Transcriptional activation of human CYP17 in H295R adrenocortical cells depends on complex formation among p54nrb/NonO, protein-associated splicing factor, and SF-1, a complex that also participates in repression of transcription
Transcriptional activation of human CYP17 in H295R adrenocortical cells depends on complex formation among p54nrb/NonO, protein-associated splicing factor, and SF-1, a complex that also participates in repression of transcription
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DOI：
10.1210/en.143.4.1280
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发表时间：
2002-04-01
期刊：
影响因子：
4.8
通讯作者：
Waterman, MR
中科院分区：
文献类型：
--
作者：
Sewer, MB;Nguyen, VQ;Waterman, MR
人CYP 17（hCYP 17）基因转录起始位点上游的前57 bp对于基础转录和cAMP依赖性转录都是必需的。用放射性标记的hCYP 17启动子的-57/-38探针孵育H295 R肾上腺皮质细胞核提取物，进行EMSA，结果显示形成了三种DNA-蛋白复合物。最快的复合物含有类固醇生成因子（SF-1）和p54（nrb）/ NonO，中间复合物含有p54（nrb）/NonO和多聚嘧啶片段结合蛋白相关剪接因子（PSF），最慢的复合物含有SF-1/PSF/p54（nrb）/NonO复合物。（BU）（2）cAMP处理导致cAMP诱导的仅上层复合物结合强度的增加，并且还激活了hCYP 17基因转录。SF-1与p54（nrb）/NonO共免疫沉淀，表明这些蛋白质之间直接相互作用。功能分析表明，PSF抑制基础转录。此外，hCYP 17启动子-报告基因构建体荧光素酶活性的抑制是由于PSF与辅阻遏物mSin 3A相互作用所致。曲古抑菌素A减弱了对基础转录的抑制，表明组蛋白脱乙酰酶与SF-1/PSF/p54（nrb）/NonO/mSin 3A复合物相互作用。我们的研究支持这样的想法，即转录激活和抑制之间的平衡是必不可少的控制肾上腺皮质类固醇激素的生物合成。
The first 57 bp upstream of the transcription initiation site of the human CYP17 (hCYP17) gene are essential for both basal and cAMP-dependent transcription. EMSA carried out by incubating H295R adrenocortical cell nuclear extracts with radiolabeled -57/-38 probe from the hCYP17 promoter showed the formation of three DNA-protein complexes. The fastest complex contained steroidogenic factor (SF-1) and p54(nrb)/ NonO, the intermediate complex contained p54(nrb)/NonO and polypyrimidine tract-binding protein-associated splicing factor (PSF), and the slowest complex contained an SF-1/PSF/p54(nrb)/NonO complex. (BU)(2)cAMP treatment resulted in a cAMP-inducible increase in the binding intensity of only the upper complex and also activated hCYP17 gene transcription. SF-1 coimmunoprecipitated with p54(nrb)/NonO, indicating direct interaction between these proteins. Functional assays revealed that PSF represses basal transcription. Further, the repression of hCYP17 promoter-reporter construct luciferase activity resulted from PSF interacting with the corepressor mSin3A. Trichostatin A attenuated the inhibition of basal transcription, suggesting that a histone deacetylase interacts with the SF-1/PSF/p54(nrb)/NonO/mSin3A complex. Our studies lend support to the idea that the balance between transcriptional activation and repression is essential in the control of adrenocortical steroid hormone biosynthesis.