Search

Human transcription co-regulator SNW1/SKIP modulates the activity of various transcription factors, the elongating complex of RNA-PolII, and the spliceosome, which suggests that SNW1/SKIP may link pre-mRNA splicing to other processes of gene expression. We identified temperature sensitive alleles of the yeast homolog of SNW1/SKIP, PRP45, which at permissive temperature elicit cell shape and cell division defects. Cwc2-TAP purified spliceosomal complexes from prp45(1-169) cells showed a decreased stoichiometry of Prp22, suggesting an aberrant association of the helicase with the spliceosome. Using a synthetic lethality screen, we isolated several alleles of PRP22 which genetically interact with the allele prp45(1-169). Splicing analysis in vivo showed that prp45(1-169) cells were less efficient in splicing of non-canonical substrates as compared to WT. The expression of Prp45(119-379) in prp45(1-169) cells restored Prp22 partition in the Cwc2-pulldowns and the splicing defects of prp45(1-169) strain. Data on splicing phenotypes of prp22 and prp45 alleles suggest that Prp45 co-modulates Prp22 in the spliceosome, contributing to splicing efficiency and proper stringency of splice site choice. This work was supported by the Czech Ministry of Education, Youth and Sports grants MSM0021620858 and LC07032. [ABSTRACT FROM AUTHOR]

Human transcription co-regulator SNW1/SKIP is implicated in the regulation of both transcription elongation and alternative splicing. Prp45, the SNW/SKIP ortholog in yeast, is assumed to be essential for pre-mRNA processing. Here, we characterize prp45(1-169), a temperature sensitive allele of PRP45, which at permissive temperature elicits cell division defects and hypersensitivity to microtubule inhibitors. Using a synthetic lethality screen, we found that prp45(1-169) genetically interacts with alleles of NTC members SYF1, CLF1/SYF3, NTC20, and CEF1, and 2nd step splicing factors SLU7, PRP17, PRP18, and PRP22. Cwc2-associated spliceosomal complexes purified from prp45(1-169) cells showed decreased stoichiometry of Prp22, suggesting its deranged interaction with the spliceosome. In vivo splicing assays in prp45(1-169) cells revealed that branch point mutants accumulated more pre-mRNA whereas 5' and 3' splice site mutants showed elevated levels of lariat-exon intermediate as compared to wild-type cells. Splicing of canonical intron was unimpeded. Notably, the expression of Prp45(119-379) in prp45(1-169) cells restored Prp22 partition in the Cwc2-pulldowns and rescued temperature sensitivity and splicing phenotype of prp45(1-169) strain. Our data suggest that Prp45 contributes, in part through its interaction with the 2nd step-proofreading helicase Prp22, to splicing efficiency of substrates non-conforming to the consensus., (2008 Wiley-Liss, Inc.)