Department of Biochemistry and Molecular Pharmacology, Chiba University Graduate School of Medicine, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670, Japan.
Cell Signal. 2011 Jun;23(6):1041-9. doi: 10.1016/j.cellsig.2011.01.023. Epub 2011 Feb 1.
Emerging evidence indicates that R4/B subfamily RGS (regulator of G protein signaling) proteins play roles in functional regulation in the cardiovascular system. In this study, we compared effects of three R4/B subfamily proteins, RGS2, RGS4 and RGS5 on angiotensin AT1 receptor signaling, and investigated roles of the N-terminus of RGS2. In HEK293T cells expressing AT1 receptor stably, intracellular Ca(2+) responses induced by angiotensin II were much more strongly attenuated by RGS2 than by RGS4 and RGS5. N-terminally deleted RGS2 proteins lost this potent inhibitory effect. Replacement of the N-terminal residues 1-71 of RGS2 with the corresponding residues (1-51) of RGS5 decreased significantly the inhibitory effect. On the other hand, replacement of the residues 1-51 of RGS5 with the residues 1-71 of RGS2 increased the inhibitory effect dramatically. Furthermore, we investigated functional contribution of N-terminal subdomains of RGS2, namely, an N-terminal region (residues 16-55) with an amphipathic α helix domain (the subdomain N1), a probable non-specific membrane-targeting subdomain, and another region (residues 56-71) between the α helix and the RGS box (the subdomain N2), a probable GPCR-recognizing subdomain. RGS2 chimera proteins with the residues 1-33 or 34-52 of RGS5 showed weak inhibitory activity, and either of RGS5 chimera proteins with residues 1-55 or 56-71 of RGS2 showed strong inhibitory effects on AT1 receptor signaling. The present study indicates the essential roles of both N-terminal subdomains for the potent inhibitory activity of RGS2 on AT1 receptor signaling.
新出现的证据表明，R4/B 亚家族 RGS（G 蛋白信号调节因子）蛋白在心血管系统的功能调节中发挥作用。在这项研究中，我们比较了三种 R4/B 亚家族蛋白，RGS2、RGS4 和 RGS5 对血管紧张素 AT1 受体信号的影响，并研究了 RGS2 N 端的作用。在稳定表达 AT1 受体的 HEK293T 细胞中，血管紧张素 II 诱导的细胞内 Ca(2+)反应被 RGS2 比 RGS4 和 RGS5 更强地减弱。截短的 RGS2 蛋白失去了这种强烈的抑制作用。用 RGS5 的相应残基（1-51）替换 RGS2 的 N 端残基 1-71，显著降低了抑制作用。另一方面，用 RGS2 的残基 1-71 替换 RGS5 的残基 1-51，则大大增加了抑制作用。此外，我们还研究了 RGS2 N 端亚域的功能贡献，即一个具有两亲性α螺旋结构域的 N 端区域（残基 16-55）（亚域 N1），一个可能的非特异性膜靶向亚域，以及另一个位于α螺旋和 RGS 盒之间的区域（残基 56-71）（亚域 N2），一个可能的 GPCR 识别亚域。用 RGS5 的残基 1-33 或 34-52 构建的 RGS2 嵌合蛋白表现出较弱的抑制活性，而用 RGS2 的残基 1-55 或 56-71 构建的任何一种 RGS5 嵌合蛋白对 AT1 受体信号都具有很强的抑制作用。本研究表明，RGS2 对 AT1 受体信号的强烈抑制活性需要两个 N 端亚域的共同作用。
