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1
Sigvardsson, Mikael. "Regulation of immunoglobulin transcription during B-cell differentiation." Lund : Lund University, 1995. http://books.google.com/books?id=TJNqAAAAMAAJ.
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Bennett, Selester. "The construction and testing of maize transcriptional fusions in yeast (Saccharomyces cerevisiae)." Thesis, This resource online, 1993. http://scholar.lib.vt.edu/theses/available/etd-10312009-020253/.
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Edelman, Lucas Brandon. "Transcriptional correlates of promoter interactions in murine cell nuclei." Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648695.
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Ezpeleta, Jessica. "The characterization of the ABF-1 promoter." Scholarly Commons, 2001. https://scholarlycommons.pacific.edu/uop_etds/559.
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The basic helix-loop-helix (bHLH) family oftranscription factors consists of proteins involved in cellular proliferation and differentiation. The HLH structure plays a key role in protein-protein dimerization and with the DNA target sites, referred to as E boxes containing the consensus DNA sequence CANNTG. One class of mammalian class I bHLH proteins includes products of the E2A gene, which result from alternative splicing (E12, E47, and ITF), E2-2 and HEB. E2A proteins have also been detected in most cell lines with high levels of expression in lymphoid- and pancreatic cells. It has also been demonstrated that E2A is required for B cell maturation, T cell development and has been shown to function as tumor suppressors. To date, an E2A-interacting bHLH transcription factor largely restricted to activated B lymphocytes, called ABF -1, was isolated using the two-hybrid system. ABF -1 is the only B cell restricted bHLH protein isolated. ABF-1/E2A heterodimers have been detected in B lymphocytes. In these studies, the mapping of the ABF-1 promoter and the critical 5' regulatory elements that control ABF-1 gene expression were analyzed through 5' deletional analysis. 5' -DNA flanking pieces of the promoter region were created through PCR and inserted into a promoterless cloning vector containing the firefly luciferase reporter gene. RT -PCR analysis and anchored PCR was utilized to demonstrate the transcriptional activity of the - promoter region of the ABF-1 gene. Transient transfections were completed to determine critical regulatory sequences. The promoter location was confirmed through computer analysis of the nucleotide sequence and deletional analysis.
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Chan, Tung-lei, and 陳冬妮. "Promoter characterization of testis specific protein, Y-linked like2 (TSPYL2)." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2008. http://hub.hku.hk/bib/B41290410.
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Chan, Tung-lei. "Promoter characterization of testis specific protein, Y-linked like 2 (TSPYL2)." Click to view the E-thesis via HKUTO, 2008. http://sunzi.lib.hku.hk/hkuto/record/B41290410.
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Schoep, Tobias Delavilla. "Isolation and characterization of Pseudobutyrivibrio ruminis gene promoters." Schoep, Tobias Delavilla (2004) Isolation and characterization of Pseudobutyrivibrio ruminis gene promoters. PhD thesis, Murdoch University, 2004. http://researchrepository.murdoch.edu.au/294/.
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A family of E. coli - P. ruminis shuttle-plasmids was constructed to allow the isolation and characterization of gene promoters from the rumen bacterium P. ruminis. The promoter rescue plasmid pBK was used to isolate a total of 4 genomic DNA fragments that promoted transcription in P. ruminis strains 0/10. These promoters, and an additional promoter, previously isolated from P. ruminis strain OR38 (Schoep, 1999), were identified by their ability to initiate expression of a promoterless ermAM gene in P. ruminis. Within 4 of the fragments, a total of 5 transcription start sites were identified in P. ruminis using a novel, fluorescent-primer extension analysis protocol. Comparison of promoters isolated in this and previous studies revealed a strong consensus RNA polymerase DNA-binding motif, including the well characterized -35 and -10 elements. Consensus sequences established for these elements were: TTgacA and AtAATAta respectively, where bold upper-case font, regular upper-case, and lower-case fonts represent conservation in 100%, 80%, and 70% of promoters respectively. The -10 and -35 motifs were interspaced by 16 - 18 nt. Among the newly identified promoters, the consensus for the -10 element was extended one nucleotide upstream and downstream of the standard hexamer (boxed). These motifs were similar to those recognized by eubacterial RNA polymerase containing the alpha -70 like factor. Promoters also contained possible UP elements, and were significantly more curved than protein-coding regions. Additional plasmid vectors were constructed, to allow the use of both the quantitative SYBR green real time PCR and beta-glucuronidase assays, to examine 4 promoters in depth. This showed a wide range of promoter strengths within the group. However, no correlation was found between the composition and context of elements within P. ruminis promoters, and promoter strength. A mutation within the -35 element of one promoter revealed that promoter strength, and the choice of transcription start site were both sensitive to single nucleotide
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Zaugg, Judith Barbara. "A computational study of promoter structure and transcriptional regulation in yeast on a genomic scale." Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609838.
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Lloyd, Amanda Lian. "Cloning, characterisation and sequencing of promoters of Helicobacter pylori 4187E /." Connect to this title, 2004. http://theses.library.uwa.edu.au/adt-WU2005.0112.
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Govender, Cindy. "Stem specific promoters from sorghum and maize for use in sugarcane." Thesis, Stellenbosch : Stellenbosch University, 2008. http://hdl.handle.net/10019.1/2374.
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Thesis (MSc (Genetics. Institute of Plant Biotechnology))--Stellenbosch University, 2008.Sugarcane (Saccharum spp.) is an important crop which is cultivated worldwide for the high sucrose content in its stem. Conventional plant breeding has proven to be very successful over the years with regard to the enhancement of yield characteristics but due to the exhaustion of genetic potential in the commercial sugarcane germplasm recent progress has been slow. Genetic engineering seems to be a more attractive approach to enhance sucrose content and pest resistance in the stems but requires appropriate transgenes and suitable promoter. A promoter is essential to drive the transcription of a gene and is therefore critical to the success of transgenic approaches in sugarcane crop improvement. A negligible number of strong stem-specific promoters is available for use in sugarcane and this is one of the major limitations to genetic engineering. The goal of this project was to isolate a stemspecific promoter from maize and sorghum to drive stem-specific transgene expression in sugarcane. The approach used was to source promoters from non-sugarcane grass species with less complex genomes to simplify isolation and possibly counteract silencing. A cDNA sequence (SS) (EST clone, Accession number AW746904) from sugarcane was shown by Northern and Southern analysis to be stem-specific and to have an appropriately low copy number. The SS gene sequence was not expressed in the leaves of maize, sorghum or the sugarcane cultivars and prominent expression was observed only in the stems of the sugarcane hybrids N19 and 88H0019. The SS gene sequence was used to isolate its upstream regions from a Lambda genomic library of maize (Zea mays) and a sorghum (Sorghum bicolor) Bacterial Artificial Chromosome library (BAC). Of the four sorghum and six maize clones obtained in this study, a 4500 bp maize genomic DNA fragment (λ5) was sub-cloned in three fragments into separate pBluescript vectors using the ‘forced’ cloning approach for sequence and database (BLASTN) analysis. This revealed the complete SS gene sequence (975 bp), the promoter and a 300 bp intron region. A stretch of DNA sequence from nucleotides 664-3194 from the maize clone 5 sequence was designated the maize5-pro. Following sequence alignment of the maize and sugarcane promoter regions, significant sequence identity (68%) was observed between nucleotide 1675 and 3194 in maize and nucleotide 1506 and 2947 in sugarcane. The distance between the putative TATA-box and the TSS for this promoter (30 bp) was found to fall within the expected range of 32± 7 bp. The promoter region was analysed for possible cis-acting regulatory elements and revealed several promoter elements that are common in other plant promoters. The comparisons made between the putative transcription factors in maizepro-5 and the sugarcane promoter show that both promoter sequences are very similar as they share ten of the same transcription factors. However, the transcriptional factors WBOX, SRE and SP8BFIBSP8BIB are unique to the maize5-pro and the TAAG motif to the sugarcane promoter. Primers were designed with appropriate restriction sites and the promoter and intron (2850 bp) region was amplified by PCR (Polymerase chain reaction). The amplified fragment was fused inframe to the GUS reporter gene encoding β-glucuronidase to produce a transformation test vector. This will be used in future work to assess the functionality of the promoter through the production of stable transformants in which GUS activity can be measured in a range of tissues.
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Soininen, Raija. "Structure of the gene for the [alpha] 1 chain of human type IV collagen." Oulu, Finland : University of Oulu, 1989. http://catalog.hathitrust.org/api/volumes/oclc/20482376.html.
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Hare, Alexander. "A mitotic bookmark containing ubiquitin marks the promoters of active genes." The Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1587125230387468.
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Venter, Mauritz. "Isolation of grapevine promoters with special emphasis on the vacuolar pyrophosphatase." Thesis, Stellenbosch : University of Stellenbosch, 2004. http://hdl.handle.net/10019.1/16078.
