DIRECT EVIDENCE THAT G(I)-COUPLED RECEPTOR STIMULATION OF MITOGEN-ACTIVATED PROTEIN-KINASE IS MEDIATED BY G(BETA-GAMMA) ACTIVATION OF P21(RAS)
DIRECT EVIDENCE THAT G(I)-COUPLED RECEPTOR STIMULATION OF MITOGEN-ACTIVATED PROTEIN-KINASE IS MEDIATED BY G(BETA-GAMMA) ACTIVATION OF P21(RAS)
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DOI：
10.1073/pnas.91.26.12706
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发表时间：
1994-12-20
影响因子：
11.1
通讯作者：
LEFKOWITZ, RJ
中科院分区：
文献类型：
--
作者：
KOCH, WJ;HAWES, BE;LEFKOWITZ, RJ
G（i）偶联受体的刺激导致促分裂原活化蛋白激酶（MAP激酶）的活化。在几种细胞类型中，这似乎依赖于p21（ras）（Ras）的激活。然而，哪个G蛋白亚基（G（alpha）或G（beta gamma）复合物）主要负责触发这种信号通路尚不清楚。我们以前已经证明，β-肾上腺素能受体激酶的羧基末端，含有其G（β-γ）结合结构域，是一种细胞G（β-γ）拮抗剂，能够特异性区分G（α）和G（β-γ）介导的过程。使用这种G（β-γ）抑制剂，我们研究了Ras和MAP激酶通过Rat-1成纤维细胞中内源性G（i）偶联受体和通过瞬时转染COS-7细胞表达的受体的激活。我们在这里报告，Ras和MAP激酶激活响应溶血磷脂酸显着衰减在大鼠-1细胞稳定转染的质粒编码这种G（β-γ）拮抗剂。同样，在用编码G（i）偶联受体（α（2）-肾上腺素能和M2毒蕈碱）的质粒转染的COS-7细胞中，在共表达的G（β-γ）拮抗剂存在下，Ras和MAP激酶的活化显著降低。通过G（q）-偶联受体（α（1）-肾上腺素能）或酪氨酸激酶表皮生长因子受体介导的Ras-MAP激酶激活不受该G（β-γ）拮抗剂的影响。这些结果确定G（β-γ）作为Ras激活的主要介质，随后响应于G（i）偶联受体的刺激通过MAP激酶进行信号传导。
Stimulation of G(i)-coupled receptors leads to the activation of mitogen-activated protein kinases (MAP kinases). In several cell types, this appears to be dependent on the activation of p21(ras) (Ras). Which G-protein subunit(s) (G(alpha) or the G(beta gamma) complex) primarily is responsible for triggering this signaling pathway, however, is unclear. We have demonstrated previously that the carboxyl terminus of the beta-adrenergic receptor kinase, containing its G(beta gamma)-binding domain, is a cellular G(beta gamma) antagonist capable of specifically distinguishing G(alpha)- and G(beta gamma)-mediated processes. Using this G(beta gamma) inhibitor, we studied Ras and MAP kinase activation through endogenous G(i)-coupled receptors in Rat-1 fibroblasts and through receptors expressed by transiently transfected COS-7 cells. We report here that both Ras and MAP kinase activation in response to lysophosphatidic acid is markedly attenuated in Rat-1 cells stably transfected with a plasmid encoding this G(beta gamma) antagonist. Likewise in COS-7 cells transfected with plasmids encoding G(i)-coupled receptors (alpha(2)-adrenergic and M2 muscarinic), the activation of Ras and MAP kinase was significantly reduced in the presence of the coexpressed G(beta gamma) antagonist. Ras-MAP kinase activation mediated through a G(q)-coupled receptor (alpha(1)-adrenergic) or the tyrosine kinase epidermal growth factor receptor was unaltered by this G(beta gamma) antagonist. These results identify G(beta gamma) as the primary mediator of Ras activation and subsequent signaling via MAP kinase in response to stimulation of G(i)-coupled receptors.