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1
Adegoke, Oluwajoba Oluwapelumi. "Transcriptional and post-transcriptional regulation in testicular toxicity". Thesis, University of Leicester, 2015. http://hdl.handle.net/2381/31979.
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The control of gene expression occurs at multiple levels one of which is controlled by epigenetic regulation. In this work, it was hypothesised that changes in DNA methylation (transcriptional level) and miRNA expression (post-transcriptional level) might be involved in the mechanism of compound-induced testicular toxicity. mRNA and miRNA analysis of mouse testis was performed following exposure to dibutyl phthalate, 17β-estradiol and doxorubicin. Pathway analysis of transcriptional changes revealed all three chemicals interfered with the steroidogenic pathway, with further modulation of oxidative stress pathways in doxorubicin models. Doxorubicin exhibited a profound effect on the testis by decreasing the expression of germ cell-specific transcripts and increasing the expression of Leydig cell transcripts, apoptotic genes and pro-apoptotic miRNAs (miR-145, miR-26a, miR29 family). The post-transcriptional regulatory activity of these proapoptotic miRNAs was demonstrated by decreased transcript expression of their target DNA-methyl transferases (Dnmt) transcripts. An extensive deregulation of DNA methylation was observed that could be a consequence of altered Dnmts levels. Hypomethylation of genes, such as Cdkn2a and Pcna2, led to activation of p53 signaling. The same experiment was repeated in in vitro models of the testis. Pathway analysis revealed miRNA-mRNAs regulation of signaling pathways between germ cell-Sertoli cell and Sertoli cell-Sertoli cell junctions. A systematic review was conducted to establish the role of epigenetic-mediated mechanisms in toxicant-induced male reproductive toxicity. The study identified that decrease in Dnmt levels following chemical exposure could play a role in germ cell apoptosis. Also, the aberrant methylation of H19 could serve as a useful biomarker in the transgenerational effects of chemicals. The findings from this project provide further insight into the mechanisms of compound-induced testicular toxicity, through the utilization of genomics and a systematic review approach to published work. It identified epigenetic mechanisms both at the transcriptional and post-transcriptional levels are involved in the mechanism of toxicity.
2
Dennis, Jonathan Hancock. "Transcriptional regulation by Brn 3 POU domain containing transcription factors". Thesis, University College London (University of London), 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.249684.
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Koutsoulidou, Andrie. "Investigation of transcriptional and post-transcriptional regulation of myogenesis". Thesis, University of Bristol, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.559081.
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Skeletal myogenesis IS a complicated and tightly regulated process, at both transcriptional and post-transcriptional levels. Muscle cells isolated from different stages of the human foetal development displayed increased capacity to differentiate in vitro at late stages of the development. Twist is an early developmental transcription factor shown to inhibit muscle differentiation in mice. Endogenous human TWIST (H-TWIST) protein levels were found to be inversely proportional to the state of foetal muscle development and the capacity of isolated myoblasts to differentiate in vitro. Investigation of H-TWIST gene transcriptional regulation revealed that endogenous MyoD binds to the H- TWIST promoter and inhibits its expression. Overexpression of MyoD increased the low capacity of human foetal myoblasts to differentiation in vitro and decreased the levels of H- TWIST protein. These results propose a mechanism by which MyoD downregulates the expression of H-TWIST gene, thus promoting myogenesis. MicroRNAs are non-coding RNA molecules that post-transcriptionally regulate many cellular processes. MiR-l, miR- 133a, miR-133b and miR-206, also known as myomiRs, are expressed in muscle tissue and induced during muscle cell differentiation. MyomiRs were found to be increased during late stages of human foetal muscle development. Increases in their expression levels were proportional to the capacity of myoblasts to differentiate in vitro. Changes in myomiR levels during human foetal development were accompanied by endogenous alterations in their known targets and also in their inducer, MyoD. Overexpression of the latter resulted in an induction of muscle cell differentiation in vitro, accompanied by an increase in the levels of miR-l, miR-133a, miR-133b and miR-206. Myotonic dystrophy type 1 is a muscular dystrophy characterized by impaired muscle cell differentiation. Muscle cells isolated from DMl patients from distinct developmental stages showed a reduction in the expression levels of myomiRs during both differentiated and undifferentiated stages, verifying their important role during myogenesis.
4
Asif, Hafiz Muhammad Shahzad. "Inference dynamics in transcriptional regulation". Thesis, University of Edinburgh, 2012. http://hdl.handle.net/1842/6238.
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Computational systems biology is an emerging area of research that focuses on understanding the holistic view of complex biological systems with the help of statistical, mathematical and computational techniques. The regulation of gene expression in gene regulatory network is a fundamental task performed by all known forms of life. In this subsystem, modelling the behaviour of the components and their interactions can provide useful biological insights. Statistical approaches for understanding biological phenomena such as gene regulation are proving to be useful for understanding the biological processes that are otherwise not comprehensible due to multitude of information and experimental difficulties. A combination of both the experimental and computational biology can potentially lead to system level understanding of biological systems. This thesis focuses on the problem of inferring the dynamics of gene regulation from the observed output of gene expression. Understanding of the dynamics of regulatory proteins in regulating the gene expression is a fundamental task in elucidating the hidden regulatory mechanisms. For this task, an initial fixed structure of the network is obtained using experimental biology techniques. Given this network structure, the proposed inference algorithms make use of the expression data to predict the latent dynamics of transcription factor proteins. The thesis starts with an introductory chapter that familiarises the reader with the physical entities in biological systems; then we present the basic framework for inference in transcriptional regulation and highlight the main features of our approach. Then we introduce the methods and techniques that we use for inference in biological networks in chapter 2; it sets the foundation for the remaining chapters of the thesis. Chapter 3 describes four well-known methods for inference in transcriptional regulation with pros and cons of each method. Main contributions of the thesis are presented in the following three chapters. Chapter 4 describes a model for inference in transcriptional regulation using state space models. We extend this method to cope with the expression data obtained from multiple independent experiments where time dynamics are not present. We believe that the time has arrived to package methods like these into customised software packages tailored for biologists for analysing the expression data. So, we developed an open-sources, platform independent implementation of this method (TFInfer) that can process expression measurements with biological replicates to predict the activities of proteins and their influence on gene expression in gene regulatory network. The proteins in the regulatory network are known to interact with one another in regulating the expression of their downstream target genes. To take this into account, we propose a novel method to infer combinatorial effect of the proteins on gene expression using a variant of factorial hidden Markov model. We describe the inference mechanism in combinatorial factorial hidden model (cFHMM) using an efficient variational Bayesian expectation maximisation algorithm. We study the performance of the proposed model using simulated data analysis and identify its limitation in different noise conditions; then we use three real expression datasets to find the extent of combinatorial transcriptional regulation present in these datasets. This constitutes chapter 5 of the thesis. In chapter 6, we focus on problem of inferring the groups of proteins that are under the influence of same external signals and thus have similar effects on their downstream targets. Main objectives for this work are two fold: firstly, identifying the clusters of proteins with similar dynamics indicate their role is specific biological mechanisms and therefore potentially useful for novel biological insights; secondly, clustering naturally leads to better estimation of the transition rates of activity profiles of the regulatory proteins. The method we propose uses Dirichlet process mixtures to cluster the latent activity profiles of regulatory proteins that are modelled as latent Markov chain of a factorial hidden Markov model; we refer to this method as DPM-FHMM. We extensively test our methods using simulated and real datasets and show that our model shows better results for inference in transcriptional regulation compared to a standard factorial hidden Markov model. In the last chapter, we present conclusions about the work presented in this thesis and propose future directions for extending this work.
5
Kang, Martin Hubert. "Post-transcriptional regulation of ABCA1". Thesis, University of British Columbia, 2012. http://hdl.handle.net/2429/43655.
