Rapid, Transcript-Specific Changes in Splicing in Response to Environmental Stress

Pleiss, Jeffrey A.; Whitworth, Gregg B.; Bergkessel, Megan; Guthrie, Christine

doi:10.1016/j.molcel.2007.07.018

Cited by 207 publications

(223 citation statements)

References 39 publications

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“…This shift is necessary to accommodate the complex landscape of cis-and trans-acting factors from a genomic perspective, which is not captured by model introns or genes. Recent RNA interference and microarray analyses in Drosophila and S. cerevisiae pointed to the possibility of a rich diversity of requirements for splicing factors/regulators (6,7,22,38,40,41). The present study underscores the importance of how weakening (or strengthening) one or more factors in a system can influence the need for others.…”

Section: Discussionsupporting

confidence: 50%

Genomic mRNA Profiling Reveals Compensatory Mechanisms for the Requirement of the Essential Splicing Factor U2AF

Sridharan

Heimiller

Singh

2011

Molecular and Cellular Biology

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The large subunit of the U2 auxiliary factor (U2AF) recognizes the polypyrimidine tract (Py-tract) located adjacent to the 3 splice site to facilitate U2 snRNP recruitment. While U2AF is considered essential for pre-mRNA splicing, its requirement for splicing on a genome-wide level has not been analyzed. Using Solexa sequencing, we performed mRNA profiling for splicing in the Schizosaccharomyces pombe U2AF 59 (prp2.1) temperature-sensitive mutant. Surprisingly, our analysis revealed that introns show a range of splicing defects in the mutant strain. While U2AF 59 inactivation (nonpermissive) conditions inhibit splicing of some introns, others are spliced apparently normally. Bioinformatics analysis indicated that U2AF 59 -insensitive introns have stronger 5 splice sites and higher A/U content. Most importantly, features that contribute to U2AF 59 insensitivity of an intron unexpectedly reside in its 5-most 30 nucleotides. These include the 5 splice site, a guanosine at position 7, and the 5 splice site-to-branch point sequence context. A differential requirement (similar to U2AF 59 ) for introns may also apply to other general splicing factors (e.g., prp10). Our combined results indicate that U2AF insensitivity is a common phenomenon and that varied intron features support the existence of unrecognized aspects of spliceosome assembly.Pre-mRNA splicing plays a major role in gene regulation (23). Furthermore, alternative splicing serves as an important mechanism to generate molecular diversity (5). In the initial stages of splicing, the U1 snRNP recognizes the 5Ј splice site (5Ј SS), and the U2 snRNP auxiliary factor (U2AF) recognizes the polypyrimidine tract (Py-tract)/3Ј splice site (3Ј SS), leading to U2 snRNP recruitment to the branch point sequence (BPS) (27).In humans, the essential splicing factor U2AF, which has been extensively studied, is a heterodimeric protein containing a large subunit (U2AF65) and a small subunit (U2AF35). These subunits bind to the Py-tract and the 3Ј SS, respectively (37,65,68,71). Both subunits of U2AF are essential for viability in many model organisms, such as zebrafish, the fruit fly, the nematode worm, and fission yeast Schizosaccharomyces pombe (U2AF59) (17,28,42,46,47,59,72). However, in the budding yeast Saccharomyces cerevisiae, the large subunit is dispensable and the small subunit is absent (1). In humans, U2AF65 interacts with several splicing factors (BBP/SF1, UAP56, SAP155 [or SF3b155], and p54) (1,14,19,43,45,70) and facilitates branch point recognition (2, 4). The mammalian U2AF65 is found to be dispensable for splicing of some introns in vitro in the presence of the SR protein SC35 (36). Microarray analyses revealed an unexpected role of the Drosophila melanogaster large subunit (dU2AF50) in the nuclear export of intronless mRNAs (8). Several splicing regulators, such as SXL, PTB, hnRNP A1, ASF/SF2, SC35, and TRA, can facilitate or antagonize U2AF activity for splicing regulation (5, 53).Biochemical and structural studies indicate that the essential splicing factor...

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Section: Discussionsupporting

confidence: 50%

Genomic mRNA Profiling Reveals Compensatory Mechanisms for the Requirement of the Essential Splicing Factor U2AF

Sridharan

Heimiller

Singh

2011

Molecular and Cellular Biology

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“…The splicing defects we observe are similar in magnitude to those observed when other histone modifications are perturbed. [27][28][29][30]64 In addition, gene-specific splicing defects in budding yeast have been described previously [12][13][14] and could possibly be a result of features of the gene or intron such as the length of the gene, and/or its transcription frequency. Nevertheless, we do observe a trend of decreased splicing efficiency upon removal of Set2, as compared to the wild-type strain ( Fig.…”

Section: H3k36 Methylation Loss Results In Gene-specific Pre-mrna Splmentioning

confidence: 98%

“…10,11 Most S. cerevisiae genes do not contain more than one intron, and thus extensive alternative splicing does not occur. However, several recent reports show that the splicing efficiency of specific subsets of transcripts changes in different environmental conditions 12 or when splicing-related proteins are mutated. 13,14 Mutations that alter transcription elongation rates or treatment with drugs that affect transcription can change alternative splicing outcomes in metazoa [15][16][17][18][19][20] and splicing efficiency in yeast.…”

