Abstract
RNA Polymerase II (RNAPII) transcription termination is regulated by the phosphorylation status of the C-terminal domain (CTD). The phosphatase Rtr1 has been shown to regulate serine 5 phosphorylation on the CTD; however, its role in the regulation of RNAPII termination has not been explored. As a consequence of RTR1 deletion, interactions within the termination machinery and between the termination machinery and RNAPII were altered as quantified by Disruption-Compensation (DisCo) network analysis. Of note, interactions between RNAPII and the cleavage factor IA (CF1A) subunit Pcf11 were reduced in rtr1Δ, whereas interactions with the CTD and RNA-binding termination factor Nrd1 were increased. Globally, rtr1Δ leads to decreases in numerous noncoding RNAs that are linked to the Nrd1, Nab3 and Sen1 (NNS) -dependent RNAPII termination pathway. Genome-wide analysis of RNAPII and Nrd1 occupancy suggests that loss of RTR1 leads to increased termination at noncoding genes. Additionally, premature RNAPII termination increases globally at protein-coding genes with a decrease in RNAPII occupancy occurring just after the peak of Nrd1 recruitment during early elongation. The effects of rtr1Δ on RNA expression levels were lost following deletion of the exosome subunit Rrp6, which works with the NNS complex to rapidly degrade a number of noncoding RNAs following termination. Overall, these data suggest that Rtr1 restricts the NNS-dependent termination pathway in WT cells to prevent premature termination of mRNAs and ncRNAs. Rtr1 facilitates low-level elongation of noncoding transcripts that impact RNAPII interference thereby shaping the transcriptome.
Highlights
The termination of transcription by eukaryotic RNA Polymerase II (RNAPII) is tightly coupled with RNA processing, including small RNA processing, splicing, and mRNA cleavage and polyadenylation at the 3’-end of protein-coding genes [1, 2]
The phosphorylation status of the RNAPII C-terminal domain (CTD) plays a major role in the regulation of the mechanisms through which transcription termination occurs [62,63,64,65]
We have recently shown that deletion of RTR1 causes global increases in CTD serine 5 (Ser5)-P [26] and it has previously been shown that loss of RTR1 results in 3’-end processing defects at the polyA-dependent gene NRD1 [13]
Summary
The termination of transcription by eukaryotic RNA Polymerase II (RNAPII) is tightly coupled with RNA processing, including small RNA processing, splicing, and mRNA cleavage and polyadenylation at the 3’-end of protein-coding genes [1, 2]. Transcription termination in yeast has been shown to be regulated through numerous termination factors as well as the phosphorylation status of the C-terminal domain (CTD) of RNAPII, which has the repetitive sequence (Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7)n [11, 12]. The exact mechanisms that underlie the role of CTD dephosphorylation in the regulation of elongation, termination, and the attenuation of these processes remain unclear. Ssu dephosphorylation of Ser serves as a prerequisite for Ser dephosphorylation by Fcp1 [14, 16] It remains unclear how temporal dephosphorylation impacts the formation and/or recruitment of RNA processing complexes during transcription and the determination of the termination pathway that will be used
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