Enrichment of K921 Acetylated Abl in chromatin fractions of γ-irradiated cells. (a) Reconstituted Abl+ 3T3 cells were exposed to IR (5 Gy) and at 18 h post-treatment, cells were harvested, lysed in hypotonic buffer and fractionated into nuclear and cytoplasmic fraction as described by Méndez et al . Nuclear fraction was again separated into chromatin bound and unbound and subjected to immunoprecipitation with anti-Abl (K-12) antibody. The immune-complex was analyzed by subjecting to western blotting with K921 Ac-Abl or Abl antibody (top panels). Input lysates were also blotted with tubulin (cytoplasmic marker) or ORC2 (chromatin marker) antibody (bottom panel). (b) Abl-/- 3T3 cells reconstituted with K921R mutant of Abl were exposed to 5 Gy of IR and nuclear and cytoplasmic fractions were formed and immune-analyzed for K921 Ac-Abl, Abl, tubulin or ORC2 proteins as described in panel a.