The Nrd1-like protein Seb1 coordinates cotranscriptional 3′ end processing and polyadenylation site selection

Abstract

Termination of RNA polymerase II (RNAPII) transcription is associated with RNA 3 ′ end formation. For coding genes, termination is initiated by the cleavage/polyadenylation machinery. In contrast, a majority of noncoding transcription events in Saccharomyces cerevisiae does not rely on RNA cleavage for termination but instead terminates via a pathway that requires the Nrd1-Nab3-Sen1 (NNS) complex. Here we show that the Schizosaccharomyces pombe ortholog of Nrd1, Seb1, does not function in NNS-like termination but promotes polyadenylation site selection of coding and noncoding genes. We found that Seb1 associates with 3 ′ end processing factors, is enriched at the 3 ′ end of genes, and binds RNA motifs downstream from cleavage sites. Importantly, a deficiency in Seb1 resulted in widespread changes in 3 ′ untranslated region (UTR) length as a consequence of increased alternative polyadenylation. Given that Seb1 levels affected the recruitment of conserved 3 ′ end processing factors, our findings indicate that the conserved RNA-binding protein Seb1 cotranscriptionally controls alternative polyadenylation.

The conserved protein Seb1 drives transcription termination by binding RNA polymerase II and nascent RNA

“…1b), resembling the Nrd1 binding motif. Consistent with our data, a similar Seb1 motif (A(U)GUA) has been identified using CRAC (crosslinking and cDNA analysis)35. Interestingly, Seb1 binding is preferentially observed downstream of PASs where the UGUA motif is also specifically enriched (annotations used from ref.…”

“…1a). In addition to expected interactors such as subunits of Pol II, several CPF components co-purify with Seb1 suggesting, in agreement with a recent publication35, that it is involved in mRNA 3′ end formation.…”

“…8f). These data suggest that Seb1 is likely needed for efficient cleavage of the pre-mRNA rather than influencing PAS choice as previously proposed35. Because 3′ extended transcripts are often unstable, a decrease in cleavage efficiency is therefore expected to result in less RNA overall.…”

“…34), the essential CID-RRM protein Seb1 from fission yeast shows preference for S2P-Pol II both in vitro and in vivo . In agreement with a recent study35, we show that Seb1 co-purifies with components of the CPF machinery and is needed for efficient pre-mRNA cleavage. Using PAR-CLIP (Photoactivatable Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation) and ChIP-Seq (chromatin immunoprecipitation-sequencing), we demonstrate that Seb1 is recruited to the 3′ ends of genes.…”

“…8e). Accumulation of 3′ extended transcripts upon loss of Seb1 was also reported by a recent study35. We performed a global analysis of the cleavage efficiency in Seb1 mutants using published PAS annotations42.…”