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Dissertation (PhD)--University of Stellenbosch, 2004.ENGLISH ABSTRACT: Understanding the complex nature of grapevine molecular biology is of great importance for viticulturists. Progress in the elucidation of key events on a genetic level could provide further insight into the underlying cues responsible for the precise control of physiological and metabolic changes during a specific condition such as fruit development. The use and analysis of molecular ‘tools’, such as promoters controlling the site and level of gene activity, could assist in the understanding of grapevine biology and serve as a platform for the future design and development of recombinant DNA protocols and strategies for Vitis vinifera L. A high-throughput gene expression system, cDNA-AFLPs, was successfully used to analyse large-scale transcriptional activity during berry ripening. Candidate cDNA fragments were selected on the basis of desired expression patterns and/or known gene function for subsequent promoter isolation. From three candidate cDNAs selected, the promoter of a gene encoding vacuolar pyrophosphatase (V-PPase) was isolated for computational and comparative analyses. Promoter activity was evaluated on a transient level using the green fluorescent protein (GFP) reporter gene. Comparative integration has allowed for putative correlation of cis-elements, acting as receptors within promoter regions, to regulate V-PPase gene expression in response to development, environmental stress and tissue-specificity. In this study, integration of genetic data have advanced the understanding and transcriptional role of a key enzyme (V-PPase) during grape ripening. Although never a replacement for experimental verification, this integrative strategy of combining gene expression profiles with bioinformatics and regulatory data will greatly assist in further elucidation of various other key components and regulatory cues associated with grapevine molecular biology. This study has allowed us to use molecular tools that could assist in gaining further insight into genetic complexities and could serve as a platform for a more refined genetic manipulation strategy in Vitis vinifera L.
AFRIKAANSE OPSOMMING: Begrip van die komplekse aard van wingerd molekulêre biologie is van groot belang vir wingerdkundiges. Vooruitgang in die begrip van belangrike gebeurtenisse op ń genetiese vlak behoort verdere insig in die onderliggende instruksies vir die noukeurige beheer van fisiologiese en metaboliese veranderinge tydens ń spesifieke kondisie soos vrug rypwording te bevorder. Die gebruik en analise van molekulêre ‘instrumente’ soos promoters, wat die posisie en vlak van geen aktiwiteit beheer, kan bydra tot n beter begrip van wingerd biologie en sodoende dien as ń platform vir die toekomstige ontwerp en ontwikkeling van rekombinante DNS (deoksiribonukleiensuur) protokolle en strategieë vir Vitis vinifera L. ń Hoë-kapasiteit geen uitdrukkings sisteem, nl. kDNS-AFLPs (komplementêre deoksiribonukleiensuur- geamplifiseerde fragment lengte polimorfisme), is suksesvol gebruik vir die analise van grootskaalse transkripsionele aktiwiteit tydens druif rypwording. Kandidaat kDNS fragmente is geselekteer, gebaseer op verlangde uitdrukkings-patrone en/of bekende geen funksie vir daaropvolgende promoter isolering. Van drie geselekteerde kandidaat kDNS fragmente, is die promoter van ń geen wat vakuolêre pirofosfatase (V-PPase) kodeer geïsoleer vir rekenaar- en vergelykende analise. Promoter aktiwiteit is op ń nie-stabiele vlak deur die gebruik van ń groen-fluoresserende proteien (GFP) verklikker geen geëvalueer. Vergelykende integrering het dit moontlik gemaak om veronderstelde korrelasies van cis-elemente, wat as reseptore binne ń promoter area dien, en die regulering van V-PPase geen uitdrukking, in reaksie tot ontwikkeling, omgewings stres en weefsel-spesifisiteit, te maak. Tydens hierdie studie, het die integrering van genetiese data gehelp om die transkripsionele rol van ń belangrike ensiem (V-PPase) tydens druif rypwording beter te verstaan. Alhoewel dit nooit ń plaasvervanger vir eksperimentele bewyse sal wees nie, kan hierdie gëintegreerde strategie, wat die kombinasie van geen-uitdrukkingsprofiele met bioinformatika en regulatoriese data behels, grootliks bydra om verskeie ander belangrike komponente en regulatorieseaanwysings geassosieërd met wingerd molekulêre biologie te ontrafel. Hierdie studie het verdere insig in genetiese kompleksiteite verleen, en kan nou dien as ń platform vir ń meer presiese genetiese manipulering strategie in Vitis vinifera L.
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Jironkin, Aleksey. "Computational and experimental analysis of plant promoters : identifying functional elements." Thesis, University of Warwick, 2013. http://wrap.warwick.ac.uk/60669/.
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Understanding the regulatory DNA sequences are becoming increasingly important in understanding the way plants integrate signalling cues mediated through the actions of the transcription factors (TFs). This thesis presents an interdisciplinary investigations into regulatory elements found in the promoter regions of a model organism Arabidopsis thaliana. The intergenic DNA sequences are studied between sets of orthologous genes in A. thaliana and 3 other related species to uncover hundreds of evolutionary conserved noncoding sequences (CNSs). The CNSs are found to be more skewed towards the annotated transcription start sites (TSSs) and enriched in previously identified transcription factors binding motifs. Furthermore, the nucleosomes are predicted to have strong presence in the uncovered CNS than random intergenic sequences alone. Altogether the evidence presented in the thesis points to the functional nature of the CNSs. Then, the promoters of genes thought to be co-regulated together and transcriptionally active during infection with fungal pathogen Botrytis cinerea are experimentally tested for direct protein-DNA interaction using high-throughput Yeast One-Hybrid (Y1H) library screens against the TFs found in A. thaliana. The resulting predictions were further validated using pairwise Y1H screen to suggest potential common regulation by ORA59, PIF7, ESE1, At4g38900 and ERF14, and uncovering a complex gene regulatory network (GRN) associated with the tested genes. The promoter fragments together with the predictions from the Y1H screens were used in the computational analysis to establish transcription factor specific binding motifs. Some of the newly predicted motifs were mutated and tested again for altered binding of the associated TFs. Furthermore, in planta mutations of the TFs predicted to be interacting with the promoters of the genes in the Y1H screens were found to have significant impact on the susceptibility of A. thaliana to infection with B. cinerea, further informing gene regulatory network active in response to biotic stress.
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Prescott, C. D. "The cloning and analysis of yeast heat shock promoters by gene : : plasmid fusion." Thesis, University of Oxford, 1985. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.370269.
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Hoo, Lai-chong Ruby, and 何麗莊. "Transcriptional regulation of the human glucose-dependent insulinotropic polypeptide gene." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2000. http://hub.hku.hk/bib/B31224428.
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Hoo, Lai-chong Ruby. "Transcriptional regulation of the human glucose-dependent insulinotropic polypeptide gene." Hong Kong : University of Hong Kong, 2000. http://sunzi.lib.hku.hk/hkuto/record.jsp?B22266665.
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Zhao, Wei, and 趙煒. "BRAF mutation and aberrant methylation of gene promoters in the pathogenesis of gastrointestinal tract adenocarcinoma." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B36718464.
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Lloyd, Amanda Lian. "Cloning, characterisation and sequencing of promoters of Helicobacter pylori 4187E." University of Western Australia. Microbiology Discipline Group, 2005. http://theses.library.uwa.edu.au/adt-WU2005.0112.
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Published information on the structure and regulation of H. pylori promoters is limited. The work presented in this thesis describes the cloning and characterisation of promoter regions from a clinical isolate of H. pylori, and the development of an alternative, non-radioactive method for verifying the location of transcriptional start sites of bacterial promoters. H. pylori 4187E promoters were randomly cloned into the promoter-trap vector pKK232-8 in Escherichia coli DH5α using two sets of restriction enzymes. Vector pKK232-8 contains a promoterless chloramphenicol acetyltransferase (CAT) gene. Seventy-four promoter-containing clones were isolated from selective media based on their resistance to chloramphenicol. The strength of each promoter was analysed qualitatively, using chloramphenicol minimum inhibitory concentrations, and quantitatively, using CAT assays following exposure of the clones to pH 4 and pH 7. Selected promoter fragments were subcloned into the GFP reporter vector pFPV25, containing a promoterless gfp gene. The subclones were exposed to buffered LB broth at pH 4, 5, 6, 7 and 8, for varying lengths of time, to study acid-induced regulation of gene expression. Subclones were examined qualitatively, using visual examination of GFP fluorescence and fluorescence microscopy, and quantitatively, using flow cytometry following acid shock. DNA sequences were determined for 61 of the 74 H. pylori promoters, and sequence alignments with the published H. pylori strains (26695 and J99) were performed. The transcriptional start site of 27 H. pylori promoter fragments was experimentally mapped using a fluorescence-based primer extension protocol developed by our group. Potential -35 and -10 sequences were identified for each promoter, and a new consensus sequence for H. pylori promoters was proposed based upon these results. This study has considerably expanded knowledge of H. pylori promoter sequences and transcriptional start sites based on those which also function in E. coli. It has also revealed several H. pylori promoters which appear to respond to acid stress
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Conradie, E. C. (Elizabeth Cornelia). "Promotor engineering in Saccharomyces cerevisiae for transcriptional control under different physiological conditions." Thesis, Stellenbosch : University of Stellenbosch, 2011. http://hdl.handle.net/10019.1/16512.
