EXTRACELLULAR MATRIX MODULATION OF CELL PHENOTYPE

描述：(改编自申请人的摘要)细胞外基质(ECM)为细胞提供线索，从而成为细胞增殖、迁移、黏附和分化的框架。细胞对基质组成和结构变化的反应是通过与整合素和其他基质蛋白受体的相互作用来解释的。细胞骨架的重组、细胞形态的改变、信号转导通路的激活、基因表达模式的调节以及细胞周期的进展都是细胞对基质信号的一些反应。这项新建议的总体目标是通过细胞外基质与整合素受体的相互作用来控制细胞表型。纤维连接蛋白(FN)是细胞外基质的重要组成部分，它与三维纤维基质中的细胞相互作用。不同重组天然和突变FN的细胞组装可以改变FN基质纤维的结构。首席研究人员发现，结构不同的FN基质对细胞生长速度和粘着斑激酶的激活有不同的影响。这些差异效应可通过抑制纤维连接原纤维的形成而消融。基质FN调节细胞功能的机制将使用结合显微镜、生化和分子方法的细胞培养系统来研究。目的1是分析局部黏附成分、细胞形状和其他细胞表面受体在FN基质结构的生长反应中的作用。目的2是研究MAP和PI3-激酶下游信号通路在细胞生长和FN组装调控中的作用。整合素胞质结构域是通过整合素将信号从基质传递到细胞内室所必需的。为了剖析细胞质结构域的功能，PI开发了一个体内的秀丽线虫模型系统。在表达含有整合素b尾巴的嵌合蛋白的转基因线虫中产生显性的负表型。目的3是研究转基因线虫中完整和突变b尾巴所引起的表型。目的4是筛选突变的转基因线虫品系，从而鉴定参与整合素功能的基因。

DESCRIPTION: (adapted from applicant's abstract) The extracellular matrix (ECM) provides cues to cells and thus serves as a framework for cell proliferation, migration, adhesion, and differentiation. Cell responses to changes in matrix composition and structure are interpreted through interactions with integrins and other receptors for matrix proteins. Cytoskeletal reorganization, changes in cell morphology, activation of signal transduction cascades, modulation of gene expression patterns, and progression through the cell cycle represent some of the cellular responses to matrix signals. The overall goal of this new proposal is to provide insights into the control of cell phenotype by ECM interactions with integrin receptors. Fibronectin (FN), an essential ECM component, interacts with cells from within a 3-dimensional fibrillar matrix. The structure of FN matrix fibrils can be varied by cell assembly of different recombinant native and mutant FNs. The principal investigator has found that structurally distinct FN matrices have different effects on cell growth rates and activation of focal adhesion kinase. These differential effects are ablated by inhibition of FN fibril formation. The mechanisms by which matrix FN modulates cell function will be investigated using cell culture systems combined with microscopic, biochemical, and molecular approaches. Aim 1 is to analyze the role of focal adhesion components, cell shape, and other cell surface receptors in the growth response to FN matrix structure. Aim 2 is to examine the contributions of downstream signaling pathways involving MAP kinase and PI3-kinase to cell growth and regulation of FN assembly. Integrin cytoplasmic domains are essential for the transmission of signals from the matrix through integrins to intracellular compartments. To dissect cytoplasmic domain function, the PI has developed an in vivo, Caenorhabditis elegans model system. Dominant negative phenotypes are generated in transgenic nematodes expressing chimeric proteins containing integrin b tails. Aim 3 is to characterize the phenotypes caused by intact and mutant b tails in such transgenic nematodes. Aim 4 is to to screen mutant transgenic nematode lines and thus identify genes that particpate in integrin function.