Cyclin A-CDK phosphorylates Sp1 and enhances Sp1-mediated transcription

Cyclin A-mediated activation of cyclin-dependent kinases (CDKs) is essentia l for cell cycle transversal. Cyclin A activity is regulated on several lev els and cyclin A elevation in a number of cancers suggests a role in tumori genesis. In the present study, we used a modified DNA binding site selectio n and PCR amplification procedure to identify DNA binding proteins that are potential substrates of cyclin A-CDK. One of the sequences identified is t he Sp1 transcription factor binding site. Co-immunoprecipitation experiment s show that cychn A and Sp1 can interact physically. In vitro and in vivo p hosphorylation studies indicate that cyclin A-CDK complexes can phosphoryla te Sp1. The phosphorylation site is located in the N-terminal region of the protein. Cells overexpressing cyclin A have elevated levels of Sp1 DNA bin ding activity, suggesting that cyclin A-CDK-mediated phosphorylation augmen ts Sp1 DNA binding properties. In co-transfection studies, cyclin A express ion stimulated transcription from an Sp1-regulated promoter. Mutation of th e phosphorylation site abrogated cyclin A-CDK-dependent phosphorylation, au gmentation of Sp1 transactivation function and DNA binding activity.