Figure 5. Phosphorylation of cJun by cellular CDK4.

A) Purified recombinant CDK4:cyclinD complex was reacted with cJun and GSK GST fusion proteins to determine specificity of phosphorylation by CDK4. Western blotting with phospho-specific Abs was used to monitor the phosphorylation of the GST-fusion proteins. These data are representative of 3 separate experiments. B) CDK4 was immunoprecipitated from differentially stimulated B lymphocytes and reacted with GST-cJun fusion protein as described in Methods. The presence of cyclin D2 and phosphorylation of c-Jun was analyzed by Western Blot using anti-Ser 73 phospho-cJun Abs. Blotting against CDK4 was used as a loading control. Quantitative analysis of chemiluminescent band development enabled comparison of relative protein amounts between treatments (bar graphs). These data are representative of 2 separate experiments.