Tumor necrosis factor alpha stimulation of Rac1 activity. Role of isoprenylcysteine carboxylmethyltransferase.
Tumor necrosis factor alpha stimulation of Rac1 activity. Role of isoprenylcysteine carboxylmethyltransferase.
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DOI：
10.1074/jbc.m410081200
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发表时间：
2005-05
期刊：
影响因子：
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通讯作者：
Christopher A Papaharalambus;W. Sajjad;A. Syed;Chen Zhang;M. Bergo;R. Alexander;Mushtaq Ahmad
中科院分区：
文献类型：
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作者：
Christopher A Papaharalambus;W. Sajjad;A. Syed;Chen Zhang;M. Bergo;R. Alexander;Mushtaq Ahmad
我们以前已经证明，异戊二烯半胱氨酸羧甲基转移酶（ICMT）和它的底物之一，RhoGTdR Rac 1，是至关重要的肿瘤坏死因子α（TNF α）刺激血管细胞粘附分子-1在内皮细胞（EC）的表达。在这里，我们已经表明，ICMT调节TNF α刺激Rac 1活性。EC的TNF α刺激增加了Rac 1的膜结合，这是Rac 1活性所必需的事件。ICMT抑制剂N-乙酰基-S-法呢基-L-半胱氨酸（AFC）在静息和TNF α刺激条件下均阻断了Rac 1在膜中的积累。同样，膜相关Rac 1在Icmt缺陷型小鼠胚胎成纤维细胞（MEFs）中低于野生型。TNF α还增加EC中Rac 1的活性形式GTP-Rac 1的水平。AFC完全抑制TNF α刺激GTP-Rac 1水平的增加。共聚焦显微镜显示静息EC Rac 1存在于质膜中，也存在于核周区域中。AFC错误定位Rac 1，无论是从质膜和核周区域。在Icmt缺陷型与野生型MEFs中也观察到Rac 1的错误定位。为了确定ICMT抑制的后果，我们研究了AFC对Rac 1下游的p38丝裂原活化蛋白（MAP）激酶磷酸化的影响。AFC抑制TNF α刺激EC中p38 MAP激酶磷酸化。TNF α刺激p38 MAP激酶磷酸化在Icmt缺陷型与野生型MEFs中也显著减弱。为了了解Rac 1活性抑制的机制，我们研究了ICMT抑制对Rac 1与其抑制剂Rho鸟嘌呤核苷酸解离抑制剂（RhoGDI）相互作用的影响。与野生型相比，在静息和TNF α刺激条件下，Icmt缺陷型MEFs中Rac 1与其抑制剂RhoGDI的结合显著增加，表明RhoGDI参与了ICMT抑制条件下Rac 1活性的抑制。这些结果表明，ICMT通过控制Rac 1与RhoGDI的相互作用来调节Rac 1的活性。我们假设ICMT调节Rac 1从RhoGDI的释放。
We have previously demonstrated that both isoprenylcysteine carboxylmethyltransferase (ICMT) and one of its substrates, the RhoGTPase Rac1, are critical for the tumor necrosis factor alpha (TNF alpha) stimulation of vascular cell adhesion molecule-1 expression in endothelial cells (EC). Here, we have shown that ICMT regulates TNF alpha stimulation of Rac1 activity. TNF alpha stimulation of EC increased the membrane association of Rac1, an event that is essential for Rac1 activity. ICMT inhibitor N-acetyl-S-farnesyl-L-cysteine (AFC) blocked the accumulation of Rac1 into the membrane both in resting and TNF alpha-stimulated conditions. Similarly, the membrane-associated Rac1 was lower in Icmt-deficient versus wild-type mouse embryonic fibroblasts (MEFs). TNF alpha also increased the level of GTP-Rac1, the active form of Rac1, in EC. AFC completely suppressed the TNF alpha stimulation of increase in GTP-Rac1 levels. Confocal microscopy revealed resting EC Rac1 was present in the plasma membrane and also in the perinuclear region. AFC mislocalized Rac1, both from the plasma membrane and the perinuclear region. Mislocalization of Rac1 was also observed in Icmt-deficient versus wild-type MEFs. To determine the consequences of ICMT inhibition, we investigated the effect of AFC on p38 mitogen-activated protein (MAP) kinase phosphorylation, which is downstream of Rac1. AFC inhibited the TNF alpha stimulation of p38 MAP kinase phosphorylation in EC. TNF alpha stimulation of p38 MAP kinase phosphorylation was also significantly attenuated in Icmt-deficient versus wild-type MEFs. To understand the mechanism of inhibition of Rac1 activity, we examined the effect of ICMT inhibition on the interaction of Rac1 with its inhibitor, Rho guanine nucleotide dissociation inhibitor (RhoGDI). The association of Rac1 with its inhibitor RhoGDI was dramatically increased in the Icmt-deficient versus wild-type MEFs both in resting as well as in TNF alpha-stimulated conditions, suggesting that RhoGDI was involved in inhibiting Rac1 activity under the conditions of ICMT inhibition. These results suggest that ICMT regulates Rac1 activity by controlling the interaction of Rac1 with RhoGDI. We hypothesize that ICMT regulates the release of Rac1 from RhoGDI.