecules that are primarily inactivated by UGTs. Deciphering the molecular mechanisms underlying tissue-specific UGT expression has been the subject of a large number of studies over the last two decades. These studies have shown that the constitutive and inducible expression of UGTs is primarily regulated by tissue-specific and ligand-activated transcription factors (TFs) via their binding to cis-regulatory elements (CREs) in UGT promoters and enhancers. This review first briefly summarizes published UGT gene transcriptional studies and the experimental models and tools utilized in these studies, and then describes in detail the TFs and their respective CREs that have been identified in the promoters and/or enhancers of individual UGT genes. PMID:25336387

  15. Human Genes Encoding Transcription Factors and Chromatin-Modifying Proteins Have Low Levels of Promoter Polymorphism: A Study of 1000 Genomes Project Data

    PubMed Central

    Ignatieva, Elena V.; Levitsky, Victor G.; Kolchanov, Nikolay A.

    2015-01-01

    The expression level of each gene is controlled by its regulatory regions, which determine the precise regulation in a tissue-specific manner, according to the developmental stage of the body and the necessity of a response to external stimuli. Nucleotide substitutions in regulatory gene regions may modify the affinity of transcription factors to their specific DNA binding sites, affecting the transcription rates of genes. In our previous research, we found that genes controlling the sensory perception of smell and genes involved in antigen processing and presentation were overrepresented significantly among genes with high SNP contents in their promoter regions. The goal of our study was to reveal functional features of human genes containing extremely small numbers of SNPs in promoter regions. Two functional groups were found to be overrepresented among genes whose promoters did not contain SNPs: (1) genes involved in gene-specific transcription and (2) genes controlling chromatin organization. We revealed that the 5′-regulatory regions of genes encoding transcription factors and chromatin-modifying proteins were characterized by reduced genetic variability. One important exception from this rule refers to genes encoding transcription factors with zinc-coordinating DNA-binding domains (DBDs), which underwent extensive expansion in vertebrates, particularly, in primate evolution. Hence, we obtained new evidence for evolutionary forces shaping variability in 5′-regulatory regions of genes. PMID:26417590

  16. Transcriptional regulation of gilthead seabream bone morphogenetic protein (BMP) 2 gene by bone- and cartilage-related transcription factors.

    PubMed

    Marques, Cátia L; Cancela, M Leonor; Laizé, Vincent

    2016-01-15

    Bone morphogenetic protein (BMP) 2 belongs to the transforming growth factor β (TGFβ) superfamily of cytokines and growth factors. While it plays important roles in embryo morphogenesis and organogenesis, BMP2 is also critical to bone and cartilage formation. Protein structure and function have been remarkably conserved throughout evolution and BMP2 transcription has been proposed to be tightly regulated, although few data is available. In this work we report the cloning and functional analysis of gilthead seabream BMP2 promoter. As in other vertebrates, seabream BMP2 gene has a 5′ non-coding exon, a feature already present in DPP gene, the fruit fly ortholog of vertebrate BMP2 gene, and maintained throughout evolution. In silico analysis of seabream BMP2 promoter revealed several binding sites for bone and cartilage related transcription factors (TFs) and their functionality was evaluated using promoter-luciferase constructions and TF-expressing vectors. Runt-related transcription factor 3 (RUNX3) was shown to negatively regulate BMP2 transcription and combination with the core binding factor β (CBFβ) further reduced transcriptional activity of the promoter. Although to a lesser extent, myocyte enhancer factor 2C (MEF2C) had also a negative effect on the regulation of BMP2 gene transcription, when associated with SRY (sex determining region Y)-box 9 (SOX9b). Finally, v-ets avian erythroblastosis virus E26 oncogene homolog 1 (ETS1) was able to slightly enhance BMP2 transcription. Data reported here provides new insights toward the better understanding of the transcriptional regulation of BMP2 gene in a bone and cartilage context. PMID:26456102

  17. Transcription factor AP-2 regulates human immunodeficiency virus type 1 gene expression.

    PubMed Central

    Perkins, N D; Agranoff, A B; Duckett, C S; Nabel, G J

    1994-01-01

    Human immunodeficiency virus type 1 (HIV-1) gene expression is regulated by an enhancer region composed of multiple potential cis-acting regulatory sites. Here, we describe binding sites for the transcription factor AP-2 in the HIV-1 long terminal repeat which modulate HIV enhancer function. One site is embedded within the two previously described kappa B elements, and a second site is detected further downstream. DNase I footprinting and electrophoretic mobility shift assay experiments demonstrated that AP-2 binds to the site between the kappa B elements. Interestingly, AP-2 and NF-kappa B bind to this region in a mutually exclusive manner. Mutations which disrupt this AP-2-binding site lower basal levels of transcription but do not affect NF-kappa B-mediated induction by tumor necrosis factor alpha in Jurkat T leukemia cells. Images PMID:8084021

  18. Genetic Diversity Affects the Daily Transcriptional Oscillations of Marine Microbial Populations

    PubMed Central

    Shilova, Irina N.; Robidart, Julie C.; DeLong, Edward F.; Zehr, Jonathan P.

    2016-01-01

    Marine microbial communities are genetically diverse but have robust synchronized daily transcriptional patterns at the genus level that are similar across a wide variety of oceanic regions. We developed a microarray-inspired gene-centric approach to resolve transcription of closely-related but distinct strains/ecotypes in high-throughput sequence data. Applying this approach to the existing metatranscriptomics datasets collected from two different oceanic regions, we found unique and variable patterns of transcription by individual taxa within the abundant picocyanobacteria Prochlorococcus and Synechococcus, the alpha Proteobacterium Pelagibacter and the eukaryotic picophytoplankton Ostreococcus. The results demonstrate that marine microbial taxa respond differentially to variability in space and time in the ocean. These intra-genus individual transcriptional patterns underlie whole microbial community responses, and the approach developed here facilitates deeper insights into microbial population dynamics. PMID:26751368

  19. Mammalian Glutaminase Gls2 Gene Encodes Two Functional Alternative Transcripts by a Surrogate Promoter Usage Mechanism

    PubMed Central

    Campos-Sandoval, José A.; Manzanares, Elisa; Lobo, Carolina; Segura, J. A.; Alonso, Francisco J.; Matés, José M.; Márquez, Javier

    2012-01-01

    Background Glutaminase is expressed in most mammalian tissues and cancer cells, but the regulation of its expression is poorly understood. An essential step to accomplish this goal is the characterization of its species- and cell-specific isoenzyme pattern of expression. Our aim was to identify and characterize transcript variants of the mammalian glutaminase Gls2 gene. Methodology/Principal Findings We demonstrate for the first time simultaneous expression of two transcript variants from the Gls2 gene in human, rat and mouse. A combination of RT-PCR, primer-extension analysis, bioinformatics, real-time PCR, in vitro transcription and translation and immunoblot analysis was applied to investigate GLS2 transcripts in mammalian tissues. Short (LGA) and long (GAB) transcript forms were isolated in brain and liver tissue of human, rat and mouse. The short LGA transcript arises by a combination of two mechanisms of transcriptional modulation: alternative transcription initiation and alternative promoter. The LGA variant contains both the transcription start site (TSS) and the alternative promoter in the first intron of the Gls2 gene. The full human LGA transcript has two in-frame ATGs in the first exon, which are missing in orthologous rat and mouse transcripts. In vitro transcription and translation of human LGA yielded two polypeptides of the predicted size, but only the canonical full-length protein displayed catalytic activity. Relative abundance of GAB and LGA transcripts showed marked variations depending on species and tissues analyzed. Conclusions/Significance This is the first report demonstrating expression of alternative transcripts of the mammalian Gls2 gene. Transcriptional mechanisms giving rise to GLS2 variants and isolation of novel GLS2 transcripts in human, rat and mouse are presented. Results were also confirmed at the protein level, where catalytic activity was demonstrated for the human LGA protein. Relative abundance of GAB and LGA transcripts was

  20. Mapping functional transcription factor networks from gene expression data

    PubMed Central

    Haynes, Brian C.; Maier, Ezekiel J.; Kramer, Michael H.; Wang, Patricia I.; Brown, Holly; Brent, Michael R.

    2013-01-01

    A critical step in understanding how a genome functions is determining which transcription factors (TFs) regulate each gene. Accordingly, extensive effort has been devoted to mapping TF networks. In Saccharomyces cerevisiae, protein–DNA interactions have been identified for most TFs by ChIP-chip, and expression profiling has been done on strains deleted for most TFs. These studies revealed that there is little overlap between the genes whose promoters are bound by a TF and those whose expression changes when the TF is deleted, leaving us without a definitive TF network for any eukaryote and without an efficient method for mapping functional TF networks. This paper describes NetProphet, a novel algorithm that improves the efficiency of network mapping from gene expression data. NetProphet exploits a fundamental observation about the nature of TF networks: The response to disrupting or overexpressing a TF is strongest on its direct targets and dissipates rapidly as it propagates through the network. Using S. cerevisiae data, we show that NetProphet can predict thousands of direct, functional regulatory interactions, using only gene expression data. The targets that NetProphet predicts for a TF are at least as likely to have sites matching the TF's binding specificity as the targets implicated by ChIP. Unlike most ChIP targets, the NetProphet targets also show evidence of functional regulation. This suggests a surprising conclusion: The best way to begin mapping direct, functional TF-promoter interactions may not be by measuring binding. We also show that NetProphet yields new insights into the functions of several yeast TFs, including a well-studied TF, Cbf1, and a completely unstudied TF, Eds1. PMID:23636944

  1. Variations in the progranulin gene affect global gene expression in frontotemporal lobar degeneration.

    PubMed

    Chen-Plotkin, Alice S; Geser, Felix; Plotkin, Joshua B; Clark, Chris M; Kwong, Linda K; Yuan, Wuxing; Grossman, Murray; Van Deerlin, Vivianna M; Trojanowski, John Q; Lee, Virginia M-Y

    2008-05-15

    Frontotemporal lobar degeneration is a fatal neurodegenerative disease that results in progressive decline in behavior, executive function and sometimes language. Disease mechanisms remain poorly understood. Recently, however, the DNA- and RNA-binding protein TDP-43 has been identified as the major protein present in the hallmark inclusion bodies of frontotemporal lobar degeneration with ubiquitinated inclusions (FTLD-U), suggesting a role for transcriptional dysregulation in FTLD-U pathophysiology. Using the Affymetrix U133A microarray platform, we profiled global gene expression in both histopathologically affected and unaffected areas of human FTLD-U brains. We then characterized differential gene expression with biological pathway analyses, cluster and principal component analyses, and subgroup analyses based on brain region and progranulin (GRN) gene status. Comparing 17 FTLD-U brains to 11 controls, we identified 414 upregulated and 210 downregulated genes in frontal cortex (P-value < 0.001). Moreover, cluster and principal component analyses revealed that samples with mutations or possibly pathogenic variations in the GRN gene (GRN+, 7/17) had an expression signature that was distinct from both normal controls and FTLD-U samples lacking GRN gene variations (GRN-, 10/17). Within the subgroup of GRN+ FTLD-U, we found >1300 dysregulated genes in frontal cortex (P-value < 0.001), many participating in pathways uniquely dysregulated in the GRN+ cases. Our findings demonstrate a distinct molecular phenotype for GRN+ FTLD-U, not readily apparent on clinical or histopathological examination, suggesting distinct pathophysiological mechanisms for GRN+ and GRN- subtypes of FTLD-U. In addition, these data from a large number of human brains provide a valuable resource for future testing of disease hypotheses. PMID:18223198

  2. Gene transcription and electromagnetic fields. Final progress report

    SciTech Connect

    Henderson, A.S.

    1992-12-31

    Our overall aim is to obtain sufficient information to allow us to ultimately determine whether ELF EM field exposure is an initiating factor in neoplastic transformation and/or if exposure can mimic characteristics of the second-step counterpart in neoplastic disease. This aim is based on our previous findings that levels of some transcripts are increased in cells exposed to EM fields. While the research is basic in nature, the ramifications have bearing on the general safety of exposure to EM fields in industrial and everyday life. A large array of diverse biological effects are reported to occur as the result of exposure to elf EM fields, suggesting that the cell response to EM fields is at a basic level, presumably initiated by molecular and/or biophysical events at the cell membrane. The hypothesized route is a signal transduction pathway involving membrane calcium fluxes. Information flow resulting from signal transduction can mediate the induction of regulatory factors in the cell, and directly affect how transcription is regulated.

  3. Zinc oxide nanoparticles cause inhibition of microbial denitrification by affecting transcriptional regulation and enzyme activity.

    PubMed

    Zheng, Xiong; Su, Yinglong; Chen, Yinguang; Wan, Rui; Liu, Kun; Li, Mu; Yin, Daqiang

    2014-12-01

    Over the past few decades, human activities have accelerated the rates and extents of water eutrophication and global warming through increasing delivery of biologically available nitrogen such as nitrate and large emissions of anthropogenic greenhouse gases. In particular, nitrous oxide (N2O) is one of the most important greenhouse gases, because it has a 300-fold higher global warming potential than carbon dioxide. Microbial denitrification is a major pathway responsible for nitrate removal, and also a dominant source of N2O emissions from terrestrial or aquatic environments. However, whether the release of zinc oxide nanoparticles (ZnO NPs) into the environment affects microbial denitrification is largely unknown. Here we show that the presence of ZnO NPs lead to great increases in nitrate delivery (9.8-fold higher) and N2O emissions (350- and 174-fold higher in the gas and liquid phases, respectively). Our data further reveal that ZnO NPs significantly change the transcriptional regulations of glycolysis and polyhydroxybutyrate synthesis, which causes the decrease in reducing powers available for the reduction of nitrate and N2O. Moreover, ZnO NPs substantially inhibit the gene expressions and catalytic activities of key denitrifying enzymes. These negative effects of ZnO NPs on microbial denitrification finally cause lower nitrate removal and higher N2O emissions, which is likely to exacerbate water eutrophication and global warming. PMID:25384038

  4. Accurate Gene Expression-Based Biodosimetry Using a Minimal Set of Human Gene Transcripts

    SciTech Connect

    Tucker, James D.; Joiner, Michael C.; Thomas, Robert A.; Grever, William E.; Bakhmutsky, Marina V.; Chinkhota, Chantelle N.; Smolinski, Joseph M.; Divine, George W.; Auner, Gregory W.

    2014-03-15

    Purpose: Rapid and reliable methods for conducting biological dosimetry are a necessity in the event of a large-scale nuclear event. Conventional biodosimetry methods lack the speed, portability, ease of use, and low cost required for triaging numerous victims. Here we address this need by showing that polymerase chain reaction (PCR) on a small number of gene transcripts can provide accurate and rapid dosimetry. The low cost and relative ease of PCR compared with existing dosimetry methods suggest that this approach may be useful in mass-casualty triage situations. Methods and Materials: Human peripheral blood from 60 adult donors was acutely exposed to cobalt-60 gamma rays at doses of 0 (control) to 10 Gy. mRNA expression levels of 121 selected genes were obtained 0.5, 1, and 2 days after exposure by reverse-transcriptase real-time PCR. Optimal dosimetry at each time point was obtained by stepwise regression of dose received against individual gene transcript expression levels. Results: Only 3 to 4 different gene transcripts, ASTN2, CDKN1A, GDF15, and ATM, are needed to explain ≥0.87 of the variance (R{sup 2}). Receiver-operator characteristics, a measure of sensitivity and specificity, of 0.98 for these statistical models were achieved at each time point. Conclusions: The actual and predicted radiation doses agree very closely up to 6 Gy. Dosimetry at 8 and 10 Gy shows some effect of saturation, thereby slightly diminishing the ability to quantify higher exposures. Analyses of these gene transcripts may be advantageous for use in a field-portable device designed to assess exposures in mass casualty situations or in clinical radiation emergencies.

  5. Precision control of recombinant gene transcription for CHO cell synthetic biology.

    PubMed

    Brown, Adam J; James, David C

    2016-01-01

    The next generation of mammalian cell factories for biopharmaceutical production will be genetically engineered to possess both generic and product-specific manufacturing capabilities that may not exist naturally. Introduction of entirely new combinations of synthetic functions (e.g. novel metabolic or stress-response pathways), and retro-engineering of existing functional cell modules will drive disruptive change in cellular manufacturing performance. However, before we can apply the core concepts underpinning synthetic biology (design, build, test) to CHO cell engineering we must first develop practical and robust enabling technologies. Fundamentally, we will require the ability to precisely control the relative stoichiometry of numerous functional components we simultaneously introduce into the host cell factory. In this review we discuss how this can be achieved by design of engineered promoters that enable concerted control of recombinant gene transcription. We describe the specific mechanisms of transcriptional regulation that affect promoter function during bioproduction processes, and detail the highly-specific promoter design criteria that are required in the context of CHO cell engineering. The relative applicability of diverse promoter development strategies are discussed, including re-engineering of natural sequences, design of synthetic transcription factor-based systems, and construction of synthetic promoters. This review highlights the potential of promoter engineering to achieve precision transcriptional control for CHO cell synthetic biology. PMID:26721629

  6. Integrated Genomic and Transcriptional Profiling Identifies Chromosomal Loci with Altered Gene Expression in Cervical Cancer

    PubMed Central

    Wilting, Saskia M.; de Wilde, Jillian; Meijer, Chris J. L. M.; Berkhof, Johannes; Yi, Yajun; van Wieringen, Wessel N.; Braakhuis, Boudewijn J. M.; Meijer, Gerrit A.; Ylstra, Bauke; Snijders, Peter J. F.; Steenbergen, Renske D. M.

    2009-01-01

    For a better understanding of the consequences of recurrent chromosomal alterations in cervical carcinomas, we integrated genome-wide chromosomal and transcriptional profiles of 10 squamous cell carcinomas (SCCs), 5 adenocarcinomas (AdCAs) and 6 normal controls. Previous genomic profiling showed that gains at chromosome arms 1q, 3q, and 20q as well as losses at 8q, 10q, 11q, and 13q were common in cervical carcinomas. Altered regions spanned multiple megabases, and the extent to which expression of genes located there is affected remains unclear. Expression analysis of these previously chromosomally profiled carcinomas yielded 83 genes with significantly differential expression between carcinomas and normal epithelium. Application of differential gene locus mapping (DIGMAP) analysis and the array CGH expression integration tool (ACE-it) identified hotspots within large chromosomal alterations in which gene expression was altered as well. Chromosomal gains of the long arms of chromosome 1, 3, and 20 resulted in increased expression of genes located at 1q32.1-32.2, 3q13.32-23, 3q26.32-27.3, and 20q11.21-13.33, whereas a chromosomal loss of 11q22.3-25 was related to decreased expression of genes located in this region. Overexpression of DTX3L, PIK3R4, ATP2C1, and SLC25A36, all located at 3q21.1-23 and identified by DIGMAP, ACE-it or both, was confirmed in an independent validation sample set consisting of 12 SCCs and 13 normal ectocervical samples. In conclusion, integrated chromosomal and transcriptional profiling identified chromosomal hotspots at 1q, 3q, 11q, and 20q with altered gene expression within large commonly altered chromosomal regions in cervical cancer. PMID:18618715

  7. Understanding the Role of Housekeeping and Stress-Related Genes in Transcription-Regulatory Networks

    NASA Astrophysics Data System (ADS)

    Heath, Allison; Kavraki, Lydia; Balázsi, Gábor

    2008-03-01

    Despite the increasing number of completely sequenced genomes, much remains to be learned about how living cells process environmental information and respond to changes in their surroundings. Accumulating evidence indicates that eukaryotic and prokaryotic genes can be classified in two distinct categories that we will call class I and class II. Class I genes are housekeeping genes, often characterized by stable, noise resistant expression levels. In contrast, class II genes are stress-related genes and often have noisy, unstable expression levels. In this work we analyze the large scale transcription-regulatory networks (TRN) of E. coli and S. cerevisiae and preliminary data on H. sapien. We find that stable, housekeeping genes (class I) are preferentially utilized as transcriptional inputs while stress related, unstable genes (class II) are utilized as transcriptional integrators. This might be the result of convergent evolution that placed the appropriate genes in the appropriate locations within transcriptional networks according to some fundamental principles that govern cellular information processing.

  8. E proteins are required to activate germline transcription of the TCR Vbeta8.2 gene.

    PubMed

    Jia, Jingquan; Dai, Meifang; Zhuang, Yuan

    2008-10-01

    Each TCR Vbeta gene is regulated by an individual Vbeta promoter, which becomes active prior to V(D) J recombination and drives germline transcription. It has been shown that Vbeta gene locus activation and recombination are dependent on the Vbeta promoter. However, transcription factors that regulate Vbeta germline transcription remain largely undefined. A major challenge in studying Vbeta gene germline transcription is the quantitative assessment of relatively low-level transcripts in T-cell progenitors. Here we used the established Vbeta8.2(CD2) knock-in mouse model to assess functions of E-protein transcription factors in Vbeta8.2 germline transcription. We show that E proteins are required for the activation but not the maintenance of the Vbeta8.2 germline transcription during thymocyte development. The activation of Vbeta8.2 germline transcription depends more on the E proteins encoded by the E2A gene than by the HEB gene. We further show that IL-7 receptor (IL-7R)-mediated signals are essential for Vbeta8.2 germline transcription. We provide evidence that IL-7R expression is only partially controlled by E2A, suggesting a role for E2A in driving Vbeta8.2 germline transcription independent of IL-7R activation. PMID:18958875

  9. Stochastic Gene Expression in Networks of Post-transcriptional Regulators

    NASA Astrophysics Data System (ADS)

    Baker, Charles; Jia, Tao; Pendar, Hodjat; Kulkarni, Rahul

    2012-02-01

    Post-transcriptional regulators, such as small RNAs and microRNAs, are critical elements of diverse cellular pathways. It has been postulated that, in several important cases, the role of these regulators is to to modulate the noise in gene expression for the regulated target. Correspondingly, general stochastic models have been developed, and results obtained, for the case in which a single sRNA regulates a single mRNA target. We generalize these results to networks containing a single mRNA regulated by multiple sRNAs and to networks containing multiple mRNAs regulated by a single sRNA. For these systems, we obtain exact expressions relating the mean levels of the sRNAs to the mean levels of the mRNAs. Additionally, we consider the convergence of the original model to an approximate model which considers sRNA concentrations to be high; for the latter model we derive an analytic form for the generating function of the protein distribution. Finally, we discuss potential experimental protocols which, in combination with the derived results, can be used to infer the underlying gene expression parameters.

  10. Identification, Phylogeny, and Transcript of Chitinase Family Genes in Sugarcane

    PubMed Central

    Su, Yachun; Xu, Liping; Wang, Shanshan; Wang, Zhuqing; Yang, Yuting; Chen, Yun; Que, Youxiong

    2015-01-01

    Chitinases are pathogensis-related proteins, which play an important role in plant defense mechanisms. The role of the sugarcane chitinase family genes remains unclear due to the highly heterozygous and aneuploidy chromosome genetic background of sugarcane. Ten differentially expressed chitinase genes (belonging to class I~VII) were obtained from RNA-seq analysis of both incompatible and compatible sugarcane genotypes during Sporisorium scitamineum challenge. Their structural properties and expression patterns were analyzed. Seven chitinases (ScChiI1, ScChiI2, ScChiI3, ScChiIII1, ScChiIII2, ScChiIV1 and ScChiVI1) showed more positive with early response and maintained increased transcripts in the incompatible interaction than those in the compatible one. Three (ScChiII1, ScChiV1 and ScChiVII1) seemed to have no significant difference in expression patterns between incompatible and compatible interactions. The ten chitinases were expressed differentially in response to hormone treatment as well as having distinct tissue specificity. ScChiI1, ScChiIV1 and ScChiVII1 were induced by various abiotic stresses (NaCl, CuCl2, PEG and 4 °C) and their involvement in plant immunity was demonstrated by over-expression in Nicotiana benthamiana. The results suggest that sugarcane chitinase family exhibit differential responses to biotic and abiotic stress, providing new insights into their function. PMID:26035173

  11. Mutations in the CRE pocket of bacterial RNA polymerase affect multiple steps of transcription

    PubMed Central

    Petushkov, Ivan; Pupov, Danil; Bass, Irina; Kulbachinskiy, Andrey

    2015-01-01

    During transcription, the catalytic core of RNA polymerase (RNAP) must interact with the DNA template with low-sequence specificity to ensure efficient enzyme translocation and RNA extension. Unexpectedly, recent structural studies of bacterial promoter complexes revealed specific interactions between the nontemplate DNA strand at the downstream edge of the transcription bubble (CRE, core recognition element) and a protein pocket formed by core RNAP (CRE pocket). We investigated the roles of these interactions in transcription by analyzing point amino acid substitutions and deletions in Escherichia coli RNAP. The mutations affected multiple steps of transcription, including promoter recognition, RNA elongation and termination. In particular, we showed that interactions of the CRE pocket with a nontemplate guanine immediately downstream of the active center stimulate RNA-hairpin-dependent transcription pausing but not other types of pausing. Thus, conformational changes of the elongation complex induced by nascent RNA can modulate CRE effects on transcription. The results highlight the roles of specific core RNAP–DNA interactions at different steps of RNA synthesis and suggest their importance for transcription regulation in various organisms. PMID:25990734

  12. Understanding Transcription Factor Regulation by Integrating Gene Expression and DNase I Hypersensitive Sites

    PubMed Central

    Wang, Guohua; Wang, Fang; Huang, Qian; Li, Yu; Liu, Yunlong; Wang, Yadong

    2015-01-01

    Transcription factors are proteins that bind to DNA sequences to regulate gene transcription. The transcription factor binding sites are short DNA sequences (5–20 bp long) specifically bound by one or more transcription factors. The identification of transcription factor binding sites and prediction of their function continue to be challenging problems in computational biology. In this study, by integrating the DNase I hypersensitive sites with known position weight matrices in the TRANSFAC database, the transcription factor binding sites in gene regulatory region are identified. Based on the global gene expression patterns in cervical cancer HeLaS3 cell and HelaS3-ifnα4h cell (interferon treatment on HeLaS3 cell for 4 hours), we present a model-based computational approach to predict a set of transcription factors that potentially cause such differential gene expression. Significantly, 6 out 10 predicted functional factors, including IRF, IRF-2, IRF-9, IRF-1 and IRF-3, ICSBP, belong to interferon regulatory factor family and upregulate the gene expression levels responding to the interferon treatment. Another factor, ISGF-3, is also a transcriptional activator induced by interferon alpha. Using the different transcription factor binding sites selected criteria, the prediction result of our model is consistent. Our model demonstrated the potential to computationally identify the functional transcription factors in gene regulation. PMID:26425553

  13. Network and pathway analysis of microRNAs, transcription factors, target genes and host genes in human glioma

    PubMed Central

    ZHANG, YING; ZHAO, SHISHUN; XU, ZHIWEN

    2016-01-01

    To date, there has been rapid development with regard to gene and microRNA (miR/miRNA) research in gliomas. However, the regulatory mechanisms of the associated genes and miRNAs remain unclear. In the present study, the genes, miRNAs and transcription factors (TFs) were considered as elements in the regulatory network, and focus was placed on the associations between TFs and miRNAs, miRNAs and target genes, and miRNAs and host genes. In order to show the regulatory correlation clearly, all the elements were investigated and three regulatory networks, namely the differentially-expressed, related and global networks, were constructed. Certain important pathways were highlighted, with analysis of the similarities and differences among the networks. Next, the upstream and downstream elements of differentially-expressed genes, miRNAs and predicted TFs were listed. The most notable aspect of the present study was the three levels of network, particularly the differentially-expressed network, since the differentially-expressed associations that these networks provide appear at the initial stages of cancers such as glioma. If the states of the differentially-expressed associations can be adjusted to the normal state via alterations in regulatory associations, which were also recorded in the study networks and tables, it is likely that cancer can be regulated or even avoided. In the present study, the differentially-expressed network illuminated the pathogenesis of glioma; for example, a TF can regulate one or more miRNAs, and a target gene can be targeted by one or more miRNAs. Therefore, the host genes and target genes, the host genes and TFs, and the target genes and TFs indirectly affect each other through miRNAs. The association also exists between TFs and TFs, target genes and target genes, and host genes and host genes. The present study also demonstrated self-adaption associations and circle-regulations. The related network further described the regulatory mechanism

  14. Post-transcriptional gene regulation in the biology and virulence of Candida albicans.

    PubMed

    Verma-Gaur, Jiyoti; Traven, Ana

    2016-06-01

    In the human fungal pathogen Candida albicans, remodelling of gene expression drives host adaptation and virulence. Recent studies revealed that in addition to transcription, post-transcriptional mRNA control plays important roles in virulence-related pathways. Hyphal morphogenesis, biofilm formation, stress responses, antifungal drug susceptibility and virulence in animal models require post-transcriptional regulators. This includes RNA binding proteins that control mRNA localization, decay and translation, as well as the cytoplasmic mRNA decay pathway. Comprehensive understanding of how modulation of gene expression networks drives C. albicans virulence will necessitate integration of our knowledge on transcriptional and post-transcriptional mRNA control. PMID:26999710

  15. PTM-Switchboard--a database of posttranslational modifications of transcription factors, the mediating enzymes and target genes.

