Abstract
RNA polymerase II synthesizes a diverse set of transcripts including both protein-coding and non-coding RNAs. One major difference between these two classes of transcripts is the mechanism of termination. Messenger RNA transcripts terminate downstream of the coding region in a process that is coupled to cleavage and polyadenylation reactions. Non-coding transcripts like Saccharomyces cerevisiae snoRNAs terminate in a process that requires the RNA–binding proteins Nrd1, Nab3, and Sen1. We report here the transcriptome-wide distribution of these termination factors. These data sets derived from in vivo protein–RNA cross-linking provide high-resolution definition of non-poly(A) terminators, identify novel genes regulated by attenuation of nascent transcripts close to the promoter, and demonstrate the widespread occurrence of Nrd1-bound 3′ antisense transcripts on genes that are poorly expressed. In addition, we show that Sen1 does not cross-link efficiently to many expected non-coding RNAs but does cross-link to the 3′ end of most pre–mRNA transcripts, suggesting an extensive role in mRNA 3′ end formation and/or termination.
Highlights
In each transcription cycle RNA polymerase II (Pol II) can follow one of two paths; terminate early through the Nrd1-Nab3Sen1 pathway or continue on to form longer, potentially coding transcripts [1,2]
Nrd1, Nab3 and Sen1 have been shown to form a complex with Pol II [3] and to direct the termination and subsequent processing of nascent transcripts [16,40]
The observation that Nrd1 ChIPs to chromatin at highly expressed genes but only cross-links to a sub-set of these transcripts (Figure 1) is consistent with a model in which Nrd1, Nab3, and Sen1 enter the early Pol II elongation complex by association with the Ser5 phosphorylated Cterminal domain (CTD) [4,6]
Summary
In each transcription cycle RNA polymerase II (Pol II) can follow one of two paths; terminate early through the Nrd1-Nab3Sen pathway or continue on to form longer, potentially coding transcripts [1,2]. In this study we have used a recently developed in vivo crosslinking approach [18] to derive high-resolution transcriptomewide maps of binding sites for Nrd, Nab, Sen, and the Pol II subunit Rpb. In this study we have used a recently developed in vivo crosslinking approach [18] to derive high-resolution transcriptomewide maps of binding sites for Nrd, Nab, Sen, and the Pol II subunit Rpb2 This approach yields a more precise picture of known Nrd and Nab binding sites on snoRNA, CUT and mRNA targets and reveals a set of previously unknown Nrd binding sites both on the 59 ends of mRNAs and on 39 antisense transcripts. We observe Sen cross-linking on mRNA transcripts, at the 39 end suggesting a potential role for Sen in mRNA 39 end formation through the cleavage and polyadenylation pathway
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