Taniyama Yoshihiro, Ushio-Fukai Masuko, Hitomi Hirofumi, Rocic Petra, Kingsley Michael J, Pfahnl Chun, Weber David S, Alexander R Wayne, Griendling Kathy K
Division of Cardiology, Department of Medicine, Emory University, 319 WMB, 1639 Pierce Drive, Atlanta, GA 30322, USA.
Am J Physiol Cell Physiol. 2004 Aug;287(2):C494-9. doi: 10.1152/ajpcell.00439.2003. Epub 2004 Apr 14.
Angiotensin II activates a variety of signaling pathways in vascular smooth muscle cells (VSMCs), including the MAPKs and Akt, both of which are required for hypertrophy. However, little is known about the relationship between these kinases or about the upstream activators of Akt. In this study, we tested the hypothesis that the reactive oxygen species (ROS)-sensitive kinase p38 MAPK and its substrate MAPKAPK-2 mediate Akt activation in VSMCs. In unstimulated VSMCs, Akt and p38 MAPK are constitutively associated and remain so after angiotensin II stimulation. Inhibition of p38 MAPK activity with SB-203580 dose-dependently inhibits Akt phosphorylation on Ser(473), but not Thr(308). Angiotensin II-induced phosphorylation of MAPKAPK-2 is also attenuated by SB-203580, as well as by inhibitors of ROS. In addition, angiotensin II stimulates the association of MAPKAPK-2 with the Akt-p38 MAPK complex, and an in vitro kinase assay shows that MAPKAPK-2 immunoprecipitates of VSMC lysates phosphorylate recombinant Akt in an angiotensin II-inducible manner. Finally, intracellular delivery of a MAPKAPK-2 peptide inhibitor blocks Akt phosphorylation on Ser(473). These results suggest that the p38 MAPK-MAPKAPK-2 pathway mediates Akt activation by angiotensin II in these cells by recruiting active MAPKAPK-2 to a signaling complex that includes both Akt and p38 MAPK. Through this mechanism, p38 MAPK confers ROS sensitivity to Akt and facilitates downstream signaling. These results provide evidence for a novel signaling complex that may help to spatially organize hypertrophy-related, ROS-sensitive signaling in VSMCs.
血管紧张素II激活血管平滑肌细胞（VSMC）中的多种信号通路，包括丝裂原活化蛋白激酶（MAPK）和Akt，二者均为细胞肥大所必需。然而，关于这些激酶之间的关系或Akt的上游激活剂，人们所知甚少。在本研究中，我们检验了以下假设：活性氧（ROS）敏感激酶p38 MAPK及其底物MAPKAPK - 2介导VSMC中Akt的激活。在未受刺激的VSMC中，Akt和p38 MAPK组成性结合，且在血管紧张素II刺激后仍保持结合状态。用SB - 203580抑制p38 MAPK活性可剂量依赖性地抑制Akt在Ser（473）位点的磷酸化，但不影响Thr（308）位点的磷酸化。SB - 203580以及ROS抑制剂也可减弱血管紧张素II诱导的MAPKAPK - 2磷酸化。此外，血管紧张素II刺激MAPKAPK - 2与Akt - p38 MAPK复合物的结合，体外激酶分析表明，VSMC裂解物的MAPKAPK - 2免疫沉淀物以血管紧张素II诱导的方式磷酸化重组Akt。最后，细胞内递送MAPKAPK - 2肽抑制剂可阻断Akt在Ser（473）位点的磷酸化。这些结果表明，p38 MAPK - MAPKAPK - 2通路通过将活性MAPKAPK - 2募集到包含Akt和p38 MAPK的信号复合物中，介导血管紧张素II在这些细胞中激活Akt。通过这种机制，p38 MAPK赋予Akt对ROS的敏感性并促进下游信号传导。这些结果为一种新型信号复合物提供了证据，该复合物可能有助于在空间上组织VSMC中与肥大相关的、对ROS敏感的信号传导。
