Academic literature on the topic 'Transcription Initiation Site'
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Journal articles on the topic "Transcription Initiation Site"
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Means, A. L., and P. J. Farnham. "Transcription initiation from the dihydrofolate reductase promoter is positioned by HIP1 binding at the initiation site." Molecular and Cellular Biology 10, no. 2 (February 1990): 653–61. http://dx.doi.org/10.1128/mcb.10.2.653.
Full textAbstract:
We have identified a sequence element that specifies the position of transcription initiation for the dihydrofolate reductase gene. Unlike the functionally analogous TATA box that directs RNA polymerase II to initiate transcription 30 nucleotides downstream, the positioning element of the dihydrofolate reductase promoter is located directly at the site of transcription initiation. By using DNase I footprint analysis, we have shown that a protein binds to this initiator element. Transcription initiated at the dihydrofolate reductase initiator element when 28 nucleotides were inserted between it and all other upstream sequences, or when it was placed on either side of the DNA helix, suggesting that there is no strict spatial requirement between the initiator and an upstream element. Although neither a single Sp1-binding site nor a single initiator element was sufficient for transcriptional activity, the combination of one Sp1-binding site and the dihydrofolate reductase initiator element cloned into a plasmid vector resulted in transcription starting at the initiator element. We have also shown that the simian virus 40 late major initiation site has striking sequence homology to the dihydrofolate reductase initiation site and that the same, or a similar, protein binds to both sites. Examination of the sequences at other RNA polymerase II initiation sites suggests that we have identified an element that is important in the transcription of other housekeeping genes. We have thus named the protein that binds to the initiator element HIP1 (Housekeeping Initiator Protein 1).
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Means, A. L., and P. J. Farnham. "Transcription initiation from the dihydrofolate reductase promoter is positioned by HIP1 binding at the initiation site." Molecular and Cellular Biology 10, no. 2 (February 1990): 653–61. http://dx.doi.org/10.1128/mcb.10.2.653-661.1990.
Full textAbstract:
We have identified a sequence element that specifies the position of transcription initiation for the dihydrofolate reductase gene. Unlike the functionally analogous TATA box that directs RNA polymerase II to initiate transcription 30 nucleotides downstream, the positioning element of the dihydrofolate reductase promoter is located directly at the site of transcription initiation. By using DNase I footprint analysis, we have shown that a protein binds to this initiator element. Transcription initiated at the dihydrofolate reductase initiator element when 28 nucleotides were inserted between it and all other upstream sequences, or when it was placed on either side of the DNA helix, suggesting that there is no strict spatial requirement between the initiator and an upstream element. Although neither a single Sp1-binding site nor a single initiator element was sufficient for transcriptional activity, the combination of one Sp1-binding site and the dihydrofolate reductase initiator element cloned into a plasmid vector resulted in transcription starting at the initiator element. We have also shown that the simian virus 40 late major initiation site has striking sequence homology to the dihydrofolate reductase initiation site and that the same, or a similar, protein binds to both sites. Examination of the sequences at other RNA polymerase II initiation sites suggests that we have identified an element that is important in the transcription of other housekeeping genes. We have thus named the protein that binds to the initiator element HIP1 (Housekeeping Initiator Protein 1).
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Blake, M. C., R. C. Jambou, A. G. Swick, J. W. Kahn, and J. C. Azizkhan. "Transcriptional initiation is controlled by upstream GC-box interactions in a TATAA-less promoter." Molecular and Cellular Biology 10, no. 12 (December 1990): 6632–41. http://dx.doi.org/10.1128/mcb.10.12.6632.
Full textAbstract:
Numerous genes contain TATAA-less promoters, and the control of transcriptional initiation in this important promoter class is not understood. We have determined that protein-DNA interactions at three of the four proximal GC box sequence elements in one such promoter, that of the hamster dihydrofolate reductase gene, control initiation and relative use of the major and minor start sites. Our results indicate that although the GC boxes are apparently equivalent with respect to factor binding, they are not equivalent with respect to function. At least two properly positioned GC boxes were required for initiation of transcription. Abolishment of DNA-protein interaction by site-specific mutation of the most proximal GC box (box I) resulted in a fivefold decrease in transcription from the major initiation site and a threefold increase in heterogeneous transcripts initiating from the vicinity of the minor start site in vitro and in vivo. Mutations that separately abolished interactions at GC boxes II and III while leaving GC box I intact affected the relative utilization of both the major and minor initiation sites as well as transcriptional efficiency of the promoter template in in vitro transcription and transient expression assays. Interaction at GC box IV when the three proximal boxes were in a wild-type configuration had no effect on transcription of the dihydrofolate reductase gene promoter. Thus, GC box interactions not only are required for efficient transcription but also regulate start site utilization in this TATAA-less promoter.
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Blake, M. C., R. C. Jambou, A. G. Swick, J. W. Kahn, and J. C. Azizkhan. "Transcriptional initiation is controlled by upstream GC-box interactions in a TATAA-less promoter." Molecular and Cellular Biology 10, no. 12 (December 1990): 6632–41. http://dx.doi.org/10.1128/mcb.10.12.6632-6641.1990.
