Fig. 2: p38α phosphorylates GLI1 on Ser937/Ser941 to destabilize GLI1.
a Co-immunoprecipitation and western blot analyses in 293T cells transfected with HA-GLI1 construct and treated with losmapimod at 10 μM for the indicated times. Western blot analyses were performed using anti-Ser/Thr and GAPDH antibodies. b Gli-luciferase reporter assays in 293T cells transiently co-transfected with Gli-luciferase/Gli(mutant)-luciferase and Renilla-luciferase reporter constructs and GLI1 variants (n = 3 independent experiments). c The conserved Ser937/Ser941 sites of GLI1 among species. d–f Western blot analyses in 293T cells transfected with HA-GLI1/HA-mGli1 or their S937A/S939A/S941A variants in combination with or without MAPK14 and treated with or without losmapimpod at 10 μM for 6 h. g–j Western blot analyses of phospho-S937-GLI1 (p-GLI1) and GLI1 in 293T cells transfected with the increasing amounts of MAPK14 or treated with losmapimpod at 10 μM, N-SHH at 100 ng/ml or SAG at 2.5 μM for the indicated times. k–n Western blot analyses in C3H10T1/2 cells transfected with or without MAPK14 or HA-GLI1 variants and treated with cycloheximide at 2 μg/ml for the indicated times or MG132 at 10 μM for 12 h. o C3H10T1/2 cells expressing Myc-GLI1, HA-ubiquitin, and Flag-MAPK14 were immunoprecipitated with anti-Myc for western blot analyses. p, q Western blot analyses of cytosolic versus nuclear fractions of the indicated proteins in C3H10T1/2 cells transfected with MAPK14 and treated with SAG at 2.5 μM for 6 h or transfected with HA-GLI1 variant alone. r Immunohistochemistry staining for phospho-S937-GLI1 in paraffin-embedded children MBSHH sections. Bar, 20 μm. Data were presented as mean ± sd, two-tailed Student’s t test for (b). A representative example of three replicates is shown for (a) and (d–r). Source data are provided as a Source Data file.
