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Neural Plate Targeting by in Utero NanoinjEction (NEPTUNE) reveals a role for Sptbn2 in neurulation and abdominal wall closure
bioRxiv - Developmental Biology Pub Date : 2020-05-28 , DOI: 10.1101/2020.05.28.115972
Katrin Mangold , Jan Mašek , Jingyan He , Urban Lendahl , Elaine Fuchs , Emma R Andersson

Gene variants associated with disease are efficiently identified with whole genome sequencing or GWAS, but validation in vivo lags behind. We developed NEPTUNE (neural plate targeting by in utero nanoinjection), to rapidly and flexibly introduce gene expression-modifying viruses to the embryonic murine neural plate prior to neurulation, to target the future adult nervous system. Stable integration in >95% of cells in the brain enabled long-term gain- or loss-of-function, and conditional expression was achieved using mini-promotors for cell types of interest. Using NEPTUNE, we silenced Sptbn2, a gene associated with Spinocerebellar ataxia type 5 (SCA5) in humans. Silencing of Sptbn2 induced severe neural tube defects and embryo resorption, suggesting that SPTBN2 in-frame and missense deletions in SCA5 reflect hypomorphic or neomorphic functions, not loss of function. In conclusion, NEPTUNE offers a novel, rapid and cost-effective technique to test gene function in brain development, and can reveal loss of function phenotypes incompatible with life.

中文翻译:

子宫纳米注射(NEPTUNE)靶向神经板揭示了Sptbn2在神经和腹壁闭合中的作用

可通过全基因组测序或GWAS有效地鉴定与疾病相关的基因变异,但体内验证滞后。我们开发了NEPTUNE(通过宫内纳米注射靶向神经板),以便在进行培养之前快速灵活地将基因表达修饰病毒引入胚胎鼠神经板,以靶向未来的成年神经系统。在大脑中> 95%的细胞中稳定整合可以实现长期的功能丧失,并且使用微型启动子针对感兴趣的细胞类型实现了条件表达。使用NEPTUNE,我们沉默了与人类小脑共济失调5型(SCA5)相关的基因Sptbn2Sptbn2的沉默引起严重的神经管缺陷和胚胎吸收,提示SCA5中的SPTBN2读框内和错义缺失反映了亚型或新态功能,而不是功能丧失。总之,NEPTUNE提供了一种新颖,快速且经济高效的技术来测试大脑发育中的基因功能,并且可以揭示与生命不相容的功能表型的丧失。
更新日期:2020-05-28
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