Figure 2.

Increasing association between p21 and cyclin B1-Cdk1 after exposure to DNA damage-inducing drugs. (A) Cell cycle distribution of asynchronously growing untreated NHFs (Cont) and cultures exposed to ICRF-193 or bleomycin for 12 h. DNA content was determined by FACS analysis of propidium iodide-stained cells. The same cell cultures were used for the biochemical analysis described below. (B) Western blot analysis showing cyclin B1, cyclin A, Cdk1, and p21 levels in p21 and cyclin B1 immunoprecipitates (IP) isolated from untreated NHFs (Con) and NHFs exposed to ICRF-193 (ICRF) or bleomycin (Bleo). For all immunoprecipitation experiments, the equivalent amounts of cell extract (150 μg) were used, and the samples were analyzed on the same immunoblots. Numbers 1, 2, and 3 indicate differentially phosphorylated Cdk1 isoforms (see text for the details). (C) Western blot analysis showing cyclin B1 levels after p21 immunodepletion in bleomycin- and ICRF-193-treated NHF. LC, loading control.