Investigation of PrPC metabolism and function in live cells

Abstract

Summary: Prion protein (PrP)C expression levels and protein localization are known to be affected by factors such as metal ions and oxidative stress. By the development of a green fluorescent protein (GFP)-PrPC fusion protein, the movement of PrP can be followed in real time. Furthermore, alterations in cellular metabolism can be detected while cells are still viable. The internalization response of PrP to 20 μM manganese (Mn) in divalent metal ion-depleted media is used to demonstrate the movement of GFP-tagged proteins in live cells and real tim0e. A live cell microtiter plate assay shows that PrP null cells are less capable of dealing with Mn-induced oxidative stress. In addition, this chapter outlines several complementary techniques for studying live cells and GFP fusion proteins. © 2008 Humana Press, a part of Springer Science + Business Media, LLC.