Tesis sobre el tema "GPCR function and regulation"
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Dromey, Jasmin Rachel. "Elucidating novel aspects of hypothalamic releasing hormone receptor regulation". University of Western Australia. School of Medicine and Pharmacology, 2008. http://theses.library.uwa.edu.au/adt-WU2008.0133.
Texto completoJama, Abdirahman Mohamud. "Functional regulation of kisspeptin receptor by calmodulin and Ca2+/calmodulin-dependent protein kinase II". Thesis, University of Edinburgh, 2015. http://hdl.handle.net/1842/15914.
Texto completoÅkerberg, Helena. "Functional Studies of the Neuropeptide Y System : Receptor-Ligand Interaction and Regulation of Food Intake". Doctoral thesis, Uppsala universitet, Institutionen för neurovetenskap, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-9533.
Texto completoRichardson, Kathryn. "Mechanisms of GPCR signal regulation in fission yeast". Thesis, University of Warwick, 2014. http://wrap.warwick.ac.uk/63554/.
Texto completoRanganathan, Anirudh. "The impact of GPCR structures on understanding receptor function and ligand binding". Doctoral thesis, Stockholms universitet, Institutionen för biokemi och biofysik, 2016. http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-129879.
Texto completoAt the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 3: Manuscript.
Hillier, Stephen Gilbert. "Regulation of ovarian function". Thesis, University of Edinburgh, 1992. http://hdl.handle.net/1842/26607.
Texto completoBittencourt, Fabiola M. "Examination of the Function of the Murine Cytomegalovirus Encoded G Protein-Coupled Receptor M33 in vivo". University of Cincinnati / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1397234044.
Texto completoMunjal, Akankshi. "Regulation of a bio-mechanical network driving shape changes during tissue morphogenesis". Thesis, Aix-Marseille, 2015. http://www.theses.fr/2015AIXM4038/document.
Texto completoForces required to power shape changes during tissue morphogenesis are generated by non-muscle MyosinII (MyoII) pulling filamentous actin. During my PhD, I investigated the role of MyoII regulation through the conserved Rho1-Rok pathway during Drosophila germband extension. The morphogenetic process is powered by cell intercalation involving shrinkage of junctions in the dorsal-ventral axis (‘vertical junctions’) followed by junction extension in the anterior-posterior axis. Advances in light microscopy revealed that the actomyosin networks exhibit pulsed contractions to power junction shrinkage, and alternate with steps of stabilization by MyoII enriched on vertical junctions (planar-polarity) to result in irreversible shape changes. Although described in many different contexts, the underlying mechanisms of this ratchet-like behavior remained unclear. Using genetic and biophysical tools, quantitative imaging and subtle perturbations, I identified 2 critical properties underlying MyoII dynamics- turnover governed by phospho-cycling of the MyoII Regulatory Light Chain, and advection due to contraction of the motors on actin networks. Spatial control over MyoII turnover establishes 2 stable regimes of high and low dissociation rates resulting in MyoII planar polarity. Pulsatility is a self-organized behavior that emerges at intermediate dissociation rates enabling advection of MyoII and upstream regulators. In the second part of my thesis, I showed that G protein coupled receptors- GRsmog and Mist, and the downstream G-protein pathway allow step-wise activation of MyoII, establishing pulsatility and stability, to drive polarized shape deformations during morphogenesis
Clay, L. "CDC20 function, regulation and proteolysis". Thesis, University of Cambridge, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597750.
Texto completoBrandao, Haga Raquel. "Function and regulation of RhoBTB1". Thesis, King's College London (University of London), 2016. https://kclpure.kcl.ac.uk/portal/en/theses/function-and-regulation-of-rhobtb1(0904ff24-d566-4987-8c61-440c09854eeb).html.
Texto completoBerry, David (David A. ). "Glycosaminoglycan regulation of cell function". Thesis, Massachusetts Institute of Technology, 2005. http://hdl.handle.net/1721.1/34153.
Texto completoIncludes bibliographical references (p. 252-285).