Full textAbstract:
Numerous genes contain TATAA-less promoters, and the control of transcriptional initiation in this important promoter class is not understood. We have determined that protein-DNA interactions at three of the four proximal GC box sequence elements in one such promoter, that of the hamster dihydrofolate reductase gene, control initiation and relative use of the major and minor start sites. Our results indicate that although the GC boxes are apparently equivalent with respect to factor binding, they are not equivalent with respect to function. At least two properly positioned GC boxes were required for initiation of transcription. Abolishment of DNA-protein interaction by site-specific mutation of the most proximal GC box (box I) resulted in a fivefold decrease in transcription from the major initiation site and a threefold increase in heterogeneous transcripts initiating from the vicinity of the minor start site in vitro and in vivo. Mutations that separately abolished interactions at GC boxes II and III while leaving GC box I intact affected the relative utilization of both the major and minor initiation sites as well as transcriptional efficiency of the promoter template in in vitro transcription and transient expression assays. Interaction at GC box IV when the three proximal boxes were in a wild-type configuration had no effect on transcription of the dihydrofolate reductase gene promoter. Thus, GC box interactions not only are required for efficient transcription but also regulate start site utilization in this TATAA-less promoter.
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Uprety, Bhawana, Amala Kaja, Jannatul Ferdoush, Rwik Sen, and Sukesh R. Bhaumik. "Regulation of Antisense Transcription by NuA4 Histone Acetyltransferase and Other Chromatin Regulatory Factors." Molecular and Cellular Biology 36, no. 6 (January 11, 2016): 992–1006. http://dx.doi.org/10.1128/mcb.00808-15.
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NuA4 histone lysine (K) acetyltransferase (KAT) promotes transcriptional initiation of TATA-binding protein (TBP)-associated factor (TAF)-dependent ribosomal protein genes. TAFs have also been recently found to enhance antisense transcription from the 3′ end of theGAL10coding sequence. However, it remains unknown whether, like sense transcription of the ribosomal protein genes, TAF-dependent antisense transcription ofGAL10also requires NuA4 KAT. Here, we show that NuA4 KAT associates with theGAL10antisense transcription initiation site at the 3′ end of the coding sequence. Such association of NuA4 KAT depends on the Reb1p-binding site that recruits Reb1p activator to theGAL10antisense transcription initiation site. Targeted recruitment of NuA4 KAT to theGAL10antisense transcription initiation site promotesGAL10antisense transcription. Like NuA4 KAT, histone H3 K4/36 methyltransferases and histone H2B ubiquitin conjugase facilitateGAL10antisense transcription, while the Swi/Snf and SAGA chromatin remodeling/modification factors are dispensable for antisense, but not sense, transcription ofGAL10. Taken together, our results demonstrate for the first time the roles of NuA4 KAT and other chromatin regulatory factors in controlling antisense transcription, thus illuminating chromatin regulation of antisense transcription.
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Yu, K., and R. T. Elder. "A region internal to the coding sequences is essential for transcription of the yeast Ty-D15 element." Molecular and Cellular Biology 9, no. 9 (September 1989): 3667–78. http://dx.doi.org/10.1128/mcb.9.9.3667.
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The major transcript of the yeast transposable element Ty1 has its 5' end in one delta and the 3' end in the opposite delta, the direct repeats of about 335 base pairs (bp) at each end of the element. The transcriptional initiation signals of the Ty-D15 element that give rise to this transcript were found to have a number of unusual characteristics. The 5' delta by itself, which contained the initiation site for Ty transcription, gave no detectable transcription. A region internal to the transcript in a translated part of the element and about 140 bp downstream of the 5' delta was essential for initiation of the major Ty transcript. This internal activating region (IAR) had several interesting properties. When the portion of the delta upstream of the initiation site was replaced with DNA fragments that did not by themselves act as promoters, initiation directed by the IAR still occurred at about the same position, 200 to 400 bp upstream of the IAR. If fragments containing the IAR were inverted, transcription could still occur. When 468 or 636 bp was inserted between the delta and the IAR, initiations occurred near the normal delta initiation site and in the inserted DNA. Therefore, the location and properties of transcription signals for Ty-D15 differ considerably from those expected for a yeast gene transcribed by RNA polymerase II.
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Yu, K., and R. T. Elder. "A region internal to the coding sequences is essential for transcription of the yeast Ty-D15 element." Molecular and Cellular Biology 9, no. 9 (September 1989): 3667–78. http://dx.doi.org/10.1128/mcb.9.9.3667-3678.1989.
Full textAbstract:
The major transcript of the yeast transposable element Ty1 has its 5' end in one delta and the 3' end in the opposite delta, the direct repeats of about 335 base pairs (bp) at each end of the element. The transcriptional initiation signals of the Ty-D15 element that give rise to this transcript were found to have a number of unusual characteristics. The 5' delta by itself, which contained the initiation site for Ty transcription, gave no detectable transcription. A region internal to the transcript in a translated part of the element and about 140 bp downstream of the 5' delta was essential for initiation of the major Ty transcript. This internal activating region (IAR) had several interesting properties. When the portion of the delta upstream of the initiation site was replaced with DNA fragments that did not by themselves act as promoters, initiation directed by the IAR still occurred at about the same position, 200 to 400 bp upstream of the IAR. If fragments containing the IAR were inverted, transcription could still occur. When 468 or 636 bp was inserted between the delta and the IAR, initiations occurred near the normal delta initiation site and in the inserted DNA. Therefore, the location and properties of transcription signals for Ty-D15 differ considerably from those expected for a yeast gene transcribed by RNA polymerase II.
