Nitric oxide-dependent pro-oxidant and pro-apoptotic effect of metallothioneins in HL-60 cells challenged with cupric nitrilotriacetate

Intracellular safeguarding functions of metallothioneins (MTs) include sequ estering transition and heavy metals, scavenging free radicals and protecti ng against electrophiles. We report that MT protection against Cu-induced c ytotoxicity can be reversed and pro-oxidant and pro-apoptotic effects can b e induced in HL-60 cells exposed to NO. We demonstrate that in ZnCl2-pretre ated HL-60 cells loaded with copper nitrilotriacetate (Cu-NTA), exposure to an NO donor, S-nitroso-N-acetyl penicillamine resulted in S-nitrosylation and oxidation of MT cysteines. This disruption of MT Cu-binding thiolate cl usters caused loosening and release of redox-active Cu, enhanced redox-cycl ing activity of Cu and increased peroxidation of major classes of membrane phospholipids. We also found that Cu-induced oxidative stress in ZnCl2-pret reated/Cu-NTA-loaded HL-60 cells was accompanied by apoptosis documented by characteristic changes of nuclear morphology, internucleosomal DNA cleavag e, externalization of phosphatidylserine, release of cytochrome c from mito chondria into cytosol and activation of caspase-3. We conclude that in Cu-c hallenged cells, NO can reverse the protective role of MTs and convert them into pro-oxidant, pro-apoptotic implements.