Protein methyltransferases as transcription coactivators

描述(申请人提供)：核受体(NR)通过募集辅活化子复合体来激活转录，该复合体局部重塑启动子中的染色质结构，并帮助招募和激活RNA聚合酶II复合体。多个共激活子的复合体参与转录起始，一些被招募的辅助激活子也可能促进随后的转录延伸和RNA加工的耦合步骤。有明确的证据表明，一个共激活复合体具有生理学意义，它包括p160家族(GRIP1、SRC-1和p/CIP)、蛋白质乙酰转移酶p300和CBP，以及蛋白质精氨酸甲基转移酶CARM1和PRMT1。P160共激活子直接与NRs结合，并招募乙酰基转移酶和甲基转移酶，它们修饰组蛋白和转录复合体中的其他蛋白质。另一种辅活化子PGC-1是一种组织特异性辅活化子，它在肝脏、棕色脂肪和肌肉中的表达水平在控制参与糖异生和产热等过程的基因表达方面发挥了关键作用。我们实验室和其他实验室对这些特定的共激活因子的研究一直处于促进我们对转录调控复杂机制的理解的前沿，但甲基转移酶及其对组蛋白和其他蛋白质的甲基化的具体机制尚不清楚。拟议的研究将集中在这些甲基转移酶作为共激活因子的作用以及它们与p300/CBP和PGC-I协同合作的机制。我们将确定：1)CARM1和PRMT1分别对组蛋白H3和H4进行精氨酸特异性甲基化，通过染色质免疫沉淀和鉴定优先与甲基化后的组蛋白结合的蛋白质，在转录激活中的作用；2)CARM1的功能结构域，它们在共激活功能中的特定作用，以及与CARM1关键结构域结合的蛋白质；3)CARM1-p300/CBP和PRMT1-PGC-1协同激活的机制以及CARM1对p300/CBP的甲基化和PRMT1对PGC-1的协同作用；4)SR及其共激活子的亚核定位和共定位，SR及其共激活子在体内稳定整合的类固醇反应基因启动子上的组装，以及在体外SR对重组染色质的染色质重塑中的作用。

DESCRIPTION (provided by applicant): Nuclear receptors (NR) activate transcription by recruiting complexes of coactivators which locally remodel chromatin structure in the promoter and help to recruit and activate an RNA polymerase II complex. Multiple complexes of coactivators participate in transcription initiation, and some recruited coactivators may also facilitate the subsequent coupled steps of transcription elongation and RNA processing. There is clear evidence for physiological relevance of one coactivator complex, which includes the p160 family (GRIP1, SRC-1, and p/CIP), the protein acetyltransferases p300 and CBP, and the protein arginine methyltransferases CARM1 and PRMT1. The p160 coactivators bind directly to NRs and recruit the acetyltransferases and methyltransferases, which modify histones and other proteins in the transcription complex. Another coactivator, PGC-1, is an example of a tissue specific coactivator whose level of expression in liver, brown fat, and muscle plays a key role in controlling the expression of genes involved in processes such as gluconeogenesis and thermogenesis. The studies of these specific coactivators, in our lab and others, have been on the forefront in advancing our understanding of the complex mechanism of transcriptional regulation, but the specific mechanistic roles of the methyltransferases and their methylation of histones and other proteins is not clear. The proposed studies will focus on the roles of these methyltransferases as coactivators and their mechanism of synergistic cooperation with p300/CBP and with PGC-I. We will define: 1) the roles of arginine-specific methylation of histones H3 and H4 by CARM1 and PRMT1, respectively, in transcriptional activation, by using chromatin immunoprecipitation and by identifying proteins which preferentially bind to the methylated histones; 2) the functional domains of CARM1, their specific roles in coactivator function, and proteins that bind to critical domains of CARM1; 3) the mechanisms of CARM1-p300/CBP and PRMT1-PGC-1 coactivator synergy and the roles which methylation of p300/CBP by CARM1 and of PGC-1 by PRMT1 play in synergy; 4) the sub-nuclear locations and co-localization of SR and their coactivators, the assembly of SR and their coactivators onto stably integrated promoters of steroid responsive genes in vivo, and the role of coactivators in chromatin remodeling by SR on reconstituted chromatin in vitro.