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. 2021 Oct 8;7(41):eabg6262. doi: 10.1126/sciadv.abg6262

Fig. 6. Stabilized β-catenin promotes Akt activation.

Fig. 6.

(A) Immunoblot analysis of phospho-Akt, pan-Akt, Foxo1, β-catenin, and β-actin in WT and stabilized β-catenin (Ctnnb1 S33Y)–transduced CD4 SP thymocytes stimulated with SDF1 for indicated amounts of time. (B) Representative immunofluorescence images of 293T cells transfected with Flag-Akt and Myc-Ctnnb1 S33Y and stained for Akt (red), β-catenin (green), and 4′,6-diamidino-2-phenylindole (DAPI; blue, nuclear staining). Scale bars, 10 μm. (C and D) 293T cells were transfected with MycHis-Ctnnb1 S33Y and/or Flag-Akt. Total cell lysates were immunoprecipitated with Flag antibody and analyzed by immunoblotting with indicated antibodies. (E) Mouse embryonic fibroblasts (MEFs) were transduced with lentiviruses encoding β-catenin S33Y (Ctnnb1 S33Y) or vector, serum-starved for 16 hours, and restimulated with 0.5% FBS for indicated amounts of time. Cytoplasmic and nuclear fractions were analyzed by immunoblotting with indicated antibodies. (F) Immunoblot analysis of cytoplasmic and nuclear fractions of CD4 SP thymocytes from WT and DKO mice. (G) MEFs were transduced with lentiviruses encoding β-catenin S33Y (Ctnnb1 S33Y) or myristoylation-tagged β-catenin S33Y (Myr-Ctnnb1 S33Y), serum-starved for 16 hours, restimulated with 0.5% FBS for 5 min, and analyzed by immunofluorescence staining for β-catenin S33Y (green) and DAPI (blue, nuclear staining). Scale bar, 10 μm. (H) Immunoblot analysis of cytoplasmic and nuclear fractions of MEFs from (G). Data are representative of at least two (D) and three (A to C and E to H) independent experiments.