Identification of an INSM1-binding site in the insulin promoter: negative regulation of the insulin gene transcription

In this study, an insulinoma-associated antigen-1 (INSM1)-binding site in the proximal promoter sequence of the Insulin gene was identified. The co-transfection of INSM1 with rat Insulin I/II promoter-driven reporter genes exhibited a 40-50% inhibitory effect on the reporter activity. Mutational experiments were performed by introducing a substitution, GG to AT, into the INSM1 core binding site of the rat Insulin I/II promoters. The mutated Insulin promoter exhibited a three- to 20-fold increase in the promoter activity over the wild-type promoter in several insulinoma cell lines. Moreover, INSM1 overexpression exhibited no inhibitory effect on the mutated Insulin promoter. Chromatin immunoprecipitation assays using beta TC-1, mouse fetal pancreas, and Ad-INSM1-transduced human islets demonstrated that INSM1 occupies the endogenous Insulin promoter sequence containing the INSM1-binding site in vivo. The binding of the INSM1 to the Insulin promoter could suppress approximately 50% of Insulin message in human islets. The mechanism for transcriptional repression of the Insulin gene by INSM1 is mediated through the recruitment of cyclin D1 and histone deacetylase-3 to the Insulin promoter. Anti-INSM1 or anti-cyclin D1 morpholino treatment of fetal mouse pancreas enhances the Insulin promoter activity. These data strongly support the view that INSM1 is a new zinc-finger transcription factor that modulates Insulin gene transcription during early pancreas development.