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Cano, A. "Characterization of the rat NHE3 promoter." American Journal of Physiology-Renal Physiology 271, no. 3 (September 1, 1996): F629—F636. http://dx.doi.org/10.1152/ajprenal.1996.271.3.f629.
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NHE3, a transmembrane protein involved in transcellular ion transport, is expressed in the apical membrane of renal and gastrointestinal epithelia. Chronic regulation by multiple stimuli, including glucocorticoid-induced transcriptional regulation, has been demonstrated. To study the tissue-specific expression and transcriptional regulation of NHE3, the 5' flanking region of the rat NHE3 gene was cloned. Two genomic libraries were screened with the 5' end of the NHE3 cDNA. The 5' flanking region and first exon were isolated. Primer extension mapped a single transcription start site in stomach, colon and kidney. The NHE3 promoter near the transcription initiation site is characterized by the absence of TATA and CAAT sequences. Two Sp1 sites, one Egr-1 site, and an initiator with the sequence GGGATTAAA mark the area of transcription initiation. Upstream sequences include multiple DNA sequence elements recognized by the glucocorticoid and thyroid receptors, Sp1, atriopeptin-2, and several other transcription factors. Transcriptional regulation by glucocorticoids and chronic acidosis was demonstrated. Promoter activity was present in OKP cells, a renal proximal tubule cell line, but not in fibroblasts. This suggests that the NHE3 promoter contains elements conferring epithelial cell-specific expression.
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Pal, S. K., S. S. Zinkel, A. A. Kiessling, and G. M. Cooper. "c-mos expression in mouse oocytes is controlled by initiator-related sequences immediately downstream of the transcription initiation site." Molecular and Cellular Biology 11, no. 10 (October 1991): 5190–96. http://dx.doi.org/10.1128/mcb.11.10.5190.
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We have employed transient expression assays to analyze the sequences that direct c-mos transcription in mouse oocytes. Plasmids containing the chloramphenicol acetyltransferase (CAT) gene fused to either a 2.4-kb or a 731-bp fragment from the 5'-flanking region of c-mos produced similar levels of CAT activity when injected into nuclei of growing oocytes. BAL 31 deletions revealed that sequences up to 20 bp upstream of the major transcription start site could be removed without any significant loss of CAT activity. Promoter activity only decreased when these deletions closely approached the transcription start site, which was mapped at 53 nucleotides upstream of the first ATG in the c-mos open reading frame. On the other hand, deletion of sequences within 20 nucleotides downstream of the transcription initiation site resulted in a 10-fold reduction in CAT expression. A similar decrease in promoter activity was observed as a result of point mutations in these 5' untranslated sequences. Thus, sequences immediately downstream of the transcription start site, including a consensus sequence (PyPyCAPyPyPyPyPy) present in the initiator elements of several genes, appear to regulate c-mos expression in mouse oocytes. Reverse transcription-polymerase chain reaction analysis of RNA from injected oocytes showed that this regulation is manifest at the transcriptional level. Expression of c-mos in mouse oocytes thus appears to be directed by a simple promoter consisting only of sequences immediately surrounding the transcription start site, including an initiator element in the untranslated leader.
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Pal, S. K., S. S. Zinkel, A. A. Kiessling, and G. M. Cooper. "c-mos expression in mouse oocytes is controlled by initiator-related sequences immediately downstream of the transcription initiation site." Molecular and Cellular Biology 11, no. 10 (October 1991): 5190–96. http://dx.doi.org/10.1128/mcb.11.10.5190-5196.1991.
Full textAbstract:
We have employed transient expression assays to analyze the sequences that direct c-mos transcription in mouse oocytes. Plasmids containing the chloramphenicol acetyltransferase (CAT) gene fused to either a 2.4-kb or a 731-bp fragment from the 5'-flanking region of c-mos produced similar levels of CAT activity when injected into nuclei of growing oocytes. BAL 31 deletions revealed that sequences up to 20 bp upstream of the major transcription start site could be removed without any significant loss of CAT activity. Promoter activity only decreased when these deletions closely approached the transcription start site, which was mapped at 53 nucleotides upstream of the first ATG in the c-mos open reading frame. On the other hand, deletion of sequences within 20 nucleotides downstream of the transcription initiation site resulted in a 10-fold reduction in CAT expression. A similar decrease in promoter activity was observed as a result of point mutations in these 5' untranslated sequences. Thus, sequences immediately downstream of the transcription start site, including a consensus sequence (PyPyCAPyPyPyPyPy) present in the initiator elements of several genes, appear to regulate c-mos expression in mouse oocytes. Reverse transcription-polymerase chain reaction analysis of RNA from injected oocytes showed that this regulation is manifest at the transcriptional level. Expression of c-mos in mouse oocytes thus appears to be directed by a simple promoter consisting only of sequences immediately surrounding the transcription start site, including an initiator element in the untranslated leader.
