FIG 8.

Phosphorylation at Ser131 by ATR is critical for RASSF1A-mediated XPA deacetylation. (A) HEK-293 cells were transiently cotransfected with expression constructs for HA-tagged XPA and GFP-tagged RASSF1A or the S(131)A nonphosphorylatable mutant of RASSF1A and left untreated (−) or exposed to 40 J/m² UV (+). One hour after UV exposure, the cells were lysed, and equal amounts of protein were immunoprecipitated with anti-acetyl-Lys beads. The immunoprecipitates were fractionated on an SDS-polyacrylamide gel and immunoblotted with anti-HA and anti-GFP antibodies. (B) Similar experiments were performed with wild-type RASSF1A except the cells were treated with 1 mM caffeine or vehicle 16 h prior to exposure to UV. One hour after UV exposure, the cells were processed as described above.