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Crapie and cellular prion proteins differ in their kinetics of synthesis and topology in cultured cells

  • Author(s): Borchelt, DR
  • Scott, M
  • Taraboulos, A
  • Stahl, N
  • Prusiner, SB
  • et al.

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Both the cellular and scrapie isoforms of the prion protein (PrP) designated PrPcand PrPScare encoded by a single-copy chromosomal gene and appear to be translated from the same 2.1-kb mRNA. PrPccan be distinguished from PrPScby limited proteolysis under conditions where PrPcis hydrolyzed and PrPScis resistant. We report here that PrPccan be released from the surface of both normal-control and scrapie-infected murine neuroblastoma (N2a) cells by phosphatidylinositol-specific phospholipase C (PIPLC) digestion and it can be selectively labeled with sulfoNHS-biotin, a membrane impermeant reagent. In contrast, PrPScwas neither released by PIPLC nor labeled with sulfo-NHS-biotin. Pulse-chase experiments showed that [35S]methionine was incorporated almost immediately into PrPcwhile incorporation into PrPScmolecules was observed only during the chase period. While PrPcis synthesized and degraded relatively rapidly (t1/2∼ 5 h), PrPScis synthesized slowly (t1/2∼ 15 h) and appears to accumulate. These results are consistent with several observations previously made on rodent brains where PrP mRNA and PrPclevels did not change throughout the course of scrapie infection, yet PrPScaccumulated to levels exceeding that of PrPc. Our kinetic studies demonstrate that PrPScis derived from a protease-sensitive precursor and that the acquisition of proteinase K resistance results from a posttranslational event. Whether or not prolonged incubation periods, which are a cardinal feature of prion diseases, reflect the slow synthesis of PrPScremains to be established.

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