A Role for Glycogen Synthase Kinase 3 beta in the Regulation of Glucagon Gene Transcription by Insulin

Dimopoulos, Nikolaos

dc.contributor.advisor	Hardeland, Rüdiger Prof. Dr.	de
dc.contributor.author	Dimopoulos, Nikolaos	de
dc.date.accessioned	2012-04-16T14:55:57Z	de
dc.date.available	2013-01-30T23:51:06Z	de
dc.date.issued	2003-11-21	de
dc.identifier.uri	http://hdl.handle.net/11858/00-1735-0000-0006-AE65-F	de
dc.identifier.uri	http://dx.doi.org/10.53846/goediss-559
dc.description.abstract	Insulin reguliert über seine Bindung an den Insulin Rezeptor in a-Zellen des endokrinen Pankreas eine Signalkaskade, die in einer Hemmung der Glucagongen-Transkription resulitert. In der vorliegenden Promotionsarbeit wurde dieser bis jetzt nicht vollständig aufgeklärte Mechanismus in Hinblick auf die Rolle von GSK3 untersucht. Es konnte gezeigt werden, dass die Überexpression von GSK3 die Glucagongen Trankription verstärkte.GSK3 Inhibitoren unterdrückten die Trankripiton des Glucagongens und zeigten somit den selben Effekt wie Insulin. Untersuchungen an Deletions- und Mutationsanalysen des Glucagonpromotors gaben Hinweise auf eine mögliche Wirkung von GSK3 über den Transkriptionsfaktor Pax6. Die transkriptionsaktivierende Wirkung von Pax6 und CBP wurde durch den GSK3 Inhibitor SB-216763 aufgehoben. Dies wurde in einem GAL4-System gezeigt. Rekombinantes GSK3b phosphorilierte die Transaktivierungsdomäne von Pax6 und ein CBP-Fragment, das die Aminosäuren 2040-2305 enthielt. Dies konnte durch in vitro Kinase assays gezeigt werden. Die Ergebnisse zeigen, das GSK3-b in die Regulation des Glucagongens involviert ist.	de
dc.format.mimetype	application/pdf	de
dc.language.iso	eng	de
dc.rights.uri	http://webdoc.sub.gwdg.de/diss/copyrdiss.htm	de
dc.title	A Role for Glycogen Synthase Kinase 3 beta in the Regulation of Glucagon Gene Transcription by Insulin	de
dc.type	doctoralThesis	de
dc.title.translated	Rolle der Glycogen Synthase Kinase 3 beta in der Regulation der Glucagongen-Transkription durch Insulin	de
dc.contributor.referee	Pieler, Tomas Prof. Dr.	de
dc.date.examination	2003-11-05	de
dc.subject.dnb	570 Biowissenschaften, Biologie	de
dc.description.abstracteng	Insulin binding to its receptor on the a-cells in endocrine pancreas initiates a signalling cascade that results in inhibition of glucagon gene transcription. Evidence suggests that the PI(3)K/PKB pathway might confer insulin-mediated inhibition, but the mechanism is still not completely understood. The present study characterized the role of GSK3b, a well known substrate of PKB, in the regulation of glucagon gene transcription.enhancement of glucagon gene transcription. The effect was due to its kinase activity, because a GSK3b kinase-dead mutant had no effect on transcription. Next, endogenous pools of GSK3 were assessed by using various recently developed selective GSK3 inhibitors. All of them inhibited transcription, thereby mimicking the effect of insulin. They could also abolish the effect of GSK3b overexpression on the stimulation of glucagon gene transcription. To validate the mode of action of the inhibitors, we could show that they modulated putative GSK3 substrates in vivo, e.g. they induced stabilization and accumulation of b-catenin levels.GSK3 might act through Pax6 and its potential coactivator CBP to regulate glucagon gene expression. Deletion and internal mutation analysis of the glucagon promoter indicated that Pax6 might play an important role to confer inhibition by the GSK3 inhibitor, SB-216763. Transcriptional activity of Pax6 and CBP was inhibited in response to SB-216763 within the glucagon promoter context, as revealed by employing the Gal4 system. Fusing full length CBP, as well as N- and C-terminal parts of CBP, with the Gal4 domain conferred transcriptional activity as well as responsiveness to GSK3b overexpression. The effect of GSK3b overexpression on the activity of the C-terminal part of CBP was further mapped.Finally, based on the above findings an in vitro kinase assay was performed. Recombinant GSK3b phosphorylated, even in the absence of priming phosphorylation by another kinase, the transactivation domain of Pax6 and a fragment of CBP consisting of amino acids 2040-2305 when fused to GST. However, the significance of this phosphorylation in vivo and in the context of the glucagon promoter remains to be shown.Taken together the results of the present study suggest the involvement of GSK3b in the regulation of glucagon gene transcription by insulin. Through inhibition of GSK3, insulin might repress gene transcription by disruption of a glucagon promoter-specific protein complex, whose critical components seem to be Pax6 and its coactivator CBP. Since hyperglucagonaemia is associated with type 2 diabetes mellitus, understanding the molecular mechanism of insulin-mediated inhibition of glucagon gene expression can prove a valuable tool to develop drugs to treat some of the aspects of the disease.	de
dc.contributor.coReferee	Gatz, Christiane Prof. Dr.	de
dc.contributor.thirdReferee	Ficner, Ralf Prof. Dr.	de
dc.subject.topic	Mathematics and Computer Science	de
dc.subject.ger	Glucagon	de
dc.subject.ger	Insulin	de
dc.subject.ger	GSK3	de
dc.subject.ger	Transkription	de
dc.subject.eng	glucagon	de
dc.subject.eng	insulin	de
dc.subject.eng	gsk3	de
dc.subject.eng	transcription	de
dc.subject.bk	30	de
dc.identifier.urn	urn:nbn:de:gbv:7-webdoc-411-8	de
dc.identifier.purl	webdoc-411	de
dc.affiliation.institute	Biologische Fakultät inkl. Psychologie	de
dc.subject.gokfull	W	de
dc.identifier.ppn	377351202	de

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