Structural Destabilization of FRMD3 by a FERM Domain Mutation Causes Hypomyelinating Disease via Oligodendrocyte Dysfunction

Abstract

Hypomyelinating diseases are a heterogeneous group of neurodevelopmental disorders caused by genetic anomalies that impair myelin formation or maintenance. Here, we investigate a novel homozygous missense variant in FRMD3 (NM_174938.6:c.898T > C; p.C300R) in a 2-year-old male presenting with global developmental delay, hypotonia, mild ataxia, and MRI features consistent with hypomyelination. The variant affects a highly conserved cysteine within the FERM domain of FRMD3, and predicted to be deleterious by multiple in silico tools. Molecular dynamics simulations and biophysical analyses revealed that p.C300R introduces steric clashes with neighboring residues, destabilizes the FERM domain, increases structural disorder, and exposes hydrophobic aggregation-prone regions. In proband fibroblasts and nonproband neuronal cells, mutant FRMD3 mislocalized from the plasma membrane to the cytosol, forming large aggregates. Thioflavin T assays confirmed elevated aggregation propensity of the mutant. In oligodendrocytes, FRMD3-p.C300R expression markedly impaired neurite formation and failed to restore proteolipid protein 1 (PLP1) and myelin basic protein (MBP) expression following FRMD3 knockdown, in contrast to wild-type rescue. Interactome and single-cell expression analyses place FRMD3 at membrane-trafficking and lipid-handling hubs in oligodendrocytes and white-matter regions, and loss of these interactions through p.C300R-driven destabilization and aggregation likely underlies the regional hypomyelination observed in the proband. Our findings establish FRMD3 as a novel candidate gene for hypomyelinating disease and reveal that structural destabilization and aggregation of FERM-domain peripheral membrane protein can disrupt oligodendrocyte function and myelin protein expression, leading to neurodevelopmental pathology.

Keywords: FERM domain; FRMD3; hypomyelinating disease; oligodendrocyte dysfunction; protein aggregation.