Dissertations / Theses on the topic "Transcription Initiation Site"
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Raborn, R. Taylor. "Genome-wide analysis of transcription initiation and promoter architecture in eukaryotes." Diss., University of Iowa, 2012. https://ir.uiowa.edu/etd/4728.
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The transcriptome represents the entirety of RNA molecules within a cell or tissue at a given time. Recent advances have facilitated the production of large-scale, global interrogations of transcriptomes, finding that genomes are extensively transcribed and contain diverse classes of RNAs (Dinger et al., 2009). Information generated by high-throughput analyses of mRNA transcription start sites (TSSs) such as CAGE (Cap Analysis of Gene Expression) indicate that eukaryotic genomes have complex landscapes of transcription initiation. The TSS is important for the annotation of cis-regulatory sequences, because it provides a link between the mRNA transcript and the promoter. The patterns of TSS distributions observed within mRNA 5' end profiling studies prevent straightforward annotation of putative promoters.
To address this challenge, we developed a method to identify- on a genome-wide basis- the putative promoter, which we define by TSS distributions and designate the transcription start region (TSR). We applied a clustering method to identify and annotate TSRs within the budding yeast Saccharomyces cerevisiae using a full-length cDNA dataset (Miura et al., 2006). To validate these TSR annotations, we performed an integrative genomic analysis using multiple datasets. Our method identified TSRs at positions consistent with bona fide promoters in S. cerevisiae. In addition, using 5'RACE, we find overall agreement between computationally-defined TSRs and TSSs identified experimentally. From this analysis, we find that a significant proportion of genes exhibiting alternative promoter usage within sporulation are associated with respiration, suggesting that this is regulated on a condition-specific basis in budding yeast.
We further developed our TSS clustering method into a bioinformatics tool called TSRchitect, which identifies and annotates TSRs from large-scale TSS profiling information. TSRchitect is capable of handling both tag and sequence-based TSS information and efficiently computes TSRs from global TSS datasets on a desktop computer. We find support for TSRchitect's annotations in human from a CAGE experiment from the ENCODE (Encyclopedia of DNA Elements) project.
Finally, we use TSRchitect to identify TSRs from the transcriptomes of diverse eukaryotes. We investigated the conservation of TSRs among orthologous genes. We frequently identify multiple TSRs for a given gene, suggesting that alternative promoter usage is widespread. Overall, using TSS profiling data derived from separate tissues within mouse and human, we find that the positions of TSRs are relatively stable across tissues surveyed; however, a small fraction of genes exhibit tissue-specific differences in TSR use.
As transcriptome profiling information continues to be generated at an rapid pace, computational approaches are increasingly important. It is anticipated that the method and approach we describe within this dissertation will contribute to an improved of gene regulation and promoter architecture in eukaryotes.
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Palii, Stela S. "Transcriptional regulation of the human system a amino acid transporter, snat2 gene by amino acid availability." [Gainesville, Fla.] : University of Florida, 2004. http://purl.fcla.edu/fcla/etd/UFE0008371.
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Thesis (Ph.D.)--University of Florida, 2004.Typescript. Title from title page of source document. Document formatted into pages; contains 210 pages. Includes Vita. Includes bibliographical references.
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Bharadwaj, Rahul. "Regulation of Higher Order Chromatin at GRIN2B and GAD1 Genetic Loci in Human and Mouse Brain: A Dissertation." eScholarship@UMMS, 2013. https://escholarship.umassmed.edu/gsbs_diss/651.
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Little is known about higher order chromatin structures in the human brain and their function in transcription regulation. We employed chromosome conformation capture (3C) to analyze chromatin architecture within 700 Kb surrounding the transcription start site (TSS) of the NMDA receptor and schizophrenia susceptibility gene, GRIN2B, in human and mouse cerebral cortex. Remarkably, both species showed a higher interaction between the TSS and an intronic sequence, enriched for (KRAB) Krueppel associated Box domain binding sites and selectively targeted by the (H3K9) histone 3 lysine 9 specific methyltransferase ESET/SETDB1. Transgenic mice brain cortical nuclei over-expressing Setdb1 showed increased heterochromatin-protein 1 signal at the interacting regions coupled with decreased Grin2b expression. 3C further revealed three long distant chromatin loop interactions enriched with functional enhancer specific (H3K27Ac) histone 3 lysine 27 acetylation signal in GRIN2B expressing tissue (human cortical nuclei and Human Embryonic Kidney - HEK cells). Doxycycline-induced SETDB1 over-expression decreased 2 out of 3 loop interaction frequencies suggesting a possible SETDB1-mediated transcription repression. We also report a specific looping interaction between a region 50Kb upstream of the (GAD1) Glutamic Acid Decarboxylase – 1 gene TSS and the GAD1 TSS in human brain nuclei. GAD1 catalyzes the rate limiting step in (GABA) gamma amino-butyric acid synthesis and is quintessential for inhibitory signaling in the human brain. Clinical studies in schizophrenia brain samples reveal a decreased looping interaction frequency in correspondence with a decrease in gene expression. Our findings provide evidence for the existence of transcription relevant higher order chromatin structures in human brain.
