The minimal promoter region of the dense-core vesicle protein IA-2: transcriptional regulation by CREB

Abstract

IA-2 is a transmembrane protein found in the dense core vesicles (DCV) of neuroendocrine cells and one of the major autoantigens in type 1 diabetes. DCV are involved in the secretion of hormones (e.g., insulin) and neurotransmitters. Stimulation of pancreatic β cells with glucose upregulates the expression of IA-2 and an increase of IA-2 results in an increase of the number of DCV. Little is known, however, about the promoter region of IA-2 or the transcriptional factors that regulate the expression of this gene. In the present study, we constructed eight deletion fragments from the upstream region of the IA-2 transcription start site and linked them to a luciferase reporter. By this approach we have identified a short bp region (−216 to +115) that has strong promoter activity. We also identified a transcription factor, CREB (cAMP Responsive Element Binding Protein), which binds to two CREB-related binding sites located in this region. The binding of CREB to these sites enhanced IA-2 transcription by more than 5 fold. We confirmed these findings by site-directed mutagenesis, chromatin immunoprecipitation (ChIP) assays and RNAi inhibition. Based on these findings we conclude that the PKA pathway is a critical, but not the exclusive signaling pathway involved in IA-2 gene expression.