Mechanical stretch-induced RhoA activation is mediated by the RhoGEF Vav2 in mesangial cells

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. (C) 2009 Elsevier B.V. All rights reserved.