Wild type vs. Nova2 KO mouse E14.5 cortex RNA

To assess the requirement of Nova2 for alternative processing of RNA in the developping brain. Neuronal migration leads to a highly organized laminar structure in the mammalian brain and its mis-regulation causes lissencephaly, behavioral and cognitive defects. Reelin signaling, mediated in part by a key adaptor, disabled-1 (Dab1), plays a critical but incompletely understood role in this process. We found that the neuron-specific RNA binding protein Nova2 regulates neuronal migration in late-generated cortical and Purkinje neurons. An unbiased HITS-CLIP and exon junction array search for Nova-dependent RNAs at E14.5 focused on components of the reelin pathway revealed only one candidate--an alternatively spliced isoform of Dab1 (Dab1.7bc). In utero electroporation demonstrated that Dab1.7bc was sufficient to induce neuronal migration defects in wild-type mice and exacerbate defects when Dab1 levels were reduced, while Dab1 overexpression mitigates defects in Nova2-null mice. Thus Nova2 regulates an RNA switch controlling the ability of Dab1 to mediate neuronal responsiveness to reelin signaling and neuronal migration, suggesting new links between splicing regulation, brain disease and development. Keywords: Comparative analysis RNA from the cortex of 3 wild type and 3 Nova2 KO E14.5 cortex. One array per biological replicate.为探究Nova2在发育大脑中对RNA可变加工的调控需求。神经元迁移可使哺乳动物大脑形成高度有序的层状结构，其调控异常会导致无脑回畸形、行为及认知缺陷。Reelin信号通路在该过程中发挥关键但尚未完全阐明的作用，其部分功能由关键接头蛋白Dab1（disabled-1）介导。本研究发现，神经元特异性RNA结合蛋白Nova2可调控晚期生成的皮层神经元与浦肯野神经元的迁移过程。本研究通过针对Reelin通路组分、在胚胎第14.5天（E14.5）开展的无偏倚HITS-CLIP与外显子连接阵列筛选，寻找Nova依赖的RNA，仅鉴定到一个候选靶点——Dab1的可变剪接异构体Dab1.7bc。子宫内电转实验证实，Dab1.7bc足以在野生型小鼠中诱导神经元迁移缺陷，并在Dab1水平降低时加剧缺陷；而Dab1过表达可缓解Nova2基因敲除小鼠的相关缺陷。综上，Nova2通过调控一种RNA开关，控制Dab1介导神经元对Reelin信号的应答能力及神经元迁移过程，这为剪接调控、脑部疾病与大脑发育之间建立了新的关联。 关键词：对3只野生型小鼠与3只Nova2敲除（Nova2 KO）小鼠胚胎第14.5天的皮层RNA开展比较分析。每个生物学重复对应一张外显子连接阵列。