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. 2018 Oct 15;7:e40474. doi: 10.7554/eLife.40474

Figure 6. Ras-driven invasion requires the two RalGEF RGL1 and RGL2.

Figure 6.

(A) HEK-HTRasV12 cells require RGL1 and RGL2 for Transwell invasion (upper panel). Depletions of the six RalGEF were validated by RT-qPCR (lower panel). Graphs show the mean ±SEM. Each dot represents one experiment. For statistics one-way ANOVA was used. *p<0.05, **p<0.01, ***p<0.001, ns not-significant. (B–C) Localization of endogenous RGL2 and RalB is increased at the cell edges of HEK-HTRasV12 transformed cells as compared to HEK-HT. Representative epifluorescence images of the immunostaining of RGL2 and RalB are depicted. Scale bar, 10 µm. Graphs show the averaged percentage ±SEM of cell perimeter which is positive for RGL2 or RalB. n > 40 cells from four independent experiments. For statistics Student t-test was used. *p<0.05, **p<0.01, ***, p<0.001, ns indicates not-significant. (D) The Ras-RGL1/2-RalB-exocyst-WRC axis drives invasion. Active Ras-GTP binds and recruit the two RalGEFs RGL1 and RGL2, which activate RalB; RalB-GTP binds to the exocyst complex, promoting its assembly and recruitment to the leading edge; by its direct association with the WRC complex, exocyst drives WRC to the leading edge, where WRC stimulates actin polymerization via the Arp2/3 complex, consequently promoting protrusion formation, motility and invasion.