FMR1 KH0-KH1 Domains Coordinate m6A Binding and Phase Separation in Fragile X Syndrome

Abstract

The Fragile X Mental Retardation Protein (FMR1) regulates neurodevelopment via m6A RNA interactions, but domain-specific roles of KH0 and KH1 in RNA binding and pathology remain unclear. Through mutagenesis and AlphaFold3 modeling, we demonstrate that KH1 serves as the primary m6A-binding interface, whereas the KH0 domain (e.g., Arg138) modulates liquid-liquid phase separation (LLPS). Pathogenic mutations in KH0 disrupt RNA binding and enhance LLPS aggregation, while m6A RNA suppresses LLPS at KH0. Structural simulations reveal synergistic cooperation between KH0 and KH1 through hydrophobic and electrostatic networks. These dual-domain interactions establish a mechanistic link between m6A dysregulation, aberrant phase separation, and Fragile X syndrome pathogenesis. Our findings propose KH0 as a therapeutic target for RNA-driven neurodevelopmental disorders.