Bioinformatic identification of altered splicing motifs resulting from the Alu insertion in exon 9 of the RP gene MAK

Abstract

Purpose: An Alu insertion into exon 9 of male germ cell-associated kinase (MAK) causes the exclusion of that exon and the retina-specific exon 12, resulting in retinitis pigmentosa. We hypothesized that insertion of the Alu element triggers exclusion of exon 9 by altering an existing exonic splicing enhancer (ESE) or by introducing an exonic splicing silencer (ESS).

Methods: Putative ESE and ESS motifs were identified using Human Splicing Finder (v2.4.1) in MAK exon 9 with and without the Alu insertion. Scores for 5’ and 3’ splice sites of MAK exons were calculated using Analyzer Splice Tool. The ExonScan tool was used to computationally predict exons from the human reference sequence (GRCh37/hg19) between the initial and terminal exons of MAK.

Results: Insertion of the Alu in exon 9 introduces a complex set of splicing motifs; however, the run of adenosines near the 3’ end of the Alu generates a strong signal for exonic splice silencing. Of note, the retina-specific exon 12 is computationally predicted by ExonScan, with splice site scores that are similar to the constitutively expressed exons.

Conclusions: Exclusion of exon 9 may result from the introduction of exonic splicing silencers within the Alu. Exon 12 can be predicted by an existing algorithm indicating that the exon shares features with other known exons. We are analyzing RNA-Seq data from retina and other tissues to characterize retina-specific splicing regulatory motifs shared by MAK exon 12 and other retina-specific exons.

Keywords: 533 gene/expression • 688 retina • 473 computational modeling