    PubMed

    Everett, Logan; Vo, Antony; Hannenhalli, Sridhar

    2009-01-01

    Gene transcription is largely regulated by sequence-specific transcription factors (TFs). The TF activity is significantly regulated by its posttranslational modifications (PTMs). TF-PTMs serve as 'molecular switchboards' that map multiple upstream signaling events, in response to various environmental perturbations, to the downstream transcriptional events. While many instances of TF-PTMs and their effect on gene regulation have been experimentally determined, a systematic meta-analysis or a quantitative model-based investigation of this process has not been undertaken. A prerequisite to such analyses is a database of known instances of TF-PTMs affecting transcriptional regulation. The PTM-Switchboard database meets this need by cataloging such instances in the model system Saccharomyces cerevisiae. The database stores triplets of genes such that the ability of one gene (TF) to regulate a target gene is dependent on one or more PTMs catalyzed by a third gene (modifying enzyme). The database currently includes a large sample of experimentally characterized instances curated from the literature. In addition to providing a framework for searching and analyzing the data, the database will serve to benchmark computational methods. In the future, the database will be expanded to mammalian organisms, and will also include triplets predicted from computational approaches. The database can be accessed at http://cagr.pcbi.upenn.edu/PTMswitchboard. PMID:18927104

  16. Regulatory network of microRNAs, target genes, transcription factors and host genes in endometrial cancer.

    PubMed

    Xue, Lu-Chen; Xu, Zhi-Wen; Wang, Kun-Hao; Wang, Ning; Zhang, Xiao-Xu; Wang, Shang

    2015-01-01

    Genes and microRNAs (miRNAs) have important roles in human oncology. However, most of the biological factors are reported in disperse form which makes it hard to discover the pathology. In this study, genes and miRNAs involved in human endometrial cancer(EC) were collected and formed into regulatory networks following their interactive relations, including miRNAs targeting genes, transcription factors (TFs) regulating miRNAs and miRNAs included in their host genes. Networks are constructed hierarchically at three levels: differentially expressed, related and global. Among the three, the differentially expressed network is the most important and fundamental network that contains the key genes and miRNAs in EC. The target genes, TFs and miRNAs are differentially expressed in EC so that any mutation in them may impact on EC development. Some key pathways in networks were highlighted to analyze how they interactively influence other factors and carcinogenesis. Upstream and downstream pathways of the differentially expressed genes and miRNAs were compared and analyzed. The purpose of this study was to partially reveal the deep regulatory mechanisms in EC using a new method that combines comprehensive genes and miRNAs together with their relationships. It may contribute to cancer prevention and gene therapy of EC. PMID:25684474

  17. The transcription factor c-Myc enhances KIR gene transcription through direct binding to an upstream distal promoter element

    PubMed Central

    Cichocki, Frank; Hanson, Rebecca J.; Lenvik, Todd; Pitt, Michelle; McCullar, Valarie; Li, Hongchuan; Anderson, Stephen K.

    2009-01-01

    The killer cell immunoglobulin-like receptor (KIR) repertoire of natural killer (NK) cells determines their ability to detect infected or transformed target cells. Although epigenetic mechanisms play a role in KIR gene expression, work in the mouse suggests that other regulatory elements may be involved at specific stages of NK-cell development. Here we report the effects of the transcription factor c-Myc on KIR expression. c-Myc directly binds to, and promotes transcription from, a distal element identified upstream of most KIR genes. Binding of endogenous c-Myc to the distal promoter element is significantly enhanced upon interleukin-15 (IL-15) stimulation in peripheral blood NK cells and correlates with an increase in KIR transcription. In addition, the overexpression of c-Myc during NK-cell development promotes transcription from the distal promoter element and contributes to the overall transcription of multiple KIR genes. Our data demonstrate the significance of the 5′ promoter element upstream of the conventional KIR promoter region and support a model whereby IL-15 stimulates c-Myc binding at the distal KIR promoter during NK-cell development to promote KIR transcription. This finding provides a direct link between NK-cell activation signals and KIR expression required for acquisition of effector function during NK-cell education. PMID:18987359

  18. Requirement of gene VII in cis for the expression of downstream genes on the major transcript of figwort mosaic virus.

    PubMed

    Gowda, S; Scholthof, H B; Wu, F C; Shepherd, R J

    1991-12-01

    The six major conserved genes of figwort mosaic virus (FMV), a caulimovirus, appear in tandem array on an RNA transcript that spans the entire viral genome. Gene VI, the only cistron that appears as a separate subgenomic RNA, has been reported to transactivate the expression of downstream genes of the full-length transcript. This transcript has a long 5'-leader of about 600 nucleotides followed by a small nonconserved region (gene VII), a smaller intergenic region (57 nucleotides), and the major conserved genes in a closely spaced array. In our present experiments we have constructed expression units containing the promoter for the full-length transcript followed by the 5' leader region, gene VII, and a reporter gene. These have been tested for expression with and without gene VI as a separate plasmid by electroporation into plant protoplasts. A series of these expression units containing truncated versions of the 5' leader region placed upstream of a reporter gene (CAT) showed that gene VI transactivation occurred only when gene VII sequences were present in cis between the leader region and the reporter gene. In addition, a more complete version of the FMV genome containing the reporter gene further downstream (in viral gene IV) showed CAT expression only when gene VII sequences were present in an upstream position. A similar construct failed to express CAT activity when gene VII was absent. PMID:1962457

  19. Induction of AhR-Mediated Gene Transcription by Coffee

    PubMed Central

    Ishikawa, Toshio; Takahashi, Satoshi; Morita, Koji; Okinaga, Hiroko; Teramoto, Tamio

    2014-01-01

    Background Aryl hydrocarbon receptor (AhR) is classically known to be activated by xenobiotics such as dioxins and polycyclic aromatic hydrocarbons (PAHs). Although it has been reported that PAHs are contained in roasted coffee beans, in general coffee beverages are not considered to be AhR activators. We tested whether exposure to coffee would activate AhR in cultured cells. Methods HepG2 cells stably expressing an AhR-responsive reporter gene were treated with coffee samples. Also, expression of CYP1A1, an endogenous AhR-responsive gene, was quantitated by RT-PCR and Western blotting in HepG2, Caco-2, and MCF-7 cells, after treatment with coffee. In order to obtain sensitive and reproducible results, all the experiments were performed with the cells placed in either phosphate-buffered saline (PBS) or pure serum, instead of routinely-used culture medium, whose intrinsic AhR-stimulating activity turned out to be so strong as to interfere with the analyses. Results All the coffee samples tested robustly stimulated AhR-mediated transcription in the reporter gene assays. Of note, to what extent coffee and other AhR agonists activated AhR was different, depending on whether the experiments were done in PBS or serum. CYP1A1 mRNA was induced by coffee, in HepG2, Caco-2, and MCF-7 cells placed in either PBS or serum. CYP1A1 protein expression, which was not detected in these cells incubated in PBS, was also increased by coffee in cells placed in serum. Conclusions By using culture medium-free experimental settings, we have shown that coffee is a strong AhR activator. Our observation may help elucidate as-yet-unrecognized effects of coffee on human health. PMID:25007155

  20. A Photo-Degradable Gene Delivery System for Enhanced Nuclear Gene Transcription

    PubMed Central

    Hoyoung, Lee; Yeji, Kim; Patrick G., Schweickert; Stephen F., Konieczny; You-Yeon, Won

    2013-01-01

    There currently exists a significant gap in our understanding of how the detailed chemical characteristics of polycation gene carriers influence their delivery performances in overcoming an important cellular-level transport barrier, i.e., intranuclear gene transcription. In this study, a UV-degradable gene carrier material (ENE4-1) was synthesized by crosslinking low molecular weight branched polyethylenimine (bPEI-2k) molecules using UV-cleavable o-nitrobenzyl urethane (NBU) as the linker molecule. NBU degrades upon exposure to mild UV irradiation. Therefore, this UV-degradable carrier allows us to control the chemical characteristics of the polymer/DNA complex (polyplex) particles at desired locations within the intracellular environment. By using this photolytic DNA carrier, we found that the exact timing of the UV degradation significantly influences the gene transfection efficiencies of ENE4-1/DNA(pGL2) polyplexes in HeLa cells. Interestingly, even if the polyplexes were UV-degraded at different intracellular locations/times, their nuclear entry efficiency was not influenced by the location/timing of UV degradation. The UV treatment did not influence the size or binding strength of the polyplexes. However, we confirmed that the degradation of the carrier molecules impacts the chemical characteristics of the polyplexes (it produces carbamic acid and nitrosobenzyl aldehyde groups on ENE4-1). We believe that these anionic acid groups enhance the interaction of the polyplexes with nuclear transcription proteins and thus the final gene expression levels; this effect was found to occur, even though UV irradiation itself has a general effect of reducing transfection efficiencies. Excess (uncomplexed) ENE4-1 polymers appear to not play any role in the UV-enhanced gene transcription phenomenon. PMID:24172855

  1. Transcriptional Profiling of Canker-Resistant Transgenic Sweet Orange (Citrus sinensis Osbeck) Constitutively Overexpressing a Spermidine Synthase Gene

    PubMed Central

    Fu, Xing-Zheng; Liu, Ji-Hong

    2013-01-01

    Citrus canker disease caused by Xanthomonas citri subsp. citri (Xcc) is one of the most devastating diseases affecting the citrus industry worldwide. In our previous study, the canker-resistant transgenic sweet orange (Citrus sinensis Osbeck) plants were produced via constitutively overexpressing a spermidine synthase. To unravel the molecular mechanisms underlying Xcc resistance of the transgenic plants, in the present study global transcriptional profiling was compared between untransformed line (WT) and the transgenic line (TG9) by hybridizing with Affymetrix Citrus GeneChip. In total, 666 differentially expressed genes (DEGs) were identified, 448 upregulated, and 218 downregulated. The DEGs were classified into 33 categories after Gene ontology (GO) annotation, in which 68 genes are in response to stimulus and involved in immune system process, 12 genes are related to cell wall, and 13 genes belong to transcription factors. These genes and those related to starch and sucrose metabolism, glutathione metabolism, biosynthesis of phenylpropanoids, and plant hormones were hypothesized to play major roles in the canker resistance of TG9. Semiquantitative RT-PCR analysis showed that the transcript levels of several candidate genes in TG9 were significantly higher than in WT both before and after Xcc inoculation, indicating their potential association with canker disease. PMID:23509803

  2. Gene transcript profiling in sea otters post-Exxon Valdez oil spill: A tool for marine ecosystem health assessment

    USGS Publications Warehouse

    Bowen, Lizabeth; Miles, A. Keith; Ballachey, Brenda E.; Waters, Shannon C.; Bodkin, James L.

    2016-01-01

    Using a panel of genes stimulated by oil exposure in a laboratory study, we evaluated gene transcription in blood leukocytes sampled from sea otters captured from 2006–2012 in western Prince William Sound (WPWS), Alaska, 17–23 years after the 1989 Exxon Valdez oil spill (EVOS). We compared WPWS sea otters to reference populations (not affected by the EVOS) from the Alaska Peninsula (2009), Katmai National Park and Preserve (2009), Clam Lagoon at Adak Island (2012), Kodiak Island (2005) and captive sea otters in aquaria. Statistically, sea otter gene transcript profiles separated into three distinct clusters: Cluster 1, Kodiak and WPWS 2006–2008 (higher relative transcription); Cluster 2, Clam Lagoon and WPWS 2010–2012 (lower relative transcription); and Cluster 3, Alaska Peninsula, Katmai and captive sea otters (intermediate relative transcription). The lower transcription of the aryl hydrocarbon receptor (AHR), an established biomarker for hydrocarbon exposure, in WPWS 2010–2012 compared to earlier samples from WPWS is consistent with declining hydrocarbon exposure, but the pattern of overall low levels of transcription seen in WPWS 2010–2012 could be related to other factors, such as food limitation, pathogens or injury, and may indicate an inability to mount effective responses to stressors. Decreased transcriptional response across the entire gene panel precludes the evaluation of whether or not individual sea otters show signs of exposure to lingering oil. However, related studies on sea otter demographics indicate that by 2012, the sea otter population in WPWS had recovered, which indicates diminishing oil exposure.

  3. Group I Metabotropic Glutamate Receptor-Mediated Gene Transcription and Implications for Synaptic Plasticity and Diseases

    PubMed Central

    Wang, Hansen; Zhuo, Min

    2012-01-01

    Stimulation of group I metabotropic glutamate receptors (mGluRs) initiates a wide variety of signaling pathways. Group I mGluR activation can regulate gene expression at both translational and transcriptional levels, and induces translation or transcription-dependent synaptic plastic changes in neurons. The group I mGluR-mediated translation-dependent neural plasticity has been well reviewed. In this review, we will highlight group I mGluR-induced gene transcription and its role in synaptic plasticity. The signaling pathways (PKA, CaMKs, and MAPKs) which have been shown to link group I mGluRs to gene transcription, the relevant transcription factors (CREB and NF-κB), and target proteins (FMRP and ARC) will be documented. The significance and future direction for characterizing group I mGluR-mediated gene transcription in fragile X syndrome, schizophrenia, drug addiction, and other neurological disorders will also be discussed. PMID:23125836

  4. Multiple transcription factor binding sites predict AID targeting in non-immunoglobulin genes

    PubMed Central

    Duke, Jamie L.; Liu, Man; Yaari, Gur; Khalil, Ashraf M.; Tomayko, Mary M.; Shlomchik, Mark J.; Schatz, David G.; Kleinstein, Steven H.

    2013-01-01

    Aberrant targeting of the enzyme Activation Induced Cytidine Deaminase (AID) results in the accumulation of somatic mutations in approximately 25% of expressed genes in germinal center B cells. Observations in Ung−/− Msh2−/− mice suggest that many other genes efficiently repair AID-induced lesions, so that up to 45% of genes may actually be targeted by AID. It is important to understand the mechanisms that recruit AID to certain genes, as this mis-targeting represents an important risk for genome instability. We hypothesize that several mechanisms will combine to target AID to each locus. In order to resolve which mechanisms affect AID targeting, we analyze 7.3Mb of sequence data, along with the regulatory context, from 83 genes in Ung−/− Msh2−/− mice to identify common properties of AID targets. This analysis identifies the involvement of three transcription factor binding sites (E-box motifs, along with YY1 and C/EBP-beta binding sites) that may work together to recruit AID. Based on previous knowledge and these newly discovered features, a classification tree model was built to predict genome-wide AID targeting. Using this predictive model we were able to identify a set of 101 high-interest genes that are likely targets of AID. PMID:23514741

  5. Mining whole genomes and transcriptomes of Jatropha (Jatropha curcas) and Castor bean (Ricinus communis) for NBS-LRR genes and defense response associated transcription factors.

    PubMed

    Sood, Archit; Jaiswal, Varun; Chanumolu, Sree Krishna; Malhotra, Nikhil; Pal, Tarun; Chauhan, Rajinder Singh

    2014-11-01

    Jatropha (Jatropha curcas L.) and Castor bean (Ricinus communis) are oilseed crops of family Euphorbiaceae with the potential of producing high quality biodiesel and having industrial value. Both the bioenergy plants are becoming susceptible to various biotic stresses directly affecting the oil quality and content. No report exists as of today on analysis of Nucleotide Binding Site-Leucine Rich Repeat (NBS-LRR) gene repertoire and defense response transcription factors in both the plant species. In silico analysis of whole genomes and transcriptomes identified 47 new NBS-LRR genes in both the species and 122 and 318 defense response related transcription factors in Jatropha and Castor bean, respectively. The identified NBS-LRR genes and defense response transcription factors were mapped onto the respective genomes. Common and unique NBS-LRR genes and defense related transcription factors were identified in both the plant species. All NBS-LRR genes in both the species were characterized into Toll/interleukin-1 receptor NBS-LRRs (TNLs) and coiled-coil NBS-LRRs (CNLs), position on contigs, gene clusters and motifs and domains distribution. Transcript abundance or expression values were measured for all NBS-LRR genes and defense response transcription factors, suggesting their functional role. The current study provides a repertoire of NBS-LRR genes and transcription factors which can be used in not only dissecting the molecular basis of disease resistance phenotype but also in developing disease resistant genotypes in Jatropha and Castor bean through transgenic or molecular breeding approaches. PMID:25106526

  6. SPL8, an SBP-box gene that affects pollen sac development in Arabidopsis.

    PubMed

    Unte, Ulrike S; Sorensen, Anna-Marie; Pesaresi, Paolo; Gandikota, Madhuri; Leister, Dario; Saedler, Heinz; Huijser, Peter

    2003-04-01

    SQUAMOSA PROMOTER BINDING PROTEIN-box genes (SBP-box genes) encode plant-specific proteins that share a highly conserved DNA binding domain, the SBP domain. Although likely to represent transcription factors, little is known about their role in development. In Arabidopsis, SBP-box genes constitute a structurally heterogeneous family of 16 members known as SPL genes. For one of these genes, SPL8, we isolated three independent transposon-tagged mutants, all of which exhibited a strong reduction in fertility. Microscopic analysis revealed that this reduced fertility is attributable primarily to abnormally developed microsporangia, which exhibit premeiotic abortion of the sporocytes. In addition to its role in microsporogenesis, the SPL8 knockout also seems to affect megasporogenesis, trichome formation on sepals, and stamen filament elongation. The SPL8 mutants described help to uncover the roles of SBP-box genes in plant development. PMID:12671094

  7. SPL8, an SBP-Box Gene That Affects Pollen Sac Development in Arabidopsis

    PubMed Central

    Unte, Ulrike S.; Sorensen, Anna-Marie; Pesaresi, Paolo; Gandikota, Madhuri; Leister, Dario; Saedler, Heinz; Huijser, Peter

    2003-01-01

    SQUAMOSA PROMOTER BINDING PROTEIN–box genes (SBP-box genes) encode plant-specific proteins that share a highly conserved DNA binding domain, the SBP domain. Although likely to represent transcription factors, little is known about their role in development. In Arabidopsis, SBP-box genes constitute a structurally heterogeneous family of 16 members known as SPL genes. For one of these genes, SPL8, we isolated three independent transposon-tagged mutants, all of which exhibited a strong reduction in fertility. Microscopic analysis revealed that this reduced fertility is attributable primarily to abnormally developed microsporangia, which exhibit premeiotic abortion of the sporocytes. In addition to its role in microsporogenesis, the SPL8 knockout also seems to affect megasporogenesis, trichome formation on sepals, and stamen filament elongation. The SPL8 mutants described help to uncover the roles of SBP-box genes in plant development. PMID:12671094

  8. Attenuation of Histone Methyltransferase KRYPTONITE-mediated transcriptional gene silencing by Geminivirus

    PubMed Central

    Sun, Yan-Wei; Tee, Chuan-Sia; Ma, Yong-Huan; Wang, Gang; Yao, Xiang-Mei; Ye, Jian

    2015-01-01

    Although histone H3K9 methylation has been intensively studied in animals and a model plant Arabidopsis thaliana, little is known about the evolution of the histone methyltransferase and its roles in plant biotic stress response. Here we identified a Nicotiana benthamiana homolog of H3K9 histone methyltransferase KRYPTONITE (NbKYP) and demonstrated its fundamental roles on methylation of plant and virus, beside of leading to the suppression of endogenous gene expression and virus replication. NbKYP and another gene encoding DNA methyltransferase CHROMOMETHYLTRANSFERASE 3 (NbCMT3-1) were further identified as the key components of maintenance of transcriptional gene silencing, a DNA methylation involved anti-virus machinery. All three types of DNA methylations (asymmetric CHH and symmetric CHG/CG) were severely affected in NbKYP-silenced plants, but only severe reduction of CHG methylation found in NbCMT3-1-silenced plants. Attesting to the importance of plant histone H3K9 methylation immunity to virus, the virulence of geminiviruses requires virus-encoded trans-activator AC2 which inhibits the expression of KYP via activation of an EAR-motif-containing transcription repressor RAV2 (RELATED TO ABI3 and VP1). The reduction of KYP was correlated with virulence of various similar geminiviruses. These findings provide a novel mechanism of how virus trans-activates a plant endogenous anti-silencing machinery to gain high virulence. PMID:26602265

  9. Mutant Huntingtin Downregulates Myelin Regulatory Factor-Mediated Myelin Gene Expression and Affects Mature Oligodendrocytes

    PubMed Central

    Huang, Brenda; Wei, Wenjie; Wang, Guohao; Gaertig, Marta A.; Feng, Yue; Wang, Wei; Li, Xiao-Jiang; Li, Shihua

    2015-01-01

    SUMMARY Growing evidence indicates that non-neuronal mutant huntingtin toxicity plays an important role in Huntington’s disease (HD); however, whether and how mutant huntingtin affects oligodendrocytes, which are vitally important for neural function and axonal integrity, remain unclear. We first verified the presence of mutant huntingtin in oligodendrocytes in HD140Q knock-in mice. We then established transgenic mice (PLP-150Q) that selectively express mutant huntingtin in oligodendrocytes. PLP-150Q mice show progressive neurological symptoms and early death, as well as age-dependent demyelination and reduced expression of myelin genes that are downstream of myelin regulatory factor (MYRF or MRF), a transcriptional regulator that specifically activates and maintains the expression of myelin genes in mature oligodendrocytes. Consistently, mutant huntingtin binds abnormally to MYRF and affects its transcription activity. Our findings suggest that dysfunction of mature oligodendrocytes is involved in HD pathogenesis and may also make a good therapeutic target. PMID:25789755

  10. Mutant huntingtin downregulates myelin regulatory factor-mediated myelin gene expression and affects mature oligodendrocytes.

    PubMed

    Huang, Brenda; Wei, WenJie; Wang, Guohao; Gaertig, Marta A; Feng, Yue; Wang, Wei; Li, Xiao-Jiang; Li, Shihua

    2015-03-18

    Growing evidence indicates that non-neuronal mutant huntingtin toxicity plays an important role in Huntington's disease (HD); however, whether and how mutant huntingtin affects oligodendrocytes, which are vitally important for neural function and axonal integrity, remains unclear. We first verified the presence of mutant huntingtin in oligodendrocytes in HD140Q knockin mice. We then established transgenic mice (PLP-150Q) that selectively express mutant huntingtin in oligodendrocytes. PLP-150Q mice show progressive neurological symptoms and early death, as well as age-dependent demyelination and reduced expression of myelin genes that are downstream of myelin regulatory factor (MYRF or MRF), a transcriptional regulator that specifically activates and maintains the expression of myelin genes in mature oligodendrocytes. Consistently, mutant huntingtin binds abnormally to MYRF and affects its transcription activity. Our findings suggest that dysfunction of mature oligodendrocytes is involved in HD pathogenesis and may also make a good therapeutic target. PMID:25789755

  11. Transcriptional Activation of Inflammatory Genes: Mechanistic Insight into Selectivity and Diversity

    PubMed Central

    Ahmed, Afsar U.; Williams, Bryan R. G.; Hannigan, Gregory E.

    2015-01-01

    Acute inflammation, an integral part of host defence and immunity, is a highly conserved cellular response to pathogens and other harmful stimuli. An inflammatory stimulation triggers transcriptional activation of selective pro-inflammatory genes that carry out specific functions such as anti-microbial activity or tissue healing. Based on the nature of inflammatory stimuli, an extensive exploitation of selective transcriptional activations of pro-inflammatory genes is performed by the host to ensure a defined inflammatory response. Inflammatory signal transductions are initiated by the recognition of inflammatory stimuli by transmembrane receptors, followed by the transmission of the signals to the nucleus for differential gene activations. The differential transcriptional activation of pro-inflammatory genes is precisely controlled by the selective binding of transcription factors to the promoters of these genes. Among a number of transcription factors identified to date, NF-κB still remains the most prominent and studied factor for its diverse range of selective transcriptional activities. Differential transcriptional activities of NF-κB are dictated by post-translational modifications, specificities in dimer formation, and variability in activation kinetics. Apart from the differential functions of transcription factors, the transcriptional activation of selective pro-inflammatory genes is also governed by chromatin structures, epigenetic markers, and other regulators as the field is continuously expanding. PMID:26569329

  12. Transcriptional Activation of Inflammatory Genes: Mechanistic Insight into Selectivity and Diversity.

    PubMed

    Ahmed, Afsar U; Williams, Bryan R G; Hannigan, Gregory E

    2015-01-01

    Acute inflammation, an integral part of host defence and immunity, is a highly conserved cellular response to pathogens and other harmful stimuli. An inflammatory stimulation triggers transcriptional activation of selective pro-inflammatory genes that carry out specific functions such as anti-microbial activity or tissue healing. Based on the nature of inflammatory stimuli, an extensive exploitation of selective transcriptional activations of pro-inflammatory genes is performed by the host to ensure a defined inflammatory response. Inflammatory signal transductions are initiated by the recognition of inflammatory stimuli by transmembrane receptors, followed by the transmission of the signals to the nucleus for differential gene activations. The differential transcriptional activation of pro-inflammatory genes is precisely controlled by the selective binding of transcription factors to the promoters of these genes. Among a number of transcription factors identified to date, NF-κB still remains the most prominent and studied factor for its diverse range of selective transcriptional activities. Differential transcriptional activities of NF-κB are dictated by post-translational modifications, specificities in dimer formation, and variability in activation kinetics. Apart from the differential functions of transcription factors, the transcriptional activation of selective pro-inflammatory genes is also governed by chromatin structures, epigenetic markers, and other regulators as the field is continuously expanding. PMID:26569329

  13. A recombinant rabies virus carrying GFP between N and P affects viral transcription in vitro.

    PubMed

    Luo, Jun; Zhao, Jing; Tian, Qin; Mo, Weiyu; Wang, Yifei; Chen, Hao; Guo, Xiaofeng

    2016-06-01

    Several studies have demonstrated the rabies virus to be a perfect potential vaccine vector to insert foreign genes into the target genome. For this study, a green fluorescent protein (GFP) gene was cloned into the rabies virus (RABV) genome between the N and P gene. CT dinucleotide was inserted as intergenic region. The recombinant high egg passage Flury strain (HEP-Flury) of RABV, carrying GFP (rHEP-NP-GFP), was generated in BHK-21 cells using reverse genetics. According to the viral growth kinetics assay, the addition of GFP between N and P gene has little effect on the viral growth compared to the parental strain HEP-Flury. Quantitative real-time PCR (qPCR) indicated that rHEP-NP-GFP showed different viral gene transcription, especially for G gene, compared to HEP-Flury. The same is true for one other recombinant RABV carrying GFP between G and L gene in NA cells. In addition, parent HEP-Flury showed more expression of innate immune-related molecules in NA cells. Compared to HEP-Flury, Western blotting (WB) indicated that insertion of a foreign gene following N gene enhanced the expression of M and G proteins. According to the qPCR and WB, GFP expression levels of rHEP-NP-GFP were significantly higher than rHEP-GFP. This study indicates HEP-Flury as valid vector to express exogenous genes between N and P. PMID:26957093

  14. Evaluation of position effect variegation of the transcription of genes from the FSHD candidate region

    SciTech Connect

    Winokur, S.T.; Wasmuth, J.J.; Altherr, M.R.

    1994-09-01

    The gene for facioscapulohumeral muscular dystrophy (FSHD) lies in close proximity to the telomere of 4q. Deletion of several copies of a 3.2 kb tandem repeat have been associated with FSHD, although no genes have been identified within this repeat. We have shown that this repeat, as well as other repeats in the FSHD region, resemble constitutive heterochromatin both by sequence analysis and FISH cross-hybridization. We hypothesize that alterations in chromatin structure near the telomere of 4q due to deletion of these heterochromatic elements may lead to a position effect variegation of nearby genes. To test this hypothesis, we have isolated exons and candidate cDNAs from the FSHD region. A 2 kb polyadenylated cDNA was isolated from both fetal and infant brain cDNA libraries. Another cDNA hybridizes to a 7 kb skeletal muscle transcript on a Northern blot. Both of these cDNAs are chromosome 4-specific and map to the FSHD region. We have examined the expression pattern of these genes by RT-PCR, RNase protection and Northern analysis. Total RNA was isolated from normal and FSHD-affected lymphoblasts and from human-hamster somatic cell hybrids in which the normal and affected chromosomes 4 from FSHD patients were segregated. RT-PCR and RNase protection were then employed as quantitive assays to evaluate the potential for position effect variegation on RNA production in FSHD patients.