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Dissertation (PhD)--University of Stellenbosch, 2005.ENGLISH ABSTRACT: To manipulate recombinant microorganisms for industrial processes, controllable genetic systems are needed that can coordinate expression of recombinant metabolic pathways. All components are sensitive to change and thus putative targets for modification and genetic elements and regulatory systems need to be understood and determined. Central in gene regulation is the transcription activators that mediate gene transcription mechanisms by binding to promoters in response to environmental signals. Promoter engineering entails the modification of transcription factors and their target promoters. In this study, a metabolic control system in Saccharomyces cerevisiae was constructed that would allow induction in response to physiological environment, specifically hypoxia and low temperature conditions. Two approaches were undertaken to find such a system. Firstly, a bi-directional reporter gene cloning vector was designed to search for novel hypoxiainducible promoters. Secondly, a transcription regulatory circuit was built, consisting of an inducible transcription regulator and promoter with a reporter gene through which it mediates transcription. Advantage was taken of the modular nature of proteins and functional domains originating from different transcriptional proteins were combined. A search for promoter elements sensitive to hypoxia from a S. cerevisiae genomic DNA (gDNA) library, using a bi-directional cloning vector, did not yield highly inducible promoters. It was concluded that a multitude of signals overlap, rendering genetic induction difficult to control. A synthetic regulatory system would minimize the impact of these multiple interactions. Such a genetic circuit was constructed, consisting of a chimeric transcription activator and a target fusion promoter. The chimeric transcription activator consisted of the GAL4 DNA binding domain, ADR1 TADIII transactivation domain and three domains of the MGA2 regulatory protein. The functional domains of Mga2p responsible for unregulated expression (at high basal levels) under both aerobic and hypoxia conditions were located, as well as a further upregulation under low temperature, and were mapped to the Nterminal and mid-Mga2p regions. A target fusion promoter consisting of a partial GAL10/1 promoter sequence and a Trichoderma reesei core xyn2 promoter were constructed as target for this chimeric transactivator. This synthetic promoter was fused to the T. reesei xyn2 open reading frame encoding for a readily assayable β-xylanase activity. Both the chimeric transactivator and fusion promoter-reporter gene cassettes were expressed from the same episomal plasmid, named pAR. Transformed into S. cerevisiae Y294, this regulatory system induced transcription under aerobic and hypoxia conditions. Furthermore, the reporter gene expression was upregulated by the chimeric transactivator at low temperatures. The chimeric transactivator mediated a seven-fold induction of the reporter gene under aerobic conditions in S. cerevisiae Y294 when transformed with plasmid AR. A two- to three-fold induction at 23ºC was reported under anaerobic conditions, relative to a reference strain expressing a transcription activator without the Mga2p domains. At 30ºC, a two- to three-fold induction under aerobic conditions and similar induction under oxygen-limited conditions were observed. Replacing the reporter gene with your favorite gene (for example a recombinant enzyme) and incorporating such a pAR system into a recombinant yeast should induce expression of the chosen gene under low temperatures, both aerobic and anaerobically (thus creating a controllable system). The system also has wider application in identifying other transcription factors’ signal-sensitive domains. The design of this system provides the ability to add a linker to a transactivator and to either create specific signal sensitivity or relieve the regulator of its signal dependence. It creates an easy system for assessing other transactivators and their domains with unknown functions and thus provides a ”workhorse and prospector in one”.
AFRIKAANSE OPSOMMING: Vir die manipulering van rekombinante mikroörganismes vir industriële prosesse word beheerbare genetiese stelsels benodig om gekoördineerde uitdrukking van rekombinante metaboliese weë teweeg te bring. Alle komponente van sulke stelsels is sensitief vir verandering en genetiese elemente en reguleerbare sisteme moet dus deeglik verstaan of bepaal word. Sentraal tot geenregulering is die transkripsie-aktiveerders wat geentranskripsie beheer deur aan promoters te bind in reaksie op eksterne omgewingsfaktore. Promotoringenieurswese behels wysigings van transkripsiefaktore en hul teikenpromotors. In hierdie studie is 'n genetiese beheerstelsel vir Saccaromyces cerevisiae ontwikkel wat induksie in reaksie tot spesifieke fisiologiese omgewingreaksies, naamlik hipoksie- en lae temperatuur, toelaat. Twee benaderings is gevolg: eerstens is ‘n tweerigting verklikker-geen vektor ontwikkel en gebruik om vir unieke induseerbare hipoksie-promoters te soek. Tweedens is ‘n transkripsie reguleringstelsel gebou wat uit ‘n induseerbare transkripsiereguleerder and promotor met ‘n verklikkergeen bestaan, waardeur transkripsie bemiddel kan word. Hierdie benadering benut die modulêre onderbou van proteïene en funksionele domeine afkomstig vanaf verskillende transkripsiefaktore is gekombineer. 'n Soektog na hipoksie-sensitiewe promotors vanuit 'n Saccharomyces cerevisiae-genoom- DNA (gDNA), deur van ‘n tweerigting verklikker-vektor gebruik te maak, het ongelukkig nie hoogs-induseerbare promotors opgelewer nie. Die gevolgtrekking was dat ‘n veelvoud van seine met mekaar oorvleuel en die beheer van genetiese induksie dus bemoeilik. Die ontwikkeling van ‘n sintetiese regulering-sisteem kan die impak van die veelvuldige interaksies verminder. Vir dié doel is ‘n sintetiese reguleringstelsel ontwerp, bestaande uit ‘n chimeriese transkripsie-aktiveerder met ‘n teiken fusie-promotor. Die chimeriese transaktiveerder bestaan uit die GAL4 DNA bindingsdomein, die ADR1 TAD III transaktiveringsdomein en drie domeine van die Mga2 reguleringsproteïen. In die studie is die funksionele domeins van Mga2p betrokke by lae temperatuur-respons en ongereguleerde uitdrukking (teen hoë basale vlakke) onder beide aërobiese en anaërobiese toestande aangedui en is tot die N-terminaal en middel-Mga2p areas gekarteer. ‘n Teiken-fusie-promoter, bestaande uit 'n gedeeltelike GAL1/10 DNA promotoropeenvolging en ‘n Trichoderma reesei kern xyn2-promoter, is as teiken vir hierdie chimeriese transaktiveerder saamgestel. Hierdie sintetiese promotor is aan die T. reesei xyn2 oopleesraam, wat vir ‘n maklik meetbare β-xylanase aktiwiteit kodeer, gekoppel. Beide die chimeriese transaktiveerder and fusie-promoter-verklikker-geenkaset word vanaf dieselfde episomale plasmied, bekend as pAR, uitgedruk. Hierdie reguleringsisteem induseer transkripsie onder aërobiese en hipoksie toestande in S. cerevisiae Y294. Verder word die verklikkergeen se uitdrukking deur die chimeriese transaktiveerder by lae temperature verhoog. Die chimeriese transaktiveerder induseer ‘n sewe-voudige induksie van die verklikkergeen onder aërobiese toestande by 23ºC vanaf die pAR-stelsel in S. cerevisiae Y294. ‘n Twee- tot drie-voudige induksie teen 23ºC is onder hipoksie toestande gevind, relatief tot induksievlakke van ‘n verwysingstam met ‘n transaktiveerder sonder die Mga2 domeine. By 30ºC is ‘n twee- tot drie-voudige induksie onder aërobiese en lae suurstofvlakke waargeneem. Deur die verklikker geen met ‘n jou-gunsteling-geen te vervang (bv. ‘n rekombinante ensiem) en so 'n pAR-sisteem in ‘n rekombinante gis te inkorporeer, word uitdrukking onder lae temperature onder beide aërobiese- en anaërobiese toestande geïnduseer (en sodoende word ‘n reguleerbare sisteem geskep). Die sisteem het wyer toepassing om sein-sensitiewe domeine van ander transkripsiefaktore te identifiseer. Die ontwerp van die stelsel maak dit moontlik om 'n skakel tot die transaktiveerder by te voeg wat óf sensitiwiteit tot 'n spesifieke sein skep, óf die reguleerder vanaf seinafhanklikheid verlos. So word ‘n bruikbare stelsel vir die bestudering van ander transaktivators en hul domeine met onbekende funksie geskep – ‘n “werksesel en prospekteerder in een”.
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Dukes, Ruth Lynn. "Comparative analysis of the promoters of the CyI-CyIIa-CyIIb actin gene cluster in the sea urchin Strongylocentrotus purpuratus." Morgantown, W. Va. : [West Virginia University Libraries], 1999. http://etd.wvu.edu/templates/showETD.cfm?recnum=709.
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Thesis (M.S.)--West Virginia University, 1999.Title from document title page. Document formatted into pages; contains iv, 64 p. : ill. Vita. Includes abstract. Includes bibliographical references (p. 61-64).
