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Please use this identifier to cite or link to this item: http://repository.li.mahidol.ac.th/dspace/handle/123456789/24162
Title: Transcriptional regulation of the distal promoter of the rat pyruvate carboxylase gene by hepatocyte nuclear factor 3β/Foxa2 and upstream stimulatory factors in insulinoma cells
Authors: Thirajit Boonsaen
Pinnara Rojvirat
Kathy H. Surinya
John C. Wallace
Sarawut Jitrapakdee
Mahidol University
University of Adelaide
Keywords: Biochemistry, Genetics and Molecular Biology;Medicine
Issue Date: 15-Jul-2007
Citation: Biochemical Journal. Vol.405, No.2 (2007), 359-367
Abstract: PC (pyruvate carboxylase) plays a crucial role in intermediary metabolism including glucose-induced insulin secretion in pancreatic islets. In the present study, we identified two regions of the 1.2 kb distal promoter, the -803/-795 site and the -408/-403 E-box upstream of the transcription start site, as the important cis-acting elements for transcriptional activation of the luciferase reporter gene. Site-directed mutagenesis of either one of these sites in the context of this 1.2 kb promoter fragment, followed by transient transfections in the insulinoma cell line, INS-1, abolished reporter activity by approx. 50%. However, disruption of either the -803/-795 or the -408/-403 site did not affect reporter gene activity in NIH 3T3 cells, suggesting that this promoter fragment is subjected to cell-specific regulation. The nuclear proteins that bound to these -803/-795 and -408/-403 sites were identified by gel retardation assays as HNF3β (hepatocyte nuclear factor 3β)/Foxa2 (forkhead/winged helix transcription factor box2) and USFs (upstream stimulatory factors), USF1 and USF2, respectively. Chromatin immunoprecipitation assays using antisera against HNF3β/Foxa2, USF1 and USF2 demonstrated that endogenous HNF3β/Foxa2 binds to the -803/-795 Foxa2 site, and USF1 and USF2 bind to the -408/-403 E-box respectively in vivo, consistent with the gel retardation assay results. Although there are weak binding sites located at regions -904 and -572 for PDX1 (pancreatic duodenal homeobox-1), a transcription factor that controls expression of β-cell-specific genes, it did not appear to regulate PC expression in INS-1 cells in the context of the 1.2 kb promoter fragment. The results presented here show that Foxa2 and USFs regulate the distal promoter of the rat PC gene in a cell-specific manner. © 2007 Biochemical Society.
URI: https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=34447116301&origin=inward
http://repository.li.mahidol.ac.th/dspace/handle/123456789/24162
ISSN: 02646021
Appears in Collections:Scopus 2006-2010

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