Transcriptional regulation of temporal dynamics in human neurogenesis and neuronal maturation [CUT&Tag]

Human pluripotent stem cells (hPSCs) provide a valuable platform for studies of human neuronal differentiation, but the mechanisms regulating the timing of this process remain poorly understood. This study compares two differentiation systems with distinct timing of differentiation, transcription factor (TF)-induced forward programming and stepwise cellular differentiation. Our analyses reveal that divergent cellular trajectories drive distinct neurogenesis timing. Multi-omic analysis identifies crucial gene regulatory networks (GRNs) that govern cell fate determination and timing control. Modulation of these GRNs modulates neurogenesis and timing of neuronal maturation. Specifically, OLIG family TFs, enriched in the TF-induced system, promoted cell cycle exit through NOTCH signaling regulation; conversely, ablation of these TFs delayed neurogenesis. In contrast, NEUROD2 overexpression accelerated neuronal maturation through precocious activation of maturation gene modules. Collectively, these findings illuminate characteristics of the cell-intrinsic mechanisms that govern timing of differentiation; the studies also offer a framework for investigation into, and the rational design of, timing-controlled in vitro differentiation strategies.