Transcriptome Sequencing During Mouse Brain Development Identifies Long Non-Coding RNAs Functionally Involved in Neurogenic Commitment

Transcriptome analysis of somatic stem cells and their progeny is fundamental to identify new factors controlling proliferation versus differentiation during tissue formation. Here we generated a combinatorial, fluorescent reporter mouse line to isolate proliferating neural stem cells, differentiating progenitors and newborn neurons that coexist as intermingled cell populations during brain development. Transcriptome sequencing revealed numerous novel long non-coding (lnc)RNAs and uncharacterized protein-coding transcripts identifying the signature of neurogenic commitment. Importantly, most lncRNAs overlapped neurogenic genes and shared with them a nearly identical expression pattern suggesting that lncRNAs control corticogenesis by tuning the expression of nearby cell fate determinants. We assessed the power of our approach by manipulating lncRNAs and protein-coding transcripts with no function in corticogenesis reported to date. This led to several evident phenotypes in neurogenic commitment and neuronal survival indicating that our study provides a remarkably high number of uncharacterized transcripts with hitherto unsuspected roles in brain development. Finally, we focussed on one lncRNA, Miat, whose manipulation was found to trigger pleiotropic effects on brain development and aberrant splicing of Wnt7b. Hence, our study suggests that lncRNA-mediated alternative splicing of cell fate determinants controls stem cell commitment during neurogenesis. "LncRNAs control neurogenesis" Aprea, Prenninger, Dori, Monasor, Wessendof, Zocher, Massalini, Ghosh, Alexopoulou, Lesche, Dahl, Groszer, Hiller, Calegari, The EMBO Journal (In Press) mRNA profiles of Proliferating Progenitors, Differentiating Progenitors and Neurons from lateral cortex of E14.5 mouse embryos. Each cell type in three biological replicates.体细胞干细胞（somatic stem cells）及其子代细胞的转录组分析，是鉴定组织形成过程中调控细胞增殖与分化的新型因子的核心研究基础。本研究构建了组合式荧光报告小鼠品系，可分离出脑发育过程中共存于混杂细胞群中的增殖态神经干细胞、分化中的前体细胞与新生神经元。通过转录组测序，本研究鉴定出大量新型长链非编码RNA（long non-coding RNAs，lncRNAs）与未注释的蛋白编码转录本，明确了神经发生定向的特征标签。值得注意的是，绝大多数lncRNAs与神经发生基因区域存在重叠，且二者的表达模式近乎一致，这提示lncRNAs通过调控邻近细胞命运决定因子的表达，参与大脑皮层发育的调控。我们通过靶向调控目前尚未见大脑皮层发育功能报道的lncRNAs与蛋白编码转录本，验证了本研究方法的有效性。该操作在神经发生定向与神经元存活层面引发了多项显著表型，表明本研究鉴定出了大量此前未被认知、且在脑发育中发挥关键作用的未注释转录本。最后，我们针对其中一种lncRNA——Miat开展了深入研究，发现靶向调控Miat可引发脑发育的多效性效应，并导致Wnt7b出现异常剪接。综上，本研究表明，lncRNAs介导的细胞命运决定因子可变剪接，在神经发生过程中调控干细胞的命运定向。《长链非编码RNAs调控神经发生（LncRNAs control neurogenesis）》，作者：Aprea、Prenninger、Dori、Monasor、Wessendof、Zocher、Massalini、Ghosh、Alexopoulou、Lesche、Dahl、Groszer、Hiller、Calegari，发表于《欧洲分子生物学组织杂志（The EMBO Journal）》（已接收待刊）。本数据集包含小鼠胚胎发育第14.5天（E14.5）胎鼠外侧皮层来源的增殖态前体细胞、分化态前体细胞与神经元的mRNA表达谱，每种细胞类型均设置3次生物学重复。