CYTOPLASMIC EFFECTOR MOLECULES FOR L-SELECTIN

描述(改编自《调查人员摘要》)：白细胞招募 是炎症性疾病发病机制的重要组成部分。 白细胞重新聚集到炎症部位的调节发生 主要是在白细胞识别内皮细胞的水平上，这是一个过程 这是由一系列被命名为选择素的分子启动的。这 该提案旨在了解人类免疫缺陷的分子和细胞基础 黏附由L-选择素介导，是白细胞上唯一表达的选择素。 这种黏附分子在所有种类的白细胞上都有表达，这些白细胞有 体内不同的迁移模式，似乎参与了许多 炎症期间白细胞募集的类型。尽管激烈的 研究发现，L-选择素介导的黏附基础只是 部分地理解了。我们的初步研究表明，相互作用 L-选择素通过α-肌动蛋白和纽蛋白与肌动蛋白细胞骨架的相互作用 L-选择素介导的黏附必不可少。在具体目标1中， L-选择素/α-肌动蛋白的分子基础及功能意义 将对相互作用进行研究。免疫沉淀和免疫印迹分析 不同L-选择素胞浆尾部突变体的细胞株 用于鉴定L-选择素胞质尾部的残留物 负责与α-肌动蛋白和纽蛋白结合。功能界别 这些相互作用的意义将通过对 这组转染组细胞系与高内皮细胞结合的能力 小静脉(HEV)的淋巴结和在体内滚动，这两个最好的特点 而最重要的功能是由L介导的-选择素。在具体目标2中， 与L-选择素胞浆尾巴相互作用的其他蛋白质将是 这些蛋白的编码基因将被克隆出来。具体而言 目的3、利用免疫电子显微镜探讨其分子基础。 L-选择素定位于血管内皮细胞微绒毛的功能意义 白细胞，一个被认为对血管功能很重要的位置 L-选择素，特别关注涉及的细胞骨架蛋白 在这种蛋白质分选现象中。这些研究应该会大大增加 为了我们了解L-选择素的工作原理，并提供了一种可能性 许多急慢性疾病的多个新的临床干预靶点 炎症性疾病。

DESCRIPTION (Adapted from Investigator's Abstract): Leukocyte recruitment is an essential component of the pathogenesis of inflammatory disorders. The regulation of leukocyte recruitment into sites of inflammation occurs principally at the level of leukocyte recognition of endothelium, a process which is initiated by a family of molecules designated selectins. This proposal is directed at understanding the molecular and cellular basis of adhesion mediated by L-selectin, the only selectin expressed on leukocytes. This adhesion molecule is expressed on all classes of leukocytes, which have distinct patterns of migration in vivo, and appears to be involved in many types of leukocyte recruitment during inflammation. In spite of intense investigation, the basis for adhesion mediated by L-selectin is only partially understood. Our preliminary studies indicate that interactions between L-selectin and the actin cytoskeleton via a-actinin and vinculin are essential for L-selectin mediated adhesion. In specific aim 1, the molecular basis and functional significance of L-selectin/a-actinin interactions will be studied. Immunoprecipitation and Western blotting from cell lines transfected with various L-selectin cytoplasmic tail mutants will be used to identify the residues of the L-selectin cytoplasmic tail responsible for binding to a-actinin and vinculin. The functional significance of these interactions will be determined by examination of the ability of this panel of transfected cell lines to bind to high endothelial venules (HEV) of lymph nodes and to roll in vivo, the two best characterized and most important functions mediated by L-selectin. In specific aim 2, other proteins which interact with the L-selectin cytoplasmic tail will be identified, and cDNA encoding these proteins will be cloned. In specific aim 3, immunoelectron microscopy will be used to explore the molecular basis and functional importance of L-selectin positioning into the microvilli of leukocytes, a location believed to be important to the function of L-selectin, with particular attention to the cytoskeletal proteins involved in this protein sorting phenomenon. These studies should add considerably to our understanding of how L-selectin works, and offer the possibility of multiple new clinical targets for intervention in many acute and chronic inflammatory disorders.