Transcription elongation is finely tuned by dozens of regulatory factors

Abstract

Understanding the complex network that regulates transcription elongation requires the quantitative analysis of RNA polymerase II (Pol II) activity in a wide variety of regulatory environments. We performed native elongating transcript sequencing (NET-seq) in 41 strains of S. cerevisiae lacking known elongation regulators, including RNA processing factors, transcription elongation factors, chromatin modifiers, and remodelers. We found that the opposing effects of these factors balance transcription elongation and antisense transcription. Different sets of factors tightly regulate Pol II progression across gene bodies so that Pol II density peaks at key points of RNA processing. These regulators control where Pol II pauses with each obscuring large numbers of potential pause sites that are primarily determined by DNA sequence and shape. Antisense transcription varies highly across the regulatory landscapes analyzed, but antisense transcription in itself does not affect sense transcription at the same locus. Our findings collectively show that a diverse array of factors regulate transcription elongation by precisely balancing Pol II activity.

Multiple direct and indirect roles of Paf1C in elongation, splicing, and histone post-translational modifications

“…29 Steady-state RNA-seq data and RNAPII profiling through NET-seq showed that paf1Δ cells exhibit widespread transcriptional misregulation of coding genes and upregulation of antisense transcripts, particularly for Set2 Repressible Antisense Transcripts (SRATs). [30][31][32] This is consistent with the loss of Set2-dependent H3 K36me3 in paf1∆, and also ctr9∆, mutants. 11,[30][31][32] While it is apparent that Paf1C broadly impacts gene expression and chromatin states, technological limitations have confounded interpretations of its functions.…”

“…[30][31][32] This is consistent with the loss of Set2-dependent H3 K36me3 in paf1∆, and also ctr9∆, mutants. 11,[30][31][32] While it is apparent that Paf1C broadly impacts gene expression and chromatin states, technological limitations have confounded interpretations of its functions. Many previous studies have relied on stable null alleles, which allow cumulative effects to accrue.…”

“…30–32 This is consistent with the loss of Set2-dependent H3 K36me3 in paf1Δ , and also ctr9Δ , mutants. 11,30–32…”

“…29 Steady-state RNA-seq data and RNAPII profiling through NET-seq showed that paf1Δ cells exhibit widespread transcriptional misregulation of coding genes and upregulation of antisense transcripts, particularly for Set2 Repressible Antisense Transcripts (SRATs). 30–32 This is consistent with the loss of Set2-dependent H3 K36me3 in paf1Δ , and also ctr9Δ , mutants. 11,30–32…”

DNA-sequence and epigenomic determinants of local rates of transcription elongation

“…Finally, in at least some cases, it is clear that elongation rates are not only indirectly influenced by structural features of DNA, RNA, or chromatin, but are actively regulated in response to various cellular stimuli, with dysregulation of elongation rates potentially contributing to disease progression [14].Other studies have focused specifically on pausing of Pol II. Aside from the pronounced pausing that occurs proximal to the promoter, many, typically more subtle, pause sites occur within gene bodies, and, in the aggregate, these sites have a major effect on the dynamics of transcription elongation [16][17][18][19] (see also [20] for a recent study in yeast). These gene-body pause sites replicate well across experiments but vary substantially in their density across genes; they also occur in divergent antisense transcripts and enhancer RNAs as well as in gene bodies [18].…”