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Bharadwaj, Rahul. "Regulation of Higher Order Chromatin at GRIN2B and GAD1 Genetic Loci in Human and Mouse Brain: A Dissertation." eScholarship@UMMS, 2002. http://escholarship.umassmed.edu/gsbs_diss/651.
Full textAbstract:
Little is known about higher order chromatin structures in the human brain and their function in transcription regulation. We employed chromosome conformation capture (3C) to analyze chromatin architecture within 700 Kb surrounding the transcription start site (TSS) of the NMDA receptor and schizophrenia susceptibility gene, GRIN2B, in human and mouse cerebral cortex. Remarkably, both species showed a higher interaction between the TSS and an intronic sequence, enriched for (KRAB) Krueppel associated Box domain binding sites and selectively targeted by the (H3K9) histone 3 lysine 9 specific methyltransferase ESET/SETDB1. Transgenic mice brain cortical nuclei over-expressing Setdb1 showed increased heterochromatin-protein 1 signal at the interacting regions coupled with decreased Grin2b expression. 3C further revealed three long distant chromatin loop interactions enriched with functional enhancer specific (H3K27Ac) histone 3 lysine 27 acetylation signal in GRIN2B expressing tissue (human cortical nuclei and Human Embryonic Kidney - HEK cells). Doxycycline-induced SETDB1 over-expression decreased 2 out of 3 loop interaction frequencies suggesting a possible SETDB1-mediated transcription repression. We also report a specific looping interaction between a region 50Kb upstream of the (GAD1) Glutamic Acid Decarboxylase – 1 gene TSS and the GAD1 TSS in human brain nuclei. GAD1 catalyzes the rate limiting step in (GABA) gamma amino-butyric acid synthesis and is quintessential for inhibitory signaling in the human brain. Clinical studies in schizophrenia brain samples reveal a decreased looping interaction frequency in correspondence with a decrease in gene expression. Our findings provide evidence for the existence of transcription relevant higher order chromatin structures in human brain.
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Pauly, Marc. "Etude structurale et fonctionnelle de la sequence tata du promoteur precoce du virus simien sv40." Université Louis Pasteur (Strasbourg) (1971-2008), 1989. http://www.theses.fr/1989STR13043.
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Dans le but d'elucider les mecanismes moleculaires de la regulation de l'expression des genes, une etude structurale et fonctionnelle detaillee de la sequence tata du promoteur precoce du virus sv40 est realisee. La mutagenese dirigee utilisant des oligodesoxynucleotides de synthese permet la localisation des domaines fonctionnels de deux elements tata situes dans la region d'origine de replication virale. Chacun des elements dirige independamment l'initiation precise et efficace de la transcription precoce in vivo a partir d'un groupe de sites definis
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Auchincloss, Andrea Helen. "Transcription initiation sites on the soybean mitochondrial genome." Thesis, McGill University, 1987. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=63914.
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Voronin, Yegor A. "Investigation of initiation of reverse transcription in retroviruses using vectors with two primer-binding sites." Morgantown, W. Va. : [West Virginia University Libraries], 2003. http://etd.wvu.edu/templates/showETD.cfm?recnum=3136.
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Yamaguchi, Shun. "Regional Expression and Regulation of Alternative Forms of mRNAs Derived from Two Distinct Transcription Initiation Sites of the Rat mGluR5 Gene." Kyoto University, 1998. http://hdl.handle.net/2433/182240.
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Myers, Stephen Anthony. "Kallikrein Gene Regulation in Hormone-Dependent Cancer Cell Lines." Queensland University of Technology, 2003. http://eprints.qut.edu.au/15842/.
