The CDK12-INTAC Axis Modulates LEO1 Phosphorylation for Processive Transcription Elongation

Cyclin-dependent kinase 12 (CDK12) interacts with Cyclin K to form a a functional nuclear kinase complex, which has been reported to phosphorylate the carboxyl-terminal domain (CTD) of RNA polymerase II (Pol II) for transcriptional regulation and co-transcriptional RNA processing. However, the precise mechanisms and targets of CDK12 action remain largely unknown. Here, we combined a chemical genetic screen and phosphoproteomic strategies and identified a landscape of nuclear CDK12 substrates, which included proteins that regulate transcription, chromatin organization, and RNA splicing. Next, we confirmed that the LEO1 subunit of the transcription elongation factor PAF1 complex (PAF1C) is a bona fide substrate of CDK12. Acute depletion of LEO1 reduces Pol II occupancy on the chromatin, while mutations of LEO1 phosphorylation sites to non-phosphorylatable alanine residues attenuated the association of PAF1C with elongating Pol II and chromatin, resulting in impaired processive transcription elongation. Furthermore, LEO1 C-terminus could interact with and be dephosphorylated by the Integrator-PP2A complex (INTAC), while acute depletion of INTAC in cells promotes the association between PAF1C and elongating Pol II on the chromatin. Together, this study provides a novel transcriptional regulatory mechanism that the CDK12-INTAC axis fine-tunes LEO1 phosphorylation for processive transcription elongation. Overall design: Determination of PAF1C and Pol II occupancy by ChIP-Rx Determination of nascent RNA synthesis by TT-seq Determination of elongating Pol II by PRO-seq