Glycosaminoglycans (GAGs) are complex polysaccharides that exist both on the cell surface and free within the extracellular matrix. The intrinsic sequence variety stemming from the large number of building blocks that compose this biopolymer leads to substantial information density as well as to the ability to regulate a wide variety of important biological processes. With the recent and progressive emergence of biochemical and analytical tools to probe GAG structure and function, efforts can be taken to understand the role of GAGs in cell biology and in disease in the various physiological locations where GAGs can exist. As a first step to probe the functions of GAGs, the heparin/heparan sulfate-GAG (HSGAG)-fibroblast growth factor (FGF) system was examined. Understanding the role of HSGAGs in inducing FGF2 dimerization led to the development of a novel engineered protein that was found to be effective at promoting functional recovery in stroke. Subsequently, methods to isolate HSGAGs from the cell surface were optimized and the ability of HSGAGs to support FGF signaling was investigated. Cell surface HSGAGs can define the responsiveness of a given cell to FGF1 and FGF2 through multiple receptor isoforms. Stromal cell derived HSGAGs were also identified as critical regulators of tumor cell growth and metastasis, effecting not only FGF2., but also 1-integrin signaling.
(cont.) Other GAGs, including dermatan sulfates, were characterized as modulators of FGFs and vascular endothelial growth factors. Finally, FGFs and HSGAGs were found to have important roles in maintaining epithelial monolayer integrity, with syndecan-l serving as a critical factor in inflammatory bowel disease. In addition to understanding HSGAGs in their normal physiological settings, techniques to internalize them were developed. Poly(3-amino ester)s were found to condense heparin and enable its endocytosis into cells. Internalized heparin is preferentially taken up by cancer cells, which often have a faster endocytic rate than non-transformed cells, and promotes apoptotic cell death. Internalized heparin can also be used as a tool to probe cell function. In Burkitt's lymphoma, poly(3-amino ester)-heparin conjugates served to identify cell surface HSGAGs as an important modulator of cell growth that can be harnessed to inhibit growth. Finally, studies that sought to broaden the scope of GAG biology were undertaken. Cell surface HSGA(:is were identified as mediators of vascular permeability. Furthermore a novel technique to immobilize GAGs was employed. The interactions between GAG and substrate were via hydrogen bonding. Immobilization of GAGs alters their properties, such that they can affect cells in ways distinct from GAGs free in the ECM.
(cont.) Furthermore, immobilized GAGs can regulate cancer cell adhesion, growth and progression, and may offer a new way to regulate the activity of cancer cells. In addition to directly providing new potential therapeutics and drug targets, these studies represent a foundation to enable additional studies of GAG function. Future work harnessing the techniques presented may open new avenues of research and facilitate the development of novel GAG-based therapeutics.
by David Berry.
Ph.D.
Vespoli, Jessica L. "Genomic Regulation of Clock Function". Kent State University / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=kent1449500602.
Texto completoTran, Stella Lê Minh. "Foxl2 regulation and function in gonadotropes". Thesis, McGill University, 2013. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=116978.
Texto completoL'hormone folliculo-stimulante de l'hypophyse (FSH) est cruciale car elle régule gamétogenèse et fonction gonadique chez les mammifères. Une diminution dans la production de FSH peut mener à l'infertilité, mais les mécanismes contrôlant la synthèse de FSH demeurent incompris. Les activines de l'hypophyse stimulent la transcription du gène FSH sous-unité β (Fshb) dans les cellules gonadotropes; ceci est l'étape limitante dans la synthèse de l'hormone FSH. Les résults provenant de lignée gonadotrope immortalisée indiquent que la transcription de Fshb est stimulée par l'activine A; ceci est dépend des protéines homologues de Drosophila mothers against decapentaplegic (SMAD). Récemment, Forkhead box L2 (FOXL2) a été décrite comme étant un facteur-clé dans la stimulation de la