  15. The yeast Hot1 transcription factor is critical for activating a single target gene, STL1

    PubMed Central

    Bai, Chen; Tesker, Masha; Engelberg, David

    2015-01-01

    Transcription factors are commonly activated by signal transduction cascades and induce expression of many genes. They therefore play critical roles in determining the cell's fate. The yeast Hog1 MAP kinase pathway is believed to control the transcription of hundreds of genes via several transcription factors. To identify the bona fide target genes of Hog1, we inducibly expressed the spontaneously active variant Hog1D170A+F318L in cells lacking the Hog1 activator Pbs2. This system allowed monitoring the effects of Hog1 by itself. Expression of Hog1D170A+F318L in pbs2∆ cells imposed induction of just 105 and suppression of only 26 transcripts by at least twofold. We looked for the Hog1-responsive element within the promoter of the most highly induced gene, STL1 (88-fold). A novel Hog1 responsive element (HoRE) was identified and shown to be the direct target of the transcription factor Hot1. Unexpectedly, we could not find this HoRE in any other yeast promoter. In addition, the only gene whose expression was abolished in hot1∆ cells was STL1. Thus Hot1 is essential for transcription of just one gene, STL1. Hot1 may represent a class of transcription factors that are essential for transcription of a very few genes or even just one. PMID:25904326

  16. From cell membrane to the nucleus: an emerging role of E-cadherin in gene transcriptional regulation

    PubMed Central

    Du, Wenjun; Liu, Xi; Fan, Guiling; Zhao, Xingsheng; Sun, Yanying; Wang, Tianzhen; Zhao, Ran; Wang, Guangyu; Zhao, Ci; Zhu, Yuanyuan; Ye, Fei; Jin, Xiaoming; Zhang, Fengmin; Zhong, Zhaohua; Li, Xiaobo

    2014-01-01

    E-cadherin is a well-known mediator of cell–cell adherens junctions. However, many other functions of E-cadherin have been reported. Collectively, the available data suggest that E-cadherin may also act as a gene transcriptional regulator. Here, evidence supporting this claim is reviewed, and possible mechanisms of action are discussed. E-cadherin has been shown to modulate the activity of several notable cell signalling pathways, and given that most of these pathways in turn regulate gene expression, we proposed that E-cadherin may regulate gene transcription by affecting these pathways. Additionally, E-cadherin has been shown to accumulate in the nucleus where documentation of an E-cadherin fragment bound to DNA suggests that E-cadherin may directly regulate gene transcription. In summary, from the cell membrane to the nucleus, a role for E-cadherin in gene transcription may be emerging. Studies specifically focused on this potential role would allow for a more thorough understanding of this transmembrane glycoprotein in mediating intra- and intercellular activities. PMID:25164084

  17. Insulin post-transcriptionally modulates Bmal1 protein to affect the hepatic circadian clock.

    PubMed

    Dang, Fabin; Sun, Xiujie; Ma, Xiang; Wu, Rong; Zhang, Deyi; Chen, Yaqiong; Xu, Qian; Wu, Yuting; Liu, Yi

    2016-01-01

    Although food availability is a potent synchronizer of the peripheral circadian clock in mammals, the underlying mechanisms are unclear. Here, we show that hepatic Bmal1, a core transcription activator of the molecular clock, is post-transcriptionally regulated by signals from insulin, an important hormone that is temporally controlled by feeding. Insulin promotes postprandial Akt-mediated Ser42-phosphorylation of Bmal1 to induce its dissociation from DNA, interaction with 14-3-3 protein and subsequently nuclear exclusion, which results in the suppression of Bmal1 transcriptional activity. Inverted feeding cycles not only shift the phase of daily insulin oscillation, but also elevate the amplitude due to food overconsumption. This enhanced and reversed insulin signalling initiates the reset of clock gene rhythms by altering Bmal1 nuclear accumulation in mouse liver. These results reveal the molecular mechanism of insulin signalling in regulating peripheral circadian rhythms. PMID:27576939

  18. Reconstitution of Heterochromatin-Dependent Transcriptional Gene Silencing

    PubMed Central

    Johnson, Aaron; Li, Geng; Sikorski, Timothy W.; Buratowski, Stephen; Woodcock, Christopher L.; Moazed, Danesh

    2009-01-01

    Summary Heterochromatin assembly in budding yeast requires the SIR complex, which contains the NAD-dependent deacetylase Sir2 and the Sir3 and Sir4 proteins. Sir3 binds to nucleosomes containing deacetylated histone H4 lysine 16 (H4K16) and, with Sir4, promotes spreading of Sir2 and deacetylation along the chromatin fiber. Combined action of histone modifying and binding activities is a conserved hallmark of heterochromatin, but the relative contribution of each activity to silencing has remained unclear. Here we reconstitute SIR-chromatin complexes using purified components and show that the SIR complex efficiently deacetylates chromatin templates and promotes the assembly of altered structures that silence Gal4-VP16-activated transcription. Silencing requires all three Sir proteins, even with fully deacetylated chromatin, and involves the specific association of Sir3 with deacetylated H4K16. These results define a minimal set of components that mediate heterochromatic gene silencing and demonstrate distinct contributions for histone deacetylation and nucleosome binding in the silencing mechanism. PMID:19782027

  19. MEF2 transcription factors: developmental regulators and emerging cancer genes

    PubMed Central

    Pon, Julia R.; Marra, Marco A.

    2016-01-01

    The MEF2 transcription factors have roles in muscle, cardiac, skeletal, vascular, neural, blood and immune system cell development through their effects on cell differentiation, proliferation, apoptosis, migration, shape and metabolism. Altered MEF2 activity plays a role in human diseases and has recently been implicated in the development of several cancer types. In particular, MEF2B, the most divergent and least studied protein of the MEF2 family, has a role unique from its paralogs in non-Hodgkin lymphomas. The use of genome-scale technologies has enabled comprehensive MEF2 target gene sets to be identified, contributing to our understanding of MEF2 proteins as nodes in complex regulatory networks. This review surveys the molecular interactions of MEF2 proteins and their effects on cellular and organismal phenotypes. We include a discussion of the emerging roles of MEF2 proteins as oncogenes and tumor suppressors of cancer. Throughout this article we highlight similarities and differences between the MEF2 family proteins, including a focus on functions of MEF2B. PMID:26506234

  20. dcp gene of Escherichia coli: cloning, sequencing, transcript mapping, and characterization of the gene product.

    PubMed Central

    Henrich, B; Becker, S; Schroeder, U; Plapp, R

    1993-01-01

    Dipeptidyl carboxypeptidase is a C-terminal exopeptidase of Escherichia coli. We have isolated the respective gene, dcp, from a low-copy-number plasmid library by its ability to complement a dcp mutation preventing the utilization of the unique substrate N-benzoyl-L-glycyl-L-histidyl-L-leucine. Sequence analysis of a 2.9-kb DNA fragment revealed an open reading frame of 2,043 nucleotides which was assigned to the dcp gene by N-terminal amino acid sequencing and electrophoretic molecular mass determination of the purified dcp product. Transcript mapping by primer extension and S1 protection experiments verified the physiological significance of potential initiation and termination signals for dcp transcription and allowed the identification of a single species of monocistronic dcp mRNA. The codon usage pattern and the effects of elevated gene copy number indicated a relatively low level of dcp expression. The predicted amino acid sequence of dipeptidyl carboxypeptidase, containing a potential zinc-binding site, is highly homologous (78.8%) to the corresponding enzyme from Salmonella typhimurium. It also displays significant homology to the products of the S. typhimurium opdA and the E. coli prlC genes and to some metalloproteases from rats and Saccharomyces cerevisiae. No potential export signals could be inferred from the amino acid sequence. Dipeptidyl carboxypeptidase was enriched 80-fold from crude extracts of E. coli and used to investigate some of its biochemical and biophysical properties. Images PMID:8226676

  1. Transcriptional Response of Selenopolypeptide Genes and Selenocysteine Biosynthesis Machinery Genes in Escherichia coli during Selenite Reduction

    PubMed Central

    Tetteh, Antonia Y.; Sun, Katherine H.; Kittur, Farooqahmed S.; Ibeanu, Gordon C.

    2014-01-01

    Bacteria can reduce toxic selenite into less toxic, elemental selenium (Se0), but the mechanism on how bacterial cells reduce selenite at molecular level is still not clear. We used Escherichia coli strain K12, a common bacterial strain, as a model to study its growth response to sodium selenite (Na2SeO3) treatment and then used quantitative real-time PCR (qRT-PCR) to quantify transcript levels of three E. coli selenopolypeptide genes and a set of machinery genes for selenocysteine (SeCys) biosynthesis and incorporation into polypeptides, whose involvements in the selenite reduction are largely unknown. We determined that 5 mM Na2SeO3 treatment inhibited growth by ∼50% while 0.001 to 0.01 mM treatments stimulated cell growth by ∼30%. Under 50% inhibitory or 30% stimulatory Na2SeO3 concentration, selenopolypeptide genes (fdnG, fdoG, and fdhF) whose products require SeCys but not SeCys biosynthesis machinery genes were found to be induced ≥2-fold. In addition, one sulfur (S) metabolic gene iscS and two previously reported selenite-responsive genes sodA and gutS were also induced ≥2-fold under 50% inhibitory concentration. Our findings provide insight about the detoxification of selenite in E. coli via induction of these genes involved in the selenite reduction process. PMID:24839442

  2. Transcription of two classes of rat growth hormone gene-associated repetitive DNA: differences in activity and effects of tandem repeat structure.

    PubMed Central

    Gutierrez-Hartmann, A; Lieberburg, I; Gardner, D; Baxter, J D; Cathala, G G

    1984-01-01

    The rat growth hormone (rGH) gene contains two classes of repetitive DNA arranged as clusters within intron B and the 3' flanking region. The major family is equivalent to the CHO type 2 DNA. The second ("truncated repeat", TR) is a truncated version of the first and occurs in certain neural-specific transcripts and genes ("identifier" elements, ID). Here we report, using the HeLa cell-free transcription assay, that RNA polymerase III (Pol III) efficiently initiates at internal promoters within a tandem array of rGH gene repetitive DNA monomers and results in a novel organization of overlapping Class III transcription units. Transcription competition studies revealed that the rat type 2 structures share Pol III transcription factors with a tRNA gene, a human Alu repeat, and a mutant VA1 gene. Also, the rGH type 2 but not the TR DNA efficiently promotes Pol III initiation, yet other TR members, which differ only in flanking DNA, are transcribed. Thus, the rGH gene is strikingly enriched with 10 repetitive DNA monomers; multimeric type 2 elements are actively transcribed; rGH-TR sequences are expressed only as part of larger transcripts promoted by type 2 DNA; and, type 2 DNA uses tRNA gene transcription factors. These studies show that flanking sequences, promoter organization and factor competition may all affect rat repetitive DNA expression. Images PMID:6091058

  3. Local potentiation of stress-responsive genes by upstream noncoding transcription

    PubMed Central

    Takemata, Naomichi; Oda, Arisa; Yamada, Takatomi; Galipon, Josephine; Miyoshi, Tomoichiro; Suzuki, Yutaka; Sugano, Sumio; Hoffman, Charles S.; Hirota, Kouji; Ohta, Kunihiro

    2016-01-01

    It has been postulated that a myriad of long noncoding RNAs (lncRNAs) contribute to gene regulation. In fission yeast, glucose starvation triggers lncRNA transcription across promoter regions of stress-responsive genes including fbp1 (fructose-1,6-bisphosphatase1). At the fbp1 promoter, this transcription promotes chromatin remodeling and fbp1 mRNA expression. Here, we demonstrate that such upstream noncoding transcription facilitates promoter association of the stress-responsive transcriptional activator Atf1 at the sites of transcription, leading to activation of the downstream stress genes. Genome-wide analyses revealed that ∼50 Atf1-binding sites show marked decrease in Atf1 occupancy when cells are treated with a transcription inhibitor. Most of these transcription-enhanced Atf1-binding sites are associated with stress-dependent induction of the adjacent mRNAs or lncRNAs, as observed in fbp1. These Atf1-binding sites exhibit low Atf1 occupancy and high histone density in glucose-rich conditions, and undergo dramatic changes in chromatin status after glucose depletion: enhanced Atf1 binding, histone eviction, and histone H3 acetylation. We also found that upstream transcripts bind to the Groucho-Tup1 type transcriptional corepressors Tup11 and Tup12, and locally antagonize their repressive functions on Atf1 binding. These results reveal a new mechanism in which upstream noncoding transcription locally magnifies the specific activation of stress-inducible genes via counteraction of corepressors. PMID:26945040

  4. Local potentiation of stress-responsive genes by upstream noncoding transcription.

    PubMed

    Takemata, Naomichi; Oda, Arisa; Yamada, Takatomi; Galipon, Josephine; Miyoshi, Tomoichiro; Suzuki, Yutaka; Sugano, Sumio; Hoffman, Charles S; Hirota, Kouji; Ohta, Kunihiro

    2016-06-20

    It has been postulated that a myriad of long noncoding RNAs (lncRNAs) contribute to gene regulation. In fission yeast, glucose starvation triggers lncRNA transcription across promoter regions of stress-responsive genes including fbp1 (fructose-1,6-bisphosphatase1). At the fbp1 promoter, this transcription promotes chromatin remodeling and fbp1 mRNA expression. Here, we demonstrate that such upstream noncoding transcription facilitates promoter association of the stress-responsive transcriptional activator Atf1 at the sites of transcription, leading to activation of the downstream stress genes. Genome-wide analyses revealed that ∼50 Atf1-binding sites show marked decrease in Atf1 occupancy when cells are treated with a transcription inhibitor. Most of these transcription-enhanced Atf1-binding sites are associated with stress-dependent induction of the adjacent mRNAs or lncRNAs, as observed in fbp1 These Atf1-binding sites exhibit low Atf1 occupancy and high histone density in glucose-rich conditions, and undergo dramatic changes in chromatin status after glucose depletion: enhanced Atf1 binding, histone eviction, and histone H3 acetylation. We also found that upstream transcripts bind to the Groucho-Tup1 type transcriptional corepressors Tup11 and Tup12, and locally antagonize their repressive functions on Atf1 binding. These results reveal a new mechanism in which upstream noncoding transcription locally magnifies the specific activation of stress-inducible genes via counteraction of corepressors. PMID:26945040

  5. Inosine monophosphate dehydrogenase expression: transcriptional regulation of the type I and type II genes.

    PubMed

    Zimmermann, A; Gu, J J; Spychala, J; Mitchell, B S

    1996-01-01

    Inosine 5'-monophosphate dehydrogenase (IMPDH) is an essential rate-limiting enzyme in the de novo guanine nucleotide synthetic pathway that catalyzes the conversion of IMP to XMP. Enzyme activity is accounted for by the expression of two distinct but closely related genes termed IMPDH I and II. Increased IMPDH activity has been linked to both cellular proliferation and neoplastic transformation and generally ascribed to an increase in the expression of the type II gene. We have characterized the type I and type II genes and identified elements important in the transcriptional regulation of both genes. The type II IMPDH gene contains a 466 bp 5' flanking region spanning the translation start site that contains several transcription factor binding sites and mediates increased transcription of a CAT reporter gene in peripheral blood T lymphocytes when these cells are induced to proliferate. The single functional IMPDH type I gene contains exon-intron boundaries and exon structures that are nearly identical to those in the type II gene. In contrast to the type II gene, however, it contains two putative promoter sites, each with the potential for transcriptional regulation. We conclude that these two genes most probably arose from an early gene duplication event and that their highly conserved structures and differential regulation at the transcriptional level argue strongly for a significant role for each gene in cellular metabolism, growth, and differentiation. PMID:8869741

  6. Transcriptional and Posttranscriptional Regulation of Dormancy-Associated Gene Expression by Afterripening in Wild Oat.

    PubMed Central

    Li, Bailin.; Foley, M. E.

    1996-01-01

    To investigate whether the afterripening-induced changes in gene expression are at the transcriptional or posttranscriptional level in wild oat (Avena fatua) seeds, we chose four dormancy-associated genes to estimate their relative transcription activities and the stability of their corresponding transcripts in afterripened and dormant embryos. The transcription activities for those genes were 1.5 to 7 times higher in dormant embryos than in afterripened embryos 24 h after incubation, as determined by nuclear run-on assays. The half-lives of the transcripts in afterripened and dormant embryos were estimated by the use of actinomycin D. The application of actinomycin D resulted in the stabilization of the transcripts. Nevertheless, the results indicated that the half-lives of the transcripts were much greater in dormant embryos than in afterripened embryos. Considering the great differences in the steady-state levels and the half-lives of the mRNAs, and the relatively small differences in transcription activities of the genes between afterripened and dormant embryos, we conclude that afterripening regulates the expression of dormancy-associated genes in excised embryos mainly at the posttranscriptional level and that transcriptional control plays a minor role. PMID:12226258

  7. Transcription of nitrification genes by the methane-oxidizing bacterium, Methylococcus capsulatus strain Bath.

    PubMed

    Poret-Peterson, Amisha T; Graham, James E; Gulledge, Jay; Klotz, Martin G

    2008-12-01

    Methylococcus capsulatus strain Bath, a methane-oxidizing bacterium, and ammonia-oxidizing bacteria (AOB) carry out the first step of nitrification, the oxidation of ammonia to nitrite, through the intermediate hydroxylamine. AOB use hydroxylamine oxidoreductase (HAO) to produce nitrite. M. capsulatus Bath was thought to oxidize hydroxylamine with cytochrome P460 (cytL), until the recent discovery of an hao gene in its genome. We used quantitative PCR analyses of cDNA from M. capsulatus Bath incubated with CH(4) or CH(4) plus 5 mM (NH(4))(2)SO(4) to determine whether cytL and hao transcript levels change in response to ammonia. While mRNA levels for cytL were not affected by ammonia, hao mRNA levels increased by 14.5- and 31-fold in duplicate samples when a promoter proximal region of the transcript was analyzed, and by sixfold when a region at the distal end of the transcript was analyzed. A conserved open reading frame, orf2, located 3' of hao in all known AOB genomes and in M. capsulatus Bath, was cotranscribed with hao and showed increased mRNA levels in the presence of ammonia. These data led to designating this gene pair as haoAB, with the role of haoB still undefined. We also determined mRNA levels for additional genes that encode proteins involved in N-oxide detoxification: cytochrome c'-beta (CytS) and nitric oxide (NO) reductase (NorCB). Whereas cytS mRNA levels increased in duplicate samples by 28.5- and 40-fold in response to ammonia, the cotranscribed norC-norB mRNA did not increase. Our results strongly suggest that M. capsulatus Bath possesses a functional, ammonia-responsive HAO involved in nitrification. PMID:18650926

  8. The roles of eighteen baculovirus late expression factor genes in transcription and DNA replication.

    PubMed Central

    Lu, A; Miller, L K

    1995-01-01

    A set of 18 plasmid subclones of the Autographa californica nuclear polyhedrosis virus genome supports expression from a late viral promoter in transient expression assays (J. W. Todd, A. L. Passarelli, and L. K. Miller, J. Virol. 69:968-974, 1995). Using this set of plasmids, we have assigned a role for each of the 18 genes required for optimal late gene expression with respect to its involvement at the levels of transcription, translation, and/or DNA replication. RNase protection analyses demonstrated that all of the known late expression factor genes (lefs) affected the steady-state level of reporter gene RNA. Thus, none of the lefs appeared to be specifically involved in translation. A subset of the lefs supported plasmid replication; ie-1, lef-1, lef-2, lef-3, p143, and p35 were essential for plasmid replication, while ie-n, lef-7, and dnapol had stimulatory effects. The predicted sequence of lef-7 suggests that it is a homolog of herpesvirus single-stranded DNA-binding protein (UL29). The role of p35 in plasmid replication appears to be suppression of apoptosis, because p35 could be functionally replaced in the replication assay by either Cp-iap or Op-iap, two heterologous baculovirus genes which suppress apoptosis by a mechanism which appears to differ from that of p35. Thus, one or more of the replication-related lefs or the process of plasmid replication appears to induce cellular apoptosis. Our results indicate that the remaining lefs, lefs 4 through 11, p47, and 39K (pp31), function either at the level of transcription or at that of mRNA stabilization. PMID:7815565

  9. Regulation of transcription of the human presenilin-1 gene by ets transcription factors and the p53 protooncogene.

    PubMed

    Pastorcic, M; Das, H K

    2000-11-10

    The expression of the human presenilin-1 cellular gene is suppressed by the p53 protooncogene. The rapid kinetic of the down-regulation has suggested that it may result from a primary mechanism. We show here that p53 also suppresses the transcription of a presenilin-1 promoter-chloramphenicol acetyltransferase reporter synthetic gene in transient infection assays in neuroblastoma (SK-N-SH) and hepatoma (HepG2) cell lines. Only a minimum promoter including sequences from -35 to + 6 from the transcription initiation is sufficient to confer down-regulation. We have previously defined a crucial DNA element controlling 90% of the expression of the gene within the same short area, and the identification of the transcription factors involved should also provide insights into the regulation of PS1 by p53. This region contains an Ets transcription factor binding motif, and a 2-base pair alteration within the core sequence (GGAA to TTAA) of the Ets consensus also reduced transcription by more than 90%. We now show that Ets1 and Ets2 indeed transactivate a PS1 promoter-chloramphenicol acetyltransferase reporter including the (-35 to +6) fragment. Furthermore, in vitro translated Ets2 binds specifically to the -10 Ets motif in electrophoretic mobility shift assays. Therefore, Ets1/2 factors bind specifically to the -10 Ets element and activate PS1 transcription. We also show that the coactivator p300 enhances the activation by Ets1 and Ets2 as well as the repression by p53. p300 is known to interact with p53 as well as with Ets1 and Ets2. We show that p53 does not bind directly to the PS1 promoter. Hence the repression of PS1 transcription by p53 is likely to be mediated through protein-protein interactions. PMID:10942770

  10. Cohesin and Polycomb Proteins Functionally Interact to Control Transcription at Silenced and Active Genes

    PubMed Central

    Schaaf, Cheri A.; Misulovin, Ziva; Gause, Maria; Koenig, Amanda; Gohara, David W.; Watson, Audrey; Dorsett, Dale

    2013-01-01

    Cohesin is crucial for proper chromosome segregation but also regulates gene transcription and organism development by poorly understood mechanisms. Using genome-wide assays in Drosophila developing wings and cultured cells, we find that cohesin functionally interacts with Polycomb group (PcG) silencing proteins at both silenced and active genes. Cohesin unexpectedly facilitates binding of Polycomb Repressive Complex 1 (PRC1) to many active genes, but their binding is mutually antagonistic at silenced genes. PRC1 depletion decreases phosphorylated RNA polymerase II and mRNA at many active genes but increases them at silenced genes. Depletion of cohesin reduces long-range interactions between Polycomb Response Elements in the invected-engrailed gene complex where it represses transcription. These studies reveal a previously unrecognized role for PRC1 in facilitating productive gene transcription and provide new insights into how cohesin and PRC1 control development. PMID:23818863

  11. Regulation of Nitrogenase Gene Expression by Transcript Stability in the Cyanobacterium Anabaena variabilis

    PubMed Central

    Pratte, Brenda S.

    2014-01-01

    The nitrogenase gene cluster in cyanobacteria has been thought to comprise multiple operons; however, in Anabaena variabilis, the promoter for the first gene in the cluster, nifB1, appeared to be the primary promoter for the entire nif cluster. The structural genes nifHDK1 were the most abundant transcripts; however, their abundance was not controlled by an independent nifH1 promoter, but rather, by RNA processing, which produced a very stable nifH1 transcript and a moderately stable nifD1 transcript. There was also no separate promoter for nifEN1. In addition to the nifB1 promoter, there were weak promoters inside the nifU1 gene and inside the nifE1 gene, and both promoters were heterocyst specific. In an xisA mutant, which effectively separated promoters upstream of an 11-kb excision element in nifD1 from the downstream genes, the internal nifE1 promoter was functional. Transcription of the nif1 genes downstream of the 11-kb element, including the most distant genes, hesAB1 and fdxH1, was reduced in the xisA mutant, indicating that the nifB1 promoter contributed to their expression. However, with the exception of nifK1 and nifE1, which had no expression, the downstream genes showed low to moderate levels of transcription in the xisA mutant. The hesA1 gene also had a promoter, but the fdxH gene had a processing site just upstream of the gene. The processing of transcripts at sites upstream of nifH1 and fdxH1 correlated with increased stability of these transcripts, resulting in greater amounts than transcripts that were not close to processing sites. PMID:25092030

  12. Regulation of nitrogenase gene expression by transcript stability in the cyanobacterium Anabaena variabilis.

    PubMed

    Pratte, Brenda S; Thiel, Teresa

    2014-10-01

    The nitrogenase gene cluster in cyanobacteria has been thought to comprise multiple operons; however, in Anabaena variabilis, the promoter for the first gene in the cluster, nifB1, appeared to be the primary promoter for the entire nif cluster. The structural genes nifHDK1 were the most abundant transcripts; however, their abundance was not controlled by an independent nifH1 promoter, but rather, by RNA processing, which produced a very stable nifH1 transcript and a moderately stable nifD1 transcript. There was also no separate promoter for nifEN1. In addition to the nifB1 promoter, there were weak promoters inside the nifU1 gene and inside the nifE1 gene, and both promoters were heterocyst specific. In an xisA mutant, which effectively separated promoters upstream of an 11-kb excision element in nifD1 from the downstream genes, the internal nifE1 promoter was functional. Transcription of the nif1 genes downstream of the 11-kb element, including the most distant genes, hesAB1 and fdxH1, was reduced in the xisA mutant, indicating that the nifB1 promoter contributed to their expression. However, with the exception of nifK1 and nifE1, which had no expression, the downstream genes showed low to moderate levels of transcription in the xisA mutant. The hesA1 gene also had a promoter, but the fdxH gene had a processing site just upstream of the gene. The processing of transcripts at sites upstream of nifH1 and fdxH1 correlated with increased stability of these transcripts, resulting in greater amounts than transcripts that were not close to processing sites. PMID:25092030

  13. Methods of Combinatorial Optimization to Reveal Factors Affecting Gene Length

    PubMed Central

    Bolshoy, Alexander; Tatarinova, Tatiana

    2012-01-01

    In this paper we present a novel method for genome ranking according to gene lengths. The main outcomes described in this paper are the following: the formulation of the genome ranking problem, presentation of relevant approaches to solve it, and the demonstration of preliminary results from prokaryotic genomes ordering. Using a subset of prokaryotic genomes, we attempted to uncover factors affecting gene length. We have demonstrated that hyperthermophilic species have shorter genes as compared with mesophilic organisms, which probably means that environmental factors affect gene length. Moreover, these preliminary results show that environmental factors group together in ranking evolutionary distant species. PMID:23300345

  14. Inhibition of human insulin gene transcription and MafA transcriptional activity by the dual leucine zipper kinase

    PubMed Central

    Stahnke, Marie-Jeannette; Dickel, Corinna; Schröder, Sabine; Kaiser, Diana; Blume, Roland; Stein, Roland; Pouponnot, Celio; Oetjen, Elke

    2016-01-01

    Insulin biosynthesis is an essential β-cell function and inappropriate insulin secretion and biosynthesis contribute to the pathogenesis of diabetes mellitus type 2. Previous studies showed that the dual leucine zipper kinase (DLK) induces β-cell apoptosis. Since β-cell dysfunction precedes β-cell loss, in the present study the effect of DLK on insulin gene transcription was investigated in the HIT-T15 β-cell line. Downregulation of endogenous DLK increased whereas overexpression of DLK decreased human insulin gene transcription. 5′- and 3′-deletion human insulin promoter analyses resulted in the identification of a DLK responsive element that mapped to the DNA binding-site for the β-cell specific transcription factor MafA. Overexpression of DLK wild-type but not its kinase-dead mutant inhibited MafA transcriptional activity conferred by its transactivation domain. Furthermore, in the non-β-cell line JEG DLK inhibited MafA overexpression-induced human insulin promoter activity. Overexpression of MafA and DLK or its kinase-dead mutant into JEG cells revealed that DLK but not its mutant reduced MafA protein content. Inhibition of the down-stream DLK kinase c-Jun N-terminal kinase (JNK) by SP600125 attenuated DLK-induced MafA loss. Furthermore, mutation of the serine 65 to alanine, shown to confer MafA protein stability, increased MafA-dependent insulin gene transcription and prevented DLK-induced MafA loss in JEG cells. These data suggest that DLK by activating JNK triggers the phosphorylation and degradation of MafA thereby attenuating insulin gene transcription. Given the importance of MafA for β-cell function, the inhibition of DLK might preserve β-cell function and ultimately retard the development of diabetes mellitus type 2. PMID:24726898

  15. Inhibition of human insulin gene transcription and MafA transcriptional activity by the dual leucine zipper kinase.

    PubMed

    Stahnke, Marie-Jeannette; Dickel, Corinna; Schröder, Sabine; Kaiser, Diana; Blume, Roland; Stein, Roland; Pouponnot, Celio; Oetjen, Elke

    2014-09-01

    Insulin biosynthesis is an essential β-cell function and inappropriate insulin secretion and biosynthesis contribute to the pathogenesis of diabetes mellitus type 2. Previous studies showed that the dual leucine zipper kinase (DLK) induces β-cell apoptosis. Since β-cell dysfunction precedes β-cell loss, in the present study the effect of DLK on insulin gene transcription was investigated in the HIT-T15 β-cell line. Downregulation of endogenous DLK increased whereas overexpression of DLK decreased human insulin gene transcription. 5'- and 3'-deletion human insulin promoter analyses resulted in the identification of a DLK responsive element that mapped to the DNA binding-site for the β-cell specific transcription factor MafA. Overexpression of DLK wild-type but not its kinase-dead mutant inhibited MafA transcriptional activity conferred by its transactivation domain. Furthermore, in the non-β-cell line JEG DLK inhibited MafA overexpression-induced human insulin promoter activity. Overexpression of MafA and DLK or its kinase-dead mutant into JEG cells revealed that DLK but not its mutant reduced MafA protein content. Inhibition of the down-stream DLK kinase c-Jun N-terminal kinase (JNK) by SP600125 attenuated DLK-induced MafA loss. Furthermore, mutation of the serine 65 to alanine, shown to confer MafA protein stability, increased MafA-dependent insulin gene transcription and prevented DLK-induced MafA loss in JEG cells. These data suggest that DLK by activating JNK triggers the phosphorylation and degradation of MafA thereby attenuating insulin gene transcription. Given the importance of MafA for β-cell function, the inhibition of DLK might preserve β-cell function and ultimately retard the development of diabetes mellitus type 2. PMID:24726898

  16. Gene activation by metazoan enhancers: Diverse mechanisms stimulate distinct steps of transcription.

    PubMed

    Beagrie, Robert A; Pombo, Ana

    2016-09-01

    Enhancers can stimulate transcription by a number of different mechanisms which control different stages of the transcription cycle of their target genes, from recruitment of the transcription machinery to elongation by RNA polymerase. These mechanisms may not be mutually exclusive, as a single enhancer may act through different pathways by binding multiple transcription factors. Multiple enhancers may also work together to regulate transcription of a shared target gene. Most of the evidence supporting different enhancer mechanisms comes from the study of single genes, but new high-throughput experimental frameworks offer the opportunity to integrate and generalize disparate mechanisms identified at single genes. This effort is especially important if we are to fully understand how sequence variation within enhancers contributes to human disease. PMID:27452946

  17. Stimulus-induced modulation of transcriptional bursting in a single mammalian gene.

    PubMed

    Molina, Nacho; Suter, David M; Cannavo, Rosamaria; Zoller, Benjamin; Gotic, Ivana; Naef, Félix

    2013-12-17

    Mammalian genes are often transcribed discontinuously as short bursts of RNA synthesis followed by longer silent periods. However, how these "on" and "off" transitions, together with the burst sizes, are modulated in single cells to increase gene expression upon stimulation is poorly characterized. By combining single-cell time-lapse luminescence imaging with stochastic modeling of the time traces, we quantified the transcriptional responses of the endogenous connective tissue growth factor gene to different physiological stimuli: serum and TGF-β1. Both stimuli caused a rapid and acute increase in burst sizes. Whereas TGF-β1 showed prolonged transcriptional activation mediated by an increase of transcription rate, serum stimulation resulted in a large and temporally tight first transcriptional burst, followed by a refractory period in the range of hours. Our study thus reveals how different physiological stimuli can trigger kinetically distinct transcriptional responses of the same gene. PMID:24297917

  18. NanoScript: A Nanoparticle-Based Artificial Transcription Factor for Effective Gene Regulation

    PubMed Central

    2015-01-01

    Transcription factor (TF) proteins are master regulators of transcriptional activity and gene expression. TF-based gene regulation is a promising approach for many biological applications; however, several limitations hinder the full potential of TFs. Herein, we developed an artificial, nanoparticle-based transcription factor, termed NanoScript, which is designed to mimic the structure and function of TFs. NanoScript was constructed by tethering functional peptides and small molecules called synthetic transcription factors, which mimic the individual TF domains, onto gold nanoparticles. We demonstrate that NanoScript localizes within the nucleus and initiates transcription of a reporter plasmid by over 15-fold. Moreover, NanoScript can effectively transcribe targeted genes on endogenous DNA in a nonviral manner. Because NanoScript is a functional replica of TF proteins and a tunable gene-regulating platform, it has great potential for various stem cell applications. PMID:25133310

  19. Expression of multiple tfb genes in different Halobacterium salinarum strains and interaction of TFB with transcriptional activator GvpE.

