Multi-omic analysis of the ciliogenic transcription factorRFX3reveals a role in promoting activity-dependent responses via enhancing CREB binding in human neurons

Heterozygous loss-of-function (LoF) variants inRFX3,a transcription factor known to play key roles in ciliogenesis, result in autism spectrum disorder (ASD) and neurodevelopmental delay. RFX binding motifs are also enriched upstream of genes found to be commonly dysregulated in transcriptomic analyses of brain tissue from individuals with idiopathic ASD. Still, the precise functions ofRFX3in the human brain is unknown. Here, we studied the impact ofRFX3deficiency using human iPSC-derived neurons and forebrain organoids. Biallelic loss ofRFX3disrupted ciliary gene expression and delayed neuronal differentiation, while monoallelic loss ofRFX3did not. Instead, transcriptomic and DNA binding analyses demonstrated that monoallelicRFX3loss disrupted synaptic target gene expression and diminished neuronal activity-dependent gene expression. RFX3 binding sites co-localized with CREB binding sites near activity-dependent genes, andRFX3deficiency led to decreased CREB binding and impaired induction of CREB targets in response to neuronal depolarization. This study demonstrates a novel role of the ASD-associated gene RFX3 in shaping neuronal synaptic development and plasticity.