Figure 7.
Blocking Rac1 function largely abolishes mutant p53 GOF in promoting migration and metastasis of tumor cells. (A–C) Blocking Rac1 activity greatly abolished mutant p53 (R175H and R273H) GOF in promoting migration of H1299 cells as measured by transwell assays. (A) H1299 mutant p53 (R175H and R273H) and H1299-Con cells were transfected with shRNA vectors against Rac1. (B) Cells were transfected with vectors expressing Rac1 DN. (C) Cells were treated with 10 µM NSC23766 or PBS. (D) Expression of Rac1-R but not Rac1ΔSUMO-R largely restored mutant p53 GOF in promoting migration in H1299 cells. (E–G) Blocking Rac1 by shRNA vectors against Rac1 (E), expression of Rac1 DN (F), or treatment with 10 µM NSC23766 (G) greatly abolished mutant p53 GOF in promoting migration of SK-BR-3 and MDA-MB468 cells as measured by transwell assays. (H) Expression of Rac1-R but not Rac1ΔSUMO-R largely restored mutant p53 GOF in promoting migration in SK-BR-3 cells with knockdown of endogenous Rac1. The left panels in A and E show representative images. Bar, 200 µm. (Right panel) Quantification of the average number of migrated cells per field. In A–H, data are presented as mean ± SD. n = 6. (#) P < 0.05; (*) P < 0.01; (**) P < 0.001, Student's t-test. (I,J) Blocking Rac1 activity by shRNA vectors against Rac1 (I) or expression of Rac1 DN (J) largely abolished mutant p53 GOF in promoting lung metastasis of H1299 cells in mice. H1299-Con and H1299 mutant p53 (R175H and R273) cells with or without Rac1 knockdown (I) or expression of Rac1 DN (J) were injected into nude mice via the tail vein. The number of lung tumors was determined after 6 wk. n = 8 mice per group. (Left panel) Representative H&E images of lung sections. Bar, 100 µm. (K) A schematic model depicting that mutant p53 inhibits SENP1-meidated Rac1 de-SUMOylation to activate Rac1 as an important mechanism of mutant p53 GOF in tumorigenesis.