Alternative splicing coupled to nonsense-mediated decay coordinates downregulation of non-neuronal genes in developing neurons
Alternative splicing coupled to nonsense-mediated decay coordinates downregulation of non-neuronal genes in developing neurons
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DOI：
10.1101/2023.09.04.556212
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发表时间：
2023-09
期刊：
影响因子：
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通讯作者：
A. Zhuravskaya;Karen Yap;F. Hamid;E. Makeyev
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文献类型：
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作者：
A. Zhuravskaya;Karen Yap;F. Hamid;E. Makeyev
前体mRNA选择性剪接（AS）和mRNA质量控制机制无义介导的衰变（NMD）之间的功能耦合可以调节转录本丰度。以前的研究已经确定了几个在发育神经元中进行这种调节的例子。然而，人们对AS-NMD在这方面的系统级影响知之甚少。我们开发了一个R软件包factR 2，它提供了一套全面的AS-NMD分析功能。使用这个工具，我们进行了一个纵向分析的基因表达的多能干细胞进行诱导神经元分化。我们的分析发现了数百个AS-NMD事件，这些事件具有调节基因表达的重要潜力。值得注意的是，这种调节在发育下调基因的特定功能组中显着过度表达。特别强的关联与基因下调检测到替代盒外显子刺激NMD（NS-CE）后，他们纳入成熟的mRNA。通过将生物信息学分析与CRISPR/Cas9基因组编辑和其他实验方法相结合，我们发现由RNA结合蛋白PTBP 1调控的NS-CE抑制了其基因在发育中的神经元中的表达。我们还提供了证据表明，NS-CE活动是暂时协调与NMD独立的基因抑制机制。我们的研究为AS-NMD目标的发现和优先级排序提供了一个可访问的工作流程。它进一步认为，AS-NMD途径通过促进功能相关的非神经元基因的下调在神经元发育中发挥着广泛的作用。
The functional coupling between alternative pre-mRNA splicing (AS) and the mRNA quality control mechanism called nonsense-mediated decay (NMD) can modulate transcript abundance. Previous studies have identified several examples of such a regulation in developing neurons. However, the systems-level effects of AS-NMD in this context are poorly understood. We developed an R package, factR2, which offers a comprehensive suite of AS-NMD analysis functions. Using this tool, we conducted a longitudinal analysis of gene expression in pluripotent stem cells undergoing induced neuronal differentiation. Our analysis uncovered hundreds of AS-NMD events with significant potential to regulate gene expression. Notably, this regulation was significantly overrepresented in specific functional groups of developmentally downregulated genes. Particularly strong association with gene downregulation was detected for alternative cassette exons stimulating NMD (NS-CEs) upon their inclusion into mature mRNA. By combining bioinformatics analyses with CRISPR/Cas9 genome editing and other experimental approaches we show that NS-CEs regulated by the RNA-binding protein PTBP1 dampen the expression of their genes in developing neurons. We also provide evidence that the NS-CE activity is temporally coordinated with NMD-independent gene repression mechanisms. Our study provides an accessible workflow for the discovery and prioritization of AS-NMD targets. It further argues that the AS-NMD pathway plays a widespread role in developing neurons by facilitating the downregulation of functionally related non-neuronal genes.