    PubMed

    Bleiholder, Anne; Frommherz, Regina; Teufel, Katharina; Pfeifer, Felicitas

    2012-04-01

    Halobacterium salinarum NRC-1 contains multiple TBP and TFB proteins required for the recruitment of RNA polymerase for transcription initiation. The presence and the expression of genes encoding TFB were investigated in the two Hbt. salinarum strains NRC-1 and PHH1 and the mutant strain PHH4. The plasmid-encoded tfbC and tfbE genes of NRC-1 were lacking in PHH1 and PHH4. The 5'-end of the tfbF transcript was determined and contained a 5'-untranslated region of 39 nucleotides able to form a stem-loop structure. The expression of these tfb genes was studied in cultures growing at 15, 37°C and under heat shock conditions. Cold temperatures reduced growth and except for tfbF also the amounts of all tfb transcripts. However, the formation of gas vesicles increased in PHH1 and NRC-1. Heat shock reduced growth of PHH1 and NRC-1, but PHH4 was not affected. A 100-fold increase in tfbA and tfbB mRNA was observed in PHH1 and PHH4, whereas NRC-1 reduced the amounts of these transcripts and increased the expression of tfbG. All TFB proteins tested were able to interact with the transcription activator GvpE involved in gas vesicle formation that thus is able to recruit TFB to the gvp promoter. PMID:21969032

  20. Carotenoid genes transcriptional regulation for astaxanthin accumulation in fresh water unicellular alga Haematococcus pluvialis by gibberellin A3 (GA3).

    PubMed

    Gao, Zhengquan; Meng, Chunxiao; Gao, Hongzheng; Li, Yan; Zhang, Xiaowen; Xu, Dong; Zhou, Shitan; Liu, Banghui; Su, Yuanfeng; Ye, Naihao

    2013-12-01

    The fresh water unicellular alga Haematococcus pluvialis is a promising natural source of astaxanthin. The present study investigated the transcriptional expression of carotenoid genes for astaxanthin accumulation in H. pluvialis using real-time fluorescence quantitative PCR (qRT-PCR). With treatments of 20 and 40 mg/L of gibberllin A3 (GA3), five genes ipi-1, ipi-2, psy, pds and bkt2 were up-regulated with different expression profiles. GA20 (20 mg/L of GA3) treatment had a greater effect on transcriptional expression of bkt2 than on ipi-1 ipi-2, psy and pds (> 4-fold up-regulation). However, GA40 (40 mg/L of GA3) induced more transcriptional expression of ipi-2, psy and bkt2 than both ipi-1 and pds. The expression of lyc, crtR-B and crtO for astaxanthin biosynthesis was not affected by GA3 in H. piuvialis. In the presence of GA3, astaxanthin biosynthesis genes of ipi-1, pds and bkt2 were up-regulated at transcriptional level, psy at post-transcriptional level, whereas ipi-2 was up-regulated at both levels. The study could potentially lead to a scale application of exogenous GA3 in astaxanthin production with H. pluvialis just like GAs perform in increasing crops production and it would provide new insight about the multifunctional roles of carotenogenesis in response to GA3. PMID:24772980

  1. TGF-β stimulation in human and murine cells reveals commonly affected biological processes and pathways at transcription level

    PubMed Central

    2014-01-01

    Background The TGF-β signaling pathway is a fundamental pathway in the living cell, which plays a key role in many central cellular processes. The complex and sometimes contradicting mechanisms by which TGF-β yields phenotypic effects are not yet completely understood. In this study we investigated and compared the transcriptional response profile of TGF-β1 stimulation in different cell types. For this purpose, extensive experiments are performed and time-course microarray data are generated in human and mouse parenchymal liver cells, human mesenchymal stromal cells and mouse hematopoietic progenitor cells at different time points. We applied a panel of bioinformatics methods on our data to uncover common patterns in the dynamic gene expression response in respective cells. Results Our analysis revealed a quite variable and multifaceted transcriptional response profile of TGF-β1 stimulation, which goes far beyond the well-characterized classical TGF-β1 signaling pathway. Nonetheless, we could identify several commonly affected processes and signaling pathways across cell types and species. In addition our analysis suggested an important role of the transcription factor EGR1, which appeared to have a conserved influence across cell-types and species. Validation via an independent dataset on A549 lung adenocarcinoma cells largely confirmed our findings. Network analysis suggested explanations, how TGF-β1 stimulation could lead to the observed effects. Conclusions The analysis of dynamical transcriptional response to TGF-β treatment experiments in different human and murine cell systems revealed commonly affected biological processes and pathways, which could be linked to TGF-β1 via network analysis. This helps to gain insights about TGF-β pathway activities in these cell systems and its conserved interactions between the species and tissue types. PMID:24886091

  2. Selective inhibition of transcription of the Ets2 gene in prostate cancer cells by a triplex-forming oligonucleotide

    PubMed Central

    Carbone, Giuseppina M.; McGuffie, Eileen M.; Collier, Angela; Catapano, Carlo V.

    2003-01-01

    The transcription factor Ets2 has a role in cancer development and represents an attractive therapeutic target. In this study, we designed a triplex-forming oligonucleotide (TFO) directed to a homopurine:homopyrimidine sequence in the Ets2 promoter. Transcription factors of the Sp family bound to this sequence and mutation of the Sp1 site reduced Ets2 promoter activity. The Ets2-TFO had high binding affinity for the target sequence and inhibited binding of Sp1/Sp3 to the overlapping site. This effect occurred with a high degree of sequence specificity. Mismatched oligonucleotides did not inhibit Sp1/Sp3 binding and mutations in the target sequence that abolished triplex formation prevented inhibition of Sp1/Sp3 binding by the TFO. The Ets2-TFO inhibited Ets2 promoter activity and expression of the endogenous gene in prostate cancer cells at nanomolar concentrations. The TFO did not affect reporter constructs with mutations in the TFO binding site and promoters of non-targeted genes. Expression of non-targeted genes was also not affected in TFO-treated cells. Collectively, these data demonstrated that the anti-transcriptional activity of the Ets2-TFO was sequence- and target-specific, and ruled out alternative, non-triplex mediated mechanisms of action. This anti-transcriptional approach may be useful to examine the effects of selective downregulation of Ets2 expression and may have therapeutic applications. PMID:12560478

  3. Fur-mediated activation of gene transcription in the human pathogen Neisseria gonorrhoeae.

    PubMed

    Yu, Chunxiao; Genco, Caroline Attardo

    2012-04-01

    It is well established that the ferric uptake regulatory protein (Fur) functions as a transcriptional repressor in diverse microorganisms. Recent studies demonstrated that Fur also functions as a transcriptional activator. In this study we defined Fur-mediated activation of gene transcription in the sexually transmitted disease pathogen Neisseria gonorrhoeae. Analysis of 37 genes which were previously determined to be iron induced and which contained putative Fur boxes revealed that only 30 of these genes exhibited reduced transcription in a gonococcal fur mutant strain. Fur-mediated activation was established by examining binding of Fur to the putative promoter regions of 16 Fur-activated genes with variable binding affinities observed. Only ∼50% of the newly identified Fur-regulated genes bound Fur in vitro, suggesting that additional regulatory circuits exist which may function through a Fur-mediated indirect mechanism. The gonococcal Fur-activated genes displayed variable transcription patterns in a fur mutant strain, which correlated with the position of the Fur box in each (promoter) region. These results suggest that Fur-mediated direct transcriptional activation is fulfilled by multiple mechanisms involving either competing with a repressor or recruiting RNA polymerase. Collectively, our studies have established that gonococcal Fur functions as an activator of gene transcription through both direct and indirect mechanisms. PMID:22287521

  4. Polycistronic transcription of fused cassettes and identification of translation initiation signals in an unusual gene cassette array from Pseudomonas aeruginosa

    PubMed Central

    Fonseca, Érica L.; Vicente, Ana Carolina Paulo

    2015-01-01

    The gene cassettes found in class 1 integrons are generally promoterless units composed by an open reading frame (ORF), a short 5’ untranslated region (UTR) and a 3’ recombination site ( attC). Fused gene cassettes are generated by partial or total loss of the attC from the first cassette in an array, creating, in some cases, a fusion with the ORF from the next cassette. These structures are rare and little is known about their mechanisms of mobilization and expression. The aim of this study was to evaluate the dynamic of mobilization and transcription of the gcu14-bla GES-1 /aacA4 gene cassette array, which harbours a fused gene cassette represented by bla GES-1 /aacA4. The cassette array was analyzed by Northern blot and real-time reverse transcription-polymerase chain reaction (RT-PCR) in order to assess the transcription mechanism of bla GES-1 /aacA4 fused cassette. Also, inverse polymerase chain reactions (PCR) were performed to detect the free circular forms of gcu14, bla GES-1 and aacA4. The Northern blot and real time RT-PCR revealed a polycistronic transcription, in which the fused cassette bla GES-1 /aacA4 is transcribed as a unique gene, while gcu14 (with a canonical attC recombination site) has a monocistronic transcription. The gcu14 cassette, closer to the weak configuration of cassette promoter (PcW), had a higher transcription level than bla GES-1/ aacA4, indicating that the cassette position affects the transcript amounts. The presence of ORF-11 at attI1, immediately preceding gcu14, and of a Shine-Dalgarno sequence upstream bla GES-1/ aacA4 composes a scenario for the occurrence of array translation. Inverse PCR generated amplicons corresponding to gcu14, gcu14-aacA4 and gcu14-bla GES-1/ aacA4 free circular forms, but not to bla GES-1 and aacA4 alone, indicating that the GES-1 truncated attC is not substrate of integrase activity and that these genes are mobilized together as a unique cassette. This study was original in showing the

  5. Dexamethasone-mediated transcriptional regulation of rat carboxylesterase 2 gene.

    PubMed

    Hori, Takeshi; Jin, Liangjing; Fujii, Ayako; Furihata, Tomomi; Nagahara, Yuko; Chiba, Kan; Hosokawa, Masakiyo

    2012-07-01

    Rat carboxylesterase 2 (rCES2), which was previously identified as a methylprednisolone 21-hemisuccinate hydrolase, is highly inducible by dexamethasone in the liver. In the present study, we investigated the molecular mechanisms by which this induction occurs. Injection of dexamethasone (1 mg/kg weight) into rats resulted in increases in the expression of rCES2 mRNA in a time-dependent manner with a peak at 12 h after injection. In primary rat hepatocytes, the expression level of rCES2 mRNA was increased by treatment with 100 nM dexamethasone, and the increase was completely blocked in the presence of 10 µM mifepristone (RU-486), a potent inhibitor of glucocorticoid receptor (GR), or 10 µg/mL cycloheximide, a translation inhibitor. Luciferase assays revealed that 100 nM dexamethasone increased rCES2 promoter activities, although the effect of dexamethasone on the promoter activity was smaller than that on rCES2 mRNA expression. The increased activities were completely inhibited by treatment of the hepatocytes with 10 µM RU-486. Based on these results, it is concluded that dexamethasone enhances transcription of the rCES2 gene via GR in the rat liver and that the dexamethasone-mediated induction of rCES2 mRNA may be dependent on de novo protein synthesis. Our results provide clues to understanding what compounds induce rCES2. PMID:22235919

  6. Transcriptional regulation of bone sialoprotein gene expression by Osx.

    PubMed

    Yang, Ya; Huang, Yehong; Zhang, Li; Zhang, Chi

    2016-08-01

    Osteoporosis is the most common metabolic bone disease characterized by decreased bone mass, decreased bone strength, and increased risk of fracture. It is due to unbalance between bone formation and bone resorption. Bone formation is a complex process which involves the differentiation of mesenchymal stem cells to osteoblasts. Osteoblasts produce a characteristic extracellular collagenous matrix that subsequently becomes mineralized. Osterix (Osx) is an osteoblast-specific transcription factor required for osteoblast differentiation. Bone sialoprotein (Bsp) is a member of the SIBLING gene family. Expression of Bsp correlates with the differentiation of osteoblasts and the onset of mineralization. Our preliminary data showed that Bsp was abolished in Osx-null mice; however, the detailed mechanism of Osx regulation on Bsp is not fully understood. In this study, regulation of Bsp expression by Osx was further characterized. It was shown that overexpression of Osx led to Bsp upregulation. Inhibition of Osx by small interfering RNA resulted in Bsp downregulation in osteoblast. Transfection assay demonstrated that Osx was able to activate Bsp promoter reporter in a dose-dependent manner. To define minimal region of Bsp promoter activated by Osx, a series of deletion mutants of Bsp promoter were generated, and the minimal region was narrowed down to the proximal 100 bp. Point-mutagenesis studies showed that one GC-rich site was required for Bsp promoter activation by Osx. ChIP assays demonstrated that endogenous Osx associated with native Bsp promoter in primary osteoblasts. Our observations provide evidence that Osx targets Bsp expression directly. PMID:27261434

  7. A yeast transcription system for the 5S rRNA gene.

    PubMed Central

    van Keulen, H; Thomas, D Y

    1982-01-01

    A cell-free extract of yeast nuclei that can specifically transcribe cloned yeast 5S rRNA genes has been developed. Optima for transcription of 5S rDNA were determined and conditions of extract preparation leading to reproducible activities and specificities established. The major in vitro product has the same size and oligonucleotide composition as in vivo 5S rRNA. The in vitro transcription extract does not transcribe yeast tRNA genes. The extract does increase the transcription of tRNA genes packaged in chromatin. Images PMID:7145700

  8. Methoprene-tolerant 1 regulates gene transcription to maintain insect larval status.

    PubMed

    Zhao, Wen-Li; Liu, Chun-Yan; Liu, Wen; Wang, Di; Wang, Jin-Xing; Zhao, Xiao-Fan

    2014-08-01

    Insect molting and metamorphosis are regulated by two hormones: 20-hydroxyecdysone (20E) and juvenile hormone (JH). The hormone 20E regulates gene transcription via the nuclear receptor EcR to promote metamorphosis, whereas JH regulates gene transcription via its intracellular receptor methoprene-tolerant (Met) to prevent larval-pupal transition. However, the function and mechanism of Met in various insect developments are not well understood. We propose that Met1 plays a key role in maintaining larval status not only by promoting JH-responsive gene transcription but also by repressing 20E-responsive gene transcription in the Lepidopteran insect Helicoverpa armigera. Met1 protein is increased during feeding stage and decreased during molting and metamorphic stages. Met1 is upregulated by JH III and a low concentration of 20E independently, but is downregulated by a high concentration of 20E. Knockdown of Met1 in larvae causes precocious pupation, decrease in JH pathway gene expression, and increase in 20E pathway gene expression. Met1 interacts with heat shock protein 90 and binds to JH response element to regulate Krüppel homolog 1 transcription in JH III induction. Met1 interacts with ultraspiracle protein 1 (USP1) to repress 20E transcription complex EcRB1/USP1 formation and binding to ecdysone response element. These data indicate that JH via Met1 regulates JH pathway gene expression and represses 20E pathway gene expression to maintain the larval status. PMID:24872508

  9. RNase L Attenuates Mitogen-stimulated Gene Expression via Transcriptional and Post-transcriptional Mechanisms to Limit the Proliferative Response*

    PubMed Central

    Brennan-Laun, Sarah E.; Li, Xiao-Ling; Ezelle, Heather J.; Venkataraman, Thiagarajan; Blackshear, Perry J.; Wilson, Gerald M.; Hassel, Bret A.

    2014-01-01

    The cellular response to mitogens is tightly regulated via transcriptional and post-transcriptional mechanisms to rapidly induce genes that promote proliferation and efficiently attenuate their expression to prevent malignant growth. RNase L is an endoribonuclease that mediates diverse antiproliferative activities, and tristetraprolin (TTP) is a mitogen-induced RNA-binding protein that directs the decay of proliferation-stimulatory mRNAs. In light of their roles as endogenous proliferative constraints, we examined the mechanisms and functional interactions of RNase L and TTP to attenuate a mitogenic response. Mitogen stimulation of RNase L-deficient cells significantly increased TTP transcription and the induction of other mitogen-induced mRNAs. This regulation corresponded with elevated expression of serum-response factor (SRF), a master regulator of mitogen-induced transcription. RNase L destabilized the SRF transcript and formed a complex with SRF mRNA in cells providing a mechanism by which RNase L down-regulates SRF-induced genes. TTP and RNase L proteins interacted in cells suggesting that RNase L is directed to cleave TTP-bound RNAs as a mechanism of substrate specificity. Consistent with their concerted function in RNA turnover, the absence of either RNase L or TTP stabilized SRF mRNA, and a subset of established TTP targets was also regulated by RNase L. RNase L deficiency enhanced mitogen-induced proliferation demonstrating its functional role in limiting the mitogenic response. Our findings support a model of feedback regulation in which RNase L and TTP target SRF mRNA and SRF-induced transcripts. Accordingly, meta-analysis revealed an enrichment of RNase L and TTP targets among SRF-regulated genes suggesting that the RNase L/TTP axis represents a viable target to inhibit SRF-driven proliferation in neoplastic diseases. PMID:25301952

  10. RNase L attenuates mitogen-stimulated gene expression via transcriptional and post-transcriptional mechanisms to limit the proliferative response.

    PubMed

    Brennan-Laun, Sarah E; Li, Xiao-Ling; Ezelle, Heather J; Venkataraman, Thiagarajan; Blackshear, Perry J; Wilson, Gerald M; Hassel, Bret A

    2014-11-28

    The cellular response to mitogens is tightly regulated via transcriptional and post-transcriptional mechanisms to rapidly induce genes that promote proliferation and efficiently attenuate their expression to prevent malignant growth. RNase L is an endoribonuclease that mediates diverse antiproliferative activities, and tristetraprolin (TTP) is a mitogen-induced RNA-binding protein that directs the decay of proliferation-stimulatory mRNAs. In light of their roles as endogenous proliferative constraints, we examined the mechanisms and functional interactions of RNase L and TTP to attenuate a mitogenic response. Mitogen stimulation of RNase L-deficient cells significantly increased TTP transcription and the induction of other mitogen-induced mRNAs. This regulation corresponded with elevated expression of serum-response factor (SRF), a master regulator of mitogen-induced transcription. RNase L destabilized the SRF transcript and formed a complex with SRF mRNA in cells providing a mechanism by which RNase L down-regulates SRF-induced genes. TTP and RNase L proteins interacted in cells suggesting that RNase L is directed to cleave TTP-bound RNAs as a mechanism of substrate specificity. Consistent with their concerted function in RNA turnover, the absence of either RNase L or TTP stabilized SRF mRNA, and a subset of established TTP targets was also regulated by RNase L. RNase L deficiency enhanced mitogen-induced proliferation demonstrating its functional role in limiting the mitogenic response. Our findings support a model of feedback regulation in which RNase L and TTP target SRF mRNA and SRF-induced transcripts. Accordingly, meta-analysis revealed an enrichment of RNase L and TTP targets among SRF-regulated genes suggesting that the RNase L/TTP axis represents a viable target to inhibit SRF-driven proliferation in neoplastic diseases. PMID:25301952

  11. Heterogeneity in the Polarity of Nra Regulatory Effects on Streptococcal Pilus Gene Transcription and Virulence▿ †

    PubMed Central

    Luo, Feng; Lizano, Sergio; Bessen, Debra E.

    2008-01-01

    Transcription of several key virulence factors of Streptococcus pyogenes is under the control of Mga and Nra/RofA. In an M serotype 49 (M49) strain, Nra is a negative regulator of pilus gene transcription; also, Nra represses mga expression, leading to downregulation of the M protein surface fibril and secreted cysteine protease SpeB. In this report, the role of Nra in the virulence of an M53 classical skin strain was investigated. In contrast to the case for the M49 strain, Nra functions as a positive regulator of pilus gene transcription in the M53 strain, and inactivation of nra leads to loss of virulence in a humanized mouse model of superficial skin infection. Furthermore, Nra has no measurable effect on mga transcription in the M53 strain; this finding is further supported by a lack of detectable Nra effects on M protein- and SpeB-dependent phenotypes. Whereas MsmR is reported to activate nra and pilus gene transcription in the M49 strain, in the M53 strain it acts as a repressor of these genes. In both strains, MsmR and Nra form a feed-forward loop network motif for pilus gene transcription, but their effects have opposite signs. The findings demonstrate key strain-specific differences in the transcriptional circuitry governing virulence gene expression in S. pyogenes and its impact on pathogenesis. PMID:18347035

  12. The evolution of gene expression and binding specificity of the largest transcription factor family in primates

    PubMed Central

    Kapopoulou, Adamandia; Mathew, Lisha; Wong, Alex; Trono, Didier; Jensen, Jeffrey D.

    2016-01-01

    The KRAB-containing zinc finger (KRAB-ZF) proteins represent the largest family of transcription factors (TFs) in humans, yet for the great majority, their function and specific genomic target remain unknown. However, it has been shown that a large fraction of these genes arose from segmental duplications, and that they have expanded in gene and zinc finger number throughout vertebrate evolution. To determine whether this expansion is linked to selective pressures acting on different domains, we have manually curated all KRAB-ZF genes present in the human genome together with their orthologous genes in three closely related species and assessed the evolutionary forces acting at the sequence level as well as on their expression profiles. We provide evidence that KRAB-ZFs can be separated into two categories according to the polymorphism present in their DNA-contacting residues. Those carrying a nonsynonymous single nucleotide polymorphism (SNP) in their DNA-contacting amino acids exhibit significantly reduced expression in all tissues, have emerged in a recent lineage, and seem to be less strongly constrained evolutionarily than those without such a polymorphism. This work provides evidence for a link between age of the TF, as well as polymorphism in their DNA-contacting residues and expression levels—both of which may be jointly affected by selection. PMID:26593440

  13. RNAi-directed post transcriptional gene silencing of an Arabidopsis Myb transgene in tobacco

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The AtMyb90 gene encodes the 'production of anthocyanin pigment 2' (PAP2) transcription factor of Arabidopsis thaliana and is able to induce a visible hyper-pigmented phenotype when expressed in tobacco. Based upon this phenotype, we have used the AtMyb90 gene as a reporter gene to examine RNAi-dire...

  14. The grape berry-specific basic helix–loop–helix transcription factor VvCEB1 affects cell size

    PubMed Central

    Lecourieux, Fatma

    2013-01-01

    The development of fleshy fruits involves complex physiological and biochemical changes. After fertilization, fruit growth usually begins with cell division, continues with both cell division and expansion, allowing fruit set to occur, and ends with cell expansion only. In spite of the economical importance of grapevine, the molecular mechanisms controlling berry growth are not fully understood. The present work identified and characterized Vitis vinifera cell elongation bHLH protein (VvCEB1), a basic helix–loop–helix (bHLH) transcription factor controlling cell expansion in grape. VvCEB1 was expressed specifically in berry-expanding tissues with a maximum around veraison. The study of VvCEB1 promoter activity in tomato confirmed its specific fruit expression during the expansion phase. Overexpression of VvCEB1 in grape embryos showed that this protein stimulates cell expansion and affects the expression of genes involved in cell expansion, including genes of auxin metabolism and signalling. Taken together, these data show that VvCEB1 is a fruit-specific bHLH transcription factor involved in grape berry development. PMID:23314819

  15. Identification of liver receptor homolog-1 as a novel regulator of apolipoprotein AI gene transcription.

    PubMed

    Delerive, Philippe; Galardi, Cristin M; Bisi, John E; Nicodeme, Edwige; Goodwin, Bryan

    2004-10-01

    The orphan nuclear receptor liver receptor homolog-1 (LRH-1) has been reported to play a role in bile acid biosynthesis and reverse cholesterol transport. In this study, we examined the role of LRH-1 in the regulation of the apolipoprotein AI (APOAI) gene. Using RNA interference and adenovirus-mediated overexpression, we show that LRH-1 directly regulates APOAI gene transcription. Transient transfection experiments and EMSAs revealed that LRH-1 directly regulates APOAI transcription by binding to an LRH-1 response element located in the proximal APOAI promoter region. Chromatin immunoprecipitation experiments revealed that LRH-1 binds to the human APO AI promoter in vivo. Finally, we show that the transcriptional repressor SHP (small heterodimer partner) suppressed APOAI gene expression by inhibiting LRH-1 transcriptional activity. Taken together, our results demonstrate that LRH-1 is a novel regulator of APOAI transcription and underscore the role of this receptor in cholesterol homeostasis. PMID:15218078

  16. Identification of a Novel Reference Gene for Apple Transcriptional Profiling under Postharvest Conditions

    PubMed Central

    Storch, Tatiane Timm; Pegoraro, Camila; Finatto, Taciane; Quecini, Vera; Rombaldi, Cesar Valmor; Girardi, César Luis

    2015-01-01

    Reverse Transcription quantitative PCR (RT-qPCR) is one of the most important techniques for gene expression profiling due to its high sensibility and reproducibility. However, the reliability of the results is highly dependent on data normalization, performed by comparisons between the expression profiles of the genes of interest against those of constitutively expressed, reference genes. Although the technique is widely used in fruit postharvest experiments, the transcription stability of reference genes has not been thoroughly investigated under these experimental conditions. Thus, we have determined the transcriptional profile, under these conditions, of three genes commonly used as reference—ACTIN (MdACT), PROTEIN DISULPHIDE ISOMERASE (MdPDI) and UBIQUITIN-CONJUGATING ENZYME E2 (MdUBC)—along with two novel candidates—HISTONE 1 (MdH1) and NUCLEOSSOME ASSEMBLY 1 PROTEIN (MdNAP1). The expression profile of the genes was investigated throughout five experiments, with three of them encompassing the postharvest period and the other two, consisting of developmental and spatial phases. The transcriptional stability was comparatively investigated using four distinct software packages: BestKeeper, NormFinder, geNorm and DataAssist. Gene ranking results for transcriptional stability were similar for the investigated software packages, with the exception of BestKeeper. The classic reference gene MdUBC ranked among the most stably transcribed in all investigated experimental conditions. Transcript accumulation profiles for the novel reference candidate gene MdH1 were stable throughout the tested conditions, especially in experiments encompassing the postharvest period. Thus, our results present a novel reference gene for postharvest experiments in apple and reinforce the importance of checking the transcription profile of reference genes under the experimental conditions of interest. PMID:25774904

  17. Alzheimer amyloid peptide aβ42 regulates gene expression of transcription and growth factors.

    PubMed

    Barucker, Christian; Sommer, Anette; Beckmann, Georg; Eravci, Murat; Harmeier, Anja; Schipke, Carola G; Brockschnieder, Damian; Dyrks, Thomas; Althoff, Veit; Fraser, Paul E; Hazrati, Lili-Naz; George-Hyslop, Peter St; Breitner, John C S; Peters, Oliver; Multhaup, Gerhard

    2015-01-01

    The pathogenesis of Alzheimer's disease (AD) is characterized by the aggregation of amyloid-β (Aβ) peptides leading to deposition of senile plaques and a progressive decline of cognitive functions, which currently remains the main criterion for its diagnosis. Robust biomarkers for AD do not yet exist, although changes in the cerebrospinal fluid levels of tau and Aβ represent promising candidates in addition to brain imaging and genetic risk profiling. Although concentrations of soluble Aβ42 correlate with symptoms of AD, less is known about the biological activities of Aβ peptides which are generated from the amyloid-β protein precursor. An unbiased DNA microarray study showed that Aβ42, at sub-lethal concentrations, specifically increases expression of several genes in neuroblastoma cells, notably the insulin-like growth factor binding proteins 3 and 5 (IGFBP3/5), the transcription regulator inhibitor of DNA binding, and the transcription factor Lim only domain protein 4. Using qRT-PCR, we confirmed that mRNA levels of the identified candidate genes were exclusively increased by the potentially neurotoxic Aβ42 wild-type peptide, as both the less toxic Aβ40 and a non-toxic substitution peptide Aβ42 G33A did not affect mRNA levels. In vivo immunohistochemistry revealed a corresponding increase in both hippocampal and cortical IGFBP5 expression in an AD mouse model. Proteomic analyses of human AD cerebrospinal fluid displayed increased in vivo concentrations of IGFBPs. IGFBPs and transcription factors, as identified here, are modulated by soluble Aβ42 and may represent useful early biomarkers. PMID:25318543

  18. Timing of flagellar gene expression in the Caulobacter cell cycle is determined by a transcriptional cascade of positive regulatory genes.

