Structural basis of PP2A phosphatase diseases

PP2A磷酸酶疾病的结构基础

基本信息

批准号：
10736835
负责人：
Yongna Xing
金额：
$ 36.2万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2027-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10736835
关键词：
Active Sites Adenine Affect Amino Acid Motifs Binding Biochemical Brain CREB1 gene Cancer Patient Catalytic Domain Cell physiology Cellular biology Common Core Complex Cryoelectron Microscopy Cyclic AMP Cyclic AMP-Dependent Protein Kinases Data Diagnosis Disease Dissection Event Family Feedback Forskolin Heart Diseases Holoenzymes Human Intellectual functioning disability Jordan Knowledge Link MAP Kinase Gene Malignant Neoplasms Missense Mutation Molecular Conformation Molecular Diagnosis Mutate Mutation Parkinsonian Disorders Patients Phase Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Play Protein Array Protein Dephosphorylation Protein Deregulation Protein Family Protein phosphatase Proteins Proteome Regulation Regulatory Element Research Resolution Role Sampling Signal Transduction Site Somatic Cell Somatic Mutation Structure Syndrome Testing Variant arm cancer type de novo mutation developmental disease disease-causing mutation early onset exome genetic regulatory protein genome sequencing human disease inhibitor insight member molecular dynamics multidisciplinary mutant nervous system disorder neurodevelopment phosphoric diester hydrolase protein phosphatase 2A regulatory subunit 65 kDa response scaffold simulation single molecule single-molecule FRET whole genome

项目摘要

PROJECT SUMMARY Protein phosphatase 2A (PP2A) is a major Ser/Thr phosphatase with complex regulation and composition. Deregulation of PP2A holoenzymes leads to devastating human diseases, including multiple types of cancer and neurological disorders. In recent years, whole exome/genome sequencing for the molecular diagnosis identified broad disease mutations in PP2A subunits and substrates in cancer and neurological disorders. The diagnosis of de novo PP2A mutations in developmental disorders enthused unprecedented multidisciplinary phosphatase research and interactions with patient families. De novo mutations in several members of B56 family of PP2A regulatory subunits, such as B56δ and B56γ, cause neurological disorders, known as Jordan Syndrome. The same somatic mutations were found in cancer patients. Built on our preliminary cryo-EM, molecular dynamic simulations, biochemical, single molecule, reverse phase protein array, and cell biology studies, here we aim to decipher the mechanisms of B56 disease mutations in cancer and intellectual disabilities. We will determine the cryo-EM structures of WT and disease variants of the PP2A-B56δ holoenzyme (Specific Aim 1), investigate how intellectual disability (ID) mutations alter holoenzyme dynamics, activation phosphorylation, and interactions with regulatory proteins, combining biochemical dissections, advanced molecular dynamic simulation approaches, and single-molecule fluorescence resonance energy transfer (smFRET) (Specific Aim 2), and test our hypothesis on the discriminating and merging mechanisms of B56δ and B56γ ID mutations in perturbing the cAMP negative feedback signaling via MAPK and CREB signaling, respectively (Specific Aim 3). These studies will shed light on a highly dynamic regulation machinery involving the folding of long disordered regions against the holoenzyme core and a super-long dynamic interface harboring the majority of residues mutated in Jordan Syndrome, multiple activation phophorylation sites, and regulatory elements that suppress both the phosphatase active site and the substrate-binding groove.

项目概要蛋白磷酸酶 2A (PP2A) 是一种主要的 Ser/Thr 磷酸酶，具有复杂的调控和组成。 PP2A 全酶的失调会导致毁灭性的人类疾病，包括多种类型的癌症和近年来，用于分子诊断的全外显子组/基因组测序已被确定。癌症和神经系统疾病中 PP2A 亚基和底物的广泛疾病突变的诊断。发育障碍中的从头 PP2A 突变引起了前所未有的多学科磷酸酶的热情 PP2A B56 家族的几个成员的新发突变的研究和互动。调节亚基，例如 B56δ 和 B56γ，会导致神经系统疾病，称为乔丹综合症。基于我们初步的冷冻电镜、分子动力学，在癌症患者中发现了相同的体细胞突变。模拟、生物化学、单分子、反相蛋白质阵列和细胞生物学研究，我们的目标是破译 B56 疾病突变在癌症和智力障碍中的机制我们将确定 WT 的冷冻电镜结构和 PP2A-B56δ 全酶的疾病变体（具体目标 1），如何研究智力障碍（ID）突变改变全酶动力学、激活磷酸化以及与调节蛋白，结合生化解剖、先进的分子动力学模拟方法，和单分子荧光共振能量转移 (smFRET)（具体目标 2），并检验我们的假设关于 B56δ 和 B56γ ID 突变干扰 cAMP 阴性的区分和合并机制分别通过 MAPK 和 CREB 信号传导反馈信号（具体目标 3）。关于高度动态的调节机制，涉及长无序区域的折叠全酶核心和超长动态界面包含约旦大部分突变残基综合征、多重激活磷酸化位点以及抑制磷酸酶的调节元件活性位点和底物结合槽。