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Epidemiological studies consistently demonstrate an inverse relationship between HDL levels and cardiovascular disease (CVD), independent of LDL and triglyceride levels. Due to the crucial role ABCA1 plays in HDL biogenesis, increasing ABCA1 expression is considered an attractive strategy to increase plasma HDL levels. In this thesis we attempt to identify novel post-transcriptional and post-translational mechanisms that regulate ABCA1 expression and/or function. Prior to translation, ABCA1 protein expression is regulated by non-coding RNA molecules known as microRNAs which bind and inhibit translation of mature mRNA transcripts in the cytoplasm. In this study we used bioinformatic prediction programs to identify potential microRNA regulators of ABCA1. Using reporter constructs, protein expression analysis by immunoblotting, and cholesterol efflux assays, we validated microRNA-145 as a novel repressor of ABCA1 translation. The inhibition of endogenous microRNA-145 in HepG2 cells increases both ABCA1 protein levels and cholesterol efflux activity. The inhibition of this microRNA in the liver is a potential strategy to increase HDL levels. Following translation, numerous post-translational modifications and protein-protein interactions are required for the ABCA1 protein to function properly. In this study we identified palmitoylation as a novel post-translational modifier of ABCA1. The majority of ABCA1-mediated cholesterol efflux and HDL biogenesis occurs at the cell surface. We show that palmitoylation is a crucial lipid addition for proper ABCA1 plasma membrane localization. We also identify a number of enzymes that mediate the incorporation of radio-labeled palmitate onto ABCA1, and demonstrate that the overexpression of the palmitoyl transferase enzyme DHHC8 increases ABCA1 palmitoylation and cholesterol efflux activity. The increase of ABCA1 palmitoylation in the liver is a novel strategy to increase HDL levels. In this thesis, we have contributed to the understanding of ABCA1 biology by the identification of two novel regulators of ABCA1 expression and/or function.
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Glasspool, Rosalind M. "The transcriptional regulation of telomerase". Thesis, University of Glasgow, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.398635.
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Weintraub, Abraham S. (Abraham Selby). "Transcriptional regulation and genome structure". Thesis, Massachusetts Institute of Technology, 2018. http://hdl.handle.net/1721.1/117886.
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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2018.Cataloged from PDF version of thesis. Page 162 blink.
Includes bibliographical references.
The regulation of gene expression is fundamental to the control of cell identity, development and disease. The control of gene transcription is a major point in the regulation of gene expression. Transcription is regulated by the binding of transcription factors to DNA regulatory elements known as enhancers and promoters. This leads to the formation of a DNA loop connecting the enhancer and the promoter resulting in the subsequent transcription of the gene. Thus the structuring of the genome into DNA loops is important in the control of gene expression. This thesis will focus on the role of genome structure in transcriptional regulation. Two key questions in this area that I have attempted to address during my PhD are "how are enhancer-promoter interactions constrained so that enhancers do not operate nonspecifically?" and "are there proteins that facilitate enhancer-promoter looping?" I will describe the identification of DNA loop structures formed by CTCF and cohesin that constrain enhancer-promoter interactions. These structures-termed insulated neighborhoods-are perturbed in cancer and this perturbation results in the inappropriate activation of oncogenes. Additionally, I will describe the identification and characterization of the transcription factor YY1 as a factor that can structure enhancer-promoter loops. Through a combination of genetics, genomics, and biochemistry, my studies have helped to identify insulated neighborhood structures, shown the importance of these structures in the control of gene expression, revealed that these structures are mutated in cancer, and identified YY1 as a structural regulator of enhancer-promoter loops. I believe these studies have produced a deeper understanding of the regulatory mechanisms that connect the control of genome structure to the control of gene transcription.
by Abraham S. Weintraub.
Ph. D.
8
McCormick, Margaret M. (Margaret Mary). "Transcriptional regulation in Corynebacterium glutamicum". Thesis, Massachusetts Institute of Technology, 1996. http://hdl.handle.net/1721.1/11197.
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Hochhauser, Daniel. "Transcriptional regulation of topoisomerase II". Thesis, University of Oxford, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.333178.
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Reid, John Edward. "Probabilistic models of transcriptional regulation". Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648864.
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Mates, Jessica Marie. "TRANSCRIPTIONAL REGULATION OF HIV-1". The Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1395845500.
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Li, Charles H. Ph D. (Charles Han)Massachusetts Institute of Technology. "Genome organization in transcriptional regulation". Thesis, Massachusetts Institute of Technology, 2020. https://hdl.handle.net/1721.1/130664.
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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, February, 2021Cataloged from the official PDF of thesis. "February 2021."
Includes bibliographical references.
Transcriptional regulation of gene expression plays critical roles in the control of cell identity, development, and disease. Genome organization contributes to transcriptional regulation in multiple ways. At a fundamental level, the genome is organized into distinct active and repressive chromatin states that facilitate transcriptional regulation. These chromatin states are established and maintained at specific genomic regions via the interconnected activities of transcription factors and epigenetic pathways. An additional layer of genome organization is the three-dimensional structure of the genome within the nucleus. Transcriptional regulation occurs within a hierarchy of genome structures that are formed by the activities of structuring factors. Studies described in this thesis identify the transcription factor YY1 as a general structural regulator of enhancer-promoter loops (Weintraub et al., 2017). In recent years, the study of biomolecular condensates has led to a dramatic shift in our understanding of the mechanisms contributing to transcriptional regulation and to genome structure. Distinct chromatin condensates organize the genome by compartmentalizing components associated with transcriptionally active euchromatin and repressive heterochromatin. Whether disruption of chromatin condensates can lead to transcriptional dysregulation in human disease is not well understood. Our finding that MeCP2 is a key component of heterochromatin condensates and that Rett syndrome patient mutations affecting MeCP2 cause condensate disruption (Li et al., 2020), demonstrates a link between chromatin condensate disruption and human disease. These studies reveal important mechanisms of genome organization contributing to transcriptional regulation, and provide new insights into human disease that might be leveraged to provide therapeutic benefit for patients in the future.
by Charles H. Li.
Ph. D.
Ph.D. Massachusetts Institute of Technology, Department of Biology
13
Karnowski, Alexander. "Post-transcriptional regulation of IgE". [S.l. : s.n.], 2004. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB10990069.
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Sodaei, Reza 1988. "Transcriptional and post-transcriptional regulation across biological units and time". Doctoral thesis, Universitat Pompeu Fabra, 2020. http://hdl.handle.net/10803/669533.
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How do genomes generate different tissues, behaviours, individuals or species? We
try to identify parts of the human transcriptome that we found only in a tissue. Then,
we focus on inter-individual extreme transcriptomic differences, and how this principle
is extendable to species variation. In the second part, we measured the transcriptomic
shifts during day-night and seasonal cycles in human postmortem tissues.Com generen diferents genomes diferents teixits, conductes, individus o espècies? Tractem d'identificar parts del transcriptoma humà que hem trovat només en un teixit. Seguidament, ens centrem en diferències transcriptòmices extremes entre individus, i en com aquest principi es pot extendre a les variacions de espècie. Al segon capítol, mesurem els canvis transcriptòmics durant cicles de dia i nit així com estacionals en teixits humans postmortem.
15
Lee, Yiu-fai Angus, i 李耀輝. "Tissue-specific transcriptional regulation of Sox2". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2007. http://hub.hku.hk/bib/B3955739X.
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Ching, Chi-yun Johannes, i 程子忻. "Transcriptional regulation of p16INK4a expression by the forkhead box transcription factor FOXM1". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2003. http://hub.hku.hk/bib/B29466192.
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Yao, Ya-Li. "Regulation of yy1, a multifunctional transciption [sic] factor /". [Tampa, Fla.] : University of South Florida, 2001. http://purl.fcla.edu/fcla/etd/SFE0000626.
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Remes, Lenicov Federico. "Transcriptional regulation of tryptophan hydroxylase-2". Thesis, University of Ottawa (Canada), 2006. http://hdl.handle.net/10393/27413.
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Dysregulation of serotonergic neurotransmission is a contributing cause of numerous pathologies. In the brain, the enzyme catalyzing the rate-limiting step in serotonin biosynthesis and thus controlling serotonin levels is tryptophan hydroxylase-2 (TPH2). The objective of this project is to study the regulation of TPH2 transcription by characterizing its promoter region.
