1-Cys peroxiredoxin overexpression protects cells against phospholipid peroxidation-mediated membrane damage.
1-Cys peroxiredoxin (1-cysPrx) is a novel antioxidant enzyme able to reduce phospholipid hydroperoxides in vitro by using glutathione as a reductant. This enzyme is widely expressed and is enriched in lungs. A fusion protein of green fluorescent protein with 1-cysPrx was stably expressed in a lung-derived cell line (NCI-H441) lacking endogenous enzyme. Overexpressing cells (C17 or C48) degraded H(2)O(2) and t-butylhydroperoxide more rapidly and showed decreased sensitivity to oxidant stress as measured by (51)Cr release. On exposure to (*)OH generated by Cu(2+)-ascorbate (Asc), overexpressing cells compared with H441 showed less increase in thiobarbituric acid-reactive substance and phosphatidylcholine hydroperoxide content. This effect was reversed by depletion of cellular glutathione. Diphenyl-1-pyrenoylphosphonium fluorescence, used as a real-time probe of membrane phospholipid peroxidation, increased immediately on exposure to Cu(2+)-Asc and was abolished by preincubation of cells with Trolox (a soluble vitamin E) or Tempol (a radical scavenger). The rate of diphenyl-1-pyrenoylphosphonium fluorescence increase with Cu(2+)-Asc exposure was markedly attenuated in C17 and C48 cells as compared with H441. Annexin V-Cy3 was used to detect phosphatidylserine translocation from the inner to outer leaflet of the plasma membrane. Cu(2+)-Asc treatment induced phosphatidylserine translocation within 2 h in H441 cells but none was observed in C48 cells up to 24 h. These results indicate that 1-cysPrx can scavenge peroxides but in addition can reduce peroxidized membrane phospholipids. Thus, the enzyme can protect cells against oxidant-induced plasma membrane damage, thereby playing an important role in cellular defense against oxidant stress.
Institute for Environmental Medicine, University of Pennsylvania Medical Center, Philadelphia, PA 19104, USA.
Yefim
Y
Manevich
Tom
T
Sweitzer
Jhang Ho
JH
Pak
Sheldon I
SI
Feinstein
Vladimir
V
Muzykantov
Aron B
AB
Fisher
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D017931:DNA Primers; D015227:Lipid Peroxidation; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010544:Peroxidases / Q000737:chemistry / Q000378:metabolism*; D054464:Peroxiredoxins; D010743:Phospholipids / Q000378:metabolism*; D051381:Rats; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism
D017931:DNA Primers; D010743:Phospholipids; D017392:Thiobarbituric Acid Reactive Substances; D010544:Peroxidases; D054464:Peroxiredoxins