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Molecular Diagnosis of Human Prion Disease

  • Protocol
Book cover Prion Protein Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 459))

Summary

Human prion diseases are associated with a range of clinical presentations, and they are classified by both clinicopathological syndrome and etiology, with subclassification according to molecular criteria. Here, we describe procedures that are used within the MRC Prion Unit to determine a molecular diagnosis of human prion disease. Sequencing of the PRNP open reading frame to establish the presence of pathogenic mutations is described, together with detailed methods for immunoblot or immunohistochemical determination of the presence of abnormal prion protein in brain or peripheral tissues.

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References

  1. Prusiner SB. (1998) Prions. Proc Natl Acad Sci U S A;95:13363–13383.

    Article  CAS  PubMed  Google Scholar 

  2. Collinge J. (2001) Prion diseases of humans and animals: their causes and molecular basis. Annu Rev Neurosci;24:519–550.

    Article  CAS  PubMed  Google Scholar 

  3. Collinge J. (2005) Molecular neurology of prion disease. J Neurol Neurosurg Psychiatry; 76:906–919.

    Article  CAS  PubMed  Google Scholar 

  4. Wadsworth JD, Hill AF, Beck JA, Collinge J. (2003) Molecular and clinical classification of human prion disease. Br Med Bull;66:241–254.

    Article  CAS  PubMed  Google Scholar 

  5. Wadsworth JD, Collinge J. (2007) Update on human prion disease. Biochim Biophys Acta; 1772:598–609.

    CAS  PubMed  Google Scholar 

  6. Weissmann C. (2004) The state of the prion. Nat Rev Microbiol;2:861–871.

    Article  CAS  PubMed  Google Scholar 

  7. Telling GC, Parchi P, DeArmond SJ, et al. (1996) Evidence for the conformation of the pathologic isoform of the prion protein enciphering and propagating prion diversity. Science;274:2079–2082.

    Article  CAS  PubMed  Google Scholar 

  8. Collinge J, Sidle KCL, Meads J, Ironside J, Hill AF. (1996) Molecular analysis of prion strain variation and the aetiology of ‘new variant’ CJD. Nature;383:685–690.

    Article  CAS  PubMed  Google Scholar 

  9. Parchi P, Castellani R, Capellari S, et al. (1996) Molecular basis of phenotypic variability in sporadic Creutzfeldt-Jakob disease. Ann Neurol;39:767–778.

    Article  CAS  PubMed  Google Scholar 

  10. Wadsworth JDF, Hill AF, Joiner S, Jackson GS, Clarke AR, Collinge J. (1999) Strain-specific prion-protein conformation determined by metal ions. Nat Cell Biol;1:55–59.

    Article  CAS  PubMed  Google Scholar 

  11. Parchi P, Giese A, Capellari S, et al. (1999) Classification of sporadic Creutzfeldt-Jakob Disease based on molecular and phenotypic analysis of 300 subjects. Ann Neurol; 46:224–233.

    Article  CAS  PubMed  Google Scholar 

  12. Hill AF, Joiner S, Wadsworth JD, et al. (2003) Molecular classification of sporadic Creutzfeldt-Jakob disease. Brain;126:1333–1346.

    Article  PubMed  Google Scholar 

  13. Zanusso G, Farinazzo A, Prelli F, et al. (2004) Identification of distinct N-terminal truncated forms of prion protein in different Creutzfeldt-Jakob disease subtypes. J Biol Chem;79: 38936–38942.

    Article  Google Scholar 

  14. Safar JG, Geschwind MD, Deering C, et al. (2005) Diagnosis of human prion disease. Proc Natl Acad Sci U S A;102:3501–3506.

    Article  CAS  PubMed  Google Scholar 

  15. Hill A, Joiner S, Beck J, et al. (2006) Distinct glycoform ratios of protease resistant prion protein associated with PRNP point mutations. Brain;129:676–685.