transcription de Fshb induite par les SMADs et l'activine A. Ici, je dissèque les mécanismes expliquant la régulation et fonction de Foxl2 dans les cellules gonadotropes. Tout d'abord, la surexpression de FOXL2 et SMAD 2, 3, et 4 confère une réactivité à l'activine au promoteur du gène murin Fshb dans les cellules hétérologues. Sous stimulation par l'activine A, FOXL2 coopère de façon synergétique avec les SMADs pour activer le promoteur Fshb; cela nécessite que FOXL2 et SMAD3 (ou SMAD4) lient l'ADN. L'l'induction via SMAD3 du gène Fshb dépend de FOXL2 endogène dans les cellules homologues. Un élément proximal forkhead binding element (FBE) et un élément adjacent SMAD binding element (SBE) sont essentiels pour l'induction de l'activité du promoteur-reporter Fshb par FOXL2/SMAD3. Basé sur ces résultats, je propose un modèle où les activines stimulent la formation de complexes FOXL2-SMAD2/3/4 induisant la transcription du gène murinFshb via liaison à un élément SBE/FBE conservé du promoteur proximal. J'ai ensuite testé l'hypothèse que FOXL2 est nécessaire pour la synthèse de FSH in vivo en utilisant une approche Cre/lox: j'ai généré une souris knock-out conditionnelle (cKO) où Foxl2 est sélectivement absent dans les cellules gonadotropes. J'ai observé que les souris cKO sont hypogonadiques et souffrent d'une baisse de fertilité à l'âge adulte. J'ai démontré que la spermatogenèse et la folliculogénèse sont altérées chez les souris cKO. En effet, les cKO mâles ont un nombre diminué de spermatozoïdes, tandis que femelles cKO ovulent moins d'ovocytes pendant leur cycle oestral. J'ai démontré que cKO mâles et femelles sont déficientes en FSH, découlant d'une diminution d'ARN Fshb dans l'hypophyse. Les cultures primaires de cellules hypophysaires où Foxl2 est absent ne réagissent pas à l'activine A : Fshb est diminuée, tant au niveau basal que dans sa réponse au ligand. Ces résultats indiquent que l'expression de Foxl2 dans les gonadotropes est requise pour l'induction sélective de la production de FSH in vivo. Outre son rôle dans les gonadotropes, FOXL2 est également détecté dans les cellules thyrotropes de l'hypophyse, dans la paupière en développement et les cellules granuleuses de l'ovaire. Cependant les mécanismes qui régissent cette expression sélective n'ont pas encore été décrits. Afin d'étudier ce qui contrôle la transcription de Foxl2, j'ai cloné la région du promoteur murin de Foxl2. Ensuite, j'ai démontré une corrélation entre l'état de méthylation des CpGs du promoteur et l'expression des gènes, par lequel la méthylation exerce une inhibition sur le gène Foxl2 dans certaines cellules hétérologues. Mes résultats indiquent que Foxl2 n'est pas seulement contrôlé par la séquence de son promoteur et dévoilent un rôle possible pour la méthylation des CpGs dans la régulation de l'expression spécifique de Foxl2 chez la souris. En résumé, ma thèse définit un rôle nécessaire pour FOXL2 exprimé dans les gonadotropes pour la synthèse de Fshb et FSH, ainsi que la reproduction in vivo. Collectivement, mes recherches sur FOXL2 hypophysaire contribueront à identifier des causes de l'infertilité idiopathique ou encore trouver de nouvelles cibles pharmacologiques.
Cheng, C. W. "Regulation of endothelial and endometrial function". Thesis, University of Cambridge, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.597573.
Texto completoEnglish, Jane Louise. "Cellular regulation of matrix metalloproteinase function". Thesis, University of East Anglia, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.247107.
Texto completoRaghavan, Srikala. "Connectin function and regulation in Drosophila". Thesis, University of Cambridge, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.624670.
Texto completoBERCLAZ, PIERRE-YVES. "REGULATION OF ALVEOLAR MACROPHAGE IMMUNE FUNCTION". University of Cincinnati / OhioLINK, 2002. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1022869185.
Texto completoRoberts, Kate. "Regulation of neutrophil function by NAMPT". Thesis, University of Liverpool, 2012. http://livrepository.liverpool.ac.uk/8355/.
Texto completoTroupiotis-Tsaïlaki, Anastassia. "Lipid-GPCR interactions: from activation of sphingosine-1-phosphate receptors to modulation of vasopressin V2 receptor function". Doctoral thesis, Universite Libre de Bruxelles, 2015. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/216727.
