TGF -induced RhoA activation and fibronectin production in mesangial cells require caveolae

Peng F, Zhang B, Wu D, Ingram AJ, Gao B, Krepinsky JC. TGF -induced RhoA activation and fibronectin production in mesangial cells require caveolae. Am J Physiol Renal Physiol 295: F153–F164, 2008. First published April 23, 2008; doi:10.1152/ajprenal.00419.2007.—Glomerular sclerosis of diverse etiologies is characterized by mesangial matrix accumulation, with transforming growth factor(TGF ) an important pathogenic factor. The GTPase RhoA mediates TGF -induced matrix accumulation in some settings. Here we study the role of the membrane microdomain caveolae in TGF -induced RhoA activation and fibronectin upregulation in mesangial cells (MC). In primary rat MC, TGF 1 time dependently increased RhoA and downstream Rho kinase activation. Rho pathway inhibition blocked TGF 1-induced upregulation of fibronectin transcript and protein. TGF 1-induced RhoA activation was prevented by disrupting caveolae with cholesterol depletion and rescued by cholesterol repletion. Compared with wild types, RhoA/ Rho kinase activation was absent in MC lacking caveolae. Reexpression of caveolin-1 (and caveolae) restored these responses. Phosphorylation of caveolin-1 on Y14, effected by Src kinases, has been implicated in signaling responses. Overexpression of nonphosphorylatable caveolin-1 Y14A prevented TGF 1-induced RhoA activation. TGF 1 also activated Src, and its inhibition blocked RhoA activation. Furthermore, TGF 1 led to association of RhoA and caveolin-1. This was prevented by Src or TGF receptor I inhibition, and by caveolin-1 Y14A overexpression. Last, fibronectin upregulation by TGF 1 was blocked by Src inhibition, not seen in caveolin-1 knockout MC, and restored by caveolin-1 reexpression in the latter. TGF 1-induced collagen I accumulation also required caveolae. TGF 1-mediated Smad2/3 activation, however, did not require caveolae. We conclude that RhoA/Rho kinase mediates TGF -induced fibronectin upregulation. This requires caveolae and caveolin-1 interaction with RhoA. Interference with caveolin/caveolae or RhoA signaling thus represents a potential target for the treatment of fibrotic renal disease.