哺乳动物大脑皮层神经前体细胞（neural progenitor cells, NPCs）的发育是一个边特化边分化的过程，随着发育时间的推移NPCs逐步拥有了形成神经元和神经胶质细胞的特性。神经元和神经胶质形成相关基因的抑制和激活对神经细胞谱系的分化尤为重要。多梳家族（PcG）和三胸家族(TrxG)就是通过对组蛋白的表观遗传修饰来调控染色质状态而起到对基因表达的抑制和激活调控的。已有研究表明，在细胞中PcG家族能够形成抑制性染色质结构域而呈现点状分布；而后的研究通过高通量染色质构象捕获技术（HiC）观察到从胚胎干细胞向神经前体细胞及神经元分化的过程中，PcG通过调控基因表达和染色质结构而起作用。这说明PcG家族对神经细胞谱系的形成很重要，而Nspc1作为PcG家族PRC1复合体的核心成员之一，其功能及行使功能的机制尚不清楚。因此，本研究聚焦Nspc1这一基因，探索其在神经细胞发育过程中的功能及发挥功能的机制。

本研究通过D6-Cre工具鼠构建了小鼠脑皮层特异性敲Nspc1基因模型。通过Pax6和Tbr2对小鼠脑皮层神经前体细胞进行特异性标记，发现从E14.5到E16.5的神经元发育时期，神经前体细胞的数量并没有明显变化，而从E18.5到P3这一神经胶质细胞形成时期，Pax6⁺神经前体细胞数量显著增多。研究进一步对神经元和神经胶质细胞的分化进行检测，通过Cux1标记Ⅱ/Ⅲ两个皮层，Ctip2标记Ⅴ皮层，Tle4标记Ⅵ皮层的神经元，发现只有Tle4⁺神经元的生成数量有轻微增加。

通过Aldh1l1，GFAP和Sox10对神经胶质的发生情况进行检测，发现神经胶质发生时期GFAP⁺信号减弱，Aldh1l1⁺细胞在皮质板区的数量减少；分离E18.5的NPCs进行体外培养，发现培养4 d的NPCs，与野生型相比，成球能力显著增强，神经球的直径明显增大。基于这些现象，研究对神经胶质发育时期的神经前体细胞进行了RNA-Seq，ATAC-Seq和BL-HiC实验，检测在敲除Nspc1后神经前体细胞的基因表达，染色质可及性和染色质结构变化情况，发现敲除Nspc1后，在神经胶质发育阶段神经元形成和神经胶质形成相关的基因受到干扰，在本该是神经胶质形成阶段，神经元形成相关的基因依然在高表达，而神经胶质形成相关的基因表达量较低。随后，我们在基因组中检测到一些与神经胶质形成密切相关的Motif可及性下调，下调最明显的Motif 是NF1A，另外，形成染色质环的重要成员CTCF可及性也下调。而且，敲除Nspc1后，神经前体细胞的染色质局部区域变得致密，染色质接触变强。

因此，本研究认为Nspc1有可能通过PcG复合体影响染色质可及性和染色质3D结构，介导神经细胞谱系形成的关键基因表达，干扰神经胶质形成相关的信号通路，进而影响NPCs的特化，神经元和神经胶质细胞的分化。

The development of neural progenitor cells (NPCs) in the mammalian cerebral cortex is a process of specialization and differentiation. With the development of time, NPCs gradually possess the characteristics of forming neurons and glial cells. Inhibition and activation of genes associated with neuronal and glial formation are particularly important for the formation of neural cell lineages. Polycomb family (PcG) and TrxG family (TrxG) inhibit and activate gene expression by regulating chromatin state through epigenetic modification of histones. It has been shown that PcG family can form an inhibitory chromatin domain and present a spot-like distribution in cells. Later studies used high-throughput chromatin conformational capture (HiC) to observe that PcG regulates gene expression and chromatin structure during differentiation from embryonic stem cells to neural progenitor cells and neurons. This indicates that PcG family plays a very important role in the formation of neural cell lineages, while Nspc1 is one of the core members of PRC1 complex of PcG family, and its function and the mechanism of its function are still unclear. Therefore, this study focused on the gene Nspc1 to explore its function in the development of neural cells and its functional mechanism.

In this study, cortical knockdown Nspc1 gene model was constructed by D6-Cre tool mice. The number of Pax6⁺ progenitors did not change significantly from E14.5 to E16.5 during neuronal development, but increased significantly from E18.5 to P3 during glial cell formation. The differentiation of neurons and glial cells was further examined by Cux1 labeling the Ⅱ/Ⅲ layer, Ctip2 labeling the Ⅴ layer, and Tle4 labeling the Ⅵ layer. Only a slight increase in the number of Tle4⁺ neurons was found.

The occurrence of glia was detected by Aldh1l1, GFAP and Sox10. It was found that GFAP⁺ signal was weakened during gliogenesis, and the number of Aldh1l1⁺ cells in cortical plate decreased. E18.5 NPCs were isolated and cultured in vitro, compared with the wild type, NPCs cultured for 4 days had significantly enhanced neurosphere forming ability and significantly increased the diameter of neurosphere. Based on these findings, RNA-seq, ATAC-seq and BL-HiC experiments were performed on neural progenitor cells during glial development to detect gene expression, chromatin accessibility and chromatin structure changes of NPCs after Nspc1 knockout. It was found that after Nspc1 was knocked out, genes related to neuronal formation and glial formation were disrupted during the glial development stage. During the supposed glial development stage, the genes related to neuronal formation was still overexpression, while the genes related to glial formation was downregulated. Subsequently, we detected accessibility downregulation of motifs in the genome that are closely related to glial formation, with NF1A being the most significant Motif and CTCF, an important member of chromatin loop formation, also downregulated. Moreover, local chromatin regions of NPCs became denser and chromatin contact increased after knockout of Nspc1.

Therefore, this study suggests that Nspc1 may influence chromatin accessibilty and chromatin 3D structure through PcG complex, mediate the expression of key genes in the formation of neural cell lineages, and interfere with gliogenesis related signaling pathways, thereby affecting the specialization of NPCs, neuron and glial cell formation.