    PubMed Central

    Ohta, N; Chen, L S; Mullin, D A; Newton, A

    1991-01-01

    The Caulobacter crescentus flagellar (fla) genes are organized in a regulatory hierarchy in which genes at each level are required for expression of those at the next lower level. To determine the role of this hierarchy in the timing of fla gene expression, we have examined the organization and cell cycle regulation of genes located in the hook gene cluster. As shown here, this cluster is organized into four multicistronic transcription units flaN, flbG, flaO, and flbF that contain fla genes plus a fifth transcription unit II.1 of unknown function. Transcription unit II.1 is regulated independently of the fla gene hierarchy, and it is expressed with a unique pattern of periodicity very late in the cell cycle. The flaN, flbG, and flaO operons are all transcribed periodically, and flaO, which is near the top of the hierarchy and required in trans for the activation of flaN and flbG operons, is expressed earlier in the cell cycle than the other two transcription units. We have shown that delaying flaO transcription by fusing it to the II.1 promoter also delayed the subsequent expression of the flbG operon and the 27- and 25-kDa flagellin genes that are at the bottom of the regulatory hierarchy. Thus, the sequence and timing of fla gene expression in the cell cycle are determined in large measure by the positions of these genes in the regulatory hierarchy. These results also suggest that periodic transcription is a general feature of fla gene expression in C. crescentus. Images PMID:1847367

  19. Control of Rta expression critically determines transcription of viral and cellular genes following gammaherpesvirus infection.

    PubMed

    Hair, James R; Lyons, Paul A; Smith, Kenneth G C; Efstathiou, Stacey

    2007-06-01

    The replication and transcriptional activator (Rta), encoded by ORF50 of gammaherpesviruses, initiates the lytic cycle of gene expression; therefore understanding the impact of Rta on viral and cellular gene expression is key to elucidating the transcriptional events governing productive infection and reactivation from latency. To this end, the impact of altering Rta transcription on viral and cellular gene expression was studied in the context of a whole virus infection. Recombinant murine gammaherpesvirus (MHV)-68 engineered to overexpress Rta greatly accelerated expression of specific lytic cycle ORFs, but repressed transcription of the major latency gene, ORF73. Increased expression of Rta accelerated the dysregulation in transcription of specific cellular genes when compared with cells infected with wild-type and revertant viruses. A subset of cellular genes was dysregulated only in cells infected with Rta-overexpressing virus, and never in those infected with non-overexpressing viruses. These data highlight the critical role of Rta abundance in governing viral and cellular gene transcription, and demonstrate the importance of understanding how the relative expression of ORF50 during the virus life cycle impacts on these processes. PMID:17485528

  20. Control of Rta expression critically determines transcription of viral and cellular genes following gammaherpesvirus infection

    PubMed Central

    Hair, James R.; Lyons, Paul A.; Smith, Kenneth G. C.; Efstathiou, Stacey

    2007-01-01

    The replication and transcriptional activator (Rta), encoded by ORF50 of gammaherpesviruses, initiates the lytic cycle of gene expression; therefore understanding the impact of Rta on viral and cellular gene expression is key to elucidating the transcriptional events governing productive infection and reactivation from latency. To this end, the impact of altering Rta transcription on viral and cellular gene expression was studied in the context of a whole virus infection. Recombinant murine gammaherpesvirus (MHV)-68 engineered to overexpress Rta greatly accelerated expression of specific lytic cycle ORFs, but repressed transcription of the major latency gene, ORF73. Increased expression of Rta accelerated the dysregulation in transcription of specific cellular genes when compared with cells infected with wild-type and revertant viruses. A subset of cellular genes was dysregulated only in cells infected with Rta-overexpressing virus, and never in those infected with non-overexpressing viruses. These data highlight the critical role of Rta abundance in governing viral and cellular gene transcription, and demonstrate the importance of understanding how the relative expression of ORF50 during the virus life cycle impacts on these processes. PMID:17485528

  1. Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors.

    PubMed

    Mercer, Andrew C; Gaj, Thomas; Sirk, Shannon J; Lamb, Brian M; Barbas, Carlos F

    2014-10-17

    The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks. PMID:24251925

  2. The Transcriptional Response in Human Umbilical Vein Endothelial Cells Exposed to Insulin: A Dynamic Gene Expression Approach

    PubMed Central

    Di Camillo, Barbara; Sanavia, Tiziana; Iori, Elisabetta; Bronte, Vincenzo; Roncaglia, Enrica; Maran, Alberto; Avogaro, Angelo; Toffolo, Gianna; Cobelli, Claudio

    2010-01-01

    Background In diabetes chronic hyperinsulinemia contributes to the instability of the atherosclerotic plaque and stimulates cellular proliferation through the activation of the MAP kinases, which in turn regulate cellular proliferation. However, it is not known whether insulin itself could increase the transcription of specific genes for cellular proliferation in the endothelium. Hence, the characterization of transcriptional modifications in endothelium is an important step for a better understanding of the mechanism of insulin action and the relationship between endothelial cell dysfunction and insulin resistance. Methodology and principal findings The transcriptional response of endothelial cells in the 440 minutes following insulin stimulation was monitored using microarrays and compared to a control condition. About 1700 genes were selected as differentially expressed based on their treated minus control profile, thus allowing the detection of even small but systematic changes in gene expression. Genes were clustered in 7 groups according to their time expression profile and classified into 15 functional categories that can support the biological effects of insulin, based on Gene Ontology enrichment analysis. In terms of endothelial function, the most prominent processes affected were NADH dehydrogenase activity, N-terminal myristoylation domain binding, nitric-oxide synthase regulator activity and growth factor binding. Pathway-based enrichment analysis revealed “Electron Transport Chain” significantly enriched. Results were validated on genes belonging to “Electron Transport Chain” pathway, using quantitative RT-PCR. Conclusions As far as we know, this is the first systematic study in the literature monitoring transcriptional response to insulin in endothelial cells, in a time series microarray experiment. Since chronic hyperinsulinemia contributes to the instability of the atherosclerotic plaque and stimulates cellular proliferation, some of the

  3. Sumoylation of Rap1 mediates the recruitment of TFIID to promote transcription of ribosomal protein genes.

    PubMed

    Chymkowitch, Pierre; Nguéa, Aurélie P; Aanes, Håvard; Koehler, Christian J; Thiede, Bernd; Lorenz, Susanne; Meza-Zepeda, Leonardo A; Klungland, Arne; Enserink, Jorrit M

    2015-06-01

    Transcription factors are abundant Sumo targets, yet the global distribution of Sumo along the chromatin and its physiological relevance in transcription are poorly understood. Using Saccharomyces cerevisiae, we determined the genome-wide localization of Sumo along the chromatin. We discovered that Sumo-enriched genes are almost exclusively involved in translation, such as tRNA genes and ribosomal protein genes (RPGs). Genome-wide expression analysis showed that Sumo positively regulates their transcription. We also discovered that the Sumo consensus motif at RPG promoters is identical to the DNA binding motif of the transcription factor Rap1. We demonstrate that Rap1 is a molecular target of Sumo and that sumoylation of Rap1 is important for cell viability. Furthermore, Rap1 sumoylation promotes recruitment of the basal transcription machinery, and sumoylation of Rap1 cooperates with the target of rapamycin kinase complex 1 (TORC1) pathway to promote RPG transcription. Strikingly, our data reveal that sumoylation of Rap1 functions in a homeostatic feedback loop that sustains RPG transcription during translational stress. Taken together, Sumo regulates the cellular translational capacity by promoting transcription of tRNA genes and RPGs. PMID:25800674

  4. A well-conserved Plasmodium falciparum var gene shows an unusual stage-specific transcript pattern

    PubMed Central

    Kyes, Sue A.; Christodoulou, Zoe; Raza, Ahmed; Horrocks, Paul; Pinches, Robert; Rowe, J. Alexandra; Newbold, Chris I.

    2010-01-01

    Summary The var multicopy gene family encodes Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) variant antigens, which, through their ability to adhere to a variety of host receptors, are thought to be important virulence factors. The predominant expression of a single cytoadherent PfEMP1 type on an infected red blood cell, and the switching between different PfEMP1 types to evade host protective antibody responses, are processes thought to be controlled at the transcriptional level. Contradictory data have been published on the timing of var gene transcription. Reverse transcription-polymerase chain reaction (RT-PCR) data suggested that transcription of the predominant var gene occurs in the later (pigmented trophozoite) stages, whereas Northern blot data indicated such transcripts only in early (ring) stages. We investigated this discrepancy by Northern blot, with probes covering a diverse var gene repertoire. We confirm that almost all var transcript types were detected only in ring stages. However, one type, the well-conserved varCSA transcript, was present constitutively in different laboratory parasites and does not appear to undergo antigenic variation. Although varCSA has been shown to encode a chondroitin sulphate A (CSA)-binding PfEMP1, we find that the presence of full-length varCSA transcripts does not correlate with the CSA-binding phenotype. PMID:12787360

  5. Mutations in the su(s) gene affect RNA processing in Drosophila melanogaster.

    PubMed Central

    Geyer, P K; Chien, A J; Corces, V G; Green, M M

    1991-01-01

    We have studied the effect of mutations in the suppressor of sable [su(s)] gene on P element-induced yellow alleles. Two independent mutations tested, y76d28 and y1#7, contain a 1.1-kilobase (kb) P element inserted in the 5' transcribed untranslated portion of the yellow gene. Sequences responsible for the y1#7 mutation are inserted in the same transcriptional orientation as yellow and cannot be processed by splicing, and this mutation is not suppressed by su(s) mutations. P element sequences are located in a transcriptional orientation opposite to that of the yellow gene in y76d28; these sequences can be spliced from a composite P element-yellow mRNA, resulting in low accumulation of a functional 1.9-kb yellow transcript. The levels of both the putative precursor P element-yellow RNA and the 1.9-kb yellow transcript increase in y76d28 su(s) flies, suggesting that mutations in su(s) do not affect the efficiency of splicing of the P element sequences. Analysis of y76d28 cDNAs isolated from flies carrying a wild-type or mutant su(s) gene demonstrates that the choice of splice junctions to process P element sequences is unchanged in these different backgrounds, suggesting that mutations in su(s) do not affect the selection of donor and acceptor splice sites. We propose that the su(s) protein functions to control the stability of unprocessed RNA during the splicing reaction. Images PMID:1714588

  6. Increased Transcript Complexity in Genes Associated with Chronic Obstructive Pulmonary Disease.

    PubMed

    Lackey, Lela; McArthur, Evonne; Laederach, Alain

    2015-01-01

    Genome-wide association studies aim to correlate genotype with phenotype. Many common diseases including Type II diabetes, Alzheimer's, Parkinson's and Chronic Obstructive Pulmonary Disease (COPD) are complex genetic traits with hundreds of different loci that are associated with varied disease risk. Identifying common features in the genes associated with each disease remains a challenge. Furthermore, the role of post-transcriptional regulation, and in particular alternative splicing, is still poorly understood in most multigenic diseases. We therefore compiled comprehensive lists of genes associated with Type II diabetes, Alzheimer's, Parkinson's and COPD in an attempt to identify common features of their corresponding mRNA transcripts within each gene set. The SERPINA1 gene is a well-recognized genetic risk factor of COPD and it produces 11 transcript variants, which is exceptional for a human gene. This led us to hypothesize that other genes associated with COPD, and complex disorders in general, are highly transcriptionally diverse. We found that COPD-associated genes have a statistically significant enrichment in transcript complexity stemming from a disproportionately high level of alternative splicing, however, Type II Diabetes, Alzheimer's and Parkinson's disease genes were not significantly enriched. We also identified a subset of transcriptionally complex COPD-associated genes (~40%) that are differentially expressed between mild, moderate and severe COPD. Although the genes associated with other lung diseases are not extensively documented, we found preliminary data that idiopathic pulmonary disease genes, but not cystic fibrosis modulators, are also more transcriptionally complex. Interestingly, complex COPD transcripts are more often the product of alternative acceptor site usage. To verify the biological importance of these alternative transcripts, we used RNA-sequencing analyses to determine that COPD-associated genes are frequently expressed in

  7. Increased Transcript Complexity in Genes Associated with Chronic Obstructive Pulmonary Disease

    PubMed Central

    Lackey, Lela; McArthur, Evonne; Laederach, Alain

    2015-01-01

    Genome-wide association studies aim to correlate genotype with phenotype. Many common diseases including Type II diabetes, Alzheimer’s, Parkinson’s and Chronic Obstructive Pulmonary Disease (COPD) are complex genetic traits with hundreds of different loci that are associated with varied disease risk. Identifying common features in the genes associated with each disease remains a challenge. Furthermore, the role of post-transcriptional regulation, and in particular alternative splicing, is still poorly understood in most multigenic diseases. We therefore compiled comprehensive lists of genes associated with Type II diabetes, Alzheimer’s, Parkinson’s and COPD in an attempt to identify common features of their corresponding mRNA transcripts within each gene set. The SERPINA1 gene is a well-recognized genetic risk factor of COPD and it produces 11 transcript variants, which is exceptional for a human gene. This led us to hypothesize that other genes associated with COPD, and complex disorders in general, are highly transcriptionally diverse. We found that COPD-associated genes have a statistically significant enrichment in transcript complexity stemming from a disproportionately high level of alternative splicing, however, Type II Diabetes, Alzheimer’s and Parkinson’s disease genes were not significantly enriched. We also identified a subset of transcriptionally complex COPD-associated genes (~40%) that are differentially expressed between mild, moderate and severe COPD. Although the genes associated with other lung diseases are not extensively documented, we found preliminary data that idiopathic pulmonary disease genes, but not cystic fibrosis modulators, are also more transcriptionally complex. Interestingly, complex COPD transcripts are more often the product of alternative acceptor site usage. To verify the biological importance of these alternative transcripts, we used RNA-sequencing analyses to determine that COPD-associated genes are frequently

  8. Selenite transiently represses transcription of photosynthesis-related genes in potato leaves.

    PubMed

    Poggi, Valeria; Del Vescovo, Valerio; Di Sanza, Claudio; Negri, Rodolfo; Hochkoeppler, Alejandro

    2008-01-01

    A striking response of potato leaves to aspersion with selenite was observed at the transcriptional level by means of cDNA microarrays analysis. This response is characterized by a general transient repression of genes coding for components of photosynthetic systems and of other light-regulated genes. In particular, maximal repression was observed 8 h after selenite aspersion, while 24 h after the treatment a complete recovery of normal transcriptional levels was detected. Another general feature of the transcriptional response to selenite is represented by the transcriptional induction of genes related to amino acid metabolism, and to stress defense; interestingly, two genes coding for glutathione S-transferases were found early-induced upon selenite treatment. PMID:17846914

  9. VITELLOGENIN GENE TRANSCRIPTION AS AN INDICATOR OF EXPOSURE TO 17-ALPHA-ETHYNYLESTRADIOL IN FATHEAD MINNOWS

    EPA Science Inventory

    Environmentally persistent chemicals that functionally mimic estrogen are ubiquitous in surface waters and have been shown to effect reproductive health of species living in these habitats. Toxicant induced transcription of specific genes is a sensitive indicator of exposure and ...

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

    PubMed

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

    2016-06-01

    Depression is a complex, heterogeneous disorder and a leading contributor to the global burden of disease. Most previous research has focused on individual brain regions and genes contributing to depression. However, emerging evidence in humans and animal models suggests that dysregulated circuit function and gene expression across multiple brain regions drive depressive phenotypes. Here, we performed RNA sequencing on four brain regions from control animals and those susceptible or resilient to chronic social defeat stress at multiple time points. We employed an integrative network biology approach to identify transcriptional networks and key driver genes that regulate susceptibility to depressive-like symptoms. Further, we validated in vivo several key drivers and their associated transcriptional networks that regulate depression susceptibility and confirmed their functional significance at the levels of gene transcription, synaptic regulation, and behavior. Our study reveals novel transcriptional networks that control stress susceptibility and offers fundamentally new leads for antidepressant drug discovery. PMID:27181059

  11. Transcriptional Regulation of Fucosyltransferase 1 Gene Expression in Colon Cancer Cells

    PubMed Central

    Taniuchi, Fumiko; Higai, Koji; Tanaka, Tomomi; Azuma, Yutaro; Matsumoto, Kojiro

    2013-01-01

    The α1,2-fucosyltransferase I (FUT1) enzyme is important for the biosynthesis of H antigens, Lewis B, and Lewis Y. In this study, we clarified the transcriptional regulation of FUT1 in the DLD-1 colon cancer cell line, which has high expression of Lewis B and Lewis Y antigens, expresses the FUT1 gene, and shows α1,2-fucosyltransferase (FUT) activity. 5′-rapid amplification of cDNA ends revealed a FUT1 transcriptional start site −10 nucleotides upstream of the site registered at NM_000148 in the DataBase of Human Transcription Start Sites (DBTSS). Using the dual luciferase assay, FUT1 gene expression was shown to be regulated at the region −91 to −81 nt to the transcriptional start site, which contains the Elk-1 binding site. Site-directed mutagenesis of this region revealed the Elk-1 binding site to be essential for FUT1 transcription. Furthermore, transfection of the dominant negative Elk-1 gene, and the chromatin immunoprecipitation (CHIp) assay, supported Elk-1-dependent transcriptional regulation of FUT1 gene expression in DLD-1 cells. These results suggest that a defined region in the 5′-flanking region of FUT1 is critical for FUT1 transcription and that constitutive gene expression of FUT1 is regulated by Elk-1 in DLD-1 cells. PMID:23533340

  12. Changes in gravity affect gene expression, protein modulation and metabolite pools of arabidopsis

    NASA Astrophysics Data System (ADS)

    Hampp, R.; Martzivanou, M.; Maier, R. M.; Magel, E.

    Callus cultures of Arabidopsis thaliana (cv. Columbia) in Petri dishes / suspension cultures were exposed to altered g-forces by centrifugation (1 to 10 g), klinorotation, and μ g (sounding rocket flights). Using semi-quantitative RT-PCR, transcripts of genes coding for metabolic key enzymes (ADP-glucose pyrophosphorylase, ADPG-PP; ß-amylase, fructose-1,6-bisphosphatase, FBPase; glyceraldehyde-P dehydrogenase, GAPDH; hydroxymethylglutaryl-CoA reductase, HMG; phenylalanine-ammonium-lyase, PAL; PEP carboxylase, PEPC) were used to monitor threshold conditions for g-number (all) and time of exposure (ß-amylase) which led to altered amounts of the gene product. Exposure to approx. 5 g and higher for 1h resulted in altered transcript levels: transcripts of ß-amylase, PAL, and PEPC were increased, those of ADPG-PP decreased, while those of FBPase, GAPDH, and HMG were not affected. This probably indicates a shift from starch synthesis to starch degradation and increased rates of anaplerosis (PEPC: supply of ketoacids for amino acid synthesis). In order to get more information about g-related effects on gene expression, we used a 1h-exposure to 7 g for a microarray analysis. Transcripts of more than 200 genes were significantly increased in amount (ratio 7g / 1g control; 21.6 and larger). They fall into several categories. Transcripts coding for enzymes of major pathways form the largest group (25%), followed by gene products involved in cellular organisation and cell wall formation / rearrangement (17%), signalling, phosphorylation/dephosphorylation (12%), proteolysis and transport (10% each), hormone synthesis plus related events (8%), defense (4%), stress-response (2%), and gravisensing (2%). Many of the alterations are part of a general stress response, but some changes related to the synthesis / rearrangement of cell wall components could be more hyper-g-specific. Using macroarrays with selected genes according to our hypergravity study (metabolism / signalling

  13. Network analysis of microRNAs, transcription factors, target genes and host genes in nasopharyngeal carcinoma

    PubMed Central

    WANG, HAO; XU, ZHIWEN; MA, MENGYAO; WANG, NING; WANG, KUNHAO

    2016-01-01

    Numerous studies on the morbidity of nasopharyngeal carcinoma (NPC) have identified several genes, microRNAs (miRNAs or miRs) and transcription factors (TFs) that influence the pathogenesis of NPC. However, summarizing all the regulatory networks involved in NPC is challenging. In the present study, the genes, miRNAs and TFs involved in NPC were considered as the nodes of the so-called regulatory network, and the associations between them were investigated. To clearly represent these associations, three regulatory networks were built seperately, namely, the differentially expressed network, the associated network and the global network. The differentially expressed network is the most important one of these three networks, since its nodes are differentially expressed genes whose mutations may lead to the development of NPC. Therefore, by modifying the aberrant expression of those genes that are differentially expressed in this network, their dysregulation may be corrected and the tumorigenesis of NPC may thus be prevented. Analysis of the aforementioned three networks highlighted the importance of certain pathways, such as self-adaptation pathways, in the development of NPC. For example, cyclin D1 (CCND1) was observed to regulate Homo sapiens-miR-20a, which in turn targeted CCND1. The present study conducted a systematic analysis of the pathogenesis of NPC through the three aforementioned regulatory networks, and provided a theoretical model for biologists. Future studies are required to evaluate the influence of the highlighted pathways in NPC. PMID:27313701

  14. Cohesin and Polycomb: Cooperative Checks and Balances in Gene Silencing and Transcription

    PubMed Central

    Dorsett, Dale; Kassis, Judith A.

    2014-01-01

    The cohesin protein complex was discovered for its roles in sister chromatid cohesion and segregation, and the Polycomb group (PcG) proteins for their roles in epigenetic gene silencing during development. Cohesin also controls gene transcription via multiple mechanisms. Genetic and molecular evidence from Drosophila argue that cohesin and the PRC1 PcG complex interact to control transcription of many active genes that are critical for development, and that via these interactions cohesin also controls the availability of PRC1 for gene silencing. PMID:24892918

  15. Virulence genes regulated at the transcriptional level by Ca2+ in Yersinia pestis include structural genes for outer membrane proteins.

    PubMed Central

    Straley, S C; Bowmer, W S

    1986-01-01

    Yersinia pestis, the causative agent of plague, has a virulence determinant called the low-Ca2+ response (Lcr+ phenotype) that confers on the bacterium Ca2+ dependence for growth at 37 degrees C and expression of V antigen. This virulence determinant is common to the three species of Yersinia and is mediated by Lcr plasmids (called pCD in Y. pestis). In this study, we generated insertions of Mu dI1(Ap lac) in pCD1 of Y. pestis KIM, screened for cells showing transcriptional regulation by Ca2+, and obtained inserts that define at least four pCD1 genes. Their patterns of transcription under different growth conditions closely paralleled the pattern of expression of the V antigen. We tested for expression of Lcr-specific yersinial outer membrane proteins (Yops) by the pCD1::Mu dI1(Ap lac) plasmids. Four of the inserts each eliminated expression of a different Yop; one of these Yops was unique to Y. pestis. Two of the insertions affecting Yops caused avirulence, and one caused strongly decreased virulence of Y. pestis in mice. These data indicate that Yops, like the V antigen, are virulence attributes regulated in the low-Ca2+ response. Images PMID:3002984

  16. Mutations in the pho2 (bas2) transcription factor that differentially affect activation with its partner proteins bas1, pho4, and swi5.

