Quantitative RNAseq Meta Analysis Of Alternative Exon Usage In C. elegans

Abstract

Almost twenty years after the completion of the C. elegans genome sequence, gene structure annotation is still an ongoing process with new evidence for gene variants still being regularly uncovered by additional in-depth transcriptome studies. While alternative splice forms can allow a single gene to encode several functional isoforms the question of how much spurious splicing is tolerated is still heavily debated.\n\nHere we gathered a compendium of 1,682 publicly available C. elegans RNA-seq datasets to increase the dynamic range of detection of RNA isoforms and obtained robust measurements of the relative abundance of each splicing event. While most of the splicing reads come from reproducibly detected splicing events, a large fraction of purported junctions are only supported by a very low number of reads. We devised an automated curation method that takes into account the expression level of each gene to discriminate robust splicing events from potential biological noise. We found that rarely used splice sites disproportionately come from highly expressed genes and are significantly less conserved in other nematode genomes than splice sites with a higher usage frequency.\n\nOur increased detection power confirmed trans-splicing for at least 84% of C. elegans protein coding genes. The genes for which trans-splicing was not observed are overwhelmingly low expression genes, suggesting that the mechanism is pervasive but not full captured by organism-wide RNA-Seq.\n\nWe generated annotated gene models including quantitative exon usage information for the entire C. elegans genome. This allows users to visualize at a glance the relative expression of each isoform for their gene of interest.