High-throughput sequence analysis of Ciona intestinalis SL trans-spliced mRNAs: Alternative expression modes and gene function correlates
- Jun Matsumoto1,
- Ken Dewar2,
- Jessica Wasserscheid2,
- Graham B. Wiley3,
- Simone L. Macmil3,
- Bruce A. Roe3,
- Robert W. Zeller4,
- Yutaka Satou5 and
- Kenneth E.M. Hastings1,6
- 1 Montreal Neurological Institute and Departments of Neurology & Neurosurgery and Biology, McGill University, Montréal, Québec H3A 2B4, Canada;
- 2 McGill University and Génome Québec Innovation Centre, Departments of Human Genetics and Experimental Medicine, McGill University, Montréal, Québec H3A 1A1, Canada;
- 3 Advanced Center for Genome Technology, Stephenson Research and Technology Center, University of Oklahoma, Norman, Oklahoma 73019-0370, USA;
- 4 Department of Biology, San Diego State University, San Diego, California 92182, USA;
- 5 Department of Zoology, Graduate School of Sciences, Kyoto University, Kyoto 606-8501, Japan
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 ∼250,000 high-quality reads corresponding to 8790 genes, ∼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 ∼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 ∼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, Ca2+ homeostasis, and actin cytoskeleton.
Footnotes
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↵6 Corresponding author.
E-mail ken.hastings{at}mcgill.ca; fax (514)398-1509.
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[Supplemental material is available online at http://www.genome.org. The sequence data from this study have been submitted to the NCBI Short Read Archive (http://www.ncbi.nlm.nih.gov/Traces/sra/sra.cgi) under accession no. SRX006190.]
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Article published online before print. Article and publication date are at http://www.genome.org/cgi/doi/10.1101/gr.100271.109.
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- Received September 4, 2009.
- Accepted February 18, 2010.
- Copyright © 2010 by Cold Spring Harbor Laboratory Press