United Graduate School of Drug Discovery and Medical Information Sciences, Gifu University, Yanagido, Gifu 501-1193, Japan.
Department of Chemistry and Biomolecular Science, Faculty of Engineering, Gifu University, Yanagido, Gifu 501-1193, Japan.
Cell Signal. 2014 Apr;26(4):691-6. doi: 10.1016/j.cellsig.2013.12.006. Epub 2013 Dec 27.
PLEKHG2/FLJ00018, a Rho family-specific guanine nucleotide exchange factor (RhoGEF), is activated by heterotrimeric GTP-binding protein (G protein) Gβγ subunits, and in turn activates the small G protein Rac and Cdc42, which have been shown to mediate signaling pathways leading to actin cytoskeletal reorganization. In the present study, we show that co-expression of the constitutively active mutant of cSrc, a non-receptor tyrosine kinase, and PLEKHG2 induced the tyrosine phosphorylation of PLEKHG2 in HEK293 cells. Through deletion and base substitution mutagenesis we have identified Tyr489 of PLEKHG2 as the site phosphorylated by cSrc. Furthermore, using a high-throughput src homology 2 (SH2) domain binding assay, the SH2 domain of ABL1 and the PI 3-kinse regulator subunit (PIK3R3) were identified as candidates for the binding partner of tyrosine-phosphorylated PLEKHG2. The interaction between PLEKHG2 and the full-length of PIK3R3, but not ABL1, occurs in a tyrosine-phosphorylation-dependent manner. Furthermore, PLEKHG2 is tyrosine phosphorylated at Tyr489 by ephrinB2 receptor signaling via cSrc. Investigation of the physiological function of tyrosine phosphorylation at Tyr489 in PLEKHG2 remains a subject for future studies.
PLEKHG2/FLJ00018 是一种 Rho 家族特异性鸟嘌呤核苷酸交换因子（RhoGEF），可被异三聚体 G 蛋白结合蛋白（G 蛋白）Gβγ亚基激活，进而激活小 G 蛋白 Rac 和 Cdc42，这些小 G 蛋白已被证明可介导导致肌动蛋白细胞骨架重排的信号通路。在本研究中，我们表明，组成型激活突变体 cSrc（一种非受体酪氨酸激酶）与 PLEKHG2 的共表达可诱导 HEK293 细胞中 PLEKHG2 的酪氨酸磷酸化。通过缺失和碱基取代突变，我们确定 PLEKHG2 的 Tyr489 是 cSrc 磷酸化的位点。此外，通过高通量 src 同源性 2（SH2）结构域结合测定法，鉴定出 ABL1 的 SH2 结构域和 PI 3-激酶调节亚基（PIK3R3）是酪氨酸磷酸化 PLEKHG2 的结合伴侣的候选物。在酪氨酸磷酸化依赖性方式下，PLEKHG2 与全长 PIK3R3 而不是 ABL1 发生相互作用。此外，通过 cSrc 介导的 EphrinB2 受体信号转导，PLEKHG2 在 Tyr489 发生酪氨酸磷酸化。在 PLEKHG2 中 Tyr489 的酪氨酸磷酸化的生理功能的研究仍是未来研究的课题。
