Figure 1

IR exposure induces c-Abl acetylation. (a) HeLa cells were exposed to 5 Gy of IR or 20 J/m2 of UV and collected at 0, 2 and 4 h later. Anti-Abl immunoprecipitates were prepared using K-12 antibody and resolved on a 5-15% SDSPAGE. The proteins were transferred onto Immobilon-P and immune-analyzed with pan-specific anti-acetyl lysine antibody (top panel). The membrane was stripped as described in methods and reprobed with anti-Abl antibody (bottom panel). (b) Abl^-/- 3T3 cells reconstituted with Flag-tagged Abl were either mock, IR (5 Gy) or UV (20 J/m2) treated and collected 4 h later. Anti-Abl immunoprecipitates were prepared and immune-analyzed with acetyl lysine (top panel) or Flag antibody (bottom panel). (c) Abl^-/- 3T3 cells reconstituted with Flag-tagged Abl were labeled with 14C acetyl CoA for 2 h and then exposed to IR (5 Gy) or UV (20 J/m2) and collected 4 h later. Anti-Abl immunoprecipitates were prepared, resolved on 8% SDS-PAGE, and subjected to fluorography followed by autoradiography (top panel). An aliquot of the immune complex was immune-analyzed with Flag antibody (bottom panel). (d) 293 cells were transfected with either vector alone or the indicated Abl constructs shown in panel e, using lipofectamine. At 36 h post-transfection, cells were exposed to 5 Gy of IR and anti-Flag immunoprecipitates were formed and immune-analyzed with anti-acetyl lysine (top panel) or anti-Flag antibody (bottom panel). (e) Schematic representation of the Abl constructs used in the experimentation described in panel d.