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Genetic interaction mapping reveals a role for the SWI/SNF nucleosome remodeler in spliceosome activation in fission yeast.

Patrick KL, Ryan CJ, Xu J, Lipp JJ, Nissen KE, Roguev A, Shales M, Krogan NJ, Guthrie C - PLoS Genet. (2015)

Bottom Line: Overexpression of SF3 components in ΔSWI/SNF cells led to inefficient splicing of many fission yeast introns, predominantly those with non-consensus splice sites.Deletion of SWI/SNF decreased recruitment of the splicing ATPase Prp2, suggesting that SWI/SNF promotes co-transcriptional spliceosome assembly prior to first step catalysis.Importantly, defects in SWI/SNF as well as SF3 overexpression each altered nucleosome occupancy along intron-containing genes, illustrating that the chromatin landscape both affects--and is affected by--co-transcriptional splicing.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, California, United States of America.

ABSTRACT
Although numerous regulatory connections between pre-mRNA splicing and chromatin have been demonstrated, the precise mechanisms by which chromatin factors influence spliceosome assembly and/or catalysis remain unclear. To probe the genetic network of pre-mRNA splicing in the fission yeast Schizosaccharomyces pombe, we constructed an epistatic mini-array profile (E-MAP) and discovered many new connections between chromatin and splicing. Notably, the nucleosome remodeler SWI/SNF had strong genetic interactions with components of the U2 snRNP SF3 complex. Overexpression of SF3 components in ΔSWI/SNF cells led to inefficient splicing of many fission yeast introns, predominantly those with non-consensus splice sites. Deletion of SWI/SNF decreased recruitment of the splicing ATPase Prp2, suggesting that SWI/SNF promotes co-transcriptional spliceosome assembly prior to first step catalysis. Importantly, defects in SWI/SNF as well as SF3 overexpression each altered nucleosome occupancy along intron-containing genes, illustrating that the chromatin landscape both affects--and is affected by--co-transcriptional splicing.

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Splicing has strong negative genetic interactions with the chromatin machinery.(A.) Hierarchical clustering of genetic interaction profile correlations between each pair of splicing factors in the Splicing E-MAP. Negative correlations were scored as zero in the heatmap. Red boxes indicate clusters of splicing factors whose correlation coefficients > 0.3. Clusters 8, 9, and 10 are highlighted. (B.) Processes (top) and complexes (bottom) enriched for negative interactions with splicing in the Splicing E-MAP. Light blue box indicates color of significance cut-off. Processes were defined using the S. pombe GO Slim database and complexes were taken from the S. pombe cellular component Gene Ontology list (PomBase). Bonferonni corrected p value <0.0001 for processes and ≤0.0005 for complexes. (C.) Genetic interactions of Splicing Cluster 8 and 10 with the chromatin machinery. Blue indicates negative genetic interactions; yellow indicates positive; grey indicates no data. (D.) Splicing U snRNPs (blue) enriched for negative genetic interactions with chromatin complexes (yellow). U snRNPs were defined as listed in S2 Table. Bonferonni corrected p-value <0.05, False Discovery Rate <10%. (E.) Serial dilutions (1:5) of WT, single, and double mutants grown at 16°, 30° and 37°C.
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pgen.1005074.g001: Splicing has strong negative genetic interactions with the chromatin machinery.(A.) Hierarchical clustering of genetic interaction profile correlations between each pair of splicing factors in the Splicing E-MAP. Negative correlations were scored as zero in the heatmap. Red boxes indicate clusters of splicing factors whose correlation coefficients > 0.3. Clusters 8, 9, and 10 are highlighted. (B.) Processes (top) and complexes (bottom) enriched for negative interactions with splicing in the Splicing E-MAP. Light blue box indicates color of significance cut-off. Processes were defined using the S. pombe GO Slim database and complexes were taken from the S. pombe cellular component Gene Ontology list (PomBase). Bonferonni corrected p value <0.0001 for processes and ≤0.0005 for complexes. (C.) Genetic interactions of Splicing Cluster 8 and 10 with the chromatin machinery. Blue indicates negative genetic interactions; yellow indicates positive; grey indicates no data. (D.) Splicing U snRNPs (blue) enriched for negative genetic interactions with chromatin complexes (yellow). U snRNPs were defined as listed in S2 Table. Bonferonni corrected p-value <0.05, False Discovery Rate <10%. (E.) Serial dilutions (1:5) of WT, single, and double mutants grown at 16°, 30° and 37°C.

Mentions: Previous work (summarized in [28]) has shown that genes belonging to the same complex or pathway tend to have similar genetic interaction profiles. To gain an unbiased overview of the splicing factors in our E-MAP we clustered them according to the similarity of their genetic interaction profiles, i.e. based on how individual splicing factor mutants genetically interact with all factors represented in the deletion library. Several clusters (correlation coefficient > 0.3, see methods) emerged containing splicing factors from multiple steps of the splicing cycle (Fig. 1A and S2 Table). Consistent with our understanding of early spliceosome assembly, our E-MAP revealed a high degree of correlation between a pair of splicing factors, prp11 (PRP5; where appropriate, S. cerevisiae orthologs will be listed parenthetically) and the branchpoint binding protein bpb1 (MSL5/BBP1), which are both known to function at the step of intron recognition (Fig. 1A, Cluster 9). Interestingly, cwf23 (CWC23) is also part of Cluster 9, perhaps indicating a role for this DNAJ domain-containing protein at an early step of spliceosome assembly.


