REST deficiency and neurogenic-to-gliogenic shift in down syndrome human cerebral organoids

Abstract Down syndrome (DS) features impaired cortical neurogenesis and excess gliogenesis, yet the temporal regulatory events driving this imbalance remain unclear. Here, we combine multi-timepoint transcriptomic analyses from publicly available datasets, network modelling, and machine-learning prioritization, with validation in isogenic human iPSC-derived cerebral organoids, to identify a discrete pathogenic window at 90 days in vitro (DIV 90). Across five developmental stages, REST target genes were preferentially dysregulated in DS organoids. WGCNA revealed a DS-associated module at DIV-90 that strongly overlapped with REST targets, and two orthogonal machine-learning approaches converged on six REST-regulated hub genes— CSTB , MCM3AP , PFKL , POFUT2 , PRMT2 , and RWDD2B . In trisomic organoids, REST mRNA and nuclear protein were markedly reduced at DIV-90, accompanied by diminished DCX expression and activation of NFIA and STAT3, suggesting a neurogenic-to-gliogenic fate transition. These findings suggest REST dysfunction as a potential temporal regulator of lineage imbalance in DS and highlight REST-linked networks as potential targets for early developmental intervention.