MOLECULAR ULTRASTRUCTURE UNDERLYING CELL ADHESION

研究表明， 各种细胞-ECM附着位点将继续，特别是 强调改变细胞粘附的分子机制， 去粘附和在转化细胞中的侵袭。 我们的工作假设 是：1）不同的，特定的细胞表面受体参与 在动态过程中组织细胞与纤连蛋白和胶原的附着 细胞粘附和侵袭的过程，2）配体如纤连蛋白可以 有助于调节受体和细胞骨架的分布， 和3）定位的细胞表面蛋白酶负责细胞 ECM的接触相关溶解，这允许侵入。 最近的研究表明， 细胞表面蛋白酶降解细胞内的纤连蛋白和胶原蛋白， 接触部位。 因此，本提案的近期目标是 使用单克隆抗体探针定位这些转化敏感的， 入侵相关蛋白酶，以鉴定蛋白酶结构域，和 干扰蛋白酶的分子相互作用。 为此，我们计划 确定这些蛋白酶在转化细胞侵袭中的可能作用 或作为肿瘤细胞转移潜能的指示剂。 拟议 研究还将确定可能的胶原附着位点， 微观水平，并确定这种附件是否通过 胶原受体 为此，我们将使用原位本地化， 抗体抑制方法类似于用于 纤连蛋白-受体结合的表征。 可能 纤维连接蛋白和胶原在组织中的调节作用 附着位点和肿瘤细胞的侵袭中， 通过让这些分子与细胞表面结合来研究， 使用特异性单克隆抗体探针抑制它们的结合， 合成肽 更具体地说，我们将寻求确定：1）分子结构 和细胞表面蛋白酶的生物化学性质 杂交瘤技术，2）细胞表面蛋白酶在 转化并侵入纤连蛋白和胶原基质，3） 细胞表面胶原受体的表达和定位， 纤连蛋白在不同的附着位点，4）的调节作用， 纤维连接蛋白和胶原蛋白在组织的粘连部位， 转化细胞，和5）细胞的体内生物学意义 表面蛋白酶或受体参与肿瘤细胞的迁移和侵袭。

Studies demonstrating the structural and functional specificities of various cell-ECM attachment sites will be continued, with particular emphasis on altered molecular mechanisms of cell adhesion, cell de-adhesion, and invasion in transformed cells. Our working hypotheses are: 1) distinct, specific cell surface receptors are involved in organizing cell attachments to fibronectin and collagen during dynamic processes of cell adhesion and invasion, 2) ligands such as fibronectin can help to regulate the distribution of their receptors and the cytoskeleton, and 3) localized cell surface proteases are responsible for cell contact-related dissolution of the ECM, which permits invasion. Recent studies have demonstrated the existence of transformation-induced cell surface proteases that degrade fibronectin and collagen at cell contact sites. Thus, the immediate goals of the present proposal are to use monoclonal antibody probes to localize these transformation-sensitive, invasion-associated proteases, to identify protease structural domains, and to perturb molecular interactions involving proteases. By this we plan to determine the possible role of these proteases in transformed cell invasion or as indicators of the metastatic potential of tumor cells. The proposed studies will also define possible collagen attachment sites at the electron microscopic level, and determine whether such attachments function via collagen receptors. To do this, we will use in situ localization and antibody inhibition approaches similar to those used for the characterization of the fibronectin-receptor association. Possible regulatory roles of fibronectin and collagen in the organization of attachment sites and in the invasion of the tumor cells will then be studied by allowing these molecules to bind to the cell surface and inhibiting their binding using specific monoclonal antibody probes and synthetic peptides. More specifically, we will seek to determine: 1) the molecular structure and biochemical properties of cell surface proteases by the use of hybridoma technology, 2) the roles of cell surface proteases in transformation and invasion into fibronectin and collagen substrata, 3) the expression and localization of cell surface receptors for collagen and fibronectin at distinct attachment sites, 4) the regulatory roles of fibronectin and collagen in the organization of adhesion sites in transformed cells, and 5) the in vivo biological significance of cell surface proteases or receptors in migration and invasion of tumor cells.