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

Figures

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  Fig. 1. The majority of detected splice variants did not contain an open reading frame (ORF) of at least 2,000 amino acids. ORFs were defined as containing a start codon, but not necessarily a stop codon. The X-axis shows the distribution of log10(ORF) lengths. The dashed blue line indicates the minimum size expected for a functional channel

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  Fig. 2. Highly expressed splice variants account for 12.5% of Cacna1e gene expression. A) Four splice variants have an expression ratio of less than 2. Each splice variant’s expression ratio is calculated by the most expressed splice variant’s expression divided by the splice variant’s expression. The dotted Line indicates where the expression ratio would equal 2. B) Top 10 most expressed Cacna1e splice variants pooled across all samples. The X-axis shows the rank of the splice variant based on expression. The Y-axis shows the proportion of the splice variant relative to Cacna1e’s total expression. C) Contribution of all 2,110 Cacna1e splice variants to the gene’s overall expression. The X-axis shows the rank of the splice variant based on expression. The Y-axis shows the proportion of the splice variant relative to Cacna1e’s total expression. Of the 2,110 variants, the 154 (cyan) have 4 annotated pore-forming domains and account for 61% of Cacna1e’s expression. We did not predict the remaining splice variants to encode for all four pore-forming domains (red) D) Cacna1e splice variant expression level ranks in each sample. X-axis show expression rank for splice variant summed across all…

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  Fig. 3. Structure of top 4 most abundantly expressed splice variants. Red indicates a pore-forming domain annotation from CDD (Ion_trans – pfam00520). Blue and purple are calmodulin-binding domain annotation from CDD (GPHH – pfam16905 and Ca_chan_IQ – pfam08763, respectively). Along the side are the number of reads for each splice variant, and long the bottom are the exon numbers. A similar visualization for all 154 splice variants that contained 4 pore-forming domains is provided in Additional File 7

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  Fig. 4. Location of most common splicing events on Cacna1e structure. Here we show where each of these six cassette exons would likely impact the channel, and the percentage of reads that do not contain the exon. Figure created with Biorender.com

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  Fig. 5. Most Cacna1e exons are well-conserved throughout the vertebrate phylogeny. The UCSC genome browser provides base-specific PhyloP scores. For each exon of Cacna1e, we annotated the average PhyloP score. Our analysis focused on prioritized splice variants with cassette exons 10, 19 and 45 (asterisk)

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  Fig. 6. Distribution of MaxEntScores for predicted donor and acceptor sites across 154 putative Cacna1e splice variants. A Donor scores for sets 9-mers found across the Cacna1e splice variants. B Acceptor scores for sets of 23-mer found across the 154 putative Cacna1e splice variants