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Angle, Jordan C. "Analysis of transcription factor promoter binding in rat hepatoma/fibroblast hybrids /." View online, 2009. http://repository.eiu.edu/theses/docs/32211131559262.pdf.
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Tsang, Shirley Xiaoman. "TATA-dependent repression of human immunodeficiency virus Type-1 transcription by the Adenovirus E1A 243R oncoprotein." Diss., Georgia Institute of Technology, 1996. http://hdl.handle.net/1853/25325.
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Ririe, Seth S. "Structure and function of the polypyrimidine region of the rat [alpha]1 (I) procollagen gene promoter." free to MU campus, to others for purchase, 2000. http://wwwlib.umi.com/cr/mo/fullcit?p9998517.
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Lavrrar, Jennifer L. "The role of regulatory proteins at the FEPDGC-ENTS promoter region in escherichia coli : a new model for the fur-DNA interaction /." free to MU campus, others may purchase free online, 2002. http://wwwlib.umi.com/cr/mo/preview?3074419.
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Wang, Ming-Bo. "Isolation of phloem specific gene promoters for use in genetic engineering of insect resistance in rice." Thesis, Durham University, 1994. http://etheses.dur.ac.uk/5146/.
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Towards the aim of producing transgenic rice with enhanced resistance to one of its phloem sap-sucking insect pests, the brown planthopper (BPH), two potential phloem-specific promoters, of the rice sucrose synthase-1 (RSsl) and the cucurbit phloem protein PP2 genes, were isolated and investigated. Using a PGR fragment of the maize sucrose synthase-1 (Shi) gene, genomic clones containing the RSsl and RSs2 (rice sucrose synthase-2) genes were isolated from a genomic library of rice (Oryza sativa L. Japonica) and characterised. A full- length RSsl gene from one of the genomic clones was sequenced, including 1756 bp of 5'-flanking sequence and 710 bp of 3'-flanking sequence. The gene had an identical intron-exon structure (16 exons and 15 introns) to the maize Shi gene. The RSsl 5'- flankmg region contained a number of promoter-like sequences, including putative exacting elements homologous to those found in several endosperm-specific, anaerobiosis-inducible, or phloem-specific promoters. The RSsl promoter region, including 1.9 kb 5'-flanking sequence, the first intron, the first exon, and the translational start codon, was fused with coding sequences for β-glucuronidase (GUS) and snowdrop (Galanthus navalis) lectin (GNA). Tobacco plants were transformed with these chimaeric genes in order to determine the expression pattern directed by the RSsl promoter. Histochemical and immunochemical assays demonstrated that the expression of both GUS and GNA was restricted to phloem cells in various tissues (i.e. stem, root, leaf, petiole) and in different transformants. In addition, GNA was detected by immunological assay in the honeydew excreted by peach potato aphids (Myzus persicae), also a phloem sap- sucking insect, feeding on RSsl-GNA transgenic tobacco plants. This provided direct evidence that GNA was not only expressed in the phloem tissue, but was also present in the phloem sap of transgenic tobacco plants. Since GNA has been shown to have antimetabolic effect against BPH, the RSsl-GNA construct is being used to transform rice plants by collaborating groups elsewhere. In order to isolate the phloem protein PP2 gene promoter, the PP2 polypeptide from 3 months old Cucurbita pepo plants was partially sequenced after in situ cleavage with cyanogen bromide vapour, giving 75 residues of sequence on two cyanogen . bromide fragments. Using an oligonucleotide probe based on this amino acid sequence, cDNA clones encoding PP2 were isolated from a C. pepo cDNA library constructed from mRNA of 3-5 days old seedling hypocotyls. A cDNA clone was used as probe to screen a C. pepo genomic library, and several genomic clones were isolated. Restriction mapping showed that these clones contained different genes, consistent with results from Southern blots of C. pepo genomic DNA probed with PP2 cDNA, in which multiple bands were detected in all restriction endonuclease digests. One of the clones was partially sequenced, and was shown to contain a gene encoding PP2. A 1.2 kb 5'-flanking region of this clone was fused with a GUS reporter gene, and this construct was used to transform tobacco. Initial histochemical analysis showed that this chimaeric gene was not expressed in the putative transgenic plants examined. Possible reasons for this failure are discussed.
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Woo, Andrew Jonghan. "Characterization and identification of transcription factors that bind to the tumor necrosis factor -308 polymorphism." University of Western Australia. School of Biomedical and Chemical Sciences, 2003. http://theses.library.uwa.edu.au/adt-WU2004.0044.
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[Formulae and special characters can only be approximated. Please see the pdf version of this abstract for an accurate reproduction.] Tumor necrosis factor (TNF) is a pleiotropic cytokine that mediates a long list of immunological and pathophysiological processes. TNF is produced by a wide variety of cells including immune and non-immune cells, however in most cell types TNF is not expressed prior to stimulation. The function of TNF is mediated via its trimeric domain by binding to TNF receptors that are found on most types of cells, especially of the haematopoietic systems, hence transpiring its effects on a wide variety of cells and organ systems. The cytotoxic (apoptosis) and pro-inflammatory (differentiation, proliferation and activation) functions of TNF are protective but can also result in pathological or deleterious consequences. A biallelic G to A transition polymorphism in the promoter region of TNF at nucleotide position 308 from the transcription start site is suggested to be involved in differential transcriptional regulation of TNF expression. The high TNF producing 308A allele is associated with susceptibility to or worse outcome of many infectious diseases in addition to autoimmune and other pathophysiological conditions. A previous study in our laboratory observed a selective affinity towards the polymorphic 308A allele by an EMSA protein(s) complex, named E. Several other protein complexes were found along with complex E and one of them was identified as Sp1. The identification of complex E was unsuccessful but it was hypothesized to play a major role as transcriptional activator in 308A allele individuals hence transpiring its effect in various pathophysiological states. In this study, the EMSA complexes observed in the TNF promoter region between nucleotides 322 to 283, encompassing the 308 polymorphism, is characterized. EMSA using mutated oligonucleotides mapped the binding sites of complexes B, C, D and E. TRANSFAC database search in addition to previous work revealed the identity of complex C as Sp1 but the rest of complexes remained unknown. Moreover, in contrast to our previous study, the protein(s) in the complex E was found to preferentially bind 308G nucleotide hence posing as a transcriptional repressor, resulting in decreased production state of TNF in 308G allele individuals than 308A allele individuals. In order to characterize putative transcription factors binding to the promoter region, first the biochemical characteristics such as the effects of temperature, salts and cations on DNA binding ability of EMSA complexes were studied. EMSA complexes B, C, DI and E required cations, probably Zn+2, to bind DNA. By optimizing a technique that couples EMSA with SDS-PAGE, the molecular weight of C, DI and E was determined. A novel technique that couples EMSA with IEF determined the pI of complexes B, C, D, DI and E. Although a commonly used technique of identifying unknown DNA-binding protein of interest, Yeast One-Hybrid assay, did not identify complex E, the novel identification method involving chromatography, two-dimensional electrophoresis, EMSA, mass spectrometry and database interrogation successfully identified TNF EMSA complex E as transcription factor Ying Yang 1 (YY1). Supershift EMSA confirmed complex E as YY1. In addition, the supershift assay showed presence of Sp1 and Sp3 in complex C. Similarly, complex DI is identified as Sp3. The novel method in identifying DNA-binding proteins is particularly useful as this technique allows identification of protein seen in EMSA without the need of extensive identification process. YY1 binds to a 6 base pair sequence, 5? TTGAGG 3?, from nt 295 to 290 of TNF promoter. The loss of affinity in 308A allele is caused by transition of underlined G nucleotide to A. The determined and described molecular weight of YY1 in literature is 60 kDa while the theoretical weight is 45 kDa. Both the determined and theoretical pI of YY1 is 5.8. YY1 is a multifunctional transcription factor implicated in both positive and negative regulation of gene expression as well as in initiation of transcription. It is ubiquitously expressed in growing, differentiated, and growth-arrested cells. Although future experiment is yet to establish in vivo presence of YY1 in TNF promoter, our study so far provides convincing evidence that the putative transcription factor that has selective affinity towards 308G allele is indeed YY1.
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Stower, Hannah Mary. "Dynamic histone modifications at the promoters of Hox genes in embryonic stem cells." Thesis, University of Birmingham, 2009. http://etheses.bham.ac.uk//id/eprint/483/.
