Wellcome Sanger Institute, Wellcome Genome Campus, Cambridge, CB10 1SA, UK.
Wellcome Trust Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QN, UK.
Genome Biol. 2021 Jan 22;22(1):43. doi: 10.1186/s13059-020-02246-2.
Microexons, exons that are ≤ 30 nucleotides, are a highly conserved and dynamically regulated set of cassette exons. They have key roles in nervous system development and function, as evidenced by recent results demonstrating the impact of microexons on behaviour and cognition. However, microexons are often overlooked due to the difficulty of detecting them using standard RNA-seq aligners.
Here, we present MicroExonator, a novel pipeline for reproducible de novo discovery and quantification of microexons. We process 289 RNA-seq datasets from eighteen mouse tissues corresponding to nine embryonic and postnatal stages, providing the most comprehensive survey of microexons available for mice. We detect 2984 microexons, 332 of which are differentially spliced throughout mouse embryonic brain development, including 29 that are not present in mouse transcript annotation databases. Unsupervised clustering of microexons based on their inclusion patterns segregates brain tissues by developmental time, and further analysis suggests a key function for microexons in axon growth and synapse formation. Finally, we analyse single-cell RNA-seq data from the mouse visual cortex, and for the first time, we report differential inclusion between neuronal subpopulations, suggesting that some microexons could be cell type-specific.
MicroExonator facilitates the investigation of microexons in transcriptome studies, particularly when analysing large volumes of data. As a proof of principle, we use MicroExonator to analyse a large collection of both mouse bulk and single-cell RNA-seq datasets. The analyses enabled the discovery of previously uncharacterized microexons, and our study provides a comprehensive microexon inclusion catalogue during mouse development.
Microexons（长度≤30 个核苷酸的外显子）是一组高度保守且动态调节的盒式外显子。它们在神经系统发育和功能中起着关键作用，最近的研究结果表明 microexons 对行为和认知有影响。然而，由于使用标准 RNA-seq 比对器检测 microexons 具有一定难度，因此常常被忽视。
在这里，我们提出了 MicroExonator，这是一种用于可重复从头发现和定量 microexons 的新方法。我们处理了来自 18 种小鼠组织的 289 个 RNA-seq 数据集，涵盖了 9 个胚胎和出生后阶段，这是对小鼠 microexons 最全面的调查。我们检测到了 2984 个 microexons，其中 332 个在小鼠胚胎大脑发育过程中存在差异剪接，包括 29 个不在小鼠转录本注释数据库中的 microexons。基于包含模式的无监督聚类将大脑组织按发育时间进行分类，进一步的分析表明 microexons 在轴突生长和突触形成中具有关键作用。最后，我们分析了来自小鼠视觉皮层的单细胞 RNA-seq 数据，首次报告了神经元亚群之间的差异包含，这表明一些 microexons 可能具有细胞类型特异性。
MicroExonator 促进了转录组研究中对 microexons 的研究，特别是在分析大量数据时。作为一个原理验证，我们使用 MicroExonator 分析了大量小鼠批量和单细胞 RNA-seq 数据集。这些分析使我们能够发现以前未被表征的 microexons，我们的研究提供了小鼠发育过程中全面的 microexon 包含目录。