    Article  PubMed  Google Scholar 

  16. Lloyd SE, Onwuazor ON, Beck JA, et al. (2001) Identification of multiple quantitative trait loci linked to prion disease incubation period in mice. Proc Natl Acad Sci U S A;98:6279–6283.

    Article  CAS  PubMed  Google Scholar 

  17. Asante EA, Linehan JM, Desbruslais M, et al. (2002) BSE prions propagate as either variant CJD-like or sporadic CJD-like prion strains in transgenic mice expressing human prion protein. EMBO J;21:6358–6366.

    Article  CAS  PubMed  Google Scholar 

  18. Collinge J. (1997) Human prion diseases and bovine spongiform encephalopathy (BSE). Hum Mol Genetics;6:1699–1705.

    Article  CAS  Google Scholar 

  19. Collinge J, Palmer.M.S. (1997) Prion Diseases. 1st ed. Oxford: Oxford University Press.

    Google Scholar 

  20. Brown P, Cathala F, Raubertas RF, Gajdusek DC, Castaigne P. (1987) The epidemiology of Creutzfeldt-Jakob disease: conclusion of a 15-year investigation in France and review of the world literature. Neurology;37:895–904.

    CAS  PubMed  Google Scholar 

  21. Collins SJ, Sanchez-Juan P, Masters CL, et al. (2006) Determinants of diagnostic investigation sensitivities across the clinical spectrum of sporadic Creutzfeldt-Jakob disease. Brain;129:2278–2287.

    Article  CAS  PubMed  Google Scholar 

  22. Kovacs GG, Trabattoni G, Hainfellner JA, Ironside JW, Knight RS, Budka H. (2002) Mutations of the prion protein gene phenotypic spectrum. J Neurol; 249:1567–1582.

    Article  CAS  PubMed  Google Scholar 

  23. Mead S. (2006) Prion disease genetics. Eur J Hum Genet;14:273–281.

    Article  CAS  PubMed  Google Scholar 

  24. Brown P, Preece MA, Will RG. (1992) “Friendly fire” in medicine: hormones, homografts, and Creutzfeldt-Jakob disease. Lancet;340:24–27.

    Article  CAS  PubMed  Google Scholar 

  25. Brown P, Preece M, Brandel JP, et al. (2000) Iatrogenic Creutzfeldt-Jakob disease at the millennium. Neurology;55:1075–1081.

    CAS  PubMed  Google Scholar 

  26. Alpers MP. Epidemiology and clinical aspects of kuru. (1987) In: Prusiner SB, McKinley MP, editors. Prions: Novel Infectious Pathogens Causing Scrapie and Creutzfeldt-Jakob Disease. San Diego: Academic Press, 451–465.

    Google Scholar 

  27. Mead S, Stumpf MP, Whitfield J, et al. (2003) Balancing selection at the prion protein gene consistent with prehistoric kurulike epidemics. Science;300:640–643.

    Article  CAS  PubMed  Google Scholar 

  28. Collinge J, Whitfield J, McKintosh E, et al. (2006) Kuru in the 21st century-an acquired human prion disease with very long incubation periods. Lancet;367:2068–2074.

    Article  PubMed  Google Scholar 

  29. Will RG, Ironside JW, Zeidler M, et al. (1996) A new variant of Creutzfeldt-Jakob disease in the UK. Lancet;347:921–925.

    Article  CAS  PubMed  Google Scholar 

  30. Hill AF, Desbruslais M, Joiner S, et al. (1997) The same prion strain causes vCJD and BSE. Nature;389:448–450.

    Article  CAS  PubMed  Google Scholar 

  31. Bruce ME, Will RG, Ironside JW, et al. (1997) Transmissions to mice indicate that ‘new variant’ CJD is caused by the BSE agent. Nature;389:498–501.

    Article  CAS  PubMed  Google Scholar 

  32. Collinge J. (1999) Variant Creutzfeldt-Jakob disease. Lancet;354:317–323.

    Article  CAS  PubMed  Google Scholar 

  33. Hilton DA, Ghani AC, Conyers L, et al. (2004) Prevalence of lymphoreticular prion protein accumulation in UK tissue samples. J Pathol;203:733–739.

