THROMBOSPONDIN 2--STRUCTURE EXPRESSION AND FUNCTION

人血小板凝血酶原蛋白(TSP)是一种高分子量 由三个相同的二硫键组成的糖蛋白(Mr 450,000) 键合的多肽链。TSP是血小板α-亚基的一种成分 颗粒和第二阶段的基本成分 血小板聚集。TSP还由多种细胞合成 包括平滑肌细胞、内皮细胞和成纤维细胞。 在这些细胞中，合成的TSP被分泌和掺入。 进入细胞外基质(ECM)。然而，有一些 最近的研究表明，TSP可能起到调节主动脉的作用 平滑肌细胞(SMC)生长。TSP也可以作为焦点 通过其结合两者的能力在ECM中产生蛋白酶 纤溶酶原和组织型纤溶酶原激活物(t-PA)。 因为TSP是血小板和血管壁的组成部分 参与了血小板-血小板和血小板血管的相互作用 它的研究应该有助于更好地理解它的作用 生理和病理状态(血栓形成、动脉粥样硬化)。 为了获得更多关于结构和 TSP在管壁中合成的调节 建议：1)研究肝素结合的作用 TSP与主动脉平滑肌细胞的结构域(HBD) 成长。这些研究将使用一种细菌表达系统， 合成HBD。隔离的HBD将被用于研究其 在主动脉平滑肌细胞增殖中的作用。2)对网络环境的研究 TSP的基因结构，以更好地了解TSP的组织和 基因的进化。解决的问题将包括：a)是否 分离功能结构域的外显子编码？B)TSP基因 通过基因复制等机制进化成现在的形式 和外显子改组？3)研究 血小板衍生生长因子(PDGF)促进TSP合成 PDGF是否通过增加血管内皮细胞TSP基因表达而增加TSP基因的表达？ 转录速率或通过稳定TSP？4的mRNA) 定义TSP基因中的PDGF响应元件 由假定的调节性基因组成的基因结构 与报告基因(细菌氯霉素乙酰基)融合的区域 转移酶(CAT))。5)研究TSP与TSP的相互作用 利用纯化蛋白和分离的主动脉SMC制备纤溶酶原 矩阵。6)开发利用TSP的癌细胞的能力 作为尝试定位细胞结合的附着因素 TSP上的域。

Human platelet thrombospondin (TSP) is a high molecular weight glycoprotein (Mr 450,000) composed of three identical disulphide bonded peptide chains. TSP is a constituent of platelet alpha- granules and an essential component of the secondary phase of platelet aggregation. TSP is also synthesized by a variety of cells including smooth muscle cells, endothelial cells and fibroblasts. In these cells the synthesized TSP is secreted and incorporated into the extracellular matrix (ECM). However there is some recent work which suggests that TSP may serve to regulate aortic smooth muscle cell (SMC) growth. TSP may also serve as a focus for protease generation in the ECM by its ability to bind both plasminogen and tissue plasminogen activator (t-PA). Since TSP is constituent of both the platelet and the vessel wall and involved in platelet-platelet and platelet vessel interactions its study should contribute to the better understanding of its role in both physiologic and pathologic states (thrombosis, atheroma). In order to gain greater knowledge about the structure and regulation of TSP synthesis in the vessel wall the following studies are proposed: 1) Investigation of the role of the heparin binding domain (HBD) of TSP with respect to aortic smooth muscle cell growth. These studies will use a bacterial expression system that synthesizes the HBD. The isolated HBD will be used to study its role in aortic smooth muscle cell proliferation. 2) Study of the gene structure of TSP to better understand the organization and evolution of the gene. Questions addressed will include, a) Do the exons code for seperate functional domains? b) Did the TSP gene evolve to its present form by mechanisms such as gene duplication and exon shuffling? 3) Investigate the mechanism by which platelet derived growth factor (PDGF) increases TSP synthesis in aortic SMC's. Does PDGF increase TSP mRNA by increasing the transcriptional rate or by stabilizing the mRNA for TSP? 4) Define the PDGF responsive elements within the TSP gene by use of gene constructions that consist of the putative regulatory region fused to a reporter gene (bacterial chloramphenicol acetyl transferase (CAT)). 5) Study the interaction of TSP with plasminogen using purified proteins and isolated aortic SMC matrices. 6) Exploit the ability of carcinoma cells which use TSP as an attachment factor in trying to localize the cell binding domain on TSP.