Regulation of heme synthesis and proteasomal activity by copper: possible implications for Wilson's disease.
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.
Hait-Darshan
Ravit
R
The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52900, Israel.
Babushkin
Tania
T
Malik
Zvi
Z
D000251:Adenosine Triphosphatases / Q000096:biosynthesis; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000378:metabolism; D027682:Cation Transport Proteins / Q000096:biosynthesis; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003300:Copper / Q000633:toxicity*; D000073840:Copper-Transporting ATPases; D004221:Disulfiram / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000633:toxicity*; D006418:Heme / Q000096:biosynthesis*; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006527:Hepatolenticular Degeneration; D006801:Humans; D011163:Hydroxymethylbilane Synthase / Q000037:antagonists & inhibitors; D008113:Liver Neoplasms / Q000188:drug therapy / Q000378:metabolism; D000623:Porphobilinogen Synthase / Q000037:antagonists & inhibitors; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects* / Q000378:metabolism; D011524:Protoporphyrins / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism
D027682:Cation Transport Proteins; D004791:Enzyme Inhibitors; D011524:Protoporphyrins; D017382:Reactive Oxygen Species; D025801:Ubiquitin; D006418:Heme; D003300:Copper; C028025:protoporphyrin IX; D011163:Hydroxymethylbilane Synthase; D046988:Proteasome Endopeptidase Complex; D000251:Adenosine Triphosphatases; D000623:Porphobilinogen Synthase; C000612464:ATP7B protein, human; D000073840:Copper-Transporting ATPases; C029892:cupric chloride; D004221:Disulfiram