Regulation of Heme Synthesis and Proteasomal Activity by Copper: Possible Implications for Wilson's Disease

The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52900, Israel.
Journal of environmental pathology, toxicology and oncology : official organ of the International Society for Environmental Toxicology and Cancer 01/2009; 28(3):209-21. DOI: 10.1615/JEnvironPatholToxicolOncol.v28.i3.20
Source: PubMed


Wilson's disease (Wd) is a genetic disorder resulting in Cu2+ accumulation, and is caused by mutations in the ATP7B gene, the copper transporter. In vivo studies show a correlation between Cu2+ accumulation and malfunction of the heme biosynthesis pathway. In this study, we describe multiple effects of Cu2+ accumulation on heme synthesis, which, in turn, affect proteasomal activity. Cu2+ toxicity was examined in two hepatocellular carcinoma cell lines, HepG2 and Hep3B, with Hep3B cells containing an integrated hepatitis B virus genome. Exposure of HepG2 and Hep3B cells to Cu2+ inhibited the enzymes PBGD and ALAD of the heme synthesis pathway and, in parallel, upregulated heme oxygenase-1 (HO-1). Proto-porphyrin IX (PpIX) and the heme pool were reduced as a result of these processes. PpIX synthesis was found to be lower in cells expressing the mutant ATP7B (P1134P), compared to those expressing the WT enzyme. Proteasomal activity was inhibited under Cu2+ treatment in HepG2 cells; however, Cu2+ induced marked proteosomal acceleration in Hep3B cells. Under these conditions, Ub-conjugated proteins were gradually accumulated, whereas treatment with bathocuproine disulfonic acid (BCS), a Cu2+ chelator, reversed this effect. In conclusion, our data suggest that copper downregulates the heme synthesis pathway in hepatocellular cells and further reduces it in the presence of mutated ATP7B.

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    • "Additional local or systemic copper dependent pathways may also affect iron absorption. We hypothesized that changes in either systemic or intestinal copper status regulate HIF-2α and as consequence iron absorption genes since: (i) we and others have shown that Cybrd1, Slc11a2 and Slc40a1 are bona fide Hif-2α targets [6], [7]; (ii) in vitro evidence suggests that changes in cellular copper levels regulate HIF-1α [20] and (iii) anemia associated with genetic disorders of copper metabolism could lead to increases in tissue hypoxia and stabilize HIF-2α [21]–[25]. We demonstrate in a mouse model of nutritional copper deficiency an increase in duodenal hypoxia and Hif-2α; i.p. copper injection alleviated copper deficiency anemia, reduced tissue hypoxia and modulated HIF-2α and iron absorption genes (Slc40a1, Slc11a2, and Cybrd1) independently of intestinal iron levels or liver hepcidin. "
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    ABSTRACT: Iron and copper are essential trace metals, actively absorbed from the proximal gut in a regulated fashion. Depletion of either metal can lead to anemia. In the gut, copper deficiency can affect iron absorption through modulating the activity of hephaestin - a multi-copper oxidase required for optimal iron export from enterocytes. How systemic copper status regulates iron absorption is unknown. Mice were subjected to a nutritional copper deficiency-induced anemia regime from birth and injected with copper sulphate intraperitoneally to correct the anemia. Copper deficiency resulted in anemia, increased duodenal hypoxia and Hypoxia inducible factor 2α (HIF-2α) levels, a regulator of iron absorption. HIF-2α upregulation in copper deficiency appeared to be independent of duodenal iron or copper levels and correlated with the expression of iron transporters (Ferroportin - Fpn, Divalent Metal transporter - Dmt1) and ferric reductase - Dcytb. Alleviation of copper-dependent anemia with intraperitoneal copper injection resulted in down regulation of HIF-2α-regulated iron absorption genes in the gut. Our work identifies HIF-2α as an important regulator of iron transport machinery in copper deficiency.
    Full-text · Article · Mar 2013 · PLoS ONE
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    ABSTRACT: Environmental pollution of heavy metals is very abundant nowadays from industry, chemicals, old paints, and pipes or resulting from previous contaminants accumulating in the food chain. Most of the iron demands of the body are needed for heme synthesis and assembly, but iron is also required for Fe-S cluster proteins and other redox enzymes. Heme is an essential, iron-binding molecule used as a prosthetic group of hemoproteins or as a regulator in multiple cellular pathways. In this review, we focused on the effect of exposure to heavy metals, such as Pb, Ga, Cu, Kd, Hg and Al, on heme synthesis as the main iron-sequestering process of the human body. These metals compete with iron on transporters, reduce the cellular iron pool and moreover, bind to proteins, and cause physical and mental disturbances. Heavy metals mainly impair various aspects of the heme synthesis pathway: gene expression, enzyme activity, and iron integration into protoporphyrin IX. Main risk factors are described as well as effects on iron dependent processes in order to increase public awareness to the distribution of heavy metals in our close environment and the harsh consequences of exposure, even in low doses.
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    ABSTRACT: We propose a conclusive difference observed between the excitation conditions required to observe porphyrins and copper-metallothioneins in cells and/or tissues using an ordinary fluorescence microscope. We have emphasized the importance of examining the spectral properties of the emissions to avoid any serious mistakes such as confusing porphyrins with copper-metallothioneins in the liver and kidneys. However, microspectrophotometry is not a conventional method for either histochemical, cytochemical, or pathological studies because microspectrophotometers are both expensive and difficult to operate. Therefore, we demonstrate a simple comparative method using ordinary excitation filter arrangements. When using our technique, it becomes possible to optically discriminate more accurately between the autofluorescence properties arising from porphyrins and those arising from copper-metallothioneins. We would like to name our simple technique "Triple Observation Method (TOM)".
    Full-text · Article · Nov 2010 · Journal of Fluorescence