Texto completoLes récepteurs couplés aux protéines G (GPCRs) forment la plus grande famille de protéines membranaires du génome humain et contribuent à une kyrielle de processus physiologiques essentiels, qui leur confèrent un intérêt pharmacologique majeur. Étudier l'interaction de ces protéines avec leurs ligands et leur environnement membranaire est primordial pour appréhender leur fonctionnement à l’échelle moléculaire. Bien que de remarquables avancées dans la détermination de structures à haute résolution de GPCRs à l'état inactif et actif aient permis de comprendre certaines bases structurales du fonctionnement des récepteurs, des approches complémentaires donnant un aperçu des aspects dynamiques et dans un environnement natif sont nécessaires pour cerner pleinement leur mécanisme d'activation. Notre travail de thèse s'inscrit dans cette problématique et s'articule autour de deux sujets: d'une part, comprendre quelles caractéristiques structurales du ligand sous-tendent l'activation de la famille des récepteurs au sphingosine-1-phosphate (S1P); d'autre part, déterminer si les lipides de la membrane plasmique modulent la structure et la fonction du récepteur à la vasopressine V2. Pour répondre à notre première question, nous avons étudié la réponse fonctionnelle en système cellulaire des récepteurs S1P1, S1P2, S1P4 et S1P5 à des composés synthétiques dérivés du S1P, portant des chaînes alkyles de longueur variable. Nos données mettent en évidence que la longueur de la chaîne hydrocarbonée du ligand est un paramètre crucial dans sa capacité d'induire l'activation du récepteur et ce pour l'ensemble des sous-types étudiés. De plus, nos résultats suggèrent que le comportement vis-à-vis de la longueur de chaîne dépend du sous-type de récepteur considéré. Nos résultats expérimentaux, combinés à une approche de modélisation dynamique, ont abouti à proposer un mécanisme d'activation pour la famille des récepteurs au S1P. Dans le second volet de notre travail, nous avons reconstitué le récepteur V2 purifié dans des systèmes de composition lipidique contrôlée, mimant la bicouche membranaire. Nous avons procédé à la caractérisation structurale et fonctionnelle du récepteur inséré dans différentes types de lipides, par des méthodes spectroscopiques infrarouge et de fluorescence. Les données obtenues suggèrent que la composition lipidique affecte la conformation et la fonction du récepteur. L'ensemble de nos travaux contribue ainsi à une meilleure compréhension du mécanisme d'activation des GPCRs et de leur régulation par l'environnement lipidique.
Doctorat en Sciences
info:eu-repo/semantics/nonPublished
Sheffler, Douglas James. "The Regulation of G Protein-Coupled Receptor (GPCR) Signal Transduction by p90 Ribosomal S6 Kinase 2 (RSK2)". Connect to text online, 2006. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=case1130777469.
Texto completoMukherjee, Abir. "ROLE OF LYSOPHOSPHATIDIC ACID IN REGULATION OF CANCER CELL METABOLISM". VCU Scholars Compass, 2012. http://scholarscompass.vcu.edu/etd/391.
Texto completoMaurel, Marion. "Les microARNs régulateurs de l’expression génique du Glypican-3 dans le Carcinome Hépatocellulaire". Thesis, Bordeaux 2, 2012. http://www.theses.fr/2012BOR21945/document.
Texto completoGlypican-3 (GPC3) is overexpressed in 72% of hepatocellular carcinoma (HCC). It is a co-receptor for WNT receptor and belongs to the heparan sulfate proteoglycans family. The general objective of my PhD thesis was to study the mechanisms by which GPC3 is post-transcriptionnally regulated in HCC. To this end, I developed a functional test that allowed me to screen a library of 876 human microRNAs. This led me to identify 5 microRNAs that regulate the expression of GPC3 mRNA through its 3’Untranslated Region (UTR). The work presented in this thesis particulary focuses on miR-1271 and miR-1291 as both microRNAs present a deregulated expression in HCC and are respectively inhibitor and activator of GPC3 mRNA expression. In a first project, I demonstrated that miR-1271 directly binds to GPC3 mRNA 3’UTR and affects its stability. This microRNA is underexpressed in HCC and its expression negatively correlates with that of GPC3 mRNA in a subgroup of HCC corresponding to those associated with hepatitis B virus infection. In a second project, I demonstrated that miR-1291 postively regulates the expression of GPC3 mRNA by targeting an intermediate factor. An in silico analysis led to the identification of the Inositol Requiring Enzyme 1 alpha (IRE1α) as a potential candidate. IRE1α is an endoplasmic reticulum (ER) resident type I transmembrane protein and contributes to the signaling of the Unfolded Protein Response (UPR). The UPR is an adaptive response activated upon accumulation of improperly folded proteins in the ER. I showed that IRE1α cleaves GPC3 mRNA through its endoribonuclease activity. Moreover I demonstrated that miR-1291 directly targets IRE1α mRNA through its 5’UTR, thereby decreasing its expression and contributing to GPC3 mRNA overexpression. MiR-1291 is overexpressed in HCC and its expression positively correlates with that of GPC3 mRNA. In summary, the work carried out during my PhD allowed the identification and the characterization of two new microRNAs (miR-1271 and miR-1291) that control the expression of GPC3 mRNA through direct or indirect mechanisms. The pathophysiological relevance of these regulatory mechanisms is in agreement with the respective expression levels of these microRNAs in HCC, which could therefore contribute to the overexpression of GPC3 in those tumors
Roux, Benoit Thomas. "Characterisation of the molecular mechanisms regulating the signalling and post-endocytic sorting of the receptors for calcitonin gene-related peptide and adrenomedullin". Thesis, University of Bath, 2013. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.604647.
