Cloning and characterization of GEF-H1, a microtubule-associated guanine nucleotide exchange factor for Rac and Rho GTPases
Cloning and characterization of GEF-H1, a microtubule-associated guanine nucleotide exchange factor for Rac and Rho GTPases
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DOI：
10.1074/jbc.273.52.34954
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发表时间：
1998-12-25
影响因子：
4.8
通讯作者：
Busch, H
中科院分区：
文献类型：
--
作者：
Ren, Y;Li, R;Busch, H
Rho相关的小G蛋白酶是参与调节细胞外刺激物到细胞核和细胞骨架的信号转导级联的关键元件，Dbl样鸟嘌呤核苷酸交换因子（GEF）参与了这些G蛋白酶的直接激活。在这里，我们已经确定了Dbl家族的新成员，GEF-H1，通过筛选人HeLa细胞cDNA文库。GEF-H1编码一种100-kDa的蛋白质，该蛋白质含有与普列克底物蛋白同源结构域串联的Db 1同源结构域的保守结构阵列，并且与lfc癌基因最密切相关，但另外它在羧基末端含有一个独特的卷曲螺旋结构域。生物化学分析表明，GEF-H1能够刺激Rac和Rho的鸟嘌呤核苷酸交换，但对Cdc 42、TC 10、此外，GEF-H1在GDP-和鸟苷5 ′-3-O-（硫代）三磷酸结合状态下结合Rac和Rho蛋白，而对Cdc 42或Ras没有可检测的亲和力。免疫荧光显示GEF-H1通过羧基末端卷曲螺旋结构域与微管共定位。GEF-H1在COS-7细胞中的过表达导致膜皱褶的诱导。这些结果表明，GEF-H1可能在Rac和/或Rho的激活中起直接作用，并将激活的GT3带到特定的靶位点，如微管。
The Rho-related small GTPases are critical elements involved in regulation of signal transduction cascades from extracellular stimuli to cell nucleus and cytoskeleton, The Dbl-like guanine nucleotide exchange factors (GEF) have been implicated in direct activation of these GTPases. Here we have identified a new member of the Dbl family, GEF-H1, by screening a human HeLa cell cDNA library. GEF-H1 encodes a 100-kDa protein containing the conserved structural array of a Dbl homology domain in tandem with a pleckstrin homology domain and is most closely related to the lfc oncogene, but additionally it contains a unique coiled-coil domain at the carboxyl terminus, Biochemical analysis reveals that GEF-H1 is capable of stimulating guanine nucleotide exchange of Rac and Rho but is inactive toward Cdc42, TC10, or Ras, Moreover, GEF-H1 binds to Rac and Rho proteins in both the GDP- and guanosine 5'-3-O-(thio)triphosphate-bound states without detectable affinity for Cdc42 or Ras. Immunofluorescence reveals that GEF-H1 colocalizes with microtubules through the carboxyl-terminal coiled-coil domain. Overexpression of GEF-H1 in COS-7 cells results in induction of membrane ruffles. These results suggest that GEF-H1 may have a direct role in activation of Rac and/or Rho and in bringing the activated GTPase to specific target sites such as microtubules.