Department of Biohealth Informatics, School of Informatics and Computing, Indiana University Purdue University, 719 Indiana Ave Ste 319, Walker Plaza Building, Indianapolis, Indiana, 46202, USA.
Department of Biological Sciences, University of Delaware, Newark, DE, 19716, USA.
Sci Rep. 2017 Sep 14;7(1):11572. doi: 10.1038/s41598-017-10615-4.
Lens development involves a complex and highly orchestrated regulatory program. Here, we investigate the transcriptomic alterations and splicing events during mouse lens formation using RNA-seq data from multiple developmental stages, and construct a molecular portrait of known and novel transcripts. We show that the extent of novelty of expressed transcripts decreases significantly in post-natal lens compared to embryonic stages. Characterization of novel transcripts into partially novel transcripts (PNTs) and completely novel transcripts (CNTs) (novelty score ≥ 70%) revealed that the PNTs are both highly conserved across vertebrates and highly expressed across multiple stages. Functional analysis of PNTs revealed their widespread role in lens developmental processes while hundreds of CNTs were found to be widely expressed and predicted to encode for proteins. We verified the expression of four CNTs across stages. Examination of splice isoforms revealed skipped exon and retained intron to be the most abundant alternative splicing events during lens development. We validated by RT-PCR and Sanger sequencing, the predicted splice isoforms of several genes Banf1, Cdk4, Cryaa, Eif4g2, Pax6, and Rbm5. Finally, we present a splicing browser Eye Splicer ( http://www.iupui.edu/~sysbio/eye-splicer/ ), to facilitate exploration of developmentally altered splicing events and to improve understanding of post-transcriptional regulatory networks during mouse lens development.
晶状体发育涉及一个复杂而高度协调的调控程序。在这里，我们使用来自多个发育阶段的 RNA-seq 数据研究了小鼠晶状体形成过程中的转录组变化和剪接事件，并构建了已知和新转录本的分子图谱。我们表明，与胚胎阶段相比，出生后晶状体中表达转录本的新颖性程度显著降低。将新转录本分为部分新转录本（PNTs）和完全新转录本（CNTs）（新颖性评分≥70%）表明，PNTs在脊椎动物中高度保守，在多个阶段高度表达。对 PNTs 的功能分析揭示了它们在晶状体发育过程中的广泛作用，而数百个 CNTs 被发现广泛表达，并预测编码蛋白质。我们在多个阶段验证了四个 CNTs 的表达。对剪接异构体的检查表明，在晶状体发育过程中，外显子跳跃和内含子保留是最丰富的可变剪接事件。我们通过 RT-PCR 和 Sanger 测序验证了 Banf1、Cdk4、Cryaa、Eif4g2、Pax6 和 Rbm5 等几个基因的预测剪接异构体。最后，我们展示了一个剪接浏览器 Eye Splicer（http://www.iupui.edu/~sysbio/eye-splicer/），以促进对发育改变的剪接事件的探索，并提高对小鼠晶状体发育过程中转录后调控网络的理解。
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