Texto completoRunne, Caitlin M. "Function and Activation Mechanism of PLEKHG2, A Novel G Beta Gamma-Activated RhoGEF in Leukemia Cells". Diss., University of Iowa, 2013. https://ir.uiowa.edu/etd/4907.
Texto completoJelic, Marko. "The function of Toc34 and its regulation". Diss., lmu, 2003. http://nbn-resolving.de/urn:nbn:de:bvb:19-14213.
Texto completoLi, Cathy Shije 1974. "Function and regulation of histone deacetylase 4". Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=98750.
Texto completoJelić, Marko. "The function of Toc34 and its regulation". [S.l.] : [s.n.], 2003. http://edoc.ub.uni-muenchen.de/archive/00001421.
Texto completoWu, Qun Garris Paul A. "Function and regulation of the dopamine transporter". Normal, Ill. Illinois State University, 1999. http://wwwlib.umi.com/cr/ilstu/fullcit?p9960430.
Texto completoTitle from title page screen, viewed July 31, 2006. Dissertation Committee: Paul A. Garris (chair), Maarten E.A. Reith, Anthony J. Otsuka, Robert L. Preston, David L. Williams. Includes bibliographical references (leaves 221-242) and abstract. Also available in print.
Wang, Chunbo. "Structure, function and regulation of TRP channels". Columbus, Ohio : Ohio State University, 2006. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1148676549.
Texto completoPyszniak, Andrew M. "Regulation of LFA-1 (CD11a/CD18) function". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq25139.pdf.
Texto completoLindebro, Maria. "Mechanisms of regulation of dioxin receptor function /". Stockholm, 2002. http://diss.kib.ki.se/2002/91-7349-231-0/.
Texto completoBahram, Fuad. "Post-translational regulation of Myc oncoprotein function /". Uppsala : Dept. of Plant Biology and Forest Genetics, Swedish Univ. of Agricultural Sciences, 2004. http://epsilon.slu.se/a467.pdf.
Texto completoLundgren, Josefin. "Studies of metazoan proteasome function and regulation". Doctoral thesis, Stockholm : Department of molecular biology and functional genomics, Stockholm university, 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:su:diva-496.
Texto completoBurnett, Amanda. "Regulation of Neutrophil Function by the Angiopoietins". Thesis, University of Sheffield, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.522497.
Texto completoSavage, Joshua S. "Protein kinase-dependent regulation of platelet function". Thesis, University of Bristol, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.559232.
Texto completoWitzel, Ini-Isabee. "Investigating the function and regulation of SNIP1". Thesis, University of Bristol, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.529875.
Texto completoJanas, Maja. "Novel Regulation of MicroRNA Biogenesis and Function". Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10121.
Texto completoCadman, Chris. "Regulation of the helicase function of PriA". Thesis, University of Nottingham, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.428956.
Texto completoClemett, Delyth A. "5-HTâ†7 receptor regulation and function". Thesis, University of Nottingham, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.262781.
Texto completoMcMullan, Rachel Jane. "Regulation of keratinocyte function by Rho kinase". Thesis, University of Birmingham, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.275126.
Texto completoHigham, Andrew Damian. "Neuroendocrine regulation of gastric endocrine cell function". Thesis, University of Liverpool, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.266054.
