Structural Dynamics of Vinculin in Adhesion Junctions

粘着连接中纽蛋白的结构动力学

基本信息

批准号：
8208009
负责人：
TINA IZARD
金额：
$ 40.33万
依托单位：
SCRIPPS FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-08-01 至 2013-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8208009
关键词：
Abbreviations Actinin Actins Adaptor Signaling Protein Address Adherens Junction Adhesions Affect Affinity Binding Biochemical Biological Cadherins Cell Nucleus Cell Survival Cell surface Cell-Cell Adhesion Cells Coin Complex Crystallization Cytoskeletal Proteins Cytoskeleton DNA Data Development Dimerization Disease Disseminated Malignant Neoplasm Environment F-Actin Focal Adhesions Foundations Funding Head Health Human Hydrophobic Interactions Integrins Ischemia Length Link Maps Messenger RNA Microfilaments Modeling Molecular Conformation Muscle Muscle Cells Mutation Myopathy N-terminal Null Lymphocytes Phosphatidylinositol 4,5-Diphosphate Phospholipids Physiological Play Plus End of the Actin Filament Polypyrimidine Tract-Binding Protein Process Production Progress Reports Proline Proline-Rich Domain Property Protein Isoforms Proteins RNA RNA Recognition Motif RNA Sequences RNA Splicing RRM1 gene Reaction Resolution Role SH3 Domains Signal Transduction Site Solutions Structure Surface Tail Talin Testing Translations Tropomyosin Vinculin cell behavior cell growth cell motility crosslink experience extracellular feeding globular protein improved insight metavinculin mutant next generation novel polymerization protein protein interaction receptor response scaffold vasodilator-stimulated phosphoprotein

项目摘要

DESCRIPTION (provided by applicant): The formation of cell-matrix (focal adhesions) and cell-cell (adherens junctions) adhesion complexes links signals at the cell surface to the actin cytoskeleton, and these direct cell migration, cell growth and survival, and the morphological changes that are needed for proper development. Signaling from focal adhesions or adherens junctions is directed by integrin or cadherin transmembrane receptors, respectively, and their links to the actin cytoskeleton require activation of the cytoskeletal protein vinculin, which binds to proteins that directly interact with these receptors, such as talin, a-actinin, and a-catenin, as well as to components of the machinery that controls cell migration. During the past funding cycle of GM071596 we defined the structure of the closed, inactive conformation of vinculin, which is compromised of five, loosely-packed helical bundle domains that are held in a closed-clamp conformation via extensive hydrophobic interactions of its N-terminal seven-helical bundle (Vh1) domain with its five-helical bundle tail (Vt) domain. Our studies also defined the atomic changes that accompany vinculin activation, where the Vh1 domain undergoes remarkable structural changes that displace Vt domain from a distance, and which release the domains of vinculin to allow binding to its partners. Finally, we demonstrated that talin and a-actinin are physiological triggers that can activate vinculin, and that they must first undergo structural alterations to bind to and activate vinculin, establishing that adhesion signaling involves a chain reaction of structural alterations. While these surprising and exciting advances defined the mechanism and structural alterations that control vinculin activation, very little is known regarding how activated vinculin binds to its numerous partners, or how it directs such diverse processes throughout the cell. Here we propose to address these important questions in a head-on fashion, together with functional studies by solving the crystal structures of activated vinculin in complex with three partners that control cell adhesion and cell migration, and the localized production of adhesion components at nascent junctions. Further, we will solve the crystal structure of metavinculin, an isoform of vinculin that is exclusively expressed in muscle tissue, and we will also define the interactions that are required for metavinculin function. Collectively, the proposed studies will resolve how activated vinculin and metavinculin direct their diverse functions, and they will lay the foundation for targeting their interactions for the treatment of diseases having vinculin or metavinculin involvement, in particular metastatic cancer, ischemia, and myopathies. PUBLIC HEALTH RELEVANCE Cells require distinct adhesion complexes at their cell surface to form contacts with their neighbors or with the extracellular environment, and the protein vinculin plays essential roles in linking these adhesion complexes to the actin cytoskeleton, and in directing the cell migration machinery. The formation of these links requires that vinculin transition from its closed, inactive conformation to its activated state, and the studies supported by R01 GM071596 defined the structure of inactive and activated vinculin, and revealed its mechanism of activation. However, essentially nothing is known regarding the interactions of activated vinculin with its binding partners in the cell, and our new studies in this revised competitive renewal application of R01 GM071596 will define the structure and function of vinculin in complex with three of its partners that play essential roles in adhesion complexes, in cell migration, and in the localized production of components of adhesion junctions. Finally, we will also define the structure and function of metavinculin, an isoform of vinculin that plays essential roles in the formation and function of muscle tissue.