Study of the 5' flanking region of the human TPH2 gene by means of reporter gene assays resulted in the finding of a core promoter and a repressing element between positions -179 and -88 relative to the transcription start site. In addition, the TPH2 promoter could be activated by Ca++ mobilization in a cell-line model of serotonergic neurons, but not in other cell lines. In agreement, Ca++ mobilization in this model also induced endogenous transcription of TPH2.
This work is the first one to identify the promoter region of the TPH2 gene and to report its activity-dependent regulation of transcription.
19
Elfving, Anna. "Transcriptional regulation of mouse ribonucleotide reductase". Licentiate thesis, Umeå universitet, Institutionen för medicinsk kemi och biofysik, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-41272.
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All living organisms are made of cells and they store their hereditary information in the form of double stranded DNA. In all organisms DNA replication and repair is essential for cell division and cell survival. These processes require deoxyribonucleotides (dNTPs), the building blocks of DNA. Ribonucleotide reductase (RNR) is catalyzing the rate limiting step in the de novo synthesis of dNTPs. Active RNR is a heterodimeric protein complex. In S phase cells, the mouse RNR consists of the R1 and the R2 proteins. The R1/R2 RNR-complex supplies the cell with dNTPs required for DNA replication. Outside S-phase or in non-proliferating cells RNR is composed of R1 and p53R2 proteins. The R1/p53R2 RNR-complex supplies cells with dNTPs required for mitochondrial DNA replication and for DNA repair. An undisturbed dNTP regulation is important since unbalanced dNTP pools results in DNA mutations and cell death. Since unbalanced pools are harmful to the cell, RNR activity is regulated at many levels. The aim of this thesis is to study how the mouse RNR genes are regulated at a transcriptional level. We have focused on the promoter regions of all three mouse RNR genes. Primer extension experiments show that the transcription start of the TATA-less p53R2 promoter colocalizes with an earlier unidentified initiator element (Inr-element). This element is similar to the known Inr-element in the mouse R1 promoter. Furthermore, functional studies of the R1 promoter revealed a putative E2F binding element. This result suggests that the S phase specific transcription of the R1 gene is regulated by a similar mechanism as the R2 promoter which contains an E2F binding site. Finally we have established a method to partially purify the transcription factor(s) binding the upstream activating region in the mouse R2 promoter by phosphocellulose chromatography and affinity purification using oligonucleotides immobilized on magnetic beads. This method will allow us to further study the transcription factors responsible for activating expression of the R2 protein. This method has a potential to be utilized as a general method when purifying unknown transcription factors.
20
Gardner-Stephen, Dione Anne, i dione bourne@flinders edu au. "Transcriptional Regulation of Human UDP-Glucuronosyltransferases". Flinders University. Medicine, 2008. http://catalogue.flinders.edu.au./local/adt/public/adt-SFU20081111.223136.
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The UDP-glucuronosyltransferases (UGTs) are a superfamily of enzymes that glucuronidate small, lipophilic molecules, thereby altering their biological activity and excretion. In humans, important examples of UGT substrates include molecules of both endogenous and xenobiotic origin; thus, UGTs are considered essential contributors to homeostatic regulation and an important defence mechanism against chemical insult. In keeping with both roles, UGTs are most strongly expressed in the liver, a predominant organ involved in detoxification.
Rates of glucuronidation in humans are neither uniform among individuals, nor constant in an individual over time. Genetic determinants and non-endogenous signals are both known to influence the expression of UGTs, which in turn may affect the efficacy of certain pharmaceutical treatments or alter long-term risk of developing disease. Thus, this thesis focuses on the transcriptional regulation of UGT genes in humans, particularly on mechanisms that are likely to be relevant to their expression and variation in the liver. Two major approaches were used: firstly, extensive studies of several UGT promoters were performed to identify and characterise transcriptional elements that are important for UGT expression; and secondly, important hepatic transcription factors were investigated as potential regulators of UGT genes.
UGT1A3, UGT1A4 and UGT1A5 are a subset of highly related, but independently regulated, genes of the human UGT1 subfamily. UGT1A3 and UGT1A4 are expressed in the liver, whereas UGT1A5 is not. The presented analysis of the UGT1A3, UGT1A4 and UGT1A5 proximal promoters demonstrates that a hepatocyte nuclear factor (HNF)1-binding site common to all three promoters is important for UGT1A3 and UGT1A4 promoter activity in vitro, but is insufficient to drive UGT1A5 expression. Two additional elements required for the maximal activity of the UGT1A3 promoter were also identified that may distinguish this gene from UGT1A4. UGT1A3 was investigated further, focusing on mechanisms that may contribute to interindividual variation in UGT1A3 expression. Polymorphisms in the UGT1A3 proximal promoter were identified and their functional consequences tested. Known variants of HNF1alpha were also tested for altered activity towards the UGT1A3 gene.
UGT1A9 is the only hepatic member of the UGT1A7-1A10 subgroup of UGT1 enzymes. Previous work had identified HNF1-binding sites in all four genes, and HNF4alpha as an UGT1A9-specific regulator. The work presented herein extends these findings to show that HNF1 factors and HNF4alpha synergistically regulate UGT1A9, and that HNF4alpha is not the only transcription factor responsible for the unique presence of UGT1A9 in the liver.
Liver-enriched transcription factors screened as potential UGT regulators were chosen from the HNF1, HNF4, HNF6, FoxA and C/EBP protein families. Functional interactions newly identified by this work were HNF4alpha with UGT1A1 and UGT1A6, HNF6 with UGT1A4 and UGT2B11, FoxA1 and FoxA3 with UGT2B11, UGT2B15 and UGT2B28 and C/EBPalpha with UGT2B17. Observations were also made regarding different patterns of interaction between each UGT and the transcription factors tested, particularly HNF1alpha.
21
Hayden, Celine. "Post-Transcriptional Gene Regulation in Plants". Diss., Tucson, Arizona : University of Arizona, 2006. http://etd.library.arizona.edu/etd/GetFileServlet?file=file:///data1/pdf/etd/azu%5Fetd%5F1684%5F1%5Fm.pdf&type=application/pdf.
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Dalton, Stephen. "Transcriptional regulation of histone gene expression /". Title page, contents and summary only, 1987. http://web4.library.adelaide.edu.au/theses/09PH/09phd152.pdf.
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Lee, Yiu-fai Angus. "Tissue-specific transcriptional regulation of Sox2". Click to view the E-thesis via HKUTO, 2007. http://sunzi.lib.hku.hk/HKUTO/record/B3955739X.
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Kasaai, Bahar. "Transcriptional regulation of the Bril gene". Thesis, McGill University, 2011. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=103567.
