Identification of a co-repressor that inhibits the transcriptional and growth-arrest activities of CCAAT/enhancer-binding protein α
Identification of a co-repressor that inhibits the transcriptional and growth-arrest activities of CCAAT/enhancer-binding protein α
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DOI：
10.1074/jbc.m512734200
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发表时间：
2006-06-30
影响因子：
4.8
通讯作者：
Roesler, William J.
中科院分区：
文献类型：
--
作者：
McFie, Pamela J.;Wang, Guo-Li;Roesler, William J.
我们使用酵母双杂交筛选方法来鉴定CCAAT/增强子结合蛋白α(C/EBPα)的新的相互作用元件，这可能为深入了解其作用和调控机制提供依据。获得的一个克隆是CA150的克隆，这是一种核蛋白，以前的特征是转录延伸因子。在这份报告中，我们证明了CA150是一种广泛表达的C/EBP蛋白的共抑制物。双杂交和免疫共沉淀分析表明CA150与C/EBPα相互作用。CA150的过表达抑制了C/EBPα介导的反式激活，并能逆转共激活子p300对C/EBPβ介导的反式激活的增强作用。对C/EBPα突变体的分析表明，CA150主要通过跨越135-150个氨基酸的结构域与C/EBPα相互作用。染色质免疫沉淀分析表明，CA150存在于C/EBPα抑制的启动子上，而不存在于C/EBPα激活的启动子上。最后，我们发现，在异位表达C/EBPα导致生长停滞的细胞中，CA150能够将它们从生长停滞中释放出来。有趣的是，CA150不能逆转C/EBPα的最小生长停滞结构域(氨基酸175-217)所产生的生长停滞，这表明CA150的作用针对这个最小结构域之外的C/EBPα区域，与我们的双杂交分析一致。综上所述，这些数据表明CA150是C/EBP蛋白的辅助抑制物，并为C/EBPα如何抑制特定基因的转录提供了可能的机制。
We used a yeast two-hybrid screening approach to identify novel interactors of CCAAT/enhancer-binding protein alpha (C/EBP alpha) that may offer insight into its mechanism of action and regulation. One clone obtained was that for CA150, a nuclear protein previously characterized as a transcriptional elongation factor. In this report, we show that CA150 is a widely expressed co-repressor of C/EBP proteins. Two-hybrid and co-immunoprecipitation analyses indicated that CA150 interacts with C/EBP alpha. Overexpression of CA150 inhibited the transactivation produced by C/EBP alpha and was also able to reverse the enhancing effect of the co-activator p300 on C/EBP beta-mediated transactivation. Analysis of C/EBP alpha mutants indicated that CA150 interacts with C/EBP alpha primarily through a domain spanning amino acids 135-150. Chromatin immunoprecipitation assays showed that CA150 was present on a promoter that is repressed by C/EBP alpha but not present on a promoter that is activated by C/EBP alpha. Finally, we showed that in cells in which growth arrest had been induced by ectopic expression of C/EBP alpha, CA150 was able to release them from growth arrest. Interestingly, CA150 could not reverse the growth arrest produced by the minimal growth-arrest domain of C/EBP alpha (amino acids 175-217), suggesting that the effect of CA150 was directed at a region of C/EBP alpha outside of this minimal domain, consistent with our two-hybrid analysis. Taken together, these data indicate that CA150 is a co-repressor of C/EBP proteins and provides a possible mechanism for how C/EBP alpha can repress transcription of specific genes.