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Characterization of factors regulating the activity of hepatocyte growth factor which is being developed for a medicine

正在开发用于药物的肝细胞生长因子活性调节因子的表征

基本信息

批准号：
09480161
负责人：
KITAMURA Naomi
金额：
$ 8.51万
依托单位：
TOKYO INSTITUTE OF TECHNOLOGY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09480161/
关键词：
hepatocyte growth factor(HGF)liver injury HGF activator HGF activator inhibitor Kunitz domain Hrs Hrs binding protein SH3 domain Hrs 結合蛋白質肝細胞増殖因子

项目摘要

Hepatocyte growth factor (HGF) is a potent factor responsible for liver regeneration, and is being developed for a medicine of liver disease. The activity of HGF is regulated by extracellular and intracellular factors. In this study, we analyzed the function of these factors in regulating the activity of HGF and obtained the following results.(1) HGF activator (HGFA) was identified as a serine protease responsible for activation of HGF after liver injury. Analysis of HGFA after liver injury revealed that amount of HGFA mRNA increased, and HGFA was activated following liver injury.(2) Two types of HGFA inhibitors (HAIs) were identified as potent inhibitors of HGFA.cDNA cloning revealed that both HAIs are serine protease inhibitors with Kunitz domains. Introduction of mutations in the Kunitz domains markedly reduced the activity of HAIs, indicating that the domains are essential for the activity. Immunoblotting analysis demonstrated that HAIs are initially produced in transmembrane forms, and then secreted by the producing cells by proteolytic processing.(3) Hrs and its binding protein (Hbp) were identified as intracellular molecules regulating the activity of HGF.Functional characterization of Hrs/Hbp complexes revealed that the complexes are localized to early endosomes and play an important role in regulating the degradation of HGF and its receptor after internalization. Further, deubiquitinating enzyme UBPY was isolated as a binding partner of the SH3 domain of Hbp, suggesting that UBPY regulates the function of the Hrs/Hbp complexes.

肝细胞生长因子（HGF）是一种有效的肝再生因子，正在开发用于治疗肝脏疾病的药物。HGF的活性受细胞外和细胞内因子的调节。本研究分析了这些因子在调节肝细胞生长因子活性中的作用，得到以下结果。(1)肝细胞生长因子激活因子（HGF activator，HGFA）是一种丝氨酸蛋白酶，在肝损伤后激活肝细胞生长因子。肝损伤后HGFA的分析显示，HGFA mRNA的量增加，并且HGFA在肝损伤后被激活。(2)两种HGFA抑制剂（HAI）被鉴定为HGFA的有效抑制剂，cDNA克隆显示这两种HAI都是具有Kunitz结构域的丝氨酸蛋白酶抑制剂。在Kunitz结构域中引入突变显著降低了HAI的活性，表明该结构域对于活性是必不可少的。免疫印迹分析表明HAI最初以跨膜形式产生，然后由产生细胞通过蛋白水解加工分泌。(3)Hrs及其结合蛋白（Hbp）被鉴定为调节HGF活性的细胞内分子，Hrs/Hbp复合物的功能研究表明，Hrs/Hbp复合物定位于早期内体，在调节HGF及其受体内化后的降解过程中起重要作用。此外，去泛素化酶UBPY被分离为Hbp的SH 3结构域的结合伴侣，表明UBPY调节Hrs/Hbp复合物的功能。