FIG. 1.

Wee1 associates with cyclin A/Cdk2 complexes. (A) Wee1 and Cdk2 complexes were immunoprecipitated from lysates of asynchronous U2-OS cells and subjected to immunoblotting (IB) for the opposite protein. Cdk2 was readily detected in Wee1 IPs, and Wee1 was readily detected in Cdk2 IPs. IPs without extract or with nonspecific immunoglobulin G (Ig) served as negative controls. (B) Same experiment as in panel A except that cells were enriched for S and G₂ phases, following treatment with HU and release. (C) Cdk2 RNAi confirms association of Wee1 with Cdk2. U2-OS cells were treated with siRNAs directed against Cdk2 or an off-target sequence (control). IB confirmed knockdown of Cdk2 but not Cdk1, thus also confirming the specificity of the respective antibodies (left). Cdk2 RNAi reduced Cdk2 detected in Wee1 IPs (right). Relative band intensity was quantitated, and values are listed below each lane. The numbers below the panels present quantitation of the main bands observed relative to the control (con).