title: Genomic mRNA profiling reveals compensatory mechanisms for the requirement of the essential splicing factor U2AF.
abstract: 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.
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO 80309, USA.
authors.forename: Vinod
authors.initials: V
authors.lastname: Sridharan
authors.forename: Joseph
authors.initials: J
authors.lastname: Heimiller
authors.forename: Ravinder
authors.initials: R
authors.lastname: Singh
mesh_terms: D001483:Base Sequence; D020869:Gene Expression Profiling; D016681:Genome, Fungal; D007438:Introns; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D022821:RNA Splice Sites; D012326:RNA Splicing; D012331:RNA, Fungal / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism*; D012568:Schizosaccharomyces / Q000235:genetics* / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics* / Q000378:metabolism*; D017381:Spliceosomes / Q000235:genetics / Q000378:metabolism; D000072668:Splicing Factor U2AF
chemical_list: D009687:Nuclear Proteins; D022821:RNA Splice Sites; D012331:RNA, Fungal; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D029702:Schizosaccharomyces pombe Proteins; D000072668:Splicing Factor U2AF