Revisiting the identification of canonical splice isoforms through integration of functional genomics and proteomics evidence

Canonical isoforms in different databases have been defined as the most prevalent, most conserved, most expressed, longest, or the one with the clearest description of domains or posttranslational modifications. In this article, we revisit these definitions of canonical isoforms based on functional genomics and proteomics evidence, focusing on mouse data. We report a novel functional relationship network‐based approach for identifying the highest connected isoforms (HCIs). We show that 46% of these HCIs are not the longest transcripts. In addition, this approach revealed many genes that have more than one highly connected isoforms. Averaged across 175 RNA‐seq datasets covering diverse tissues and conditions, 65% of the HCIs show higher expression levels than nonhighest connected isoforms at the transcript level. At the protein level, these HCIs highly overlap with the expressed splice variants, based on proteomic data from eight different normal tissues. These results suggest that a more confident definition of canonical isoforms can be made through integration of multiple lines of evidence, including HCIs defined by biological processes and pathways, expression prevalence at the transcript level, and relative or absolute abundance at the protein level. This integrative proteogenomics approach can successfully identify principal isoforms that are responsible for the canonical functions of genes.