The pancreatic islet β-cell-enriched transcription factor Pdx-1 regulates Slc30a8 gene transcription through an intronic enhancer.
The pancreatic islet β-cell-enriched transcription factor Pdx-1 regulates Slc30a8 gene transcription through an intronic enhancer.
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DOI：
10.1042/bj20101488
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发表时间：
2011-01-01
期刊：
影响因子：
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通讯作者：
O'Brien RM
中科院分区：
文献类型：
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作者：
Pound LD;Hang Y;Sarkar SA;Wang Y;Milam LA;Oeser JK;Printz RL;Lee CE;Stein R;Hutton JC;O'Brien RM
SLC30A8基因编码锌转运体ZnT-8，该转运体为胰岛素六聚体的形成提供锌。全基因组关联研究表明，SLC30A8的多态性变异与2型糖尿病易感性的改变有关，我们最近报道了从SLC30A8基因敲除小鼠分离的胰岛中葡萄糖刺激的胰岛素分泌减少。本研究探讨了Slc30a8在胰岛特异性表达的分子基础。VISTA分析在Slc30a8内含子2和3上发现了两个保守区域，分别指定为增强子A和B。转染实验表明，增强子B在βTC-3细胞和αTC-6细胞中均能提高融合基因的表达。相比之下，增强子A在βTC-3而非αTC-6细胞中选择性地提高融合基因的表达。这些数据表明增强子A是胰岛β细胞特异性增强子，并且在α和β细胞中控制Slc30a8表达的机制重叠但不同。凝胶阻滞和染色质免疫沉淀（ChIP）实验显示，富集胰岛的转录因子Pdx-1分别在体外和原位结合增强子A。增强子A中两个Pdx-1结合位点的突变显著降低了融合基因的表达，提示该因子促进了Slc30a8在β细胞中的表达，这一结论与发育研究一致，表明限制Pdx-1进入胰岛β细胞与体内诱导Slc30a8基因和ZnT-8蛋白表达相关。
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.