Selective Roles of Vertebrate PCF11 in Premature and Full-Length Transcript Termination

Abstract

Summary The pervasive nature of RNA polymerase II (Pol II) transcription requires efficient termination. A key player in this process is the cleavage and polyadenylation (CPA) factor PCF11, which directly binds to the Pol II C-terminal domain and dismantles elongating Pol II from DNA in vitro . We demonstrate that PCF11-mediated termination is essential for vertebrate development. A range of genomic analyses, including mNET-seq, 3′ mRNA-seq, chromatin RNA-seq, and ChIP-seq, reveals that PCF11 enhances transcription termination and stimulates early polyadenylation genome-wide. PCF11 binds preferentially between closely spaced genes, where it prevents transcriptional interference and consequent gene downregulation. Notably, PCF11 is sub-stoichiometric to the CPA complex. Low levels of PCF11 are maintained by an auto-regulatory mechanism involving premature termination of its own transcript and are important for normal development. Both in human cell culture and during zebrafish development, PCF11 selectively attenuates the expression of other transcriptional regulators by premature CPA and termination.

Transcriptional control by premature termination: a forgotten mechanism

“…Therefore, it will be important that future PTT studies also assay the nascent transcriptome. Notably, the genes undergoing PCF11-mediated PTT are enriched for transcriptional regulators, both in humans and in zebrafish [61]. Similarly, in primary human tissues, IPA has also been found to occur preferentially on genes encoding transcriptional regulators [41].…”

“…This is triggered by a 3' processing and termination factor called PCF11 [61]. Like CSTF3, PCF11 employs PTT as an autoregulation mechanism [61,62]. However, we have additionally identified 218 other human genes, which are attenuated by PCF11-mediated PTT.…”

“…Recently, we have established in human cells, as well as during zebrafish embryogenesis, that a subset of protein-coding genes are downregulated by PTT under physiological conditions. This is triggered by a 3' processing and termination factor called PCF11 [61]. Like CSTF3, PCF11 employs PTT as an autoregulation mechanism [61,62].…”

“…We predict that this is likely an underestimate, because many genes undergoing PCF11-mediated PTT might also be dependent on PCF11 for efficient full-length transcript expression. Interestingly, half of PCF11attenuated genes show PTT without detectable IPA products [61]. This suggests that many transcripts attenuated by PTT under physiological conditions are unstable, and that IPA corresponds to only a subset of intragenic PTT.…”

Transcript isoform sequencing reveals widespread promoter-proximal transcriptional termination

“…Notably, the extent to which stalled RNAPII near metazoan TSSs represent transient transcriptional pausing or dynamic RNAPII turn-over, perhaps through transcriptional termination, is now actively debated 25 . The sppRNA biogenesis mechanism likely differs from shorter RNAs (~20-65 nts) detected near initiation regions that may depend on metazoan pausing factors 24,[26][27][28][29] . The conspicuous absence of pausing factor homologs in plants 16 supports transcriptional termination as the outcome of RNAPII stalling near many plant promoters.…”