Fig. 5.

Mus81 knockdown partially rescues CDK1^AF-induced defects in MEFs. a MEFs were prepared as described in Materials and methods section. Cdk1^flox/SAF and Cdk1^null/AF cells were harvested in S phase (18 h) or mitosis/mitotic-like state (noco). Noco – nocodazole. Cdk1^flox/SAF MEFs were treated with nocodazole from 21 to 26 h to arrest them at metaphase–anaphase transition, while the drug was added to Cdk1^null/AF MEFs from 16 to 21 h to capture cells in mitotic-like state. Protein extracts from Cdk1^flox/SAF and Cdk1^null/AF MEFs were subjected to immunoprecipitation with antibodies against SLX4 (IP: SLX4) followed by immunoblotting using the indicated antibodies. The endogenous levels of MUS81 and SLX4 were verified by western blotting (input). HSP90 served as a loading control. Results are representative of three independent experiments. b S phase failure and c the levels of DNA damage were determined by FACS as previously described (Figs. 2b and 4a, respectively). Striped red column indicates Cdk1^AF MEFs infected with an empty vector. Quantitative analyses of FACS data are shown. Results are represented as mean ± SD. d Chromosomal fragmentation was analyzed by PFGE as described in Fig. 4c. Results (b–d) are representative of three independent experiments