FIGURE 4.

TWEAK-Fn14 activation of Rac1 is dependent upon SGEF and RhoG. A, U87 and U118 glioma cells stably expressing either control empty vector (Ctrl) or shRNA targeting SGEF (SGEF-12) were cultured overnight in reduced serum medium (0.5% FBS). Cells were left untreated or treated with TWEAK for 5 min. Cells were lysed and assessed for levels of active Rac1. WB, Western blot. B, U87 glioma cells were transfected with one of two independent siRNA oligonucleotides targeting RhoG (RhoG-1 and RhoG-2), with siRNA targeting non-mammalian luciferase (Ctrl), or left untreated (NT). After 72 h, cell lysates were immunoblotted with the indicated antibodies. C, U87 glioma cells were transfected with siRNA targeting non-mammalian luciferase (Ctrl) or with a siRNA oligonucleotide targeting RhoG (RhoG-2). 48 h after transfection, cells were cultured an additional 16 h overnight in reduced-serum medium (0.5% FBS) prior to addition of TWEAK treatment for 5 min in the indicated samples. Cells were lysed and assessed for levels of Rac1 activity. Data are representative of two independent experiments.