Fig 7.

Role of Rb and p53 pathways in senescence induction by Wnt inhibition. (A) Rb dephosphorylation upon SFRP1 expression. IMR-90 cells were infected with vector or SFRP1 lentivirus, and 5 days after infection, Rb, p53, p21, p16, and tubulin expression was analyzed by immunoblotting. SFRP1 induced Rb dephosphorylation but did not affect p53, p21, or p16 expression. For comparison, Rb, p16, and tubulin expression upon Ras-induced senescence was also analyzed by immunoblotting. (B) Knockdown of Rb and p53 in IMR-90 cells. IMR-90 cells were infected with the indicated lentiviruses. Five days after infection, Rb, p53, nucleolin, and tubulin expression was analyzed by immunoblotting. sh, shRNA. (C) Knockdown of Rb or p53 abolishes SFRP1-induced senescence. IMR-90 cells were coinfected with the indicated lentiviruses. Five days postinfection, the cells were stained for SA-β-Gal. *, P < 0.05 compared to SFRP1 plus shLuc. (D) p53 knockdown attenuates Rb dephosphorylation by SFRP1. IMR-90 cells were coinfected with the indicated lentiviruses, and 5 days after infection, Rb expression was analyzed by immunoblotting. (E) β-Catenin knockdown results in Rb dephosphorylation. IMR-90 cells were infected with lentiviruses expressing scrambled shRNA (sh scr), shRNA against β-catenin-1 (sh β-cat 1), or shRNA against β-catenin-2 (sh β-cat#2), and 5 days after infection, Rb, p53, and p21 expression was analyzed by immunoblotting. Nucleolin served as a loading control. (F) Knockdown of Rb or p53 abolishes β-catenin RNAi-induced senescence. IMR-90 cells were coinfected with the indicated lentiviruses and 5 days after infection stained for SA-β-Gal. *, P < 0.05 compared to sh β-catenin 2 plus shLuc.