1887

Abstract

This study describes the conversion of murine PrP by PrP from three different mouse scrapie strains (ME7, 87V and 22A) and from a mouse-passaged bovine spongiform encephalopathy (BSE) strain (BSE/Bl6). This was demonstrated by a modified, non-radioactive, cell-free conversion assay using bacterial prion protein, which was converted into a proteinase K (PK)-resistant fragment designated PrP. Using this assay, newly formed PrP could be detected by an antibody that discriminated PrP and the original PrP seed. The results suggested that PrP formation occurs in three phases: the first 48 h when PrP formation is delayed, followed by a period of substantially accelerated PrP formation and a plateau phase when a maximum concentration of PrP is reached after 72 h. The conversion of prokaryotically expressed PrP by ME7 and BSE prions led to unglycosylated, PK-digested, abnormal PrP fragments, which differed in molecular mass by 1 kDa. Therefore, prion strain phenotypes were retained in the cell-free conversion, even when recombinant PrP was used as the substrate. Moreover, co-incubation of ME7 and BSE prions resulted in equal amounts of both ME7- and BSE-derived PrP fragments (as distinguished by their different molecular sizes) and also in a significantly increased total amount of -generated PrP. This was found to be more than twice the amount of either strain when incubated separately. This result indicates a synergistic effect of both strains during cell-free conversion. It is not yet known whether such a cooperative action between BSE and scrapie prions also occurs .

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2006-12-01
2019-11-15
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References

  1. Aguzzi, A. & Polymenidou, M. ( 2004; ). Mammalian prion biology: one century of evolving concepts. Cell 116, 313–327.[CrossRef]
    [Google Scholar]
  2. Baron, T. G. M. & Bicabe, A.-G. ( 2001; ). Molecular analysis of the abnormal prion protein during coinfection of mice by bovine spongiform encephalopathy and a scrapie agent. J Virol 75, 107–114.[CrossRef]
    [Google Scholar]
  3. Bartz, J. C., Bessen, R. A., McKenzie, D., Marsh, R. F. & Aiken, J. M. ( 2000; ). Adaption and selection of prion protein strain conformations following interspecies transmission of transmissible mink encephalopathy. J Virol 74, 5542–5547.[CrossRef]
    [Google Scholar]
  4. Baskakov, I. V. & Bocharova, O. V. ( 2005; ). In vitro conversion of mammalian prion protein into amyloid fibrils displays unusual features. Biochemistry 44, 2339–2348.[CrossRef]
    [Google Scholar]
  5. Bessen, R. A. & Marsh, R. F. ( 1992; ). Biochemical and physical properties of the prion protein from two strains of the transmissble mink encephalopathy agent. J Virol 66, 2096–2101.
    [Google Scholar]
  6. Bessen, R. A., Kocisko, D. A., Raymond, G. J., Nandan, S., Lansbury, P. T. & Caughey, B. ( 1995; ). Non-genetic propagation of strain-specific properties of scrapie prion potein. Nature 375, 698–700.[CrossRef]
    [Google Scholar]
  7. Bian, J., Nazor, K. E., Angers, R., Jernigan, M., Seward, T., Centers, A., Green, M. & Telling, G. C. ( 2006; ). GFP-tagged PrP supports compromised prion replication in transgenic mice. Biochem Biophys Res Commun 340, 894–900.[CrossRef]
    [Google Scholar]
  8. Bocharova, O. V., Breydo, L., Parfenov, A. S., Salnikov, V. V. & Baskakov, I. V. ( 2005; ). In vitro conversion of full-length mammalian prion protein produces amyloid form with physical properties of PrPSc. J Mol Biol 346, 645–659.[CrossRef]
    [Google Scholar]
  9. Bossers, A., Schreuder, B. E., Muileman, I. H., Belt, P. B. & Smits, M. A. ( 1996; ). PrP genotype contributes to determining survival times of sheep with natural scrapie. J Gen Virol 77, 2669–2673.[CrossRef]
    [Google Scholar]
  10. Bossers, A., Belt, P. B. G. M., Raymond, G. J., Caughey, B., de Vries, R. & Smits, M. A. ( 1997; ). Scrapie susceptibility-linked polymorphisms modulate the in vitro conversion of sheep prion protein to protease-resistant forms. Proc Natl Acad Sci U S A 94, 4931–4936.[CrossRef]
    [Google Scholar]
  11. Bossers, A., de Vries, R. & Smits, M. A. ( 2000; ). Susceptibility of sheep for scrapie as assessed by in vitro conversion of nine naturally occurring variants of PrP. J Virol 74, 1407–1414.[CrossRef]
    [Google Scholar]
  12. Bossers, A., Righter, A., de Vries, R. & Smits, M. A. ( 2003; ). In vitro conversion of normal prion protein into pathologic isoforms. Clin Lab Med 23, 227–247.[CrossRef]
    [Google Scholar]
  13. Butler, D. A., Scott, M. R. D., Bockmann, J. M., Borchelt, D. R., Taraboulos, A., Hsiao, K. K., Kingsbury, D. T. & Prusiner, S. B. ( 1988; ). Scrapie-infected murine neuroblastoma cells produce protease-resistant prion proteins. J Virol 62, 1558–1564.
