Response

Abstract

Two observations led us to test the expression of alternative prion protein (AltPrP) (1). First, many functions were attributed to PrP, a protein encoded in the PRNP gene (2,3). How one gene could encode one protein with such a variety of function is puzzling. Second, we noticed a putative overlapping reading frame in the +3 reading frame of PRNP. This coding sequence overlaps the octapeptide repeat region, a domain well conserved across species. Based on these two considerations only, a strategy was used that undoubtedly established that PRNP encodes two proteins: PrP and AltPrP (1). AltPrP is a tryptophan-rich mitochondrial protein, and its physiological function or its role in prion diseases is still unknown. Previously, a hypothetical-induced frameshifting mechanism was proposed to explain the replication of prions, and the following model was suggested (4). First, a frameshift followed by a compensating frameshift during the translation of PrP results in the production of hybrid PrP molecules with sequence elements of AltPrP. Second, hybrid AltPrP-PrP molecules are capable of stimulating these frameshifting errors. Third, hybrid AltPrP-PrP molecules are the infectious agent and provide nuclei for PrP aggregation. In our experiments, we have had no evidence that AltPrP synthesis involves ribosomal frameshifting. Furthermore, we could not detect hybrid AltPrP-PrP molecules by Western blot, and no experimental data indicated the possibility of frameshifting errors during PrP translation. However, levels of hybrid molecules may be too low to be detected by Western blot. Alternatively, the expression of hybrid molecules may require specific experimental conditions. As noted by Dr Wills, some unresolved gaps persist in the current model of prion replication, despite major breakthroughs in recent years. Nonetheless, the existence and the role for hybrid AltPrP-PrP molecules in prion disease remain speculative. Sensitive proteomic techniques on infectious prion particles may help address this issue. Another strategy to challenge the frameshifting model would be to test if expression of artificial hybrid AltPrP-PrP molecules in cultured cells or in transgenic animals results in spontaneous production of infectious prions.