Regulation of Protein Dephosphorylation

蛋白质去磷酸化的调节

基本信息

批准号：
6930984
负责人：
Marc C. MUMBY
金额：
$ 31.34万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
1994
资助国家：
美国
起止时间：
1994-08-01 至 2007-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Protein phosphatase 2A (PP2A) is a ubiquitous member of the protein serine/threonine phosphatase gene family functions in fundamental cellular processes including metabolism and the cell cycle. PP2A is composed of core complex containing the catalytic subunit (C) and the scaffold subunit (A), which interacts with a wide variety of regulatory subunits and interacting proteins that target the AC core dimer to specific substrates and intracellular locations. The assembly of many different types of heterotrimeric holoenzymes accounts for the ability of PP2A to regulate a wide range of biological processes. Important targets for PP2A include the neuronal microtubule cytoskeleton, the Cdc6 protein involved in DNA replication, and components of the Wnt signaling pathway. One overall goal of this project is to determine the molecular basis for the actions of PP2A in these processes. Another major goal is to identify novel functions of PP2A by identifying new phosphoprotein targets for the enzyme. The project has four specific aims. 1. Aim 1 will complete the analysis of transgenic mice in which targeting of PP2A in the brain is disrupted by expression of SV4O small tumor antigen. The experiments will test the hypothesis that forms of PP2A targeted to microtubules play an important role in regulating the phosphorylation of the neuron-specific microtubule-associated protein tau. The experiments will also test whether increased phosphorylation of tau can lead to neuronal degeneration. 2. Aim 2 will determine the role of the PR48 subunit in targeting PP2A to the Cdc6 protein. The experiments will test the hypothesis that the interaction of PP2A with Cdc6 is required to maintain Cdc6 in a dephosphorylated state during the GI phase of the cell cycle. Experiment in this aim will also determine the role of calcium in regulating Cdc6 dephosphorylation. 3. Aim 3 is designed to identify novel PP2A substrates and functions using a proteomic approach. Individual PP2A subunits will be knocked out using RNAi. The consequences of the loss of these subunits on cellular signaling will be examined by two-dimensional electrophoresis, identification of proteins by mass spectrometry, and DNA microarrays. 4. Aim 4 will test whether the ABeta subunit of PP2A is a tumor suppressor protein. Wild type Abeta protein and forms of the protein containing mutations identified in lung and colon tumors will be tested for their abilities to downregulate the Wnt signaling pathway and to reverse the tumorigenic potential of lung cancer cell lines

描述（由申请人提供）：蛋白质磷酸酶2a（PP2A）是蛋白质丝氨酸/苏氨酸磷酸酶基因家族的无处不在的成员，包括代谢和细胞周期，包括代谢和细胞周期。 PP2A由包含催化亚基（C）和支架亚基（a）的核心复合物组成，它们与各种调节亚基相互作用和相互作用的蛋白质相互作用，这些蛋白将AC核心二聚体靶向特定的底物和细胞内位置。许多不同类型的异三聚体全酶的组装说明了PP2A调节广泛生物过程的能力。 PP2A的重要靶标包括神经元微管细胞骨架，参与DNA复制的CDC6蛋白以及Wnt信号通路的成分。该项目的总体目标之一是确定PP2A在这些过程中的作用的分子基础。另一个主要目标是通过鉴定酶的新磷蛋白靶标识别PP2A的新功能。该项目具有四个具体目标。 1. AIM 1将完成对脑中pp2a靶向大脑中pp2a的分析，而SV4O小肿瘤抗原的表达破坏了。实验将检验以下假设：针对微管的PP2A形式在调节神经元特异性微管相关蛋白TAU的磷酸化方面起着重要作用。实验还将测试tau的磷酸化增加是否会导致神经元变性。 2. AIM 2将确定PR48亚基在将PP2A靶向CDC6蛋白中的作用。实验将检验以下假设：在细胞周期的GI阶段，PP2A与Cdc6的相互作用与Cdc6的相互作用保持在去磷酸化状态。在此目标中的实验还将决定钙在调节CDC6去磷酸化中的作用。 3. AIM 3旨在使用蛋白质组学方法识别新型PP2A底物和功能。单个PP2A亚基将使用RNAi淘汰。这些亚基在细胞信号传导上丢失的后果将通过二维电泳，通过质谱鉴定蛋白质和DNA微阵列来检查。 4。AIM4将测试PP2A的ABETA亚基是否是肿瘤抑制蛋白。将测试野生型Abeta蛋白和含有肺和结肠肿瘤中鉴定的突变的蛋白质形式，以便其下调Wnt信号传导途径的能力并逆转肺癌细胞系的致瘤潜力