Mammalian species vary widely in their apparent susceptibility to prion diseases. For example, several felid species developed prion disease (feline spongiform encephalopathy or FSE) during the bovine spongiform encephalopathy (BSE) epidemic in the United Kingdom, whereas no canine BSE cases were detected. Whether either of these or other groups of carnivore species can contract other prion diseases (e.g. chronic wasting disease or CWD) remains an open question. Variation in the host-encoded prion protein (PrPC) largely explains observed disease susceptibility patterns within ruminant species, and may explain interspecies differences in susceptibility as well. We sequenced and compared the open reading frame of the PRNP gene encoding PrPC protein from 609 animal samples comprising 29 species from 22 genera of the Order Carnivora; amongst these samples were 15 FSE cases. Our analysis revealed that FSE cases did not encode an identifiable disease-associated PrP polymorphism. However, all canid PrPs contained aspartic acid or glutamic acid at codon 163 which we propose provides a genetic basis for observed susceptibility differences between canids and felids. Among other carnivores studied, wolverine (Gulo gulo) and pine marten (Martes martes) were the only non-canid species to also express PrP-Asp163, which may impact on their prion diseases susceptibility. Populations of black bear (Ursus americanus) and mountain lion (Puma concolor) from Colorado showed little genetic variation in the PrP protein and no variants likely to be highly resistant to prions in general, suggesting that strain differences between BSE and CWD prions also may contribute to the limited apparent host range of the latter.
Expression of the cellular prion protein (PrP(C)) is crucial for the development of prion diseases. Resistance to prion diseases can result from reduced availability of the prion protein or from amino acid changes in the prion protein sequence. We propose here that increased production of a natural PrP α-cleavage fragment, C1, is also associated with resistance to disease. We show, in brain tissue, that ARR homozygous sheep, associated with resistance to disease, produced PrP(C) comprised of 25% more C1 fragment than PrP(C) from the disease-susceptible ARQ homozygous and highly susceptible VRQ homozygous animals. Only the C1 fragment derived from the ARR allele inhibits in-vitro fibrillisation of other allelic PrP(C) variants. We propose that the increased α-cleavage of ovine ARR PrP(C) contributes to a dominant negative effect of this polymorphism on disease susceptibility. Furthermore, the significant reduction in PrP(C) β-cleavage product C2 in sheep of the ARR/ARR genotype compared to ARQ/ARQ and VRQ/VRQ genotypes, may add to the complexity of genetic determinants of prion disease susceptibility.
This paper describes the generation, characterisation and potential applications of a panel of novel anti-prion protein monoclonal antibodies (mAbs). The mAbs were generated by immunising PRNP null mice, using a variety of regimes, with a truncated form of recombinant ovine prion protein spanning residues 94–233. Epitopes of specific antibodies were mapped using solid-phase Pepscan analysis and clustered to four distinct regions within the PrP molecule. We have demonstrated the utility of these antibodies by use of Western blotting and immunohistochemistry in tissues from a range of different species affected by transmissible spongiform encephalopathy (TSE). In comparative tests against extensively-used and widely-published, commercially available antibodies, similar or improved results can be obtained using these new mAbs, specifically in terms of sensitivity of detection. Since many of these antibodies recognise native PrPC, they could also be applied to a broad range of immunoassays such as flow cytometry, DELFIA analysis or immunoprecipitation. We are using these reagents to increase our understanding of TSE pathogenesis and for use in potential diagnostic screening assays.
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