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Hormone-dependent cancers (HDCs), such as those of the prostate, ovary, breast and endometrium, share characteristics that indicate similar underlying mechanisms of carcinogenesis. Through steroid hormone signalling on "down-stream" target genes, the growth, development and progression of HDCs are regulated. One such family of target genes, highly expressed in HDCs and regulated by steroid hormones, are the tissue kallikreins (KLKs). The KLKs are a multigene family of serine proteases involved in physiological processes such as blood pressure regulation, inflammation, and tumour development and progression via the hydrolysis of specific substrates. Although the KLK gene family is clearly implicated in tumourigenesis, the precise roles played by these genes are largely unknown. Additionally, except for the androgen-responsive genes, KLK2 and KLK3, the mechanisms underlying their hormonal regulation in HDCs are yet to be identified. The initial focus of this thesis was to examine the regulation of the kallikreins, KLK1 and KLK4, by estradiol and progesterone in endometrial and breast cancer cell lines. From these studies, progesterone clearly regulated KLK4 expression in T47D cells and therefore, the focus of the remaining studies was to further examine this regulation at the transcriptional level. An overview of the results obtained is detailed below. Human K1 and hK4 protein levels were increased by 10 nmol/L estradiol benzoate, progesterone, or a combination of the two, over 48 hours in the endometrial cancer cell line, KLE. However, these same treatments resulted in no change in KLK1 gene or hK1 protein levels in the endometrial cancer cell lines, HEC1A or HEC1B (only hK1 analysed). Progesterone treatment (0-100 nmol/L) over 24 hours resulted in a clear increase in KLK4 mRNA at the 10 nmol/L dose in the breast cancer cell line, T47D. Additionally, treatment of T47D cells with 10 nmol/L progesterone over 0-48 hr, resulted in the rapid expression of the hK4 protein at 2 hr which was sustained for 24 hr. Further analysis of this latter progesterone regulation with the antiprogesterone, RU486, over 24 hours, resulted in an observable decrease in hK4 levels at 1 µmol/L RU486. Although the estrogen and progesterone regulation of the hK1 protein was not further analysed, the data obtained for hK4 regulation in T47D cell lines, supported the premise that this gene was progesterone-responsive. The rapid expression of hK4 protein by progesterone at two hours suggests that KLK4 transcription is directly coupled to progesterone regulation, perhaps through progesterone receptor (PR) binding to progesterone-responsive regions within the KLK4 promoter or far "up-stream" regions. Thus, the following further studies were performed. To test this hypothesis, the transcription initiation site (TIS) and 5' flanking regions of the KLK4 gene in T47D cells were interrogated. Primer extension and 5' RACE identified the TIS 78 bp 5' of the putative ATG site for translation as identified by Korkmaz et al. (2001). This KLK4 gene transcript consists of only four exons, and thus excludes the pre/pro signal peptide. Although a TATA-box is not present within -25 to -30 bp 5' of the identified TIS, a number of consensus binding motifs for Sp1 and estrogen receptor half-sites were identified. It is possible that the Sp1 sites are involved in the basal levels of transcription for this gene. Additionally, a putative progesterone response element (PRE) was identified in the far "up-stream" regions of the KLK4 gene. Basal levels of transcription were observed within the KLK4 proximal promoter region when coupled to a luciferase reporter gene and transfected into T47D cell lines. Additionally, the KLK4 proximal promoter region did not induce the luciferase reporter gene expression when progesterone was added to the system, however, estradiol was inhibitory for luciferase gene expression. This suggests that the proximal promoter region of the KLK4 gene could contain functional EREs but not PREs. In keeping with this hypothesis, some ER half-sites were identified, but PR sites were not obvious within this region. The identified PRE in the far "up-stream" region of the KLK4 gene assembled the progesterone receptor in vitro, and in vivo, as assessed by electromobility shift assays and chromatin immunoprecipitation assays (EMSAs and ChIPs), respectively. The binding of the PR to the KLK4 PRE was successfully competed out by a PR antibody and not by an androgen receptor antibody, and thus confirms the specificity of the KLK4 PRE-PR complex. Additionally, the PR was recruited and assembled onto and off the progesterone-responsive KLK4 region in a cyclic fashion. Thus, these data strongly suggest that the PR represents one of the core components of a transcription complex for the KLK4 gene, and presumably also contributes to the expression of this gene. Moreover, these data suggest a functional coordination between the PR and the KLK4 progesterone-responsive region in T47D cells, and thus, provide a model system to further study these events in vivo.
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Sidarovich, Viktoryia. "Transcription of the human plasma prekallikrein gene : demonstration of alternative promoters, multiple initiation sites, and alternative splicing and identification of cis-acting DNA elements." kostenfrei, 2008. http://mediatum2.ub.tum.de/doc/645828/645828.pdf.
Full textBook chapters on the topic "Transcription Initiation Site"
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Zhang, Xuming. "Identification of a Noncanonical Transcription Initiation Site for Transcription of a Subgenomic mRNA of Mouse Hepatitis Virus." In Advances in Experimental Medicine and Biology, 563–70. Boston, MA: Springer US, 2001. http://dx.doi.org/10.1007/978-1-4615-1325-4_83.
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Naryshkin, Nikolai, Sergei Druzhinin, Andrei Revyakin, Younggyu Kim, Vladimir Mekler, and Richard H. Ebright. "Static and Kinetic Site-Specific Protein-DNA Photocrosslinking: Analysis of Bacterial Transcription Initiation Complexes." In Methods in Molecular Biology™, 403–37. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60327-015-1_25.
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Mulligan, R. Michael. "In vitro Capping of Maize Mitochondrial RNA and Transcription Initiation Site Characterization by RNase Protection." In The Maize Handbook, 559–65. New York, NY: Springer New York, 1994. http://dx.doi.org/10.1007/978-1-4612-2694-9_96.
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Forget, Diane, Céline Domecq, and Benoit Coulombe. "Use of Site-Specific Protein-DNA Photocrosslinking of Purified Complexes to Analyze the Topology of the RNA Polymerase II Transcription Initiation Complex." In Methods in Molecular Biology™, 439–51. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60327-015-1_26.
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Steege, D. A., and M. Ivey-Hoyle. "The Phage f1 Gene VII Start Site and its Mutants Reveal that Translational Coupling can Confer Function to Inherently Inactive Initiation Sites." In Post-Transcriptional Control of Gene Expression, 197–206. Berlin, Heidelberg: Springer Berlin Heidelberg, 1990. http://dx.doi.org/10.1007/978-3-642-75139-4_19.