    PubMed

    Bhoite, Leena T; Allen, Jason M; Garcia, Emily; Thomas, Lance R; Gregory, I David; Voth, Warren P; Whelihan, Kristen; Rolfes, Ronda J; Stillman, David J

    2002-10-01

    The yeast PHO2 gene encodes a homeodomain protein that exemplifies combinatorial control in transcriptional activation. Pho2 alone binds DNA in vitro with low affinity, but in vivo it activates transcription with at least three disparate DNA-binding proteins: the zinc finger protein Swi5, the helix-loop-helix factor Pho4, and Bas1, an myb-like activator. Pho2 + Swi5 activates HO, Pho2 + Pho4 activates PHO5, and Pho2 + Bas1 activates genes in the purine and histidine biosynthesis pathways. We have conducted a genetic screen and identified 23 single amino acid substitutions in Pho2 that differentially affect its ability to activate its specific target genes. Analysis of the mutations suggests that the central portion of Pho2 serves as protein-protein interactive surface, with a requirement for distinct amino acids for each partner protein. PMID:12145299

  17. Mitotic genes are transcriptionally upregulated in the fibroblast irradiated with very low doses of UV-C.

    PubMed

    Takeuchi, Seiji; Matsuda, Toshiro; Ono, Ryusuke; Tsujimoto, Mariko; Nishigori, Chikako

    2016-01-01

    Ultraviolet (UV) radiation induces a variety of biological effects, including DNA damage response and cell signaling pathways. We performed transcriptome analysis using microarray in human primary cultured fibroblasts irradiated with UV-C (0.5 or 5 J/m(2)) and harvested at 4 or 12 h following UV exposure. All transcript data were analyzed by comparison with the corresponding results in non-irradiated (control) cells. The number of genes with significantly altered expression (≥2-fold difference relative to the control) is higher in the sample irradiated with high dose of UV, suggesting that gene expression was UV dose-dependent. Pathway analysis on the upregulated genes at 12 h indicates that the expression of some cell cycle-related genes was predominantly induced irrespective of UV-dose. Interestingly, almost all the genes with significant altered expression were cell cycle-related genes designated as 'Mitotic Genes', which function in the spindle assembly checkpoint. Therefore, even a low dose of UV could affect the transcriptional profile. PMID:27378355

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

    SciTech Connect

    Karen S. Browning; Marie Petrocek; Bonnie Bartel

    2006-06-01

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

  19. Assessment of anaerobic toluene biodegradation activity by bssA transcript/gene ratios.

    PubMed

    Brow, Christina N; O'Brien Johnson, Reid; Johnson, Richard L; Simon, Holly M

    2013-09-01

    Benzylsuccinate synthase (bssA) genes associated with toluene degradation were profiled across a groundwater contaminant plume under nitrate-reducing conditions and were detected in significant numbers throughout the plume. However, differences between groundwater and core sediment samples suggested that microbial transport, rather than local activity, was the underlying cause of the high copy numbers within the downgradient plume. Both gene transcript and reactant concentrations were consistent with this hypothesis. Expression of bssA genes from denitrifying toluene degraders was induced by toluene but only in the presence of nitrate, and transcript abundance dropped rapidly following the removal of either toluene or nitrate. The drop in bssA transcripts following the removal of toluene could be described by an exponential decay function with a half-life on the order of 1 h. Interestingly, bssA transcripts never disappeared completely but were always detected at some level if either inducer was present. Therefore, the detection of transcripts alone may not be sufficient evidence for contaminant degradation. To avoid mistakenly associating basal-level gene expression with actively degrading microbial populations, an integrated approach using the ratio of functional gene transcripts to gene copies is recommended. This approach minimizes the impact of microbial transport on activity assessment and allows reliable assessments of microbial activity to be obtained from water samples. PMID:23811506

  20. Dysregulation of the homeobox transcription factor gene HOXB13: role in prostate cancer

    PubMed Central

    Decker, Brennan; Ostrander, Elaine A

    2014-01-01

    Prostate cancer (PC) is the most common noncutaneous cancer in men, and epidemiological studies suggest that about 40% of PC risk is heritable. Linkage analyses in hereditary PC families have identified multiple putative loci. However, until recently, identification of specific risk alleles has proven elusive. Cooney et al used linkage mapping and segregation analysis to identify a putative risk locus on chromosome 17q21-22. In search of causative variant(s) in genes from the candidate region, a novel, potentially deleterious G84E substitution in homeobox transcription factor gene HOXB13 was observed in multiple hereditary PC families. In follow-up testing, the G84E allele was enriched in cases, especially those with an early diagnosis or positive family history of disease. This finding was replicated by others, confirming HOXB13 as a PC risk gene. The HOXB13 protein plays diverse biological roles in embryonic development and terminally differentiated tissue. In tumor cell lines, HOXB13 participates in a number of biological functions, including coactivation and localization of the androgen receptor and FOXA1. However, no consensus role has emerged and many questions remain. All HOXB13 variants with a proposed role in PC risk are predicted to damage the protein and lie in domains that are highly conserved across species. The G84E variant has the strongest epidemiological support and lies in a highly conserved MEIS protein-binding domain, which binds cofactors required for activation. On the basis of epidemiological and biological data, the G84E variant likely modulates the interaction between the HOXB13 protein and the androgen receptor, as well as affecting FOXA1-mediated transcriptional programming. However, further studies of the mutated protein are required to clarify the mechanisms by which this translates into PC risk. PMID:25206306

  1. Human transcriptional interactome of chromatin contribute to gene co-expression

    PubMed Central

    2010-01-01

    Background Transcriptional interactome of chromatin is one of the important mechanisms in gene transcription regulation. By chromatin conformation capture and 3D FISH experiments, several chromatin interactions cases among sequence-distant genes or even inter-chromatin genes were reported. However, on genomics level, there is still little evidence to support these mechanisms. Recently based on Hi-C experiment, a genome-wide picture of chromatin interactions in human cells was presented. It provides a useful material for analysing whether the mechanism of transcriptional interactome is common. Results The main work here is to demonstrate whether the effects of transcriptional interactome on gene co-expression exist on genomic level. While controlling the effects of transcription factors control similarities (TCS), we tested the correlation between Hi-C interaction and the mutual ranks of gene co-expression rates (provided by COXPRESdb) of intra-chromatin gene pairs. We used 6,084 genes with both TF annotation and co-expression information, and matched them into 273,458 pairs with similar Hi-C interaction ranks in different cell types. The results illustrate that co-expression is strongly associated with chromatin interaction. Further analysis using GO annotation reveals potential correlation between gene function similarity, Hi-C interaction and their co-expression. Conclusions According to the results in this research, the intra-chromatin interactome may have relation to gene function and associate with co-expression. This study provides evidence for illustrating the effect of transcriptional interactome on transcription regulation. PMID:21156067

  2. Pharmacological and Genetic Modulation of REV-ERB Activity and Expression Affects Orexigenic Gene Expression.

    PubMed

    Amador, Ariadna; Wang, Yongjun; Banerjee, Subhashis; Kameneka, Theodore M; Solt, Laura A; Burris, Thomas P

    2016-01-01

    The nuclear receptors REV-ERBα and REV-ERBβ are transcription factors that play pivotal roles in the regulation of the circadian rhythm and various metabolic processes. The circadian rhythm is an endogenous mechanism, which generates entrainable biological changes that follow a 24-hour period. It regulates a number of physiological processes, including sleep/wakeful cycles and feeding behaviors. We recently demonstrated that REV-ERB-specific small molecules affect sleep and anxiety. The orexinergic system also plays a significant role in mammalian physiology and behavior, including the regulation of sleep and food intake. Importantly, orexin genes are expressed in a circadian manner. Given these overlaps in function and circadian expression, we wanted to determine whether the REV-ERBs might regulate orexin. We found that acute in vivo modulation of REV-ERB activity, with the REV-ERB-specific synthetic ligand SR9009, affects the circadian expression of orexinergic genes in mice. Long term dosing with SR9009 also suppresses orexinergic gene expression in mice. Finally, REV-ERBβ-deficient mice present with increased orexinergic transcripts. These data suggest that the REV-ERBs may be involved in the repression of orexinergic gene expression. PMID:26963516

  3. Pharmacological and Genetic Modulation of REV-ERB Activity and Expression Affects Orexigenic Gene Expression

    PubMed Central

    Amador, Ariadna; Wang, Yongjun; Banerjee, Subhashis; Kameneka, Theodore M.; Solt, Laura A.; Burris, Thomas P.

    2016-01-01

    The nuclear receptors REV-ERBα and REV-ERBβ are transcription factors that play pivotal roles in the regulation of the circadian rhythm and various metabolic processes. The circadian rhythm is an endogenous mechanism, which generates entrainable biological changes that follow a 24-hour period. It regulates a number of physiological processes, including sleep/wakeful cycles and feeding behaviors. We recently demonstrated that REV-ERB-specific small molecules affect sleep and anxiety. The orexinergic system also plays a significant role in mammalian physiology and behavior, including the regulation of sleep and food intake. Importantly, orexin genes are expressed in a circadian manner. Given these overlaps in function and circadian expression, we wanted to determine whether the REV-ERBs might regulate orexin. We found that acute in vivo modulation of REV-ERB activity, with the REV-ERB-specific synthetic ligand SR9009, affects the circadian expression of orexinergic genes in mice. Long term dosing with SR9009 also suppresses orexinergic gene expression in mice. Finally, REV-ERBβ-deficient mice present with increased orexinergic transcripts. These data suggest that the REV-ERBs may be involved in the repression of orexinergic gene expression. PMID:26963516

  4. Modelling transcriptional interference and DNA looping in gene regulation.

    PubMed

    Dodd, Ian B; Shearwin, Keith E; Sneppen, Kim

    2007-06-22

    We describe a hybrid statistical mechanical and dynamical approach for modelling the formation of closed, open and elongating complexes of RNA polymerase, the interactions of these polymerases to produce transcriptional interference, and the regulation of these processes by a DNA-binding and DNA-looping regulatory protein. As a model system, we have used bacteriophage 186, for which genetic, biochemical and structural studies have suggested that the CI repressor binds as a 14-mer to form alternative DNA-looped complexes, and activates lysogenic transcription indirectly by relieving transcriptional interference caused by the convergent lytic promoter. The modelling showed that the original mechanisms proposed to explain this relief of transcriptional interference are not consistent with the available in vivo reporter data. However, a good fit to the reporter data was given by a revised model that incorporates a novel predicted regulatory mechanism: that RNA polymerase bound at the lysogenic promoter protects itself from transcriptional interference by recruiting CI to the lytic promoter. This mechanism and various estimates of in vivo biochemical parameters for the 186 CI system should be testable. Our results demonstrate the power of mathematical modelling for the extraction of detailed biochemical information from in vivo data. PMID:17498740

  5. Validation of Reference Genes for Transcriptional Analyses in Pleurotus ostreatus by Using Reverse Transcription-Quantitative PCR

    PubMed Central

    Castanera, Raúl; López-Varas, Leticia; Pisabarro, Antonio G.

    2015-01-01

    Recently, the lignin-degrading basidiomycete Pleurotus ostreatus has become a widely used model organism for fungal genomic and transcriptomic analyses. The increasing interest in this species has led to an increasing number of studies analyzing the transcriptional regulation of multigene families that encode extracellular enzymes. Reverse transcription (RT) followed by real-time PCR is the most suitable technique for analyzing the expression of gene sets under multiple culture conditions. In this work, we tested the suitability of 13 candidate genes for their use as reference genes in P. ostreatus time course cultures for enzyme production. We applied three different statistical algorithms and obtained a combination of stable reference genes for optimal normalization of RT-quantitative PCR assays. This reference index can be used for future transcriptomic analyses and validation of transcriptome sequencing or microarray data. Moreover, we analyzed the expression patterns of a laccase and a manganese peroxidase (lacc10 and mnp3, respectively) in lignocellulose and glucose-based media using submerged, semisolid, and solid-state fermentation. By testing different normalization strategies, we demonstrate that the use of nonvalidated reference genes as internal controls leads to biased results and misinterpretations of the biological responses underlying expression changes. PMID:25862220

  6. Pleiohomeotic Interacts with the Core Transcription Elongation Factor Spt5 to Regulate Gene Expression in Drosophila

    PubMed Central

    Jennings, Barbara H.

    2013-01-01

    The early elongation checkpoint regulated by Positive Transcription Elongation Factor b (P-TEFb) is a critical control point for the expression of many genes. Spt5 interacts directly with RNA polymerase II and has an essential role in establishing this checkpoint, and also for further transcript elongation. Here we demonstrate that Drosophila Spt5 interacts both physically and genetically with the Polycomb Group (PcG) protein Pleiohomeotic (Pho), and the majority of Pho binding sites overlap with Spt5 binding sites across the genome in S2 cells. Our results indicate that Pho can interact with Spt5 to regulate transcription elongation in a gene specific manner. PMID:23894613

  7. The Role of Transcription Factors at Antisense-Expressing Gene Pairs in Yeast

    PubMed Central

    Mostovoy, Yulia; Thiemicke, Alexander; Hsu, Tiffany Y.; Brem, Rachel B.

    2016-01-01

    Genes encoded close to one another on the chromosome are often coexpressed, by a mechanism and regulatory logic that remain poorly understood. We surveyed the yeast genome for tandem gene pairs oriented tail-to-head at which expression antisense to the upstream gene was conserved across species. The intergenic region at most such tandem pairs is a bidirectional promoter, shared by the downstream gene mRNA and the upstream antisense transcript. Genomic analyses of these intergenic loci revealed distinctive patterns of transcription factor regulation. Mutation of a given transcription factor verified its role as a regulator in trans of tandem gene pair loci, including the proximally initiating upstream antisense transcript and downstream mRNA and the distally initiating upstream mRNA. To investigate cis-regulatory activity at such a locus, we focused on the stress-induced NAD(P)H dehydratase YKL151C and its downstream neighbor, the metabolic enzyme GPM1. Previous work has implicated the region between these genes in regulation of GPM1 expression; our mutation experiments established its function in rich medium as a repressor in cis of the distally initiating YKL151C sense RNA, and an activator of the proximally initiating YKL151C antisense RNA. Wild-type expression of all three transcripts required the transcription factor Gcr2. Thus, at this locus, the intergenic region serves as a focal point of regulatory input, driving antisense expression and mediating the coordinated regulation of YKL151C and GPM1. Together, our findings implicate transcription factors in the joint control of neighboring genes specialized to opposing conditions and the antisense transcripts expressed between them. PMID:27190003

  8. The WRKY Transcription Factor Genes in Eggplant (Solanum melongena L.) and Turkey Berry (Solanum torvum Sw.)

    PubMed Central

    Yang, Xu; Deng, Cao; Zhang, Yu; Cheng, Yufu; Huo, Qiuyue; Xue, Linbao

    2015-01-01

    WRKY transcription factors, which play critical roles in stress responses, have not been characterized in eggplant or its wild relative, turkey berry. The recent availability of RNA-sequencing data provides the opportunity to examine WRKY genes from a global perspective. We identified 50 and 62 WRKY genes in eggplant (SmelWRKYs) and turkey berry (StorWRKYs), respectively, all of which could be classified into three groups (I–III) based on the WRKY protein structure. The SmelWRKYs and StorWRKYs contain ~76% and ~95% of the number of WRKYs found in other sequenced asterid species, respectively. Positive selection analysis revealed that different selection constraints could have affected the evolution of these groups. Positively-selected sites were found in Groups IIc and III. Branch-specific selection pressure analysis indicated that most WRKY domains from SmelWRKYs and StorWRKYs are conserved and have evolved at low rates since their divergence. Comparison to homologous WRKY genes in Arabidopsis revealed several potential pathogen resistance-related SmelWRKYs and StorWRKYs, providing possible candidate genetic resources for improving stress tolerance in eggplant and probably other Solanaceae plants. To our knowledge, this is the first report of a genome-wide analyses of the SmelWRKYs and StorWRKYs. PMID:25853261

  9. Transcriptional Profiling Reveals Differential Gene Expression of Amur Ide (Leuciscus waleckii) during Spawning Migration

    PubMed Central

    Cui, Jun; Xu, Jian; Zhang, Songhao; Wang, Kai; Jiang, Yanliang; Mahboob, Shahid; Al-Ghanim, Khalid A.; Xu, Peng

    2015-01-01

    Amur ide (Leuciscus waleckii), an important aquaculture species, inhabits neutral freshwater but can tolerate high salinity or alkalinity. As an extreme example, the population in Dali Nor lake inhabits alkalized soda water permanently, and migrates from alkaline water to neutral freshwater to spawn. In this study, we performed comparative transcriptome profiling study on the livers of Amur ide to interrogate the expression differences between the population that permanently inhabit freshwater in Ganggeng Nor lake (FW) and the spawning population that recently migrated from alkaline water into freshwater (SM). A total of 637,234,880 reads were generated, resulting in 53,440 assembled contigs that were used as reference sequences. Comparisons of these transcriptome files revealed 444 unigenes with significant differential expression (p-value ≤ 0.01, fold-change ≥ 2), including 246 genes that were up-regulated in SM and 198 genes that were up-regulated in FW. The gene ontology (GO) enrichment analysis and KEGG pathway analysis indicated that the mTOR signaling pathway, Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway, and oxidative phosphorylation were highly likely to affect physiological changes during spawning migration. Overall, this study demonstrates that transcriptome changes played a role in Amur ide spawning migration. These results provide a foundation for further analyses on the physiological and molecular mechanisms underlying Amur ide spawning migration. PMID:26096003

  10. The WRKY transcription factor genes in eggplant (Solanum melongena L.) and Turkey Berry (Solanum torvum Sw.).

    PubMed

    Yang, Xu; Deng, Cao; Zhang, Yu; Cheng, Yufu; Huo, Qiuyue; Xue, Linbao

    2015-01-01

    WRKY transcription factors, which play critical roles in stress responses, have not been characterized in eggplant or its wild relative, turkey berry. The recent availability of RNA-sequencing data provides the opportunity to examine WRKY genes from a global perspective. We identified 50 and 62 WRKY genes in eggplant (SmelWRKYs) and turkey berry (StorWRKYs), respectively, all of which could be classified into three groups (I-III) based on the WRKY protein structure. The SmelWRKYs and StorWRKYs contain ~76% and ~95% of the number of WRKYs found in other sequenced asterid species, respectively. Positive selection analysis revealed that different selection constraints could have affected the evolution of these groups. Positively-selected sites were found in Groups IIc and III. Branch-specific selection pressure analysis indicated that most WRKY domains from SmelWRKYs and StorWRKYs are conserved and have evolved at low rates since their divergence. Comparison to homologous WRKY genes in Arabidopsis revealed several potential pathogen resistance-related SmelWRKYs and StorWRKYs, providing possible candidate genetic resources for improving stress tolerance in eggplant and probably other Solanaceae plants. To our knowledge, this is the first report of a genome-wide analyses of the SmelWRKYs and StorWRKYs. PMID:25853261

  11. Involvement of several transcriptional regulators in the differential expression of tfd genes in Cupriavidus necator JMP134.

    PubMed

    Trefault, Nicole; Guzmán, Leda; Pérez, Heidi; Godoy, Margarita; González, Bernardo

    2009-06-01

    Cupriavidus necator JMP134 has been extensively studied because of its ability to degrade chloroaromatic compounds, including the herbicides 2,4-dichlorophenoxyacetic acid (2,4-D) and 3-chlorobenzoic acid (3-CB), which is achieved through the pJP4-encoded chlorocatechol degradation gene clusters: tfdCIDIEIFI and tfdDIICIIEIIFII. The present work describes a different tfd-genes expression profile depending on whether C. necator cells were induced with 2,4-D or 3-CB. By contrast, in vitro binding assays of the purified transcriptional activator TfdR showed similar binding to both tfd intergenic regions; these results were confirmed by in vivo studies of the expression of transcriptional lacZ fusions for these intergenic regions. Experiments aimed at investigating whether other pJP4 plasmid or chromosomal regulatory proteins could contribute to the differences in the response of both tfd promoters to induction by 2,4-D and 3-CB showed that the transcriptional regulators from the benzoate degradation pathway, CatR1 and CatR2, affected 3-CB- and 2,4-D-related growth capabilities. It was also determined that the ISJP4-interrupted protein TfdT decreased growth on 3-CB. In addition, an ORF with 34% amino acid identity to IclR-type transcriptional regulator members and located near the tfdII gene cluster module was shown to modulate the 2,4-D growth capability. Taken together, these results suggest that tfd transcriptional regulation in C. necator JMP134 is far more complex than previously thought and that it involves proteins from different transcriptional regulator families. PMID:19784929

  12. Microprocessor mediates transcriptional termination in long noncoding microRNA genes

    PubMed Central

    Dhir, Ashish; Dhir, Somdutta; Proudfoot, Nick J.; Jopling, Catherine L.

    2015-01-01

    MicroRNA (miRNA) play a major role in the post-transcriptional regulation of gene expression. Mammalian miRNA biogenesis begins with co-transcriptional cleavage of RNA polymerase II (Pol II) transcripts by the Microprocessor complex. While most miRNA are located within introns of protein coding genes, a substantial minority of miRNA originate from long non coding (lnc) RNA where transcript processing is largely uncharacterized. We show, by detailed characterization of liver-specific lnc-pri-miR-122 and genome-wide analysis in human cell lines, that most lnc-pri-miRNA do not use the canonical cleavage and polyadenylation (CPA) pathway, but instead use Microprocessor cleavage to terminate transcription. This Microprocessor inactivation leads to extensive transcriptional readthrough of lnc-pri-miRNA and transcriptional interference with downstream genes. Consequently we define a novel RNase III-mediated, polyadenylation-independent mechanism of Pol II transcription termination in mammalian cells. PMID:25730776

  13. Fail-Safe Transcriptional Termination for Protein-Coding Genes in S. cerevisiae

    PubMed Central

    Rondón, Ana G.; Mischo, Hannah E.; Kawauchi, Junya; Proudfoot, Nick J.

    2009-01-01

    Summary Transcription termination of RNA polymerase II (Pol II) on protein-coding genes in S. cerevisiae relies on pA site recognition by 3′ end processing factors. Here we demonstrate the existence of two alternative termination mechanisms that rescue polymerases failing to disengage from the template at pA sites. One of these fail-safe mechanisms is mediated by the NRD complex, similar to termination of short noncoding genes. The other termination mechanism is mediated by Rnt1 cleavage of the nascent transcript. Both fail-safe termination mechanisms trigger degradation of readthrough transcripts by the exosome. However, Rnt1-mediated termination can also enhance the usage of weak pA signals and thereby generate functional mRNA. We propose that these alternative Pol II termination pathways serve the dual function of avoiding transcription interference and promoting rapid removal of aberrant transcripts. PMID:19818712

  14. Androgen Receptor Repression of GnRH Gene Transcription

    PubMed Central

    Brayman, Melissa J.; Pepa, Patricia A.; Berdy, Sara E.

    2012-01-01

    Alterations in androgen levels lead to reproductive defects in both males and females, including hypogonadotropic hypogonadism, anovulation, and infertility. Androgens have been shown to down-regulate GnRH mRNA levels through an androgen receptor (AR)-dependent mechanism. Here, we investigate how androgen regulates expression from the GnRH regulatory region in the GT1-7 cell line, a model of GnRH neurons. A synthetic androgen, R1881, repressed transcription from the GnRH promoter (GnRH-P) in an AR-dependent manner, and liganded AR associated with the chromatin at the GnRH-P in live GT1-7 cells. The three known octamer-binding transcription factor-1 (Oct-1) binding sites in GnRH-P were required for AR-mediated repression, although other sequences were also involved. Although a multimer of the consensus Oct-1 binding site was not repressed, a multimer of the cluster of Oct-1, Pre-B cell leukemia transcription factor (Pbx)/Prep, and NK2 homeobox 1 (Nkx2.1) binding sites, found at −106/−91 in GnRH-P, was sufficient for repression. In fact, overexpression of any of these factors disrupted the androgen response, indicating that a balance of factors in this tripartite complex is required for AR repression. AR bound to this region in EMSA, indicating a direct interaction of AR with DNA or with other transcription factors bound to GnRH-P at this sequence. Collectively, our data demonstrate that GnRH transcription is repressed by AR via multiple sequences in GnRH-P, including three Oct-1 binding sites, and that this repression requires the complex interaction of several transcription factors. PMID:22074952

  15. Extracellular Matrix-Regulated Gene Expression RequiresCooperation of SWI/SNF and Transcription Factors

    SciTech Connect

    Xu, Ren; Spencer, Virginia A.; Bissell, Mina J.

    2006-05-25

    Extracellular cues play crucial roles in the transcriptional regulation of tissue-specific genes, but whether and how these signals lead to chromatin remodeling is not understood and subject to debate. Using chromatin immunoprecipitation (ChIP) assays and mammary-specific genes as models, we show here that extracellular matrix (ECM) molecules and prolactin cooperate to induce histone acetylation and binding of transcription factors and the SWI/SNF complex to the {beta}- and ?-casein promoters. Introduction of a dominant negative Brg1, an ATPase subunit of SWI/SNF complex, significantly reduced both {beta}- and ?-casein expression, suggesting that SWI/SNF-dependent chromatin remodeling is required for transcription of mammary-specific genes. ChIP analyses demonstrated that the ATPase activity of SWI/SNF is necessary for recruitment of RNA transcriptional machinery, but not for binding of transcription factors or for histone acetylation. Coimmunoprecipitation analyses showed that the SWI/SNF complex is associated with STAT5, C/EBP{beta}, and glucocorticoid receptor (GR). Thus, ECM- and prolactin-regulated transcription of the mammary-specific casein genes requires the concerted action of chromatin remodeling enzymes and transcription factors.

  16. Transcription Profile of Aging and Cognition-Related Genes in the Medial Prefrontal Cortex

    PubMed Central

    Ianov, Lara; Rani, Asha; Beas, Blanca S.; Kumar, Ashok; Foster, Thomas C.

    2016-01-01

    Cognitive function depends on transcription; however, there is little information linking altered gene expression to impaired prefrontal cortex function during aging. Young and aged F344 rats were characterized on attentional set shift and spatial memory tasks. Transcriptional differences associated with age and cognition were examined using RNA sequencing to construct transcriptomic profiles for the medial prefrontal cortex (mPFC), white matter, and region CA1 of the hippocampus. The results indicate regional differences in vulnerability to aging. Age-related gene expression in the mPFC was similar to, though less robust than, changes in the dorsolateral PFC of aging humans suggesting that aging processes may be similar. Importantly, the pattern of transcription associated with aging did not predict cognitive decline. Rather, increased mPFC expression of genes involved in regulation of transcription, including transcription factors that regulate the strength of excitatory and inhibitory inputs, and neural activity-related immediate-early genes was observed in aged animals that exhibit delayed set shift behavior. The specificity of impairment on a mPFC-dependent task, associated with a particular mPFC transcriptional profile indicates that impaired executive function involves altered transcriptional regulation and neural activity/plasticity processes that are distinct from that described for impaired hippocampal function. PMID:27242522

  17. Helix-loop-helix transcription factors mediate activation and repression of the p75LNGFR gene.

    PubMed Central

    Chiaramello, A; Neuman, K; Palm, K; Metsis, M; Neuman, T

    1995-01-01

    Sequence analysis of rat and human low-affinity nerve growth factor receptor p75LNGFR gene promoter regions revealed a single E-box cis-acting element, located upstream of the major transcription start sites. Deletion analysis of the E-box sequence demonstrated that it significantly contributes to p75LNGFR promoter activity. This E box has a dual function; it mediates either activation or repression of the p75LNGFR promoter activity, depending on the interacting transcription factors. We showed that the two isoforms of the class A basic helix-loop-helix (bHLH) transcription factor ME1 (ME1a and ME1b), the murine homolog of the human HEB transcription factor, specifically repress p75LNGFR promoter activity. This repression can be released by coexpression of the HLH Id2 transcriptional regulator. In vitro analyses demonstrated that ME1a forms a stable complex with the p75LNGFR E box and likely competes with activating E-box-binding proteins. By using ME1a-overexpressing PC12 cells, we showed that the endogenous p75LNGFR gene is a target of ME1a repression. Together, these data demonstrate that the p75LNGFR E box and the interacting bHLH transcription factors are involved in the regulation of p75LNGFR gene expression. These results also show that class A bHLH transcription factors can repress and Id-like negative regulators can stimulate gene expression. PMID:7565756

  18. FoxO1 Deacetylation Regulates Thyroid Hormone-induced Transcription of Key Hepatic Gluconeogenic Genes*

    PubMed Central

    Singh, Brijesh Kumar; Sinha, Rohit Anthony; Zhou, Jin; Xie, Sherwin Ying; You, Seo-Hee; Gauthier, Karine; Yen, Paul Michael

    2013-01-01

    Hepatic gluconeogenesis is a concerted process that integrates transcriptional regulation with hormonal signals. A major regulator is thyroid hormone (TH), which acts through its nuclear receptor (TR) to induce the expression of the hepatic gluconeogenic genes, phosphoenolpyruvate carboxykinase (PCK1) and glucose-6-phosphatase (G6PC). Forkhead transcription factor FoxO1 also is an important regulator of these genes; however, its functional interactions with TR are not known. Here, we report that TR-mediated transcriptional activation of PCK1 and G6PC in human hepatic cells and mouse liver was FoxO1-dependent and furthermore required FoxO1 deacetylation by the NAD+-dependent deacetylase, SirT1. siRNA knockdown of FoxO1 decreased, whereas overexpression of FoxO1 increased, TH-dependent transcriptional activation of PCK1 and G6PC in cultured hepatic cells. FoxO1 siRNA knockdown also decreased TH-mediated transcription in vivo. Additionally, TH was unable to induce FoxO1 deacetylation or hepatic PCK1 gene expression in TH receptor β-null (TRβ−/−) mice. Moreover, TH stimulated FoxO1 recruitment to the PCK1 and G6PC gene promoters in a SirT1-dependent manner. In summary, our results show that TH-dependent deacetylation of a second metabolically regulated transcription factor represents a novel mechanism for transcriptional integration of nuclear hormone action with cellular energy status. PMID:23995837

  19. A discrete region centered 22 base pairs upstream of the initiation site modulates transcription of Drosophila tRNAAsn genes.