    Article  CAS  PubMed  Google Scholar 

  34. Frosh A, Smith LC, Jackson CJ, et al. (2004) Analysis of 2000 consecutive UK tonsillectomy specimens for disease-related prion protein. Lancet;364:1260–1262.

    Article  PubMed  Google Scholar 

  35. Hilton DA. (2005) Pathogenesis and prevalence of variant Creutzfeldt-Jakob disease. J Pathol;208:134–141.

    Article  Google Scholar 

  36. Wadsworth JDF, Joiner S, Hill AF, et al. (2001) Tissue distribution of protease resistant prion protein in variant CJD using a highly sensitive immuno-blotting assay. Lancet;358:171–180.

    Article  CAS  PubMed  Google Scholar 

  37. Llewelyn CA, Hewitt PE, Knight RS, et al. (2004) Possible transmission of variant Creutzfeldt-Jakob disease by blood transfusion. Lancet;363:417–421.

    Article  CAS  PubMed  Google Scholar 

  38. Peden AH, Head MW, Ritchie DL, Bell JE, Ironside JW. (2004) Preclinical vCJD after blood transfusion in a PRNP codon 129 heterozygous patient. Lancet; 364:527–529.

    Article  PubMed  Google Scholar 

  39. Wroe SJ, Pal S, Siddique D, et al. (2006) Clinical presentation and pre-mortem diagnosis of variant Creutzfeldt-Jakob disease associated with blood transfusion: a case report. Lancet;368:2061–2067.

    Article  PubMed  Google Scholar 

  40. Collinge J, Palmer MS, Dryden AJ. (1991) Genetic predisposition to iatrogenic Creutzfeldt-Jakob disease. Lancet;337:1441–1442.

    Article  CAS  PubMed  Google Scholar 

  41. Palmer MS, Dryden AJ, Hughes JT, Collinge J. (1991) Homozygous prion protein genotype predisposes to sporadic Creutzfeldt-Jakob disease. Nature; 352:340–342.

    Article  CAS  PubMed  Google Scholar 

  42. Windl O, Dempster M, Estibeiro JP, et al. (1996) Genetic basis of Creutzfeldt-Jakob disease in the United Kingdom: a systematic analysis of predisposing mutations and allelic variation in the PRNP gene. Hum Genet;98:259–264.

    Article  CAS  PubMed  Google Scholar 

  43. Lee HS, Brown P, Cervenáková L, et al. (2001) Increased susceptibility to Kuru of carriers of the PRNP 129 methionine/methionine genotype. J Infect Dis;183:192–196.

    Article  CAS  PubMed  Google Scholar 

  44. Wadsworth JD, Asante EA, Desbruslais M, et al. (2004) Human prion protein with valine 129 prevents expression of variant CJD phenotype. Science;306:1793–1796.

    Article  CAS  PubMed  Google Scholar 

  45. Collinge J, Harding AE, Owen F, et al. (1989) Diagnosis of Gerstmann-Sträussler syndrome in familial dementia with prion protein gene analysis. Lancet;2:15–17.

    Article  CAS  PubMed  Google Scholar 

  46. Collinge J, Owen F, Poulter M, et al. (1990) Prion dementia without characteristic pathology. Lancet;336:7–9.

    Article  CAS  PubMed  Google Scholar 

  47. Collinge J, Brown J, Hardy J, et al. (1992) Inherited prion disease with 144 base pair gene insertion: II: clinical and pathological features. Brain;115:687–710.

    Article  PubMed  Google Scholar 

  48. Mallucci GR, Campbell TA, Dickinson A, et al.(1999) Inherited prion disease with an alanine to valine mutation at codon 117 in the prion protein gene. Brain;122:1823–1837.

