Regulation of aldosterone synthase by activator transcription factor/cAMP response element-binding protein family members.
Regulation of aldosterone synthase by activator transcription factor/cAMP response element-binding protein family members.
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DOI：
10.1210/en.2009-0977
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发表时间：
2010-01
期刊：
影响因子：
4.8
通讯作者：
E. Nogueira;W. Rainey
中科院分区：
文献类型：
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作者：
E. Nogueira;W. Rainey
醛固酮合成受血管紧张素II（Ang II）和K（+）作用于肾上腺肾小球的调节，部分通过调节醛固酮合成酶（CYP 11 B2）。在这里，我们分析了cAMP反应元件（CRE）结合蛋白（CREB）在CYP 11B 2调节中的作用。使用定量实时PCR对H295 R细胞和正常人肾上腺组织中的激活转录因子（ATF）/CREB家族成员（即ATF 1和ATF 2、CREB和CRE调节剂）的表达进行分析。通过Western印迹分析分析Ang II诱导的ATF/CREB成员的磷酸化，并使用染色质免疫沉淀分析其随后与CYP 11B 2启动子的结合。在小干扰RNA转染细胞中测定醛固酮产生和CYP 11B 2表达，以敲低ATF/CREB成员的表达。CYP 11B 2启动子活性测定在H295 R细胞中与NR 4A 2共转染或与ATF/CREB成员的组成型活性载体共转染。Ang II以时间依赖性方式诱导ATF 1、ATF 2和CRE调节剂的磷酸化。基于染色质免疫沉淀分析，在Ang II和K（+）处理后，这些蛋白与CYP 11B 2启动子的结合增加。磷酸化ATF/CREB成员也结合CYP 11B 2启动子。ATF/CREB成员的敲除减少了Ang II和K（+）对肾上腺细胞CYP 11 B2 mRNA表达和醛固酮产生的诱导。组成型活性ATF/CREB载体增加了CYP 11B 2的启动子活性，并与CYP 11 R1具有协同作用。总之，这些结果表明ATF/CREB和NGFI-B家族成员在CYP 11B 2的转录调节和肾上腺细胞产生醛固酮的能力中起着至关重要的作用。
Aldosterone synthesis is regulated by angiotensin II (Ang II) and K(+) acting in the adrenal zona glomerulosa, in part through the regulation of aldosterone synthase (CYP11B2). Here, we analyzed the role of cAMP response element (CRE)-binding proteins (CREBs) in the regulation of CYP11B2. Expression analysis of activator transcription factor (ATF)/CREB family members, namely the ATF1 and ATF2, the CREB, and the CRE modulator, in H295R cells and normal human adrenal tissue was performed using quantitative real-time PCR. Ang II-induced phosphorylation of ATF/CREB members was analyzed by Western blot analysis, and their subsequent binding to the CYP11B2 promoter using chromatin immunoprecipitation assay. Aldosterone production and CYP11B2 expression were measured in small interfering RNA-transfected cells to knockdown the expression of ATF/CREB members. CYP11B2 promoter activity was measured in H295R cells cotransfected with NURR1 (NR4A2) alone or with constitutively active vectors for ATF/CREB members. Ang II induced phosphorylation of ATF1, ATF2, and CRE modulator in a time-dependent manner. Based on chromatin immunoprecipitation analysis, there was an increased association of these proteins with the CYP11B2 promoter after Ang II and K(+) treatment. Phosphorylated ATF/CREB members also bound the CYP11B2 promoter. Knockdown of ATF/CREB members reduced Ang II and K(+) induction of adrenal cell CYP11B2 mRNA expression and aldosterone production. The constitutively active ATF/CREB vectors increased the promoter activity of CYP11B2 and had a synergistic effect with NURR1. In summary, these results suggest that ATF/CREB and NGFI-B family members play a crucial role in the transcriptional regulation of CYP11B2 and adrenal cell capacity to produce aldosterone.