Abstract

Prion diseases propagate by converting a normal glycoprotein of the host, PrP^C, into a pathogenic 'prion' conformation. Several misfolding mutants of PrP^C are degraded through the ER-associated degradation (ERAD)–proteasome pathway. In their infectious form, prion diseases such as bovine spongiform encephalopathy involve PrP^C of wild-type sequence. In contrast to mutant PrP, wild-type PrP^C was hitherto thought to be stable in the ER and thus immune to ERAD. Using proteasome inhibitors, we now show that 10% of nascent PrP^C molecules are diverted into the ERAD pathway. Cells incubated with N-acetyl-leucinal-leucinal-norleucinal (ALLN), lactacystin or MG132 accumulated both detergent-soluble and insoluble PrP species. The insoluble fraction included an unglycosylated 26 kDa PrP species with a protease-resistant core, and a M_r 'ladder' that contained ubiquitylated PrP. Our results show for the first time that wild-type PrP^C molecules are subjected to ERAD, in the course of which they are dislocated into the cytosol and ubiquitylated. The presence of wild-type PrP molecules in the cytosol may have potential pathogenic implications.