Grant S L, Lassègue B, Griendling K K, Ushio-Fukai M, Lyons P R, Alexander R W
Department of Medicine, Division of Cardiology, Emory University, Atlanta, Georgia, USA.
Mol Pharmacol. 2000 Mar;57(3):460-7. doi: 10.1124/mol.57.3.460.
The effects of angiotensin II (Ang II) are mediated primarily by Ang II type 1 receptors, which in turn are coupled to heterotrimeric G proteins. After receptor activation, the G(alpha) and G(betagamma) subunits dissociate, contributing to the signaling cascades involving protein kinase C (PKC) activation. Regulators of G protein signaling (RGS proteins) comprise a class of proteins that have been shown to negatively regulate the G(alpha) subunit. We examined which RGS sequences were expressed in vascular smooth muscle cells and which of these were regulated by Ang II. Reverse transcription-polymerase chain reaction showed that of 16 RGS sequences screened, six RGS transcripts (RGS2, 3, 10, 11, and 12 and GAIP) were present. Northern blot analysis demonstrated that RGS3, 10, and 12 and GAIP were not regulated by Ang II at the mRNA level. In contrast, RGS2 mRNA was rapidly and dose dependently increased (395 +/- 24% peak, 45 min) by Ang II but returned to baseline level by 6 to 8 h. Phorbol-12-myristate-13-acetate, a PKC activator, robustly increased RGS2. This signal was attenuated by the PKC inhibitor GF 109203X (50 +/- 4%) and by phorbol-12, 13-dibutyrate-mediated down-regulation of PKC (48 +/- 13%). Tyrosine kinase inhibition and calcium deprivation did not affect the up-regulation of RGS2 mRNA after Ang II stimulation. Actinomycin D treatment inhibited both Ang II- and phorbol-12-myristate-13-acetate-stimulated RGS2 up-regulation, suggesting activation of transcription by these agonists. The stability of RGS2 mRNA did not appear to be affected by Ang II. Thus, RGS2 is a likely candidate for negative regulation of the G proteins coupled to the Ang II type 1 receptor in vascular smooth muscle cells. Regulation of this protein may be of critical importance in modulating the role of Ang II in vascular disease.
血管紧张素II（Ang II）的作用主要由血管紧张素II 1型受体介导，该受体又与异源三聚体G蛋白偶联。受体激活后，G（α）和G（βγ）亚基解离，参与涉及蛋白激酶C（PKC）激活的信号级联反应。G蛋白信号调节剂（RGS蛋白）包括一类已被证明对G（α）亚基具有负调节作用的蛋白质。我们研究了哪些RGS序列在血管平滑肌细胞中表达，以及其中哪些受Ang II调节。逆转录-聚合酶链反应显示，在筛选的16个RGS序列中，有6个RGS转录本（RGS2、3、10、11和12以及GAIP）存在。Northern印迹分析表明，RGS3、10和12以及GAIP在mRNA水平不受Ang II调节。相反，Ang II可使RGS2 mRNA迅速且呈剂量依赖性增加（峰值为395±24%，45分钟），但在6至8小时后恢复到基线水平。佛波醇-12-肉豆蔻酸酯-13-乙酸酯（一种PKC激活剂）可显著增加RGS2。该信号被PKC抑制剂GF 109203X减弱（50±4%），并被佛波醇-12,13-二丁酸酯介导的PKC下调减弱（48±13%）。酪氨酸激酶抑制和钙剥夺不影响Ang II刺激后RGS2 mRNA的上调。放线菌素D处理可抑制Ang II和佛波醇-12-肉豆蔻酸酯-13-乙酸酯刺激的RGS2上调，提示这些激动剂可激活转录。RGS2 mRNA的稳定性似乎不受Ang II影响。因此，RGS2可能是血管平滑肌细胞中与血管紧张素II 1型受体偶联的G蛋白负调节的候选者。该蛋白的调节可能对调节Ang II在血管疾病中的作用至关重要。
