A handful of intron-containing genes produces the lion's share of yeast mRNA

Ares, Manuel; Grate, Leslie R.; Pauling, Michelle H.

doi:10.1017/s1355838299991379

Cited by 143 publications

(119 citation statements)

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“…The in vivo rate of transcription by RNAP II is estimated at ∼30 nt/sec, indicating that the time advantage enjoyed by the intron 1 branchpoint sequence over its counterpart in intron 2 is ∼3 sec. The kinetics of spliceosome assembly in vivo are unknown, but it has been estimated based on the number of introns that must be spliced per hour by the 500 molecules of U2 snRNA that a complete cycle of intron removal and U2 snRNP regeneration must take <3 min (Ares et al 1999). Depending on the fraction of the average cycle time devoted to branchpoint recognition, the 3 sec that the intron 1 branchpoint spends in the nucleoplasm before the intron 2 branchpoint is made could provide a significant advantage in the competition to pair with the 5Ј splice site.…”

Section: Can Intron 1 Be Recognized Before Rnap II Gets To the End Ofmentioning

confidence: 99%

Perturbation of transcription elongation influences the fidelity of internal exon inclusion in Saccharomyces cerevisiae

Howe

Kane²,

Ares³

2003

RNA

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149

131

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Unknown mechanisms exist to ensure that exons are not skipped during biogenesis of mRNA. Studies have connected transcription elongation with regulated alternative exon inclusion. To determine whether the relative rates of transcription elongation and spliceosome assembly might play a general role in enforcing constitutive exon inclusion, we measured exon skipping for a natural two-intron gene in which the internal exon is constitutively included in the mRNA. Mutations in this gene that subtly reduce recognition of the intron 1 branchpoint cause exon skipping, indicating that rapid recognition of the first intron is important for enforcing exon inclusion. To test the role of transcription elongation, we treated cells to increase or decrease the rate of transcription elongation. Consistent with the "first come, first served" model, we found that exon skipping in vivo is inhibited when transcription is slowed by RNAP II mutants or when cells are treated with inhibitors of elongation. Expression of the elongation factor TFIIS stimulates exon skipping, and this effect is eliminated when lac repressor is targeted to DNA encoding the second intron. A mutation in U2 snRNA promotes exon skipping, presumably because a delay in recognition of the first intron allows elongating RNA polymerase to transcribe the downstream intron. This indicates that the relative rates of elongation and splicing are tuned so that the fidelity of exon inclusion is enhanced. These findings support a general role for kinetic coordination of transcription elongation and splicing during the transcription-dependent control of splicing.

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Section: Can Intron 1 Be Recognized Before Rnap II Gets To the End Ofmentioning

confidence: 99%

Perturbation of transcription elongation influences the fidelity of internal exon inclusion in Saccharomyces cerevisiae

Howe

Kane²,

Ares³

2003

RNA

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149

131

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“…Because ribosome biogenesis is so energetically costly, eukaryotes have evolved multiple means to regulate rRNA and ribosomal protein production in response to changes in cellular demand and surveillance systems to remove aberrant ribosomal protein complexes formed during assembly or after environmental insult (Jorgensen et al 2002;Fingerman et al 2003; FromontRacine et al 2003;Jorgensen et al 2004;Marion et al 2004;Henras et al 2008;Kressler et al 2010;Lafontaine 2010). The coordination of pre-mRNA processing with ribosome biogenesis is especially relevant in the intron-poor environment of the yeast genome, where the highly expressed ribosomal protein transcripts represent a disproportionate amount of the spliced mRNA (Ares et al 1999;Staley and Woolford 2009). Our understanding of how this coordination is accomplished is limited, however, to general principles supported by a few specific examples where individual ribosomal proteins act as feedback regulators to inhibit the processing or stability of cognate ribosomal protein transcripts (Li et al 1995(Li et al , 1996Vilardell et al 2000;Pleiss et al 2007;Gudipati et al 2012).…”

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confidence: 99%

Limited Portability of G-Patch Domains in Regulators of the Prp43 RNA Helicase Required for Pre-mRNA Splicing and Ribosomal RNA Maturation in Saccharomyces cerevisiae

