FIG. 2.

Substrate palmitoylation is not required for MIR2-mediated downregulation of MHC-I. (A) HeLa cells stably transduced with HLA-A2 or HLA-B7 were transfected with MIR2-EGFP or EGFP alone, treated with 100 μM 2-Br or vehicle alone, stained with mouse-raised primary antibody against HLA-A2 or HLA-B7, stained with PE-conjugated secondary antibody against mouse IgG, and analyzed by flow cytometry. The level of MHC-I expression in MIR2-expressing cells was calculated by determining the ratio of MHC-I expression (mean PE value in GFP⁺ cells) in MIR2-EGFP-expressing cells to that in EGFP alone. Bar graphs represent the results of three consecutive experiments. (B) HeLa cells stably transduced with wild-type HLA-B7, HLA-B7-C309S, or HLA-B7-C309S C326W (unpalmitoylated) were transfected with MIR2-EGFP or EGFP alone. Samples were stained with a mouse-raised primary antibody against HLA-B7, stained with PE-conjugated secondary antibody against mouse IgG, and analyzed by flow cytometry.