Sen1p Performs Two Genetically Separable Functions in Transcription and Processing of U5 Small Nuclear RNA in<i>Saccharomyces cerevisiae</i>

Finkel, Jonathan S.; Chinchilla, Karen; Ursic, Doris; Culbertson, Michael R.

doi:10.1534/genetics.109.110031

Cited by 39 publications

(57 citation statements)

References 46 publications

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“…Sen1 is required for termination and processing of a number of snoRNAs (Ursic et al 1997;Rasmussen and Culbertson 1998;Steinmetz et al 2001;Finkel et al 2010). Surprisingly, we observe efficient cross-linking of Sen1 only to box H/ACA snoRNAs.…”

Section: Resultsmentioning

confidence: 61%

Yeast Nrd1, Nab3, and Sen1 transcriptome-wide binding maps suggest multiple roles in post-transcriptional RNA processing

Jamonnak

Creamer

Darby

et al. 2011

RNA

102

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RNA polymerase II transcribes both coding and noncoding genes, and termination of these different classes of transcripts is facilitated by different sets of termination factors. Pre-mRNAs are terminated through a process that is coupled to the cleavage/ polyadenylation machinery, and noncoding RNAs in the yeast Saccharomyces cerevisiae are terminated through a pathway directed by the RNA-binding proteins Nrd1, Nab3, and the RNA helicase Sen1. We have used an in vivo cross-linking approach to map the binding sites of components of the yeast non-poly(A) termination pathway. We show here that Nrd1, Nab3, and Sen1 bind to a number of noncoding RNAs in an unexpected manner. Sen1 shows a preference for H/ACA over box C/D snoRNAs. Nrd1, which binds to snoRNA terminators, also binds to the upstream region of some snoRNA transcripts and to snoRNAs embedded in introns. We present results showing that several RNAs, including the telomerase RNA TLC1, require Nrd1 for proper processing. Binding of Nrd1 to transcripts from tRNA genes is another unexpected observation. We also observe RNA polymerase II binding to transcripts from RNA polymerase III genes, indicating a possible role for the Nrd1 pathway in surveillance of transcripts synthesized by the wrong polymerase. The binding targets of Nrd1 pathway components change in the absence of glucose, with Nrd1 and Nab3 showing a preference for binding to sites in the mature snoRNA and tRNAs. This suggests a novel role for Nrd1 and Nab3 in destruction of ncRNAs in response to nutrient limitation.

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Section: Resultsmentioning

confidence: 61%

Yeast Nrd1, Nab3, and Sen1 transcriptome-wide binding maps suggest multiple roles in post-transcriptional RNA processing

Jamonnak

Creamer

Darby

et al. 2011

RNA

102

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“…A plasmid-based system for characterizing yeast SEN1 function Previous genetic studies of S. cerevisiae SEN1 used primarily chromosomal mutations and tested a small number of alleles (Winey and Culbertson 1988;Steinmetz and Brow 1996;Finkel et al 2010). The essential chromosomal SEN1 gene was deleted previously (DeMarini et al 1992), but the complementing plasmid used, although functional, was subsequently found to lack the SEN1 promoter, 59-UTR, and part of the protein-coding region.…”

Section: Resultsmentioning

confidence: 99%

“…Functions of Sen1 in other nuclear processes have been proposed, including Pol I transcription termination, DNA repair, RNA processing, and maintenance of genome stability (Ursic et al 2004;Kawauchi et al 2008;Finkel et al 2010;Mischo et al 2011;Alzu et al 2012).…”

Section: Terminator Readthrough Is a Common Effect Of The Aoa2 Mutatimentioning

confidence: 99%

Saccharomyces cerevisiae Sen1 as a Model for the Study of Mutations in Human Senataxin That Elicit Cerebellar Ataxia

