Identification of Splice Variants and Isoforms in Transcriptomics and Proteomics

Alternative splicing is pivotal to the regulation of gene expression and protein diversity in eukaryotic cells. The detection of alternative splicing events requires specific omics technologies. Although short-read RNA sequencing has successfully supported a plethora of investigations on alternative splicing, the emerging technologies of long-read RNA sequencing and top-down mass spectrometry open new opportunities to identify alternative splicing and protein isoforms with less ambiguity. Here, we summarize improvements in short-read RNA sequencing for alternative splicing analysis, including percent splicing index estimation and differential analysis. We also review the computational methods used in top-down proteomics analysis regarding proteoform identification, including the construction of databases of protein isoforms and statistical analyses of search results. While many improvements in sequencing and computational methods will result from emerging technologies, there should be future endeavors to increase the effectiveness, integration, and proteome coverage of alternative splicing events.

Automated summary

Alternative splicing plays a crucial role in regulating gene expression and protein diversity in eukaryotic cells. Traditional short-read RNA sequencing has been widely used for studying alternative splicing, but the emergence of long-read RNA sequencing and top-down mass spectrometry offers new opportunities to identify alternative splicing events and protein isoforms with higher accuracy. This review summarizes the recent improvements in short-read RNA sequencing and computational methods for alternative splicing analysis, as well as the approaches used in top-down proteomics to identify protein isoforms. However, future efforts are needed to further enhance the effectiveness, integration, and proteome coverage of alternative splicing events.