Importance of being non-triplet in alternative splicing
Alternative mRNA splicing can be used by a cell to regulate gene expression and to generate transcriptomic diversity. Most cases of alternative splicing studied to date are triplet in nature, meaning that both isoforms retain the same translational reading frame. Indeed, non-triplet alternative splicing is sometimes considered evidence of splicing errors or splicing noise. Nevertheless, some examples of functionally important non-triplet alternative splicing exist. We set out to determine the global prevalence, regulation, and function of non-triplet alternative splicing in vivo in C. elegans. We performed RNA-Seq on NMD-deficient mutants and bioinformatically categorized the molecular consequences of non-triplet alternative splicing into three categories: NMD-sensitive isoforms, alternative C-terminal length isoforms, and dual-coding isoforms. We identify hundreds of non-triplet alternative splicing events across these three categories. Genetic and molecular analysis reveals cases of developmental regulation, splicing factor autoregulation, cell-specific splicing patterns, and multiple non-triplet alternative splicing events with physiologically important functions. Together these experiments reveal the importance of non-triplet alternative splicing, a large but underappreciated class of alternative splicing events, for regulating gene expression and generating protein-coding diversity Overall design: L4 staged animals from wild-type and mutant conditions. RNA extracted, and selected for polyA+ mRNA prior to library generation and short-read sequencing.