Sugimoto Naotoshi, Takuwa Noriko, Okamoto Hiroyuki, Sakurada Sotaro, Takuwa Yoh
Department of Physiology, Kanazawa University Graduate School of Medicine, Kanazawa, Ishikawa 920-8640, Japan.
Mol Cell Biol. 2003 Mar;23(5):1534-45. doi: 10.1128/MCB.23.5.1534-1545.2003.
The G protein-coupled receptors S1P2/Edg5 and S1P3/Edg3 both mediate sphingosine-1-phosphate (S1P) stimulation of Rho, yet S1P2 but not S1P3 mediates downregulation of Rac activation, membrane ruffling, and cell migration in response to chemoattractants. Specific inhibition of endogenous Galpha12 and Galpha13, but not of Galphaq, by expression of respective C-terminal peptides abolished S1P2-mediated inhibition of Rac, membrane ruffling, and migration, as well as stimulation of Rho and stress fiber formation. Fusion receptors comprising S1P2 and either Galpha12 or Galpha13, but not Galphaq, mediated S1P stimulation of Rho and also inhibition of Rac and migration. Overexpression of Galphai, by contrast, specifically antagonized S1P2-mediated inhibition of Rac and migration. The S1P2 actions were mimicked by expression of V14Rho and were abolished by C3 toxin and N19Rho, but not Rho kinase inhibitors. In contrast to S1P2, S1P3 mediated S1P-directed, pertussis toxin-sensitive chemotaxis and Rac activation despite concurrent stimulation of Rho via G12/13. Upon inactivation of Gi by pertussis toxin, S1P3 mediated inhibition of Rac and migration just like S1P2. These results indicate that integration of counteracting signals from the Gi- and the G12/13-Rho pathways directs either positive or negative regulation of Rac, and thus cell migration, upon activation of a single S1P receptor isoform.
G蛋白偶联受体S1P2/Edg5和S1P3/Edg3均介导1-磷酸鞘氨醇（S1P）对Rho的刺激作用，但S1P2而非S1P3介导对趋化因子应答时Rac激活、膜皱褶和细胞迁移的下调。通过表达各自的C末端肽特异性抑制内源性Gα12和Gα13而非Gαq，消除了S1P2介导的对Rac的抑制、膜皱褶和迁移，以及对Rho的刺激和应力纤维形成。包含S1P2与Gα12或Gα13而非Gαq的融合受体介导了S1P对Rho的刺激以及对Rac和迁移的抑制。相比之下，Gαi的过表达特异性拮抗了S1P2介导的对Rac和迁移的抑制。S1P2的作用可被V14Rho的表达模拟，并被C3毒素和N19Rho消除，但不能被Rho激酶抑制剂消除。与S1P2不同，S1P3介导了S1P导向的、百日咳毒素敏感的趋化作用和Rac激活，尽管同时通过G12/13刺激了Rho。在用百日咳毒素使Gi失活后，S1P3介导了对Rac和迁移的抑制，就像S1P2一样。这些结果表明，来自Gi和G12/13-Rho途径的拮抗信号的整合在单个S1P受体亚型激活时指导Rac的正向或负向调节，从而指导细胞迁移。
