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Fig. 5. Interaction between Cdk5, Bem1 and Cdc24. (A) For immunoprecipitation analysis, whole cell extracts (WCEs) were prepared from cdk5-3MYC bem1-VSV cdc24-3HA (UMI83) and cdk5^ts-3MYC bem1-VSV cdc24-3HA (UMI82) cells incubated for 4 hours at 34°C. HA, MYC and VSV immunoprecipitates were isolated and blotted with antibodies to recognize the HA, MYC and VSV epitopes. Bem1 associated with Cdk5 independently of Cdk5 function. However, the formation of the Bem1-Cdc24 complex was dependent on Cdk5 function. (B) Bimolecular fluorescence complementation (BiFC) assay. Tip-associated YFP signal is clearly seen in NYFP-bem1 cdc24-CYFP (UMI127) cells but no YFP signal is detected in NYFP-bem1 cdc24-CYFP cdk5^ts (UMI138) cells grown for 4 hours at 34°C. Scale bars: 15 µm.

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