    PubMed Central

    Lofquist, A K; Garcia, A D; Sharp, S J

    1988-01-01

    We have studied the mechanism by which 5'-flanking sequences modulate the in vitro transcription of eucaryotic tRNA genes. Using deletion and linker substitution mutagenesis, we have found that the 5'-flanking sequences responsible for the different in vitro transcription levels of three Drosophila tRNA5Asn genes are contained within a discrete region centered 22 nucleotides upstream from the transcription initiation site. In conjunction with the A-box intragenic control region, this upstream transcription-modulatory region functions in the selection mechanism for the site of transcription initiation. Since the transcription-modulatory region directs the position of the start site and the actual sequence of the transcription-modulatory region determines the level of tRNAAsn gene transcription, the possibility is raised that the transcription-modulatory region directs a transcription initiation event similar to open complex formation at procaryotic promoters. Images PMID:3141790

  20. Genetic variations in GPSM3 associated with protection from rheumatoid arthritis affect its transcript abundance

    PubMed Central

    Gall, BJ; Wilson, A; Schroer, AB; Gross, JD; Stoilov, P; Setola, V; Watkins, CM; Siderovski, DP

    2015-01-01

    G protein signaling modulator 3 (GPSM3) is a regulator of G protein-coupled receptor signaling, with expression restricted to leukocytes and lymphoid organs. Previous genome-wide association studies have highlighted single-nucleotide polymorphisms (SNPs rs204989, rs204991) in a region upstream of the GPSM3 transcription start site as being inversely correlated to the prevalence of rheumatoid arthritis (RA) -- this association is supported by the protection afforded to Gpsm3-deficient mice in models of inflammatory arthritis. Here, we assessed the functional consequences of these polymorphisms. We collected biospecimens from 50 volunteers with RA diagnoses, 50 RA-free volunteers matched to the aforementioned group, and 100 unmatched healthy young volunteers. We genotyped these individuals for GPSM3 (rs204989, rs204991), CCL21 (rs2812378), and HLA gene region (rs6457620) polymorphisms, and found no significant differences in minor allele frequencies between the RA and disease-free cohorts. However, we identified that individuals homozygous for SNPs rs204989 and rs204991 had decreased GPSM3 transcript abundance relative to individuals homozygous for the major allele. In vitro promoter activity studies suggest that SNP rs204989 is the primary cause of this decrease in transcript levels. Knockdown of GPSM3 in THP-1 cells, a human monocytic cell line, was found to disrupt ex vivo migration to the chemokine MCP-1. PMID:26821282

  1. Close association of RNA polymerase II and many transcription factors with Pol III genes.

    PubMed

    Raha, Debasish; Wang, Zhong; Moqtaderi, Zarmik; Wu, Linfeng; Zhong, Guoneng; Gerstein, Mark; Struhl, Kevin; Snyder, Michael

    2010-02-23

    Transcription of the eukaryotic genomes is carried out by three distinct RNA polymerases I, II, and III, whereby each polymerase is thought to independently transcribe a distinct set of genes. To investigate a possible relationship of RNA polymerases II and III, we mapped their in vivo binding sites throughout the human genome by using ChIP-Seq in two different cell lines, GM12878 and K562 cells. Pol III was found to bind near many known genes as well as several previously unidentified target genes. RNA-Seq studies indicate that a majority of the bound genes are expressed, although a subset are not suggestive of stalling by RNA polymerase III. Pol II was found to bind near many known Pol III genes, including tRNA, U6, HVG, hY, 7SK and previously unidentified Pol III target genes. Similarly, in vivo binding studies also reveal that a number of transcription factors normally associated with Pol II transcription, including c-Fos, c-Jun and c-Myc, also tightly associate with most Pol III-transcribed genes. Inhibition of Pol II activity using alpha-amanitin reduced expression of a number of Pol III genes (e.g., U6, hY, HVG), suggesting that Pol II plays an important role in regulating their transcription. These results indicate that, contrary to previous expectations, polymerases can often work with one another to globally coordinate gene expression. PMID:20139302

  2. Transcription factor C/EBPβ promotes the transcription of the porcine GPR120 gene.

    PubMed

    Chen, Kun; Zhou, Ji-Dan; Zhang, Feng; Zhang, Fang; Zhang, Rui-Rui; Zhan, Meng-Si; Tang, Xiao-Yin; Deng, Bing; Lei, Ming-Gang; Xiong, Yuan-Zhu

    2016-02-01

    G protein-coupled receptor 120 (GPR120), an adipogenic receptor critical for the differentiation and maturation of adipocytes, plays an important role in controlling obesity in both humans and rodents and, thus, is an attractive target of obesity treatment studies. However, the mechanisms that regulate the expression of porcine GPR120 remain unclear. In this study, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) techniques were used to analyze and identify the binding of C/EBPβ (transcription factor CCAAT/enhancer binding protein beta) to the GPR120 promoter. C/EBPβ overexpression and RNA interference studies showed that C/EBPβ regulated GPR120 promoter activity and endogenous GPR120 expression. The binding site of C/EBPβ in the GPR120 promoter region from -101 to -87 was identified by promoter deletion analysis and site-directed mutagenesis. Overexpression of C/EBPβ increased endogenous GPR120 expression in pig kidney cells (PK). Furthermore, when endogenous C/EBPβ was knocked down, GPR120 mRNA and protein levels were decreased. The stimulatory effect of C/EBPβ on GPR120 transcription and its ability to bind the transcription factor-binding site were confirmed by luciferase, ChIP, and EMSA. Moreover, the mRNA and protein expression levels of C/EBPβ were induced by high fat diet feeding. Taken together, it can be concluded that C/EBPβ plays a vital role in regulating GPR120 transcription and suggests HFD-feeding induces GPR120 transcription by influencing C/EBPβ expression. PMID:26576644

  3. The transcriptional activities and cellular localization of the human estrogen receptor alpha are affected by the synonymous Ala87 mutation.

    PubMed

    Fernández-Calero, Tamara; Astrada, Soledad; Alberti, Alvaro; Horjales, Sofía; Arnal, Jean Francois; Rovira, Carlos; Bollati-Fogolín, Mariela; Flouriot, Gilles; Marin, Mónica

    2014-09-01

    Until recently, synonymous mutations (which do not change amino acids) have been much neglected. Some evidence suggests that this kind of mutations could affect mRNA secondary structure or stability, translation kinetics and protein structure. To explore deeper the role of synonymous mutations, we studied their consequence on the functional activity of the estrogen receptor alpha (ERα). The ERα is a ligand-inducible transcription factor that orchestrates pleiotropic cellular effects, at both genomic and non-genomic levels in response to estrogens. In this work we analyzed in transient transfection experiments, the activity of ERα carrying the synonymous mutation Ala87, a polymorphism involving about 5-10% of the population. In comparison to the wild type receptor, our results show that ERαA87 mutation reduces the transactivation efficiency of ERα on an ERE reporter gene while its expression level remains similar. This mutation enhances 4-OHT-induced transactivation of ERα on an AP1 reporter gene. Finally, the mutation affects the subcellular localization of ERα in a cell type specific manner. It enhances the cytoplasmic location of ERα without significant changes in non-genomic effects of E2. The functional alteration of the ERαA87 determined in this work highlights the relevance of synonymous mutations for biomedical and pharmacological points of view. PMID:24607813

  4. What's that gene (or protein)? Online resources for exploring functions of genes, transcripts, and proteins

    PubMed Central

    Hutchins, James R. A.

    2014-01-01

    The genomic era has enabled research projects that use approaches including genome-scale screens, microarray analysis, next-generation sequencing, and mass spectrometry–based proteomics to discover genes and proteins involved in biological processes. Such methods generate data sets of gene, transcript, or protein hits that researchers wish to explore to understand their properties and functions and thus their possible roles in biological systems of interest. Recent years have seen a profusion of Internet-based resources to aid this process. This review takes the viewpoint of the curious biologist wishing to explore the properties of protein-coding genes and their products, identified using genome-based technologies. Ten key questions are asked about each hit, addressing functions, phenotypes, expression, evolutionary conservation, disease association, protein structure, interactors, posttranslational modifications, and inhibitors. Answers are provided by presenting the latest publicly available resources, together with methods for hit-specific and data set–wide information retrieval, suited to any genome-based analytical technique and experimental species. The utility of these resources is demonstrated for 20 factors regulating cell proliferation. Results obtained using some of these are discussed in more depth using the p53 tumor suppressor as an example. This flexible and universally applicable approach for characterizing experimental hits helps researchers to maximize the potential of their projects for biological discovery. PMID:24723265

  5. FAK and HAS inhibition synergistically decrease colon cancer cell viability and affect expression of critical genes.

    PubMed

    Heffler, Melissa; Golubovskaya, Vita M; Conroy, Jeffrey; Liu, Song; Wang, Dan; Cance, William G; Dunn, Kelli B

    2013-05-01

    Focal adhesion kinase (FAK), hyaluronan (HA), and hyaluronan synthase-3 (HAS3) have been implicated in cancer growth and progression. FAK inhibition with the small molecule inhibitor Y15 decreases colon cancer cell growth in vitro and in vivo. HAS3 inhibition in colon cancer cells decreases FAK expression and activation, and exogenous HA increases FAK activation. We sought to determine the genes affected by HAS and FAK inhibition and hypothesized that dual inhibition would synergistically inhibit viability. Y15 (FAK inhibitor) and the HAS inhibitor 4-methylumbelliferone (4-MU) decreased viability in a dose dependent manner; viability was further inhibited by treatment with Y15 and 4-MU in colon cancer cells. HAS inhibited cells treated with 2 μM of Y15 showed significantly decreased viability compared to HAS scrambled cells treated with the same dose (p < 0.05) demonstrating synergistic inhibition of viability with dual FAK/HAS inhibition. Microarray analysis showed more than 2-fold up- or down-regulation of 121 genes by HAS inhibition, and 696 genes by FAK inhibition (p < 0.05) and revealed 29 common genes affected by both signaling. Among the genes affected by FAK or HAS3 inhibition were genes, playing role in apoptosis, cell cycle regulation, adhesion, transcription, heatshock and WNT pathways. Thus, FAK or HAS inhibition decreases SW620 viability and affects several similar genes, which are involved in the regulation of tumor survival. Dual inhibition of FAK and HAS3 decreases viability to a greater degree than with either agent alone, and suggests that synergistic inhibition of colon cancer cell growth can result from affecting similar genetic pathways. PMID:22934709

  6. Transcriptionally targeted gene therapy to detect and treat cancer

    PubMed Central

    Wu, Lily; Johnson, Mai; Sato, Makoto

    2010-01-01

    The greatest challenge in cancer treatment is to achieve the highest levels of specificity and efficacy. Cancer gene therapy could be designed specifically to express therapeutic genes to induce cancer cell destruction. Cancer-specific promoters are useful tools to accomplish targeted expression; however, high levels of gene expression are needed to achieve therapeutic efficacy. Incorporating an imaging reporter gene in tandem with the therapeutic gene will allow tangible proof of principle that gene expression occurs at the correct location and at a sufficient level. Gene-based imaging can advance cancer detection and diagnosis. By combining the cancer-targeted imaging and therapeutic strategies, the exciting prospect of a ‘one-two punch’ to find hidden, disseminated cancer cells and destroy them simultaneously can potentially be realized. PMID:14557054

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

    PubMed

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

    2006-07-01

    Identification of transcriptional regulatory sites plays an important role in the investigation of gene regulation. For this propose, we designed and implemented a data warehouse to integrate multiple heterogeneous biological data sources with data types such as text-file, XML, image, MySQL database model, and Oracle database model. The utility of the biological data warehouse in predicting transcriptional regulatory sites of coregulated genes was explored using a synexpression group derived from a microarray study. Both of the binding sites of known transcription factors and predicted over-represented (OR) oligonucleotides were demonstrated for the gene group. The potential biological roles of both known nucleotides and one OR nucleotide were demonstrated using bioassays. Therefore, the results from the wet-lab experiments reinforce the power and utility of the data warehouse as an approach to the genome-wide search for important transcription regulatory elements that are the key to many complex biological systems. PMID:16871724

  8. Circadian Enhancers Coordinate Multiple Phases of Rhythmic Gene Transcription In Vivo

    PubMed Central

    Fang, Bin; Everett, Logan J.; Jager, Jennifer; Briggs, Erika; Armour, Sean M.; Feng, Dan; Roy, Ankur; Gerhart-Hines, Zachary; Sun, Zheng; Lazar, Mitchell A.

    2014-01-01

    SUMMARY Mammalian transcriptomes display complex circadian rhythms with multiple phases of gene expression that cannot be accounted for by current models of the molecular clock. We have determined the underlying mechanisms by measuring nascent RNA transcription around the clock in mouse liver. Unbiased examination of eRNAs that cluster in specific circadian phases identified functional enhancers driven by distinct transcription factors (TFs). We further identify on a global scale the components of the TF cistromes that function to orchestrate circadian gene expression. Integrated genomic analyses also revealed novel mechanisms by which a single circadian factor controls opposing transcriptional phases. These findings shed new light on the diversity and specificity of TF function in the generation of multiple phases of circadian gene transcription in a mammalian organ. PMID:25416951

  9. Distinct DNA-based epigenetic switches trigger transcriptional activation of silent genes in human dermal fibroblasts

    PubMed Central

    Pandian, Ganesh N.; Taniguchi, Junichi; Junetha, Syed; Sato, Shinsuke; Han, Le; Saha, Abhijit; AnandhaKumar, Chandran; Bando, Toshikazu; Nagase, Hiroki; Vaijayanthi, Thangavel; Taylor, Rhys D.; Sugiyama, Hiroshi

    2014-01-01

    The influential role of the epigenome in orchestrating genome-wide transcriptional activation instigates the demand for the artificial genetic switches with distinct DNA sequence recognition. Recently, we developed a novel class of epigenetically active small molecules called SAHA-PIPs by conjugating selective DNA binding pyrrole-imidazole polyamides (PIPs) with the histone deacetylase inhibitor SAHA. Screening studies revealed that certain SAHA-PIPs trigger targeted transcriptional activation of pluripotency and germ cell genes in mouse and human fibroblasts, respectively. Through microarray studies and functional analysis, here we demonstrate for the first time the remarkable ability of thirty-two different SAHA-PIPs to trigger the transcriptional activation of exclusive clusters of genes and noncoding RNAs. QRT-PCR validated the microarray data, and some SAHA-PIPs activated therapeutically significant genes like KSR2. Based on the aforementioned results, we propose the potential use of SAHA-PIPs as reagents capable of targeted transcriptional activation. PMID:24457603

  10. Molecular cloning and expression of a novel MYB transcription factor gene in rubber tree.

    PubMed

    Qin, Bi; Zhang, Yu; Wang, Meng

    2014-12-01

    MYB family proteins regulate a variety of cellular processes in plants. Tapping panel dryness (TPD) in rubber tree (Hevea brasiliensis Muell. Arg.) affects latex biosynthesis and causes serious losses to rubber producers. In this study, a novel SANT/MYB transcription factor gene down-regulated in TPD rubber tree, named as HbSM1, was isolated from rubber tree. The complete HbSM1 open reading frame (ORF) was 948 bp in length. The deduced HbSM1 protein is 315 amino acids. HbSM1 belonged to 1RMYB subfamily with a single SANT domain. Sequence alignment revealed that HbSM1 had high homology with MYB members from Ricinus communis and Manihot esculenta, with 72 and 78 % identity, respectively. Moreover, HbSM1 shared 56 % identity with Glycine max GmMYB176. Phylogenetic analysis revealed that HbSM1, GmMYB176, rice OsMYBS2, and OsMYBS3 fell into the same cluster with 93 % bootstrap support value. Comparing expression among different tissues demonstrated that HbSM1 was ubiquitously expressed in all tissues, but it appeared to be preferentially expressed in leaf and latex. Furthermore, HbSM1 transcripts were significantly induced by various phytohormones (including gibberellic acid, ethephon, methyl jasmonate, salicylic acid, and abscisic acid) and wounding treatments. These results suggested that HbSM1 might play multiple roles in plant development via different phytohormones signaling pathways. PMID:25195053

  11. Gene characterization and transcription analysis of two new ammonium transporters in pear rootstock (Pyrus betulaefolia).

    PubMed

    Li, Hui; Han, Jin-Long; Chang, You-Hong; Lin, Jing; Yang, Qing-Song

    2016-07-01

    Ammonium is the primarily nitrogen source for plant growth, but the molecular basis of ammonium acquisition in fruit species remains poorly understood. In this study, we report on the characterization of two new ammonium transporters (AMT) in the perennial tree Pyrus betulaefolia. In silico analyses and yeast complementation assays revealed that both PbAMT1;3 and PbAMT1;5 can be classified in the AMT1 sub-family. The specific expression of PbAMT1;3 in roots and of PbAMT1;5 in leaves indicates that they have diverse functions in ammonium uptake or transport in P. betulaefolia. Their expression was strongly influenced by ammonium availability. In addition, the transcript level of PbAMT1;5 was significantly affected by the diurnal cycle and senescence hormones. They conferred the ability to uptake nitrogen to the yeast strain 31019b; however, the (15)NH4 (+) uptake kinetics of PbAMT1;3 were different from those of PbAMT1;5. Indeed, PbAMT1;3 had a higher affinity for (15)NH4 (+), and pH changes were associated with this substrates' transport in yeast. The present study provides basic gene features and transcriptional information for the two new members of the AMT1 sub-family in P. betulaefolia and will aid in decoding the precise roles of AMTs in P. betulaefolia physiology. PMID:26943161

  12. Ribosome profiling reveals post-transcriptional buffering of divergent gene expression in yeast

    PubMed Central

    McManus, C. Joel; May, Gemma E.; Spealman, Pieter; Shteyman, Alan

    2014-01-01

    Understanding the patterns and causes of phenotypic divergence is a central goal in evolutionary biology. Much work has shown that mRNA abundance is highly variable between closely related species. However, the extent and mechanisms of post-transcriptional gene regulatory evolution are largely unknown. Here we used ribosome profiling to compare transcript abundance and translation efficiency in two closely related yeast species (S. cerevisiae and S. paradoxus). By comparing translation regulatory divergence to interspecies differences in mRNA sequence features, we show that differences in transcript leaders and codon bias substantially contribute to divergent translation. Globally, we find that translation regulatory divergence often buffers species differences in mRNA abundance, such that ribosome occupancy is more conserved than transcript abundance. We used allele-specific ribosome profiling in interspecies hybrids to compare the relative contributions of cis- and trans-regulatory divergence to species differences in mRNA abundance and translation efficiency. The mode of gene regulatory divergence differs for these processes, as trans-regulatory changes play a greater role in divergent mRNA abundance than in divergent translation efficiency. Strikingly, most genes with aberrant transcript abundance in F1 hybrids (either over- or underexpressed compared to both parent species) did not exhibit aberrant ribosome occupancy. Our results show that interspecies differences in translation contribute substantially to the evolution of gene expression. Compensatory differences in transcript abundance and translation efficiency may increase the robustness of gene regulation. PMID:24318730

  13. Encoding four gene expression programs in the activation dynamics of a single transcription factor.

    PubMed

    Hansen, Anders S; O'Shea, Erin K

    2016-04-01

    Cellular signaling response pathways often exhibit a bow-tie topology [1,2]: multiple upstream stress signals converge on a single shared transcription factor, which is thought to induce different downstream gene expression programs (Figure 1A). However, if several different signals activate the same transcription factor, can each signal then induce a specific gene expression response? A growing body of literature supports a temporal coding theory where information about environmental signals can be encoded, at least partially, in the temporal dynamics of the shared transcription factor [1,2]. For example, in the case of the budding yeast transcription factor Msn2, different stresses induce distinct Msn2 activation dynamics: Msn2 shows pulsatile nuclear activation with dose-dependent frequency under glucose limitation, but sustained nuclear activation with dose-dependent amplitude under oxidative stress [3]. These dynamic patterns can then lead to differential gene expression responses [3-5], but it is not known how much specificity can be obtained. Thus, a major question of this temporal coding theory is how many gene response programs or cellular functions can be robustly encoded by dynamic control of a single transcription factor. Here we provide the first direct evidence that, simply by regulating the activation dynamics of a single transcription factor, it is possible to preferentially induce four distinct gene expression programs. PMID:27046808

  14. African Swine Fever Virus Multigene Family 360 and 530 Genes Affect Host Interferon Response

    PubMed Central

    Afonso, C. L.; Piccone, M. E.; Zaffuto, K. M.; Neilan, J.; Kutish, G. F.; Lu, Z.; Balinsky, C. A.; Gibb, T. R.; Bean, T. J.; Zsak, L.; Rock, D. L.

    2004-01-01

    African swine fever virus (ASFV) multigene family 360 and 530 (MGF360/530) genes affect viral growth in macrophage cell cultures and virulence in pigs (L. Zsak, Z. Lu, T. G. Burrage, J. G. Neilan, G. F. Kutish, D. M. Moore, and D. L. Rock, J. Virol. 75:3066-3076, 2001). The mechanism by which these novel genes affect virus-host interactions is unknown. To define MGF360/530 gene function, we compared macrophage transcriptional responses following infection with parental ASFV (Pr4) and an MGF360/530 deletion mutant (Pr4Δ35). A swine cDNA microarray containing 7,712 macrophage cDNA clones was used to compare the transcriptional profiles of swine macrophages infected with Pr4 and Pr4Δ35 at 3 and 6 h postinfection (hpi). While at 3 hpi most (7,564) of the genes had similar expression levels in cells infected with either virus, 38 genes had significantly increased (>2.0-fold, P < 0.05) mRNA levels in Pr4Δ35-infected macrophages. Similar up-regulation of these genes was observed at 6 hpi. Viral infection was required for this induced transcriptional response. Most Pr4Δ35 up-regulated genes were part of a type I interferon (IFN) response or were genes that are normally induced by double-stranded RNA and/or viral infection. These included monocyte chemoattractant protein, transmembrane protein 3, tetratricopeptide repeat protein 1, a ubiquitin-like 17-kDa protein, ubiquitin-specific protease ISG43, an RNA helicase DEAD box protein, GTP-binding MX protein, the cytokine IP-10, and the PKR activator PACT. Differential expression of IFN early-response genes in Pr4Δ35 relative to Pr4 was confirmed by Northern blot analysis and real-time PCR. Analysis of IFN-α mRNA and secreted IFN-α levels at 3, 8, and 24 hpi revealed undetectable IFN-α in mock- and Pr4-infected macrophages but significant IFN-α levels at 24 hpi in Pr4Δ35-infected macrophages. The absence of IFN-α in Pr4-infected macrophages suggests that MGF360/530 genes either directly or indirectly suppress a type

  15. African swine fever virus multigene family 360 and 530 genes affect host interferon response.

    PubMed

    Afonso, C L; Piccone, M E; Zaffuto, K M; Neilan, J; Kutish, G F; Lu, Z; Balinsky, C A; Gibb, T R; Bean, T J; Zsak, L; Rock, D L

    2004-02-01

    African swine fever virus (ASFV) multigene family 360 and 530 (MGF360/530) genes affect viral growth in macrophage cell cultures and virulence in pigs (L. Zsak, Z. Lu, T. G. Burrage, J. G. Neilan, G. F. Kutish, D. M. Moore, and D. L. Rock, J. Virol. 75:3066-3076, 2001). The mechanism by which these novel genes affect virus-host interactions is unknown. To define MGF360/530 gene function, we compared macrophage transcriptional responses following infection with parental ASFV (Pr4) and an MGF360/530 deletion mutant (Pr4 Delta 35). A swine cDNA microarray containing 7,712 macrophage cDNA clones was used to compare the transcriptional profiles of swine macrophages infected with Pr4 and Pr4 Delta 35 at 3 and 6 h postinfection (hpi). While at 3 hpi most (7,564) of the genes had similar expression levels in cells infected with either virus, 38 genes had significantly increased (>2.0-fold, P < 0.05) mRNA levels in Pr4 Delta 35-infected macrophages. Similar up-regulation of these genes was observed at 6 hpi. Viral infection was required for this induced transcriptional response. Most Pr Delta 35 up-regulated genes were part of a type I interferon (IFN) response or were genes that are normally induced by double-stranded RNA and/or viral infection. These included monocyte chemoattractant protein, transmembrane protein 3, tetratricopeptide repeat protein 1, a ubiquitin-like 17-kDa protein, ubiquitin-specific protease ISG43, an RNA helicase DEAD box protein, GTP-binding MX protein, the cytokine IP-10, and the PKR activator PACT. Differential expression of IFN early-response genes in Pr4 Delta 35 relative to Pr4 was confirmed by Northern blot analysis and real-time PCR. Analysis of IFN-alpha mRNA and secreted IFN-alpha levels at 3, 8, and 24 hpi revealed undetectable IFN-alpha in mock- and Pr4-infected macrophages but significant IFN-alpha levels at 24 hpi in Pr4 Delta 35-infected macrophages. The absence of IFN-alpha in Pr4-infected macrophages suggests that MGF360/530 genes

  16. Chromosomal clustering and GATA transcriptional regulation of intestine-expressed genes in C. elegans.

    PubMed

    Pauli, Florencia; Liu, Yueyi; Kim, Yoona A; Chen, Pei-Jiun; Kim, Stuart K

    2006-01-01

    We used mRNA tagging to identify genes expressed in the intestine of C. elegans. Animals expressing an epitope-tagged protein that binds the poly-A tail of mRNAs (FLAG::PAB-1) from an intestine-specific promoter (ges-1) were used to immunoprecipitate FLAG::PAB-1/mRNA complexes from the intestine. A total of 1938 intestine-expressed genes (P<0.001) were identified using DNA microarrays. First, we compared the intestine-expressed genes with those expressed in the muscle and germline, and identified 510 genes enriched in all three tissues and 624 intestine-, 230 muscle- and 1135 germ line-enriched genes. Second, we showed that the 1938 intestine-expressed genes were physically clustered on the chromosomes, suggesting that the order of genes in the genome is influenced by the effect of chromatin domains on gene expression. Furthermore, the commonly expressed genes showed more chromosomal clustering than the tissue-enriched genes, suggesting that chromatin domains may influence housekeeping genes more than tissue-specific genes. Third, in order to gain further insight into the regulation of intestinal gene expression, we searched for regulatory motifs. This analysis found that the promoters of the intestine genes were enriched for the GATA transcription factor consensus binding sequence. We experimentally verified these results by showing that the GATA motif is required in cis and that GATA transcription factors are required in trans for expression of these intestinal genes. PMID:16354718

  17. Cytogenetic and molecular localization of tipE: A gene affecting sodium channels in Drosophila melanogaster

    SciTech Connect

    Feng, G.; Deak, P.; Hall, L.M.

    1995-04-01

    Voltage-sensitive sodium channels play a key role in nerve cells where they are responsible for the increase in sodium permeability during the rising phase of action potentials. In Drosophila melanogaster a subset of temperature-sensitive paralytic mutations affect sodium channel function. One such mutation is temperature-induced paralysis locus E (tipE), which has been shown by electrophysiology and ligand binding studies to reduce sodium channel numbers. Three new {gamma}-ray-induced tipE alleles associated with either visible deletions in 64AB or a translocation breakpoint within 64B2 provide landmarks for positional cloning of tipE. Beginning with the flanking cloned gene Ras2, a 140-kb walk across the translocation breakpoint was completed. Germline transformation using a 42-kb cosmid clone and successively smaller subclones localized the tipE gene within a 7.4-kb genomic DNA segment. Although this chromosome region is rich in transcripts, only three overlapping mRNAs (5.4, 4.4, and 1.7 kb) lie completely within the smallest rescuing construct. The small sizes of the rescuing construct and transcripts suggests that tipE does not encode a standard sodium channel {alpha}-subunit with four homologous repeats. Sequencing these transcripts will elucidate the role of the tipE gene product in sodium channel functional regulation. 55 refs., 4 figs., 2 tabs.

  18. Non-Viral Gene Transfer as a Tool for Studying Transcription Regulation of Xenobiotic Metabolizing Enzymes

    PubMed Central

    Bonamassa, Barbara; Liu, Dexi

    2010-01-01

    Numerous xenobiotic metabolizing enzymes are regulated by nuclear receptors at transcriptional level. The challenge we currently face is to understand how a given nuclear receptor interacts with its xenobiotics, migrates into nucleus, binds to the xenobiotic response element of a target gene, and regulates transcription. Toward this end, new methods have been developed to introduce the nuclear receptor gene into appropriate cells and study its activity in activating reporter gene expression under the control of a promoter containing xenobiotic response elements. The goal of this review is to critically examine the gene transfer methods currently available. We concentrate on the gene transfer mechanism, advantages and limitations of each method when employed for nuclear receptor-mediated gene regulation studies. It is our hope that the information provided highlights the importance of gene transfer in studying the mechanisms by which our body eliminates the potentially harmful substances and maintains the homeostasis. PMID:20713102

  19. Bypassing the Requirements for Epigenetic Modifications in Gene Transcription by Increasing Enhancer Strength▿

    PubMed Central

    Koutroubas, George; Merika, Menie; Thanos, Dimitris

    2008-01-01

    Our current concept postulates that histone acetylation is required for the recruitment of bromodomain-containing transcription complexes, such as the chromatin-remodeling machine SWI/SNF and the basal transcription factor TFIID. We generated simple NF-κB-dependent enhancers of increasing transcriptional strengths and found that the histone acetylation requirements for activation of transcription depended on the strengths of these enhancers. All enhancers function by recruiting SWI/SNF and TFIID to induce nucleosome sliding, a prerequisite for transcriptional activation. However, histone acetylation, although it occurs, is dispensable for TFIID and SWI/SNF recruitment by the strong enhancers, indicating that strong activators can overcome the chromatin barrier by directly recruiting the necessary transcriptional complexes. Weak enhancers depend on histone acetylation for recruitment, and this requirement is independent of a histone acetylation code. Thus, the need for nucleosome modifications is imposed on genes and translated according to the quality and strengths of the activators. PMID:18025106

  20. Biomarkers and transcription levels of cancer-related genes in cockles Cerastoderma edule from Galicia (NW Spain) with disseminated neoplasia.

