The Regulation of PP1 in the Nucleus

PP1在细胞核中的调控

基本信息

批准号：
8728948
负责人：
Rebecca Page
金额：
$ 29.08万
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/8728948
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 Targeting 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均由所有真核细胞中存在的Ser/Thr蛋白磷酸酶1（PP1）控制。 PP1是一种单域金属结合蛋白，缺乏任何内在特异性。相反，它与> 200个确认的靶向蛋白的相互作用密切调节，该蛋白将PP1定位在细胞的不同区域并调节其底物特异性。虽然1000个细胞生物学和生化报告描述了PP1的关键生物学作用，但到目前为止，只有很少的结构努力才成功。在这里，我们描述了一个完整的研究计划，以了解核中PP1的调节。提出的研究项目采用了强大的集成方法，该方法将NMR光谱，X射线晶体学和SAX与生化和体内实验结合在一起，以获得对调节PP1的分子机制的新见解。具体而言，我们专注于两个靶向PP1的蛋白质：1）PP1（NIPP1）和2）PP1核靶向亚基（Pnuts）的核抑制剂。 PP1的核池超过1/3，与Nipp1形成全酶。缺乏NIPP1的小鼠是胚胎致死的，而Nipp1：pp1的几种底物是癌基因。 NIPP1：PP1全酶调节细胞周期的进展，通过染色质重塑和前MRNA剪接以及其他必要的生物学功能来调节细胞周期的进展，表观遗传沉默。因此，对NIPP1：PP1全酶导致疾病的放松管制毫不奇怪。 Pnuts与染色质相关并促进染色体脱凝结。它还通过p53和MDM2的翻译后修饰来控制细胞死亡，以响应细胞应激，并且它在响应细胞应激的调节性切诺细胞瘤蛋白的调节中起着至关重要的作用，这是癌症调节的关键过程。在我们的综合努力中，我们将：1）确定这些生物学上关键的PP1调节剂的自由形式的结构，2）确定PP1全酶的结构，3）确定这些复合物如何直接和调节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方式。该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，并利用了最好的国家和国际合作者。此外，它具有初步数据，该数据表明这项工作将为PP1的分子调节提供独特的新颖见解。