Molecular Mechanism Controlling Acceleration Patterns of G-protein Signaling Induced by RGS Proteins.

控制 RGS 蛋白诱导的 G 蛋白信号传导加速模式的分子机制。

• 批准号: 11680647
• 负责人: SAITOH Osamu
• 金额: $ 2.11万
• 依托单位: Tokyo Metropolitan Organization for Medical Research
• 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-11680647/
• 关键词: G-protein; RGS; desensitization; k+channel; receptor; transmitter; brain; neuron

The recently discovered family of RGS (regulators of G protein signaling) proteins acts as GTPase activating proteins which bind to α subunits of heterotrimeric G proteins. We previously showed that a brain specific RGS, RGS8 speeds up the activation and deactivation kinetics of the G protein- coupled inward rectifier K+ channel (GIRK) upon receptor stimulation. Further, the acute desensitization of receptor-activated GIRK was observed with RGS8.1. We isolated a full-length rat cDNA of another brain specific RGS, RGS7. RGS7 cDNA encoded a protein of 477 amino acids with a long N-terminus containing 330 residues. RGS8 is a small RGS protein of 180 amino acids with a short N-terminus. When co-expressed with GIRK1/2 in Xenopus oocytes, RGS7 and RGS8 differentially accelerate G protein-mediated modulation of GIRK.RGS7 clearly accelerated activation of GIRK current similarly with RGS8 but the acceleration effect of deactivation was significantly weaker than that os RGS8. The acute desensiti … More zation of receptor-activated GIRK observed with RGS8 was not apparent with RGS7. These acceleration properties of RGS proteins may play important roles in the rapid regulation of neuronal excitability and the cellular responses to short-lived stimulations. The N-terminal domains of RGS proteins might function as a regulatory element for these diffenrential acceleration of G protein signaling.2. Functional roles of the N-terminal region of RGS8 in G-protein signaling were studied. The deletion of the N-terminal region of RGS8 (ΔNRGS8) resulted in a partial loss of the inhibitory function in pheromone response of yeasts, although Gα binding was not affected. To examine roles in subcellular distribution, we co-expressed two fusion proteins of RGS8-RFP and ΔNRGS8-GFP in DDT1MF2 cells. RGS8-RFP was highly concentrated in nuclei of unstimulated cells. Co-expression of constitutively active Gαo resulted in translocation of RGS8 protein to the plasma membrane. In contrast, ΔNRGS8-GFP was distributed diffusely through the cytoplasm in the presence or absence of active Gαo. When co-expressed with GIRK channels, ΔNRGS8 accelerated both turning-on and -off similar to RGS8. Acute desensitization of GIRK current observed in RGS8, however, was not induced by ΔNRGS8. Thus, we, for the first time, showed that the N-terminus of RGS8 contributes to the subcellular localization and to the desensitization of the G-protein-coupled response. Less

最近发现的RGS（regulators of G protein signaling）蛋白家族作为GT3激活蛋白与异源三聚体G蛋白的α亚基结合。我们先前表明，脑特异性RGS，RGS 8在受体刺激时加速G蛋白偶联的内向整流K+通道（GIRK）的激活和失活动力学。此外，用RGS8.1观察到受体激活的GIRK的急性脱敏。我们分离了另一个脑特异性RGS，RGS 7的全长大鼠cDNA。RGS 7 cDNA编码477个氨基酸，N端长330个残基。RGS 8是一种小的RGS蛋白，具有180个氨基酸和短的N-末端。当与GIRK 1/2在爪蟾卵母细胞中共表达时，RGS 7和RGS 8差异性地加速G蛋白介导的GIRK调节。RGS 7与RGS 8类似地明显加速GIRK电流的激活，但失活的加速效应显著弱于RGS 8。急性感觉缺失 ...更多信息 用RGS 8观察到的受体活化的GIRK的化用RGS 7不明显。RGS蛋白的这些加速特性可能在快速调节神经元兴奋性和细胞对短暂刺激的反应中起重要作用。RGS蛋白的N端结构域可能是G蛋白信号传导差异性加速的调控元件.研究了RGS 8的N端区域在G蛋白信号传导中的功能作用。RGS 8的N端区域（Δ NRGS 8）的缺失导致酵母对信息素反应的抑制功能部分丧失，但Gα结合不受影响。为了研究在亚细胞分布中的作用，我们在DDT 1 MF 2细胞中共表达RGS 8-RFP和Δ NRGS 8-GFP的两种融合蛋白。RGS 8-RFP在未刺激的细胞的细胞核中高度浓缩。组成型活性Gαo的共表达导致RGS 8蛋白易位至质膜。与此相反，在存在或不存在活性Gαo的情况下，Δ NRGS 8-GFP在细胞质中弥散分布。当与GIRK通道共表达时，Δ NRGS 8与RGS 8类似地加速开启和关闭。然而，在RGS 8中观察到的GIRK电流的急性脱敏不是由Δ NRGS 8诱导的。因此，我们，第一次，表明RGS 8的N-末端有助于亚细胞定位和G-蛋白偶联反应的脱敏。少

项目成果

Saitoh, O., Masuho, I., Terakawa, I., nomoto, S., Asano, T., and Kubo, Y.: "Regulator of G protein signaling 8 (RS8) requires its N-terminus for subcellular localization and acute desensitization of G protein-gated K^+ channels."J.Biol. Chem.. 276. 5052-5

Saitoh, O.、Masuho, I.、Terakawa, I.、nomoto, S.、Asano, T. 和 Kubo, Y.：“G 蛋白信号传导调节器 8 (RS8) 需要其 N 末端进行亚细胞定位，并且

Osamu Saitoh et al.: "Molecular Cloning and Characterization of Xenopus RGS5"Biochem.Biophys.Res.Comm.. 270.1. 34-39 (2000)

Osamu Saitoh 等人：“爪蟾 RGS5 的分子克隆和表征”Biochem.Biophys.Res.Comm.. 270.1。

Osamu Saitoh et al.: "Regulator of G Protein Signaling 8(RGS8) Requires Its NH_2 Terminusfor Subcellular Localization and Acute Desensitization..."J.Biol.Chem.. 276.7. 5052-5058 (2001)

Osamu Saitoh 等人：“G 蛋白信号传导调节器 8 (RGS8) 需要其 NH_2 末端进行亚细胞定位和急性脱敏......”J.Biol.Chem.. 276.7。