Cyclin-dependent kinase 12 influences protein kinase D1 kinase activity

A unique and aggressive molecular subtype of prostate cancer is driven by recurrent mutations in the cyclin-dependent kinase 12 (CDK12) gene, which occur exclusive of other common genetic alterations. Protein Kinase D1 (PrKD1) is a well-established tumor suppressor in prostate cancer. Phosphoproteomics studies have identified serines 681 and 685 as putative PrKD1 phosphorylation sites in CDK12-mutated tumors; however, whether these proteins interact directly or the potential impact on either protein's function remains unknown. In this study, we demonstrate a direct interaction between PrKD1 and CDK12 in a prostate cancer cell line using co-immunoprecipitation and a bimolecular fluorescence complementation (BiFC) assay. Site-directed mutagenesis of serines 681 and 685, the putative PrKD1 phosphorylation sites in CDK12, did not alter the phosphorylation of the well-established CDK12 substrates, serines 2 and 5 on RNA polymerase II. Interestingly, these site-directed mutagenesis experiments resulted in altered PrKD1 kinase activity. Molecular modeling studies suggest that phosphorylation at serine 681, or both serines 681 and 685, releases PrKD1 from an autoinhibitory conformation, promoting its kinase activity. These findings suggest a potential regulatory role of CDK12 in modulating PrKD1 kinase function in prostate cancer.