MiTF phosphorylation on serine 73 by Erk1/2 kinase was required for its proteasome-mediated degradation after UVR. A, c83-2C cells were exposed to UVC in the presence of MG132 and collected for western blot analysis at the indicated time points. B, transient expression of MiTF-WT, MiTF-S73A in A375 melanoma cell line. C83-2C cells served as a positive control for MiTF western blot. GFP is the control cells transfected with GFP in the same QCXIP vector that carried MiTF-WT or MiTF-S73A coding sequence. C, Cells expressing MiTF-S73A were exposed to UVC and collected for western blot analysis. D, Cells expressing MiTF-WT were exposed to UVC and collected for western blot analysis. E, MiTF was poly-ubiquitinated after UVC radiation. NHMs were exposed to UVC (3 mJ/cm2) and collected for immunoprecipitation by anti-MiTF antibodies (un-irradiated cells and anti-GFP antibodies were used as controls), then probed with anti-ubiquitin antibodies (top panel). The membrane was stripped and blotted with anti-MiTF antibodies for a loading control (bottom). The IgG label indicates antibody heavy chain of IgG proteins.