Nearly ubiquitous tissue distribution of the scrapie agent precursor protein.

The "modified host protein" model of scrapie proposes that the transmissible agent is composed of the degradation-resistant protein, Sp33-37, and that clinical and pathologic signs result from neurotoxic accumulations of this protein. Sp33-37 is an abnormal, amyloidogenic isoform of the normally occurring cellular protein Cp33-37. This study investigated the tissue distribution of Cp33-37 in hamster. In brain, Cp33-37 was most concentrated in the hippocampal formation. Immunohistochemical studies localized Cp33-37 to neurons and surrounding neuropil in hippocampus; septal, caudate, and thalamic nuclei; dorsal root ganglia cells; and large-diameter dorsal root axons. In non-neuronal hamster tissues, Cp33-37 was detected in circulating leukocytes, heart, skeletal muscle, lung, intestinal tract, spleen, testis, ovary, and some other organs. The presence of Cp33-37 in extracerebral tissues indicates that its function is not unique to brain. These results indicate that the molecular substrate for the production of Sp33-37, the scrapie agent, and scrapie amyloid is present in a variety of cerebral and extracerebral sites.

Molecular location of a species-specific epitope on the hamster scrapie agent protein.

Scrapie is a transmissible neurodegenerative disease of sheep and goats. An abnormal host protein, Sp33-37, is the major protein component of the scrapie agent and the only known disease- or agent-specific macromolecule. Two monoclonal antibodies (MAbs), 4H8 (immunoglobulin G2b [IgG2b]) and 6B11 (IgG1), produced by immunizing mice with the intact hamster 263K scrapie agent protein, Sp33-37Ha, were found to have species specificity similar to that reported previously for MAb 3F4 (IgG2a), which was produced by using PrP-27-30 as the immunogen (R. J. Kascsak, R. Rubenstein, P. A. Merz, M. Tonna-DeMasi, R. Fersko, R. I. Carp, H. M. Wisniewski, and H. Diringer, J. Virol. 61:3688-3693, 1987). These antibodies all bound to Sp33-37 derived from hamster but not from mouse cells. Competitive binding assays demonstrated that all three MAbs bound to the same or overlapping sites on Sp33-37Ha. The molecular location of the epitope for these antibodies was determined to within 10 residues by using an antigen competition enzyme-linked immunosorbent assay in which synthetic peptides spanning Sp33-37Ha residues 79 to 93 or 84 to 93 specifically inhibited binding of these antibodies to plates coated with purified Sp33-37Ha. A synthetic peptide with the mouse-specific sequence (83 to 92) that differed from the hamster sequence by substitution at two positions (MetHa-87----LeuMo-86 and MetHa-90----ValMo-89) did not inhibit antibody binding to Sp33-37Ha. MAb 3F4 binding to hamster Sp33-37 was eliminated by chemical modification of Sp33-37Ha with diethylpyrocarbonate or succinic anhydride and by cleavage with CNBr or trypsin. The effect of diethylpyrocarbonate on MAb 3F4 binding was not reversed by hydroxylamine treatment. MAb 3F4 binding was not affected by prolonged exposure of Sp33-37Ha to 70% formic acid or by boiling in sodium dodecyl sulfate. We conclude that the epitope for these MAbs is a linear determinant that includes Met-87, Lys-88, and Met-90 and that Met-90 is probably the major species-specific determinant.