Functional Screen of Wilson Disease ATP7B Variants Reveals Residual Transport Activities

Wilson disease is a disorder of copper (Cu) homeostasis caused by the malfunction of Cu transporter ATP7B and associated Cu accumulation in tissues. The existence of over 700 disease-associated variants in the ATP7B gene and a broad spectrum of disease manifestations complicate the analysis of genotype-phenotype correlations and the development of better treatments for this disorder. To assist such studies, we screen 101 variants of ATP7B for expression and Cu transport activity in human fibroblasts lacking active ATP-dependent Cu transporters. The ClinVar database classified 59 of these as variants of uncertain significance or having conflicting pathogenicity classifications; six variants were not in the database. Thirty-three of the variants have been previously characterized by other assays. Only three variants (S657R, G1061E, and G1266R) resulted in the complete inactivation of Cu transport. The in silico analysis of these mutants was used to rationalize this drastic effect on ATP7B activity. The remaining ATP7B variants showed a range of Cu transport activities. Coexpression of variants with different properties yielded activity values different from the simple average. The advantages and limitations of this functional screen are discussed.