Copper trafficking to the secretory pathway

Abstract

Copper (Cu) is indispensible for growth and development of human organisms. It is required for such fundamental and ubiquitous processes as respiration and protection against reactive oxygen species. Cu also enables catalytic activity of enzymes that critically contribute to the functional identity of many cells and tissues. Pigmentation, production of norepinephrine by the adrenal gland, the key steps in the formation of connective tissue, neuroendocrine signaling, wound healing - all these processes require Cu and depend on Cu entering the secretory pathway. To reach the Cu-dependent enzymes in a lumen of the trans-Golgi network and various vesicular compartments, Cu undertakes a complex journey crossing the extracellular and intracellular membranes and staying firmly on course while traveling in a cytosol. The proteins that assist Cu in this journey by mediating its entry, distribution, and export, have been identified. The accumulating data also indicate that the current model of cellular Cu homeostasis is still a "skeleton" that has to be fleshed out with many new details. This review summarizes recent data on the mechanisms responsible for Cu transfer to the secretory pathway. The emerging new concepts and gaps in our knowledge are discussed.

Fig. 1

The major pathways of Cu distribution in human cells. The cartoon illustrates the main Cu distribution pathways in hepatocytes. The high affinity Cu transporter CTR1 at the basolateral membrane is responsible for at least 50% of Cu entering the cell. The low affinity transporters are unknown. Upon entry, Cu may bind transiently to glutathione and be retrieved from glutathione or directly from CTR1 by cytosolic Cu chaperones. Cytosolic Cu chaperone for SOD1 in a Cu-bound form facilitates Cu incorporation into SOD1 and the formation of an important disulfide bond. Atox1 binds Cu within the Cys-Gly-Gly-Cys metal binding site, which is sensitive to changes in redox environment. Increase in the GSH : GSSG ratio increases the reduction of Atox metal-binding site, enabling more Cu binding to Atox1 and thus increasing the delivery of Cu to the secretory pathway. In the TGN, Cu-transporting ATPase ATP7B transfers Cu to ceruloplasmin, which is then secreted across the basolateral membrane. Cu elevation triggers trafficking of ATP7B from the trans-Golgi network toward the apical membrane (ATP7A in other cells traffics to basolateral membrane in response to high Cu), thus facilitating export of excess Cu. Complex set of soluble and membrane-bound chaperones facilitates Cu transfer to mitochondria.

Fig. 2

Structural model of CTR1. Left: A side view of the CTR trimer with the Met residues shown in purple, Cys in yellow, and His in cyan. Extracellular Cu (green ball) enters the transporter through the Met-rich selectivity filter (either directly or via intermediate binding to the HisHiscontaining site in the N-terminus of CTR1) and is directed through the wide vestibule of the transporter towards the narrow Cys/His-based ring at the cytoplasmic side. The release of Cu from CTR1 is driven either by conformational changes that disrupt the Cis/His ring or by interaction with the cytosolic molecule(s). Right: The top view of the Ctr1 trimer. The HisHis pair located in the vicinity of Met-rich entry is indicated by dark cyan and is circled. The images are generated using CTR1 pdb coordinates generated by Tsigelny and co-workers.

Fig. 3

Changes in the intracellular levels of metals in response to down regulation of specific transporters. The Z-score higher than plus 2 or lower than minus 2 indicates a significant change; a variation within this range (marked by a gray block) is insignificant. Inactivation of CTR1 (positive control) is associated with a selective decrease of Cu levels in cells. Inactivation of CLC7 (the intracellular chloride transporter) lowers Mg²⁺, iron, and Cu levels; inactivation of Zip5 (Zn²⁺-uptake transporter) significantly decreases the cellular Cu content and zinc levels, but also results in increases of iron and Mg²⁺ levels. The data illustrate importance of determining the status of entire metallome following inactivation of transporters with an unknown specificity.