Abstract
Human peroxiredoxins serve dual roles as anti-oxidants and regulators of H(2)O(2)-mediated cell signaling. The functional versatility of peroxiredoxins depends on progressive oxidation of key cysteine residues. The sulfinic or sulfonic forms of peroxiredoxin lose their peroxidase activity, which allows cells to accumulate H(2)O(2) for signaling or pathogenesis in inflammation, cancer, and other disorders. We report that arachidonic acid lipid hydroperoxide metabolites of 5-, 12-, 15-lipoxygenase-1, and cyclooxygenase-2 oxidize the 2-Cys-peroxiredoxins 1, 2, and 3 to their sulfinic and sulfonic forms. When added exogenously to cells, 5-, 12- and 15-hydroperoxy-eicosatetraenoic acids also over-oxidized peroxiredoxins. Our results suggest that lipoxygenases and cyclooxygenases may affect 2-Cys peroxiredoxin signaling, analogous to NADPH oxidases in the "floodgate" model (Wood, Z. A., Poole, L. B, and Karplus P. A. (2003) Science 300, 600-653). Peroxiredoxin-dependent mechanisms may modulate the receptor-dependent actions of autocoids derived from cellular lipoxygenase and cyclooxygenase catalysis.
Highlights
Switch between an ancestral anti-oxidant role and a more intricate, acquired role as a transducer of H2O2 redox signaling (12– 15)
We report that lipid hydroperoxide metabolites of arachidonic acid (HpETEs) formed by cellular 5-LOX, 12-LOX, 15-LOX-1, and COX-2 oxidize Prx1, 2, and 3 to PrxSO2/SO3
Oxidation of Prx to PrxSO3 by LOX and COX-2 Catalysis of Exogenous Arachidonic acid (AA)—After 48 h, the medium consisting of 10% v/v fetal calf serum (FCS), 1ϫ Dulbecco’s modified eagle’s medium (DMEM) with 10 M ponasterone was removed from cells expressing 5-LOX, 12-LOX, 15-LOX-1, or ⌬Ile662 15-LOX-1 and replaced with 1% FCS, DMEM with 10 M ponasterone
Summary
Materials—The COX inhibitor, aspirin, the LOX inhibitor, nordihydroguauretic acid (NDGA), and porcine pancreatic phospholipase A2 (PLA2) were from Sigma. Oxidation of Prx to PrxSO3 by LOX and COX-2 Catalysis of Exogenous AA—After 48 h, the medium consisting of 10% v/v FCS, 1ϫ DMEM with 10 M ponasterone was removed from cells expressing 5-LOX, 12-LOX, 15-LOX-1, or ⌬Ile662 15-LOX-1 and replaced with 1% FCS, DMEM with 10 M ponasterone. Oxidation of Prx to PrxSO3 in Cells Treated with Phospholipase A2—293 EcR cells expressing 15-LOX-1 or 12-LOX were incubated for 30, 60, and 90 min in 1% v/v FCS and DMEM with 20 units/ml pancreatic PLA2, to determine whether metabolism of endogenous fatty acids released from cellular phospholipids leads to PrxSO3 formation. B, immunoblots of 15-LOX-1, Prx, and PrxSO3 protein (arrows) in 293 EcR cells harboring inducible inactive mutant ⌬ ile662 15-LOX-1 (lanes 1– 4) or wild type 15-LOX-1 (lanes 5–10) treated with 10 M ponasterone for 48 h (lanes 1, 2, and 5–10), incubated with 60 M AA (lanes 2, 4, and 5–10).
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