Improving the Annotation of <i>Arabidopsis lyrata</i> Using RNA-Seq Data

<div><p>Gene model annotations are important community resources that ensure comparability and reproducibility of analyses and are typically the first step for functional annotation of genomic regions. Without up-to-date genome annotations, genome sequences cannot be used to maximum advantage. It is therefore essential to regularly update gene annotations by integrating the latest information to guarantee that reference annotations can remain a common basis for various types of analyses. Here, we report an improvement of the <i>Arabidopsis lyrata</i> gene annotation using extensive RNA-seq data. This new annotation consists of 31,132 protein coding gene models in addition to 2,089 genes with high similarity to transposable elements. Overall, ~87% of the gene models are corroborated by evidence of expression and 2,235 of these models feature multiple transcripts. Our updated gene annotation corrects hundreds of incorrectly split or merged gene models in the original annotation, and as a result the identification of alternative splicing events and differential isoform usage are vastly improved.</p></div

Heat stress induces alternative splicing events.

<p><b>(A)</b> Examples of differentially expressed isoforms in response to heat stress in <i>A</i>. <i>lyrata</i>. AL3G42820 expresses a second isoform that lacks the middle exon in heat-treated samples (HS). Transcripts from wild-type (WT) and recovery (REC) samples contain all three exons. AL2G15640 retains an intron in response to heat stress (HS) while wild-type (WT) and recovery (REC) samples show partial intron splicing. <b>(B)</b> Number of differential splicing events, including alternative 5’ and 3’ splice sites, mutually exclusive exons, intron retention, and exon skipping events identified with MATs based on version-1 and version-2 annotations.</p

Examples of version-1 gene models split and merged in <i>A</i>. <i>lyrata</i> gene annotation version-2.

<p><b>(A)</b> Example of a gene model that was split into two gene models in version-2. Reverse transcription-PCR could not confirm the connection of both. <b>(B)</b> Example of version-1 gene models that were merged during the annotation update. Reverse transcription-PCR confirmed presence of a transcript bridging the two version-1 genes.</p

Updating the gene model annotation of <i>A</i>. <i>lyrata</i>.

<p><b>(A)</b> Left, version-2 gene models predicted by Augustus [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137391#pone.0137391.ref028" target="_blank">28</a>]. Number of gene models overlapping with version-1 (yellow), genes predicted with Cufflinks (red), and genes with expression evidence (blue). Right, gene models of the version-1 annotation. Number of models without overlap to version-2 models (yellow), without orthologs in five other Brassicaceae (red), and without significant expression evidence (blue). <b>(B)</b> Correlation of the lengths of <i>A</i>. <i>lyrata</i> gene models with the length of their orthologous gene models in <i>A</i>. <i>thaliana</i>. Left, <i>A</i>. <i>lyrata</i> version-1 gene models. Correlations using version-1 gene models (left), version-2 gene models before (middle) and after (right) the homology-based correction of gene models. <b>(C)</b> Length distribution of gene models including genes that were removed or newly added in the version-2.</p

Comparing the <i>A</i>. <i>lyrata</i> gene annotation version-2 with the annotations of five other Brassicaceae.

<p><b>(A)</b> Orthologous gene models shared between <i>A</i>. <i>lyrata</i> (version-2), <i>A</i>. <i>thaliana</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137391#pone.0137391.ref029" target="_blank">29</a>], <i>A</i>. <i>alpina</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137391#pone.0137391.ref005" target="_blank">5</a>], <i>B</i>. <i>rapa</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137391#pone.0137391.ref010" target="_blank">10</a>], <i>C</i>. <i>rubella</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137391#pone.0137391.ref004" target="_blank">4</a>] and <i>S</i>. <i>parvula</i> [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0137391#pone.0137391.ref011" target="_blank">11</a>]. <b>(B)</b> Gene, Protein and UTR length distributions of above-mentioned species including the new and old <i>A</i>. <i>lyrata</i> annotations. UTR distribution is only shown for <i>A</i>. <i>lyrata</i> and <i>A</i>. <i>thaliana</i> because of poor UTR annotation in some of the other species.</p

Comparison of version-1 and version-2 annotations.

<p>Comparison of version-1 and version-2 annotations.</p