PHYSIOLOGICAL MECHANISMS OF INTEGRIN AFFINITY MODULATION

描述（改编自研究者摘要）：血细胞，如 血小板、淋巴细胞和中性粒细胞发挥其生理功能 通过与细胞外基质或与其他细胞因子的粘附相互作用， 细胞整合素是跨膜α，β异二聚体家族 介导许多这些细胞类型的粘附，值得注意的是 的动态调节。为了响应外部刺激，整合素 上调配体结合亲和力并改变细胞的亲和力。 初步数据表明，整合素的“激活”或亲和力 调节涉及通过整合素的细胞特异性信号传导 胞质结构域。在本建议书中，主要研究者将阐明 生理途径首先，他将识别α和β亚基 参与的细胞质残基。使用细胞外α IIb， β 3作为报告基团，他将产生变异的胞质结构域， 将这些构建体转染到异源细胞中，并分析它们的表达。 结合抗体或蛋白质配体的能力。第二，他会检查 整合素胞质结构域的过表达对 生理激活机制。嵌合体由 IL-2受体的细胞外和跨膜区连接到 已经获得了特异性整联蛋白胞质结构域。这些 构建体将与组成型活性整联蛋白共转染 并分析对结合的抑制作用。最后，他会尝试 鉴定与整合素相互作用的假设胞质因子 胞质结构域。新的因素将被分离和鉴定， 利用上述抑制性嵌合体的交联研究， 或通过在双杂交系统中再生GAL 4转录因子。 确定亲和力调节的机制对于 了解整合素在肿瘤的生理和病理中的作用 血细胞

DESCRIPTION (Adapted from investigator's abstract): Blood cells such as platelets, lymphocytes and neutrophils exert their physiological function through adhesive interactions with the extracellular matrix or with other cells. The integrins, a family of transmembrane alpha, beta heterodimers which mediate the adhesion of many of these cell types, are notable for their dynamic regulation. In response to external stimuli, integrins upregulate ligand binding affinity and alter cellular adhesiveness. Preliminary data suggest that integrin "activation" or affinity modulation involves cell- specific signalling through integrin cytoplasmic domains. In this proposal, the PI will elucidate this physiological pathway. First, he will identify alpha and beta subunit cytoplasmic residues which are involved. Using extracellular alphaIIb, beta3 as a reporter group, he will generate variant cytoplasmic domains, transfect these constructs into heterologous cells and analyze their ability to bind antibody or protein ligands. Second, he will examine the effect of overexpression of integrin cytoplasmic domains on physiological activation mechanisms. Chimera consisting of the extracellular and transmembrane regions of the IL-2 receptor joined to specific integrin cytoplasmic domains have been obtained. These constructs will be co-transfected with constitutively active integrins and the inhibitory effect on binding analyzed. Finally, he will attempt to identify hypothetical cytosolic factors which interact with integrin cytoplasmic domains. Novel factors will be isolated and identified in crosslinking studies utilizing the inhibitory chimera identified above, or by regenerating the GAL4 transcription factor in a two hybrid system. Defining the mechanisms of affinity modulation is crucial to understanding the role of integrins in the physiology and pathology of blood cells.