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Autor: Grafiati

Publicado: 26 de julio de 2024

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1

Quehl, Eike. "Etablierung transgener Zelllinien zur Visualisierung der Aktivität des Doublecortin-Promotors als Modell der Neurogenese in vitro". kostenfrei, 2008. http://www.opus-bayern.de/uni-regensburg/volltexte/2009/1149/.

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2

Hohenstatt, Antonia. "Hypoxia inducible factor 1 (HIF 1) alpha- und beta-vermittelte Induktion der ABCA1-Promotor-Aktivität". kostenfrei, 2009. http://www.opus-bayern.de/uni-regensburg/volltexte/2009/1306/.

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3

Sommer, Heide. "Der Übergang von der Latenz zur lytischen Replikation des Epstein-Barr-Virus vergleichende Analysen zur Bedeutung regulatorischer HI-Motive im Promotor des viralen Gens BZLF-1 /". [S.l.] : [s.n.], 2001. http://deposit.ddb.de/cgi-bin/dokserv?idn=962765007.

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4

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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5

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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6

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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7

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 Alvares
Tese (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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8

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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9

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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10

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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11

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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12

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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13

Chu, Ying-ying Jamie y 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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14

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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15

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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16

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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17

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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18

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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19

Zhao, Wei y 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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20

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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21

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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22

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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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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23

FOLEY, SUSAN MARIE. "HEALTH PROMOTION AND PRESYMPTOMATIC GENETIC TESTING". University of Cincinnati / OhioLINK, 2001. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1005250853.

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24

Wells, Carol Dawn. "The functional significance of the G to A point mutation in the promoter region of the Apolipoprotein AI gene". Master's thesis, University of Cape Town, 1993. http://hdl.handle.net/11427/27143.

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AG to A transition at position -76 in the promoter region of the apoAI gene was previously identified, and the A-76 has been shown to be associated with high apoAI levels. The functional significance of the point mutation was assessed by analysing the DNA-protein binding and promoter activities of the different alleles. This data would suggest that the point mutation alters the function of the apoAI promoter as gel retention assays revealed that the G fragment (-140 to +10) formed an extra DNA-protein complex compared to the A fragment (-140 to +10). Concurrent with the altered DNA-protein interaction between the G and the A fragments, the transcriptional activities of the apoAI gene were found to also be altered. CAT assays have indicated a 1.91 fold increase in promoter activity of the A fragment as compared to the G fragment (-256 to +397). The difference in promoter activity was, however, highly dependent on the particular fragment used, as no difference was observed between the alleles when a fragment {-256 to +68) was used. In this study elements were identified in the region +68 to +397 that causes a reduction in the promoter activity of the G allele by 3.6 fold, whilst reducing the A allele activity by 2 fold. This data would suggest that the point mutation functionally alters the apoAI promoter activity via its interaction with other sequences especially in the region +68 to +397.

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25

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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26

Chim, Chor-sang James y 詹楚生. "The role of aberrant gene promoter methylation in multiple myeloma". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B36898430.

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27

Yan, Shao-feng. "Development of an inducible promoter system in Leishmania donovani /". Thesis, Connect to this title online; UW restricted, 2000. http://hdl.handle.net/1773/9306.

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28

馮家禮 y Ka-lai Fung. "Patterns of gene promoter methylation in malignant lymphoma". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2001. http://hub.hku.hk/bib/B3122734X.

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29

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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30

Baynton, Clair Elizabeth. "Isolation and characterisation of a maize promoter". Thesis, University of Reading, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.239171.

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31

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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32

Chan, Ching Eunice. "Pathogenetic role of aberrant promoter methylation in lung cancer". Click to view the E-thesis via HKUTO, 2007. http://sunzi.lib.hku.hk/HKUTO/record/B39557819.

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33

Johnson, Sabrina D. "Characterization of the Pichia pastoris alcohol oxidase I promoter". Scholarly Commons, 2003. https://scholarlycommons.pacific.edu/uop_etds/575.

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The methylotrophic yeast, Pichia past oris, is one of the most respected and widely used systems today. The ability of this yeast to produce large masses of protein and metabolize methanol as a sole source of carbon and energy is attributed to the highly induceable Alcohol Oxidase I promoter (AOXI). Despite of the disperse popularity and use of this promoter over the last 15 years, little is known about the transcription controls at a molecular level. A 5'>3' deletion analysis of the AOXI promoter was perrormed to gain understanding of the promoter's regulation and provided insight to the approximate locations of the important regulatory regions. A total of 10 truncations were made unveiling two areas ofhigh activity located between positions, -257 to-235, and, -235 to -188. In addition, a 14-base pair internal deletion was made between positions, -215 to -201. This region was shown to be necessary for transcriptional activation by deletion analysis. Sufficiency studies suggested that this 14-base pair element could serve as an activator sequence in both glucose and methanol.