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"Determination of Transcription Initiation Site." In Techniques for Molecular Biology, 119–20. CRC Press, 2006. http://dx.doi.org/10.1201/9781482294460-38.
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Milhé, C. "Determination by 1H NMR of a Slow Conformational Transition and Hydration Change in the Consensus TATAAT Prsbnow Box." In Biological NMR Spectroscopy. Oxford University Press, 1997. http://dx.doi.org/10.1093/oso/9780195094688.003.0027.
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The conformational dynamics and hydration of a DNA 14-mer containing the consensus Pribnow box sequence TATAAT have been measured using rotating frame T1 measurements and NOESY and ROESY in water. The H2 proton resonances of adenines show fast intermediate exchange behavior which can be attributed to a conformational transition that affects the distances between H2 protons of neighboring adenine residues, both sequential and cross-strand. The relaxation rate constant of the transition was measured at 4000s-1 at 25°C. Bound water close to the H2 proton of adenines was observed with residence times of >lns. At low temperature (5°C), the Pribnow box is in a closed state in which hydration water in the minor groove is tightly bound. At higher temperatures, the conformation opens up as judged by the increase in separation between sequential H2 protons of adenines and water exchanges freely from the minor groove. The conformational transition and the altered hydration pattern may be related to promoter function. The control of gene expression in procaryotes depends on the specific recognition by RNA polymerase of a six base-pair sequence (consensus: TTGACA) located at -35 from the transcription site, and a second one, named the Pribnow box (consensus: TATAAT) at about 10 base-pairs upstream the initiation site (Rosenberg and Court, 1979). It has been shown (Hawley and McClure, 1983) that strong promoters exhibit a high degree of homology with the consensus sequences, separated by an optimum consensus spacer length of 17 base pairs. The strength of a promoter depends on, among other thing, the rate of the initiation of transcription. This rate depends on the product between the thermodynamic and kinetic constants KB and k2 (McClure, 1980). The initial binding of RNA polymerase to the promoter results in the formation of a transcriptionally inactive ‘closed’ complex, characterized by the association constant KB. Isomerization to the active ‘open’ complex then occurs, and is characterized by the first order rate constant k2. Hence, the frequency of transcription initiation depends both on the strength of the polymerase-promoter interaction, and the ease with which this complex can isomerize to the productive state. Both of these events are likely to depend on the physical properties of the promoter.
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Forget, Diane, and Benoit Coulombe. "Site-Specific Protein-DNA Photocross-Linking of Purified Complexes: Topology of the RNA Polymerase II Transcription Initiation Complex." In Methods in Enzymology, 701–12. Elsevier, 2003. http://dx.doi.org/10.1016/s0076-6879(03)70057-3.
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Lucchesi, John C. "Epigenetic chromatin changes and the transcription cycle." In Epigenetics, Nuclear Organization & Gene Function, 57–68. Oxford University Press, 2019. http://dx.doi.org/10.1093/oso/9780198831204.003.0005.
Full textAbstract:
In order to allow transcription to occur, the association of DNA with histone octamers and the compacted physical state of the chromatin fiber must be modified by the opportunistic binding of pioneer transcription factors to their cognate DNA binding sites. Once bound, pioneer factors recruit chromatin remodelers and histone-modifying enzymes for the purpose of repositioning nucleosomes and exposing regulatory regions (enhancers and gene promoters) to the components necessary for the initiation of transcription. Histone modifications, such as acetylation, methylation and ubiquitination, and the dynamic phosphorylation of specific amino acids on the major RNA polymerase II subunit activate transcription and attract the factors necessary to eliminate the pausing that normally occurs soon after initiation. Further histone modifications and the replacement of certain core histones by histone variants facilitate transcript elongation and termination. Two additional major epigenetic modifications that impact the process of transcription consist of the action of non-coding RNAs and DNA methylation.
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Lucchesi, John C. "The basic mechanism of gene transcription." In Epigenetics, Nuclear Organization & Gene Function, 17–32. Oxford University Press, 2019. http://dx.doi.org/10.1093/oso/9780198831204.003.0003.
Full textAbstract:
Transcription is initiated by factors that interact with RNA polymerases and recruit them to specific sites, unwind the DNA molecules and allow the synthesis of RNA transcripts complementary to one of the single DNA strands. RNA polymerase II (RNAPII) transcribes genes that encode proteins and some non-coding RNAs; RNAPI transcribes ribosomal RNA genes; RNAPIII transcribes genes that encode tRNAs and other non-coding RNAs. The transcription process starts with a pre-initiation complex (PIC), its activation and promoter clearance. Activation involves chromatin looping, usually promoted by the large multiprotein Mediator complex. RNAPII often makes a promoter-proximal pause, then resumes productive elongation of the transcript. Transition through the different phases of transcription is orchestrated by the phosphorylation of the main subunit of RNAPII. The 5´ end of many transcripts is protected by a methylated guanosine “cap,” and the 3´ end by the addition of a chain of adenosine monophosphates (polyadenylation). Many transcripts undergo splicing to remove regions that interrupt the coding sequence.
