Uncovering the role of FOXA2 in the Development of Human Serotonin Neurons

Abstract Directed differentiation of serotonin neurons (SNs) from human pluripotent stem cells (hPSCs) provides a valuable tool for uncovering the mechanism of human SN development and the associated neuropsychiatric disorders. Previous studies report that FOXA2 is expressed by serotonergic progenitors (SNPs) and functioned as a serotonergic fate determinant in mouse. However, in the routine differentiation experiments, it is accidentally found that less SNs and more non‐neuronal cells are obtained from SNP stage with higher percentage of FOXA2‐positive cells. This phenomenon prompted them to question the role of FOXA2 as an intrinsic fate determinant for human SN differentiation. Herein, by direct differentiation of engineered hPSCs into SNs, it is found that the SNs are not derived from FOXA2‐lineage cells; FOXA2‐knockout hPSCs can still differentiate into mature and functional SNs with typical serotonergic identity; FOXA2 overexpression suppresses the SN differentiation, indicating that FOXA2 is not intrinsically required for human SN differentiation. Furthermore, repressing FOXA2 expression by retinoic acid (RA) and dynamically modulating Sonic Hedgehog (SHH) signaling pathway promotes human SN differentiation. This study uncovers the role of FOXA2 in human SN development and improves the differentiation efficiency of hPSCs into SNs by repressing FOXA2 expression.