Novel interactions between FOXP2 and PAX6: Implications for neural development and autism spectrum disorders

FOXP2 and PAX6 are transcription factors essential for neural development, with mutations in both linked to autism spectrum disorders (ASDs). Their DNA-binding domains include a forkhead domain (FHD) for FOXP2 and a paired domain (PD) plus homeodomain (HD) for PAX6. We investigated whether the FOXP2 FHD interacts directly with PAX6 PD or HD, and how such interactions influence DNA binding. Fluorescence anisotropy showed that all three domains bind specifically to their respective DNA targets with similar affinities. The FOXP2 FHD also interacts directly with both PAX6 PD and HD, with low micromolar binding affinities. Despite its stronger intrinsic DNA affinity, the FHD was displaced from its target DNA by both PAX6 domains, suggesting that protein-protein interactions can override DNA affinity under competitive conditions. In contrast, FOXP2 could not displace PD or HD from their DNA targets. Molecular docking supported these findings: DNA-protein interfaces were largely unchanged by the second protein, but protein-protein interfaces were strongly influenced by DNA occupancy. The H3 helix of FHD was identified as a central point for assembly, contributing to both DNA and protein interfaces. When FHD was bound to DNA, H3 was occupied, forcing PD or HD to dock at alternative, less optimal sites. HD maintained stronger contacts in these rearranged states, consistent with its greater competitive strength. This asymmetric interplay indicates competitive dominance by PAX6 and suggests mechanisms that could underlie transcriptional regulation in neurodevelopment.