Directed Differentiation of Human Embryonic Stem Cells into Corticofugal Neurons Uncovers Heterogeneous <i>Fezf2</i>-Expressing Subpopulations

<div><p>Understanding how neuronal diversity is achieved within the cerebral cortex remains a major challenge in neuroscience. The advent of human embryonic stem cells (hESCs) as a model system provides a unique opportunity to study human corticogenesis <i>in vitro</i> and to identify the mechanisms that promote neuronal differentiation to achieve neuronal diversity in human brain. The transcription factor <i>Fezf2</i> is necessary and sufficient for the specification of subcerebral projection neurons in mouse. However, its function during human corticogenesis is poorly understood. This study reports the differentiation of a h<i>Fezf2</i>-YFP hESC reporter line into corticofugal projection neurons capable of extending axons toward the spinal cord upon transplantation into neonatal mouse brains. Additionally, we show that triple inhibition of the TGFß/BMP/Wnt-Shh pathway promotes the generation of h<i>Fezf2</i>-expressing cells <i>in vitro</i>. Finally, this study unveils the isolation of two novel and distinct populations of h<i>Fezf2</i>-YFP expressing cells reminiscent of the distinct <i>Fezf2</i>-expressing neuronal subtypes in the developing mouse brain. Overall our data suggest that the directed differentiation of hESCs into corticofugal neurons provides a useful model to identify the molecular mechanisms regulating human corticofugal differentiation and survival.</p>