GSE99955
Knockdown of CFIm25 or CFIm68 decreases RNA polymerase II pausing and transcription
Transcription of eukaryotic protein-coding genes by RNA polymerase II (pol II) is highly regulated at initiation, elongation and termination and these steps are coordinated with co-transcriptional processing of the emerging pre-mRNA by capping, splicing, and cleavage and polyadenylation. Polyadenylation (Poly(A)) site recognition, which defines the end of the mRNA, relies on the cleavage and polyadenylation (CPA) complex. It was previously observed that knocking-down proteins of the CPA complex affects not only recognition of the poly(A) site but also results in increased pausing of pol II at the beginning of genes. This finding suggests that the CPA complex plays a role in regulating pol II turnover after transcription initiation. To explore this possibility, we knocked-down two subunits of the cleavage factor I (CFIm), CFIm25 and CFIm68, which are part of the CPA complex and involved in alternative polyadenylation, and performed pol II ChIP-seq. Surprisingly, we found that contrary to the effect of knocking down of other CPA factors, reduction of the level of CFIm25 or CFIm68 decreases pol II pausing and transcription genome-wide in HEK293 cells. These findings indicate that different subunits of the CPA complex have opposite effects on pol II pausing at the start of genes. Overall design: Examination of the effect of CFIm68 knockdown on RNA polymerase II in HEK293 cells after 30 minutes treatment with DMSO or DRB.