Pancreatic β cell-specific transcription of the pdx-1 gene -: The role of conserved upstream control regions and their hepatic nuclear factor 3β sites
Pancreatic β cell-specific transcription of the pdx-1 gene -: The role of conserved upstream control regions and their hepatic nuclear factor 3β sites
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DOI：
10.1074/jbc.275.5.3485
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发表时间：
2000-02-04
影响因子：
4.8
通讯作者：
Stein, R
中科院分区：
文献类型：
--
作者：
Gerrish, K;Grannon, M;Stein, R
为了确定PDX-1的潜在反式激活因子，我们对人和鸡的同源物的5‘启动子区域进行了大约4.5kb的测序，假设与小鼠基因保守的序列将包含关键的顺式调节元件。与超敏位点1(HSS1)相关的序列代表了主要的同源性区域，其中明显存在三个保守的亚区：区域I(-2694到-2561个碱基对(BP))、区域II(-2139到-1958个碱基对)和区域III(-1879到-1799个碱基对)，尽管在鸡的这个区域中只存在区域I和III，但小鼠和鸡/人的基因之间的同源性很高，从78%到89%不等。从PDX-1驱动的报告活性分析显示，胰岛β细胞的选择性表达受鼠和人的I区或II区控制，但不受III区的控制。对位于I区和II区的保守的肝核因子3(HNF3)样位点的突变和功能分析表明，这些区域的激活是由HNF3β介导的，为了确定内源基因背景下是否存在类似的调控关系，在HNF3β等位基因之一或两个失活的胚胎干细胞中检测到PDX-1的表达，在纯合子缺失的HNF3β细胞中诱导向类胚体分化时PDX-1mRNA水平下调。综上所述，这些结果表明，区域I和II所代表的保守序列定义了HNF3β等因子的结合位点，这些因子控制着PDX-1基因的胰岛β细胞选择性表达。
To identify potential transactivators of pdx-1, we sequenced approximately 4.5 kilobases of the 5' promoter region of the human and chicken homologs, assuming that sequences conserved with the mouse gene would contain critical cis-regulatory elements. The sequences associated with hypersensitive site 1 (HSS1) represented the principal area of homology within which three conserved subdomains were apparent: area I (-2694 to -2561 base pairs (bp)), area II (-2139 to -1958 bp), and area III (-1879 to -1799 bp), The identities between the mouse and chicken/human genes are very high, ranging from 78 to 89%, although only areas I and III are present within this region in chicken. Pancreatic beta cell-selective expression was shown to be controlled by mouse and human area I or area II, but not area III, from an analysis of pdx-1-driven reporter activity in transfected beta- and non-beta cells. Mutational and functional analyses of conserved hepatic nuclear factor 3 (HNF3)-like sites located within area I and area II demonstrated that activation by these regions was mediated by HNF3 beta, To determine if a similar regulatory relationship might exist within the context of the endogenous gene, pdx-1 expression was measured in embryonic stem cells in which one or both alleles of HNF3 beta were inactivated, pdx-1 mRNA levels induced upon differentiation to embryoid bodies were down-regulated in homozygous null HNF3 beta cells. Together, these results suggest that the conserved sequences represented by areas I and II define the binding sites for factors such as HNF3 beta, which control islet beta cell-selective expression of the pdx-1 gene.