Destabilizing polymorphism in cervid prion protein hydrophobic core determines prion conformation and conversion efficiency

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Abstract
Prion diseases are infectious neurodegenerative disorders of humans and animals caused by misfolded forms of the cellular prion protein PrPC. Prions cause disease by converting PrPC into aggregation-prone PrPSc. Chronic wasting disease (CWD) is the most contagious prion disease with substantial lateral transmission, affecting free-ranging and farmed cervids. Although the PrP primary structure is highly conserved among cervids, the disease phenotype can be modulated by species-specific polymorphisms in the prion protein gene. How the resulting amino-acid substitutions impact PrPC and PrPSc structure and propagation is poorly understood. We investigated the effects of the cervid 116A>G substitution, located in the most conserved PrP domain, on PrPC structure and conversion and on 116AG-prion conformation and infectivity. Molecular dynamics simulations revealed structural de-stabilization of 116G-PrP, which enhanced its in vitro conversion efficiency when used as recombinant PrP substrate in real-time quaking-induced conversion (RT-QuIC). We demonstrate that 116AG-prions are conformationally less stable, show lower activity as a seed in RT-QuIC and exhibit reduced infectivity in vitro and in vivo. Infectivity of 116AG-prions was significantly enhanced upon secondary passage in mice, yet conformational features were retained. These findings indicate that structurally de-stabilized PrPC is readily convertible by cervid prions of different genetic background and results in a prion conformation adaptable to cervid wild-type PrP. Conformation is an important criterion when assessing transmission barrier, and conformational variants can target a different host range. Therefore, a thorough analysis of CWD isolates and re-assessment of species-barriers is important in order to fully exclude a zoonotic potential of CWD. Chronic wasting disease (CWD) is a prion disease which affects wild and captive cervids. Prion diseases are infectious neurodegenerative disorders, and the causative agent consists of abnormally folded prion protein termed PrPSc. Prions replicate without genetic information, and their three-dimensional structure is thought to encode heritable information necessary to propagate using the cellular prion protein PrPC as a substrate for conversion. In this study, we use in vitro and in vivo techniques to analyze the effect of a polymorphism at codon 116 (A>G) of the white-tailed deer prion protein on CWD prion conformation, propagation and pathogenesis. We observed differences in conformation, infectivity and seeding activity in vitro between CWD prions isolated from white-tailed deer encoding wild-type (116AA) PrPC or 116AG-PrPC. In mouse bioassays conformational differences are retained, however, 116AG CWD prions resulted in significantly shortened incubation times upon passages. Molecular dynamics simulations suggest that the structure of 116G-PrPC is more flexible, which is supported by an improved convertibility in an in vitro conversion assay. Altogether these data indicate the importance of a variation in the most conserved PrP domain, and highlight the relationship between PrPC structural flexibility, prion conformation and conversion, and pathogenesis of prion disease in vivo.
Funding Information
  • Alberta Prion Research Institute and Alberta Livestock and Meat Agency (CA) (201300027)
  • Alberta Prion Research Institute and Alberta Livestock and Meat Agency (CA) (20160012)
  • Canada Research Chairs
  • Natural Sciences and Engineering Research Council of Canada
  • Genome Canada
  • Genome Canada (CA)
  • Genome Canada

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