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Bril is a 134 amino acid-long protein compartmentalized exclusively on the osteoblast cell surface, with two transmembrane domains. Its osteoblast-specific expression coincides with the onset of bone matrix formation and mineralization and has been shown to have an essential role in bone mineralization in vitro, although the mechanism involved is still unknown. The purpose of this study was to characterize the regulation pattern of the Bril gene by identifying the cis-elements on the promoter and the transcription factors that modulate its activation. The promoter for the mouse, rat, and human Bril gene were independently cloned into a luciferase (Luc) reporter plasmid. Transient transfection studies were then performed to compare the activity of the Luc reporter in permissive (UMR106, MC3T3-E1) and non-expressing cell lines (HEK293). Deletion mutant analyses revealed that the proximal 300bp promoter was sufficient to confer highest activation in the three species studied. In silico analysis, EMSA and DNAseI footprinting studies revealed that this region is rich in GC-boxes and contains many potential regulatory elements known to be involved in osteoblastic differentiation (e.g. Runx2, TCF1/LEF, Osx/Sp7/Sp1) that are fully functional. Co-transfection experiments in HEK293 and MC3T3-E1 cells showed that the Bril promoter is most strongly trans-activated by forced expression of Sp1 and the long variant of Sp3, moderately by members of the Wnt pathway (i.e. β-catenin and TCF/LEF), yet is not considerably affected by bone-specific factors. Additionally, shRNA-mediated knockdown of Sp1 (but not Sp3) in UMR106 cells resulted in complete abrogation of Bril protein. In vitro interference of the GC-rich boxes via GC-bisintercalating agents and CpG methylation resulted in a dramatic drop in Bril promoter activity; which highlights the importance of these GC-sequences in promoting Bril regulation. In a search for molecules that might regulate Bril expression, the parathyroid hormone (PTH) was found to be a potent negative regulator. Endogenous Bril expression in UMR106 was dramatically downregulated by PTH in a time- and dose-dependent manner, which coincided with suppressed osteoblast mineralization. These results provide the first mechanistic evidence for regulation of the Bril gene, which could help situate its function in the bone mineralization process in vitro.Bril est une protéine de 134 acides aminés, localisée exclusivement sur la membrane des ostéoblastes avec deux domaines transmembranaires. Son expression exclusive dans les ostéoblastes coïncide avec le début de la formation osseuse et la minéralisation de la matrice. Ainsi, un rôle essentiel a été démontré dans la minéralisation osseuse in vitro, bien que le mécanisme en cause n'est pas encore connu. L'objectif de cette étude était de caractériser le mode de régulation du gène Bril en identifiant les éléments cis sur le promoteur et les facteurs de transcription qui modulent son activation. Le promoteur du gène Bril de la souris, du rat et de l'homme ont été indépendamment clonés dans un plasmide rapporteur de la luciférase (Luc). Des transfections transitoires ont ensuite été effectuées pour comparer l'activité Luc dans des cellules qui expriment (UMR106, MC3T3-E1) ou non (HEK293) le gène Bril de manière constitutive. L'analyse des mutants de délétion a révélé que le promoteur proximal 300 pb est suffisant pour conférer la majorité d'activation dans les trois espèces étudiées. Ainsi, les analyses in silico ont révélé que cette région est riche en séquences de GC et contient des éléments de régulation putatifs connus pour être impliqués dans la différenciation ostéoblastique (par exemple Runx2, TCF1/LEF, Osterix ou Sp1). Des analyses de retard sur gel (EMSA) et de cartographie à la DNAseI ont indiqué que beaucoup d'entre eux sont des sites fonctionnels. Les co-transféctions dans des cellules HEK293 et MC3T3-E1 ont montré que le promoteur Bril est le plus fortement trans-activé par l'expression forcée de Sp1 et la variante longue du Sp3, modérément par des membres de la voie Wnt (β-caténine et TCF /LEF), mais ne sont pas encore fortement influencés par des facteurs spécifiques à l'os. En plus, l'invalidation dans les cellules UMR106 de Sp1 (et non Sp3) par les shRNAs mène à l'abrogation complète de la protéine Bril. In vitro, l'interférence des séquences GC (via les drogues MMA et WP631 et la méthylation) a entraîné une forte baisse de l'activité promoteur de Bril; ce qui souligne l'importance de ces GC-séquences dans la promotion et la réglementation du Bril et propose la méthylation de l'ADN comme un mécanisme à considérer dans la spécificité d'expression cellulaire de Bril. La recherche de molécules qui pourraient réguler l'expression de Bril a identifié l'hormone parathyroïde (PTH) comme un puissant régulateur négatif. L'expression endogène de Bril dans les UMR106 a été considérablement diminuée par la PTH dans une manière dose-dépendante, qui coïncide avec une reduction de la minéralisation des ostéoblastes. Ces résultats fournissent les premières preuves d'un mécanisme pour la régulation du gène Bril, qui pourrait aider à démontrer sa fonction dans le processus de minéralisation osseuse in vitro.
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Binda, Olivier. "Regulation of RBP1BCAA transcriptional repression activities". Thesis, McGill University, 2007. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=103023.
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Normal cellular proliferation is controlled by two major pathways, the pRB and the p53 pathways. It is believed that one or both of these two pathways is disrupted in all tumour cells. The functions of pRB are disrupted by viral oncoproteins, which displace important cellular components from a highly conserved region known as the "pocket". The first cellular "pocket"-binding protein identified is the retinoblastoma binding protein 1 (RBP1). RBP1 and the RBP1-like protein BCAA are inherent subunits of the chromatin modifying transcription repression complex mSIN3A/HDAC. By linking the pRB protein to this repression complex, RBP1 is believed to mediate repression of E2F-dependent transcription, thereby regulating cellular proliferation.RBP1 and BCAA repress transcription in both HDAC-independent (R1) and HDAC-dependent (R2) manners. I show in this work that, as with RBP1, BCAA can associate with the mSIN3A/HDAC complex via a direct interaction with the SAP30 subunit. The region responsible for this interaction (R2) mediates HDAC-dependent transcriptional repression. The latter is regulated by the NAD+-dependent enzymatic activity of the class III histone deacetylase longevity factor SIRT1, which is recruited to the mSIN3A/HDAC complex via the tumour suppressors ING1b/ING2 and probably acts by deacetylating HDAC1.
The HDAC-independent repression activity of both RBP1 and BCAA is regulated by post-translational modifications. The R1 repression activity can be further dissected into a domain that targets basal transcription (R1alpha) and one that represses both basal and activated transcription (R1sigma). SUMOylation of the R1sigma region is essential for its repression activities. SUMOylation of R1sigma is itself regulated by the overall local amino acids charge. In agreement with published results demonstrating that a negative charge in the vicinity of the PsiKxE site allows interaction with a positively charged patch on the SUMO E2 ligase enzyme UBC9, neutralization of lysine positive charge by acetylation of R1sigma allows SUMOylation whereas mutation of lysines to arginines conserves the positive charge and hinders SUMOylation.
The biological relevance of RBP1 and BCAA transcriptional repression activities is highlighted by their requirement for induction of cell growth arrest and terminal cell cycle withdrawal or cellular senescence.
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Goodison, Steven. "Transcriptional regulation of the insulin gene". Thesis, University of Oxford, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.316878.
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Chapman, Andrew Glen. "Transcriptional regulation of the XIST locus". Thesis, University of British Columbia, 2013. http://hdl.handle.net/2429/45021.
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X-chromosome inactivation is a mechanism that has evolved in mammalian females
allowing dosage compensation of X-linked genes. A region of the X chromosome called the Xinactivation
centre (XIC) is required for X inactivation to occur. Within this region is a long noncoding
RNA, XIST/Xist, which is upregulated on the future inactive X and initiates silencing. A
major questions in the field of X inactivation is how XIST/Xist is regulated, becoming expressed
on the inactive X and silenced on the active X. Much of what we currently know about XIST/Xist
regulation comes from studies using mice, however, differences in conservation of the XIC and
expression patterns of the major mouse Xist regulator, Tsix, indicate that humans and mice may
regulate XIST/Xist differently. The objective of this thesis was to identify regulatory elements
that are important for regulation of XIST in humans.
Since regulatory elements controlling XIST are believed to reside within the XIC, we
searched the XIC and identified two inactive X specific regulatory elements within the 5’ end of
XIST using DNase I hypersensitivity mapping. We found one of the hypersensitive sites to be
acting as an alternative P2 promoter for XIST which contains an upstream antisense promoter,
P2as. The second hypersensitive site was associated with alternative splicing and inclusion of
two novel exons for XIST. Interestingly, both P2 and the novel alternative splicing result in
transcripts that lack functional domains of XIST. An additional candidate regulator is the region
3’ of XIST due to the importance of Tsix in mice. We found that transcription 3’ of XIST in
somatic cells is low level sense transcription so we believe this to be leaky XIST rather than
TSIX. In human embryonic stem cells we found an antisense transcript that extends the full
length of XIST providing the first evidence for mouse-like TSIX in humans but very low-levels of
this transcript argue against regulatory ability. Taken together, our results highlight the
differences between mouse and human X inactivation and indicate that XIST transcription is
more complex than previously thought, generating XIST molecules that lack functional domains.
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Grubisic, Ivan. "Evaluating Transcriptional Regulation Through Multiple Lenses". Thesis, University of California, Berkeley, 2014. http://pqdtopen.proquest.com/#viewpdf?dispub=3640448.