Conference papers on the topic "Transcription Initiation Site"
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Collins, Corolyn J., Richard B. Levene, Christina P. Ravera, Marker J. Dombalagian, David M. Livingston, and Dennis C. Lynch. "MOLECULAR CLONING OF THE HUMAN GENE FOR VON WILLEBRAND FACTOR AND IDENTIFICATION OF THE TRANSCRIPTION INITIATION SITE." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642830.
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Most patients with von Willebrand's disease appear to have a defect affecting the level of expression of the von Willebrand factor (vWf) gene. Thus, an understanding of the pathogenesis of von Willebrand's disease will require an analysis of the structure and function of the vWf gene in normals and in patients. To begin such analyses, we have screened a human genomic cosmid library with probes obtained from vWf cDNA and isolated a colinear segment spanning ≈175 kb in five overlapping clones. This segment extends ≈25 kb upstream and ≈5 kb downstream of the transcription start and stop sites for vWf mRNA, implying the vWf gene has a length of ≈150 kb. Within one of these clones, the vWf transcription initiation sites have been mapped. A portion of the promoter region has been sequenced, revealing a typical TATA box, a downstream CCAAT box, and a perfect downstream repeat of the 8 base pairs containing the major transcription start site. Primer extension analysis suggests that sequences contained within the downstream repeat of the transcription start site may be used as minor initiation sites in endothelial cells. Transfection studies are underway to evaluate the role of sequences within this promoter region in gene regulatory activity. Comparative restriction analyses of cloned and chromosomal DNA segments strongly suggests that no major alterations ocurred during cloning and that there is only one complete copy of the vWf gene in the human haploid genome. Similar analyses of DNA from vWf-expressing endothelial cells and non-expressing white blood cells suggests that no major rearrangements are associated with vWf gene expression. Finally, cross hybridization patterns among seven mammalian species suggests a strong conservation of genomic sequences encoding the plasma portion of vWf, but a lower degree of conservation of sequences encoding the N terminal region of provWf.
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Bosma, P. J., E. A. van den Berg, and T. Kooistra. "ISOLATION OF THE GENE CODING FOR HUMAN PLASMINOGEN ACTIVATOR INHIBITOR TYPE 1 (PAI-1)." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644440.
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A human placenta genomic DNA cosmid library was screened for the presence of the PAI-1 gene using a cDNA probe coding for PAI-1. Two overlapping recombinant cosmids were obtained that contain human DNA spanning 55 kb. The cosmids were mapped using 3' and 5' end probes isolated from an almost full-length cDNA clone of 2.5 kb. The two cosmids were found to contain the entire structural PAI-1 gene (approximately 15 kb) and also included 25 kb 5' flanking sequences. The transcription initiation site was identified by SI nuclease protection experiments and the promotor region was sequenced. Further experiments will be directed at characterizing the regulatory elements of the PAI-1 gene.In order to determine the chromosomal localization of the PAI-1 gene we have hybridized our genomic clones in situ to metaphase chromosomes of a human blood cell culture. Preliminary experiments show a specific hybridization signal which will enable us to sublocalize the chromosomal position of the PAI-1 gene.
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Huber, P., J. Dalmon, M. Laurent, G. Courtois, D. Thevenon, and G. Marguerie. "CHARACTERIZATION OFTHE 5’FLANKING REGION FOR THE HUMAN FIBRINOGEN β GENE." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642889.
Full textAbstract:
Fibrinogen is coded by three separate genes located in a 50kb region of chromosome 4 and organized in a α - β - γ orientation with an inversion of the gene 3- A human genomic library was constructed using the EMBL4 phage and screened with cDNA probes coding for human fibrinogen Aα, Bβ and γ chains. Clones, covering the fibrinogen locus,were identified, and their organization was analyzed by means of hybridization and restriction mapping. Among these clones one recombinant phage containing the β gene and large 5’ and 3’ -flanking sequences was isolated.To identify the regulatory sequences Dpstream from the human β gene, a 1.5 kb fragment of the immediate 5’-flanking region was sequenced. The SI mapping experiments revealed three transcription initiation sites. PotentialTATA and CAAT sequences were identified upstream the initiation start points at the positions -21 and -58 from the first initiation start point.Comparison of this sequence with that previously reported for the same region upstream from the human γ gene revealed no significant homology which suggests that the potential promoting sequences of these genes are different. In contrast, comparison of the 5’flanking regions of human and rat β genes showed more than 80% homology for 142 bp upstream from the gene. This highly conserved region is a potential candidate for a regulatory sequence of the human β gene.To verify this activity, a β fibrinogen minigene was constructed by deletion of the internal part of the normal gene and including 3.4kb of the 5’flanking region and 1.4kb of the 3’flanking region. The minigene was transfected into HepG2, a human hepatoma cell line, to show whether the 5’flanking region of the human fibrinogen gene contains DNA sequences sufficient for efficient transcription in HepG2. Constructions of several parts of the sequenced 5’flanking region of the human β gene with the gene of the chloramphenical acetyl transferase have been also transfected in the HepG2 cells to determine the specificity of the gene expression and to localize the sequences controlling the transcription of the gene.