PrP C -related signal transduction is influenced by copper, membrane integrity and the alpha cleavage site

Abstract

The copper-binding, membrane-anchored, cellular prion protein (PrP C) has two constitutive cleavage sites producing distinct N- and C-terminal fragments (N1/C1 and N2/C2). Using RK13 cells expressing either human PrP C, mouse PrP C or mouse PrP C carrying the 3F4 epitope, this study explored the influence of the PrP C primary sequence on endoproteolytic cleavage and one putative PrP C function, MAP kinase signal transduction, in response to exogenous copper with or without a perturbed membrane environment. PrP C primary sequence, especially that around the N1/C1 cleavage site, appeared to influence basal levels of proteolysis at this location and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation, with increased processing demonstrating an inverse relationship with basal ERK1/2 activation. Human PrP C showed increased N1/C1 cleavage in response to copper alone, accompanied by specific p38 and JNK/SAPK phosphorylation. Combined exposure to copper plus the cholesterol-sequestering antibiotic filipin resulted in a mouse PrP C -specific substantial increase in signal protein phosphorylation, accompanied by an increase in N1/C1 cleavage. Mouse PrP C harboring the human N1/C1 cleavage site assumed more human-like profiles basally and in response to copper and altered membrane environments. Our results demonstrate that the PrP C primary sequence around the N1/C1 cleavage site influences endoproteolytic processing at this location, which appears linked to MAP kinase signal transduction both basally and in response to copper. Further, the primary sequence appears to confer a mutual dependence of N1/C1 cleavage and membrane integrity on the fidelity of PrP C -related signal transduction in response to exogenous stimuli. © 2009 IBCB, SIBS, CAS All rights reserved.