Spongiform Encephalopathies

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					   The Role of Prions in
Spongiform Encephalopathies

Spongiform Encephalopathies (SEs)

• Fatal neurodegenerative disorders: infectious,
  inherited, or sporadic
• Animal species:
   – Humans: Creutzfeldt-Jakob disease (CJD); kuru, Gerstmann-
      Straussler-Sheinker disease (GSS); Familial Fatal Insomnia (FFI)

   – Sheep: Scrapie
   – Cattle: Bovine Spongiform Encephalopathy (BSE)

• Symptoms: progressive dementia, motor dysfunction (ataxia;
  myoclonus; pyramidal/extra-pyramidal signs)
The origin of Kuru
GSS autosomal inheritance of a P102L mutation in the PrPc protein
Both Kuru and GSS were transmissible by injecting
       infected brain tissue into Monkeys

  Iatrogenic CJD developed from using contaminated surgical instruments,
                        or human growth hormone

• Astrogliosis        astrocytic hypertrophy

• Loss of neurones

• Vacuolation        Spongiform degeneration

• Amyloid plaques (occasionally)

Neuronal Vacuolation   Spongiform Change
British beef is safe! John Gummer, 1990

• First demonstrated in 1939: sheep scrapie   goat

• Transmissibility of kuru and CJD shown

  What is the nature of the infectious
   Purification of Scrapie Agent

• By Stanley B. Prusiner

• Infectivity   by proteinases

• Infectivity unchanged by nucleases and UV

• ‘Prion’= proteinaceous infectious particle
  lacking nucleic acid
            PrPC and PrPSc

• Scrapie agent found to contain 27-30 kDa
  protease resistant protein: PrP27-30

• PrP27-30 = protease resistant core of abnormal
  isoform of normal host protein PrPC

• Abnormal isoform termed PrPSc: only
  macromolecule specific for SEs
       Histopathology of vCJD

Section from frontal cortex     PrP immunopositivity in
showing aggregates of plaques       multiple plaques
surrounded by spongiform
  Prions are composed of PrPSc

• PrPSc and scrapie infectivity co-purify
• Properties of PrPSc mimic those of prions
• PrPSc levels directly proportional to prion
• PrPSc accumulation invariably associated
  with SE pathology
              PrP Genetics
• PrPC and PrPSc have identical 1o structure
• Encoded by single copy gene on chro. 20
Differences in         PrPC    and       PrP Sc

                          C         Sc
                    PrP       PrP
     Resistance       -         +
     Solubility       +         -
    Propensity to
     aggregate        -         +
     Structure      -helix   -sheet
Structures of   PrPC
Postulated Structure of   PrPSc
           Function of PrPC

• Unknown function - found on cell surface
  of neurones and glia & peripherally

• Possibly a modulator in apoptosis
Agent Structure Hypotheses

  ‘Protein-Only Hypothesis’


     ‘Virino Hypothesis’
        Conversion Reaction

• Two main models of molecular conversion
  of PrPC   PrPSc

     1. Template-directed refolding

     2. Nucleation-Dependent Polymerisation
 Template-Directed Refolding Model

• PrPC unfolded and refolded using PrPSc as template
• High activation energy barrier          molecular
  chaperone (protein X)
• RLS: binding of PrPC and chaperone
     Evidence for Template Model

• Chaperone studies: increase formation of
Nucleation-Dependent Polymerisation

• PrPC and PrPSc in reversible monomeric equilibrium
• Several PrPSc monomers needed to form stable
  nucleus or ‘seed’
• Rapid autocatalytic growth of PrPSc     polymer
• Infective material contains PrPSc seed
    Evidence for Nucleation Model

• Excess PrPSc can convert recombinant PrPC to
  protease resistant form

• PrPSc levels in transgenic mice

• Recombinant PrP converted reversibly between -
  helical and -sheet folds
     Conditions for PrP conversion

• Neutral or basic pH: -helical structure

• Reduction of disulphide bond and acidic pH: -
  sheet structure

• Residues had preference for -conformation

• Gel filtration of -PrP eluted monomers
      Critical Appraisal of Models
• Both models explain three important
             1. Transmissibility
            2. Species Barrier
            3. PrPSc sequence corresponds to last
               hosts PrP gene
• Transgenic PrP knockout mice are resistant to
  development of SE
• Synthesised peptide refolded can accelerate or ?
  initiate disease
        Both BSE and nvCJD are Type 4

Strains differ by the glycosylation profiles of PrPSc
                Inherited SE

• Pathognomic mutation in PRNP gene
• ~15% of all human SEs: 10% CJD, all cases GSS
  and FFI
• Mechanism: ?thermodynamic instability of PrPC
• Disease can occur without PrPSc
• Working instead by CtmPrP

• Certain amino acid changes CtmPrP levels -
• CtmPrP and not PrPSc in some human brains
•   CtmPrP   causes neurodegeneration
      CtmPrP        and Transmissible SEs

• Higher CtmPrP generators develop disease at lower
  PrPSc levels and vice versa

• Three lines of evidence:
  1. Increasing CtmPrP beyond threshold leads to neurodegeneration
     without PrPSc
  2. Amount of PrPSc needed to cause neurodegeneration influenced by
     hosts propensity to generate CtmPrP
  3. Brain contains increasing CtmPrP during course of PrPSc
CtmPrP:   Proposed Mechanism
    PrPSc: the true pathogenic factor?

• Not all PrPSc associated with infectivity

• 2o transmission elicited despite low levels of
  protease-resistant PrP (e.g. PrPSc)

        Is there another transmissible agent?
          Concluding Remarks

* Can species specific information be propagated by
  protein alone?
* Is SE pathogenesis due to neurotoxicity of
  converted product or simply loss of function of
* Are the transmissible and inherited SEs a result of
  the same molecular mechanism?
                          Laboratory Safety

 Prions can be destroyed by 150 degrees autoclaving or treatment with
                        1M sodium hydroxide,
           not eliminated by normal sterilisation procedures;
tonsilectomy (FDCs) or any nervous tissue (FDCs) contain much PrPSc
                         in infected individuals;
            B-cells in human blood could be contaminated!!
                Handled under P3 containment conditions