Prp45 affects Prp22 partition in spliceosomal complexes and splicing efficiency of non-consensus substrates

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. J. Cell. Biochem. 106: 139–151, 2009. © 2008 Wiley‐Liss, Inc.