Several models have been proposed for the infectious agents that cause human Creutzfeldt-Jakob disease (CJD) and sheep scrapie. Purified

A class of viruslike agents that induces Creutzfeldt-Jakob Disease (CJD) and scrapie remains undefined at the molecular level. Several investigators believe this infectious agent is constituted by a single host protein or 'prion', and have emphasized data that would seem to exclude the presence of any viral nucleic acids. However, more rigorous evaluations in scrapie have shown reasonably abundant nucleic acids. Additionally, in highly purified 120S CJD preparations that have been treated with nucleases, RNAs as long as 6,000 bases have been detected. Few nucleic acids have been characterized in either scrapie or CJD, but previous cloning experiments delineated relatively short LTR regions of the endogenous IAP retrovirus in 120S CJD preparations. We therefore used specific primers encompassing the entire IAP genome to test for the presence of long viral RNAs, and here show approximately 5,000 contiguous bases of the IAP RNA genome can be recovered from reasonable amounts of starting brain. The 3' env region of IAP is comparably truncated in CJD and normal preparations, and we find no evidence for IAP transduction of CJD-specific sequences. Because IAP cores can coencapsidate unrelated sequences, and are unusually resistant to physical and chemical treatments, it was relevant to find if cosedimenting cognate proteins of the IAP core, such as gag, could be detected. The predicted approximately 65 kd acidic gag protein, showing appropriate antigenic and nucleic acid binding features, was apparent in both one and 2-D Western blots. This data strongly indicates specific viral complexes cofractionate with the CJD agent. Interestingly, these nuclease resistant IAPs do not appear to be in morphologically recognizable 'R' particles. This cosedimenting viral assembly therefore provides a paradigm for non-particulate CJD complexes in infectious preparations. In developing strategies to identify a CJD specific sequence, cosedimenting IAPs can be used to assess the quality, length and recovery of RNAs extracted from highly resistant viral complexes.

No abstract

Representative preparations of partially purified Creutzfeldt-Jakob disease (CJD), including disaggregated density gradient fractions, were treated with a variety of nucleases. RNases as well as exhaustive digestions with micrococcal nuclease did not significantly diminish infectivity, but resulted in an approximately 7,000-fold specific purification of infectivity with respect to nucleic acid. Protected nucleic acids included species of up to 2,000 bases in length. After nuclease treatment, infectivity co-migrated with nucleic acid-protein complexes at a density of 1.27 g/cm3 in sucrose. Substantial specific protein purification were also achieved in the gradient step (approximately 11,000-fold), where 70% the host Gp34 ("prion protein") as well as other free proteins separated from infectivity. These CJD purifications are better than those previously attained in scrapie, and may be useful for further studies of non-host protein and nucleic acid species. The data are consistent with the hypothesis that CJD-like agents are composed of nucleic acid-protein complexes.

No abstract