描述（由申请人提供）：形成细胞 - 矩阵（焦点粘附）和细胞细胞（粘附连接）粘附复合物将细胞表面的信号与肌动蛋白细胞骨架联系起来，以及这些直接的细胞迁移，细胞生长和存活以及正确发育所需的形态变化。 Signaling from focal adhesions or adherens junctions is directed by integrin or cadherin transmembrane receptors, respectively, and their links to the actin cytoskeleton require activation of the cytoskeletal protein vinculin, which binds to proteins that directly interact with these receptors, such as talin, a-actinin, and a-catenin, as well as to components of the machinery that controls cell迁移。 During the past funding cycle of GM071596 we defined the structure of the closed, inactive conformation of vinculin, which is compromised of five, loosely-packed helical bundle domains that are held in a closed-clamp conformation via extensive hydrophobic interactions of its N-terminal seven-helical bundle (Vh1) domain with its five-helical bundle tail (Vt) domain.我们的研究还定义了伴随Vinculin激活的原子变化，其中VH1域经历了显着的结构变化，从远处将VT域置换，并释放Vinculin的域以允许与其伴侣结合。最后，我们证明塔林和a-肌动蛋白是可以激活vinculin的生理触发因素，并且必须首先进行结构改变以结合和激活vinculin，以确定粘附信号涉及结构改变的链反应。尽管这些令人惊讶且令人兴奋的进步定义了控制Vinculin激活的机制和结构变化，但对于激活的Vinculin如何与其众多伴侣结合，或者它如何指导整个细胞中的这种多样化过程。在这里，我们建议以正面方式解决这些重要问题，以及功能研究，通过与控制细胞粘附和细胞迁移的三个伴侣以及在新生交界处的粘附成分的局部生产来解决复合物中激活的vinculin的晶体结构。此外，我们将求解元环蛋白的晶体结构，Metavinculin是一种仅在肌肉组织中表达的Vinculin的同工型，我们还将定义元蛋白素功能所需的相互作用。总的来说，拟议的研究将解决激活的质蛋白和元元素如何指导其多种功能，并将为目标相互作用奠定基础，以治疗患有vinculin或metavinculin受累的疾病，特别是转移性癌症，缺血和肌病。公共卫生相关性细胞在其细胞表面需要不同的粘附复合物，以与邻居或细胞外环境形成接触，并且蛋白质vinculin在将这些粘附复合物与肌动蛋白细胞骨架之间以及指导细胞迁移机械方面起着至关重要的作用。这些链接的形成要求vinculin从其封闭的，无活跃的构象转变为其活化状态，而R01 GM071596支持的研究定义了非活性和活化的Vinculin的结构，并揭示了其激活的机制。但是，基本上，关于活化的杂种蛋白与其结合伴侣在细胞中的相互作用的相互作用以及我们在此修订的R01 gm071596的竞争更新应用中的新研究中，尚无任何了解，将定义vinculin在粘附复合物中在粘附复合物中起着重要作用，在迁移和本地生产中起作用的成分的基本伴侣的结构和功能。最后，我们还将定义元环蛋白的结构和功能，metavinculin是一种在肌肉组织的形成和功能中起着至关重要的作用。