Project 2- Mechanistic Role of Talin in Cellular Signaling

项目 2 - Talin 在细胞信号转导中的机制作用

• 批准号: 10268698
• 负责人: JUN QIN
• 金额: $ 52.81万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10268698
• 关键词: Ablation; Actin-Binding Protein; Actins; Adhesions; Affinity; Binding; Biochemical; Blood; Blood Vessels; Blood coagulation; C-terminal; Cell Adhesion; Cells; Communication; Coupled; Cytoskeleton; Cytosol; Data; Development; Diagnosis; Disease; Event; Extracellular Matrix; Face; Genetic study; Head; Impairment; Integrin Binding; Integrin alpha Chains; Integrins; Laboratories; Lead; Length; Ligands; Link; Mechanics; Mediating; Membrane; Microfilaments; Molecular Conformation; Monomeric GTP-Binding Proteins; N-terminal; Pathologic Processes; Phosphatidylinositol 4,5-Diphosphate; Physiological Processes; Pilot Projects; Process; Proteins; PubMed; Rod; Role; Signal Transduction; Stroke; Structure; Talin; Testing; Therapeutic; Thrombosis; Ubiquitin; angiogenesis; cell motility; insight; mutant; novel; paxillin; receptor; recruit

Project Summary The long-term objective of this proposal is to understand the activation and signaling mechanisms of integrins – a class of heterodimeric transmembrane receptors that control cell-extracellular matrix (ECM) adhesion and diverse adhesion-dependent physiological and pathological processes such as angiogenesis, blood clotting, thrombosis, and stroke. Discovered four decades ago, integrins have been intensively studied with nearly 75,000 articles in PubMed. Tremendous effort has been made by many laboratories to investigate the mechanism of integrin activation – the first key step to initiate the cell-ECM adhesion, which led to the discovery and elucidation of a key integrin activator talin – a large actin-binding protein containing an N-terminal head domain (talin-H) and a C-terminal rod domain (Talin-R). It is now widely believed that talin utilizes its talin-H to bind integrin cytoplasmic face to elicit an “inside-out” conformational signal to activate the receptor. However, a recent pilot study from our laboratory revealed unexpectedly that a constitutively active full length talin is much more potent than talin-H to activate integrins, challenging the talin-H-only premise for controlling the integrin activation. Moreover, intact talin is autoinhibited in the cytosol and how it is recruited/activated to activate integrin and link integrin to actin (outside-in signaling) for promoting cytoskeleton reassembly and dynamic adhesion remains elusive. We propose to resolve these puzzling issues in three highly integrated aims. Aim1 will elucidate how a small GTPase Rap1 cooperatively binds talin F0 and F1 domains of talin-H to facilitate the talin recruitment and how a nascent adhesion adaptor paxillin synergizes with PIP2 to activate talin. Aim2 will investigate that after activating talin, how paxillin further regulates the talin function by bridging its talin-H as well as talin-R to another integrin activator kindlin-2 to promote potent integrin activation. The results would provide novel insight into how talin R contributes significantly to the integrin activation. Aim3 will examine how talin mediates integrin signaling to actin cytoskeleton – a long speculated integrin outside-in process that has remained poorly elucidated. Together, our proposed studies are expected to lead to a new paradigm for understanding the cell adhesion mechanisms by providing a comprehensive view of how talin is regulated to undergo a sequence of dynamic events to activate integrin and link it to actin filaments leading to cytoskeleton reassembly and dynamic adhesion. Given the critical involvement of talin and its associated proteins in thrombosis, stroke, etc., our studies may ultimately lead to the development of better diagnosis and therapeutics for these diseases.

项目摘要 本提案的长期目标是了解整合素的激活和信号传导机制- 一类控制细胞-细胞外基质（ECM）粘附的异二聚体跨膜受体， 多种粘附依赖性生理和病理过程如血管生成，血液凝固， 血栓和中风整合素是四十年前发现的，已经被深入研究了近75，000种整合素。 PubMed上的文章许多实验室已经做出了巨大的努力来研究 整合素活化-启动细胞-ECM粘附的第一个关键步骤，导致发现和阐明 一种关键的整合素激活剂talin -一种含有N-末端头部结构域的大肌动蛋白结合蛋白（talin-H）， C-末端杆状结构域（Talin-R）。现在普遍认为talin利用其talin-H结合整合素 细胞质面的“内-外”构象信号，以激活受体。然而，最近一名飞行员 我们实验室的研究出乎意料地揭示了组成型活性全长talin的效力要强得多， 比talin-H激活整联蛋白，挑战了仅talin-H控制整联蛋白激活的前提。 此外，完整的talin在胞质溶胶中是自抑制的，以及它如何被募集/激活以激活整合素和连接蛋白。 用于促进细胞骨架重组和动态粘附的整联蛋白到肌动蛋白（由外向内信号传导）仍然存在 难以捉摸。我们建议在三个高度综合的目标中解决这些令人困惑的问题。AIM 1将阐明 小的GTdR Rap 1协同结合talin-H的talin F0和F1结构域以促进talin募集， 新生粘附适配器桩蛋白如何与PIP 2协同激活塔林蛋白。AIM 2将在 激活talin，桩蛋白如何通过桥接talin-H和talin-R进一步调节talin功能， 整合素激活剂Kindlin-2促进有效的整合素激活。这些结果将为我们提供新的见解， talin R对整联蛋白活化有显著贡献。aim 3将研究talin如何介导整合素信号传导 到肌动蛋白细胞骨架-一个长期推测的整合素由外向内的过程，仍然很难阐明。 总之，我们提出的研究有望导致一个新的范式，了解细胞粘附 通过提供全面的看法，塔林是如何调节，以经历一系列的动态机制， 事件激活整联蛋白并将其连接到肌动蛋白丝，导致细胞骨架重组和动态粘附。 考虑到talin及其相关蛋白质在血栓形成、中风等中的重要参与，我们的研究可能 最终导致对这些疾病更好的诊断和治疗的发展。