Chen¹,

Müller²,

Sundling³

et al. 2014

Genetics

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The nuclear RNA and DNA helicase Sen1 is essential in the yeast Saccharomyces cerevisiae and is required for efficient termination of RNA polymerase II transcription of many short noncoding RNA genes. However, the mechanism of Sen1 function is not understood. We created a plasmid-based genetic system to study yeast Sen1 in vivo. Using this system, we show that (1) the minimal essential region of Sen1 corresponds to the helicase domain and one of two flanking nuclear localization sequences; (2) a previously isolated terminator readthrough mutation in the Sen1 helicase domain, E1597K, is rescued by a second mutation designed to restore a salt bridge within the first RecA domain; and (3) the human ortholog of yeast Sen1, Senataxin, cannot functionally replace Sen1 in yeast. Guided by sequence homology between the conserved helicase domains of Sen1 and Senataxin, we tested the effects of 13 missense mutations that cosegregate with the inherited disorder ataxia with oculomotor apraxia type 2 on Sen1 function. Ten of the disease mutations resulted in transcription readthrough of at least one of three Sen1-dependent termination elements tested. Our genetic system will facilitate the further investigation of structure-function relationships in yeast Sen1 and its orthologs.T RANSCRIPTION termination by eukaryotic RNA polymerase II (Pol II) uses at least two pathways, one that is coupled to cleavage and polyadenylation of the nascent transcript [the poly(A)-dependent pathway] and one that involves the activity of the RNA/DNA helicase Sen1 (the Sen1-dependent pathway) (Kuehner et al. 2011). The Sen1-dependent pathway was first identified in the budding yeast Saccharomyces cerevisiae and is responsible for transcription termination of many short, noncoding RNA genes, including small nuclear (sn) and small nucleolar (sno) RNAs (Winey and Culbertson 1988;Steinmetz and Brow 1996;Rasmussen and Culbertson 1998;Steinmetz et al. 2001). It also restricts the elongation and accumulation of pervasive cryptic unstable transcripts (Arigo et al. 2006b;Thiebaut et al. 2006) and regulates transcription of some protein-coding genes by premature termination, i.e., attenuation (Steinmetz et al. 2001;Arigo et al. 2006a;Steinmetz et al. 2006b;Jenks et al. 2008;Kuehner and Brow 2008). A set of core factors distinguish the Sen1-dependent pathway from the poly(A)-dependent pathway, including Sen1 and the RNA-binding proteins Nrd1 and Nab3 Brow 1996, 1998;Conrad et al. 2000;Steinmetz et al. 2001;Carroll et al. 2007). However, some short messenger RNA (mRNA) genes, such as CYC1, may have hybrid terminators that require factors from both pathways (Steinmetz et al. 2006b).S. cerevisiae Sen1 is a 252-kDa superfamily 1 helicase encoded by the essential SEN1 gene. Its helicase domain is located in the C-terminal half of the protein ( Figure 1A), and the N-terminal 975 amino acids are dispensable for viability (DeMarini et al. 1992). The ortholog of Sen1 in the fission yeast Schizosaccharomyces pombe has ATP-dependent, 59-to-39 DNA and RNA unw...

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“…First, we tested Sen1-Pol II interaction under lowered CTD Ser2 phosphorylation in pcf11-13. We used the sen1-R302W mutant, which harbors a point mutation in the N terminus that weakens Sen1-Pol II interaction (Finkel et al 2010;Chinchilla et al 2012). Western blot analysis revealed reduced Pol II coimmunoprecipitated with Sen1-R302W in pcf11-13, while the level of Nab3 coimmunoprecipitation with Sen1-R302W was unaffected as compared with wild type (Fig.…”

Section: Pcf11 Promotes Sen1 Functionmentioning

confidence: 99%

“…Moreover, inactivation of Ser2 kinase Bur1 or Ctk1 results in transcription readthrough at NRD terminators (Tomson et al 2012;Lenstra et al 2013). Ser2-P marks are also required for Sen1-Pol II binding, which is necessary for NRD-dependent termination in vivo (Finkel et al 2010;Chinchilla et al 2012;Chen et al 2014) but not in vitro (Porrua and Libri 2013).…”

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confidence: 99%

Pcf11 orchestrates transcription termination pathways in yeast

2015

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In Saccharomyces cerevisiae, short noncoding RNA (ncRNA) generated by RNA polymerase II (Pol II) are terminated by the NRD complex consisting of Nrd1, Nab3, and Sen1. We now show that Pcf11, a component of the cleavage and polyadenylation complex (CPAC), is also generally required for NRD-dependent transcription termination through the action of its C-terminal domain (CTD)-interacting domain (CID). Pcf11 localizes downstream from Nrd1 on NRD terminators, and its recruitment depends on Nrd1. Furthermore, mutation of the Pcf11 CID results in Nrd1 retention on chromatin, delayed degradation of ncRNA, and restricted Pol II CTD Ser2 phosphorylation and Sen1-Pol II interaction. Finally, the pcf11-13 and sen1-1 mutant phenotypes are very similar, as both accumulate RNA:DNA hybrids and display Pol II pausing downstream from NRD terminators. We predict a mechanism by which the exchange of Nrd1 and Pcf11 on chromatin facilitates Pol II pausing and CTD Ser2-P phosphorylation. This in turn promotes Sen1 activity that is required for NRD-dependent transcription termination in vivo.

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Sen1p Performs Two Genetically Separable Functions in Transcription and Processing of U5 Small Nuclear RNA inSaccharomyces cerevisiae

Cited by 39 publications

References 46 publications

Yeast Nrd1, Nab3, and Sen1 transcriptome-wide binding maps suggest multiple roles in post-transcriptional RNA processing

Yeast Nrd1, Nab3, and Sen1 transcriptome-wide binding maps suggest multiple roles in post-transcriptional RNA processing

Saccharomyces cerevisiae Sen1 as a Model for the Study of Mutations in Human Senataxin That Elicit Cerebellar Ataxia

Pcf11 orchestrates transcription termination pathways in yeast

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