Spatial control of actin assembly by phosphoinositides

磷酸肌醇对肌动蛋白组装的空间控制

基本信息

批准号：
8962478
负责人：
Paul A Janmey
金额：
$ 44.35万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2019-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8962478
关键词：
Actin-Binding Protein Actins Affect Atomic Force Microscopy Binding Binding Proteins Binding Sites Biochemical Biochemistry Biological Biophysics Brain Cell membrane Cell physiology Cells Cellular biology Cereals Cholesterol Collaborations Computer Analysis Cytoskeleton Data Defect Disease Electron Microscopy Electrostatics Elements Employee Strikes Enzymes Event Fluorescence Microscopy Gelsolin Goals In Vitro Isomerism Lateral Lead Ligands Link Lipids Liquid substance Malignant Neoplasms Mechanics Mediating Membrane Membrane Lipids Metabolism Methods Microfilaments Modeling Mole the mammal Molecular Conformation Mutation Nerve Degeneration Neurons Neuropathy Phase Phase Transition Phosphatidylinositol 4,5-Diphosphate Phosphatidylinositols Phospholipids Physiological Production Property Protein Binding Proteins Regulation Resolution Signal Transduction Specificity Structure Surface System Temperature Testing Vesicle Work aqueous base biophysical analysis cell motility cell type computer studies design experience human disease in vivo membrane model molecular dynamics monolayer nanoscale novel strategies polymerization pressure protein activation public health relevance quantum spectroscopic imaging spectroscopic survey

项目摘要

DESCRIPTION (provided by applicant): Phosphoinositides are membrane phospholipids that control many cellular events and bind with variable levels of specificity to dozens of intracellula proteins. Defects in production or metabolism of these anionic phospholipids are associated with cancer, neuronal defects, and other diseases. The most abundant phosphoinositide in most cells types, PI4,5P2, is particularly important for regulation of cytoskeletal assembly during cell motility, differentiation and proliferation largely through its effects on numerous actin binding proteins including gelsolin, N-WASP, and formins, which are the focus of this project. Other isomers of PI4,5P2, such as PI3,5P2, are associated with neurodegeneration in several genetically distinct diseases. How specific phosphoinositides affect their ligands is much less understood than are the mutations that produce abnormal phosphoinositide production. Defining how these lipids exert their biological control at the membrane-cytoskeletal interface could lead to new approaches to limiting or reversing the abnormal function of these lipids in disease. Previous work and preliminary data show that PI4,5P2 at low mole fraction in membranes forms nanoscale clusters in the presence of physiologically relevant levels of Ca2+ and partitions into the liquid disordered phase when membranes undergo fluid phase transitions due to changes in cholesterol content or temperature. Redistribution of PI4,5P2 into these nanodomains alters its ability to nucleate actin assembly from brain extracts and to inhibit gelsolin, the actin filament severing protein. The goal of this project is to quantitatively define the conditions under which PI4,5P2 reorganizes into nanoscale membrane domains using a combination of high resolution imaging, spectroscopy, and molecular dynamics computations and to relate changes in PI4,5P2 membrane distribution to its ability to inhibit actin assembly. Biochemical analyses will test which elements of the actin regulatory system are affected when PI4,5P2 redistributes in membranes. This project involves collaboration among three groups with complementary experience in membrane biophysics, gelsolin biochemistry and the mechanics of actin polymerization; computational studies of membrane structure and mechanics; and electron microscopy with emphasis on high resolution studies of actin assembly at membranes. The multi-disciplinary study will lead to an atomic level understanding of phosphoinositide-protein interactions that will help direct strategies designed to alter phosphoinositide production, distribution, and signaling in the numerous contexts where their altered expression or distribution is linked to disease.

描述（由申请人提供）：磷脂肌醇是控制许多细胞事件的膜磷脂，并以不同水平的特异性与数十种细胞内蛋白质结合。这些阴离子磷脂的产生或代谢缺陷与癌症、神经元缺陷和其他疾病有关。大多数细胞类型中最丰富的磷酸肌醇 PI4、5P2，对于细胞发育过程中细胞骨架组装的调节特别重要 PI4,5P2 的其他异构体（例如 PI3,5P2）与神经变性相关，主要通过其对多种肌动蛋白结合蛋白（包括凝溶胶蛋白、N-WASP 和福尔明）的影响来实现运动、分化和增殖。与确定这些脂质如何发挥其生物控制作用相比，对特定磷酸肌醇如何影响其配体的了解要少得多。膜-细胞骨架界面上的PI4,5P2可能会导致限制或逆转这些脂质在疾病中的异常功能的新方法，先前的工作和初步数据表明，膜中低摩尔分数的PI4,5P2在生理相关水平存在的情况下形成纳米级簇。当膜因胆固醇含量或温度的变化而发生流体相变时，Ca2+ 会重新分布到液体无序相中，PI4,5P2 重新分布到这些纳米域中会改变其能力。使脑提取物中的肌动蛋白组装成核并抑制凝溶胶蛋白（肌动蛋白丝切断蛋白）该项目的目标是结合高分辨率成像、光谱学和技术来定量定义 PI4,5P2 重组为纳米级膜结构域的条件。分子动力学计算并将 PI4,5P2 膜分布的变化与其抑制肌动蛋白组装的能力联系起来。生化分析将测试肌动蛋白调节系统的哪些元件受到影响。该项目涉及膜生物物理学、凝溶胶蛋白生物化学和肌动蛋白聚合力学方面经验互补的三个小组的合作；膜结构和力学以及电子显微镜的计算研究，重点是肌动蛋白组装的高分辨率研究；这项多学科研究将导致对磷酸肌醇-蛋白质相互作用的原子水平理解，这将有助于指导旨在改变磷酸肌醇产生、分布和作用的策略。在许多情况下，它们的表达或分布改变与疾病相关。