Long-Range Enhancers Are Required to Maintain Expression of the Autoantigen Islet-Specific Glucose-6-Phosphatase Catalytic Subunit–Related Protein in Adult Mouse Islets In Vivo

Abstract

OBJECTIVE-Islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) is selectively expressed in islet ␤-cells and is a major autoantigen in both mouse and human type 1 diabetes. This study describes the use of a combination of transgenic and transfection approaches to characterize the gene regions that confer the islet-specific expression of IGRP.RESEARCH DESIGN AND METHODS-Transgenic mice were generated containing the IGRP promoter sequence from Ϫ306, Ϫ911, or Ϫ3911 to ϩ3 ligated to a LacZ reporter gene. Transgene expression was monitored by 5-bromo-4-chloro-3-indolyl-␤-D-galactopyranoside staining of pancreatic tissue.RESULTS-In all the transgenic mice, robust LacZ expression was detected in newborn mouse islets, but expression became mosaic as animals aged, suggesting that additional elements are required for the maintenance of IGRP gene expression. VISTA analyses identified two conserved regions in the distal IGRP promoter and one in the third intron. Transfection experiments demonstrated that all three regions confer enhanced luciferase reporter gene expression in ␤TC-3 cells when ligated to a minimal IGRP promoter. A transgene containing all three conserved regions was generated by using a bacterial recombination strategy to insert a LacZ cassette into exon 5 of the IGRP gene. Transgenic mice containing a 15-kbp fragment of the IGRP gene were then generated. This transgene conferred LacZ expression in newborn mouse islets; however, expression was still suppressed as animals aged. CONCLUSIONS-The

The pancreatic islet β-cell-enriched transcription factor Pdx-1 regulates Slc30a8 gene transcription through an intronic enhancer

“…Intronic enhancers have previously been identified in several genes whose expression are enriched in islets including those encoding glucagon [26], islet amyloid polypeptide [27] and G6pc2 [28]. To determine whether intronic enhancers may also exist in the human SLC30A8 and mouse Slc30a8 genes a sequence alignment was performed using the VISTA program [29] focusing on the region between the translation start site and the last exon [7].…”

Moving on from GWAS: Functional Studies on the G6PC2 Gene Implicated in the Regulation of Fasting Blood Glucose

“…Furthermore, the catalytically more active G6pc3 isoform was expressed at much higher levels [79] such that the loss of barely detectable G6pc2 expression would likely have little impact on total hypothalamic glucose-6-phosphatase activity. Of note, an analysis of multiple G6pc2 - LacZ transgenes failed to detect hypothalamic expression [80, 27, 81]. Finally, a third possibility to explain the connection between G6PC2 and body weight relates to the kinetics of GSIS.…”

Glucose-6-phosphatase Catalytic Subunit Gene Family

“…However, unlike the endogenous G6pc2 gene, transgene expression decreases after birth, indicating that this Ϫ306/ϩ3 promoter region is unable to confer transgene expression in adult mice (39). The search for the transcriptional elements that mediate sustained G6pc2 expression in adult animals has led to the identification of multiple enhancers 5Ј, within, and 3Ј of the G6pc2 gene; however, the transcriptional boundaries of the G6pc2 locus remain to be defined (40).…”