Texto completoFinkelstein, Erik I. "Regulation of neutrophil function by toxic aldehydes /". For electronic version search Digital dissertations database. Restricted to UC campuses. Access is free to UC campus dissertations, 2002. http://uclibs.org/PID/11984.
Texto completoHillwig, Melissa S. "Regulation, function, and evolution of T2 RNases". [Ames, Iowa : Iowa State University], 2009. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3389103.
Texto completoMeng, Meng. "Plant UDP-glucose Pyrophosphorylase : Function and Regulation". Doctoral thesis, Umeå : Department of Plant Physiology, Umeå University, 2008. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-1796.
Texto completoGlanz, Anna Nicole. "Regulation of the Antiviral Function of IRF3". University of Toledo Health Science Campus / OhioLINK, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=mco1596792904962609.
Texto completoLindås, Ann-Christin. "Tripeptidyl-Peptidase II : Structure, Function and Gene Regulation". Doctoral thesis, Uppsala University, Department of Biochemistry, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-7345.
Texto completoThe protein degradation process is of vital importance for the cell to maintain cellular functions. An important enzyme in this process is the multimeric tripeptidyl-peptidase II (TPP II). It removes tripeptides from a free N-terminus of the substrates. TPP II has broad substrate specificity and wide-spread distribution, suggesting that the TPP II gene is a house-keeping gene. However, the levels of both mRNA and TPP II protein varies during different conditions and the TPP II gene promoter was therefore identified and characterized. It is a 215 bp fragment just upstream of the coding sequence. This fragment lacks a TATA-box but contains an initiator, two inverted CCAAT-boxes and an E-box. The CCAAT-boxes and the E-box were found to bind the nuclear factor Y (NF-Y) and upstream stimulatory factor-1 (USF-1) respectively. The CCAAT-boxes appear to be most important for the transcriptional activation. Furthermore, several silencer element were identified further upstream of the 215 bp promoter and the octamer binding factor Oct-1 was found to bind one of these fragments. If Oct-1 is responsible for the inhibition of the transcription of the TPP II gene remains to be investigated. In addition, the substrate specificity was investigated. For this purpose an expression system using Pichia pastoris was developed. The purified recombinant TPP II was found to have the same enzymatic properties as the native enzyme. In order to identify the amino acids involved in the binding of the N-terminus of the substrate, wild-type murine TPP II and four mutants E305Q, E305K, E331Q and E331K were purified. Steady-state kinetic analysis clearly demonstrated that both Glu-305 and Glu-331 are important for this binding as the KMapp is more than 102 higher for the mutants than wild-type. Finally, the pH-dependence for cleavage of two chromogenic substrates was compared for TPP II from different species.
Holmqvist, Marie. "The Cyanobacterial Uptake Hydrogenase : Regulation, Maturation and Function". Doctoral thesis, Uppsala universitet, Mikrobiell Kemi, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-129223.
Texto completoEriksson, Anna S. "Syndecan - Regulation and Function of its Glycosaminoglycan Chains". Doctoral thesis, Uppsala universitet, Institutionen för medicinsk biokemi och mikrobiologi, 2013. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-197691.
Texto completoAfar, Ronith. "Regulation and function of neuronal nicotinic acetylcholine receptors". Thesis, McGill University, 1993. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=41288.
Texto completoInitial studies involved thymopentin (TP-5), a 5 amino-acid peptide which may represent the active site of TPO. TP-5 inhibited nicotinic receptor-induced release of catecholamines in bovine adrenal medullary cells in culture, a function mediated through the $ alpha$-BGT-insensitive nAChR. On the other hand, TP-5 did not inhibit either ($ sp3$H) (-)nicotine or ($ sp{125}$I) $ alpha$-BGT binding to rat brain membranes. These results suggested that TP-5 interacted in a non-competitive manner with the $ alpha$-BGT-insensitive neuronal nAChR.
