Involvement of PDX-1 in activation of human insulin gene transcription.
Islet beta cell-specific transcription of the insulin gene is mediated through the binding of the islet-enriched PDX-1, BETA2, and MafA transcription factors to conserved 5'-flanking region regulatory elements. However, additional non-conserved sequences within this region are also significant in regulating expression. Thus, PDX-1 binds to and activates the GG2 element located between nucleotides -145 and -140 of the human gene, while the corresponding, but non-identical, site in the rodent insulin genes are negatively regulated by the Nkx2.2 transcription factor. Here, we show that despite binding PDX-1 approximately 20-fold less effectively than the conserved insulin A3 and A1 sites in gel mobility shift assays, human GG2 appears to be more important for the activation of transfected human insulin enhancer-driven reporter constructs in beta cell lines. Furthermore, functional interaction analysis in non-islet cell lines demonstrated that PDX-1 binding to GG2, A1, and A3 contributes to synergistic activation of insulin gene expression with MafA. Our analysis also illustrated the requirement of poorly conserved human sequences between -293 and -251 in mediating activity through the more upstream A3 binding site. Collectively these experiments have revealed distinct features in control of the human and rodent insulin genes by PDX-1, processes that may be involved in regulating insulin expression under both normal and diabetic conditions in humans.
Le Lay
John
J
Department of Molecular Physiology and Biophysics, Vanderbilt University, 2001 West End Ave Nashville, Tennessee 37235, USA.
Stein
Roland
R
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017124:Conserved Sequence / Q000235:genetics; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics; D006367:HeLa Cells; D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D007328:Insulin / Q000235:genetics*; D050417:Insulin-Secreting Cells / Q000502:physiology; D051473:Maf Transcription Factors, Large / Q000235:genetics; D051379:Mice; D009687:Nuclear Proteins; D015534:Trans-Activators / Q000235:genetics*; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000235:genetics; D014162:Transfection
D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins; D007328:Insulin; C485159:MAFA protein, human; D051473:Maf Transcription Factors, Large; C000715631:NKX2-2 protein, human; C000715627:Nkx2-2 protein, mouse; D009687:Nuclear Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C084010:pancreatic and duodenal homeobox 1 protein