Symmetric neural progenitor divisions require chromatin-mediated homologous recombination DNA repair by Ino80 [RNA-seq]

Chromatin regulates spatiotemporal gene expression during neurodevelopment, but it also mediates DNA damage repair crucial to proliferating neural progenitor cells (NPCs). Here, we uncover molecularly dissociable roles for nucleosome remodeler INO80 in chromatin-mediated transcriptome and genome maintenance in corticogenesis. We find that conditional Ino80 deletion from cortical NPCs impairs DNA double-strand break repair, triggering p53 activation, robust apoptosis, and microcephaly, whereas co-deletion of Trp53 with Ino80 leads to remarkable reversal of these phenotypes. Using an in vivo DNA repair assay, we find that Ino80 is selectively required for homologous recombination (HR) DNA repair, which is mechanistically distinct from INO80 function in YY1-associated transcription. Unexpectedly, sensitivity to loss of INO80-mediated HR is dependent on the mode of NPC division: Ino80 deletion causes extensive DNA damage and apoptosis in symmetric NPC-NPC divisions, but not in asymmetric neurogenic divisions. Thus, distinct modes of NPC division have divergent requirements for Ino80-dependent HR DNA repair. We analyzed a total of 19 samples of rRNA-depleted RNA from E13.5 mouse cortex using UMI RNA-seq. Libraries were generated using Click-seq. 7 samples were controls, 5 samples were Emx1-Cre Ino80 conditional mutants, and 7 samples were Emx1-Cre Ino80/Trp53 conditional double mutants.染色质在神经发育进程中调控基因表达的时空特异性，同时还介导对增殖性神经前体细胞（neural progenitor cells，NPCs）至关重要的DNA损伤修复过程。本研究揭示了核小体重塑因子INO80在大脑皮层发育过程中，通过染色质调控转录组与基因组维持的两种分子层面可分离的功能。研究发现，从皮层神经前体细胞中条件性敲除Ino80会损伤DNA双链断裂修复过程，进而激活p53通路、引发严重的细胞凋亡与小头畸形；而同时敲除Ino80与Trp53则可显著逆转上述表型。通过体内DNA修复实验检测，我们发现Ino80仅对同源重组（homologous recombination，HR）型DNA修复具有特异性需求，这一功能与INO80在YY1相关转录过程中的作用机制完全不同。出乎意料的是，细胞对INO80介导的同源重组缺失的敏感性取决于神经前体细胞的分裂模式：在神经前体细胞的对称分裂过程中，敲除Ino80会引发广泛的DNA损伤与细胞凋亡，但在不对称神经源性分裂中则无此效应。由此可见，神经前体细胞的不同分裂模式，对依赖Ino80的同源重组DNA修复有着截然不同的需求。本研究采用唯一分子标识符RNA测序（UMI RNA-seq）技术，对19份取自胚胎13.5天小鼠皮层的核糖体RNA去除型RNA样本进行了分析。所有文库均通过Click-seq方法构建：其中7份为对照组样本，5份为Emx1-Cre介导的Ino80条件性敲除突变体样本，剩余7份为Emx1-Cre介导的Ino80与Trp53双条件性敲除突变体样本。