title: Cell-lysate conversion of prion protein into its protease-resistant isoform suggests the participation of a cellular chaperone.
abstract: A conformational transition between the normal cellular prion protein (PrPC) and the beta-sheet-rich pathological isoform (PrPSc) is a central event in the pathogenesis of spongiform encephalopathies. The prion infectious agent seems to contain mainly, if not exclusively, PrPSc, which has the ability to propagate its abnormal conformation by transforming the host PrPC into the pathological isoform. We have developed an in vitro system to induce the PrPC --> PrPSc conversion by incubating a cell-lysate containing mouse PrPC with partially purified mouse PrPSc. After 48 h of incubation with a 10-fold molar excess of PrPSc, the cellular protein acquired PK-resistance resembling a PrPSc-like state. Time course experiments suggest that the conversion follows a stepwise mechanism involving kinetic intermediates. The conversion was induced by PrPSc extracted from mice infected with two different prion strains, each propagating its characteristic Western blot profile. The latter results and the fact that all the cellular components are present in the conversion reaction suggest that PrPC-PrPSc interaction is highly specific and required for the conversion. No transformation was observed under the same conditions using purified proteins without cell-lysate. However, when PrPC-depleted cell-lysate was added to the purified proteins the conversion was recovered. These findings provide direct evidence for the participation of a chaperone-like activity involved in catalyzing the conversion of PrPC into PrPSc.
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D010450:Endopeptidases / Q000378:metabolism*; D051379:Mice; D018832:Molecular Chaperones / Q000378:metabolism*; D011328:Prions / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism*