Further Characterization of Glycoform-Selective Prions of Variably Protease-Sensitive Prionopathy

Abstract

Prion is an infectious protein (PrPSc) that is derived from a cellular glycoprotein (PrPC) through a conformational transition and associated with a group of prion diseases in animals and humans. Characterization of proteinase K (PK)-resistant PrPSc by western blotting has been critical to diagnosis and understanding of prion diseases including Creutzfeldt-Jakob disease (CJD) and Gerstmann-Sträussler-Scheinker (GSS) disease in humans. However, formation as well as biochemical and biological properties of the glycoform-selective PrPSc in variably protease-sensitive prionopathy (VPSPr) remain poorly understood. Here we reveal that formation of the ladder-like PrPSc in VPSPr is a PK-dependent two-step process, which is enhanced by basic pH. Two sets of PrPSc fragments can be identified with antibodies directed against an intermediate or a C-terminal domain of the protein. Moreover, antibodies directed against specific PrP glycoforms reveal faster electrophoretic migrations of PrP fragments mono-glycosylated at residue 181 and 197 in VPSPr than those in sporadic CJD (sCJD). Finally, RT-QuIC assay indicates that PrPSc-seeding activity is lower and its lag time is longer in VPSPr than in sCJD. Our results suggest that the glycoform-selective PrPSc in VPSPr is associated with altered glycosylation, resulting in different PK-truncation and aggregation seeding activity compared to PrPSc in sCJD.

Phenotypic Heterogeneity of Variably Protease-Sensitive Prionopathy: A Report of Three Cases Carrying Different Genotypes at PRNP Codon 129

“…In line with previous studies [11,30], we found that VPSPr PrP Sc shows the highest sensitivity to PK digestion in individuals 129VV, the lowest in those carrying 129MM, and intermediate values in those with 129MV. We also confirmed that VPSPr PrP Sc shows an overall distinctive immunoblot profile, but with an additional significant heterogeneity among cases depending on the PK activity and the codon 129 genotype [31]. Overall, both 3F4 and T2 antibodies detect five PrP Sc fragments independent from the codon 129 genotype, including mono-and un-glycosylated forms of a 19 kDa fragment corresponding to CJD PrP Sc type 2, monoglycosylated and unglycosylated forms of a 17 kDa fragment lacking the GPI anchor and a 7 kDa fragment ragged at both N-and C-termini.…”

Biochemical and Neuropathological Findings in a Creutzfeldt–Jakob Disease Patient with the Rare Val180Ile-129Val Haplotype in the Prion Protein Gene

“…An RT-QuIC assay was performed to detect the presence of the pathological misfolded PrP in CSF and the OE collected by nasal brushing, using standard protocols in the presence of the recombinant full-length hamster prion protein (Ha rPrP 23–231) as a substrate for the reaction [ 29 , 30 ]. Moreover, additional tests were run with either the recombinant, truncated hamster prion protein (Ha rPrP 90–231) or the recombinant bank vole full-length prion protein (BV rPrP 23–230).…”