Transcriptional repression of human cad gene by hypoxia inducible factor-1alpha.
Transcriptional repression of human cad gene by hypoxia inducible factor-1alpha.
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DOI：
10.1093/nar/gki839
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发表时间：
2005
影响因子：
14.9
通讯作者：
Tsai SJ
中科院分区：
文献类型：
--
作者：
Chen KF;Lai YY;Sun HS;Tsai SJ
嘧啶核苷酸的从头生物合成为DNA合成和细胞增殖提供了必要的前体。嘧啶从头合成的前三个步骤由称为CAD（氨甲酰磷酸合成酶-天冬氨酸氨甲酰转移酶-二氢乳清酸酶）的多功能酶催化。在这项工作中，在缺氧或去铁胺（DFO）诱导的HIF-1α积累下孵育的许多细胞系和原代培养人基质细胞中检测到CAD表达的减少。通过生物信息学方法分析人cad基因启动子，鉴定了一个假定的缺氧反应元件（HRE）结合基质。启动子活性测定，分别使用含有cad启动子（−710/+122）和−67/HRE片段（25个碱基）的构建体，证明了缺氧条件下对p53基因表达的抑制。cad启动子活性的抑制通过野生型HIF-1α的强制表达得到证实，但通过显性负性HIF-1α的过表达被消除。染色质免疫沉淀试验进一步证明了HIF-1α在体内与cad启动子结合。这些数据表明，cad基因的表达受到HIF-1α的抑制，这代表了缺氧和细胞周期停滞之间的功能联系。
De novo biosynthesis of pyrimidine nucleotides provides essential precursors for DNA synthesis and cell proliferation. The first three steps of de novo pyrimidine biosynthesis are catalyzed by a multifunctional enzyme known as CAD (carbamoyl phosphate synthetase-aspartate carbamoyltransferase-dihydroorotase). In this work, a decrease in CAD expression is detected in numerous cell lines and primary culture human stromal cells incubated under hypoxia or desferrioxamine (DFO)-induced HIF-1α accumulation. A putative hypoxia response element (HRE) binding matrix is identified by analyzing human cad-gene promoter using a bioinformatic approach. Promoter activity assays, using constructs harboring the cad promoter (−710/+122) and the −67/HRE fragment (25-bases), respectively, demonstrate the suppression of reporter-gene expression under hypoxia. Suppression of cad-promoter activity is substantiated by forced expression of wild-type HIF-1α but abolished by overexpression of dominant-negative HIF-1α. A chromatin immunoprecipitation assay provides further evidence that HIF-1α binds to the cad promoter in vivo. These data demonstrate that the cad-gene expression is repressed by HIF-1α, which represents a functional link between hypoxia and cell-cycle arrest.
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影响因子：
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