Focal adhesion kinase in the cardiovascular system

描述（由申请人提供）： 本研究的长期目标是了解黏着斑激酶（FAK）及其信号通路在心血管系统中的作用机制。FAK是一种细胞质酪氨酸激酶，在整联蛋白介导的信号转导途径以及生长因子受体（包括在心血管系统中起关键调节作用的那些）的信号转导中起关键作用。与这些体外研究结果一致，小鼠中FAK基因敲除导致胚胎致死表型，轴向中胚层组织和心血管系统存在重大缺陷。尽管对FAK与其他蛋白质的相互作用及其在体外细胞信号传导中的作用有了丰富的了解，但对FAK在胚胎发育或成年生物体中的体内功能仍然知之甚少。尽管FAK基因完全敲除小鼠提示FAK在心血管系统中的潜在作用，但胚胎致死性使得其对于研究FAK在成年动物心血管发育或功能中的作用不是很有用。为了克服FAK全基因敲除小鼠的胚胎致死性问题，我们在初步研究中产生了FAK基因侧翼有两个loxP位点的FAK floxed小鼠。我们还建立了FAK转基因在内皮细胞（EC）中表达的转基因小鼠，并获得了Cre重组酶在EC或心肌细胞中特异性表达的转基因小鼠。在本申请中，目的1将通过使FAK/IoxP小鼠与TIE 2-Cre小鼠繁殖以产生EC特异性FAK敲除小鼠并检查对胚胎和成年小鼠中的血管发生和血管生成的影响来研究FAK在EC中在血管发生和血管生成中的作用。目的2通过FAK条件性基因敲除小鼠与FAK基因或其突变体转基因小鼠杂交，分离培养内皮细胞，研究FAK与其他蛋白的相互作用、细胞内信号通路以及细胞迁移和增殖等功能，探讨FAK在内皮细胞中发挥功能的分子和细胞机制。目的3将FAK/loxP小鼠与心肌细胞特异性Cre小鼠杂交，建立心脏特异性FAK条件性基因敲除小鼠，分析FAK基因敲除对心脏发育和功能的影响，探讨FAK在心脏发育和功能中的作用及其机制。这些研究不仅将产生对体内FAK信号通路的生理功能的重要见解，而且还为肿瘤血管生成和心脏肥大和心力衰竭的病理改变的潜在新疗法提供关键信息。

DESCRIPTION (provided by applicant): The long-term goals of this proposal are to understand the mechanisms and roles of focal adhesion kinase (FAK) and its signaling pathways in the cardiovascular system. FAK is a cytoplasmic tyrosine kinase that plays a key role in integrin-mediated signal transduction pathways as well as in signal transduction by growth factor receptors including those that play critical regulatory roles in the cardiovascular system. Consistent with these results from in vitro studies FAK gene knockout in mice resulted in an embryonic lethal phenotype with major defects in the axial mesodermal tissues and cardiovascular system. Despite the abundant knowledge of FAK interaction with other proteins and its roles in cell signaling in vitro, still relatively little is known about the in vivo Functions of FAK in embryonic development or in the adult organisms. Although the FAK total knockout mice suggested a potential role for FAK in the cardiovascular system, the embryonic lethality made it not very useful for studies on the role of FAK in cardiovascular development or functions in the adult animal. To overcome the problem of embryonic lethality of the FAK total knockout mice, we have generated FAK floxed mice with the FAK gene flanked by two loxP sites in preliminary studies. We have also generated transgenic mice with FAK transgene expression in the endothelial cells (ECs) and obtained transgenic mice with specific expression of Cre recombinase in ECs or cardiomyocytes. In this application, Aim 1 will investigate the role of FAK in ECs in [vasculogenesis and angiogenesis by breeding the FAK/IoxP mice with the TIE2-Cre mice to create EC-specific FAK knockout mice and examine the effects on vasculogenesis and angiogenesis in the embryo and in adult mice. Aim 2 will study molecular and cellular mechanism of FAK function in ECs by crossing the FAK conditional knockout mice with transgenic mice expressing FAK or its mutants and by isolating ECs from these mice to examine FAK interaction with other proteins, intracellular signaling pathways and cellular functions including migration and proliferation. Aim 3 will determine the role and mechanism of FAK in cardiac development and function by crossing FAK/loxP mice with cardiomyocyte specific Cre mice to generate heart specific FAK conditional knockout mice and analyzing the effects on the development and function of the heart. These studies will generate not only significant insights into the physiological function of FAK signaling pathways in vivo but also critical information for potential new therapies for pathological alterations in tumor angiogenesis and cardiac hypertrophy and heart failure.