COMMD1 and PtdIns(4,5)P2 interaction maintain ATP7B copper transporter trafficking fidelity in HepG2 cells

Abstract

Copper-responsive intracellular ATP7B trafficking is critical to maintain copper balance in mammalian hepatocytes and thus organismal copper levels. The COMMD1 protein binds both the ATP7B copper transporter and phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2), while COMMD1 loss causes hepatocyte copper accumulation. Although it is clear that COMMD1 is localized to endocytic trafficking complexes, a direct function for COMMD1 in ATP7B trafficking is not defined. In this study, experiments using quantitative colocalization analysis reveal that COMMD1 modulates the copper-responsive ATP7B trafficking through recruitment to PtdIns(4,5)P2. Decreased COMMD1 abundance results in loss of ATP7B from lysosomes and the trans-Golgi network (TGN) in high copper conditions, while excess expression of COMMD1 also disrupts ATP7B trafficking and TGN structure. Overexpression of COMMD1 mutated to inhibit PtdIns(4,5)P2 binding has little impact on ATP7B trafficking. A mechanistic PtdIns(4,5)P2-mediated function for COMMD1 is proposed that is consistent with decreased cellular copper export due to disruption of the ATP7B trafficking itinerary and early endosome accumulation when COMMD1 is depleted. PtdIns(4,5)P2 interaction with COMMD1 as well as COMMD1 abundance may both be important in maintenance of specific membrane protein trafficking pathways.

Membrane Heterogeneity Controls Cellular Endocytic Trafficking

“…PI(4,5)P 2 must be generated on the Rab7 + endosomal membrane by the phophoinositide-5 kinase PIPKIγi5 for cargo entry and direct retromer recruitment (Sun et al, 2020). Furthermore, binding of the COMMD1 subunit of the CCC complex to PI(4,5)P 2 is required for cargo recovery to the trans-Golgi network (Stewart et al, 2019), providing a potential link between retriever and the PI(3)P/PI(4,5)P 2 -enriched membrane domain utilized by retromer for cargo recovery (Figures 3D,E -left).…”

Copper Homeostasis in Mammals, with Emphasis on Secretion and Excretion. A Review

“…Another protein (COMMD1, formerly MURR1) is also necessary for biliary Cu excretion, as first demonstrated in Bedlington terriers suffering from liver Cu toxicosis [5,28]. COMMD1 was the first member of what is now recognized as a family of ten proteins involved in endosomal sorting, and also plays a role in endosomal recycling [27]. As a complex with two other proteins (CCDC22-CCDC93), it binds to ATP7B in early endosomes, and somehow assists its return to the TGN [26] (Figure 4).…”

“…When levels of excess Cu in the hepatocyte subside, ATP7B recycles back into the TGN, via a clathrin-dependent endocytic process (Figure 4). This depends on a leucine triad located in the C-terminal portion of the ATP7B that is recognized by adaptins [27]. Another protein (COMMD1, formerly MURR1) is also necessary for biliary Cu excretion, as first demonstrated in Bedlington terriers suffering from liver Cu toxicosis [5,28].…”

“…These processes are summarized in Figure 4. (For further details, please see reviews by Polishchuk and Lutsenko [25], and Polishchuk and Polishchuk [26], as well as Stewart et al [27]. )…”

Investigation of Genetic Modifiers of Copper Toxicosis in Labrador Retrievers

“…COMMD1 and ATP7B are involved in copper excretion into the bile. Under high intracellular copper conditions, ATP7B traffics into late endosomes/lysosomes, and COMMD1, as a complex with CCD22-CCDC93, binds to ATP7B and mediates ATP7B trafficking and plays a role in ATP7B stability and fidelity [4,5]. Additionally, ATP7B incorporates copper into apo-ceruloplasmin to form holo-ceruloplasmin in the TGN, which is subsequently excreted into the plasma [4].…”