Cell-specific expression of glucose-dependent-insulinotropic polypeptide is regulated by the transcription factor PDX-1
Cell-specific expression of glucose-dependent-insulinotropic polypeptide is regulated by the transcription factor PDX-1
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DOI：
10.1210/en.2004-0223
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发表时间：
2005-01-01
期刊：
影响因子：
4.8
通讯作者：
Wolfe, MM
中科院分区：
文献类型：
--
作者：
Jepeal, LI;Fujitani, Y;Wolfe, MM
葡萄糖依赖性促胰岛素多肽（GIP）是一种有效的胰岛素分泌刺激剂，是肠胰岛轴的重要组成部分。GIP在主要位于小肠上部的肠内分泌K细胞中合成。含有同源框的基因PDX-1也在小肠中表达，并且在胰腺发育和胰腺特异性基因的表达中起关键作用。先前的研究确定转录因子加塔-4和ISL-1对GIP的表达是重要的。在这项研究中，我们证明PDX-1也参与调节K细胞中GIP的表达。使用免疫组织化学，我们验证了表达GIP的小鼠K细胞核中PDX-1蛋白的表达，并在受孕后18.5天评估了野生型和PDX-1（-/-）小鼠中PDX-1、5-羟色胺和GIP的表达。虽然我们证明了PDX-1（-/-）小鼠中表达GIP的细胞数量减少了97.8%;但在降钙素阳性细胞数量方面没有统计学差异。此外，PDX-1的转录本和蛋白质在表达GIP的神经内分泌细胞系STC-1中被检测到。使用STC-1核提取物的电迁移率变化测定证明了PDX-1蛋白与GIP启动子中的特定调控区的特异性结合。使用染色质免疫沉淀分析，我们证明了在完整细胞中PDX-1与GIP启动子的相同区域的结合。最后，在瞬时转染试验中PDX-1的过表达导致GIP/Luc报告基因构建体的活性特异性增加。这些研究的结果表明，转录因子PDX-1在GIP基因的细胞特异性表达中起着关键作用。
Glucose-dependent insulinotropic polypeptide (GIP) is a potent stimulator of insulin secretion and comprises an important component of the enteroinsular axis. GIP is synthesized in enteroendocrine K-cells located principally in the upper small intestine. The homeobox-containing gene PDX-1 is also expressed in the small intestine and plays a critical role in pancreatic development and in the expression of pancreatic-specific genes. Previous studies determined that the transcription factors GATA-4 and ISL-1 are important for GIP expression. In this study, we demonstrate that PDX-1 is also involved in regulating GIP expression in K-cells. Using immunohistochemistry, we verified the expression of PDX-1 protein in the nucleus of GIP-expressing mouse K-cells and evaluated the expression of PDX-1, serotonin, and GIP in wild-type and PDX-1(-/-) mice at 18.5 d after conception. Although we demonstrated a 97.8% reduction in the number of GIP-expressing cells in PDX-1(-/-) mice; there was no statistical difference in the number of serotonin-positive cells. Additionally, PDX-1 transcripts and protein were detected in a GIP-expressing neuroendocrine cell line, STC-1. Electromobility shift assays using STC-1 nuclear extracts demonstrated the specific binding of PDX-1 protein to a specific regulatory region in the GIP promoter. Using chromatin immunoprecipitation analysis, we demonstrated binding of PDX-1 to this same region of the GIP promoter in intact cells. Lastly, overexpression of PDX-1 in transient transfection assays led to a specific increase in the activity of GIP/Luc reporter constructs. The results of these studies indicate that the transcription factor PDX-1 plays a critical role in the cell-specific expression of the GIP gene.