MECP2 MBD-ID Module: A Unified DNA/RNA Binding Interface Disrupted in Rett Syndrome

ABSTRACT Rett syndrome neurodevelopmental disorder is caused by mutations in the gene encoding the epigenetic regulator MECP2. While the MECP2 methyl-CpG binding domain (MBD) is well-characterized, the function of the adjacent intervening domain (ID) remains largely understudied. The ID has been described as a distinct RNA-binding region, yet evidence also suggests RNA competitively displaces MECP2 from DNA. Here, we address these conflicting findings by demonstrating the MBD and ID do not function in isolation but as a synergistic functional unit, establishing a new model for MECP2 function. We show the ID significantly enhances affinity of the MBD for methylated DNA by ∼35-fold. Moreover, together these two subdomains form a high-affinity, promiscuous RNA-binding module, with affinity for structured RNAs increased over 1,000-fold compared to the MBD or ID alone. We find binding to RNA precludes binding to DNA, such that the integrated MBD-ID unit explains the competition phenomenon. Analysis of Rett syndrome-associated ID mutations (R167W, K174Q, and R190H) and a therapeutic MiniGene reveals they do not disrupt methyl-DNA binding but instead selectively weaken RNA and non-methylated DNA binding, thereby disrupting the competitive balance between nucleic acid ligands. Our work establishes the MBD-ID module as MECP2’s central nucleic acid interaction hub, whose disruption provides a potential molecular etiology of Rett syndrome due to mutations in the intervening domain. GRAPHICAL ABSTRACT