Amino acid sequence and prion strain specific effects on the in vitro and in vivo convertibility of ovine/murine and bovine/murine prion protein chimeras

Abstract

Prion diseases are characterised by the conversion of a cellular prion protein (PrP(C)) by its misfolded, hence pathogenic, isoform (PrP(Sc)). The efficiency of this transition depends on the molecular similarities between both interaction partners and on the intrinsic convertibility of PrP(C). Transgenic mice expressing chimeric murine/ovine PrP(C) (Tgmushp mice) are susceptible to BSE and/or scrapie prions of bovine or ovine origin while transgenic mice expressing similar murine/bovine PrP(C) chimera (Tgmubo mice) are essentially resistant. We have studied this phenomenon by cell-free conversion on procaryotically expressed chimeric PrP(C). Mouse passaged scrapie or BSE PrP(Sc) was used as a seed and the conversion reaction was carried out under semi-native conditions. The results obtained in this assay were similar to those of our in vivo experiments. Since mubo- and mushp-PrP(C) differ only at four amino acid positions (S96G, N142S, Y154H and Q185E), single or double point mutations of mushp-PrP(C) were examined in the cell-free conversion assay. While the scrapie Me7 prion induced conversion was largely reduced by the N142S and Q185E but not by the S96G and Y154H mutation, the BSE induced conversion was retained in all mutants. Newly formed PrP(res) exhibited strain specific characteristics, such as the localisation of the proteinase K cleavage site, even in the chimeric PrP(C) mutants. We therefore postulate that the efficiency of the conversion of chimeric PrP(C) depends on the amino acid sequence as well as on prion strain specific effects.

Preclinical Detection of Variant CJD and BSE Prions in Blood

“…For instance, ovine BSE propagates with an apparently similar efficiency to cattle BSE prions in bovine transgenic mice (tgBov) [36] and with an higher efficiency in human transgenic mice (tg650) [51]. BSE/vCJD agents propagate with little or no transmission barrier in transgenic mice expressing the ovine ARQ PrP [33], [52], [53] and can be passaged in those expressing the ovine VRQ PrP variant (tg338 mice) [54].…”

Influence of Breed and Genotype on the Onset and Distribution of Infectivity and Disease-associated Prion Protein in Sheep Following Oral Infection with the Bovine Spongiform Encephalopathy Agent

“…Five Suffolk sheep, four ARQ/ARQ (three in the 16 month cull group and one in the clinical disease group) and one ARQ/ARR were methionine/threonine (M/T) heterozygous at codon 112 of Prnp (MARQ/TARQ or TARQ/MARR, respectively). This additional polymorphism of the ARQ allotype of Suffolk sheep is associated with resistance to scrapie (Laegreid et al, 2008; and to BSE (Saunders et al, 2009); it was absent from all Romney sheep in the experiment. ARQ/ARR and ARR/ARR sheep were culled at 10, 22, 34, 46 and 85e122 mpi.…”

“…During the course of these experiments, novel transgenic murine models were developed, which were deemed more sensitive for detection of BSE infectivity. Accordingly, selected inocula were re-assayed in TgshpXI mice, which overexpress ARQ/ARQ ovine PrP on a murine PrP null background (Kupfer et al, 2007). Each inoculum was injected intracerebrally to a group of 10 TgshpXI mice, each of which received 20 ml of a specified inoculum.…”

Mammalian prions

“…38 F and T residues conservation in front of H3 might be necessary proteins permissive to hamster, human and sheep prions respectively. [22][23][24] These findings, along the pivotal role of the Met/ Val polymorphism at PrP codon 129 in the human susceptibility to prions, 25 suggested that the region upstream from the last two helices is a major domain involved in the conformational change. The successful, cell-free amplification of a PK-resistant entity from the short PrP segment corresponding to the disease-associated human mutation Y145Stop recently further highlighted the potential role of this region.…”