The Regulation of PP1 in the Nucleus

PP1在细胞核中的调控

基本信息

批准号：
8326580
负责人：
Rebecca Page
金额：
$ 29.14万
依托单位：
BROWN UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-01 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8326580
关键词：
Accounting Active Sites Apoptosis Applications Grants Binding Binding Proteins Biochemical Biological Biological Assay Biological Process Cell Cycle Progression Cell Death Cell Nucleus Cells Cellular Stress Cellular biology Chromatin Chromosomes Complex Data Disease Drug Delivery Systems EZH2 gene Embryo Epigenetic Process Eukaryota Eukaryotic Cell Event FHA Domain Genes Genetic Transcription Hand Histone H3 Holoenzymes Human Genome International Investigation MDM2 gene Malignant Neoplasms Metals Molecular Mus N-terminal NMR Spectroscopy Nuclear Nuclear Protein Phosphatase Oncogenes Phosphoric Monoester Hydrolases Phosphorylase a Phosphorylation Phosphotransferases Physical condensation Play Polycomb Post-Translational Protein Processing Process Protein Dephosphorylation Protein Serine/Threonine Phosphatase Protein phosphatase Protein-Serine-Threonine Kinases Proteins RNA Splicing Reaction Recruitment Activity Regulation Reporting Research Research Personnel Research Project Grants Retinoblastoma Protein Roentgen Rays Role Serine Solutions Specificity Spliceosome Assembly Pathway Structure Substrate Specificity Techniques Tertiary Protein Structure Threonine Transcription Repressor/Corepressor Tyrosine Work X-Ray Crystallography base chromatin remodeling genetic regulatory protein histone methyltransferase human disease in vivo inhibitor/antagonist insight mRNA Precursor novel programs protein complex protein phosphatase inhibitor-1 research study response three dimensional structure

项目摘要

DESCRIPTION (provided by applicant): 1/3 of all dephosphorylation reactions are controlled by ser/thr protein phosphatase 1 (PP1), which is present in all eukaryotic cells. PP1 is a single domain metal-binding protein, which lacks any intrinsic specificity. Rather, it is closely regulated by its interaction with >200 confirmed targeting proteins, which localize PP1 to distinct regions of the cell and modulate its substrate specificity. While 1000's of cell biology and biochemical reports describe key biological roles for PP1, only very few structural efforts have so far been successful. Here we describe a complete research plan to understand the regulation of PP1 in the nucleus. The presented research project uses a powerful integrated approach that combines NMR spectroscopy, X-ray crystallography and SAXS with biochemical and in vivo experiments to obtain novel insights into the molecular mechanisms that regulate PP1. Specifically we are focusing on two PP1-targeting proteins: 1) the nuclear inhibitor of PP1 (NIPP1) & 2) the PP1 nuclear targeting subunit (PNUTS). More than 1/3 of the nuclear pool of PP1 forms a holoenzyme with NIPP1. Mice lacking NIPP1 are embryonic lethal and several substrate of NIPP1:PP1 are oncogenes. The NIPP1:PP1 holoenzyme regulates cell cycle progression, epigenetic silencing through chromatin remodeling and pre-mRNA splicing, among other essential biological functions. Thus it is of no surprise that deregulation of the NIPP1:PP1 holoenzyme leads to disease. PNUTS associates with chromatin and promotes chromosome de-condensation. It also controls cell death in response to cellular stresses through the post-translational modification of p53 and MDM2 and it plays essential role in the regulation of the retinoblastoma protein in response to cellular stress, critical processes for the regulation of cancer. In our combined efforts, we will: 1) determine the structures of the free form of these biologically critical PP1 regulators, 2) determine the structures of the PP1 holoenzymes and 3) determine how these complexes direct and regulate PP1 activity. Furthermore, we will leverage these protein and protein complex structures to elucidate, at a molecular level, the biological functions and modes of action of these key nuclear PP1 holoenzymes. The research described in this proposal leverages the extensive expertise of the investigators in the PP1 research field, as well as takes advantage of the best possible national and international collaborators. Furthermore, it has the preliminary data that demonstrates that this work will provide unique, novel insights into the molecular regulation of PP1.

描述（由申请人提供）：所有去磷酸化反应的 1/3 由丝氨酸/苏氨酸蛋白磷酸酶 1 (PP1) 控制，该酶存在于所有真核细胞中。 PP1 是一种单域金属结合蛋白，缺乏任何内在特异性。相反，它受到与超过 200 个已确认的靶向蛋白的相互作用的密切调节，这些蛋白将 PP1 定位到细胞的不同区域并调节其底物特异性。虽然数以千计的细胞生物学和生化报告描述了 PP1 的关键生物学作用，但迄今为止只有极少数的结构研究取得了成功。在这里，我们描述了一个完整的研究计划，以了解 PP1 在细胞核中的调节。所提出的研究项目采用了一种强大的综合方法，将 NMR 波谱学、X 射线晶体学和 SAXS 与生化和体内实验相结合，以获得对调节 PP1 的分子机制的新见解。具体来说，我们关注两种 PP1 靶向蛋白：1) PP1 核抑制剂 (NIPP1) 和 2) PP1 核靶向亚基 (PNUTS)。超过1/3的PP1核库与NIPP1形成全酶。缺乏 NIPP1 的小鼠是胚胎致死的，并且 NIPP1:PP1 的几种底物是癌基因。 NIPP1:PP1 全酶可调节细胞周期进程、通过染色质重塑和前 mRNA 剪接实现表观遗传沉默以及其他重要的生物学功能。因此，NIPP1:PP1 全酶的失调导致疾病也就不足为奇了。 PNUTS 与染色质结合并促进染色体解凝。它还通过 p53 和 MDM2 的翻译后修饰来控制响应细胞应激的细胞死亡，并且它在响应细胞应激的视网膜母细胞瘤蛋白的调节中发挥着重要作用，这是调节癌症的关键过程。在我们的共同努力中，我们将：1) 确定这些生物学关键 PP1 调节剂的游离形式的结构，2) 确定 PP1 全酶的结构，3) 确定这些复合物如何指导和调节 PP1 活性。此外，我们将利用这些蛋白质和蛋白质复合物结构在分子水平上阐明这些关键核 PP1 全酶的生物学功能和作用模式。本提案中描述的研究利用了 PP1 研究领域研究人员的广泛专业知识，并利用了最好的国内和国际合作者。此外，初步数据表明这项工作将为 PP1 的分子调控提供独特、新颖的见解。