Individual letters of the RNA polymerase II CTD code govern distinct gene expression programs in fission yeast

Schwer, Beate; Bitton, Danny A.; Sánchez, Ana M.; Bähler, Jürg; Shuman, Stewart

doi:10.1073/pnas.1321842111

Cited by 52 publications

(90 citation statements)

References 30 publications

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“…Publicly available data sets were downloaded from GEO, SRA, or ArrayExpress repositories (accessions indicated in Supplemental Table S1). Because the exon-skipping discovery pipeline does not support paired-end reads, only the 5 ′ -end of the read was used for all paired-end data sets (Livesay et al 2013;Schwer et al 2014). An earlier version of our intron discovery pipeline supported paired-end data sets; thus, both ends of the paired-end reads from Rhind et al (2011) were used for novel intron discovery.…”

Section: Methodsmentioning

confidence: 99%

Widespread exon skipping triggers degradation by nuclear RNA surveillance in fission yeast

et al. 2015

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Exon skipping is considered a principal mechanism by which eukaryotic cells expand their transcriptome and proteome repertoires, creating different splice variants with distinct cellular functions. Here we analyze RNA-seq data from 116 transcriptomes in fission yeast (Schizosaccharomyces pombe), covering multiple physiological conditions as well as transcriptional and RNA processing mutants. We applied brute-force algorithms to detect all possible exon-skipping events, which were widespread but rare compared to normal splicing events. Exon-skipping events increased in cells deficient for the nuclear exosome or the 5 ′ -3 ′ exonuclease Dhp1, and also at late stages of meiotic differentiation when nuclear-exosome transcripts decreased. The pervasive exon-skipping transcripts were stochastic, did not increase in specific physiological conditions, and were mostly present at less than one copy per cell, even in the absence of nuclear RNA surveillance and during late meiosis. These exon-skipping transcripts are therefore unlikely to be functional and may reflect splicing errors that are actively removed by nuclear RNA surveillance. The average splicing rate by exon skipping was ∼0.24% in wild type and ∼1.75% in nuclear exonuclease mutants. We also detected approximately 250 circular RNAs derived from single or multiple exons. These circular RNAs were rare and stochastic, although a few became stabilized during quiescence and in splicing mutants. Using an exhaustive search algorithm, we also uncovered thousands of previously unknown splice sites, indicating pervasive splicing; yet most of these splicing variants were cryptic and increased in nuclear degradation mutants. This study highlights widespread but low frequency alternative or aberrant splicing events that are targeted by nuclear RNA surveillance.[Supplemental material is available for this article.]Splicing is a fundamental step in gene expression, which determines the information content of messenger RNAs (mRNAs). It is a highly accurate process that relies on the spliceosome to identify sequence signals and remove noncoding introns from precursor mRNAs (pre-mRNAs) and thus generate translatable mRNAs. In its simplest form, splicing requires several intronic sequence elements for intron excision, including splice donor and acceptor sites, a branch-site, and a polypyrimidine tract (Wang and Burge 2008). Another layer of complexity, alternative splicing, results in the production of multiple transcripts from a single gene, some of which may be condition-or tissue-specific . These transcripts, referred to as alternative splice variants, carry nonconsecutive combinations of exons and are thought to be a major source of transcriptome and proteome diversity in multicellular eukaryotes ( Alternative splice variants may arise via single or multiple exon-skipping or intron-retention events or via alternative 5 ′ -or 3 ′ -splice sites . The extent of alternative splicing in the human transcriptome is potentially immense, with >90% of genes showing more than one isoform...

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Section: Methodsmentioning

confidence: 99%

Widespread exon skipping triggers degradation by nuclear RNA surveillance in fission yeast

et al. 2015

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“…Induction of the phosphate-regulated genes is strictly dependent on transcription factor Pho7 (Henry et al 2011;Carter-O'Connell et al 2012). Repression of pho1 under phosphate replete conditions is itself an active process involving protein kinases Csk1 and Cdk9 and the phosphorylation of the carboxyl-terminal domain (CTD) of the Rpb1 subunit of RNA polymerase II (Pol2) (Carter-O'Connell et al 2012;Schwer et al 2014Schwer et al , 2015.…”

Section: Introductionmentioning

confidence: 99%

“…Phosphorylation of the CTD Tyr1, Ser2, Thr4, Ser5, and Ser7 residues inscribes a dynamic "CTD code" that orchestrates transcription, cotranscriptional RNA processing, and chromatin modifications (Schwer and Shuman 2011;Schwer et al 2012;Corden 2013;Eick and Geyer 2013;Doamekpor et al 2014;Mbogning et al 2015). Deletion of Csk1, missense mutations of the Cdk9 T-loop threonine, and alanine mutations of Pol2 CTD phospho-sites Ser5, Pro6, or Ser7 cause derepression of pho1 in phosphate replete cells (Carter-O'Connell et al 2012;Schwer et al 2014Schwer et al , 2015. In contrast, CTD mutations T4A and S7E, and inactivation of CTD phosphatase Ssu72, hyper-repress pho1 in phosphate replete cells .…”

