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An emerging model organism Caenorhabditis elegans for alternative pre‐mRNA processing in vivo

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A nematode Caenorhabditis elegans is an intron‐rich organism and up to 25% of its pre‐mRNAs are estimated to be alternatively processed. Its compact genomic organization enables construction of fluorescence splicing reporters with intact genomic sequences and visualization of alternative processing patterns of interest in the transparent living animals with single‐cell resolution. Genetic analysis with the reporter worms facilitated identification of trans‐acting factors and cis‐acting elements, which are highly conserved in mammals. Analysis of unspliced and partially spliced pre‐mRNAs in vivo raised models for alternative splicing regulation relying on specific order of intron excision. RNA‐seq analysis of splicing factor mutants and CLIP‐seq analysis of the factors allow global search for target genes in the whole animal. An mRNA surveillance system is not essential for its viability or fertility, allowing analysis of unproductively spliced noncoding mRNAs. These features offer C. elegans as an ideal model organism for elucidating alternative pre‐mRNA processing mechanisms in vivo. Examples of isoform‐specific functions of alternatively processed genes are summarized. WIREs RNA 2017, 8:e1428. doi: 10.1002/wrna.1428 This article is categorized under: RNA Processing > Splicing Regulation/Alternative Splicing RNA in Disease and Development > RNA in Development RNA Methods > RNA Analyses in Cells
Fluorescence splicing reporters enable visualization of tissue‐specific alternative splicing patterns in living worms. (a) A trio of unc‐32 exon 4 splicing reporter minigenes. A genomic fragment of interest is cloned upstream of fluorescent protein cDNA cassettes and artificial termination codons and/or frameshifts are introduced so that expression of each fluorescent protein indicates a specific alternative splicing pattern. (b) A three‐color unc‐32 exon 4 reporter worm. Expression of Venus (green), mRFP1 (red), and ECFP (blue) indicates selection of exon 4a, 4b, and 4c, respectively.
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Germline‐specific selection of the upstream spliced sites from adjacent alternative 3′ splice sites separated by ≤ 18 nt. See text for details.
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Three splicing factors A, B and C with partially overlapping expression patterns regulate overlapping exon networks (red, green, and blue circles). The repertoire of splicing factors in each neuronal class together with the combination of cis‐elements in each gene creates the neuron‐class‐specific isoform repertoires.
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Three models for mutually exclusive selection of alternative exons. (a) Muscle‐specific selection of egl‐15 exon 5A. The RBFOX family and SUP‐12 cooperatively bind to the intron upstream of the upstream exon and represses the upstream exon in the muscles. The exons are mutually exclusive because the intervening sequence between them is too short for splicing machinery. (b) Muscle‐specific developmental switch from exon 9 to exon 10 in the let‐2 gene. Downstream introns are removed first. ASD‐2 binds to the intron downstream of the downstream exon to promote splicing between exon 10 and exon 11. The exons are mutually exclusive because the intervening sequence between them is too short for splicing machinery. (c) Neuron‐specific selection of unc‐32 exon 7a. UNC‐75 represses exon 7b and the RBFOX family promotes splicing between exon 7a and exon 8. The exons are mutually exclusive because the first splicing removes one of the exons.
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RNA Methods > RNA Analyses in Cells
RNA in Disease and Development > RNA in Development
RNA Processing > Splicing Regulation/Alternative Splicing

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