REGULATION OF PLASMA PROTEIN BIOSYNTHESIS

血浆蛋白生物合成的调节

• 批准号: 3352537
• 负责人: GERALD M FULLER
• 金额: $ 13.05万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-01-01 至 1987-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3352537
• 关键词: blood coagulation disorders; cyclic nucleoside monophosphate; fibrinogen; genetic regulation; haptoglobins; hemoprotein biosynthesis; hepatocellular carcinoma; immunologic techniques; liver cells; messenger RNA; molecular cloning; monoclonal antibody; plasma; plasminogen; polysomes; radiotracer; stoichiometry; tissue /cell culture

Fibrinogen and plasminogen are two hepatically derived plasma protein that play critical but opposite roles in hemostasis. Activated fibrinogen (fibrin) is clot forming and activated plasminogen (plasmin) is clot dissolving. Even though much is known about the chemistry of these two proteins little is known about how their synthesis is regulated or how the polypeptides are assembled and processed into functional proteins. The long range goal of this proposal is to elucidate at the cellular and molecular level these regulatory processes. Fibrinogen is a plasma protein whose synthesis is greatly increased during an inflammatory reaction. We have isolated a factor from monocytes that causes hepatocytes in culture to greatly increase their synthesis of fibrinogen thus mimicing the acute-phase response in culture. This cell culture will be used as a model system to investigate the intracellular events which leads to increased transcription of the fibrinogen mRNA's. We will use well characterized highly specific cDNA's as probes to determine both the kinetics of transcription of the fibrinogen mRNA's and the mRNA half-lives. We will also use the hepatocyte culture and stimulating factor to investigate what intracellular signals are operant that leads to the increased transcription of fibrinogen. We will attempt to determine exactly how the six polypeptides of fibrinogen are assembled into a functional protein. These experiments will be carried out by translating fibrinogen mRNA's in vitro and using very specific (monoclonal) antibodies as probes to determine the way the subunits link together. Elucidating the molecular origin of the two forms of plasminogen is one of the goals of this proposal. Plasminogen mRNA will be isolated and used in an in vitro translation system and the products analyzed to determine if two different translational forms of plasminogen exist. These results should give information on whether two mRNA's code for the two forms. We will use the purified mRNA to construct cDNA clones. Selected clones will be isolated and characterized as a first step to understanding the plasminogen gene structure. It is anticipated that information derived from these studies will significantly increase our understanding of how these important molecules are regulated. Once the control systems are fully understood then there is a possibility of being able to regulate their synthesis which can be of enormous therapeutic value in vascular diseases.

纤维蛋白原和纤溶酶原是两种肝源性血浆蛋白， 在止血中发挥关键但相反的作用。 活化纤维蛋白原 纤溶酶原（纤维蛋白）是凝块形成的，而活化的纤溶酶原（纤溶酶）是凝块形成的。 溶解 尽管我们对这两种物质的化学性质了解很多 蛋白质的合成是如何被调节的， 多肽被组装并加工成功能蛋白。 的 该提议的长期目标是阐明蜂窝和 分子水平上的这些调节过程。 纤维蛋白原是血浆蛋白 其合成在炎症反应期间大大增加。 我们 已经从单核细胞中分离出一种因子，该因子可使培养的肝细胞 大大增加了纤维蛋白原的合成， 培养物中的急性期反应。 这种细胞培养物将被用作模型 系统来研究导致增加的细胞内事件 纤维蛋白原mRNA的转录。 我们将使用特征良好的 高度特异性的cDNA作为探针，以确定 纤维蛋白原mRNA的转录和mRNA半衰期。 我们将 并采用肝细胞培养和刺激因子研究 细胞内信号是导致转录增加的操纵性信号， 纤维蛋白原 我们将试图确定这六个人 纤维蛋白原的多肽被组装成功能蛋白。 这些 将通过在体外翻译纤维蛋白原mRNA来进行实验 并使用非常特异的（单克隆）抗体作为探针来确定 子单元连接的方式。 阐明了 两种形式的纤溶酶原是本发明的目标之一。 酶原 mRNA将被分离并用于体外翻译系统， 分析产品，以确定是否有两种不同的翻译形式 存在纤溶酶原。 这些结果应该提供信息， 这两种形式的mRNA的代码。 我们将使用纯化的mRNA构建 cDNA克隆。 将分离选定的克隆并作为第一个 了解纤溶酶原基因结构的第一步。 预计 从这些研究中获得的信息将显着增加我们的 了解这些重要的分子是如何调节的。 一旦 控制系统被完全理解，那么就有可能被 能够调节它们的合成，这可能具有巨大的治疗价值， 在血管疾病中。