A Proximal Sox2 Enhancer Cluster is Required for the Anterior Regional Identity of Neural Progenitors

Embryonic development depends on spatially and temporally orchestrated gene regulatory networks. Expressed in neural stem and progenitor cells (NSPCs), the transcription factor sex-determining region Y box 2 (Sox2) is critical for embryogenesis and stem cell maintenance in neural development. Whereas the enhancers of Sox2 are well defined in early embryogenesis, little is known about Sox2 gene regulation in the neural lineage. Using functional genome data, we identify an enhancer cluster region that regulates Sox2 transcription in NSPCs derived from mouse embryonic stem cells (ESCs). By generating allelic mutants using CRISPR-Cas9 mediated deletions, we show that this proximal enhancer cluster, termed Sox2 regulatory regions 2-18 (SRR2-18), controls Sox2 allelic dosage during neural differentiation. Transcriptome analyses demonstrate that SRR2-18 is necessary for maintaining the anterior regional identity of ESC-generated NSPCs. Biallelic deletion of the Sox2 neural enhancer cluster perturbs the regulation of neurodevelopmental and regionalization genes. Furthermore, homozygous NSPC deletants exhibit cell-type autonomous self-renewal defects and altered developmental potency. Altogether, our data define a cis-regulatory enhancer cluster controlling Sox2 in the neural lineage and highlight how the allelic Sox2 dosage is critical for the anterior-posterior regionalization of NSPCs.