    PubMed

    Ruiz, Pamela; Díaz, Seila; Orbea, Amaia; Carballal, Maria J; Villalba, Antonio; Cajaraville, Miren P

    2013-07-15

    Disseminated neoplasia (DN) is a pathological condition reported for several species of marine bivalves throughout the world, but its aetiology has not yet been satisfactorily explained. It has been suggested that chemical contamination could be a factor contributing to neoplasia. The aim of the present study was to compare cell and tissue biomarkers and the transcription level of cancer-related genes in cockles (Cerastoderma edule) affected by DN with those of healthy cockles in relation to chemical contaminant burdens. For this, cockles were collected from a natural bed in Cambados (Ria de Arousa, Galicia) in May 2009. The prevalence of DN was 12.36% and 3 degrees of DN severity were distinguished. No significant differences in metal accumulation, non-specific inflammatory responses and parasites were observed between healthy and DN-affected cockles. Lysosomal membrane stability was significantly reduced in cockles affected by DN, which indicates a poorer health condition. Very low frequencies of micronuclei were recorded and no significant differences were detected between DN severity groups. Haemolymph analyses showed a higher frequency of mitotic figures and binucleated cells in cockles affected by moderate and heavy DN than in healthy ones. Neoplastic animals showed significantly higher transcription levels of p53 and ras than healthy cockles and mutational alterations in ras gene sequence were detected. Low concentrations of metals, polycyclic aromatic hydrocarbons, polychlorinated biphenyls and phthalate esters were measured in cockles from Cambados. In conclusion, cockles affected by DN suffer a general stress situation and have altered patterns of cancer-related gene transcription. Further studies are in progress to elucidate mechanisms of carcinogenesis in this species. PMID:23665240

  1. Transcription of Interleukin-8: How Altered Regulation Can Affect Cystic Fibrosis Lung Disease.

    PubMed

    Jundi, Karim; Greene, Catherine M

    2015-01-01

    Interleukin-8 (IL-8) is a neutrophil chemokine that is encoded on the CXCL8 gene. Normally CXCL8 expression is repressed due to histone deacetylation, octamer-1 binding to the promoter and the inhibitory effect of nuclear factor-κB repressing factor (NRF). However, in response to a suitable stimulus, the human CXCL8 gene undergoes transcription due to its inducible promoter that is regulated by the transcription factors nuclear factor-κB (NF-κB), activating protein (AP-1), CAAT/enhancer-binding protein β (C/EBPβ, also known as NF-IL-6), C/EBP homologous protein (CHOP) and cAMP response element binding protein (CREB). CXCL8 mRNA is then stabilised by the activity of p38 mitogen-activated protein kinase (p38 MAPK). Cystic fibrosis (CF) lung disease is characterised by a neutrophil-dominated airway inflammatory response. A major factor contributing to the large number of neutrophils is the higher than normal levels of IL-8 that are present within the CF lung. Infection and inflammation, together with intrinsic alterations in CF airway cells are responsible for the abnormally high intrapulmonary levels of IL-8. Strategies to inhibit aberrantly high CXCL8 expression hold therapeutic potential for CF lung disease. PMID:26140537

  2. Replication-induced transcription of an autorepressed gene: The replication initiator gene of plasmid P1

    PubMed Central

    Mukhopadhyay, Suman; Chattoraj, Dhruba K.

    2000-01-01

    The replication origin of plasmid P1 contains an array of five repeats (iterons) that bind the plasmid-encoded initiator RepA. Within the array lies the repA promoter, which becomes largely repressed on RepA binding (autorepression). One might expect that extra iterons produced on plasmid replication would titrate RepA and release the repression. The promoter, however, is induced poorly by extra iterons. The P1 copy number is reduced by extra iterons in the presence of the autorepressed repA gene but not when additional RepA is provided from constitutive sources. It has been proposed that the iteron-bound RepA couples with the promoter-bound RepA and thereby maintains repression. Although not the product of replication, we find that the act of replication itself can renew RepA synthesis. Replication apparently cleans the promoter of bound RepA and provides a window of opportunity for repA transcription. We propose that replication-induced transcription is required to ensure initiator availability in a system that is induced poorly when challenged with additional initiator binding sites. PMID:10840063

  3. Environmental associations with gene transcription in Babine Lake rainbow trout: evidence for local adaptation

    PubMed Central

    Wellband, Kyle W; Heath, Daniel D

    2013-01-01

    The molecular genetic mechanisms facilitating local adaptation in salmonids continue to be poorly characterized. Gene transcription is a highly regulated step in the expression of a phenotype and it has been shown to respond to selection and thus may be one mechanism that facilitates the development of local adaptation. Advances in molecular genetic tools and an increased understanding of the functional roles of specific genes allow us to test hypotheses concerning the role of variable environments in shaping transcription at known-function candidate loci. To address these hypotheses, wild rainbow trout were collected in their first summer and subjected to metabolic and immune challenges. We assayed gene transcription at candidate loci that play a role in the molecular genetic response to these stresses, and correlated transcription with temperature data from the streams and the abundance and diversity of bacteria as characterized by massively parallel pyrosequencing. Patterns of transcriptional regulation from resting to induced levels varied among populations for both treatments. Co-inertia analysis demonstrated significant associations between resting levels of metabolic gene transcription and thermal regime (R2 = 0.19, P = 0.013) as well as in response to challenge (R2 = 0.39, P = 0.001) and resting state and challenged levels of cytokine gene transcription with relative abundances of bacteria (resting: R2 = 0.25, P = 0.009, challenged: R2 = 0.65, P = 0.001). These results show that variable environments, even within a small geographic range (<250 km), can drive divergent selection among populations for transcription of genes related to surviving stress. PMID:23762507

  4. The β-cell specific transcription factor Nkx6.1 inhibits glucagon gene transcription by interfering with Pax6

    PubMed Central

    Gauthier, Benoit R.; Gosmain, Yvan; Mamin, Aline; Philippe, Jacques

    2007-01-01

    The transcription factor Nkx6.1 is required for the establishment of functional insulin-producing β-cells in the endocrine pancreas. Overexpression of Nkx6.1 has been shown to inhibit glucagon gene expression while favouring insulin gene activation. Down-regulation resulted in the opposite effect, suggesting that absence of Nkx6.1 favours glucagon gene expression. To understand the mechanism by which Nkx6.1 suppresses glucagon gene expression, we studied its effect on the glucagon gene promoter activity in non-islet cells using transient transfections and gel-shift analyses. In glucagonoma cells transfected with an Nkx6.1-encoding vector, the glucagon promoter activity was reduced by 65%. In BHK21 cells, Nkx6.1 inhibited by 93% Pax6-mediated activation of the glucagon promoter, whereas Cdx2/3 and Maf stimulations were unaltered. Although Nkx6.1 could interact with both the G1 and G3 element, only the former displayed specificity for Nkx6.1. Mutagenesis of the three potential AT-rich motifs within the G1 revealed that only the Pax6-binding site preferentially interacted with Nkx6.1. Chromatin immunoprecipitation confirmed interaction of Nkx6.1 with the glucagon promoter and revealed a direct competition for binding between Pax6 and Nkx6.1. A weak physical interaction between Pax6 and Nkx6.1 was detected in vitro and in vivo suggesting that Nkx6.1 predominantly inhibits glucagon gene transcription through G1-binding competition. We suggest that cell-specific expression of the glucagon gene may only proceed when Nkx6.1, in combination with Pdx1 and Pax4, are silenced in early α-cell precursors. PMID:17263687

  5. Nature of bacterial colonization influences transcription of mucin genes in mice during the first week of life

    PubMed Central

    2012-01-01

    Background Postnatal regulation of the small intestinal mucus layer is potentially important in the development of adult gut functionality. We hypothesized that the nature of bacterial colonization affects mucus gene regulation in early life. We thus analyzed the influence of the presence of a conventional microbiota as well as two selected monocolonizing bacterial strains on the transcription of murine genes involved in mucus layer development during the first week of life. Mouse pups (N = 8/group) from differently colonized dams: Germ-free (GF), conventional specific pathogen free (SPF), monocolonized with either Lactobacillus acidophilus NCFM (Lb) or Escherichia coli Nissle (Ec) were analyzed by qPCR on isolated ileal tissue sections from postnatal days 1 and 6 (PND1, PND6) after birth with respect to: (i) transcription of specific genes involved in mucus production (Muc1-4, Tff3) and (ii) amounts of 16S rRNA of Lactobacillus and E. coli. Quantification of 16S rRNA genes was performed to obtain a measure for amounts of colonized bacteria. Results We found a microbiota-independent transcriptional increase of all five mucus genes from PND1 to PND6. Furthermore, the relative level of transcription of certain mucus genes on PND1 was increased by the presence of bacteria. This was observed for Tff3 in the SPF, Ec, and Lb groups; for Muc2 in SPF; and for Muc3 and Muc4 in Ec and Lb, respectively. Detection of bacterial 16S rRNA genes levels above the qPCR detection level occurred only on PND6 and only for some of the colonized animals. On PND6, we found significantly lower levels of Muc1, Muc2 and Muc4 gene transcription for Lb animals with detectable Lactobacillus levels as compared to animals with Lactobacillus levels below the detection limit. Conclusions In summary, our data show that development of the expression of genes encoding secreted (Muc2/Tff3) and membrane-bound (Muc1/Muc3/Muc4) mucus regulatory proteins, respectively, is distinct and that the onset of

  6. Mitotic genes are transcriptionally upregulated in the fibroblast irradiated with very low doses of UV-C

    PubMed Central

    Takeuchi, Seiji; Matsuda, Toshiro; Ono, Ryusuke; Tsujimoto, Mariko; Nishigori, Chikako

    2016-01-01

    Ultraviolet (UV) radiation induces a variety of biological effects, including DNA damage response and cell signaling pathways. We performed transcriptome analysis using microarray in human primary cultured fibroblasts irradiated with UV-C (0.5 or 5 J/m2) and harvested at 4 or 12 h following UV exposure. All transcript data were analyzed by comparison with the corresponding results in non-irradiated (control) cells. The number of genes with significantly altered expression (≥2-fold difference relative to the control) is higher in the sample irradiated with high dose of UV, suggesting that gene expression was UV dose-dependent. Pathway analysis on the upregulated genes at 12 h indicates that the expression of some cell cycle-related genes was predominantly induced irrespective of UV-dose. Interestingly, almost all the genes with significant altered expression were cell cycle-related genes designated as ‘Mitotic Genes’, which function in the spindle assembly checkpoint. Therefore, even a low dose of UV could affect the transcriptional profile. PMID:27378355

  7. Chemosensory mediated behaviors and gene transcription profiles in wild yellow perch (Perca flavescens) from metal contaminated lakes.

    PubMed

    Azizishirazi, Ali; Dew, William A; Bougas, Berenice; Dashtban, Mehdi; Bernatchez, Louis; Pyle, Greg G

    2014-08-01

    The olfactory system of fish is sensitive to the toxic effects of low concentrations of contaminants. To investigate the effects of long-term metal exposure on olfaction in wild yellow perch (Perca flavescens), fish from one clean (Geneva Lake) and two metal-contaminated lakes (Ramsey and Hannah lakes) were collected in and around the metal-mining district of Sudbury, ON. Two different techniques were used to measure the effects of exposure to environmental contamination: (i) behavioral responses were recorded in response to conspecific skin extract and (ii) gene transcription differences in olfactory rosettes were characterized using a novel, 1000-candidate gene yellow perch microarray. Behavioral assays performed on fish from the clean lake demonstrated avoidance of a conspecific skin extract, while fish from metal contaminated lakes showed no avoidance response. A total of 109 out of the 1000 genes were differentially transcribed among the lakes. Most of the differentially transcribed genes were between the two metal contaminated lakes relative to either of the contaminated lakes and the reference lake. No genes were differentially expressed between Geneva Lake (clean) and Hannah Lake (metal contaminated). These results demonstrated that even though the different populations of fish from both Hannah and Ramey lakes were affected at the behavioral level, the impairment of olfaction was not measurable using gene transcriptional changes in olfactory rosettes. PMID:24859710

  8. A Myb-related transcription factor is involved in the phytochrome regulation of an Arabidopsis Lhcb gene.

    PubMed Central

    Wang, Z Y; Kenigsbuch, D; Sun, L; Harel, E; Ong, M S; Tobin, E M

    1997-01-01

    We have isolated the gene for a protein designated CCA1. This protein can bind to a region of the promoter of an Arabidopsis light-harvesting chlorophyll a/b protein gene, Lhcb1*3, which is necessary for its regulation by phytochrome. The CCA1 protein interacted with two imperfect repeats in the Lhcb1*3 promoter, AAA/cAATCT, a sequence that is conserved in Lhcb genes. A region near the N terminus of CCA1, which has some homology to the repeated sequence found in the DNA binding domain of Myb proteins, is required for binding to the Lhcb1*3 promoter. Lines of transgenic Arabidopsis plants expressing antisense RNA for CCA1 showed reduced phytochrome induction of the endogenous Lhcb1*3 gene, whereas expression of another phytochrome-regulated gene, rbcS-1A, which encodes the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, was not affected. Thus, the CCA1 protein acts as a specific activator of Lhcb1*3 transcription in response to brief red illumination. The expression of CCA1 RNA was itself transiently increased when etiolated seedlings were transferred to light. We conclude that the CCA1 protein is a key element in the functioning of the phytochrome signal transduction pathway leading to increased transcription of this Lhcb gene in Arabidopsis. PMID:9144958

  9. Exploiting single-molecule transcript sequencing for eukaryotic gene prediction.

    PubMed

    Minoche, André E; Dohm, Juliane C; Schneider, Jessica; Holtgräwe, Daniela; Viehöver, Prisca; Montfort, Magda; Sörensen, Thomas Rosleff; Weisshaar, Bernd; Himmelbauer, Heinz

    2015-01-01

    We develop a method to predict and validate gene models using PacBio single-molecule, real-time (SMRT) cDNA reads. Ninety-eight percent of full-insert SMRT reads span complete open reading frames. Gene model validation using SMRT reads is developed as automated process. Optimized training and prediction settings and mRNA-seq noise reduction of assisting Illumina reads results in increased gene prediction sensitivity and precision. Additionally, we present an improved gene set for sugar beet (Beta vulgaris) and the first genome-wide gene set for spinach (Spinacia oleracea). The workflow and guidelines are a valuable resource to obtain comprehensive gene sets for newly sequenced genomes of non-model eukaryotes. PMID:26328666

  10. Directional gene expression and antisense transcripts in sexual and asexual stages of Plasmodium falciparum

    PubMed Central

    2011-01-01

    Background It has been shown that nearly a quarter of the initial predicted gene models in the Plasmodium falciparum genome contain errors. Although there have been efforts to obtain complete cDNA sequences to correct the errors, the coverage of cDNA sequences on the predicted genes is still incomplete, and many gene models for those expressed in sexual or mosquito stages have not been validated. Antisense transcripts have widely been reported in P. falciparum; however, the extent and pattern of antisense transcripts in different developmental stages remain largely unknown. Results We have sequenced seven bidirectional libraries from ring, early and late trophozoite, schizont, gametocyte II, gametocyte V, and ookinete, and four strand-specific libraries from late trophozoite, schizont, gametocyte II, and gametocyte V of the 3D7 parasites. Alignment of the cDNA sequences to the 3D7 reference genome revealed stage-specific antisense transcripts and novel intron-exon splicing junctions. Sequencing of strand-specific cDNA libraries suggested that more genes are expressed in one direction in gametocyte than in schizont. Alternatively spliced genes, antisense transcripts, and stage-specific expressed genes were also characterized. Conclusions It is necessary to continue to sequence cDNA from different developmental stages, particularly those of non-erythrocytic stages. The presence of antisense transcripts in some gametocyte and ookinete genes suggests that these antisense RNA may play an important role in gene expression regulation and parasite development. Future gene expression studies should make use of directional cDNA libraries. Antisense transcripts may partly explain the observed discrepancy between levels of mRNA and protein expression. PMID:22129310

  11. Transcriptional Repression by the BRG1-SWI/SNF Complex Affects the Pluripotency of Human Embryonic Stem Cells

    PubMed Central

    Zhang, Xiaoli; Li, Bing; Li, Wenguo; Ma, Lijuan; Zheng, Dongyan; Li, Leping; Yang, Weijing; Chu, Min; Chen, Wei; Mailman, Richard B.; Zhu, Jun; Fan, Guoping; Archer, Trevor K.; Wang, Yuan

    2014-01-01

    Summary The SWI/SNF complex plays an important role in mouse embryonic stem cells (mESCs), but it remains to be determined whether this complex is required for the pluripotency of human ESCs (hESCs). Using RNAi, we demonstrated that depletion of BRG1, the catalytic subunit of the SWI/SNF complex, led to impaired self-renewing ability and dysregulated lineage specification of hESCs. A unique composition of the BRG1-SWI/SNF complex in hESCs was further defined by the presence of BRG1, BAF250A, BAF170, BAF155, BAF53A, and BAF47. Genome-wide expression analyses revealed that BRG1 participated in a broad range of biological processes in hESCs through pathways different from those in mESCs. In addition, chromatin immunoprecipitation sequencing (ChIP-seq) demonstrated that BRG1 played a repressive role in transcriptional regulation by modulating the acetylation levels of H3K27 at the enhancers of lineage-specific genes. Our data thus provide valuable insights into molecular mechanisms by which transcriptional repression affects the self-renewal and differentiation of hESCs. PMID:25241744

  12. A Novel PCR Assay for Listeria welshimeri Targeting Transcriptional Regulator Gene lwe1801

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transcriptional regulator genes encode a group of specialized molecules that play essential roles in microbial responses to changing external conditions. These genes have been shown to possess species or group specificity and are useful as detection targets for diagnostic application. The present st...

  13. Autogenous Regulation of Splicing of the Transcript of a Yeast Ribosomal Protein Gene

    NASA Astrophysics Data System (ADS)

    Dabeva, Mariana D.; Post-Beittenmiller, Martha A.; Warner, Jonathan R.

    1986-08-01

    The gene for a yeast ribosomal protein, RPL32, contains a single intron. The product of this gene appears to participate in feedback control of the splicing of the intron from the transcript. This autogenous regulation of splicing provides a striking analogy to the autogenous regulation of translation of ribosomal proteins in Escherichia coli.

  14. Transcriptional activation of jun and actin genes by estrogen during mitogenic stimulation of rat uterine cells.

    PubMed

    Cicatiello, L; Ambrosino, C; Coletta, B; Scalona, M; Sica, V; Bresciani, F; Weisz, A

    1992-03-01

    Estrogens induce transcriptional activation of c-fos and c-myc proto-oncogenes during mitogenic stimulation of human, chicken, mouse and rat cells in vivo and in vitro. In this paper we show that 17 beta-estradiol injected into adult ovariectomized rats increases c-jun, jun-B and jun-D gene transcription in the uterus. Kinetics and amplitude of response are different for each gene, since c-jun is activated first, within 30 min after injection, followed by jun-D and jun-B, 60 and 90 min after injection, respectively. Maximal activation of jun-B marks a drop in transcription of all the jun genes. Furthermore, transcriptional activation by 17 beta-estradiol of the growth-regulated beta- and gamma-cytoskeletal actin genes is prevented by an inhibitor of protein synthesis, indicating that it is a secondary response to the hormone. These data support the hypothesis that during growth stimulation of target cells the estrogen receptor induces transcription of regulatory genes, triggering in this way a cascade of gene regulation events that results in progression through the cell cycle. PMID:1373300

  15. Master transcription factors and mediator establish super-enhancers at key cell identity genes.

    PubMed

    Whyte, Warren A; Orlando, David A; Hnisz, Denes; Abraham, Brian J; Lin, Charles Y; Kagey, Michael H; Rahl, Peter B; Lee, Tong Ihn; Young, Richard A

    2013-04-11

    Master transcription factors Oct4, Sox2, and Nanog bind enhancer elements and recruit Mediator to activate much of the gene expression program of pluripotent embryonic stem cells (ESCs). We report here that the ESC master transcription factors form unusual enhancer domains at most genes that control the pluripotent state. These domains, which we call super-enhancers, consist of clusters of enhancers that are densely occupied by the master regulators and Mediator. Super-enhancers differ from typical enhancers in size, transcription factor density and content, ability to activate transcription, and sensitivity to perturbation. Reduced levels of Oct4 or Mediator cause preferential loss of expression of super-enhancer-associated genes relative to other genes, suggesting how changes in gene expression programs might be accomplished during development. In other more differentiated cells, super-enhancers containing cell-type-specific master transcription factors are also found at genes that define cell identity. Super-enhancers thus play key roles in the control of mammalian cell identity. PMID:23582322

  16. Scaling of Gene Expression with Transcription-Factor Fugacity

    NASA Astrophysics Data System (ADS)

    Weinert, Franz M.; Brewster, Robert C.; Rydenfelt, Mattias; Phillips, Rob; Kegel, Willem K.

    2014-12-01

    The proteins associated with gene regulation are often shared between multiple pathways simultaneously. By way of contrast, models in regulatory biology often assume these pathways act independently. We demonstrate a framework for calculating the change in gene expression for the interacting case by decoupling repressor occupancy across the cell from the gene of interest by way of a chemical potential. The details of the interacting regulatory architecture are encompassed in an effective concentration, and thus, a single scaling function describes a collection of gene expression data from diverse regulatory situations and collapses it onto a single master curve.

  17. Scaling of gene expression with transcription-factor fugacity.

    PubMed

    Weinert, Franz M; Brewster, Robert C; Rydenfelt, Mattias; Phillips, Rob; Kegel, Willem K

    2014-12-19

    The proteins associated with gene regulation are often shared between multiple pathways simultaneously. By way of contrast, models in regulatory biology often assume these pathways act independently. We demonstrate a framework for calculating the change in gene expression for the interacting case by decoupling repressor occupancy across the cell from the gene of interest by way of a chemical potential. The details of the interacting regulatory architecture are encompassed in an effective concentration, and thus, a single scaling function describes a collection of gene expression data from diverse regulatory situations and collapses it onto a single master curve. PMID:25554908

  18. Scaling of Gene Expression with Transcription-Factor Fugacity

    PubMed Central

    Weinert, Franz M.; Brewster, Robert C.; Rydenfelt, Mattias; Phillips, Rob; Kegel, Willem K.

    2015-01-01

    The proteins associated with gene regulation are often shared between multiple pathways simultaneously. By way of contrast, models in regulatory biology often assume these pathways act independently. We demonstrate a framework for calculating the change in gene expression for the interacting case by decoupling repressor occupancy across the cell from the gene of interest by way of a chemical potential. The details of the interacting regulatory architecture are encompassed in an effective concentration, and thus, a single scaling function describes a collection of gene expression data from diverse regulatory situations and collapses it onto a single master curve. PMID:25554908

  19. Patterns of Transcript Abundance of Eukaryotic Biogeochemically-Relevant Genes in the Amazon River Plume.

    PubMed

    Zielinski, Brian L; Allen, Andrew E; Carpenter, Edward J; Coles, Victoria J; Crump, Byron C; Doherty, Mary; Foster, Rachel A; Goes, Joaquim I; Gomes, Helga R; Hood, Raleigh R; McCrow, John P; Montoya, Joseph P; Moustafa, Ahmed; Satinsky, Brandon M; Sharma, Shalabh; Smith, Christa B; Yager, Patricia L; Paul, John H

    2016-01-01

    The Amazon River has the largest discharge of all rivers on Earth, and its complex plume system fuels a wide array of biogeochemical processes, across a large area of the western tropical North Atlantic. The plume thus stimulates microbial processes affecting carbon sequestration and nutrient cycles at a global scale. Chromosomal gene expression patterns of the 2.0 to 156 μm size-fraction eukaryotic microbial community were investigated in the Amazon River Plume, generating a robust dataset (more than 100 million mRNA sequences) that depicts the metabolic capabilities and interactions among the eukaryotic microbes. Combining classical oceanographic field measurements with metatranscriptomics yielded characterization of the hydrographic conditions simultaneous with a quantification of transcriptional activity and identity of the community. We highlight the patterns of eukaryotic gene expression for 31 biogeochemically significant gene targets hypothesized to be valuable within forecasting models. An advantage to this targeted approach is that the database of reference sequences used to identify the target genes was selectively constructed and highly curated optimizing taxonomic coverage, throughput, and the accuracy of annotations. A coastal diatom bloom highly expressed nitrate transporters and carbonic anhydrase presumably to support high growth rates and enhance uptake of low levels of dissolved nitrate and CO2. Diatom-diazotroph association (DDA: diatoms with nitrogen fixing symbionts) blooms were common when surface salinity was mesohaline and dissolved nitrate concentrations were below detection, and hence did not show evidence of nitrate utilization, suggesting they relied on ammonium transporters to aquire recently fixed nitrogen. These DDA blooms in the outer plume had rapid turnover of the photosystem D1 protein presumably caused by photodegradation under increased light penetration in clearer waters, and increased expression of silicon transporters as

  20. Insight on Genes Affecting Tuber Development in Potato upon Potato spindle tuber viroid (PSTVd) Infection

    PubMed Central

    Zhang, Runxuan; Bonar, Nicola; Morris, Jenny; Hedley, Pete E.; Bryan, Glenn J.; Kalantidis, Kriton; Hornyik, Csaba

    2016-01-01

    Potato (Solanum tuberosum L) is a natural host of Potato spindle tuber viroid (PSTVd) which can cause characteristic symptoms on developing plants including stunting phenotype and distortion of leaves and tubers. PSTVd is the type species of the family Pospiviroidae, and can replicate in the nucleus and move systemically throughout the plant. It is not well understood how the viroid can affect host genes for successful invasion and which genes show altered expression levels upon infection. Our primary focus in this study is the identification of genes which can affect tuber formation since viroid infection can strongly influence tuber development and especially tuber shape. In this study, we used a large-scale method to identify differentially expressed genes in potato. We have identified defence, stress and sugar metabolism related genes having altered expression levels upon infection. Additionally, hormone pathway related genes showed significant up- or down-regulation. DWARF1/DIMINUTO, Gibberellin 7-oxidase and BEL5 transcripts were identified and validated showing differential expression in viroid infected tissues. Our study suggests that gibberellin and brassinosteroid pathways have a possible role in tuber development upon PSTVd infection. PMID:26937634

  1. Insight on Genes Affecting Tuber Development in Potato upon Potato spindle tuber viroid (PSTVd) Infection.

    PubMed

    Katsarou, Konstantina; Wu, Yun; Zhang, Runxuan; Bonar, Nicola; Morris, Jenny; Hedley, Pete E; Bryan, Glenn J; Kalantidis, Kriton; Hornyik, Csaba

    2016-01-01

    Potato (Solanum tuberosum L) is a natural host of Potato spindle tuber viroid (PSTVd) which can cause characteristic symptoms on developing plants including stunting phenotype and distortion of leaves and tubers. PSTVd is the type species of the family Pospiviroidae, and can replicate in the nucleus and move systemically throughout the plant. It is not well understood how the viroid can affect host genes for successful invasion and which genes show altered expression levels upon infection. Our primary focus in this study is the identification of genes which can affect tuber formation since viroid infection can strongly influence tuber development and especially tuber shape. In this study, we used a large-scale method to identify differentially expressed genes in potato. We have identified defence, stress and sugar metabolism related genes having altered expression levels upon infection. Additionally, hormone pathway related genes showed significant up- or down-regulation. DWARF1/DIMINUTO, Gibberellin 7-oxidase and BEL5 transcripts were identified and validated showing differential expression in viroid infected tissues. Our study suggests that gibberellin and brassinosteroid pathways have a possible role in tuber development upon PSTVd infection. PMID:26937634

  2. Trichostatin A selectively suppresses the cold-induced transcription of the ZmDREB1 gene in maize.

    PubMed

    Hu, Yong; Zhang, Lu; Zhao, Lin; Li, Jun; He, Shibin; Zhou, Kun; Yang, Fei; Huang, Min; Jiang, Li; Li, Lijia

    2011-01-01

    Post-translational modifications of histone proteins play a crucial role in responding to environmental stresses. Histone deacetylases (HDACs) catalyze the removal of an acetyl group from histones and are generally believed to be a transcriptional repressor. In this paper, we report that cold treatment highly induces the up-regulation of HDACs, leading to global deacetylation of histones H3 and H4. Treatment of maize with the HDAC inhibitor trichostatin A (TSA) under cold stress conditions strongly inhibits induction of the maize cold-responsive genes ZmDREB1 and ZmCOR413. However, up-regulation of the ZmICE1 gene in response to cold stress is less affected. The expression of drought and salt induced genes, ZmDBF1 and rab17, is almost unaffected by TSA treatment. Thus, these observations show that HDACs may selectively activate transcription. The time course of TSA effects on the expression of ZmDREB1 and ZmCOR413 genes indicates that HDACs appear to directly activate the ZmDREB1 gene, which in turn modulates ZmCOR413 expression. After cold treatment, histone hyperacetylation and DNA demethylation occurs in the ICE1 binding region, accompanied by an increase in accessibility to micrococcal nuclease (MNase). The two regions adjacent to the ICE1 binding site remain hypoacetylated and methylated. However, during cold acclimation, TSA treatment increases the acetylation status and accessibility of MNase and decreases DNA methylation at these two regions. However, TSA treatment does not affect histone hyperacetylation and DNA methylation levels at the ICE1 binding regions of the ZmDREB1 gene. Altogether, our findings indicate that HDACs positively regulate the expression of the cold-induced ZmDREB1 gene through histone modification and chromatin conformational changes and that this activation is both gene and site selective. PMID:21811564

  3. Transcriptional and post-transcriptional regulation of HIV-1 gene expression: role of cellular factors for Tat and Rev.

    PubMed

    Nekhai, Sergei; Jeang, Kuan-Teh

    2006-12-01

    The emergence of drug-resistant HIV-1 strains presents a challenge for the design of new therapy. Targeting host cell factors that regulate HIV-1 replication might be one way to overcome the propensity for HIV-1 to mutate in order to develop resistance to antivirals. This article reviews the interplay between viral proteins Tat and Rev and their cellular cofactors in the transcriptional and post-transcriptional regulation of HIV-1 gene expression. HIV-1 Tat regulates viral transcription by recruiting cellular factors to the HIV promoter. Tat interacts with protein kinase complexes Cdk9/cyclin T1 and Cdk2/cyclin E; acetyltransferases p300/CBP, p300/CBP-associated factor and hGCN5; protein phosphatases and other factors. HIV-1 Rev regulates post-transcriptional processing of viral mRNAs. Rev primarily functions to export unspliced and partially spliced viral RNAs fr