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34

Lee, Sun K. "Genetic transformation of broccoli and promoter tagging in Brassica species". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1996. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq24027.pdf.

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35

Gardella, Thomas James. "A Genetic Analysis of RNA Polymerase-Promoter Interactions: A Thesis". eScholarship@UMMS, 1988. http://escholarship.umassmed.edu/gsbs_diss/200.

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Transcription initiation is a key step at which gene expression can be regulated. The sigma subunit of RNA polymerase provides the enzyme with the ability to recognize promoter sequences and initiate transcription at specific sites on the chromosome. The molecular basis of sigma function is not well known. It has been suggested that sigma factors confer promoter specificty by making direct contacts to the promoter DNA (Losick and Pero, 1981). To test this idea, suppressors of promoter down mutations were sought that affected the promoter recogniton properties of the σ70 subunit of E. coli RNA polymerase. Four such sigma mutants were obtained, two of which are allele-specific. One of these mutants has a change at a position in the predicted helix-turn-helix DNA binding structure which lies in a conserved region of the protein (region 4). This mutant specifically suppresses promoter down mutations in the -35 region of the promoter. The other mutant has a change at a residue that lies in a predicted α-helix of conserved region 2. This mutant specifically suppresses promoter mutations in the -10 region of the promoter. These data support the idea that regions 2 and 4 of sigma interact with the -10 and -35 regions of the promoter, respectively.

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36

Burbridge, Stephen Anthony. "Analysis of the Xenopus N-cadherin promoter region". Thesis, University of Warwick, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.362548.

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37

Rojas, Carlos Hernán Barrera. "Estudo da interação entre a via genética do microRNA156/squamosa promoter-binding protein-like e brassinosteróides durante o desenvolvimento de Arabidopsis thaliana /". Botucatu, 2015. http://hdl.handle.net/11449/128123.

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Orientador: Fábio Tebaldi Silveira Nogueira
Banca: Luiz Fernando Rolim de Almeida
Banca: Daniel Scherer de Moura
Banca: Maria Isabel Nogueira Cano
Banca: Paulo Mazzafera
Resumo: 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
Abstract: 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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38

Rundlöf, Anna-Klara. "Expression of thioredoxin reductase 1 in mammalian cells with regulation by the core promoter and use of alternative splice variants /". Stockholm, 2003. http://diss.kib.ki.se/2003/91-7349-757-6/.

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39

Wong, Li-Chuen. "IL-10 promoter polymorphisms in atopic dermatitis". Thesis, The University of Sydney, 2002. https://hdl.handle.net/2123/27849.

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Atopic dermatitis (AD) is one of the many clinical manifestations of atopy. It is one of the most common chronic inflammatory skin diseases with increasing prevalence over the last century. The aetiology of this disease is multifactorial with a complex interaction of genetic, environmental and immunologic factors.

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40

Chim, Chor-sang James y 詹楚生. "Study of gene promoter methylation in acute promyelocytic leukaemia". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2002. http://hub.hku.hk/bib/B25256725.

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41

Chan, Ching Eunice y 陳清. "Pathogenetic role of aberrant promoter methylation in lung cancer". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B39557819.

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42

McCarthy, Shane. "Comparative sequencing of candidate genes in complex disease /". Stockholm, 2006. http://diss.kib.ki.se/2006/91-7140-663-8/.

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43

Kim, S. "Structural analysis of the TRPI promoter in Saccharomyces cerevisiae". Thesis, University of Oxford, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.306685.

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44

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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45

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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46

Chim, Chor-sang James. "The role of aberrant gene promoter methylation in multiple myeloma". Click to view the E-thesis via HKUTO, 2006. http://sunzi.lib.hku.hk/hkuto/record/B36898430.

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47

Macleod, Donald T. "A molecular analysis of a promoter trap in embryonic stem systems". Thesis, University of Edinburgh, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.280628.

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48

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 Alvares
Dissertação (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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Deljou, Ali. "Tissue culture and genetic transformation in potato breeding". Thesis, University of Nottingham, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.339661.

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Holmqvist, Per-Henrik. "Transcription factor effects on chromatin organisation and gene regulation /". Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-453-8/.

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