RAS RECRUITS RAF-1 TO THE PLASMA-MEMBRANE FOR ACTIVATION BY TYROSINE PHOSPHORYLATION
RAS RECRUITS RAF-1 TO THE PLASMA-MEMBRANE FOR ACTIVATION BY TYROSINE PHOSPHORYLATION
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DOI：
10.1002/j.1460-2075.1995.tb07316.x
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发表时间：
1995-07-03
期刊：
影响因子：
11.4
通讯作者：
MARSHALL, CJ
中科院分区：
文献类型：
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作者：
MARAIS, R;LIGHT, Y;MARSHALL, CJ
受体和致癌酪氨酸激酶下游信号转导的中心特征是由Raf-1、Mek（MAP激酶激酶）和ERK（MAP激酶）组成的蛋白激酶级联的Ras依赖性活化。为了研究酪氨酸激酶活性在Raf-1激活中的作用，我们研究了p74 Raf-1和致癌Src的特性，这些特性是p74 Raf-1激活所必需的。我们发现，在哺乳动物细胞中，致癌Src激活p74 Raf-1需要pp 60 Src被豆蔻酰化，并且p74 Raf-1能够与p21 Ras-GTP相互作用。Ras/Raf相互作用是p21 Ras-GTP将p74 Raf-1带到质膜上进行酪氨酸340或341磷酸化所必需的，可能是通过膜结合的pp 60 Src。当致癌Src与Raf-1一起表达时，p74 Raf-1被激活5倍;然而，当与致癌Ras和Src共表达时，Raf-1被激活25倍，这与酪氨酸磷酸化进一步增加3倍有关。因此，p21 Ras-GTP是将p74 Raf-1带到质膜进行酪氨酸磷酸化的限制性组分。使用突变体的Raf-1在Tyr 340/341，我们表明，除了在这些网站的酪氨酸磷酸化，有一个额外的激活步骤导致从p21 Ras-GTP招聘p74 Raf-1的质膜。因此，Ras在Raf-1活化中的作用是将p74 Raf-1带到质膜上，进行至少两个不同的活化步骤。
A central feature of signal transduction downstream of both receptor and oncogenic tyrosine kinases is the Ras-dependent activation of a protein kinase cascade consisting of Raf-1, Mek (MAP kinase kinase) and ERKs (MAP kinases). To study the role of tyrosine kinase activity in the activation of Raf-1, we have examined the properties of p74Raf-1 and oncogenic Src that are necessary for activation of p74Raf-1. We show that in mammalian cells activation of p74Raf-1 by oncogenic Src requires pp60Src to be myristoylated and the ability of p74Raf-1 to interact with p21Ras-GTP. The Ras/Raf interaction is required for p21Ras-GTP to bring p74Raf-1 to the plasma membrane for phosphorylation at tyrosine 340 or 341, probably by membrane-bound pp60Src. When oncogenic Src is expressed with Raf-1, p74Raf-1 is activated 5-fold; however, when co-expressed with oncogenic Ras and Src, Raf-1 is activated 25-fold and this is associated with a further 3-fold increase in tyrosine phosphorylation. Thus, p21Ras-GTP is the Limiting component in bringing p74Raf-1 to the plasma membrane for tyrosine phosphoryIation. Using mutants of Raf-1 at Tyr340/341, we show that in addition to tyrosine phosphorylation at these sites, there is an additional activation step resulting from p21Ras-GTP recruiting p74Raf-1 to the plasma membrane. Thus, the role of Ras in Raf-1 activation is to bring p74Raf-1 to the plasma membrane for at least two different activation steps.