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Physiological role of PrP in anti-oxidant defence 3

To investigate the role of PrP and domains within the protein in the cellular response to oxidative stress we help different partner generating neuronal cells stably transfected with constructs of PrP with deletions, mutations or insertions in the octapeptide repeat region and elsewhere in the N-terminus. These cell lines are being challenged with hydrogen peroxide in the absence and presence of copper ions, and the cell viability, intracellular radical formation, and the activities of superoxide dismutase and glutathione peroxidase measured.

We investigated with the other partners the effects of cellular prion protein (PrPC) overexpression on paraquat-induced toxicity by using our established model rabbit kidney epithelial A74 cells, which express a doxycycline-inducible murine PrPC gene. PrPC overexpression was found to significantly reduce paraquat-induced cell toxicity, DNA damage, and malondialdehyde acid levels. Superoxide dismutase (total SOD and CuZn-SOD) and glutathione peroxidase activities were higher in doxycycline-stimulated cells. Our findings clearly show that PrPC overexpression plays a protective role against paraquat toxicity, probably by virtue of its superoxide dismutase-like activity (Senator et al).

We participated to a work with a Japanese group were copper was quantified using Zeeman graphite furnace atomic absorption spectroscopy, in immortalised PrP gene (Prnp)-deficient neuronal cells transfected with Prnp and/or Prnd, which encodes PrP-like protein (PrPLP/Dpl), in the presence or absence of oxidative stress induced by serum deprivation. In the presence of serum, copper levels were not significantly affected by the expression of PrP and/or PrPLP/Dpl, whereas serum deprivation induced a decrease in copper levels that was inhibited by PrP but not by PrPLP/Dpl. The inhibitory effect of PrP on the decrease of copper levels was prevented by overexpression of PrPLP/Dpl. These findings indicate that PrP specifically stabilises copper homeostasis, which is perturbed under oxidative conditions, while PrPLP/Dpl overexpression prevents PrP function in copper homeostasis, suggesting an interaction of PrP and PrPLP/Dpl and distinct functions between PrP and PrPLP/Dpl on metal homeostasis. Taken together, these results strongly suggest that PrP, in addition to its antioxidant properties, plays a role in stabilising cellular copper homeostasis under oxidative conditions.

Senator A., Rachidi W., Lehmann S., Favier A. & Benboubetra M. (2004) Prion protein protects against DNA damage induced by Paraquat in cultured cells. Free Radical Biology & Medicine. 37, 1224-30.

Sakudo A, Lee D, Yoshimura E, Nagasaka S, Nitta K, Saeki K, Matsumoto Y, Lehmann S, Itohara S, Sakaguchi S, & Onodera T. (2004) Prion protein suppresses perturbation of cellular copper homeostasis under oxidative conditions. Biochem Biophys Res Commun. 313, 850-5.

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