Regulation of G protein-signaling by RGS proteins in cardiovascular system

心血管系统中 RGS 蛋白对 G 蛋白信号传导的调节

• 批准号: 11680623
• 负责人: KIMURA Sadao
• 金额: $ 2.37万
• 依托单位: CHIBA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11680623/
• 关键词: RGS; G protein; angiotensin; endothelin; G蛋白質αサブユニット; 脱感作; G蛋白質共役受容体キナーゼ

RGS proteins (regulators of G protein signaling) serve as GTPase-activating proteins (GAPs) for Gα subunits and negatively regulate G protein-coupled receptor signaling. In this study, we characterized biochemical properties of RGS5 and its N terminal (1-33)-deleted mutant (ΔN-RGS5). RGS5 bound to Gαi1, Gαi2, Gαi3, Gαo and Gαq but not to Gαs and Gα13 in the presence of GDP/AlF, and accelerated the catalytic rate of GTP hydrolysis of Gαi3 subunit. When expressed in 293T cells stably expressing angiotensin (Ang) AT1a receptors (AT1a-293T cells), RGS5 suppressed Ang II-and endothelin (ET)-1-induced intracellular Ca transients. The effect of RGS5 was concentration-dependent, and the slope of the concentration-response relationship showed that a 10-fold increase in amounts of RGS5 induced about 20-25% reduction of the Ca signaling. Furthermore, a comparison study of three sets of 293T cells with different expression levels of AT1a receptors showed that RGS5 inhibited Ang II-induced responses more effectively in 293T cells with the lower density of AT1a receptors, suggesting that the degree of inhibition by RGS proteins reflects the ratio of amounts of RGS proteins to those of activated Gα subunits after receptor stimulation by agonists. When expressed in AT1a-293T cells, ΔN-RGS5 was localized almost exclusively in the cytosolic fraction, and exerted the inhibitory effects as potently as RGS5 which was present in both membrane and cytosolic fractions. Studies on relationship between subcellular localization and inhibitory effects of RGS5 and ΔN-RGS5 revealed that the N terminal (1-33) of RGS5 plays a role in targeting this protein to membranes, and that the N terminal region of RGS5 is not essential for exerting activities.

RGS蛋白（G蛋白信号调节剂）作为Gα亚基的GTP酶激活蛋白（GAP），负调节G蛋白偶联受体信号。本研究对RGS 5及其N端（1-33）缺失突变体（Δ N-RGS 5）的生化特性进行了研究。在GDP/AlF存在下，RGS 5与Gαi1、Gαi2、Gαi3、Gαo和Gαq结合，而不与Gαs和Gα13结合，并加速Gαi3亚基的GTP水解速率。当在稳定表达血管紧张素（Ang）AT 1a受体的293 T细胞（AT 1a-293 T细胞）中表达时，RGS 5抑制Ang II和内皮素（ET）-1诱导的细胞内Ca瞬变。RGS 5的作用是浓度依赖性的，并且浓度-响应关系的斜率显示RGS 5的量增加10倍诱导Ca信号传导减少约20-25%。此外，对三组具有不同AT 1a受体表达水平的293 T细胞的比较研究表明，RGS 5在AT 1a受体密度较低的293 T细胞中更有效地抑制Ang II诱导的反应，这表明RGS蛋白的抑制程度反映了激动剂刺激受体后RGS蛋白与活化Gα亚基的比例。当在AT 1a-293 T细胞中表达时，Δ N-RGS 5几乎仅定位于胞质组分，并且发挥与存在于膜和胞质组分中的RGS 5一样强的抑制作用。对RGS 5和Δ N-RGS 5的亚细胞定位与抑制作用关系的研究表明，RGS 5的N端（1-33）在将该蛋白定位于细胞膜中起作用，而N端区域不是发挥活性所必需的。

项目成果

T. Shibasaki, 他: "Characterization of the carboxyl-terminal truncated endothelin B receptor coexpressed with G protein-coupled receptor kinase 2"Biochem. Mol. Biol. Int.. 47. 569-577 (1999)

T. Shibasaki 等：“与 G 蛋白偶联受体激酶 2 共表达的羧基末端截短的内皮素 B 受体的表征”Biochem. Int. 47. 569-577 (1999)

Zhou J: "Characterization of RGS5 in regulation of G protein-coupled receptor signaling."Life Sciences. 68. 1457-1469 (2001)

Zhou J：“RGS5 在 G 蛋白偶联受体信号传导调节中的表征。”生命科学。

H. Usui, 他: "RGS domain in the amino-terminus of G protein-coupled receptor kinase-2 inhibits Gq-mediated signaling"Int. J. Mol. Med.. (印刷中). (2000)

H. Usui 等人：“G 蛋白偶联受体激酶 2 的氨基末端的 RGS 结构域抑制 Gq 介导的信号”Int. Mol.（出版中）。

Usui H: "RGS domain in the amino-terminus of G protein-coupled receptor kinase-2 inhibits Gqmediated signaling."Int.J.Mol.Med.. 5. 335-340 (2000)

Usui H：“G 蛋白偶联受体激酶 2 氨基末端的 RGS 结构域抑制 Gq 介导的信号传导。”Int.J.Mol.Med.. 5. 335-340 (2000)

J.Zhou, et al.: "Characterization of RGS5 in regulation of G protein-coupled receptor signaling."Life Sciences. 68. 1457-1469 (2001)

J.Zhou 等人：“RGS5 在 G 蛋白偶联受体信号传导调节中的表征。”生命科学。