title: High-throughput sequence analysis of Ciona intestinalis SL trans-spliced mRNAs: alternative expression modes and gene function correlates.
abstract: Pre-mRNA 5' spliced-leader (SL) trans-splicing occurs in some metazoan groups but not in others. Genome-wide characterization of the trans-spliced mRNA subpopulation has not yet been reported for any metazoan. We carried out a high-throughput analysis of the SL trans-spliced mRNA population of the ascidian tunicate Ciona intestinalis by 454 Life Sciences (Roche) pyrosequencing of SL-PCR-amplified random-primed reverse transcripts of tailbud embryo RNA. We obtained approximately 250,000 high-quality reads corresponding to 8790 genes, approximately 58% of the Ciona total gene number. The great depth of this data revealed new aspects of trans-splicing, including the existence of a significant class of "infrequently trans-spliced" genes, accounting for approximately 28% of represented genes, that generate largely non-trans-spliced mRNAs, but also produce trans-spliced mRNAs, in part through alternative promoter use. Thus, the conventional qualitative dichotomy of trans-spliced versus non-trans-spliced genes should be supplanted by a more accurate quantitative view recognizing frequently and infrequently trans-spliced gene categories. Our data include reads representing approximately 80% of Ciona frequently trans-spliced genes. Our analysis also revealed significant use of closely spaced alternative trans-splice acceptor sites which further underscores the mechanistic similarity of cis- and trans-splicing and indicates that the prevalence of +/-3-nt alternative splicing events at tandem acceptor sites, NAGNAG, is driven by spliceosomal mechanisms, and not nonsense-mediated decay, or selection at the protein level. The breadth of gene representation data enabled us to find new correlations between trans-splicing status and gene function, namely the overrepresentation in the frequently trans-spliced gene class of genes associated with plasma/endomembrane system, Ca(2+) homeostasis, and actin cytoskeleton.
authors.affiliation: Department of Neurology & Neurosurgery, McGill University, Montreal Neurological Institute, Montréal, Québec H3A 2B4, Canada.
authors.forename: Jun
authors.initials: J
authors.lastname: Matsumoto
authors.forename: Ken
authors.initials: K
authors.lastname: Dewar
authors.forename: Jessica
authors.initials: J
authors.lastname: Wasserscheid
authors.forename: Graham B
authors.initials: GB
authors.lastname: Wiley
authors.forename: Simone L
authors.initials: SL
authors.lastname: Macmil
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Roe
authors.forename: Robert W
authors.initials: RW
authors.lastname: Zeller
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Satou
authors.forename: Kenneth E M
authors.initials: KE
authors.lastname: Hastings
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D002874:Chromosome Mapping; D002938:Ciona intestinalis / Q000196:embryology / Q000235:genetics / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D008957:Models, Genetic; D016133:Polymerase Chain Reaction; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016053:RNA, Protozoan / Q000235:genetics / Q000378:metabolism; D020038:RNA, Spliced Leader / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA / Q000379:methods; D020040:Trans-Splicing
chemical_list: D018076:DNA, Complementary; D015800:Protozoan Proteins; D012322:RNA Precursors; D012333:RNA, Messenger; D016053:RNA, Protozoan; D020038:RNA, Spliced Leader