THROMBOSPONDIN 2; STRUCTURE EXPRESSION AND FUNCTION

血小板反应蛋白 2；

• 批准号: 3202426
• 负责人: VISHVA M DIXIT
• 金额: $ 25.14万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-08-04 至 1997-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3202426
• 关键词: RNA biosynthesis; affinity chromatography; arteriosclerosis; autoradiography; binding proteins; biological products; biotechnology; cell growth regulation; chemical binding; extracellular matrix; fibroblasts; gel electrophoresis; gene expression; gene mutation; genetic manipulation; genetic mapping; genetic promoter element; genetic recombination; genetic transcription; heparin; immunoprecipitation; laboratory mouse; laboratory rabbit; messenger RNA; molecular cloning; monoclonal antibody; neoplastic cell culture for noncancer research; nucleic acid hybridization; nucleic acid probes; nucleic acid sequence; online computer; plasmids; plasminogen; platelet derived growth factor; platelets; protein biosynthesis; protein sequence; protein structure; radiotracer; regulatory gene; reporter genes; thrombosis; thrombospondins; tissue /cell culture; vascular smooth muscle

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.

人血小板血小板反应蛋白（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） 键合的肽链。 TSP 是血小板α-的组成部分 颗粒和第二相的主要成分 血小板聚集。 TSP也由多种细胞合成 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 被分泌并掺入 进入细胞外基质（ECM）。 然而有一些 最近的研究表明 TSP 可能有助于调节主动脉 平滑肌细胞（SMC）生长。 TSP 也可作为焦点 通过其结合两者的能力，在 ECM 中生成蛋白酶 纤溶酶原和组织纤溶酶原激活剂（t-PA）。 由于 TSP 是血小板和血管壁的组成部分 并参与血小板-血小板和血小板血管相互作用 其研究应有助于更好地理解其作用 生理和病理状态（血栓形成、动脉粥样硬化）。 为了更深入地了解其结构和 血管壁中 TSP 合成的调节如下研究 建议：1）研究肝素结合的作用 TSP 与主动脉平滑肌细胞相关的结构域 (HBD) 生长。 这些研究将使用细菌表达系统 合成HBD。 分离的 HBD 将用于研究其 主动脉平滑肌细胞增殖的作用。 2）研究 TSP 的基因结构，以更好地了解组织和 基因的进化。 解决的问题包括：a) 不同功能域的外显子代码？ b) TSP 基因是否 通过基因复制等机制进化到现在的形式 和外显子改组？ 3）研究其机制 血小板衍生生长因子 (PDGF) 增加 TSP 合成 主动脉 SMC。 PDGF 是否通过增加 TSP mRNA 转录率还是通过稳定 TSP 的 mRNA？ 4） 通过使用定义 TSP 基因内的 PDGF 响应元件 由假定的调控组成的基因构建 与报告基因融合的区域（细菌乙酰氯霉素 转移酶（CAT））。 5) 研究TSP与 使用纯化蛋白和分离的主动脉 SMC 进行纤溶酶原 矩阵。 6) 利用癌细胞利用TSP的能力 作为尝试定位细胞结合的附着因子 TSP 上的域。