Nuclear Receptor Coactivators GRIP1, CoCoA and Flil

描述(申请人提供)：激素激活的核受体(NR)与目标基因启动子结合，并招募辅助激活子的复合体来帮助激活转录。P160共激活子复合体由一个直接与激活的NRs结合的p160蛋白和几个次级共激活子组成。我们的总体策略是确定辅激活子的功能结构域，并寻找与其激活结构域结合的蛋白质，并在辅激活子信号通路中介导下游事件。通过重复应用这一过程，我们将发现p160共激活因子在染色质或基础转录机制中的最终靶点。先前研究表明，p160蛋白有两个C端激活结构域用于信号传递：AD1与CBP/p300结合，而AD2与CARM1结合。我们最近在p160共激活子(AD3)的N-末端发现了一个新的激活域，并发现了两种与p160结合并协同作用的蛋白质：卷曲共激活子(CocoA)和无飞行I(Flil)。在这里，我们将确定这些蛋白中的每一个与其他NR辅助激活子合作并促进NRs转录激活的机制。我们将通过定义可可和Flil与NRs、p160共激活子和/或其他已知蛋白质相互作用伙伴的相互作用位点来定义可可和Flil的功能亚域。我们还将定义它们的激活域，用于将激活信号传递到转录机制，并识别与激活域结合的蛋白质，从而在激活信号通路的下游。可可和Flil作为共激活因子与其他类别的转录因子的参与也将被调查。最后，我们将研究A549细胞对可可和Flil的不同需求以及它们在多个激素调节启动子(一些激素诱导的和一些激素抑制的)启动子上的不同功能域，以探讨共激活功能的机制与启动子结构的关系。

DESCRIPTION (provided by applicant): Hormone-activated nuclear receptors (NR) bind to target gene promoters and recruit complexes of coactivators to help activate transcription. The p160 coactivator complex consists of a p160 protein, which binds directly to activated NRs, and several secondary coactivators. Our global strategy is to characterize the functional domains of coactivators and search for proteins that bind to their activation domains and mediate downstream events in the coactivator signaling pathway. By reiterative application of this process, we will find the ultimate targets of the p160 coactivators in the chromatin or basal transcription machinery. The p160 proteins were previously shown to have two C-terminal activation domains for signal transmission: AD1 binds CBP/p300, while AD2 binds CARM1. We recently identified a new activation domain in the N-terminal region of p160 coactivators (AD3) and identified two proteins which bind to the p160 N-terminus and cooperate synergistically with p160 proteins as coactivators for NRs: Coiled-coil Coactivator (CoCoA) and Flightless I (Flil). Here, we will determine the mechanisms by which each of these proteins cooperates with other NR coactivators and contributes to transcriptional activation by NRs. We will define the functional subdomains of CoCoA and Flil by defining their sites of interaction with NRs, p160 coactivators, and/or other known protein interaction partners. We will also define their activation domains, which are used to transmit the activating signal to the transcription machinery, and identify proteins that bind to the activation domains and are thus downstream in the activation signaling pathway. The involvement of CoCoA and Flil as coactivators with other classes of transcription factors will also be investigated. Finally, we will study the variable requirements for CoCoA and Flil and their various functional domains on multiple steroid hormone-regulated promoters (some hormone inducible and some hormone repressible) in the A549 cell line to investigate the relationship between mechanism of coactivator function and promoter architecture.