Characterization of detergent-insoluble complexes containing the cellular prion protein and its scrapie isoform.
Cells infected with prions contain both prion protein isoforms cellular prion protein (PrPC) and scrapie prion protein (PrPSc). PrPSc is formed posttranslationally through the pathological refolding of PrPC. In scrapie-infected ScN2a cells, the metabolism of both PrP isoforms involves cholesterol-dependent pathways. We show here that both PrPC and PrPSc are attached to Triton X-100-insoluble, low-density complexes or "rafts." These complexes are sensitive to saponin and thus probably contain cholesterol. This finding suggests that the transformation PrPC --> PrPSc occurs within rafts. It also reveals the existence of rafts in late compartments of the endocytic pathway, where most PrPSc resides. When Triton X-100 lysates of cells were incubated at 37 degrees C prior to density analysis, PrPC was still found in buoyant complexes, although it now failed to sediment at high speed. This property was shared by another glycophosphatidyl inositol protein, Thy-1, and also by the raft resident GM1. In one ScN2a clone and in the brain of a Syrian hamster with scrapie, Triton X-100 extraction at 37 degrees C permitted resolution of PrPC and PrPSc into two distinct peaks of different densities. This suggests that there are two populations of PrP-containing rafts and may permit isolation of PrPC-specific rafts from those containing PrPSc. Our findings reinforce the contention that rafts are involved in various aspects of PrP metabolism and in the "life cycle" of prions.
Naslavsky
N
N
Department of Molecular Biology, the Hebrew University-Hadassah Medical School, P. O. Box 12272, Jerusalem 91120, Israel.
Stein
R
R
Yanai
A
A
Friedlander
G
G
Taraboulos
A
A
D000818:Animals; D002784:Cholesterol / Q000737:chemistry; D006224:Cricetinae; D003902:Detergents; D005677:G(M1) Ganglioside / Q000737:chemistry; D017261:Glycosylphosphatidylinositols; D008647:Mesocricetus; D051379:Mice; D011328:Prions / Q000737:chemistry*; D012503:Saponins / Q000737:chemistry; D012608:Scrapie; D012995:Solubility; D018800:Thy-1 Antigens / Q000737:chemistry
D003902:Detergents; D017261:Glycosylphosphatidylinositols; D011328:Prions; D012503:Saponins; D018800:Thy-1 Antigens; D005677:G(M1) Ganglioside; D002784:Cholesterol