Kindlin-2 in Cell-Matrix Adhesion and Signaling

描述(申请人提供)：这项竞争性更新申请的长期目标是阐明细胞-细胞外基质(ECM)黏附和调节的分子基础，以及它们控制细胞行为、组织完整性、生长和再生的机制。申请人和其他人最近的研究表明，Kindlin-2(也称为Mig-2)，一种广泛表达的膜-细胞骨架连接蛋白，在整合素激活和细胞-ECM黏附中发挥关键作用。然而，Kindlin-2如何调控这些过程尚不清楚。根据以前项目期间获得的研究结果，申请人假设kindlin-2通过与细胞-细胞外基质黏附的膜脂和蛋白质成分相互作用来调节这些过程。为了验证这一假设，他提出了以下三个目标的研究。目的1是研究Kindlin-2与膜脂的相互作用，并评价其在整合素和整合素依赖过程中的调节作用。为此，他将采用遗传、药理学和显性负抑制策略来消除这种相互作用，并确定后果。目的2确定Kindlin-2与焦点黏附蛋白相互作用在调节细胞-ECM黏附中的作用。他将定义调节相互作用的位点，并使用“敲入”策略，用缺乏特定蛋白质结合活性的突变体取代野生型kindlin-2，并确定后果。目的3研究Kindlin-2及其与ILK在肝脏结构、生长和再生中的相互作用，已知这些功能受ECM黏附和ILK信号的调节。他将建立肝细胞特异性的kindlin-2基因敲除和“敲入”小鼠，其中野生型kindlin-2被缺乏特定结合活性的kindlin-2突变体取代，并确定kindlin-2及其相互作用在调节肝细胞行为、肝脏结构、生长和再生中的作用。这些研究将填补我们对细胞-细胞外基质黏附和细胞外基质依赖的组织过程调控机制的理解上的重要空白。鉴于细胞-ECM黏附在人类疾病中的重要性，这些研究也可能导致控制与细胞-ECM黏附和信号异常相关的疾病的新方法。 公共卫生相关性：细胞-ECM黏附的改变在包括癌症在内的人类疾病的发病机制中起着至关重要的作用。该项目旨在确定最近发现的一种细胞-ECM黏附调节因子如何影响细胞-ECM黏附、组织结构、生长和再生。这些研究可能导致确定新的治疗靶点，以控制与异常细胞-ECM黏附和生长相关的疾病。

DESCRIPTION (provided by applicant): The long-term goal of this competing renewal application is to elucidate the molecular basis underlying cell- extracellular matrix (ECM) adhesion and regulation, and the mechanism whereby they control cell behavior, tissue integrity, growth and regeneration. Recent studies by the applicant and others have demonstrated a critical role of kindlin-2 (also known as Mig-2), a widely expressed membrane-cytoskeleton junctional protein, in integrin activation and cell-ECM adhesion. How kindlin-2 regulates these processes, however, is not known. Based on findings obtained during previous project periods, the applicant hypothesizes that kindlin-2 regulates these processes through interacting with membrane lipids and protein components of cell-ECM adhesions. To test this hypothesis, he proposes studies with the following three aims. Aim 1 is to characterize the interaction of kindlin-2 with membrane lipids and assess its role in regulation of integrins and integrin-dependent processes. To this end, he will employ genetic, pharmacological and dominant negative inhibition strategies to ablate this interaction, and determine the consequences. Aim 2 is to determine the functions of kindlin-2 interactions with focal adhesion proteins in regulation of cell-ECM adhesion. He will define the sites mediating the interactions and use a "knock-in" strategy to replace wild type kindlin-2 with mutants lacking specific protein-binding activity and determine the consequences. Aim 3 is to investigate the functions of kindlin-2 and its interplay with ILK in liver structure, growth and regeneration, which are known to be regulated by ECM adhesion and ILK signaling. He will generate hepatocyte-specific kindlin-2 knockout and "knock-in" mice, in which wild type kindlin-2 is substituted with kindlin-2 mutants lacking specific binding activities, and determine contributions of kindlin-2 and its interactions to regulation of hepatocyte behavior, liver structure, growth and regeneration. These studies will fill important gaps in our understanding of the mechanism whereby cell-ECM adhesion and ECM-dependent tissue processes are regulated. Given the importance of cell-ECM adhesion in human diseases, these studies may also lead to novel approaches to control diseases associated with abnormal cell-ECM adhesion and signaling. PUBLIC HEALTH RELEVANCE: Alteration of cell-ECM adhesion is critically involved in the pathogenesis of human diseases including cancer. This project seeks to determine how a recently identified regulator of cell-ECM adhesion influences cell-ECM adhesion, tissue structure, growth and regeneration. These studies may lead to identification of novel therapeutic targets to control diseases associated with abnormal cell-ECM adhesion and growth.