The scrapie-associated form of PrP is made from a cell surface precursor that is both protease- and phospholipase-sensitive.
A common feature of scrapie and related transmissible spongiform encephalopathies is the accumulation of an abnormal protease-resistant form of PrP which may be the major component of the infectious agent. While it is known that both the normal (protease-sensitive) PrP and protease-resistant PrP are encoded by the same endogenous gene, the nature of the disease-associated modification of PrP is not understood. To study the cellular events leading to the formation of protease-resistant PrP, we have compared its biosynthesis to that of its normal isoform in scrapie-infected mouse neuroblastoma cells. In pulse-chase labeling experiments, the protease-resistant PrP was synthesized and degraded much more slowly than the normal PrP, suggesting that protease-resistant PrP is made from a protease-sensitive precursor. More significantly, we found that the precursor of protease-resistant PrP was eliminated from intact cells by treatments with phosphatidylinositol-specific phospholipase C and trypsin. This demonstrated that, unlike the protease-resistant PrP itself, the precursor is phospholipase- and protease-sensitive and at least transiently found on the cell surface. By these criteria, the precursor of protease-resistant PrP is indistinguishable from the normal PrP isoform. These results indicate that the conversion of PrP to the protease- and phospholipase-resistant state is a post-translational event that occurs after the precursor reaches the cell surface.
Caughey
B
B
National Institutes of Health, National Institute of Allergy and Infectious Diseases, Rocky Mountain Laboratories, Hamilton, Montana 59840.
Raymond
G J
GJ
D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D009447:Neuroblastoma / Q000382:microbiology / Q000473:pathology; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010740:Phospholipases / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D018620:PrPSc Proteins; D011328:Prions / Q000096:biosynthesis / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D014357:Trypsin; D014407:Tumor Cells, Cultured
D008565:Membrane Proteins; D018620:PrPSc Proteins; D011328:Prions; D011498:Protein Precursors; D010740:Phospholipases; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D014357:Trypsin; D043265:Phosphatidylinositol Diacylglycerol-Lyase