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

Cyclin A-mediated activation of cyclin-dependent kinases (CDKs) is essential for cell cycle transversal. Cyclin A activity is regulated on several levels and cyclin A elevation in a number of cancers suggests a role in tumorigenesis. In the present study, we used a modified DNA binding site selection and PCR amplification procedure to identify DNA binding proteins that are potential substrates of cyclin A-CDK. One of the sequences identified is the Sp1 transcription factor binding site. Co-immunoprecipitation experiments show that cyclin A and Sp1 can interact physically. In vitro and in vivo phosphorylation studies indicate that cyclin A-CDK complexes can phosphorylate Sp1. The phosphorylation site is located in the N-terminal region of the protein. Cells overexpressing cyclin A have elevated levels of Sp1 DNA binding activity, suggesting that cyclin A-CDK-mediated phosphorylation augments Sp1 DNA binding properties. In co-transfection studies, cyclin A expression stimulated transcription from an Sp1-regulated promoter. Mutation of the phosphorylation site abrogated cyclin A-CDK-dependent phosphorylation, augmentation of Sp1 transactivation function and DNA binding activity.