Isoform Age - Splice Isoform Profiling Using Long-Read Technologies

Alternative splicing (AS) of RNA is a key mechanism that results in the expression of multiple transcript isoforms from single genes and leads to an increase in the complexity of both the transcriptome and proteome. Regulation of AS is critical for the correct functioning of many biological pathways, while disruption of AS can be directly pathogenic in diseases such as cancer or cause risk for complex disorders. Current short-read sequencing technologies achieve high read depth but are limited in their ability to resolve complex isoforms. In this review we examine how long-read sequencing (LRS) technologies can address this challenge by covering the entire RNA sequence in a single read and thereby distinguish isoform changes that could impact RNA regulation or protein function. Coupling LRS with technologies such as single cell sequencing, targeted sequencing and spatial transcriptomics is producing a rapidly expanding suite of technological approaches to profile alternative splicing at the isoform level with unprecedented detail. In addition, integrating LRS with genotype now allows the impact of genetic variation on isoform expression to be determined. Recent results demonstrate the potential of these techniques to elucidate the landscape of splicing, including in tissues such as the brain where AS is particularly prevalent. Finally, we also discuss how AS can impact protein function, potentially leading to novel therapeutic targets for a range of diseases.

Automated summary

Alternative splicing of RNA is a key mechanism leading to the expression of multiple transcript isoforms from a single gene, increasing the complexity of the transcriptome and proteome. Long-read sequencing technologies can address the limitations of short-read sequencing in resolving complex isoforms and distinguish changes that may impact RNA regulation or protein function. Integration of long-read sequencing with other technologies allows for detailed profiling of alternative splicing at the isoform level and determination of the impact of genetic variation on isoform expression.