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Scientific research, especially within the space of translational research is becoming increasingly multidisciplinary. With the development of each new method there is not only a need for a broad fundamental understanding of all the sciences and mathematics, but also an acute awareness of how errors propagate across methods, the limitation of the methods and what contextual frameworks need to be used for the interpretation. The ability to understand transcriptional mechanisms and the affect that subtle changes in equilibrium may have on cell fate decisions has been greatly advanced by next generation sequencing and subsequent tools that have been developed. Bioinformatic techniques can serve multiple roles. They fundamentally provide a global picture of what is happening within an experimental condition which can then be used to either confirm individual experimental findings as globally relevant, or to discover new insights to inform the next iteration of experiments. Many of the experiments are done in in vitro conditions and therefore I have also focused energy on trying to understand how the mechanical inputs, largely not representative of what is occurring in vivo, from these methods affect transcriptional regulation. Much of this research requires the switching of frameworks to understand how results from disparate data sources can be correlated. I then applied a similar thought process to the development of Lens. Without an effective means of communicating research findings in an elegant and streamline mannered, we are slowing down the ability for researchers to learn new frame- works to efficiently approach the next research questions. In addition to better methods of communicating, we also need more modular and simplified tools that can be applied to various experimental systems to increase the speed and efficiency of translational research.
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Smirlis, Despina. "Transcriptional regulation of rat CYP2B genes". Thesis, University College London (University of London), 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.271548.
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Abson, Nicole Claire. "Transcriptional regulation in developing sensory neurones". Thesis, University College London (University of London), 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.286308.
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Li, I. M. "Transcriptional regulation of the aggrecan gene". Thesis, University of Liverpool, 2016. http://livrepository.liverpool.ac.uk/3004940/.
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Aggrecan is a large- aggregating proteoglycan that is essential for the function of articular cartilage. A loss of aggrecan is one of the major event in osteoarthritis, a debilitating and degenerative joint disease. To understand how a gene is lost and what fails to prevent transcription in disease state, an understanding of the bases of transcription must first be sought. Transcription is the first step in gene regulation and the chromatin plays an important role in blocking or allowing transcription to occur. Enhancers are non-coding DNA sequences that allow the binding of proteins, such as transcription factors that drive transcription irrespective of distance or orientation. Enhancers are marked by histone modifications that distinguish them from promoters and divide them into poised or active and are generally highly evolutionary conserved. Using publically available data on histone modifications found on ENCODE and transgenic mice this study has added to the understanding of the transcriptional regulation of aggrecan along with the known enhancer at -10kb. 3 intergenic regions, -35kb, -65kb and -87kb and one intronic (+26kb) from the transcription start site of aggrecan expresses primarily in the chondrocytes at E15.5. The -87kb enhancer marks chondrocytes that commit to an articular cartilage fate, and is strongly active in adult mice. The -65kb appears developmentally active, marking hypertrophic chondrocytes. The -35kb and +26kb are expressed in all chondrocytes at E15.5 mice and the -35kb is active in adult tissue. These enhancers bind to Sox9 and a loss of Sox9 in the -35kb enhancer shifts the expression from chondrocytes to fibroblast or perichondrium cells. These enhancers may play a role in response to diseases such as OA, as when these enhancers are transfected into differentiated murine ADTC5 cells and treated with Il-1β or hOSM there is a reduced level of expression from the -87kb element and an increase expression of the -65kb in response to Il-1β. A highly conserved intronic sequences (+55kb) was identified that does not express in chondrocytes, rather in kidney and lungs and may play a role outside of aggrecan gene regulation. Using the Acan promoter and the -35kb enhancer a cartilage-specific Cre recombinase line was generated providing a tool to explore the pathogenesis of cartilage genes in disease such as OA or in lineage tracing.
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Lo, Kin Yui Alice. "Transcriptional regulation of adipose insulin resistance". Thesis, Massachusetts Institute of Technology, 2012. http://hdl.handle.net/1721.1/71466.
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Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biological Engineering, 2012.Page 168 blank. Cataloged from PDF version of thesis.
Includes bibliographical references (p. 155-167).
Insulin resistance is a condition that underlies type 2 diabetes and various cardiovascular diseases. It is highly associated with obesity, making it a pressing medical problem in face of the obesity epidemic. The obesity association also makes adipose tissue the target of interest for ongoing research. Previous work on adipose insulin resistance has largely been focused on deciphering the signaling defects and abnormal adipokine secretion profiles. There is increasing awareness that transcriptional control is a source of dysregulation as well as an avenue of therapeutic intervention for insulin resistance. However, knowledge of transcriptional regulation and dysregulation of adipose insulin resistance remains fragmentary. Here, we present a genome-wide perspective on transcriptional regulation of adipocyte biology and adipose insulin resistance. We made use of the latest high-throughput sequencing technology to interrogate different aspects of transcriptional regulation, namely, histone modifications, protein-DNA interactions, and chromatin accessibility in adipocytes. In combination with the transcriptional outcomes measured by microarray and RNA-sequencing, we (1) characterized a largely unknown histone modification, H3K56 acetylation, in human adipocytes, and (2) set up four diverse in vitro insulin resistance models in mouse adipocytes and analyzed them in parallel with mouse adipose tissues from diet-induced obese mice. In both cases, through computational analysis of the experimentally identified cis-regulatory regions, we identified existing and novel trans-regulators responsible for adipose transcriptional regulation. Furthermore, by comprehensive pathway analysis of the in vitro models and mouse models, we identified aspects of in vivo adipose insulin resistance that are captured by the different in vitro models. Taken together, our studies present a systems view on adipose transcriptional regulation, which provides a wealth of novel resources for gaining insights into adipose biology and insulin resistance.
by Kin Yui Alice Lo.
Ph.D.
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Jani, Meghna. "Transcriptional regulation of LAMB3 by p53". Wright State University / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=wright1229958362.
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Palencia, Desai Sharina. "Transcriptional Regulation of Early Endocardial Development". University of Cincinnati / OhioLINK, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1378195299.
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Cockman, Eric Michael. "Post-Transcriptional Regulation of Selenoprotein S". Case Western Reserve University School of Graduate Studies / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=case1562593531805034.
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Ng, Tien Haeng Sky. "Mechanisms of IL-10 transcriptional regulation". Thesis, University of Bristol, 2015. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.702135.
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Previous work has shown that subcutaneous (s.c) specific immunotherapy (SIT) using the myelin protein peptide MBP Acl-9 [4Y] induces protection against EAE in the TCR transgenic Tg4 mice. Repeated administration of MBP Ac1-9 [4Y] was found to induce IL-1O expressing TH1 like cells which were termed IL-l0 Tregs. To further the understanding of how IL-1O transcription is regulated, the expression of IFN-y, IL-4, IL-1O, c-Maf, NFIL3 protein and mRNA, Gata3 mRNA and epigenetic modifications at the Il10 promoter were characterised for each dose of s.c MBP Ac 1-9 [4 Y] administration. The analysis revealed that c-Maf and NFIL3 positively correlated with Il-IO expression, whilst levels of H3K27me3 at the Il10 promoter, an epigenetic mark associated with transcriptional repression, correlated inversely with IL-IO expression. To investigate the therapeutic potential of non-antigen specific immunoregulatory small molecules, GSK3 inhibitors were co-cultured with murine TH1, TH2 and human memory T cells. The analysis shows that GSK3 inhibitors also induce IL-10 expression via transcriptional mechanisms in both murine Th cells and human memory T cells. Similar to IL-l0 Tregs, murine TH1 GSK3 inhibitor treated cells expressed higher levels of NFIL3 and c-Maf whilst murine TH2 GSK3 inhibitor treated cells expressed higher levels of NFIL3 only. In contrast to IL-10 Tregs both THI and TH2 cells expressed higher levels of Gata3 in response to GSK3 inhibitor treatment. In addition, epigenetic changes also take place in the Il10 promoter of GSK3 inhibitor treated murine TH1 and human memory T cells. Collectively these results show that it is possible to alter epigenetic modifications and induce IL-1O expression in murine TH1 and TH1-like cells with in vivo and in vitro methods. Furthermore, we propose combining the antigen specific s.c SIT with GSK3 inhibitors to improve the efficacy and safety of both treatments.
