Cataloging the potential functional diversity of Cacna1e splice variants using long-read sequencing

Abstract

Background: The degree to which alternative RNA splicing influences the function and structure of voltage gated calcium channel (VGCC) splice variants is poorly understood. Here we used long-read RNA-sequencing to catalog rat Cacna1e (Cav2.3) splice variants, and computationally prioritize which are likely to impact channel function.

Result: We sequenced Cacna1e transcripts from rat thalamus using Oxford Nanopore sequencing yielding the structure of 2,110 Cacna1e splice variants. Of these, up to 154 had the potential encode for a functional channel based on predicted amino acid sequences. Our analysis revealed a total of 31 cassette splicing events (in various combinations) potentially affecting channel function, with three cassette exons appreciably expressed and conserved.

Conclusion: Our work both provides the first long-read sequencing of Cacna1e and the first computational evaluation of Cacna1e splice variants for future follow-up. This overall strategy to provide the field with prioritized transcripts will improve our understanding of Cacna1e function, its role in disease pathophysiology, and serve as a general approach to evaluate splice variant function across multiple ion channel types.

Keywords: Alternative splicing; Calcium channels; Nanopore sequencing; Noisy splicing model.