FIG. 5.

p125 is activated by wild-type p48, but not by p48 from XPE⁻ cells. Wild-type p48 cDNAs (p48 and FLAG-p48) and p48 cDNAs encoding amino acid changes found in XPE⁻ cells (2415 and 82TO) were cloned into an expression vector and transfected into V79 (hamster) and 293T (human) cells. The expression vector in lanes vec contained no cDNA. After transfection, cell extracts (2 μg) were assayed for UV-specific binding activity with DNA probe damaged with UV at 5,000 J/m² (two upper panels). A protein-DNA complex in 293T cells migrates to slightly below B1 but is present whether or not the probe DNA is damaged with UV. Transfection efficiency was measured by analyzing the activity from a cotransfected luciferase reporter gene (10²/μg). Levels of p48 protein were measured by immunoblotting with anti-FLAG antibody (2 μg/ml) after SDS-PAGE of V79 (50 μg) and 293T (10 μg) extracts. F, free DNA probe migration; B1 and B2, UV damage-specific protein-DNA complex mobilities.