Inhibition of Gαi activity by Gβγ is mediated by PI 3-kinase-γ- and cSrc-dependent tyrosine phosphorylation of Gαi and recruitment of RGS12
Inhibition of Gαi activity by Gβγ is mediated by PI 3-kinase-γ- and cSrc-dependent tyrosine phosphorylation of Gαi and recruitment of RGS12
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DOI：
10.1152/ajpgi.00440.2013
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发表时间：
2014-05-01
影响因子：
4.5
通讯作者：
Murthy, Karnam S.
中科院分区：
文献类型：
--
作者：
Huang, Jiean;Nalli, Ancy D.;Murthy, Karnam S.
其他人和我们已经表征了平滑肌中的几种G β γ依赖性效应物，包括G蛋白偶联受体激酶2（GRK 2）、PLC β（3）和磷脂酰肌醇（PI）3-激酶-γ，并鉴定了PI 3-激酶-γ下游的各种信号传导靶点，包括cSrc、整合素连接激酶和Rac 1-Cdc 42/p21激活激酶/p38 MAP激酶。这项研究确定了一种新的机制，即G β γ通过PI 3-激酶-γ和cSrc作用，对G α（i）活性产生抑制作用。G（i2）-偶联δ-阿片受体激动剂D-青霉胺（2，5）-脑啡肽（DPDPE）激活cSrc，刺激G α（i2）的酪氨酸磷酸化，并诱导G蛋白信号转导调节因子12（RGS 12）结合;所有三个事件都被PI 3-激酶阻断（LY 294002）和cSrc（PP 2）抑制剂，并通过表达GRK 2-（495-689）（一种G β γ-清除肽）的COOH末端序列。PP 2、LY 294002和选择性PI 3-激酶-γ抑制剂AS-605420增强了DPDPE对毛喉素刺激的cAMP和肌肉松弛的抑制作用。酪氨酸缺陷型（Y 69 F、Y231 F或Y321 F）G α（i2）突变体的表达或RGS 12的敲低阻断了G α（i2）磷酸化和G α（i2）-RGS 12缔合，并引起cAMP的更大抑制。使用生长抑素、环戊基腺苷或ACh分别激活G（i1）偶联生长抑素（sstr（3））受体和G（i3）偶联腺苷A（1）或毒蕈碱m（2）受体的平行研究引起cSrc激活、G α（i1）或G α（i3）磷酸化、G α（i1）-RGS 12或G α（i3）-RGS 12缔合以及cAMP抑制。AS-605240和PP 2对cAMP和肌肉松弛的抑制作用显著增加。结果表明，通过cSrc的G α（i1/2/3）的G β γ依赖性酪氨酸磷酸化促进了RGS 12的募集，RGS 12是一种具有独特磷酸酪氨酸结合结构域的G α（i）特异性RGS蛋白，导致G α（i）快速失活并促进平滑肌松弛。
Others and we have characterized several G beta gamma-dependent effectors in smooth muscle, including G protein-coupled receptor kinase 2 (GRK2), PLC beta(3), and phosphatidylinositol (PI) 3-kinase-gamma, and have identified various signaling targets downstream of PI 3-kinase-gamma, including cSrc, integrin-linked kinase, and Rac1-Cdc42/p21-activated kinase/p38 MAP kinase. This study identified a novel mechanism whereby G beta gamma acting via PI 3-kinase-gamma and cSrc exerts an inhibitory influence on G alpha(i) activity. The G(i2)-coupled delta-opioid receptor agonist D-penicillamine (2,5)-enkephalin (DPDPE) activated cSrc, stimulated tyrosine phosphorylation of G alpha(i2), and induced regulator of G protein signaling 12 (RGS12) association; all three events were blocked by PI 3-kinase (LY294002) and cSrc (PP2) inhibitors and by expression of the COOH-terminal sequence of GRK2-(495-689), a G beta gamma-scavenging peptide. Inhibition of forskolin-stimulated cAMP and muscle relaxation by DPDPE was augmented by PP2, LY294002, and a selective PI 3-kinase-gamma inhibitor, AS-605420. Expression of tyrosine-deficient (Y69F, Y231F, or Y321F) G alpha(i2) mutant or knockdown of RGS12 blocked G alpha(i2) phosphorylation and G alpha(i2)-RGS12 association and caused greater inhibition of cAMP. Parallel studies using somatostatin, cyclopentyl adenosine, or ACh to activate, respectively, G(i1)-coupled somatostatin (sstr(3)) receptors, and G(i3)-coupled adenosine A(1) or muscarinic m(2) receptors elicited cSrc activation, G alpha(i1) or G alpha(i3) phosphorylation, G alpha(i1)-RGS12 or G alpha(i3)-RGS12 association, and inhibition of cAMP. Inhibition of cAMP and muscle relaxation was greatly increased by AS-605240 and PP2. The results demonstrate that G beta gamma-dependent tyrosine phosphorylation of G alpha(i1/2/3) by cSrc facilitated recruitment of RGS12, a G alpha(i)-specific RGS protein with a unique phosphotyrosine-binding domain, resulting in rapid deactivation of G alpha(i) and facilitation of smooth muscle relaxation.