Analysis of the signal transduction pathway through the thromboxane A_2 receptor using the receptor-G-protein reconstitution system

使用受体-G-蛋白重建系统分析血栓素 A_2 受体的信号转导途径

基本信息

批准号：
05670121
负责人：
USHIKUBI Fumitaka
金额：
$ 1.28万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05670121/
关键词：
Thromboxane Receptor GTP-binding protein Reconstitution

项目摘要

Thromboxane (TX) A_2 is a metabolite of arachidonic acid. It induces activation of platelets and constriction of vascular- and bronchial smooth muscles. It is well known that TXA_2 has physiologically and path-physologically important roles in these tissues. On the other hand, we have recently cloned a cDNA of the mouse TXA_2 receptor and have revealed its primary structure. We have also examined the distribution of the mouse TXA_2 receptor mRNA and found its broad and specific expressions in various mouse organs. In this study, we intended to construct the TXA_2 receptor-G-protein reconstitution system and to clarify the signal transduction mechanisms through the receptor using this system.We have purified Gi and Gq proteins and partially purified the TXA_2 receptor from human platelets. We also succeeded in reconstituting these proteins in phospholipid vesicles last year.This year, we examined the ligand binding characteristics of the reconstituted receptor and the agonist-induced activation of the reconstituted G-proteins. We found that the TXA_2 receptor couples to both Gq and Gi proteins. We also evaluated the efficacies and the extent of the receptor-G-protein coupling. These results explained at least partially the multiplicity of signal transduction pathways through the TAX_2 receptor and suggested the participation of the Gi protein in the signal transduction, with which TXA_2 induces cell proliferation.

血栓烷（TX）A_2是花生四烯酸的代谢产物。它诱导血小板的激活以及血管和支气管平滑肌肉的收缩。众所周知，TXA_2在这些组织中具有生理和路径生理上重要的作用。另一方面，我们最近克隆了小鼠TXA_2受体的cDNA，并揭示了其主要结构。我们还检查了小鼠TXA_2受体mRNA的分布，并在各种小鼠器官中发现了其广泛而特定的表达。在这项研究中，我们打算构建TXA_2受体-G蛋白重建系统，并使用该系统通过受体阐明信号转导机制。我们已经纯化了GI和GQ蛋白，并从人类Platelets中部分纯化了TXA_2受体。去年，我们还成功地在磷脂囊泡中重建了这些蛋白质。这一年，我们研究了重构受体的配体结合特性，以及激动剂诱导的重构G蛋白的激活。我们发现TXA_2受体夫妇均与GQ和GI蛋白质夫妇伴侣。我们还评估了受体-G蛋白偶联的功效和程度。这些结果至少部分解释了通过税收_2受体的信号转导途径的多样性，并提出了GI蛋白参与信号转导中的参与，TXA_2诱导细胞增殖。