Effects of Dlx2 Overexpression on the Genome of Maxillary Process in Early Embryo of Mice [WT Bulk RNA-seq]

Transcription factor Dlx2 plays an important role in craniomaxillofacial development. Overexpression or null mutation can lead to craniomaxillofacial malformation in mice. Although some in vivo or in vitro experiments have explored part information about the mechanism of Dlx2 regulation, there is still a lack of complete description. Using a mouse model that can stably overexpress Dlx2 in neural crest cells, the authors conducted bulk RNA-Seq, scRNA-Seq and CUT&Tag on the early maxillary processes of mice, and comprehensively described the effects of Dlx2 overexpression on the early development of maxillary processes. Bulk RNA-Seq results showed that Dlx2 had greater interference on nerve development at E10.5, and gradually affected bone development at E12.5. ScRNA-Seq proved that overexpression of Dlx2 did not change the differentiation type of mesenchymal cells during this development process, but it would restrict the proliferation of cells and make mesenchymal cells differentiate prematurely, thus limiting the development of maxillary. CUT&Tag suggested that this regulatory process is closely related to Notch signaling pathway, and MNT and RUNX2, as direct downstream regulatory target genes of Dlx2, also play a very critical role. The authors conducted bulk RNA Seq on the maxillary process of E10.5 wildtype mouse and Dlx2 overexpression mouse, analyzed the impact of Dlx2 overexpression on genome expression, and compared it with the existing data of E12.5 mouse maxillary process in GEO database.转录因子Dlx2在颅颌面发育中发挥关键调控作用。在小鼠体内，其过表达或功能缺失突变（null mutation）均可引发颅颌面畸形。尽管已有部分体内、体外实验对Dlx2的调控机制进行了探索，但目前仍缺乏完整的机制阐释。研究团队利用可在神经嵴细胞（neural crest cells）中稳定过表达Dlx2的小鼠模型，对小鼠早期上颌突进行了批量RNA测序（bulk RNA-Seq）、单细胞RNA测序（scRNA-Seq）以及CUT&Tag实验，并全面阐释了Dlx2过表达对上颌突早期发育的影响。批量RNA测序结果显示，Dlx2在胚胎发育第10.5天（E10.5）对神经发育的干扰更为显著，而在胚胎发育第12.5天（E12.5）则逐渐影响骨组织发育。单细胞RNA测序结果证实，在该发育过程中，Dlx2过表达并未改变间充质细胞（mesenchymal cells）的分化类型，但会抑制细胞增殖并促使间充质细胞过早分化，进而限制上颌突的发育。CUT&Tag实验结果表明，该调控过程与Notch信号通路（Notch signaling pathway）密切相关，而作为Dlx2直接下游调控靶基因的MNT与RUNX2同样发挥着极为关键的作用。研究团队对胚胎发育第10.5天的野生型小鼠与Dlx2过表达小鼠的上颌突进行了批量RNA测序，分析了Dlx2过表达对基因组表达的影响，并将该数据与GEO数据库（Gene Expression Omnibus database）中已公开的胚胎发育第12.5天小鼠上颌突相关数据进行了比对。