Section: Introductionmentioning

confidence: 99%

Transcription of lncRNA prt, clustered prt RNA sites for Mmi1 binding, and RNA polymerase II CTD phospho-sites govern the repression of pho1 gene expression under phosphate-replete conditions in fission yeast

Chatterjee

Sánchez

Goldgur

et al. 2016

RNA

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Expression of fission yeast Pho1 acid phosphatase is repressed during growth in phosphate-rich medium. Repression is mediated by transcription of the prt locus upstream of pho1 to produce a long noncoding (lnc) prt RNA. Repression is also governed by RNA polymerase II CTD phosphorylation status, whereby inability to place a Ser7-PO 4 mark (as in S7A) derepresses Pho1 expression, and inability to place a Thr4-PO 4 mark (as in T4A) hyper-represses Pho1 in phosphate replete cells. Here we find that basal pho1 expression from the prt-pho1 locus is inversely correlated with the activity of the prt promoter, which resides in a 110-nucleotide DNA segment preceding the prt transcription start site. CTD mutations S7A and T4A had no effect on the activity of the prt promoter or the pho1 promoter, suggesting that S7A and T4A affect post-initiation events in prt lncRNA synthesis that make it less and more repressive of pho1, respectively. prt lncRNA contains clusters of DSR (determinant of selective removal) sequences recognized by the YTH-domain-containing protein Mmi1. Altering the nucleobase sequence of two DSR clusters in the prt lncRNA caused hyper-repression of pho1 in phosphate replete cells, concomitant with increased levels of the prt transcript. The isolated Mmi1 YTH domain binds to RNAs with single or tandem DSR elements, to the latter in a noncooperative fashion. We report the 1.75 Å crystal structure of the Mmi1 YTH domain and provide evidence that Mmi1 recognizes DSR RNA via a binding mode distinct from that of structurally homologous YTH proteins that recognize m 6 Amodified RNA.

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“…Upregulation of the expression of these genes during phosphate starvation depends on the transcription factor Pho7 (Henry et al 2011;Carter-O'Connell et al 2012), a member of the zinc binuclear cluster family of fungal DNA-binding transcription regulators (MacPherson et al 2006). The protein kinase Csk1 is a negative regulator of the phosphate response, as judged by the fact that pho1 expression is constitutively turned on in csk1Δ cells under phosphate-replete conditions (Henry et al 2011;Carter-O'Connell et al 2012;Schwer et al 2014). Csk1 is a CDK-activating kinase with several physiological targets, including the kinases Cdc2, Mcs6, and Cdk9 (Saiz and Fisher 2002;Pei et al 2006).…”

Section: Introductionmentioning

confidence: 99%

RNA polymerase II CTD phospho-sites Ser5 and Ser7 govern phosphate homeostasis in fission yeast

Schwer

Sánchez

Shuman

2015

RNA

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Phosphorylation of the tandem YSPTSPS repeats of the RNA polymerase II CTD inscribes an informational code that orchestrates eukaryal mRNA synthesis. Here we interrogate the role of the CTD in phosphate homeostasis in fission yeast. Expression of Pho1 acid phosphatase, which is repressed during growth in phosphate-rich medium and induced by phosphate starvation, is governed strongly by CTD phosphorylation status, but not by CTD repeat length. Inability to place a Ser7-PO 4 mark (as in S7A) results in constitutive derepression of Pho1 expression in phosphate-replete medium. In contrast, indelible installation of a Ser7-PO 4 mimetic (as in S7E) hyper-represses Pho1 in phosphate-replete cells and inhibits Pho1 induction during starvation. Pho1 phosphatase is derepressed by ablation of the CTD Ser5-PO 4 mark, achieved either by mutating Ser5 in all consensus heptads to alanine, or replacing all Pro6 residues with alanine. We find that Ser5 status is a tunable determinant of Pho1 regulation, i.e., serial decrements in the number of consensus Ser5 heptads from seven to two elicits a progressive increase in Pho1 expression in phosphate-replete medium. Pho1 is also derepressed by hypomorphic mutations of the CTD kinase Cdk9. Inactivation of the CTD phosphatase Ssu72 attenuates Pho1 induction in wild-type cells and blocks Pho1 derepression in S7A cells. These experiments implicate Ser5, Pro6, and Ser7 as component letters of a CTD coding "word" that transduces a repressive transcriptional signal via serine phosphorylation.

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Individual letters of the RNA polymerase II CTD code govern distinct gene expression programs in fission yeast

Cited by 52 publications

References 30 publications

Widespread exon skipping triggers degradation by nuclear RNA surveillance in fission yeast

Widespread exon skipping triggers degradation by nuclear RNA surveillance in fission yeast

Transcription of lncRNA prt, clustered prt RNA sites for Mmi1 binding, and RNA polymerase II CTD phospho-sites govern the repression of pho1 gene expression under phosphate-replete conditions in fission yeast

RNA polymerase II CTD phospho-sites Ser5 and Ser7 govern phosphate homeostasis in fission yeast

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