Peng Fangfang, Zhang Baifang, Ingram Alistair J, Gao Bo, Zhang Ying, Krepinsky Joan C
Division of Nephrology, McMaster University, Hamilton, Canada.
Cell Signal. 2010 Jan;22(1):34-40. doi: 10.1016/j.cellsig.2009.09.003. Epub 2009 Sep 13.
Increased intraglomerular pressure is an important hemodynamic determinant of glomerulosclerosis, and can be modelled in vitro by exposing mesangial cells (MC) to cyclic mechanical stretch. We have previously shown that the GTPase RhoA mediates stretch-induced fibronectin production. Here we investigate the role of the RhoGEF Vav2 in the activation of RhoA by stretch. Primary rat MC were exposed to 1 Hz cyclic stretch, previously shown to induce maximal RhoA activation at 1 min. Total Vav2 tyrosine phosphorylation and specific phosphorylation on Y172, required for activation, were increased by 1 min of stretch. Overexpression of dominant-negative Vav2 Y172/159F in COS-1 cells or downregulation of Vav2 by siRNA in MC prevented stretch-induced RhoA activation. Vav2 is known to be activated in response to growth factors, and we have previously shown the epidermal growth factor receptor (EGFR) to be transactivated by stretch in MC. Both Vav2 Y172 phosphorylation and RhoA activation were blocked by the EGFR inhibitor AG1478 and prevented in MC overexpressing kinase inactive EGFR. Stretch led to physical association between the EGFR and Vav2, and this was dependent on EGFR activation. EGFR Y992 phosphorylation, required for growth factor-induced Vav2 phosphorylation, was also induced by stretch. Activation of both Src and PI3K were necessary upstream mediators of stretch-induced Vav2 Y172 phosphorylation and RhoA activation. In summary, stretch-induced RhoA activation is dependent on transactivation of the EGFR and activation of the RhoGEF Vav2. Src and PI3K are both required upstream of Vav2 and RhoA activation.
肾小球内压升高是肾小球硬化的一个重要血流动力学决定因素，并且可以通过将系膜细胞（MC）暴露于周期性机械牵张在体外进行模拟。我们之前已经表明，GTP酶RhoA介导牵张诱导的纤连蛋白产生。在此，我们研究Rho鸟嘌呤核苷酸交换因子Vav2在牵张激活RhoA中的作用。原代大鼠MC暴露于1 Hz的周期性牵张，先前已表明该牵张在1分钟时可诱导最大程度的RhoA激活。牵张1分钟可使激活所需的总Vav2酪氨酸磷酸化以及Y172位点的特异性磷酸化增加。在COS-1细胞中过表达显性负性Vav2 Y172/159F或在MC中通过小干扰RNA下调Vav2可阻止牵张诱导的RhoA激活。已知Vav2可响应生长因子而被激活，并且我们之前已经表明表皮生长因子受体（EGFR）在MC中可被牵张反式激活。EGFR抑制剂AG1478可阻断Vav2 Y172磷酸化和RhoA激活，并且在过表达激酶失活型EGFR的MC中可阻止这种情况发生。牵张导致EGFR与Vav2之间发生物理结合，并且这依赖于EGFR激活。生长因子诱导的Vav2磷酸化所需的EGFR Y992磷酸化也可由牵张诱导。Src和PI3K的激活都是牵张诱导的Vav2 Y172磷酸化和RhoA激活的必要上游介质。总之，牵张诱导的RhoA激活依赖于EGFR的反式激活和Rho鸟嘌呤核苷酸交换因子Vav2的激活。Src和PI3K都是Vav2和RhoA激活上游所必需的。
