Molecular mechanisms of transcription activation by HLF and HIF1α in response to hypoxia:: their stabilization and redox signal-induced interaction with CBP/p300
Molecular mechanisms of transcription activation by HLF and HIF1α in response to hypoxia:: their stabilization and redox signal-induced interaction with CBP/p300
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DOI：
10.1093/emboj/18.7.1905
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发表时间：
1999-04-01
期刊：
影响因子：
11.4
通讯作者：
Fujii-Kuriyama, Y
中科院分区：
文献类型：
--
作者：
Ema, M;Hirota, K;Fujii-Kuriyama, Y
缺氧诱导因子1 α (hypoxia -inducible factor 1 α, HIF1 α)及其相关因子HLF在低氧环境下激活促红细胞生成素等基因的表达，利用GAL4DBD与这两个因子不同片段的融合基因转染分析，确定了两个在缺氧条件下可诱导的转录激活结构域，它们的序列在HLF和HIF1 α之间保守。一个被命名为NAD （n端激活域），另一个被命名为CAD （c端激活域）。在哺乳动物双杂交系统中，CAD和NAD诱饵通过与creb结合蛋白（CBP）猎物共转染，增强了报告基因荧光素酶的表达，而CAD诱饵通过CBP共表达，增强了酵母中β -半乳糖苷酶的表达，而NAD诱饵通过CBP共表达，增强了酵母中β -半乳糖苷酶的表达。共转染实验表明，Ref-1和硫氧还蛋白的表达进一步增强了CAD表达的荧光素酶活性，而NAD则没有。CADs序列的氨基酸替换显示，一种特定的半胱氨酸是它们与CBP缺氧诱导的相互作用所必需的，当O-2浓度降低时，细胞质中硫氧还蛋白发生核易位。
Hypoxia-inducible factor 1 alpha (HIF1 alpha) and its related factor, HLF, activate expression of a group of genes such as erythropoietin in response to low oxygen, Transfection analysis using fusion genes of GAL4DBD with various fragments of the two factors delineated two transcription activation domains which are inducible in response to hypoxia and are localized in the C-terminal half, Their sequences are conserved between HLF and HIF1 alpha. One is designated NAD (N-terminal activation domain), while the other is CAD (C-terminal activation domain). Immunoblot analysis revealed that NADs, which were rarely detectable at normoxia, became stabilized and accumulated at hypoxia, whereas CADs were constitutively expressed, In the mammalian two-hybrid system, CAD and NAD baits enhanced the luciferase expression from a reporter gene by co-transfection with CREB-binding protein (CBP) prey, whereas CAD, but not NAD, enhanced beta-galactosidase expression in yeast by CBP co-expression, suggesting that NAD and CAD interact with CBP/p300 by a different mechanism, Co-transfection experiments revealed that expression of Ref-1 and thioredoxin further enhanced the luciferase activity expressed by CAD, but not by NAD, Amino acid replacement in the sequences of CADs revealed a specific cysteine to be essential for their hypoxia-inducible interaction with CBP, Nuclear translocation of thioredoxin from cytoplasm was observed upon reducing O-2 concentrations.