A Cdk9-PP1 switch regulates the elongation-termination transition of RNA polymerase II

摘要

The end of the RNA polymerase II (Pol II) transcription cycle is strictly regulated to prevent interference between neighbouring genes and safeguard transcriptome integrity. Pol II accumulation downstream of the cleavage and polyadenylation signal (CPS) can facilitate recruitment of factors involved in mRNA 3’-end formation and termination, but how this sequence is initiated remains unclear. In a chemical-genetic screen, we identified human protein phosphatase 1 (PP1) isoforms as substrates of positive transcription elongation factor b (P-TEFb), the cyclin-dependent kinase 9 (Cdk9)-cyclin T1 complex. Here we show that Cdk9 and PP1 govern phosphorylation of the conserved elongation factor Spt5 in the fission yeast Schizosaccharomyces pombe. Cdk9 phosphorylates both Spt5 and a negative regulatory site on the PP1 isoform Dis2. Sites targeted by Cdk9 in the Spt5 carboxy-terminal domain (CTD) can be dephosphorylated by Dis2 in vitro, and dis2 mutations retard Spt5 dephosphorylation after Cdk9 inhibition in vivo. Chromatin immunoprecipitation and sequencing (ChIP-seq) analysis indicates that Spt5 is dephosphorylated as transcription complexes traverse the CPS, concomitant with accumulation of Pol II phosphorylated at CTD repeat residue Ser2. A conditionally lethal Dis2-inactivating mutation attenuates the drop in Spt5 phosphorylation (pSpt5) on chromatin, promotes transcription beyond the normal termination zone detected by precision run-on transcription and sequencing (PRO-seq), and is genetically suppressed by ablation of Cdk9 target sites in Spt5. These results suggest that the transition from elongation to termination by Pol II coincides with Dis2-dependent reversal of Cdk9 signaling—a switch analogous to a Cdk1-PP1 circuit that controls mitotic progression.