Signals from the Ras, Rac, and Rho GTPases converge on the Pak protein kinase in Rat-1 fibroblasts.
Signals from the Ras, Rac, and Rho GTPases converge on the Pak protein kinase in Rat-1 fibroblasts.
复制标题
来自 Ras、Rac 和 Rho GTPases 的信号汇聚到 Rat-1 成纤维细胞中的 Pak 蛋白激酶上。
DOI：
10.1128/mcb.19.3.1881
复制
发表时间：
1999
影响因子：
5.3
通讯作者：
Field,J
中科院分区：
文献类型：
--
作者：
Tang,Y;Yu,J;Field,J
Ras在调节细胞增殖、分化和转化中起关键作用。在Ras > Raf > Mek >细胞外信号激活激酶（ERK）级联中，Raf是Ras的主要效应子。第二个效应子是磷脂酰肌醇3-OH激酶（PI 3-激酶），其反过来激活小G蛋白Rac。Rac还具有多种效应子，其中之一是丝氨酸苏氨酸激酶Pak（p65 Pak）。在这里，我们表明，Ras，而不是Raf，激活Pak 1在大鼠-1细胞的共转染试验，但不是NIH 3 T3细胞。我们测试了激活或阻断Ras下游特定组分的试剂，并证明Ras > PI 3-激酶> Rac/Cdc 42> Pak信号。虽然这些研究表明，信号从Ras通过PI 3-激酶是足以激活Pak，其他研究表明，其他效应器有助于Pak激活。RasV 12 S35和RasV 12 G37是两种不能激活PI 3-激酶的效应突变体蛋白，它们单独转染时不能激活Pak，但共转染时能激活Pak。类似地，不结合Pak的效应突变体RacV 12 H40和Rho都与Raf合作激活Pak。显性负性Rho突变体也抑制Pak的Ras激活。Rac/Raf和Ras/Raf和Rho/Raf效应突变体的所有组合协同转化细胞刺激ERK。合作是Pak依赖的，因为所有的组合被抑制激酶缺陷型Pak突变体在转化试验和ERK激活试验。这些数据表明，其他Ras效应物可以与PI 3-激酶合作，并相互合作，以激活Pak。此外，Pak激活和合作转化之间的强相关性表明，Pak激活是必要的，虽然不是足够的，Rat-1成纤维细胞的Ras，Rac，和Rho的合作转化。
Ras plays a key role in regulating cellular proliferation, differentiation, and transformation. Raf is the major effector of Ras in the Ras > Raf > Mek > extracellular signal-activated kinase (ERK) cascade. A second effector is phosphoinositide 3-OH kinase (PI 3-kinase), which, in turn, activates the small G protein Rac. Rac also has multiple effectors, one of which is the serine threonine kinase Pak (p65Pak). Here we show that Ras, but not Raf, activates Pak1 in cotransfection assays of Rat-1 cells but not NIH 3T3 cells. We tested agents that activate or block specific components downstream of Ras and demonstrate a Ras > PI 3-kinase > Rac/Cdc42 > Pak signal. Although these studies suggest that the signal from Ras through PI 3-kinase is sufficient to activate Pak, additional studies suggested that other effectors contribute to Pak activation. RasV12S35and RasV12G37, two effector mutant proteins which fail to activate PI 3-kinase, did not activate Pak when tested alone but activated Pak when they were cotransfected. Similarly, RacV12H40, an effector mutant that does not bind Pak, and Rho both cooperated with Raf to activate Pak. A dominant negative Rho mutant also inhibited Ras activation of Pak. All combinations of Rac/Raf and Ras/Raf and Rho/Raf effector mutants that transform cells cooperatively stimulated ERK. Cooperation was Pak dependent, since all combinations were inhibited by kinase-deficient Pak mutants in both transformation assays and ERK activation assays. These data suggest that other Ras effectors can collaborate with PI 3-kinase and with each other to activate Pak. Furthermore, the strong correlation between Pak activation and cooperative transformation suggests that Pak activation is necessary, although not sufficient, for cooperative transformation of Rat-1 fibroblasts by Ras, Rac, and Rho.