<p> Prion protein protects against zinc-mediated cytotoxicity by modifying intracellular exchangeable zinc compartmentation in cultured cells. </p><p>Walid Rachidi1, Fabrice Chimienti2, M’hammed Aouffen1, Abderrahmane Senator3, Pascale Guiraud3, Michel Seve4 and Alain Favier#1 </p><p>1 DRFMC/SCIB/LAN, CEA de Grenoble, 17, rue des Martyrs 38054, Grenoble, France. 2 MELLITECH SAS France, 17, rue des Martyrs 38054 Grenoble, France 3 ORSOX, Faculté de Pharmacie, Domaine de la Merci, 38706 La Tronche-Grenoble Cedex 9 4 Centre d’Innovation en Biologie, CHU Grenoble, BP 217, 38043 Grenoble, France </p><p>Summary </p><p>PrPC contains several octapeptide repeats sequences toward the N-terminus which have binding affinity for divalent metals such as copper, zinc, nickel and manganese. However, up to now the link between PrPC expression and zinc metabolism remains elusive. In this study, we have explored in detail the relationship between PrPC and zinc ions using a cell line expressing a doxycycline-inducible PrPC gene. Radioactive zinc (65Zn2+) was used to explore the impact of PrPC expression on cellular zinc uptake. No significant difference in 65Zn2+ uptake was observed in cells expressing PrPC when compared with control cells. This result was confirmed by flow cytometry analysis, using the zinc-specific probe zinpyr-1, which shows no global difference in their intracellular exchangeable zinc content. Furthermore, spectrofluorometry quantification of total zinc corroborates the absence of PrPC expression impact on the zinc accumulation. However, PrPC-expressing cells were more resistant to zinc-induced toxicity, suggesting an adaptative mechanism induced by PrPC. Using zinquin-ethyl-ester, a specific fluorophore for vesicular free zinc, we observed a significant re-localization of intracellular exchangeable zinc in vesicles after PrPC expression. Finally, we demonstrated that PrPC expression induces metallothionein expression, a zinc-upregulated zinc-binding protein. Taken together, these results suggest that PrPC modifies the intracellular localization of zinc rather than the cellular content. Neuronal zinc dyshomeostasis appears to be a common phenomenon in several neurodegenerative disorders, and prion protein may play a crucial role in zinc homeostasis within the brain. </p>