Ezrin is highly expressed in glomerular podocytes and is reported to form a multi-protein complex with scaffold protein Na+/H+ exchanger regulatory factor 2 (NHERF2) and podocalyxin, a major sialoprotein. Podocalyxin-knockout mice died within 24 h of birth with anuric renal failure, whereas NHERF2-knockout mice show no apparent changes in the glomerular functions. However, the physiological roles of ezrin in glomerular podocytes remain unclear. Here, we investigated the importance of ezrin in the regulation of glomerular podocyte function using ezrin-knockdown mice (Vil2 kd/kd ). The Vil2 kd/kd mice did not exhibit apparent glomerular dysfunction, morphological defects or abnormal localisation of podocalyxin and NHERF2 in podocytes. Thus, we investigated the influence of ezrin defects on Rho-GTPase activity, as ezrin interacts with the Rho-GTPase dissociation inhibitor (Rho-GDI), which plays a key role in the regulation of podocyte actin organisation. In Vil2 kd/kd glomeruli, Rac1 activity was significantly reduced compared to wildtype (WT) glomeruli at baseline. Furthermore, Vil2 kd/kd mice showed reduced susceptibility to glomerular injury. In WT glomeruli, Rac1 activity was enhanced in nephrotic conditions, but remained at baseline levels in Vil2 kd/kd glomeruli, suggesting that loss of ezrin protects podocytes from injury-induced morphological changes by suppressing Rac1 activation.