Brain calcification‐associated JAM2 pathogenic mutations impair JAM2 – JAM3 tight junction structure and the blood–brain barrier integrity

Primary brain calcification (PBC) is a neurodegenerative disorder characterized by bilateral brain calcification. JAM2 is a PBC causative gene, encoding the tight junction protein JAM2 and critical for blood-brain barrier (BBB) integrity, with an uncertain pathogenic mechanism. We analyzed two JAM2 missense mutations identified in PBC patients using plasmid constructs, immunoprecipitation, flow cytometry, and structural modeling. A Jam2 knockout (KO) mouse model was generated to assess behavioral deficits, brain calcification, and BBB integrity via histology, Evans blue assay, and in situ hybridization. JAM2-W108C mutants showed abnormal localization, while JAM2-R108H specifically disrupted JAM2-JAM3 interaction, mirroring the pathogenic JAM3-E116K mutation. Jam2 KO mice exhibited midbrain calcification (von Kossa staining) and motor impairments (rotarod/beam-walking tests) at 6-12 months. JAM2 and JAM3 co-localized in brain endothelial cells, and KO mice demonstrated BBB leakage (Evans blue extravasation). The main pathogenic mechanism of JAM2-PBC should be associated with the impaired JAM2-JAM3 heterodimer formation, compromising tight junctions and BBB integrity. Our study indicates JAM2-JAM3 interaction as a potential therapeutic target for PBC and related brain calcification disorders.