TBR2 COORDINATES ATP-DEPENDENT CHROMATIN REMODELING REQUIRED FOR NEURONAL DIFFERENTIATION IN HUMAN CORTICAL ORGANOIDS

The T-box transcription factor TBR2 (EOMES) plays a central role in intermediate progenitor (IP) dynamics during cortical development, yet its mechanistic contribution to human neuronal differentiation remains incompletely understood due to early embryonic lethality associated with total TBR2 loss in vivo. To address this limitation, we generated CRISPR/Cas9-mediated TBR2 knockout (KO) human induced pluripotent stem cells (hiPSC) and differentiated them into forebrain organoids to model early stages of human corticogenesis. Immunofluorescence, metabolic assays, and single-nucleus transcriptomic and chromatin accessibility profiling revealed an accumulation of neural progenitors accompanied by a profound impairment in neuronal differentiation, with particularly strong effects on subplate and cortical layer neuron populations. While chromatin accessibility at neurogenic loci remained largely permissive, multiomic integration uncovered widespread, neuron-biased transcriptional and epigenetic alterations. Notably, TBR2-deficient organoids failed to execute the normal developmental switch from progenitor-associated to neuronal ATPase subunits within ATP-dependent chromatin remodeling complexes, identifying components of the NuRD (CHD3, CHD5) and neuronal BAF (nBAF; SMARCA2) complexes as candidate downstream targets of TBR2 regulation. In parallel, TBR2 loss was associated with altered mitochondrial morphology and downregulation of translational and oxidative phosphorylation pathways. Pharmacological enhancement of translation using the integrated stress response inhibitor ISRIB rescued progenitor-associated and mitochondrial phenotypes but did not restore neuronal differentiation, indicating that metabolic and translational defects reflect downstream or compensatory responses to TBR2 loss. Together, these findings position TBR2 as a key regulator of chromatin remodeling dynamics during the progenitor-to-neuron transition and highlight how disruption of this regulatory axis may contribute to neurodevelopmental disorders.