    Article  PubMed  Google Scholar 

  49. Mead S, Poulter M, Beck J, et al. (2006) Inherited prion disease with six octapeptide repeat insertional mutation—molecular analysis of phenotypic heterogeneity. Brain; 129:2297–2317.

    Article  PubMed  Google Scholar 

  50. World Health Organisation. (2003) WHO manual for surveillance of human transmissible spongiform encephalopathies. http://www.who.int/bloodproducts/TSE-manual2003.pdf

  51. Collinge J, Poulter M, Davis MB, et al. (1991) Presymptomatic detection or exclusion of prion protein gene defects in families with inherited prion diseases. Am J Hum Genet;49:1351–1354.

    CAS  PubMed  Google Scholar 

  52. Budka H, Aguzzi A, Brown P, et al. (1995) Neuropathological diagnostic criteria for Creutzfeldt-Jakob disease (CJD) and other human spongiform encephalopathies (prion diseases). Brain Pathol;5:459–466.

    Article  CAS  PubMed  Google Scholar 

  53. Budka H. Neuropathology of prion diseases. (2003) Br Med Bull;66:121–130.

    Article  CAS  PubMed  Google Scholar 

  54. Hainfellner JA, Brantner-Inthaler S, Cervenáková L, et al. (1995) The original Gerstmann-Sträussler-Scheinker family of Austria: divergent clinicopathological phenotypes but constant PrP genotype. Brain Pathol;5:201–211.

    Article  CAS  PubMed  Google Scholar 

  55. Ironside JW, Head MW. Neuropathology and molecular biology of variant Creutzfeldt-Jakob disease. (2004) Curr Top Microbiol Immunol;284:133–159.

    CAS  PubMed  Google Scholar 

  56. Hill AF, Butterworth RJ, Joiner S, et al. (1999) Investigation of variant Creutzfeldt-Jakob disease and other human prion diseases with tonsil biopsy samples. Lancet;353:183–189.

    Article  CAS  PubMed  Google Scholar 

  57. Glatzel M, Abela E, Maissen M, Aguzzi A. (2003) Extraneural pathologic prion protein in sporadic Creutzfeldt-Jakob disease. N Engl J Med;349:1812–1820.

    Article  CAS  PubMed  Google Scholar 

  58. Hilton DA, Sutak J, Smith ME, et al. (2004) Specificity of lymphoreticular accumulation of prion protein for variant Creutzfeldt-Jakob disease. J Clin Pathol;57:300–302.

    Article  CAS  PubMed  Google Scholar 

  59. Head MW, Ritchie D, Smith N, et al. (2004) Peripheral tissue involvement in sporadic, iatrogenic, and variant Creutzfeldt-Jakob disease: an immunohistochemical, quantitative, and biochemical study. Am J Pathol;164:143–153.

    Article  CAS  PubMed  Google Scholar 

  60. Joiner S, Linehan JM, Brandner S, Wadsworth JD, Collinge J. (2005) High levels of disease related prion protein in the ileum in variant Creutzfeldt-Jakob disease. Gut;54:1506–1508.

    Article  CAS  PubMed  Google Scholar 

  61. Wadsworth JD, Joiner S, Fox K, et al. (2007) Prion infectivity in vCJD rectum. Gut;56:90–94.

    Article  CAS  PubMed  Google Scholar 

  62. Siddique D, Kennedy A, Thomas D, et al. (2005) Tonsil biopsy in the investigation of suspected variant Creutzfeldt-Jakob disease—a cohort study of 50 patients. J Neurol Sci;238:S1–S570.

    Google Scholar 

  63. Advisory Committee on Dangerous Pathogens and the Spongiform Encephalopathy Advisory Committee. (2003) Transmissible spongiform encephalopathy agents: safe working and the prevention of infection. UK Department of Health. http://www.advisorybodies.doh.gov.uk/ acdp/tseguidance/Index.htm

  64. Safar J, Wille H, Itri V, et al. (1998) Eight prion strains have PrPSc molecules with different conformations. Nat Med; 4:1157–1165.

