Fig. 5

SRGN induces cytoskeleton reorganization and Rho-family GTPase activation. a-c Phalloidin staining of actin. H1299/Mock and H1299/SRGN cells (a) were incubated in serum-free medium for 24 h and H460/sh-CTRL, and H460/sh-SRGN cells (b) were incubated in 2% FBS-containing medium for 72 h, followed by Alexa Fluor™ 594 Phalloidin staining of filamentous actin. c HuTu80/Mock and HuTu80/CD44 cells were seeded into the Culture-Insert 2 Well on slide and incubated overnight to form the gap for cell migration. After removing the insert, cells were washed with 1X PBS twice and incubated with CM harvested from H1299/SRGN cells that had been treated with or without ChaseABC for 24 h, followed by Alexa Fluor™ 594 Phalloidin staining of filamentous actin. Filopodia (arrows) and lamellipodia (arrowheads) structures were observed using confocal microscopy. The circumscribed rectangular regions are further enlarged for better viewing. d H1299/Mock, H1299/SRGN, H460/sh-CTRL, and H460/sh-SRGN cells were seeded in FN-coated dishes. In 30 min, attached cells were lysed and subjected for pulling down of Rac1-GTP, Cdc42-GTP and RhoA-GTP. Western blot analysis was performed for total and active Rac1, Cdc42, and RhoA as indicated