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

Abstract

Synopsis The SLC30A8 gene encodes the zinc transporter ZnT-8, which provides zinc for insulin-hexamer formation. Genome-wide association studies have shown that a polymorphic variant in SLC30A8 is associated with altered susceptibility to type 2 diabetes and we recently reported that glucose-stimulated insulin secretion is decreased in islets isolated from Slc30a8 knockout mice. The present study examines the molecular basis for the islet-specific expression of Slc30a8. VISTA analyses identified two conserved regions in Slc30a8 introns 2 and 3, designated enhancers A and B, respectively. Transfection experiments demonstrated that enhancer B confers elevated fusion gene expression in both βTC-3 cells and αTC-6 cells. In contrast, enhancer A confers elevated fusion gene expression selectively in βTC-3 and not αTC-6 cells. These data suggest that enhancer A is an islet beta cell-specific enhancer and that the mechanisms controlling Slc30a8 expression in alpha and beta cells are overlapping but distinct. Gel retardation and chromatin immunoprecipitation (ChIP) assays revealed that the islet-enriched transcription factor Pdx-1 binds enhancer A in vitro and in situ, respectively. Mutation of two Pdx-1 binding sites in enhancer A markedly reduces fusion gene expression suggesting that this factor contributes to Slc30a8 expression in beta cells, a conclusion consistent with developmental studies showing that restriction of Pdx-1 to pancreatic islet beta cells correlates with the induction of Slc30a8 gene and ZnT-8 protein expression in vivo.

Zinc transporter 8 (ZnT8) and β cell function

“…Pound and colleagues identified a strong transcriptional enhancer in the second intron of the mouse Scl30a8 gene [49] that is highly conserved (>80%) in humans [33]. This enhancer is active in islet-derived β cells but not α cells [33, 49].…”

“…Pound and colleagues identified a strong transcriptional enhancer in the second intron of the mouse Scl30a8 gene [49] that is highly conserved (>80%) in humans [33]. This enhancer is active in islet-derived β cells but not α cells [33, 49]. Analysis of the enhancer by scanning mutagenesis indicates that it binds multiple transcription factors, one of which was identified through gel retardation and ChIP analyses as Pdx-1 [49].…”

“…This enhancer is active in islet-derived β cells but not α cells [33, 49]. Analysis of the enhancer by scanning mutagenesis indicates that it binds multiple transcription factors, one of which was identified through gel retardation and ChIP analyses as Pdx-1 [49]. …”

The Physiological, Biochemical, and Molecular Roles of Zinc Transporters in Zinc Homeostasis and Metabolism

“…ZnT8 transcription in ␤-cells is regulated by the ␤-cell-enriched transcription factor Pdx-1 through an intronic enhancer (326). ZnT8 plays an indispensable role in supplying zinc to insulin granules in ␤-cells to form insulin-zinc crystals.…”

4. New Insights of Islet Biology and the Pathophysiology of Type 2 Diabetes

“…The schematic is based on data, concepts, and figures from several sources (1,23,24,(56)(57)(58)(59)62). Pdx-1, which is broadly expressed in the developing pancreas and then becomes restricted to beta cells and a subset of other endocrine cell types in adult islets, regulates a broad range of processes critical for functional beta cell mass, including those essential to glucose-stimulated insulin secretion (GSIS) (such as GLUT2 and glucokinase), cell survival, replication, and the response to insulin resistance [(51, 52) and references therein; (66)(67)(68)(69)(70)(71)(72)(73)(74)(75)]. In contrast, the MafA transcription factor is expressed very late in islet beta cell development and exclusively in this cell type, but also regulates genes significant to adult islet function (76)(77)(78)(79)(80)(81)(82)(83)(84) (Figure 4-2).…”