SIRT7-dependent deacetylation of CDK9 activates RNA polymerase II transcription

Blank, Maximilian F.; Chen, Sifan; Poetz, Fabian; Schnölzer, Martina; Voit, Renate; Grummt, Ingrid

Previous results have established that SIRT7 activates transcription of rRNA genes by enhancing Pol I occupancy at rDNA (1, 12) . We therefore reasoned that overexpression of SIRT7 may increase binding of Pol II to genes that are regulated by SIRT7. Indeed, Pol II occupancy at selected Pol II-dependent genes was enhanced after overexpression of wildtype SIRT7 but not mutant SIRT7/H187Y ( Figure 4D ). Conversely, knockdown of SIRT7 decreased Pol II occupancy at SIRT7-responsive genes to a similar level as Pol I at the rDNA locus, while binding of Pol II to actin and U1 snRNA genes, which are not occupied by SIRT7, was not altered ( Figure 4E and Supplementary Figures S4A and S4D ). ChIP assays with antibodies against Pol II-pSer5 and Pol II-pSer2 revealed that knockdown of SIRT7 affected the occupancy of both initiating and elongating Pol II, supporting that hyperacetylation of CDK9 prevents CTD phosphorylation and impairs transcription elongation ( Figure 4H and Supplementary Figure S4F ). This view is substantiated by ChIP assays showing increased occupancy of both SIRT7 and CDK9 at the promoterproximal region of target genes ( Figure 4F) . Significantly, the association of CDK9 with these genes was abolished upon knockdown of SIRT7, confirming that downregulation of SIRT7 correlates with abrogation of CDK9 binding ( Figure 4G and Supplementary Figure S4F ). To further strengthen the functional link between SIRT7 and CDK9 activity, we determined Pol II occupancy at target genes in SIRT7-depleted cells that overexpress CDK9/K48R or CDK9/K48Q. While decreased Pol II occupancy in SIRT7 knockdown cells could be relieved by overexpression of Nucleic Acids Research, 2017 , Vol. 45, No. 5 2683 Figure S4G) . Taken together, these results uncover the mechanism underlying SIRT7-dependent activation of Pol II, demonstrating that SIRT7 enhances transcription elongation by deacetylation of lysine 48 of CDK9, which is required for CTD phosphorylation and transcription activation.