Ovine plasma prion protein levels show genotypic variation detected by C-terminal epitopes not exposed in cell-surface PrP^C

Ovine PBMCs (peripheral blood mononuclear cells) express PrP^C [cellular PrP (prion-related protein)] and have the potential to harbour and release disease-associated forms of PrP during scrapie in sheep. Cell-surface PrP^C expression by PBMCs, together with plasma PrP^C levels, may contribute to the regulatory mechanisms that determine susceptibility and resistance to natural scrapie in sheep. Here, we have correlated cell-surface PrP^C expression on normal ovine PBMCs by FACS with the presence of PrP^C in plasma measured by capture–detector immunoassay. FACS showed similar levels of cell-surface PrP^C on homozygous ARR (Ala¹³⁶-Arg¹⁵⁴-Arg¹⁷¹), ARQ (Ala¹³⁶-Arg¹⁵⁴-Gln¹⁷¹) and VRQ (Val¹³⁶-Arg¹⁵⁴-Gln¹⁷¹) PBMCs. Cell-surface ovine PrP^C showed modulation of N-terminal epitopes, which was more evident on homozygous ARR cells. Ovine plasma PrP^C levels showed genotypic variation and the protein displayed C-terminal epitopes not available in cell-surface PrP^C. Homozygous VRQ sheep showed the highest plasma PrP^C level and homozygous ARR animals the lowest. For comparison, similar analyses were performed on normal bovine PBMCs and plasma. PrP^C levels in bovine plasma were approx. 4-fold higher than ovine homozygous ARQ plasma despite similar levels of PBMC cell-surface PrP^C expression. Immunoassays using C-terminal-specific anti-PrP monoclonal antibodies as capture and detector reagents revealed the highest level of PrP^C in both ovine and bovine plasma, whilst lower levels were detected using N-terminal-specific monoclonal antibody FH11 as the capture reagent. This suggested that a proportion of plasma PrP^C was N-terminally truncated. Our results indicate that the increased susceptibility to natural scrapie displayed by homozygous VRQ sheep correlates with a higher level of plasma PrP^C.