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Meadows, Stryder. "Transcriptional Regulation In Early Embryonic Development". Diss., The University of Arizona, 2008. http://hdl.handle.net/10150/194034.
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Transcription factors are a class of proteins that function to regulate the expression of genes. During early development, their role is to provide the precise order of activation or suppression of genes that are required for the formation of an embryo. A major goal of a developing embryo is to establish a complete body plan that includes the development of all of the organ systems. Thus it is paramount that the correct genes are switched on or off to insure that all organ systems form.Our studies investigate the role of several transcription factors involved in coordinating the expression of genes that are essential for the development of skeletal muscle and blood vessels.In the formation of skeletal muscle, a class of transcription factors called the myogenic regulatory factors (MRFs) is known to promote the induction of the structural genes that comprise the skeletal muscle. In fact, the MRF family member, MyoD, has been termed the "master regulator" of skeletal muscle gene expression. However, a recently discovered transcription factor, MASTR, has been suggested to play a role in skeletal muscle development. Our studies of MASTR are the first to demonstrate that, in vivo, MASTR is necessary and sufficient to activate genes involved in the formation of skeletal muscle. Furthermore, MASTR cooperates with MRFs to induce skeletal muscle genes and therefore places MASTR among a group of transcription factors, such as the MRFs, that are essential regulators of skeletal muscle development.In vascular development, the Flk-1 gene is critical to the formation of blood vessels. Mice lacking Flk-1 do not produce angioblasts, the precursor cells that give rise to the endothelial cells that make up blood vessels. In our efforts to understand the regulation of this important vascular gene, we have discovered a new function of the Kruppel-like transcription factor 2 (KLF2) to activate Flk-1 expression. Moreover, we have identified a new Ets transcription factor (Etsrp) capable of inducing Flk-1 expression alone and in cooperation with KLF2. These findings uncover a novel mechanism by which KLF2 and Etsrp act to promote the expression of Flk-1 during embryonic vascular development.
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MORALES, A. ZULUETA. "TRANSCRIPTIONAL REGULATION OF THE B3GALT5 GENE". Doctoral thesis, Università degli Studi di Milano, 2014. http://hdl.handle.net/2434/231099.
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RIASSUNTO
Introduzione. La β1,3 galattosiltransferasi (B3GALT5) è responsabile della sintesi della catena oligosaccaridica di tipo 1, tra cui gli antigeni Lewis come il sialil-Lewis a, epitope del marcatore tumorale CA19.9 e ligando della E-selectina, potenzialmente coinvolto nella malignità tumorale. La sua trascrizione è regolata da molteplici promotori. In alcuni epiteli essa è sotto il controllo di un promotore debole, chiamato nativo, modulato epigeneticamente e tramite il fattore nucleare NF-Y. In alcuni organi e cellule di origine gastrointestinale è attivo inoltre un altro promotore, più forte e chiamato LTR per la sua origine retrovirale, che secondo la letteratura dovrebbe essere regolato attraverso il fattore nucleare epatocitario HNF1α/β e quello analogo al Caudale di drosofila Cdx1/2. Tuttavia la B3GALT5 è repressa nel cancro del colon, il trascritto LTR non è rilevante nell’intestino tenue, e Cdx1/2 risultano assenti in una linea cellulare che esprime grandi quantità di tale trascritto.
Scopi. Scoprire i meccanismi che controllano la trascrizione di B3GALT5 attraverso il suo promotore LTR, con l’obiettivo di spiegare la specificità tissutale e la repressione negli adenocarcinomi del colon, nonché di capire il processo di stabilizzazione evolutiva del trasposone in alcuni primati.
Metodi. A questo scopo abbiamo quantificato l’espressione di HNF1α/β e Cdx1/2, tramite Western Blot, e quella del trascritto B3GALT5 LTR, tramite RT-PCR competitiva, in tessuti tumorali e linee cellulari. Inoltre, abbiamo silenziato HNF1α o β in diverse cellule, tramite shRNA, e li abbiamo espressi in un’altra, tramite trasfezione del cDNA. Abbiamo quindi trattato delle cellule con l'agente demetilante il DNA 5'-AZA-2'-desossicitidina, misurandone poi i livelli di trascritto LTR. Abbiamo infine valutato il promotore LTR in vitro, mediante saggi di EMSA e di luciferasi. Risultati. Cdx1/2 risultano immisurabili in cellule e tessuti che pur esprimono alti livelli di trascritto LTR, mentre HNF1α/β sono presenti anche in cellule e tumori che esprimono una quantità bassa o nulla del trascritto. Nelle cellule che non esprimono HNF1α/β però non c’è espressione alcuna del trascritto LTR. Tra queste, le MDA-MB-231, dopo transfezione con HNF1α o β, esprimono il trascritto LTR, ma a bassi livelli, paragonabili a quelli dei tumori del colon. Il silenziamento di HNF1α in una linea cellulare che esprime entrambi i fattori HNF1α e β non ha effetti nell’espressione del trascritto LTR, mentre quello di HNF1β in un’altra linea che esprime solo HNF1β produce forte riduzione del trascritto. Pure il trattamento con 5'-AZA-2'-desossicitidina riduce il trascritto in cellule che ne esprimono alti livelli, portandolo ai livelli dei tumori del colon, e senza effetto sulla quantità di HNF1. Usando i saggi delle luciferasi, abbiamo visto che la luciferasi sotto il controllo del promotore LTR è più attiva in cellule e cloni che esprimono alte quantità di HNF1, anche se non esprimono o esprimono poco trascritto LTR, che in quelle cellule che esprimono poco HNF1 ma magari una grande quantità di trascritto. Usando la sequenza del promotore LTR in saggi di EMSA, abbiamo visto che forma dei complessi specifici con estratti di proteine nucleari di tutte le linee cellulari che esprimono HNF1, indipendentemente dei livelli di espressione del trascritto B3GALT5 LTR.