Banerjee¹,

McDaniel²,

Rymond

2015

Genetics

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The Prp43 DExD/H-box protein is required for progression of the biochemically distinct pre-messenger RNA and ribosomal RNA (rRNA) maturation pathways. In Saccharomyces cerevisiae, the Spp382/Ntr1, Sqs1/Pfa1, and Pxr1/Gno1 proteins are implicated as cofactors necessary for Prp43 helicase activation during spliceosome dissociation (Spp382) and rRNA processing (Sqs1 and Pxr1). While otherwise dissimilar in primary sequence, these Prp43-binding proteins each contain a short glycine-rich G-patch motif required for function and thought to act in protein or nucleic acid recognition. Here yeast two-hybrid, domain-swap, and site-directed mutagenesis approaches are used to investigate G-patch domain activity and portability. Our results reveal that the Spp382, Sqs1, and Pxr1 G-patches differ in Prp43 two-hybrid response and in the ability to reconstitute the Spp382 and Pxr1 RNA processing factors. G-patch protein reconstitution did not correlate with the apparent strength of the Prp43 two-hybrid response, suggesting that this domain has function beyond that of a Prp43 tether. Indeed, while critical for Pxr1 activity, the Pxr1 G-patch appears to contribute little to the yeast two-hybrid interaction. Conversely, deletion of the primary Prp43 binding site within Pxr1 (amino acids 102-149) does not impede rRNA processing but affects small nucleolar RNA (snoRNA) biogenesis, resulting in the accumulation of slightly extended forms of select snoRNAs, a phenotype unexpectedly shared by the prp43 loss-of-function mutant. These and related observations reveal differences in how the Spp382, Sqs1, and Pxr1 proteins interact with Prp43 and provide evidence linking G-patch identity with pathway-specific DExD/H-box helicase activity.KEYWORDS Saccharomyces cerevisiae; spliceosome; rRNA processing; DExD/H-box protein; pre-mRNA splicing N UMEROUS genome-wide interaction and gene expression studies identify ribosome biogenesis as a central integrative feature of Saccharomyces cerevisiae metabolism (see, e.g ., Mnaimneh et al. 2004;Davierwala et al. 2005;Gavin et al. 2006;Collins et al. 2007;Hu et al. 2007;Magtanong et al. 2011). In rapidly dividing organisms such as this budding yeast, ribosomal RNAs (rRNAs) make up the lion's share of cellular nucleic acid by mass, while ribosomal protein transcripts can account for more than half the transcribed messenger RNA (mRNA). Because ribosome biogenesis is so energetically costly, eukaryotes have evolved multiple means to regulate rRNA and ribosomal protein production in response to changes in cellular demand and surveillance systems to remove aberrant ribosomal protein complexes formed during assembly or after environmental insult (Jorgensen et al. 2002;Fingerman et al. 2003; FromontRacine et al. 2003;Jorgensen et al. 2004;Marion et al. 2004;Henras et al. 2008;Kressler et al. 2010;Lafontaine 2010). The coordination of pre-mRNA processing with ribosome biogenesis is especially relevant in the intron-poor environment of the yeast genome, where the highly expressed ribosomal protein transcr...

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“…Alexandrov et al pared to tRNA (Waldron & Lacroute, 1975;Ares et al+, 1999), and because our preparations are enriched for low molecular weight RNA+ However, it is formally possible that residual m 7 G in RNA from trm8-⌬/trm8-⌬ and trm82-⌬/trm82-⌬ strains could arise from one or more of the 10 other tRNA species known to have m 7 G in their tRNA (Sprinzl et al+, 1998), or from rRNA (Zueva et al+, 1985b), as we have not explicitly assessed their m 7 G content+ Nonetheless, it seems more likely to us that the m 7 G modification at position 46 in different tRNAs is mediated by one Trm8/ Trm82 protein complex, much as i 6 A (Dihanich et al+, 1987), m 2 2 G (Ellis et al+, 1986), and m 5 C (Motorin & Grosjean, 1999) modifications are each formed in different tRNAs by a single protein+…”

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confidence: 99%

Two proteins that form a complex are required for 7-methylguanosine modification of yeast tRNA

Alexandrov

Martzen

Phizicky

2002

RNA

281

278

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ABSTRACT7-methylguanosine (m 7 G) modification of tRNA occurs widely in eukaryotes and bacteria, is nearly always found at position 46, and is one of the few modifications that confers a positive charge to the base. Screening of a Saccharomyces cerevisiae genomic library of purified GST-ORF fusion proteins reveals two previously uncharacterized proteins that copurify with m 7 G methyltransferase activity on pre-tRNA Phe . ORF YDL201w encodes Trm8, a protein that is highly conserved in prokaryotes and eukaryotes and that contains an S-adenosylmethionine binding domain. ORF YDR165w encodes Trm82, a less highly conserved protein containing putative WD40 repeats, which are often implicated in macromolecular interactions. Neither protein has significant sequence similarity to yeast Abd1, which catalyzes m 7 G modification of the 59 cap of mRNA, other than the methyltransferase motif shared by Trm8 and Abd1. Several lines of evidence indicate that both Trm8 and Trm82 proteins are required for tRNA m 7 G-methyltransferase activity: Extracts derived from strains lacking either gene have undetectable m 7 G methyltransferase activity, RNA from strains lacking either gene have much reduced m 7 G, and coexpression of both proteins is required to overproduce activity. Aniline cleavage mapping shows that Trm8/Trm82 proteins modify pre-tRNA Phe at G46, the site that is modified in vivo. Trm8 and Trm82 proteins form a complex, as affinity purification of Trm8 protein causes copurification of Trm82 protein in approximate equimolar yield. This functional two-protein family appears to be retained in eukaryotes, as expression of both corresponding human proteins, METTL1 and WDR4, is required for m 7 Gmethyltransferase activity.

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A handful of intron-containing genes produces the lion's share of yeast mRNA

Cited by 143 publications

References 9 publications

Perturbation of transcription elongation influences the fidelity of internal exon inclusion in Saccharomyces cerevisiae

Perturbation of transcription elongation influences the fidelity of internal exon inclusion in Saccharomyces cerevisiae

Limited Portability of G-Patch Domains in Regulators of the Prp43 RNA Helicase Required for Pre-mRNA Splicing and Ribosomal RNA Maturation in Saccharomyces cerevisiae

Two proteins that form a complex are required for 7-methylguanosine modification of yeast tRNA

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