Interaction of the copper chaperone HAH1 with the Wilson disease protein is essential for copper homeostasis

The delivery of copper to specific sites within the cell is mediated by dis tinct intracellular carrier proteins termed copper chaperones. Previous stu dies in Saccharomyces cerevisiae suggested that the human copper chaperone HAH1 may play a role in copper trafficking to the secretory pathway of the cell. In this current study, HAH1 was detected in lysates from multiple hum an cell lines and tissues as a single-chain protein distributed throughout the cytoplasm and nucleus. Studies with a glutathione S-transferase-HAH1 fu sion protein demonstrated direct protein-protein interaction between HAH1 a nd the Wilson disease protein, which required the cysteine copper ligands i n the amino terminus of HAH1, Consistent with these in vitro observations, coimmunoprecipitation experiments revealed that HAH1 interacts with both th e Wilson and Menkes proteins in vivo and that this interaction depends on a vailable copper. When these studies were repeated utilizing three disease-a ssociated mutations in the amino terminus of the Wilson protein, a marked d iminution in HAH1 interaction was observed, suggesting that impaired copper delivery by HAH1 constitutes the molecular basis of Wilson disease in pati ents harboring these mutations. Taken together, these data provide a mechan ism for the function of HAH1 as a copper chaperone in mammalian cells and d emonstrate that this protein is essential for copper homeostasis.