Recent studies showed that RNA binding motif protein 20 (RBM20) is involved in the regulation of the alternative splicing of several genes both in rats and humans. Mutations in RBM20 have been shown to cause human dilated cardiomyopathy. RBM20 RNA-binding site maps in the intronic sequence of the target genes in close proximity to differentially spliced exons and an overrepresentation of RBM20 RNA-binding sites in these regions was noticed. However, a detailed rule is missing. The aim of our ongoing investigation is to find exons whose alternative splicing is affected by RBM20, and then to define sequence patterns of the RBM20 regulated exons. RNA sequencing data of human and rat cardiomyocytes made avaible by Guo were used. Rat samples showed 3 different conditions: 3 wild type (RBM20+/+), 3 heterozygosity (RBM20+/-) and 3 deletion (RBM20-/-) samples. Human samples showed 2 different conditions: 2 control individuals (RBM20+/+) and 1 RBM20 mutated individual (RBM20-/-). The rat RGSC3.4 and human GRCh37 reference genomes were used to map the rat and the human samples, respectively, and the expression of each individual exon has been measured. An analysis was carried out to identify differentially spliced (DS) exons between RBM20 positive and negative samples. The differential analysis between RBM20+/+ and RBM20-/- samples showed 96 DS exons (p. adj < 0.05) belonging to 33 different genes, and 1816 DS exons belonging to 657 different genes, in rat and humans, respectively. We studied the 400 nt preceding and following the sequence of the affected exons. The exon following sequence showed a greater number of RBM20 RNA-binding sites than the exon preceding sequence, and the opposite was observed for the analyzed control sequences. The enrichment analysis of novel motifs showed 2 motifs more common in case than in control sequences. We are now evaluating the statistical significance of these observations and we are testing novel hypothesis for a more accurate definition of the RBM20 RNA-binding site.
Searching for DNA motifs that affect splicing of exons regulated by RBM20
Dal Molin, Anna;LORENZI, Pamela;ROMANELLI, Maria;MALERBA, Giovanni
2015-01-01
Abstract
Recent studies showed that RNA binding motif protein 20 (RBM20) is involved in the regulation of the alternative splicing of several genes both in rats and humans. Mutations in RBM20 have been shown to cause human dilated cardiomyopathy. RBM20 RNA-binding site maps in the intronic sequence of the target genes in close proximity to differentially spliced exons and an overrepresentation of RBM20 RNA-binding sites in these regions was noticed. However, a detailed rule is missing. The aim of our ongoing investigation is to find exons whose alternative splicing is affected by RBM20, and then to define sequence patterns of the RBM20 regulated exons. RNA sequencing data of human and rat cardiomyocytes made avaible by Guo were used. Rat samples showed 3 different conditions: 3 wild type (RBM20+/+), 3 heterozygosity (RBM20+/-) and 3 deletion (RBM20-/-) samples. Human samples showed 2 different conditions: 2 control individuals (RBM20+/+) and 1 RBM20 mutated individual (RBM20-/-). The rat RGSC3.4 and human GRCh37 reference genomes were used to map the rat and the human samples, respectively, and the expression of each individual exon has been measured. An analysis was carried out to identify differentially spliced (DS) exons between RBM20 positive and negative samples. The differential analysis between RBM20+/+ and RBM20-/- samples showed 96 DS exons (p. adj < 0.05) belonging to 33 different genes, and 1816 DS exons belonging to 657 different genes, in rat and humans, respectively. We studied the 400 nt preceding and following the sequence of the affected exons. The exon following sequence showed a greater number of RBM20 RNA-binding sites than the exon preceding sequence, and the opposite was observed for the analyzed control sequences. The enrichment analysis of novel motifs showed 2 motifs more common in case than in control sequences. We are now evaluating the statistical significance of these observations and we are testing novel hypothesis for a more accurate definition of the RBM20 RNA-binding site.
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