Figure 8. Serines 332 /335 in the HLA-C cytoplasmic tail are phosphorylated.

(A) CEM T cells or (B) U937 cells expressing the indicated MHC-I were treated with 100nM bafilomycin or DMSO overnight and metabolically labeled with ³²P orthophosphate as described in Materials and Methods. MHC-I molecules were immunoprecipitated from lysates with an antibody directed against HLA-A2 (BB7.2) and separated by SDS PAGE. In the lower panel, western blots of lysates from cells grown in parallel are shown. (C) Treatment of U937 cells with LPS and PMA to induce differentiation into macrophage-like cells results in a reduction of phosphorylation and a stabilization of A2/C protein. U937 cells were treated with DMSO or PMA and ionomycin as described in Figure 7. Cells were labeled with ³²P orthophosphate and immunoprecipitated as described above. In the lower panel, western blots of lysates from cells grown in parallel are shown. “Control” indicates results obtained with the parental cell line that lacked expression of HLA-A2.