SCRAPIE PRION PROTEIN FORMATION IN HUMAN PRION DISEASES

Case Western Reserve University, Cleveland OH

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Abstract

GRANT=6546045;P01AG Scrapie prion protein (PrP-Sc) is now recognized as a major player in the pathogenesis of prion diseases. However, important features of this critical molecule remain obscure or controversial. The overall objective of this Research Project is to identify the mechanisms that lead to the formation of distinct PrP-Sc species or strains, which in turn determine different phenotypes in human prion diseases. Four specific aims are proposed. Specific Aim 1 will determine whether an 8kDa PrP internal fragment, that is resistant to proteases and is prominently present in a transmissible form of human prion disease, can propagate the disease. Transgenic (Tg) mice expressing P102L mutated human PrP will be inoculated with homogenates containing the PrP 8kDa peptide alone or together with PrP-Sc 27-30 from brains affected by a familial prion disease linked to the P102L mutation. Specific aims 2 and 3 address the controversial issue of the role of glycans in determining PrP-Sc strain diversity. In Specific Aim 2, two-dimensional gel electrophoresis (2-D PAGE) will be applied to the study of PrP-Sc species that are indistinguishable as for the size of the protease resistant fragments but have different ratios of the glycoforms such as the variant Creutzfeldt-Jakob disease and fatal familial insomnia. This aim is based on the finding that two PrP-Sc associated with two different disease phenotypes but with protease-resistant fragment of the same size were found to carry distinct glycans by 2-D PAGE. Specific Aim 3 is designed to assess whether glycans alone can confer diversity to PrP-Sc strains. PrP-Sc strains, which apparently differ only on the types of their glycans but are associated with two distinct phenotypes, will be inoculated to humanized Tg mice with and without ablation of the glycans. They will also be inoculated to Tg mice expressing only unglycosylated human PrP. Specific Aim 4 deals with the mechanism of PrP to PrP-Sc conversion in cell models of human genetic prion diseases. This aim is based on previous studies showing that PrP gene mutations profoundly affect the metabolism of mutant PrP, which, in turn, likely affects the modalities of prion formation and prion diversity in the still poorly known field of human prion diseases.