Conclusione. I nostri risultati suggeriscono che HNF1α e -β sono necessari ma non sufficienti a regolare l'espressione del promotore LTR, mentre Cdx1/2 non sono coinvolti. HNF1α/β svolgono un ruolo intercambiabile e non cumulativo, e non sono immediatamente responsabili della regolazione negativa che avviene nel cancro, che invece dipende da elementi regolatori distanti attivi solo se metilati. Il successo della inserzione e l'attivazione del promotore B3GALT5 LTR durante l'evoluzione dipendono quindi non solo dal suo sito di legame a HNF1, ma anche da questi elementi distanti attualmente sconosciuti.ABSTRACT Background. β1,3 galactosyltransferase (B3GALT5) is responsible for the synthesis of type 1 chain oligosaccharides, including Lewis antigens as sialyl-Lewis a, the epitope of tumor marker CA19.9 and an E-selectin ligand potentially involved in cancer malignancy. Transcription occurs through multiple promoters. In some epithelia it is driven by a weak promoter, known as the native promoter that is epigenetically modulated and sensitive to nuclear factor NF-Y. In some organs of the gastrointestinal tract (as the colon, stomach, pancreas and related cell lines) another stronger promoter is active and named the LTR promoter after its retroviral origin. It was supposed to be regulated through a set of homeoproteins: hepatocyte nuclear factor HNF1α/β and caudal-related homeobox Cdx1/2. Surprisingly, B3GALT5 is strongly down regulated in colon cancer, the LTR transcript is not relevant in the small intestine, and Cdx1/2 were reported absent from a cell line expressing large amount of such transcript. Aims. To elucidate the mechanisms controlling transcription of B3GALT5 through its retroviral LTR promoter, in order to explain the tissue specificity and down-regulation in colon adenocarcinomas, and to understand the evolutionary stabilization of the transposon in some primates. Methods. To this aim, we determined the expression levels of putative transcription factors by western blot and the amounts of B3GALT5 LTR transcript by competitive RT-PCR in cancer tissues and cell lines. Moreover, we silenced HNF1α or β in different cell lines, through an shRNA approach, expressed them in another by permanent cDNA transfection, and treated cells with the DNA demethylating agent 5’-AZA-2’-deoxycitydine and in all cases, we measure the effects on LTR transcript levels. We also evaluated the behavior of the LTR promoter in vitro, through electrophoresis mobility shift and reporter luciferase assays. Results. We found that Cdx1/2 are not detectable in cells and tissues expressing high amount of B3GALT5 LTR transcript, while HNF1α/β are well detectable, but even in cells and cancers expressing very low or undetectable levels of the transcript, which is absent in all cells lacking HNF1α/β. Among them, the cell line MDA-MB-231, upon transfection with HNF1α or β, became able to express B3GALT5 LTR transcript, but a very low levels, similar to those found in colon cancers. Transient silencing of HNF1α in cells expressing both HNF1α and β, has no effect on LTR transcript, while similar silencing of HNF1β in cells expressing HNF1β only, determines strong reduction of the transcript. Cell lines expressing high levels of B3GALT5 LTR transcript are affected by the demethylating agent 5AZA that determines strong down regulation of the transcript, falling down to the amounts found in colon cancers, while HNF1 levels remain unaffected. In vitro, luciferase placed under the control of LTR promoter is more active in cells or clones expressing high HNF1 and low or no LTR transcript than in those expressing low HNF1 and high transcript. The same promoter, when used as a probe in EMSA, forms specific complex with nuclear protein extracted from all cells expressing HNF1, irrespectively of the levels of B3GALT5 LTR transcript. Conclusion. Our results suggest that HNF1α and HNF1β are necessary but not sufficient to drive expression of LTR promoter, while Cdx1/2 are not involved. HNF1α/β play an interchangeable and not cumulative role and are not immediately responsible for cancer down-regulation, which depends on a distal regulatory element(s) active when methylated. The successful insertion and activation of B3GALT5 LTR promoter during evolution depended not only on its HNF1 binding site, but even on such distal element(s) unknown at present.
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Ruiz, Carlos Ariel. "Transcriptional and Post-Transcriptional Regulation of Synaptic Acetylcholinesterase in Skeletal Muscle". Scholarly Repository, 2009. http://scholarlyrepository.miami.edu/oa_dissertations/370.
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myotubesProper muscle function depends upon the fine tuning of the different molecular components of the neuromuscular junction (NMJ). Synaptic acetylcholinesterase (AChE) is responsible for rapidly terminating neurotransmission. Neuroscientists in the field have elucidated many aspects of synaptic AChE structure, function, and localization during the last 75 years. Nevertheless, how the enzyme is regulated and targeted to the NMJ is not completely understood. In skeletal muscle the synaptic AChE form derives from two separate genes encoding the catalytic and the collagenic tail (ColQ) subunits respectively. ColQ-AChE expression is regulated by muscle activity; however, how this regulation takes place remains poorly understood. We found that over or down-regulation of ColQ is sufficient to change the levels of AChE activity by promoting assembly of higher order oligomeric forms including the collagen-tailed forms. Furthermore, when peptides containing the Proline Rich Attachment Domain (PRAD), the region of ColQ that interacts with the AChE, are fed to muscle cells or cell lines expressing AChE, they are taken up by the cells and retrogradely transported to the endoplasmic reticulum (ER)/Golgi network where they induce assembly of newly synthesize AChE into tetramers. This results in an increase, as a consequence, in total cell associated AChE activity and active tetramer secretion, making synthetic PRAD peptides potential candidates for the treatment of organophosphate pesticides and nerve gas poisoning. To study the developmental regulation of ColQ-AChE we determined the levels of ColQ and ColQ mRNA in primary quail muscle cells in culture and as a function of muscle activity. Surprisingly, we found dissociation between transcription and translation of ColQ from its assembly into ColQ-AChE indicating the importance of posttranslational controls in the regulation of AChE folding and assembly. Furthermore, we found that the vast majority of the ColQ molecules in QMCs are not assembled into ColQ-AChE, suggesting that they can have alternative function(s). Finally, we found that the levels of ER molecular chaperones calnexin, calreticulin, and particularly protein disulfide isomerase are regulated by muscle activity and they correlate with the levels of ColQ-AChE. More importantly, our results suggest that newly synthesized proteins compete for chaperone assistance during the folding process.
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Begbie, Megan Elaine. "Transcriptional and post-transcriptional regulation of the human factor VIII gene". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0021/NQ45262.pdf.
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Liu, Jun-Li. "Transcriptional and post-transcriptional regulation of somatostatin gene expression by glucocorticoids". Thesis, McGill University, 1995. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=28826.
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Glucocorticoids and somatostatin both influence a broad spectrum of biological activities and their actions are cooperative in growth control, pancreatic islet function, immune suppression, and stress response, e.g. in vivo studies indicate that glucocorticoids may act through somatostatin to suppress growth, growth hormone secretion and inflammation. Recent studies have suggested that glucocorticoids influence somatostatin production but the precise nature of this effect has remained unclear. In this thesis, I characterized the actions of glucocorticoids on somatostatin gene expression and their molecular mechanisms of action in three consecutive studies. (1) I started with an investigation of the in vivo and in vitro effects of glucocorticoids and found that dexamethasone exerts significant effects on somatostatin peptide and steady state mRNA levels in normal somatostatinoma (1027B$ sb2$) cells. Glucocorticoids stimulate somatostatin production in peripheral tissues (stomach, pancreas, and jejunum) and suppress its biosynthesis in cerebral cortex and hypothalamus. Glucocorticoids induce dose-dependent biphasic effects on steady state somatostatin-mRNA levels in normal rat islet and 1027B$ sb2$ cells, characterized by stimulation at low doses (10$ sp{-10}$ M) and marked inhibition at high doses ($ geq$10$ sp{-7}$ M). This suggests a complex molecular mechanisms of glucocorticoid action on the somatostatin gene involving multi-level regulation. (2) I further discovered that glucocorticoids stimulate somatostatin gene transcription in PC12 (pheochromocytoma) cells transfected with somatostatin promoter-CAT (chloramphenicol acetyl transferase) reporter gene. Dexamethasone induces a dose-dependent 2.2 fold stimulation of somatostatin-CAT expression in PC12 cells and exerts an additive effect on cAMP-induced gene transcription. The dexamethasone effect is abolished in A126-1B2 (protein kinase A-deficient mutant PC12) cells and with CRE (cAMP response element) mutant
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Toedling, Joern Michael. "Comprehensive analysis of high-throughput experiments for investigating transcription and transcriptional regulation". Thesis, University of Cambridge, 2009. https://www.repository.cam.ac.uk/handle/1810/267885.
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As the number of fully sequenced genomes grows, efforts are shifted towards investigation of functional aspects. One research focus is the transcriptome, the set of all transcribed genomic features. We aspire to understand what features constitute the transcriptome, in which context these are transcribed and how their transcription is regulated. Studies that aim to answer these questions frequently make use of high-throughput technologies that allow for investigation of multiple genomic regions, or transcribed copies of genomic regions, in parallel. In this dissertation, I present three high-throughput studies I have been involved in, in which data gained from oligo-nucleotide tiling microarrays or large-scale cDNA sequencing provided insights into the transcriptome and transcriptional regulation in the model organisms Saccharomyces cerevisiae and Mus musculus. Interpretation of such high-throughput data poses two major computational tasks. The primary statistical analysis includes quality assessment, data normalisation and identification of significantly affected targets, i.e. regions of the genome deemed transcribed or involved in transcriptional regulation. Second, in an integrative bioinformatic analysis, the identified targets need to be interpreted in context of the current genome annotation and related experimental results. I provide details of these individual steps as they were conducted in the three studies. For both primary and integrative analysis, functional, extensible and welldocumented software is required, which implements individual analysis steps, allows for concise visualisation of intermittent and final results and facilitates the construction of automated, programmed workflows. Ideally such software is optimised with respect to scalability, reproducibility and methodical scope of the analyses. This dissertation contains details of two such software packages in the Bioconductor project, which I (co-)developed.