Section: Introductionmentioning

confidence: 99%

Histone H3K36 methylation regulates pre-mRNA splicing inSaccharomyces cerevisiae

et al. 2016

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Co-transcriptional splicing takes place in the context of a highly dynamic chromatin architecture, yet the role of chromatin restructuring in coordinating transcription with RNA splicing has not been fully resolved. To further define the contribution of histone modifications to pre-mRNA splicing in Saccharomyces cerevisiae, we probed a library of histone point mutants using a reporter to monitor pre-mRNA splicing. We found that mutation of H3 lysine 36 (H3K36) -a residue methylated by Set2 during transcription elongation -exhibited phenotypes similar to those of pre-mRNA splicing mutants. We identified genetic interactions between genes encoding RNA splicing factors and genes encoding the H3K36 methyltransferase Set2 and the demethylase Jhd1 as well as point mutations of H3K36 that block methylation. Consistent with the genetic interactions, deletion of SET2, mutations modifying the catalytic activity of Set2 or H3K36 point mutations significantly altered expression of our reporter and reduced splicing of endogenous introns. These effects were dependent on the association of Set2 with RNA polymerase II and H3K36 dimethylation. Additionally, we found that deletion of SET2 reduces the association of the U2 and U5 snRNPs with chromatin. Thus, our study provides the first evidence that H3K36 methylation plays a role in co-transcriptional RNA splicing in yeast.Abbreviations: (H3K36), histone H3 lysine 36; (ChIP), chromatin immunoprecipitations; (RT-qPCR), reverse transcription PCR; (snRNP), small nuclear ribonucleprotein; (RNA Pol II), RNA polymerase II

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“…Geranylgeranyl substrates include various Ras, Rho, Rab, and Rac signaling proteins (Dogbo et al 1988;Kuzuguchi et al 1999). While the biochemical basis of these genetic interactions needs additional study, it is enticing to speculate that geranylgeranyl addition may influence signal transduction pathway(s) shown to promote changes in splicing efficiency (Pleiss et al 2007;see Lynch 2007).…”

Section: Discussionmentioning

confidence: 99%

Spp382p Interacts with Multiple Yeast Splicing Factors, Including Possible Regulators of Prp43 DExD/H-Box Protein Function

Pandit

Paul

Zhang³

et al. 2009

Genetics

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Prp43p catalyzes essential steps in pre-mRNA splicing and rRNA biogenesis. In splicing, Spp382p stimulates the Prp43p helicase to dissociate the postcatalytic spliceosome and, in some way, to maintain the integrity of the spliceosome assembly. Here we present a dosage interference assay to identify Spp382p-interacting factors by screening for genes that when overexpressed specifically inhibit the growth of a conditional lethal prp38-1 spliceosome assembly mutant in the spp382-1 suppressor background. Identified, among others, are genes encoding the established splicing factors Prp8p, Prp9p, Prp11p, Prp39p, and Yhc1p and two poorly characterized proteins with possible links to splicing, Sqs1p and Cwc23p. Sqs1p copurifies with Prp43p and is shown to bind Prp43p and Spp382p in the two-hybrid assay. Overexpression of Sqs1p blocks pre-mRNA splicing and inhibits Prp43p-dependent steps in rRNA processing. Increased Prp43p levels buffer Sqs1p cytotoxicity, providing strong evidence that the Prp43p DExD/H-box protein is a target of Sqs1p. Cwc23p is the only known yeast splicing factor with a DnaJ motif characteristic of Hsp40-like chaperones. We show that similar to SPP382, CWC23 activity is critical for efficient pre-mRNA splicing and intron metabolism yet, surprisingly, this activity does not require the canonical DnaJ/Hsp40 motif. These and related data establish the value of this dosage interference assay for finding genes that alter cellular splicing and define Sqs1p and Cwc23p as prospective modulators of Spp382p-stimuated Prp43p function. E IGHT phylogenetically conserved DExD/H-box proteins act at discrete steps to regulate the assembly, activation, and dissociation of the splicing apparatus (reviewed in Brow 2002; Konarska and Query 2005;Linder 2006). How these RNA-dependent ATPases are temporally and functionally regulated remains poorly understood. One putative regulator, the 83-kDa Spp382/Ntr1 protein (henceforth referred to by the Saccharomyces Genome Database standard name, Spp382p), was discovered in a screen for mutants capable of suppressing defects in yeast spliceosome assembly (Pandit et al. 2006). While spp382 null alleles are lethal, partial loss of function suppresses mutations in several other splicing factors, including the genes encoding the essential spliceosomal proteins Prp8p and Prp38p. Spp382p is a spliceosomal protein that binds the Prp43p DExD/H-box protein to promote efficient dissociation of spliceosomal factors after completion of splicing in vitro (Tsai et al. 2005). Some but not all spp382 mutants also accumulate the excised intron product of splicing in vivo, ostensibly due to protection of the intron within a hyperstabilized spliceosome (Pandit et al. 2006;Tanaka et al. 2007). The suppression of spliceosome assembly defects by spp382 mutation is proposed to occur by impairing Spp382p-stimulated dissociation of kinetically impaired or otherwise inefficient spliceosomes by Prp43p (Pandit et al. 2006). Consistent with this hypothesis, prp43 mutations also suppress spliceosome a...

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Rapid, Transcript-Specific Changes in Splicing in Response to Environmental Stress

Cited by 207 publications

References 39 publications

Genomic mRNA Profiling Reveals Compensatory Mechanisms for the Requirement of the Essential Splicing Factor U2AF

Genomic mRNA Profiling Reveals Compensatory Mechanisms for the Requirement of the Essential Splicing Factor U2AF

Histone H3K36 methylation regulates pre-mRNA splicing inSaccharomyces cerevisiae

Spp382p Interacts with Multiple Yeast Splicing Factors, Including Possible Regulators of Prp43 DExD/H-Box Protein Function

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