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 这项建议的长期目标是了解整合素的激活和信号传递机制。 一类异二聚体跨膜受体，控制细胞-细胞外基质(ECM)黏附和 不同的黏附依赖的生理和病理过程，如血管生成，血液凝结， 血栓形成和中风。整合素在40年前被发现，目前已有近75,000人进行了深入的研究 发表在PubMed上的文章。许多实验室已经做出了巨大的努力来研究 整合素激活-启动细胞-ECM黏附的第一个关键步骤，这导致了发现和阐明 关键的整合素激活剂talin-一种包含N-末端头域(talin-H)的大肌动蛋白结合蛋白和 C-末端杆状结构域(Talin-R)。现在人们普遍认为talin利用其talin-H结合整合素 细胞质表面产生一个“由内而外”的构象信号来激活受体。然而，最近的一项试点 我们实验室的研究出人意料地显示，具有结构性活性的全长Talin的效力要强得多 而不是talin-H来激活整合素，挑战了talin-H唯一控制整合素激活的前提。 此外，完整的talin在胞浆中被自动抑制，以及它是如何被招募/激活来激活整合素和连接的。 整合素转肌动蛋白(Outside-In Signal)促进细胞骨架重组和动态黏附残留 难以捉摸。我们建议通过三个高度整合的目标来解决这些令人困惑的问题。Aim1将阐明一个 小GTP酶Rap1协同结合talin-H的talin F0和F1结构域以促进talin的募集和 新生的黏附适配器paxlin如何与PIP2协同激活talin。AIM2将在之后对此进行调查 激活talin，paxlin如何通过连接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。这一结果将为我们提供新的洞察力 Talin R对整合素的激活有重要作用。Aim3将研究talin如何介导整合素信号 肌动蛋白细胞骨架--一种长期推测的整合素外向内过程，至今仍未得到很好的阐明。 总之，我们提出的研究有望带来一种理解细胞黏附的新范式。 通过提供Talin如何被调节以经历一系列动态的机制的综合视图 激活整合素并将其连接到肌动蛋白细丝，导致细胞骨架重组和动态黏附的事件。 鉴于Talin及其相关蛋白在血栓形成、中风等方面的关键作用，我们的研究可能 最终导致对这些疾病更好的诊断和治疗方法的发展。