Coupling of endothelin receptors to the ERK/MAP kinase pathway -: Roles of palmitoylation and Gαq
Coupling of endothelin receptors to the ERK/MAP kinase pathway -: Roles of palmitoylation and Gαq
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DOI：
10.1046/j.0014-2956.2001.02486.x
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发表时间：
2001-10-01
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影响因子：
--
通讯作者：
Schroeder, C
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文献类型：
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作者：
Cramer, H;Schmenger, K;Schroeder, C
内皮素是有效的有丝分裂原，其通过其同源G蛋白偶联受体ETA和ETB刺激细胞外信号调节激酶（ERK/MAP激酶）。为了解决翻译后ET受体修饰的作用，如酰化对ERK激活，并确定相关的下游效应器耦合ET受体的ERK信号转导级联，我们构建了一个面板的棕榈酰化缺陷ET受体突变体与差异Got蛋白结合能力。内皮素-1刺激野生型ETA或ETB诱导5倍至6倍增加ERK在COS-7和CHO细胞，而全长nonpalmitoylated ETA和ETB突变体不能刺激ERK。缺少C-末端尾部结构域（包括推定的磷酸化和抑制蛋白结合位点）但保留关键棕榈酰化位点的截短的ETB仍然能够完全刺激ERK活化。使用具有选择性G蛋白偶联的突变的ET受体，我们发现内皮素诱导的G α（q）的刺激，而不是G α（i）或G α（s）的刺激，对于内皮素介导的ERK激活是必需的。抑制蛋白激酶A和C或表皮生长因子受体激酶未能阻止ETA和ETB介导的ERK激活，而磷脂酶C-P的阻断完全废除了重组COS-7和天然C6细胞中通过ETA和ETB的内皮素促进的ERK激活。Ca ~（2+）复合物的形成或Src家族酪氨酸激酶的抑制可阻止ET-1诱导的C6-细胞ERK-2活化。我们的研究结果表明，内皮素促进的ERK/MAPK激活关键取决于棕榈酰化，而不是ET受体的磷酸化，和G α（q）/磷脂酶C-β/Ca 2 +/Src信号级联是必要的有效耦合ET受体的ERK/MAPK途径。
Endothelins are potent mitogens that stimulate extracellular signal-regulated kinases (ERK/MAP kinases) through their cognate G-protein-coupled receptors, ETA and ETB. To address the role of post-translational ET receptor modifications such as acylation on ERK activation and to identify relevant downstream effectors coupling the ET receptor to the ERK signaling cascades we have constructed a panel of palmitoylation-deficient ET receptor mutants with differential Got protein binding capacity. Endothelin-1 stimulation of wild-type ETA or ETB induced a fivefold to sixfold increase in ERK in COS-7 and CHO cells whereas full-length nonpalmitoylated ETA and ETB mutants failed to stimulate ERK. A truncated ETB lacking the C-terminal tail domain including putative phosphorylation and arrestin binding site(s) but retaining the critical palmitoylation site(s) was still able to fully stimulate ERK activation. Using mutated ET receptors with selective G-protein-coupling we found that endothelin-induced stimulation of G alpha (q), but not of G alpha (i) or G alpha (s), is essential for endothelin-mediated ERK activation. Inhibition of protein kinases A and C or epidermal growth factor receptor kinase failed to prevent ETA- and ETB-mediated ERK activation whereas blockage of phospholipase C-P completely abrogated endothelin-promoted ERK activation through ETA and ETB in recombinant COS-7 and native C6 cells. Complex formation of Ca2+ or inhibition of Src family tyrosine kinases prevented ET-1-induced ERK-2 activation in C6-cells. Our results indicate that endothelin-promoted ERK/MAPK activation criticially depends on palmitoylation but not on phosphorylation of ET receptors, and that the G alpha (q)/phospholipase C-beta /Ca2+/Src signaling cascade is necessary for efficient coupling of ET receptors to the ERK/MAPK pathway.