    [Google Scholar]
  14. Callahan, M. A., Xiong, L. & Caughey, B. ( 2001; ). Reversibility of scrapie-associated prion protein aggregation. J Biol Chem 276, 28022–28028.[CrossRef]
    [Google Scholar]
  15. Castilla, J., Saá, P., Hetz, C. & Soto, C. ( 2005; ). In vitro generation of infectious scrapie prions. Cell 121, 195–206.[CrossRef]
    [Google Scholar]
  16. Caughey, B., Horiuchi, M., Demaimay, R. & Raymond, G. J. ( 1999a; ). Assays of protease-resistant prion protein and its formation. Methods Enzymol 309, 122–133.
    [Google Scholar]
  17. Caughey, B., Raymond, G. J., Priola, S. A., Kocisko, D. A., Race, R. E., Bessen, R. A., Lansbury, J. R. & Chesebro, B. ( 1999b; ). Methods for studying prion protein (PrP) metabolism and the formation of protease-resistant PrP in cell culture and cell-free systems. An update Mol Biotechnol 13, 45–55.[CrossRef]
    [Google Scholar]
  18. Chabry, J., Priola, S. A., Wehrly, K., Nishio, J., Hope, J. & Chesebro, B. ( 1999; ). Species-independent inhibition of abnormal prion protein (PrP) formation by a peptide containing a conserved PrP sequence. J Virol 73, 6245–6250.
    [Google Scholar]
  19. Gauczynski, S., Krasemann, S., Bodemer, W. & Weiss, S. ( 2002; ). Recombinant human prion protein mutants huPrP D178N/M129 (FFI) and huPrP+9OR (fCjD) reveal proteinase K resistance. J Cell Sci 115, 4025–4036.[CrossRef]
    [Google Scholar]
  20. Groschup, M. H. & Kuczius, T. ( 2002; ). Die TSE-Erregerstämme. In Prionen und Prionenkrankheiten, pp. 117–131. Edited by B. Hörnlimann, D. Riesner & H. Kretzschmar. Berlin & New York: Walter de Gruyter.
  21. Groschup, M. H., Langeveld, J. & Pfaff, E. ( 1994; ). The major species specific epitope in prion proteins of ruminants. Arch Virol 136, 423–431.[CrossRef]
    [Google Scholar]
  22. Groschup, M. H., Harmeyer, S. & Pfaff, E. ( 1997; ). Antigenic features of prion proteins of sheep and other mammalian species. J Immunol Methods 207, 89–101.[CrossRef]
    [Google Scholar]
  23. Groschup, M. H., Junghans, F., Eiden, M. & Kuczius, T. ( 2001; ). Characterization of bovine spongiform encephalopathy and scrapie strains/isolates by immunochemical analysis of PrPSc. In Molecular Pathology of the Prions. Methods in Molecular Medicine, vol. 59, pp. 71–83. Edited by H. F. Baker. Totowa, NJ: Humana Press.
  24. Harmeyer, S., Pfaff, E. & Groschup, M. H. ( 1998; ). Synthetic vaccines yield monoclonal antibodies to cellular and pathological prion proteins of ruminants. J Gen Virol 79, 937–945.
    [Google Scholar]
  25. Hayashi, H. K., Yokoyama, T., Takata, M., Iwamaru, Y., Imamura, M., Ushiki, Y. K. & Shinagawa, M. ( 2005; ). The N-terminal cleavage site of PrPSc from BSE differs from that of PrPSc from scrapie. Biochem Biophys Res Commun 328, 1024–1027.[CrossRef]
    [Google Scholar]
  26. Horiuchi, M., Priola, S. A., Chabry, J. & Caughey, B. ( 2000; ). Interactions between heterologous forms of prion protein: binding, inhibition of conversion, and species barriers. Proc Natl Acad Sci U S A 97, 5836–5841.[CrossRef]
    [Google Scholar]
  27. Kascsak, R. J., Rubenstein, R., Merz, P. A., Carp, R. I., Robakis, N. K., Wisniewski, H. M. & Diringer, H. ( 1986; ). Immunological comparison of scrapie-associated fibrils isolated from animals infected with four different scrapie strains. J Virol 59, 676–683.
