Cyclin E but not cyclin A selectively interacts with phosphorylated p27^kip1 isoforms in vivo. 2D gel analysis shows that cyclin E but not cyclin A or cdk2 selectively binds phosphorylated p27^kip1 in vivo (A). The isoforms of p27^kip1 coimmunoprecipitated with cyclin A, cyclin E, and cdk2 are indicated with arrows. Rabbit polyclonal antibodies specific to cyclin A, cyclin E, cdk2, and p27^kip1 were used to carry out immunoprecipitation. Coimmunoprecipitated p27^kip1 isoforms were detected with a mouse monoclonal anti-p27^kip1 antibody SX53G8. Similar results were also obtained with another anti–p27^kip1 mouse monoclonal antibody with a different epitope from SX53G8 (data not shown). The patterns of p27^kip1 isoforms associated with cyclin E in Cal51 cells treated with 30 μM roscovitine or DMSO for 24 hours are shown (B). The ability of 10 or 30 μM roscovitine to inhibit the cdk2 kinase activity obtained from the anti-cdk2 immunoprecipitate of Cal51 cells in vitro is shown (C). Panel D shows the patterns of p27^kip1 isoforms detected from the anti–cyclin E immunoprecipitates of Cal51 cell lysates incubated with DMSO or 60 μM roscovitine throughout the immunoprecipitation process (indicated as DMSO or roscovitine, respectively).