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Bhatia, Deepak. "Transcriptional and post-transcriptional regulation of Gadd45[alpha] in response to arsenic". Morgantown, W. Va. : [West Virginia University Libraries], 2007. https://eidr.wvu.edu/etd/documentdata.eTD?documentid=5311.
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Thesis (Ph. D.)--West Virginia University, 2007.Title from document title page. Document formatted into pages; contains xv, 156 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references (p. 112-153).
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POURPIRALI, SOMAYEH. "Transcriptional and post-transcriptional regulation of Ambra1 in physiological and pathological conditions". Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2012. http://hdl.handle.net/2108/202211.
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(Macro)autophagy is the process of self-digestion of cellular compartments through lysosomal pathway. Ambra1 is one of the genes involved in this process which is regulated at both transcriptional and post-translational levels. Considering the involvement of Ambra1 in various disease conditions, it is of great significance to identify the factors involved in its regulation. In this study we analyzed Amba1 promoter and identified two fragments with inducer and repressor activities. We also analyzed the effect of different stressors such as starvation, DNA damage, and hypoxia on Ambra1. Starvation could upregulate Ambra1 both at RNA and protein levels. Induction of DNA damage by etoposide resulted in an increase in Ambra1 protein levels without affecting RNA levels. The stressor which was studied most extensively was hypoxia induced by hypoxia mimetic, CoCl2. Interestingly, while some autophagy genes were up-regulated upon treatment, a decrease in amount of Ambra1 protein levels was detected. In the meantime, qRT-PCR analysis showed only a slight increase in Amrba1 mRNA levels at the early stages of hypoxia, but no significant changes were observed at the later time points. Long-term stress such as starvation or staurosporin treatment leads to Ambra1 protein degradation through caspase and calpain cleavage, therefore inhibiting the pro-survival role of autophagic machinery. After CoCl2 treatment for 18 hours, caspase 3 was cleaved, this showing the activation of apoptotic machinery. Still, the inhibition of caspases along with hypoxia did not completely restore the levels of Ambra1 protein, indicating that other mechanisms are possibly involved. This led us to investigate the translation rate of Ambra1 mRNA. We observed translocation of Ambra1 mRNA from polysomes to non-translating fractions. Besides, we analyzed the mRNA localization in the cytoplasm. Interestingly, Ambra1 mRNA was present in distinct foci in the cell under normal conditions while CoCl2 treatment resulted in more intense and larger foci. Co-detection of mRNA and protein experiments revealed the re-localization of Ambra1 to cytoplasmic P-bodies which results in translational repression. This data suggests a possible new mechanism regulating autophagy. Still, further experiments such as RNA-IP and localization of the mRNA of other autophagy genes are required to confirm this hypothesis.
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Yin, Shiyi. "Transcriptional Regulation of CFTR in the Intestinal Epithelium". Case Western Reserve University School of Graduate Studies / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=case1625503766675073.
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SICILIANO, DILETTA. "ANALYSIS OF THE TRANSCRIPTIONAL REGULATION OF MTORC1 ACTIVITY BY MIT/TFE TRANSCRIPTION FACTORS". Doctoral thesis, Università degli Studi di Milano, 2019. http://hdl.handle.net/2434/607642.
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The mechanistic Target Of Rapamycin Complex 1 (mTORC1) regulates cellular biosynthetic pathways in response to variations in nutrient availability. Activation of mTORC1 is mediated by Rag GTPases, that act as heterodimers and promote mTORC1 recruitment to the lysosome. Many studies have clarified the post-translational control of mTORC1, but little is known about its transcriptional regulation. Our study demonstrates that TFEB, TFE3 and MITF, members of the MiT/TFE family of transcription factors and master regulators of lysosomal and melanosomal biogenesis and autophagy, are nutrient-sensitive transcriptional activators of mTORC1 signaling. During starvation they induce the expression of the RagD gene and this enhances mTORC1 recruitment to the lysosome and its reactivation when nutrients become available. Thus, in periods of nutrient deprivation, this mechanism allows the cell to rapidly reactivate anabolic pathways and turn off catabolism when nutrient levels are restored. Furthermore this mechanism plays an important role in cancer growth. Up-regulation of the MiT/TFE genes in renal cell carcinoma and melanoma is associated to RagD-induced mTORC1 activation, causing cell hyperproliferation and cancer progression.
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Sasmono, R. Tedjo. "Transcriptional regulation of c-fms gene expression /". [St. Lucia, Qld.], 2003. http://www.library.uq.edu.au/pdfserve.php?image=thesisabs/absthe17479.pdf.
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Xiao, Lei. "Transcriptional Regulation of the Xenopus MyoD Gene". Diss., lmu, 2003. http://nbn-resolving.de/urn:nbn:de:bvb:19-11960.
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Zadissa, Amonida, i n/a. "Inferring transcriptional regulation in mammals using bioinformatics". University of Otago. Department of Biochemistry, 2007. http://adt.otago.ac.nz./public/adt-NZDU20070614.111340.
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Gene expression and its regulation is a highly coordinated system, involved in many biological processes such as cell growth, division and differentiation. Transcriptional regions, involved in gene regulation, consist of a heterogeneous collection of smaller regulatory elements. In some cases, co-regulated genes contain a common set of transcription factor binding sites (TFBS).
Analysis of promoter regions is the major approach in understanding the transcriptional regulatory mechanisms. It is also useful for interpretation of mammalian gene expression studies, where co-expressed genes may share motifs representing putative TFBS. Motif identification also has the advantage that it can predict control regions in genes that have not been measured experimentally. However, a common problem is incomplete genomic sequence for the experimental species of interest. The approach here is to identify and use orthologous gene promoter sequences from a related and well-characterised species.
The primary aim of this study was to identify and predict regulatory TFBS in species where promoter sequence does not exist or is incomplete. The MEME programme was employed for the motif prediction step. The predicted elements were subsequently compared to known TFBS using TRANSFAC and JASPAR databases for identification. A methodology based on relative entropy was used. The validity of the method was confirmed as the predicted motifs in the training set were the expected sites involved in regulation of muscle development. The technique was applied to two data sets, generated from expressed sequence tag (EST) clustering analysis and microarray experiments. All data sets, software and results are available on the accompanying CD.
Bovine expression data was analysed for cardiac-specific expression using two separate approaches, combining bovine library EST frequency and human gene expression ratios. For each approach, the orthologous human and bovine promoter sequences were analysed for common motifs. Across all comparisons, 37% of motifs were identified as known TFBS using the TRANSFAC and JASPAR databases. As the human comparison had more promoter sequences available, this was the main limiting factor for the corresponding bovine analysis, rather than cross-species divergence or accuracy of gene expression measurement. Results from this study demonstrate that using promoter sequences from a related species is a viable approach when studying gene expression in species with limited amount of genomic sequence. As the bovine genome becomes more complete, it can in turn serve as the reference genome for other agriculturally important ruminants, such as sheep, goat and deer.
The second application concerned in silico analysis of gene regulation patterns in response to stimuli. Recently it has been shown that a mutation in the bone morphogenetic receptor IB leads to an increased ovulation rate in sheep. The objective of this study was to analyse gene expression patterns in cultured cells in response to four members of the BMP family, i.e. BMP2, BMP4, BMP6 and BMP7 and the control TGFβ. Microarray data was provided by J. Young. Twelve highly upregulated genes were stimulated by all BMPs, seven of which are known BMP target genes. Analysis of the predicted motifs identified four elements that may be involved in the regulation process. Cross-species comparison for one of the genes, ID1, showed high conservation of one of the motifs across 11 mammalian genomes. This particular motif had not been identified as a known binding site. In summary, the analysis of the expression data suggest an extension of the list of BMP targets.
The proposed method is relatively robust when sufficiently co-expressed (co-regulated) sequences can be identified, whether from the same or another species.
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Stramwasser, Melissa P. "Investigations into the transcriptional regulation of BRCA1". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ37982.pdf.
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