Copper-dependent interaction of dynactin subunit p62 with the N terminus of ATP7B but not ATP7A.
The P-type ATPase affected in Wilson disease, ATP7B, is a key liver protein required to regulate and maintain copper homeostasis. When hepatocytes are exposed to elevated copper levels, ATP7B traffics from the trans-Golgi network toward the biliary canalicular membrane to excrete excess copper into bile. The N-terminal region of ATP7B contains six metal-binding sites (MBS), each with the copper-binding motif MXCXXC. These sites are required for the activity and copper-regulated intracellular redistribution of ATP7B. Two proteins are known to interact with the ATP7B N-terminal region: the copper chaperone ATOX1 that delivers copper to ATP7B, and COMMD1 (MURR1) that is potentially involved in vesicular copper sequestration. To identify additional proteins that interact with ATP7B and hence are involved in copper homeostasis, a yeast two-hybrid approach was employed to screen a human liver cDNA library. The dynactin subunit p62 (dynactin 4; DCTN4) was identified as an interacting partner, and this interaction was confirmed by co-immunoprecipitation from mammalian cells. The dynactin complex binds cargo, such as vesicles and organelles, to cytoplasmic dynein for retrograde microtubule-mediated trafficking and could feasibly be involved in the copper-regulated trafficking of ATP7B. The ATP7B/p62 interaction required copper, the metal-binding CXXC motifs, and the region between MBS 4 and MBS 6 of ATP7B. The p62 subunit did not interact with the related copper ATPase, ATP7A. We propose that the ATP7B interaction with p62 is a key component of the copper-induced trafficking pathway that delivers ATP7B to subapical vesicles of hepatocytes for the removal of excess copper into bile.
Centre for Cellular and Molecular Biology, School of Life and Environmental Sciences, Deakin University, Burwood, Victoria 3125, Australia.
Chris M
CM
Lim
Michael A
MA
Cater
Julian F B
JF
Mercer
Sharon
S
La Fontaine
D000251:Adenosine Triphosphatases / Q000737:chemistry*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001646:Bile / Q000378:metabolism; D027682:Cation Transport Proteins / Q000737:chemistry*; D003300:Copper / Q000737:chemistry* / Q000378:metabolism; D000073840:Copper-Transporting ATPases; D000072159:Dynactin Complex; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism
D027682:Cation Transport Proteins; C000606363:DCTN4 protein, human; D000072159:Dynactin Complex; D008869:Microtubule-Associated Proteins; D003300:Copper; D000251:Adenosine Triphosphatases; C081184:ATP7A protein, human; C000612464:ATP7B protein, human; D000073840:Copper-Transporting ATPases