Studies were subsequently done with thymic preparations presumed to be purified TPO ('TPO'), the native polypeptide containing the TP-5 amino acid sequence. In contrast to the effect of TP-5, 'TPO' preparations did not alter nicotinic receptor mediated catecholamine release from neuronal cells in culture. However, 'TPO' preparations selectively decreased ($ sp{125}$I) $ alpha$-BGT binding to brain membranes suggesting an interaction between this polypeptide and $ alpha$-BGT receptors. Quantitative autoradiography revealed that 'TPO' inhibited specific ($ sp{125}$I) $ alpha$-BGT binding uniformly and with similar potency in different brain regions. As $ alpha$-BGT binding sites are highly expressed in the hippocampal formation, primary cultures of fetal rat hippocampal cells were used next to investigate regulation of the $ alpha$-BGT receptor by 'TPO'. 'TPO' caused a dose-dependent and slowly reversible decrease in the density of $ alpha$-BGT receptors. After completion of this work with 'TPO', studies by Quik and coworkers (1993) showed that $ alpha$-Naja toxin (or $ alpha$-cobratoxin) from Naja naja siamensis snake venom was present in the 'TPO' preparations; furthermore, this toxin component appeared to be responsible for the reported effects of 'TPO' on $ alpha$-BGT receptors. Therefore, the above results which had initially been interpreted to occur as a consequence of the interaction of the thymic polypeptide TPO at the nicotinic $ alpha$-BGT site, must now be attributed to the presence of $ alpha$-Naja toxin contaminant in the 'TPO' preparations.
Studies were also undertaken to identify a nicotinic function for $ alpha$-BGT receptors in neuronal cells. Intracellular calcium levels were measured in response to nicotine as recent work using parasympathetic neurons showed that this may represent an $ alpha$-BGT-sensitive response. In contrast to earlier findings, the present work indicates that nAChR-mediated calcium fluxes in cultured chromaffin cells do not reveal an $ alpha$-BGT-sensitive component. These results thus suggest that nicotinic $ alpha$-BGT receptors mediate their response by altering intracellular calcium levels in some but not all neuronal preparations.
Unterberger, Alexander. "The role of DNMT1 regulation in cellular function". Thesis, McGill University, 2010. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=92154.
Texto completoDuring the cell cycle DNMT1 levels increase as the cell enters into S-phase. It has previously been shown that this cyclical regulation of DNMT1 occurs by destabilization of DNMT1 mRNA in G0/G1 through the action of a protein, identified to be the mRNA binding protein AUF1. AUF1 binds a regulator element located in the 3'UTR of DNMT1 mRNA and recruits the exosome, the RNA degradation complex, to degrade it.
When AUF1 is depleted in these cells, DNMT1 mRNA is stabilized which leads to increased DNMT1 protein levels, methyltransferase activity and genomic methylation. The changes of DNMT1 mRNA levels in the cell cycle were determined to occur as an inverse function of AUF1 protein levels. AUF1 levels were observed to decrease in S-phase which lead to increased stability in DNMT1 mRNA. This cell cycle regulation of AUF1 was determined to occur as a function of Rb. Rb actively stabilizes AUF1 protein. Indeed, upon elimination of Rb, AUF1 is degraded through the function of Hsp70 and the proteasome. This consequently leads to an elevation in DNMT1 protein levels which in turn increases genomic methylation levels. Elevated DNMT1 levels resulted in greater association with EZH2, which in turn leads to increased methylation of EZH2 targeted promoters, including p16 and CNR1. This promoter hypermethylation occurred as a function of DNMT1 and EZH2.These observations indicate that regulation of DNMT1 is tied into the cell cycle function of Rb and upon disruption of this system, a characteristic of cancer, site-specific methylation occurs at tumour suppressors, another characteristic of cancer.
Furthermore, we examined the effect of depleting DNMT1 in cancer cells. Upon depletion of DNMT1, a signaling pathway known as the replication arrest/DNA damage checkpoint was induced. Activation of this pathway results in arrest of cell growth and cell cycle blockage and occurred independently of the catalytic activity of DNMT1 and instead responded to the absence of DNMT1. This supports a role for DMNT1 as a negative regulator of the replication arrest/DNA damage checkpoint through the action of interaction with an unknown protein. Moreover, suppression of the replication arrest/DNA damage checkpoint has been determined to be a necessary step in the proliferation of cancer cells. Taken together, the data from this thesis determined that common events in cancer, such as inactivation of Rb, lead to deregulation of DNMT1 mRNA, through AUF1, leading to site-specific methylation of tumour suppressors and could potentially serve to block growth arresting checkpoints like the replication arrest/DNA damage checkpoint. The novel functions of DNMT1, such as cell cycle regulation, site-specific methylation and role in the replication arrest/DNA damage checkpoint discovered in this thesis could serve to help better understand how cancer develops. The results of this thesis could serve to develop novel strategies to target these events and better treat cancer.