    [Google Scholar]
  28. Kimberlin, R. H. & Walker, C. A. ( 1985; ). Competition between strains of scrapie depends on the blocking agent being infectious. Intervirology 23, 74–81.[CrossRef]
    [Google Scholar]
  29. Kirby, L., Birkett, C. R. H., Rudyk, H., Gilbert, I. H. & Hope, J. ( 2003; ). In vitro cell-free conversion of bacterial recombinant PrP to PrPres as a model for conversion. J Gen Virol 84, 1013–1020.[CrossRef]
    [Google Scholar]
  30. Kocisko, D. A., Come, J. H., Priola, S. A., Chesebro, B., Raymond, G. J., Lansbury, P. T. & Caughey, B. ( 1994; ). Cell-free formation of protease-resistant prion protein. Nature 370, 471–474.[CrossRef]
    [Google Scholar]
  31. Kocisko, D. A., Priola, S. A., Raymond, G. J., Chesebro, B., Lansbury, P. T., Jr & Caughey, B. ( 1995; ). Species specificity in the cell-free conversion of prion protein to protease-resistant forms: a model for the scrapie species barrier. Proc Natl Acad Sci U S A 92, 3923–3927.[CrossRef]
    [Google Scholar]
  32. Kuczius, T. & Groschup, M. H. ( 1999; ). Differences in proteinase K resistance and neuronal deposition of abnormal prion proteins characterize bovine spongifrom encephalopathy (BSE) and scrapie strains. Mol Med 5, 406–418.
    [Google Scholar]
  33. Mulcahy, E. R. & Bessen, R. A. ( 2004; ). Strain-specific kinetics of prion protein formation in vitro and in vivo. J Biol Chem 279, 1643–1649.[CrossRef]
    [Google Scholar]
  34. Prusiner, S. B. ( 1984; ). Prions: novel infectious pathogens. Adv Virus Res 29, 1–56.
    [Google Scholar]
  35. Race, R. E., Fadness, L. H. & Chesebro, B. ( 1987; ). Characterization of scrapie infection in mouse neuroblastoma cells. J Gen Virol 68, 1391–1399.[CrossRef]
    [Google Scholar]
  36. Raymond, G. J., Hope, J., Kocisko, D. A. & 12 other authors ( 1997; ). Molecular assessment of the transmissibilities of BSE and scrapie to humans. Nature 388, 285–288.[CrossRef]
    [Google Scholar]
  37. Raymond, G. J., Bossers, A., Raymond, L. D. & 7 other authors ( 2000; ). Evidence of a molecular barrier limiting susceptibility of humans, cattle and sheep to chronic wasting disease. EMBO J 19, 4425–4430.[CrossRef]
    [Google Scholar]
  38. Saborio, G. P., Permanne, B. & Soto, C. ( 2001; ). Sensitive detection of pathological prion protein by cyclic amplification of protein misfolding. Nature 411, 810–813.[CrossRef]
    [Google Scholar]
  39. Silveira, J. R., Raymond, G. J., Hughson, A. G., Race, R. E., Sim, V. L., Hayes, S. F. & Caughey, B. ( 2005; ). The most infectious prion protein particles. Nature 437, 257–261.[CrossRef]
    [Google Scholar]
  40. Telling, G. C., Tremblay, P., Torchia, M., DeArmond, S. J., Cohen, F. E. & Prusiner, S. B. ( 1997; ). N-terminally tagged prion protein supports prion propagation in transgenic mice. Protein Sci 6, 825–833.
    [Google Scholar]
  41. Vorberg, I. & Priola, S. A. ( 2002; ). Molecular basis of scrapie strain glycoform variation. J Biol Chem 277, 36775–36781.[CrossRef]
    [Google Scholar]
  42. Vorberg, I., Pfaff, E. & Groschup, M. H. ( 2000; ). The use of monoclonal antibody epitopes for tagging PrP in conversion experiments. Arch Virol Suppl, 285–290.
    [Google Scholar]
  43. Zahn, R., von Schroetter, C. & Wuthrich, K. ( 1997; ). Human prion proteins expressed in Escherichia coli and purified by high-affinity column refolding. FEBS Lett 417, 400–404.[CrossRef]
    [Google Scholar]
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