GRIP1, a novel mouse protein that serves as a transcriptional coactivator in yeast for the hormone binding domains of steroid receptors
GRIP1, a novel mouse protein that serves as a transcriptional coactivator in yeast for the hormone binding domains of steroid receptors
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DOI：
10.1073/pnas.93.10.4948
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发表时间：
1996-05-14
影响因子：
11.1
通讯作者：
Stallcup, MR
中科院分区：
文献类型：
--
作者：
Hong, H;Kohli, K;Stallcup, MR
利用酵母双杂交系统从17日龄小鼠胚胎cDNA文库中克隆了一个编码糖皮质激素受体相互作用蛋白1（GRIP 1）的新的812-aa长的蛋白片段，该蛋白可与糖皮质激素受体的激素结合域（HBD）相互作用。在酵母双杂交系统和体外实验中，GRIP 1与糖皮质激素、雌激素和雄激素受体的HBDs相互作用，并受酵母双杂交系统的调控。当与异源蛋白的DNA结合结构域融合时，GRIP 1片段在酵母细胞和哺乳动物细胞中激活了含有合适增强子位点的报告基因，这表明GRIP 1含有转录激活结构域。GRIP 1片段在哺乳动物细胞中的过表达干扰了小鼠乳腺肿瘤病毒氯霉素酰基转移酶基因的表达和巨细胞病毒β-半乳糖苷酶报告基因的组成型表达，但不干扰tRNA基因启动子的组成型表达。这种选择性的抑制活性表明GRIP 1可以与RNA聚合酶II转录机制的重要组成部分相互作用。最后，虽然类固醇受体HBD与GAL 4 DNA结合域融合本身并不激活酵母中报告基因的转录，但该融合蛋白与GRIP 1的共表达强烈激活了报告基因。因此，在酵母中，GRIP 1可以作为共激活因子，增强类固醇受体HBD中的反式激活功能，可能是通过充当受体HBD和基础转录机制之间的桥梁。
The yeast two-hybrid system was used to isolate a clone from a 17-day-old mouse embryo cDNA library that codes for a novel 812-aa long protein fragment, glucocorticoid receptor-interacting protein 1 (GRIP1), that can interact with the hormone binding domain (HBD) of the glucocorticoid receptor. In the yeast two-hybrid system and in vitro, GRIP1 interacted with the HBDs of the glucocorticoid, estrogen, and androgen receptors in a hormone-regulated manner. When fused to the DNA binding domain of a heterologous protein, the GRIP1 fragment activated a reporter gene containing a suitable enhancer site in yeast cells and in mammalian cells, indicating that GRIP1 contains a transcriptional activation domain. Overexpression of the GRIP1 fragment in mammalian cells interfered with hormone-regulated expression of mouse mammary tumor virus-chloramphenicol acyltransferase gene and constitutive expression of cytomegalovirus-beta-galactosidase reporter gene, but not constitutive expression from a tRNA gene promoter. This selective squelching activity suggests that GRIP1 can interact with an essential component of the RNA polymerase II transcription machinery. Finally, while a steroid receptor HBD fused with a GAL4 DNA binding domain did not, by itself, activate transcription of a reporter gene in yeast, coexpression of this fusion protein with GRIP1 strongly activated the reporter gene. Thus, in yeast, GRIP1 can serve as a coactivator, potentiating the transactivation functions in steroid receptor HBDs, possibly by acting as a bridge between HBDs of the receptors and the basal transcription machinery.