    Article  CAS  PubMed  Google Scholar 

  65. Riesner D. (2003) Biochemistry and structure of PrPC and PrPSc. Br Med Bull;66:21–33.

    Article  CAS  PubMed  Google Scholar 

  66. Parchi P, Capellari S, Chen SG, et al. (1997) Typing prion isoforms. Nature;386:232–233.

    Article  CAS  PubMed  Google Scholar 

  67. Piccardo P, Dlouhy SR, Lievens PMJ, et al. (1998) Phenotypic variability of Gerstmann-Sträussler-Scheinker disease is associated with prion protein heterogeneity. J Neuropathol Exp Neurol;57:979–988.

    Article  CAS  PubMed  Google Scholar 

  68. Parchi P, Chen SG, Brown P, et al. (1998) Different patterns of truncated prion protein fragments correlate with distinct phenotypes in P102L Gerstmann-Sträussler-Scheinker disease. Proc Natl Acad Sci U S A;95:8322–8327.

    Article  CAS  PubMed  Google Scholar 

  69. Furukawa H, Doh-ura K, Kikuchi H, Tateishi J, Iwaki T. (1998) A comparative study of abnormal prion protein isoforms between Gerstmann-Sträussler-Scheinker syndrome and Creutzfeldt-Jakob disease. J Neurol Sci;158:71–75.

    Article  CAS  PubMed  Google Scholar 

  70. Piccardo P, Liepnieks JJ, William A, et al. (2001) Prion proteins with different conformations accumulate in Geustmann-Sträussler-Scheinker disease caused by A117V and F198S mutations. Am J Pathol;158:2201–2207.

    Article  CAS  PubMed  Google Scholar 

  71. Tagliavini F, Lievens PMJ, Tranchant C, et al. (2001) A 7-kDa prion protein (PrP) fragment, an integral component of the PrP region required for infectivity, is the major amyloid protein in Gerstmann-Sträussler-Scheinker disease A117V. J Biol Chem;276:6009–6015.

    Article  CAS  PubMed  Google Scholar 

  72. Wadsworth JD, Joiner S, Linehan JM, et al. (2006) Phenotypic heterogeneity in inherited prion disease (P102L) is associated with differential propagation of protease-resistant wild-type and mutant prion protein. Brain;129:1557–1569.

    Article  PubMed  Google Scholar 

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Acknowledgments

We especially thank all patients and their families for generously consenting to use of human tissues in this research, and the UK neuropathologists who have kindly helped in providing these tissues. We are grateful to R. Young for preparation of the figures. This work was funded by the UK Medical Research Council and the European Commission, and it was performed under approval from the Institute of Neurology/National Hospital for Neurology and Neurosurgery Local Research Ethics Committee.

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Authors and Affiliations

  1. MRC Prion Unit, University College London Institute of Neurology, London, UK

    Jonathan D. F. Wadsworth, Caroline Powell, Jonathan A. Beck, Susan Joiner, Jacqueline M. Linehan, Sebastian Brandner, Simon Mead & John Collinge

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  1. Jonathan D. F. Wadsworth
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  2. Caroline Powell
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  3. Jonathan A. Beck
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  4. Susan Joiner
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  5. Jacqueline M. Linehan
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Editors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute; Department of Pathology and Mental Health Institute of Victoria, University of Melbourne, 30 Flemington Road Parkville, Melbourne, VIC, 3010, Australia

    Andrew F. Hill

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Wadsworth, J.D.F. et al. (2008). Molecular Diagnosis of Human Prion Disease. In: Hill, A.F. (eds) Prion Protein Protocols. Methods in Molecular Biology™, vol 459. Humana Press. https://doi.org/10.1007/978-1-59745-234-2_14

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  • DOI: https://doi.org/10.1007/978-1-59745-234-2_14

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-897-3

  • Online ISBN: 978-1-59745-234-2

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