title: The Fission Yeast Pre-mRNA-processing Factor 18 (prp18+) Has Intron-specific Splicing Functions with Links to G1-S Cell Cycle Progression.
abstract: The fission yeast genome, which contains numerous short introns, is an apt model for studies on fungal splicing mechanisms and splicing by intron definition. Here we perform a domain analysis of the evolutionarily conserved Schizosaccharomyces pombe pre-mRNA-processing factor, SpPrp18. Our mutational and biophysical analyses of the C-terminal α-helical bundle reveal critical roles for the conserved region as well as helix five. We generate a novel conditional missense mutant, spprp18-5 To assess the role of SpPrp18, we performed global splicing analyses on cells depleted of prp18+and the conditional spprp18-5 mutant, which show widespread but intron-specific defects. In the absence of functional SpPrp18, primer extension analyses on a tfIId+intron 1-containing minitranscript show accumulated pre-mRNA, whereas the lariat intron-exon 2 splicing intermediate was undetectable. These phenotypes also occurred in cells lacking both SpPrp18 and SpDbr1 (lariat debranching enzyme), a genetic background suitable for detection of lariat RNAs. These data indicate a major precatalytic splicing arrest that is corroborated by the genetic interaction between spprp18-5 and spprp2-1, a mutant in the early acting U2AF59 protein. Interestingly, SpPrp18 depletion caused cell cycle arrest before S phase. The compromised splicing of transcripts coding for G1-S regulators, such as Res2, a transcription factor, and Skp1, a regulated proteolysis factor, are shown. The cumulative effects of SpPrp18-dependent intron splicing partly explain the G1arrest upon the loss of SpPrp18. Our study using conditional depletion of spprp18+and the spprp18-5 mutant uncovers an intron-specific splicing function and early spliceosomal interactions and suggests links with cell cycle progression.
authors.affiliation: From the Department of Microbiology and Cell Biology and.
authors.forename: Nagampalli
authors.initials: N
authors.lastname: Vijaykrishna
authors.affiliation: From the Department of Microbiology and Cell Biology and.
authors.forename: Geetha
authors.initials: G
authors.lastname: Melangath
authors.affiliation: From the Department of Microbiology and Cell Biology and.
authors.forename: Rakesh
authors.initials: R
authors.lastname: Kumar
authors.affiliation: From the Department of Microbiology and Cell Biology and.
authors.forename: Piyush
authors.initials: P
authors.lastname: Khandelia
authors.affiliation: From the Department of Microbiology and Cell Biology and.
authors.forename: Pushpinder
authors.initials: P
authors.lastname: Bawa
authors.affiliation: the Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Raghavan
authors.initials: R
authors.lastname: Varadarajan
authors.affiliation: From the Department of Microbiology and Cell Biology and uvr@mcbl.iisc.ernet.in.
authors.forename: Usha
authors.initials: U
authors.lastname: Vijayraghavan
mesh_terms: D002453:Cell Cycle / Q000235:genetics*; D018360:Crystallography, X-Ray; D007438:Introns / Q000235:genetics*; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D011487:Protein Conformation; D012322:RNA Precursors / Q000235:genetics*; D012326:RNA Splicing / Q000235:genetics*; D012331:RNA, Fungal / Q000235:genetics*; D012568:Schizosaccharomyces / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017381:Spliceosomes
chemical_list: D012322:RNA Precursors; D012331:RNA, Fungal; D029702:Schizosaccharomyces pombe Proteins
keywords: RNA splicing; Schizosaccharomyces pombe; SpPrp18; functional genomics; intron-specific splicing; molecular genetics; mutagenesis; splicing microarrays