Modelling zoonotic transmission using protein misfolding cyclic amplification

The vast majority of cases of human prion diseases world-wide are either idiopathic (apparently sporadic) or associated with mutations in the PRNP gene. Nevertheless, human prion diseases can be acquired and the experience of bovine spongiform encephalopathy (BSE) and variant Creutzfeldt-Jakob disease in the United Kingdom clearly demonstrates the profound and long-term effects that zoonotic prion disease can have. Whether or not an animal prion disease presents a risk to human health is multifactorial, but one fundamental factor may be molecular compatibility. Specifically, compatibility between the prion protein conformers and glycotypes associated with individual animal prion diseases and the normal prion protein and its genotypic variants in humans.
We have sought to examine this hypothesis using a cell-free conversion system, protein misfolding cyclic amplification (PMCA) which is thought to mimic in accelerated form the fundamental event in prion pathogenesis, namely the conversion of the normal prion protein from one source (in this case human) by abnormal prion protein from another source (in this case animal). We have explored the effects of species, prion strain and Prnp genotype using brain samples from BSE, experimental BSE in sheep, scrapie, atypical scrapie, H- and L-type BSE and chronic wasting disease. We have also explored the effect of the M/V polymorphism at codon 129 of the human prion protein gene using homogenates of human brain, humanised transgenic mouse brain or human cell lines as the PMCA substrate.
The results show that the animal prion diseases samples tested differed in their ability to convert human prion protein. Conversion of human prion protein was associated with samples of BSE and experimental sheep BSE. In contrast, samples of scrapie in sheep and H- and L-type BSE in cattle failed to result in detectable conversion of the human prion protein, whereas a sample of elk CWD did effect some limited conversion of human prion protein. Conversion was more efficient when M rather than V was present at position 129 in the human prion protein. Serial PMCA of experimental sheep BSE showed an unexpected sheep genotype-dependent switch in the conformer of abnormal prion protein present. When this occurred, the ability to convert human prion protein was lost. The results from these experiments suggest that cell-free prion protein conversion assays, such as PMCA might play a useful role in the assessment of zoonotic potential and that they can contribute to a better understanding of mechanisms involved in prion propagation.