RNA-seq analysis of Scn1a +/- mouse

In this study we recapitulated in the mouse, an SCN1A mutation found in a human Dravet syndrome (DS) patient. The targeted mutation, NC_000068.7:g.66293870C>G (GRCm38.p6) lies in a highly conserved alternate poison exon (20N) of the mouse Scn1a. We performed molecular and behavioral analysis of Scn1a +/- mice and Scn1a +/+ littermate controls. We found that the mutation causes Scn1a mRNA and protein levels to be reduced by about 50% in brain compared to control mice. In addition, the Scn1a +/- mice exhibit behavioral phenotypes seen in previous DS model mice models. We performed qPCR and RNA-seq analysis of the brains of four Scn1a +/- mice and four Scn1a +/+ littermate controls. There was a ~50% reduction in Scn1a RNA-seq counts in Scn1a +/- mice. Our data provides evidence that the mutation causes increased inclusion of the poison exon 20N in Scn1a transcripts leading to nonsense mediated decay and reduction in protein levels. RNA-seq analysis of four Scn1a +/- mice and four Scn1a+/+ littermate controls (brain tissue).本研究在小鼠模型中重现了人类德拉韦综合征（Dravet syndrome, DS）患者携带的SCN1A突变。本次引入的定点突变NC_000068.7:g.66293870C>G（GRCm38.p6参考基因组版本）位于小鼠Scn1a基因高度保守的可变毒外显子（20N）区域内。我们对Scn1a+/-杂合子小鼠与Scn1a+/+同窝野生型对照小鼠开展了分子与行为学分析。研究发现，相较于对照小鼠，该突变可使小鼠脑内Scn1a的mRNA与蛋白表达水平降低约50%。此外，Scn1a+/-杂合子小鼠呈现出既往德拉韦综合征小鼠模型中已报道的行为学表型。我们对4只Scn1a+/-杂合子小鼠与4只Scn1a+/+同窝野生型对照小鼠的脑组织开展了定量聚合酶链反应（qPCR）与RNA测序（RNA-seq）分析，结果显示Scn1a+/-杂合子小鼠的Scn1a基因RNA-seq测序reads计数降低约50%。本研究数据表明，该突变可促使Scn1a转录本中20N毒外显子的剪接保留，进而引发无义介导的mRNA衰变（nonsense mediated decay, NMD），最终导致蛋白表达水平降低。