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Histone modifications have been closely associated with changing levels of gene expression, but their role in determining, or possibly predicting, patterns of expression is uncertain. Here, the link between histone modifications and Hoxb gene expression in mouse embryonic stem (ES) cells was explored. Levels of the “active” modifications H3K9ac and H3K4me3 at Hoxb promoters varied widely from gene to gene, but were closely correlated in ES cells. Contrastingly, the repressive modification H3K27me3 was found at equivalent levels across the cluster. Treatment with the histone deacetylase inhibitor valproate induced a coordinate increase in the levels of H3K9ac and H3K4me3 at all Hoxb promoters, but not other genes, whilst H3K27me3 was unaffected. Such increases were not maintained upon removal of the inhibitor. All Hoxb genes were silent in undifferentiated ES cells, but expression was activated at defined times of differentiation in the expected 3’ to 5’ sequence. The valproate induced increase in active modifications did not induce Hoxb expression from the cluster in undifferentiated cells, nor was there any major shift in the timing of Hoxb expression in cells transiently exposed to valproate (ie. hyperacetylated) during the start of differentiation. Thus, active histone modifications at the Hox genes are uncoupled from transcription.
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Lorson, Christian. "An analysis of transcriptional regulation of the MVM capsid gene promoter." free to MU campus, to others for purchase, 1997. http://wwwlib.umi.com/cr/mo/fullcit?p9841319.
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Oppon, Ekow CruickShank. "Synergistic use of promoter prediction algorithms: a choice of small training dataset?" Thesis, University of the Western Cape, 2000. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_8222_1185436339.
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Promoter detection, especially in prokaryotes, has always been an uphill task and may remain so, because of the many varieties of sigma factors employed by various organisms in transcription. The situation is made more complex by the fact, that any seemingly unimportant sequence segment may be turned into a promoter sequence by an activator or repressor (if the actual promoter sequence is made unavailable). Nevertheless, a computational approach to promoter detection has to be performed due to number of reasons. The obvious that comes to mind is the long and tedious process involved in elucidating promoters in the &lsquo
wet&rsquo
laboratories not to mention the financial aspect of such endeavors. Promoter detection/prediction of an organism with few characterized promoters (M.tuberculosis) as envisaged at the beginning of this work was never going to be easy. Even for the few known Mycobacterial promoters, most of the respective sigma factors associated with their transcription were not known. If the information (promoter-sigma) were available, the research would have been focused on categorizing the promoters according to sigma factors and training the methods on the respective categories. That is assuming that, there would be enough training data for the respective categories. Most promoter detection/prediction studies have been carried out on E.coli because of the availability of a number of experimentally characterized promoters (+- 310). Even then, no researcher to date has extended the research to the entire E.coli genome.
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Zhao, Wei. "BRAF mutation and aberrant methylation of gene promoters in the pathogenesis of gastrointestinal tract adenocarcinoma /." View the Table of Contents & Abstract, 2006. http://sunzi.lib.hku.hk/hkuto/record/B35880867.
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Penniket, Carolyn Renee. "Tissue-specific gene expression and promoter characterization in triticale." Thesis, Lethbridge, Alta. : University of Lethbridge, Dept. of Biological Sciences, c2013, 2013. http://hdl.handle.net/10133/3373.
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Triticale (x Triticosecale Whitm.) is a cereal with favorable agronomic traits for a Canadian bioproduction platform crop. Appropriate tissue sampling times were determined and gene expression profiles were evaluated in five triticale seed tissues and eleven vegetative tissues using the Affymetrix Wheat GeneChip®. Genes that were expressed, not expressed, tissue-specific, tissue-enriched and developmentally regulated were identified. The percentage of probe sets on the wheat GeneChip with gene ontology annotations was improved from less than 3% to over 76% using homologous sequence identification and annotation transfer. This information was used to determine functions and processes over-represented within the identified gene lists and provide biological meaning to the results. Expression of candidate genes was further evaluated using qRT-PCR, RNA in situ hybridization and promoter characterization. This study has provided a comprehensive triticale gene expression atlas; knowledge regarding triticale development, gene function, expression and regulation; and tools enabling further triticale research and development.xxiii, 425 leaves : col. ill. ; 29 cm
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Hagen, Darren Erich. "Identification and characterization of germline-specific promoters for remobilization of transgenes in the mosquitoes, Aedes aegypti and Anopheles gambiae." [College Station, Tex. : Texas A&M University, 2007. http://hdl.handle.net/1969.1/ETD-TAMU-1513.
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Morris, Terry Lynn. "Molecular characterization of the fepA-fes bidirectional promoter in escherichia coli." free to MU campus, to others for purchase, 2001. http://wwwlib.umi.com/cr/mo/fullcit?p3025640.
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Curach, Natalie Claire. "An investigation into the hex1 gene and gene promoter for the enhancement of protein production in Trichoderma reesei." Phd thesis, Australia : Macquarie University, 2005. http://hdl.handle.net/1959.14/71199.
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Supplementary material to figures contained on DVD only available with manuscript.Thesis (PhD)--Macquarie University, Division of Environmental & Life Sciences, Dept. of Biological Sciences, 2005.
Bibliography: p. 221-244.
Introduction -- Materials and methods -- Isolation of the hex1 gene from Trichoderma reesei and Ophiostoma floccosum -- Expression of DsRed under the cbh1 promoter and the hex1 promoter with random integration -- Modified expression vectors containing a fusion to a portion of hex1 gene sequence -- Expression of DsRed from the hex1 locus and the phenotypic characteristics of a hex1 deletion mutant -- Summary and concluding discussion.
For Trichoderma reesei to be developed as an effiecient producer of a large variety of proteins, the expression system requires diversification. In particular, the choice of promoters available needs to be broadened to include promoters which are active in conditions other than those conducive to induction of cellulase expression. Using proteomics, the HEX1 protein was identified as an abundant protein of the cell envelope of T. reesei when grown on a range of carbon sources, suggesting that a strong constitutive promoter drives the expression of this physiologically important protein. This thesis is an exploration into the hex1 gene promoter and the role of hex1 in the maintenance of mycelium integrity in T. reesei with consideration for the application of this gene in the further development of filamentous fungi as protein expression systems. -- The single copy hex1 gene and flanking regions were isolated from T. reesei and another biotechnologically important fungus, Ophiostoma floccosum. The fluorescent reporter protein DsRed1-E5 was expressed under the T. reesei hex1 promoter and promoter activity was monitored by fluorescence CLSM and RNA analysis. During the rapid growth phase of a culture, the hex1 promoter was active in a range of carbon sources and three transcipt types with alternative tsp and splicing sites were discovered for the hex1 gene. The distribution of fluorescence throughout the mycelium suggested spatial regulation of the hex1 promoter as well as temporal regulation. The promoter was continually active in the absence of a functional hex1 gene product suggesting that the hex1 promoter is regulated in part, by negative feedback from the endogenous gene product. Interruption of the hex1 gene produced hyphae that leaked excessive volumes of cytoplasm when physically damaged which may be advantageous for the externalisation of selected protein products. The results indicate that the regulation of the hex1 hene promoter is complex and that the hex1 gene is integral to the maintenance of the integrity of the fungal mycelium.
Mode of access: World Wide Web.
xv, 244 p. ill
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Hartt, Gregory Thomas. "Regulation of the human neuronal nitric oxide synthase gene via alternate promoters." Columbus, OH : Ohio State University, 2003. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1056034844.
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Thesis (Ph. D.)--Ohio State University, 2003.Title from first page of PDF file. Document formatted into pages; contains xii, 152 p. : ill., (some col.). Includes abstract and vita. Advisor: Anthony Young, Molecular, Cellular, and Developmental Biology Program. Includes bibliographical references (p. 137-150).
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Grade, Carla Vermeulen Carvalho 1983. "Caracterização funcional do promotor gênico da miostatina." [s.n.], 2013. http://repositorio.unicamp.br/jspui/handle/REPOSIP/317671.