Genetic interaction mapping reveals a role for the SWI/SNF nucleosome remodeler in spliceosome activation in fission yeast.

Patrick KL, Ryan CJ, Xu J, Lipp JJ, Nissen KE, Roguev A, Shales M, Krogan NJ, Guthrie C - PLoS Genet. (2015)

Splicing has strong negative genetic interactions with the chromatin machinery.(A.) Hierarchical clustering of genetic interaction profile correlations between each pair of splicing factors in the Splicing E-MAP. Negative correlations were scored as zero in the heatmap. Red boxes indicate clusters of splicing factors whose correlation coefficients > 0.3. Clusters 8, 9, and 10 are highlighted. (B.) Processes (top) and complexes (bottom) enriched for negative interactions with splicing in the Splicing E-MAP. Light blue box indicates color of significance cut-off. Processes were defined using the S. pombe GO Slim database and complexes were taken from the S. pombe cellular component Gene Ontology list (PomBase). Bonferonni corrected p value <0.0001 for processes and ≤0.0005 for complexes. (C.) Genetic interactions of Splicing Cluster 8 and 10 with the chromatin machinery. Blue indicates negative genetic interactions; yellow indicates positive; grey indicates no data. (D.) Splicing U snRNPs (blue) enriched for negative genetic interactions with chromatin complexes (yellow). U snRNPs were defined as listed in S2 Table. Bonferonni corrected p-value <0.05, False Discovery Rate <10%. (E.) Serial dilutions (1:5) of WT, single, and double mutants grown at 16°, 30° and 37°C.
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pgen.1005074.g001: Splicing has strong negative genetic interactions with the chromatin machinery.(A.) Hierarchical clustering of genetic interaction profile correlations between each pair of splicing factors in the Splicing E-MAP. Negative correlations were scored as zero in the heatmap. Red boxes indicate clusters of splicing factors whose correlation coefficients > 0.3. Clusters 8, 9, and 10 are highlighted. (B.) Processes (top) and complexes (bottom) enriched for negative interactions with splicing in the Splicing E-MAP. Light blue box indicates color of significance cut-off. Processes were defined using the S. pombe GO Slim database and complexes were taken from the S. pombe cellular component Gene Ontology list (PomBase). Bonferonni corrected p value <0.0001 for processes and ≤0.0005 for complexes. (C.) Genetic interactions of Splicing Cluster 8 and 10 with the chromatin machinery. Blue indicates negative genetic interactions; yellow indicates positive; grey indicates no data. (D.) Splicing U snRNPs (blue) enriched for negative genetic interactions with chromatin complexes (yellow). U snRNPs were defined as listed in S2 Table. Bonferonni corrected p-value <0.05, False Discovery Rate <10%. (E.) Serial dilutions (1:5) of WT, single, and double mutants grown at 16°, 30° and 37°C.
Mentions: Previous work (summarized in [28]) has shown that genes belonging to the same complex or pathway tend to have similar genetic interaction profiles. To gain an unbiased overview of the splicing factors in our E-MAP we clustered them according to the similarity of their genetic interaction profiles, i.e. based on how individual splicing factor mutants genetically interact with all factors represented in the deletion library. Several clusters (correlation coefficient > 0.3, see methods) emerged containing splicing factors from multiple steps of the splicing cycle (Fig. 1A and S2 Table). Consistent with our understanding of early spliceosome assembly, our E-MAP revealed a high degree of correlation between a pair of splicing factors, prp11 (PRP5; where appropriate, S. cerevisiae orthologs will be listed parenthetically) and the branchpoint binding protein bpb1 (MSL5/BBP1), which are both known to function at the step of intron recognition (Fig. 1A, Cluster 9). Interestingly, cwf23 (CWC23) is also part of Cluster 9, perhaps indicating a role for this DNAJ domain-containing protein at an early step of spliceosome assembly.

Bottom Line: Overexpression of SF3 components in ΔSWI/SNF cells led to inefficient splicing of many fission yeast introns, predominantly those with non-consensus splice sites.Deletion of SWI/SNF decreased recruitment of the splicing ATPase Prp2, suggesting that SWI/SNF promotes co-transcriptional spliceosome assembly prior to first step catalysis.Importantly, defects in SWI/SNF as well as SF3 overexpression each altered nucleosome occupancy along intron-containing genes, illustrating that the chromatin landscape both affects--and is affected by--co-transcriptional splicing.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, California, United States of America.

ABSTRACT
Although numerous regulatory connections between pre-mRNA splicing and chromatin have been demonstrated, the precise mechanisms by which chromatin factors influence spliceosome assembly and/or catalysis remain unclear. To probe the genetic network of pre-mRNA splicing in the fission yeast Schizosaccharomyces pombe, we constructed an epistatic mini-array profile (E-MAP) and discovered many new connections between chromatin and splicing. Notably, the nucleosome remodeler SWI/SNF had strong genetic interactions with components of the U2 snRNP SF3 complex. Overexpression of SF3 components in ΔSWI/SNF cells led to inefficient splicing of many fission yeast introns, predominantly those with non-consensus splice sites. Deletion of SWI/SNF decreased recruitment of the splicing ATPase Prp2, suggesting that SWI/SNF promotes co-transcriptional spliceosome assembly prior to first step catalysis. Importantly, defects in SWI/SNF as well as SF3 overexpression each altered nucleosome occupancy along intron-containing genes, illustrating that the chromatin landscape both affects--and is affected by--co-transcriptional splicing.

Show MeSH