Abstract
Physiologic Cu levels regulate the intracellular location of the Cu ATPase ATP7B. Here, we determined the routes of Cu-directed trafficking of endogenous ATP7B in the polarized hepatic cell line WIF-B and in the liver in vivo. Copper (10 µm) caused ATP7B to exit the trans-Golgi network (TGN) in vesicles, which trafficked via large basolateral endosomes to the apical domain within 1 h. Although perturbants of luminal acidification had little effect on the TGN localization of ATP7B in low Cu, they blocked delivery to the apical membrane in elevated Cu. If the vesicular proton-pump inhibitor bafilomycin-A1 (Baf) was present with Cu, ATP7B still exited the TGN, but accumulated in large endosomes located near the coverslip, in the basolateral region. Baf washout restored ATP7B trafficking to the apical domain. If ATP7B was staged apically in high Cu, Baf addition promoted the accumulation of ATP7B in subapical endosomes, indicating a blockade of apical recycling, with concomitant loss of ATP7B at the apical membrane. The retrograde pathway to the TGN, induced by Cu removal, was far less affected by Baf than the anterograde (Cu-stimulated) case. Overall, loss of acidification-impaired Cu-regulated trafficking of ATP7B at two main sites: (i) sorting and exit from large basolateral endosomes and (ii) recycling via endosomes near the apical membrane.
Highlights
Copper (Cu) is essential for life because it is a cofactor for certain oxidative enzymes, including superoxide dismutase, cytochrome oxidase, tyrosinase, lysyl oxidase, and ceruloplasmin
Cu levels regulate the reversible trafficking of endogenous ATP7B in polarized WIF-B cells
Endogenous ATP7B should respond to Cu concentrations typically seen in the blood, and the redistribution towards the apical domain should occur with sufficiently rapid kinetics to avoid Cu toxicity
Summary
Copper (Cu) is essential for life because it is a cofactor for certain oxidative enzymes, including superoxide dismutase, cytochrome oxidase, tyrosinase, lysyl oxidase, and ceruloplasmin. Excess Cu in cells is toxic [1]. Cellular Cu homeostasis is highly regulated and is achieved in part by two intracellular Cu-transporting P-Type ATPases, ATP7A and ATP7B [2]. Menkes and Wilson's Diseases result from loss-of-function mutations in ATP7A and ATP7B, respectively [3]. In Wilson's Disease, loss of ATP7B function leads to excess Cu accumulation in the brain, kidney and the liver, due to defective biliary Cu excretion across the apical surface of hepatocytes. ATP7B in hepatocytes shows Curegulated localization in vivo [4]. We recently identified a Wilson's Disease mutation that affects the intracellular trafficking of ATP7B, while having little effect on ATPase activity itself, indicating that a mis-localization of ATP7B is sufficient to cause Wilson's Disease [5]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