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Orientador: Lucia Elvira AlvaresTese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A Miostatina é um regulador negativo da deposição de musculatura esquelética e mutações no gene que codifica esta proteína têm sido associadas a um aumento marcante na massa muscular de organismos vertebrados, resultado de hiperplasia e hipertrofia das fibras musculares. Nosso grupo identificou previamente o promotor basal do gene da Miostatina e análises de bioinformática revelaram a presença de sítios evolutivamente conservados para a ligação de CREB, Meis, FXR e NFY, além de um sítio TATA. No presente trabalho nós utilizamos mutagênese sítio-dirigida para gerar diversas construções delecionais que possuem um ou mais sítios mutados, e testamos sua atividade in vitro usando mioblastos C2C12 de camundongo sob condições de proliferação e diferenciação, para analisar o papel destes sítios de ligação sobre a atividade do promotor. Os resultados mostraram que FXR aparentemente não confere efeito na atividade transcricional do promotor da Miostatina em ambos os momentos analisados, indicando que o papel regulador desta proteína pode estar relacionado ao controle da expressão da Miostatina em outro tipo celular, que não o mioblasto. O NFY apresentou um papel de ativador transcricional, enquanto CREB e Meis atuaram inicialmente como repressores durante a proliferação, passando a relaxar esta repressão durante a diferenciação dos mioblastos, permitindo que a atividade do promotor aumentasse significativamente. Trabalhando juntos, estes fatores de transcrição são capazes de manter a atividade do promotor em níveis mais baixos durante a proliferação dos mioblastos e, com o início da diferenciação, a repressão é liberada, e os níveis de atividade podem aumentar. Este padrão está de acordo com o padrão de expressão dinâmico observado para a proteína da Miostatina durante o desenvolvimento da musculatura esquelética em vertebrados
Abstract: Myostatin is a negative regulator of skeletal muscle deposition and mutations in the gene that encodes this protein have been associated to a remarkable increase in skeletal muscle mass, attributable to both hyperplasia and hypertrophy. We have previously identified Myostatin's basal promoter and bioinformatic analyses revealed the presence of evolutionarily conserved binding sites for CREB, Meis, FXR and NFY, besides a TATA box. In the present study we used site-directed mutagenesis to generate several expression constructs possessing one or more mutated sites, and tested their activity in vitro using mouse C2C12 myoblasts in proliferation and differentiation conditions, to analyze the role of these sites on the activity of the promoter. The results show that FXR appears not to confer any effect on the transcriptional activity of the promoter in both conditions, indicating that the regulatory role of this protein might be involved in the control of Myostatin expression in another cell type. NFY presents a role as transcriptional activator, while CREB and Meis act initially as repressors during proliferation, releasing this repression upon differentiation, which allows the activity of the promoter to significantly increase. Working together, these transcription factors are capable of maintaining the promoter activity at lower levels during the proliferation of myoblasts and, upon differentiation, the repression is released, and activity levels can be increased. This pattern is in agreement with the dynamic expression pattern observed for Myostatin during the skeletal muscle development in vertebrates
Doutorado
Histologia
Doutor em Biologia Celular e Estrutural
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Warshamana, Gnana Sakuntala. "Interactions of T7 RNA polymerase with its promoters : Part I: T7 promoter contacts essential for promoter activity in vivo ; Part II: Isolation and characterization of a mutant T7 RNA polymerase with altered promoter specificity." Diss., Georgia Institute of Technology, 1992. http://hdl.handle.net/1853/26303.
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Silvert, Martin. "Origines et conséquences des variants régulateurs chez l'humain." Electronic Thesis or Diss., Sorbonne université, 2019. http://www.theses.fr/2019SORUS359.
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L'expression des gènes est le résultat de nombreuses interactions. De la régulation de la transcription par les promoters et les enhancers à de nombreuses formes de régulations post-transcriptionelles, chaque région régulatrice étant soumise à des pressions sélectives différentes. Dans ce contexte, l’étude de l’évolution des différentes régions régulatrices au sein des populations humaines, ainsi que l’évaluation de leurs contributions respectives à la variabilité de l’expression génique sont essentielles à la compréhension de la variabilité phénotypique humaine. Ce manuscrit se propose donc d’étudier la contribution de la variabilité génétique à la régulation de l’expression génique sous deux angles différents. Je me suis penché sur les conséquences de l'introgression néandertalienne sur la diversité au sein des régions régulatrices dans les populations eurasiennes. Pour cela, j’ai déterminé non seulement quelles sont les régions régulatrices dont la diversité provient de l'introgression néandertalienne de manière disproportionnée, mais j’ai également d'identifié si la source du probable évènement de sélection associé. Je me suis aussi intéressé plus précisément au fonctionnement d'un type de régulation particulier, la régulation par les miARN. En utilisant les résultats de séquençage des petits ARN dans les monocytes, immuno-stimulés ou non, de 100 individus d'ascendance européenne et 100 individus d'ascendance africaine, j'ai pu étudier à la fois la diversité de l'expression des miARN au sein de ces individus, mais également comment ceux-ci participent à la régulation de l'expression des gènes dans un contexte immunitaireGene expression is the result of numerous interactions, from transcription regulation by promoters and enhancers, to several forms of post-transcriptionnal regulation. But each regulatory region is under different selective constraint. In this context, the study of the evolution of regulatory regions in human populations and their relative contribution to the variability of gene expression are vital to the understanding of the human phenotypic diversity. This thesis studies the contribution of the genetic diversity to gene regulation following to different path of investigations. I investigated the consequences of the Neanderthal introgression on diversity of regulatory regions in Eurasians populations. I identified which are the regulatory regions that which diversity is unexpectedly influenced by Neanderthal introgressed mutations, and also identified the sources of the associated selection event. I also focused on the specific case of the regulation by miRNA. Using small RNA sequencing in monocytes, either resting or immune-stimulated, of 100 individual of European ancestry and 100 individuals of African ancestry. I studied the diversity of miRNA expression within this cohort, but also how miRNA participate to the regulation of genes in an immune context
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Zhang, Xiao-Qun. "Functional Studies on the PDGFR α gene promoter and effects of autocrine PDGF-A stimulation in vivo." Doctoral thesis, Uppsala universitet, Institutionen för genetik och patologi, 2001. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-1455.
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Platelet-derived growth factor receptor α (PDGFRα) plays an important role during embryogenesis. After implantation, the patterns of expression of Pdgfrα and its ligand Pdgf-A undergo an "autocrine-paracrine transition", in that Pdgf-A becomes expressed in the ectoderm and epithelia, while Pdgfrα is expressed in the adjacent mesenchymal tissue. In human tumors, such as malignant glioma, both PDGF and PDGFRα are overexpressed within the same tissue, indicating that an autocrine PDGF loop is generated in the tumors. This thesis is focused on the in vivo functionality of the PDGFRα gene (PDGFRA) promoter, arid on the effect of autocrine PDGF-A stimulation in transgenic n-iice during embryogenesis. To test the in vivo promoter function of a human PDGFRA 2.2 kb 5' flanking fragment, we generated transgenic mouse lines and found that the 2.2 kb fragment was able to promote lacZ reporter gene expression in most of the endogenous Pdgfra expressing tissues. Absence of expression and "ectopic" expression of the transgenic lacZ were also observed. To investigate the autocrine PDGF effect, we produced autocrine PDGF-As (A short-chain) transient transgenic embryos. These transgenic embryos carried a 6 kb mouse Pdgfra 5' flanking sequence linked to a human PDGF-As cDNA. The pattern of expression of the PDGF-As transgene mRNA was similar to that of lacZ. Some of the transgenic embryos exhibited severe abnormal phenotypes, such as midline fusion defects in the cephalic and craniofacial region and small body size, and these embryos die at mid-gestation stage. These findings indicate that a paracrine pattern of expression and the dosage of PDGF are important for sustaining normal embryo development, especially with regard to the middline fusion in craniofacial regions. The possible signaling pathways that may be involved in regulating Pdgfra activity were also studied by comparison of patterns of mRNA expression of Gli, Ptc, and Paxl with that of Pdgfra. The results pointed to the possibility that the Shh signaling pathway may be involved in the regulation of Pdgfra expression for example during early bone and foregut development. The specific regulatory mechanisms may vary for different tissues.
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Santos, Railson Schreinert dos. "Regulação transcricional e epigenética de ERFs em arroz sob estresse abiótico." Universidade Federal de Pelotas, 2012. http://guaiaca.ufpel.edu.br/handle/123456789/1207.
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Previous issue date: 2012-07-27Rice (Oryza sativa L.) is one of the most important cereals in the world being cultivated in lowland ecosystems, which have a common characteristic: lack of drainage conditions. The lack of oxygen, due to submergence of seedlings, and iron toxicity are two of the major abiotic stresses which rice plants are subjected. Recently it was realized the importance of some ethylene responsive transcription factors (ERFs) in plant response to different stresses, especially to oxygen depletion. Considering it, two studies were made in order to evaluate the expression of ERFs under different stresses. In a first study seven genes were selected, based on previous results obtained from Genevestigator analysis, and had their transcriptional expression evaluated trough quantitative PCR (qPCR) in rice seedlings under oxygen depletion and iron overload. In the second study thirteen ERFs, which had no information available in the literature, also had their transcriptional levels evaluated in seedlings under conditions of hypoxia and anoxia through qPCR in a cultivar susceptible to flooding (Bonança) and a tolerant one (Nipponbare). As general conclusions it was once more emphasized the importance of ERF genes on plant response to environmental stresses. The specific function of each of these genes is yet to be studied. In silico analysis suggests the methylation of promoters as a possible way to regulate them. More studies about the function of these ERFs and about the role of methylation in plant stress response should be done.