L'altération de l'épigénome et de ses composants est une marque caractéristique de tous types de cancer. Une altération des profils de méthylation de l'ADN, associée à une inactivation de gènes suppresseurs de tumeurs ainsi qu'une augmentation de l'(activité/expression) de la méthyltransférase de l'ADN (DNMT1) sont largement observés dans les cancers. Cependant, les causes de cette augmentation de DNMT1 (expression/activité) dans le cancer et l'utilisation potentielle de cette augmentation comme cible thérapeutique n'ont pas encore été déterminées.
Au cours du cycle cellulaire, le niveau de DNMT1 augmente dès lors que la cellule entre en phase S. Il a été montré précédemment qu'une régulation cyclique de DNMT1 se met en place grâce à une déstabilisation de son ARN messager en phase G0/G1 sous l'action d'une protéine non identifiée. Cette protéine a été identifié comme AUF1. AUF1 interagit avec un élément régulateur situé dans la partie 3'-UTR de l'ARNm de DNMT1 et entraîne la dégradation de cet ARNm en recrutant l'exosome, un complexe de dégradation de l'ARN. La déplétion d'AUF1 stabilise l'ARNm de DNMT1 ce qui conduit à une augmentation de l'expression de cette protéine, de son activité méthyltransférase ainsi que de la méthylation du génome. Il a été également montré que le niveau d'expression de l'ARNm de DNMT1 au cours du cycle cellulaire est inversement corrélé à celui de la protéine AUF1. Ce niveau d'AUF1 est diminué en phase S ce qui traduit par une stabilité accrue de l'ARNm de DNMT1. Il a été montré que cette régulation d'AUF1 au cours du cycle cellulaire est fonction de la protéine Rb. Rb stabilise activement la protéine AUF1. En effet, AUF1 est dégradée par l'intermédiaire de la protéine Hsp70 et du protéasome. Cette dégradation a pour conséquence une augmentation du niveau d'expression de DNMT1 lequel conduit à une augmentation du niveau de méthylation du génome. De plus, cette augmentation de DNMT1 résulte en une plus grande association avec la protéine EZH2 entraînant une hyperméthylation de promoteurs de gènes ciblés par EZH2 (ex : p16, CNR1 et PCNA). Ces observations démontrent que la régulation de DNMT1 est étroitement liée aux fonctions de Rb dans le cycle cellulaire. Caractéristique dans les cancers, une rupture de cette relation DNMT1-Rb, entraîne ainsi une méthylation site-spécifique de gènes suppresseurs de tumeurs, une autre caractéristique des cancers.
En parallèle, nous avons étudié l'effet d'une déplétion de DNMT1 dans des cellules cancéreuses. Suite à une déplétion de DNMT1, une voie de signalisation connue comme un point de contrôle de l'arrêt de la réplication/lésions de l'ADN est induite. L'activation de cette voie de signalisation entraîne l'arrêt de la croissance cellulaire et le blocage du cycle cellulaire. L'activation de cette voie répond à l'absence de DNMT1 et de façon indépendante de son activité catalytique. Ceci est en faveur d'un rôle pour DNMT1 de régulateur négatif du contrôle de l'arrêt de la réplication/lésions de l'ADN via l'interaction avec une protéine qui reste encore à identifier. De plus, la suppression des points de contrôle de l'arrêt de la réplication/lésion de l'ADN a été montré comme étant une étape nécessaire à la prolifération des cellules cancéreuses. L'ensemble des données de cette thèse démontre que des événements communs aux cancers, telle que l'inactivation de Rb, peuvent conduire à la dérégulation, via AUF1, de l'ARNm de DNMT1, laquelle entraîne la méthylation site-spécifique de gènes suppresseurs de tumeurs. Cette dérégulation de DNMT1 pourrait potentiellement servir à bloquer les points de contrôle d'arrêt du cycle cellulaire/lésions de l'ADN.
Les nouvelles fonctions de DNMT1, telles que la régulation du cycle cellulaire, la méthylation site-spécifique et le contrôle de la réplication/lésions de l'ADN découverts dans cette thèse devraient permettre de mieux comprendre le développement cancéreux et de développer de nouvelles stratégies thérapeutiques.