P.293Novel splicing events in skeletal muscle revealed by RNA sequencing

Abstract

Most human genes consist of more than one coding exon and therefore undergo splicing to remove the intronic areas. Alternative splicing gives rise to alternative isoforms. Many alternative splicing events (ASEs) have been shown to be developmental stage and/or tissue specific. Hence a comprehensive understanding of ASEs is of great importance for understanding diseases showing tissue and/or developmental stage specificity, but also for discovery of aberrant splicing. RNA sequencing (RNA-seq) enables detailed studies of the transcriptome and also reveals the detailed ASE landscape in the cells at the time of sampling. As such RNA-seq is an invaluable tool for splicing detection and direct discovery of aberrant splicing when samples of relevant tissues are available. Using RNA-seq we have studied ASEs in 42 samples from muscles of the lower leg from patients without muscle disorders. A large amount of unexpected splicing events were detected. This also included hitherto undescribed exons even in identified genes causing neuromuscular disorders. The findings were validated by other sequencing methods (PCR and the long-read technology PacBio) and also by comparison with public RNA-seq data from the ENCODE project. Our results are important for understanding of the complex splicing observed in skeletal muscle. Further, for correct interpretation and annotation of mutations affecting splicing in these genes, a comprehensive understanding of the ASEs is crucial.