O arroz (Oryza sativa L.) é um dos cereais mais importantes no mundo, sendo muito cultivado em ecossistemas de várzea, os quais apresentam uma característica comum: condições de deficiência de drenagem. A falta de oxigênio, devido à submergência das plântulas, e a toxidez por excesso de ferro são dois dos maiores estresses abióticos dentre os quais as plantas de arroz são submetidas. Recentemente percebeu-se a importância de alguns fatores de transcrição responsivos ao etileno (ERFs) na resposta vegetal à diferentes estresses, em especial à deficiência de oxigênio. Considerando-se o exposto efetuaram-se basicamente dois estudos visando avaliar a expressão de ERFs em diferentes estresses. Em um primeiro estudo sete genes foram selecionados a partir de resultados anteriores obtidos de análise no Genevestigator e estes tiveram sua expressão transcricional avaliada por PCR quantitativa (qPCR) em plântulas de arroz sob deficiência de oxigênio e excesso de ferro. No segundo estudo treze ERFs, os quais não apresentavam informações disponíveis na literatura, também tiveram seus níveis de expressão transcricional avaliados em plântulas sob condições de estresse por hipoxia e anoxia, também através de qPCR em uma cv. sensível à inundação (Bonança) e uma tolerante (Nipponbare). Como conclusões gerais se ressalta, novamente, a importância dos ERFs na resposta vegetal frente a estresses ambientais. A função específica de cada um destes genes ainda necessita ser estudada. Uma análise in silico nos promotores destes genes indica a metilação como possível forma de regulação transcricional. Mais estudos sobre a função destes ERFs e sobre o papel da metilação na resposta de arroz a estresses deverão ser feitas.
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Chu, Ying-ying Jamie. "Characterization of the promoter of dehalogenase IVa gene of Burkholderia sp. MBA4." Click to view the E-thesis via HKUTO, 2006. http://sunzi.lib.hku.hk/hkuto/record/B37840150.
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Chu, Ying-ying Jamie, and 朱盈盈. "Characterization of the promoter of dehalogenase IVa gene of Burkholderia sp. MBA4." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B37840150.
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Grade, Carla Vermeulen Carvalho 1983. "Identificação e caracterização funcional dos elementos cis-regulatorios da miostatina." [s.n.], 2009. http://repositorio.unicamp.br/jspui/handle/REPOSIP/317672.
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Orientador: Lucia Elvira AlvaresDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: A proteina Miostatina (tambem conhecida como GDF8) e um membro da superfamilia de crescimento e diferenciacao ß (TGF- ß) e e expressa quase que exclusivamente em musculatura esqueletica, tanto no embriao em desenvolvimento quanto no individuo adulto, onde circula livre pela corrente sanguinea. A Miostatina foi inicialmente identificada em 1997 por MCPHERRON et al. e, desde entao, muitos estudos tem demonstrado seu papel essencial na regulacao do desenvolvimento de musculatura esqueletica de aves e mamiferos. O nocaute genico da Miostatina causa hiperplasia e hipertrofia das fibras musculares, resultando em musculos individuais ate duas vezes maiores do que em animais selvagens. Isso demonstra que a Miostatina e um regulador negativo da deposicao de musculatura esqueletica. A estrutura e a funcao desta proteina sao conservadas em diversas especies, incluindo humanos, onde os niveis de Miostatina circulante no sangue se encontram aumentados durante condicoes de distrofia e na caquexia que acompanha alguns tipos de cancer e a AIDS. Um melhor entendimento dos mecanismos que regem a expressao da Miostatina e essencial para o desenvolvimento de estrategias que possam regular sua atividade durante tais condicoes. No presente trabalho, nos identificamos, com o uso de ferramentas de Bioinformatica, elementos cisregulatorios putativos (promotor e enhancers) que possivelmente regulam a transcricao do gene da Miostatina. Inicialmente foi realizada uma comparacao dos loci do GDF8, incluindo as regioes intergenicas adjacentes, provenientes dos genomas de Humano, Camundongo e Galinha. Essa analise revelou a presenca de diferentes regioes evolutivamente conservadas (RECs) adjacentes a sequencia codificadora desta proteina, sete downstream e uma upstream ao gene. Por terem sido mantidas relativamente conservadas ao longo da evolucao, essas regioes supostamente possuem um papel funcional, possivelmente como elementos cis-regulatorios do gene da Miostatina. Em seguida, com o intuito de entender as funcoes que cada uma dessas regioes possa estar exercendo sobre a regulacao da atividade transcricional do gene da Miostatina, foi realizada uma busca por sitios de ligacao para fatores transcricionais que tenham sido conservados evolutivamente nessas RECs. Muitos sitios conservados foram observados nas sete RECs downstream ao gene da Miostatina, entre eles estao sitios para fatores relacionados ao desenvolvimento de musculatura esqueletica (MyoD, Myogenin, E47, EN1), membros (Pax3, Tbx5) e coracao (Nkx2.5, Pitx2). Juntos, esses dados sugerem uma regulacao modular do gene da Miostatina durante a embriogenese dos vertebrados. A unica REC localizada upstream ao GDF8 representa o promotor minimo putativo deste gene. Essa hipotese e reforcada pela presenca de um sitio de ligacao conservado para a Proteina de Ligacao ao sitio TATA. Com o intuito de validar as hipoteses formuladas com base nas analises de Bioinformatica, no presente trabalho buscamos caracterizar funcionalmente o promotor minimo do gene da Miostatina. Para tanto, a regiao do promotor minimo foi inicialmente clonada em um vetor que nao contem promotor e possui como gene reporter o GFP. Essa construcao de expressao foi entao testada atraves de experimentos de eletroporacao em embrioes de galinha in ovo. A analise dos embrioes eletroporados revelou que a regiao de DNA elegida para as analises funcionais e capaz de dirigir a transcricao do gene reporter, indicando que ela corresponde ao promotor minimo do gene da Miostatina. Alem do sitio TATA, ha, na regiao do promotor, diversos sitios conservados para a ligacao de proteinas envolvidas na via de sinalizacao mediada por cAMP (CREB, ATF, NFY). Esse achado esta de acordo com estudos recentes que demonstram o envolvimento do cAMP na regulacao dos fatores miogenicos Myf5 e MyoD, bem como de Pax3, sugerindo que a atividade do gene da Miostatina tambem possa estar sendo regulada por essa via de sinalizacao. Outras regioes do genoma humano que possuem arquitetura semelhante a observada no promotor da Miostatina foram identificadas, demonstrando que outros genes podem estar sob influencia da mesma via de sinalizacao que regula a atividade do promotor da Miostatina, dentre eles genes envolvidos na miogenese e neurogenese.
Abstract: The Myostatin protein (also known as GDF8) is a member of the transforming growth factor-ß (TGF-ß) superfamily and is expressed almost exclusively in skeletal muscle, both in the embryo and in the adult, where the protein circulates in the blood flow. It was initially identified in 1997 by MCPHERRON et al., and since then many studies have been demonstrating its essential role in the regulation of the development of skeletal muscle from birds and mammals. The knockout of the Myostatin gene causes both hyperplasia and hypertrophy of the skeletal muscle fibers, resulting in muscles twice as big as the wildtype ones, thus showing that Myostatin is a negative regulator of skeletal muscle deposition. The GDF8 structure and function is conserved in many species, including humans where the Myostatin levels are increased during dystrophy conditions and in the cachexia that accompanies some types of cancer and AIDS. A better understanding of the mechanisms that rule the Myostatin expression is essential for the development of strategies that might regulate its activity during such conditions. In this research, we have identified, with the use of bioinformatic tools, the cis-regulatory elements (promoter and enhancers) that regulate the Myostatin gene transcription. We compared the GDF8 loci from human, chicken and mouse and found different evolutionary conserved regions (ECRs), adjacent to the GDF8 coding sequence. Because these intergenic sequences remained relatively conserved throughout evolution, they supposedly have a functional role, possibly as cis-regulatory elements for the Myostatin gene. Our analyses revealed the presence of seven possible enhancers downstream of the GDF8 gene and one conserved region upstream of it. In order to understand the role these regions might have in the regulation of Myostatin's transcription activity, we searched for binding sites that were also evolutionary conserved. Many conserved binding sites were observed in the RECs downstream to the Myostatin gene, and among them are sites for factors related to the development of the skeletal muscle (MyoD, Myogenin, E47, EN1), limbs (Pax3, Tbx5) and heart (Nkx2.5, AREB6, Pitx2). Together, these data suggest a modular regulation of the Myostatin gene during vertebrates' embryogenesis. The only REC observed upstream of the Myostatin locus represents the putative basal gene promoter. This hypothesis is strengthened by the presence of a binding site for the Tata Binding Protein conserved for the studied species. In this research, we aimed at functionally characterizing the Myostatin gene basal promoter. For that purpose, we cloned the studied region in a promoterless vector, which contains GFP as a reporter gene. This expression construct was then tested through in ovo electroporation assays. The analysis of the electroporated embryos revealed that the cloned DNA region is capable of driving the transcription of the reporter gene, which indicates that it truly corresponds to the basal promoter of the Myostatin gene. Moreover, there are conserved binding sites for the CREB and ATF1 transcription factors in the basal promoter, which are activated by the cAMP signaling path. This finding is in agreement with recent studies that demonstrate the involvement of cAMP in the regulation of the myogenic factors Myf5 and MyoD, as well as Pax3, thus suggesting that the activity of the Myostatin gene might be under the influence of this signaling path. Other regions of the human genome that have a similar architecture to the one observed in the Myostatin promoter were identified. This demonstrates that other genes are possibly under the influence of the same signaling path regulating the activity of the Myostatin promoter, among them genes involved in myogenesis and neurogenesis.
