Reactive oxygen species-mediated beta-cleavage of the prion protein in the cellular response to oxidative stress.
The cellular prion protein (PrP(C)) is critical for the development of prion diseases. However, the physiological role of PrP(C) is less clear, although a role in the cellular resistance to oxidative stress has been proposed. PrP(C) is cleaved at the end of the copper-binding octapeptide repeats through the action of reactive oxygen species (ROS), a process termed beta-cleavage. Here we show that ROS-mediated beta-cleavage of cell surface PrP(C) occurs within minutes and was inhibited by the hydroxyl radical quencher dimethyl sulfoxide and by an antibody against the octapeptide repeats. A construct of PrP lacking the octapeptide repeats, PrPDeltaoct, failed to undergo ROS-mediated beta-cleavage, as did two mutant forms of PrP, PG14 and A116V, associated with human prion diseases. As compared with cells expressing wild type PrP, when challenged with H2O2 and Cu2+, cells expressing PrPdeltaoct, PG14, or A116V had reduced viability and glutathione peroxidase activity and increased intracellular free radicals. Thus, lack of ROS-mediated beta-cleavage of PrP correlated with the sensitivity of the cells to oxidative stress. These data indicate that the beta-cleavage of PrP(C) is an early and critical event in the mechanism by which PrP protects cells against oxidative stress.
Watt
Nicole T
NT
Proteolysis Research Group, School of Biochemistry and Microbiology, Leeds Institute of Genetics, Health and Therapeutics, University of Leeds, Leeds LS2 9JT, United Kingdom.
Taylor
David R
DR
Gillott
Andrew
A
Thomas
Daniel A
DA
Perera
W Sumudhu S
WS
Hooper
Nigel M
NM
D000818:Animals; D001562:Benzimidazoles / Q000494:pharmacology; D019921:Biotinylation; D015153:Blotting, Western; D002154:Calpain / Q000737:chemistry; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D004396:Coloring Agents / Q000494:pharmacology; D003300:Copper / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004121:Dimethyl Sulfoxide / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis; D019286:Endopeptidase K / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry; D005456:Fluorescent Dyes / Q000494:pharmacology; D005609:Free Radicals; D005979:Glutathione Peroxidase / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D006861:Hydrogen Peroxide / Q000737:chemistry; D047468:Immunoprecipitation; D051379:Mice; D008856:Microscopy, Fluorescence; D009154:Mutation; D018384:Oxidative Stress; D010100:Oxygen / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D011328:Prions / Q000737:chemistry*; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins / Q000737:chemistry; D013997:Time Factors; D015854:Up-Regulation
D001562:Benzimidazoles; D004396:Coloring Agents; D018076:DNA, Complementary; D000939:Epitopes; D005456:Fluorescent Dyes; D005609:Free Radicals; D010455:Peptides; D011328:Prions; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D003300:Copper; D006861:Hydrogen Peroxide; D005979:Glutathione Peroxidase; D019286:Endopeptidase K; D002154:Calpain; C017807:bisbenzimide ethoxide trihydrochloride; D010100:Oxygen; D004121:Dimethyl Sulfoxide