Base editing reveals an essential role for NANOG in human embryo lineage specification

Understanding the mechanisms that regulate how the first cell lineages in human development are specified and maintained is of fundamental importance and has clinical implications for regenerative medicine, infertility and pregnancy loss. Decades of research in mouse models have provided valuable insights into the role of transcription factors that regulate early development, however translating these findings to human embryos has been limited by ethical, technical and biological constraints. Functional studies of key transcription factors in human embryos have been hindered by nuclease-based genome-editing approaches that often induce genotoxicity, such as aneuploidy1-3. To overcome this limitation, we applied adenine base editing (ABE8e)4,5 to efficiently and precisely target an exon splice donor site, resulting in a splicing defect and a functional knockout of the transcription factor NANOG. This work represents the first application of base editing to study a developmental regulator in human embryos. Importantly, we found this approach did not trigger genotoxicity. Loss of NANOG disrupts specification of the pluripotent epiblast and instead cells differentiate toward a yolk sac or placental-like transcriptional programme. The retention of yolk-sac progenitor cell differentiation in NANOG-edited human embryos reveals a functional compensation that is distinct from our observations in the mouse, underscoring the importance of direct investigation of human development. Altogether, our findings demonstrate an essential role for NANOG in human pluripotency and epiblast specification and highlights the utility of adenine base editing for precise functional interrogation of human development.