Mestrado
Histologia
Mestre em Biologia Celular e Estrutural
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Riddihough, Guy. "The Drosophila hsp27 promoter." Thesis, University of Cambridge, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.258159.
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Rojas, Carlos Hernán Barrera [UNESP]. "Estudo da interação entre a via genética do microRNA156/squamosa promoter-binding protein-like e brassinosteróides durante o desenvolvimento de Arabidopsis thaliana." Universidade Estadual Paulista (UNESP), 2015. http://hdl.handle.net/11449/128123.
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000847745.pdf: 1720621 bytes, checksum: 5da3023273e6a040671b72235040def6 (MD5)O desenvolvimento vegetal é influenciado por diferentes fatores como os microRNAs (miRNAs) e os fitohormônios, os quaisinteragem numa complexa rede de regulação. Entre os miRNAs, o miRNA156 (miR156) regula os fatores de transcrição SQUAMOSA Promoter-Binding Protein-Like(SPL) afetando diferentes processos do desenvolvimento vegetal. Entre os fitohormônios, os brassinosteróides (BRs) participam na regulação dos eventos associados à fase juvenil da planta. A interação miR156/SPL-BRs não é conhecida, pelo qual, este estudo avaliou a interação destas duas vias durante o desenvolvimento juvenil de Arabidopsis thaliana. Formacaco da raiz principal (RP) e número de raizes laterais (RL) bem como o crescimento do hipocótilo foram utilizados como marcadores desta possivel interação. Foram utilizadas plantas de A. thaliana (Ecotipo Col-0) que expressam constitutivamente o miR156 (miR156-OE), plantas com níveis reduzidos do miR156 (Mimicry-156) e plantas selvagens (WT). Entre os BRs foi escolhido o 24-EpiBrassinolídeo (24-EBL) por ser o BR mais ativo. Plântulas miR156-OE apresentam maior comprimento da RP, maior número de RL e maior sensibilidade aos tratamentos com 24-EBL; fenótipos e comportamento opostosforam observados nas plântulas Mimicry-156. Além disso, plântulas miR156-OE apresentam maior comprimento do hipocótilo, enquanto as plântulas Mimicry-156 apresentam reduzido comprimento. Entre os genes SPLs que respondem ao tratamento com 24-EBL se encontram os SPL2, - 3, -4, -5, e -6. Entre os genes da via dos BRs foram observados alterações na expressão dos genes CPD, BZR1, BES1 e BAS1. Estes dados sugerem que a via genética do miR156/SPL interage com os BRs, e também contribuem para um melhor conhecimento da genética molecular do desenvolvimento de arabidopsis
Plant development is affected by different factors such as micro-RNAs (miRNAs) and phytohormones which interact in a complex regulation network. Among miRNAS, miRNA156 (miR156) regulates SQUAMOSA Promoter- Binding Protein-Like (SPL) transcription factor family affecting different plant development processes. Among phytohormones, brassinosteroids (BRs) participate in regulation of vegetal juvenile processes. miR156/SPL-BRs interaction is unknown whereby the aim of this work was to evaluate the interaction between those two pathways during Arabidopsis thaliana juvenile development. Main root (RP), lateral root number (RL) and hypocotyl length were selected as markers of this interaction. A. thaliana (Col-0 ecotype) overexpressing miR156 (miR156-OE), plants with miR156 reduced activity (Mimicry-156) and wild type (WT) plants were used. 24-Epibrassinolide (24- EBL), the most active BRs, was selected. miR156-OE plants have longer RP length, more RL and 24-EBL sensitivity. Opossite phenotypes were observed on Mimicry-156 plants. Besides, miR156-OE plants have longer hypocotyl length while Mimicry-156 plants have shorter. SPLs genes, SPL2, -3, -4, -5, and -6 responded to 24-EBL treatment. BRs pathway genes, CPD, BZR1, BES1 and BAS1 had changes in gene expression. Our data suggest a interaction between the miR156/SPL and BRs pathways and help to understand the molecular genetics of Arabidopsis development
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Cowie, Philip David. "Analysis of the effects of disease-associated variation within a cis-regulatory element of the CNR1 locus on CNR1 promoter dynamics." Thesis, University of Aberdeen, 2014. http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=225652.
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Genetic variation within the cannabinoid 1 receptor (CB1R) locus (CNR1) has been repeatedly associated with drug addiction pathologies. Genomic annotation of CNR1 indicates the vast majority of this genetic variation likely results in altered transcriptional regulation of the CNR1 gene as a mechanistic link to the disease phenotype. There is a lack of information describing the regulation of CNR1 transcription and the potential impact of disease-associated variation within the CNR1 locus on its transcriptional regulation. This study investigates the impact of an evolutionary conserved regulatory region of CNR1, termed ECR1, and the disease-associated variation contained within, on the transcriptional activity of the cognate CNR1 promoter region. Reporter assays conducted in primary hippocampal cells demonstrate that CNR1 promoter exhibits variable transcriptional activity during periods of CB1R signalling and cell depolarisation. Coupled to allelic variants of ECR1, the CNR1 promoter shows significant changes in transcriptional activity under resting conditions indicating that disease-associated variation within ECR1 may decrease CNR1 transcription. Further, alleles of ECR1 can drive allele-specific transcriptional responses from the CNR1 promoter during periods of CB1R stimulation and cell depolarisation. The results highlight the potential for disease-associated regulatory variation of the CNR1 locus to create stratified transcriptional responses to specific cell signalling scenarios and putatively to clinical strategies employing pharmacological agents. Furthermore, investigation of DNA-protein interactions at the allelic ECR1 region demonstrate that disease-associated variation within ECR1 alters DNA-protein interactions within the nucleus consistent with a decrease in transcriptional activity in the disease-associated allele variant. Collectively the current work supports the hypothesis that disease-associated variation within the ECR1 regulatory region of the CNR1 locus has the capacity to significantly impact on CNR1 promoter transcriptional activity. It is posited that allele-specific transcriptional effects may have a major impact on the susceptibility of individuals to drug addiction or on responses to clinical pharmacological treatments.
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Stamey, Jessica Reńee. "Isolation and characterization of the tubuliform spidroin 1 promoter from the black widow spider, Latrodectus Hesperus." Scholarly Commons, 2007. https://scholarlycommons.pacific.edu/uop_etds/670.
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Little is actually known about the transcriptional regulation of spider silk as most studies have focused on the material properties of silks. We isolated and mapped the TuSp1 core promoter from the black widow spider, Latrodectus hesperus. Using a genomic DNA walking strategy, we have isolated an upstream segment (581 bp) of genomic DNA containing the promoter as well as the first exon of the TuSp1 gene. This upstream regulatory element was able to initiate transcription in insect cells when placed upstream the promoterless firefly luciferase reporter gene. Initiation of transcription was orientation dependent, as insertion of this upstream regulatory module in the reverse orientation led to inefficient transcriptional initiation. Only 170 bp of upstream sequence was required for strong transcriptional initiation, showing that core promoter resides within the first 170 bp of upstream 5' -flanking DNA. We also demonstrate the bHLH factor SGSF1 can repress gene transcription of the TuSp1 core promoter, implying SGSF I might participate in the transcriptional regulation of the TuSp1 gene in vivo.
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Loewy, Amanda Duvall 1981. "Hypermethylation of the MMACHC promoter is associated with methionine dependence in the human malignant melanoma cell line Me-Wo-LC1." Thesis, McGill University, 2008. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=116118.
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Methionine dependence, the inability of cells to grow when the amino acid methionine is replaced in culture medium by its metabolic precursor homocysteine, is characteristic of many cancer cell lines. Most cells proliferate normally under these conditions. The methionine dependent tumorigenic human melanoma cell line MeWo-LC1 was derived from the methionine independent non-tumorigenic line MeWo. The MeWo-LC1 cell line has been shown to have a cellular phenotype similar to that of cells from patients with the cblC inborn error of cobalamin metabolism, with decreased synthesis of cobalamin coenzymes and decreased activity of the cobalamin dependent enzymes methionine synthase and methylmalonyl-CoA mutase. Inability of cblC cells to complement the defect in cobalamin metabolism in MeWo-LC1 suggested that the defect was caused by decreased activity of the MMACHC gene product. However, no potentially disease causing mutations could be detected in the coding sequence of MMACHC in MeWo-LC1. No MMACHC expression could be detected in MeWo-LC1, and there was virtually complete methylation of a CpG island at the 5' end of the MMACHC gene in MeWo-LC1, consistent with inactivation of the gene by methylation; the CpG island was partially methylated in MeWo and only lightly methylated in control fibroblasts. Transfection of MeWo-LC1 with wild type MMACHC with a constitutive promoter resulted in correction of the defect in cobalamin metabolism and restoration of the ability of cells to grow in medium containing homocysteine. We conclude that epigenetic inactivation of the MMACHC gene is responsible for methionine dependence in MeWo-LC1.
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Phillips, Julian Peter. "Promoter analysis in transgenic sugar beet." Thesis, De Montfort University, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.391084.
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