Synthesis of high titer infectious prions with cofactor molecules

Abstract

Recently, synthetic prions with a high level of specific infectivity have been produced from chemically defined components in vitro. A major insight arising from these studies is that various classes of host-encoded cofactor molecules such as phosphatidylethanolamine and RNA molecules are required to form and maintain the specific conformation of infectious prions. Synthetic mouse prions formed with phosphatidylethanolamine exhibit levels of specific infectivity ∼1 million-fold greater than "protein-only" prions (Deleault, N. R., Walsh, D. J., Piro, J. R., Wang, F., Wang, X., Ma, J., Rees, J. R., and Supattapone, S. (2012) Proc. Natl. Acad. Sci. U.S.A. 109, E1938-E1946). Moreover, cofactor molecules also appear to regulate prion strain properties by limiting the potential conformations of the prion protein (see Deleault et al. above). The production of fully infectious synthetic prions provides new opportunities to study the mechanism of prion infectivity directly by structural and biochemical methods.

Keywords: Cofactor; Infectivity; Phosphatidylethanolamine; Phospholipid; Prion; Protein Misfolding; RNA; Synthetic.

FIGURE 1.

Schematic diagram showing the effect of cofactor withdrawal during serial propagation of purified recombinant PrP^Sc molecules in vitro. Removal of cofactor from substrate mixture results in PrP^Sc conformation, as judged by an ∼2-kDa shift in the mobility of the protease-resistant core and ∼10⁶-fold loss of specific infectivity, as judged by end-point dilution bioassay.

FIGURE 2.

Can a single cofactor maintain prion strain diversity? A, hypothetical model illustrating how the existence of multiple cofactor molecules could encipher prion strain diversity. B, schematic diagram showing the experimental paradigm used to test whether a single cofactor (PE) is able to maintain strain diversity. Strain typing assays showed that three different strains converged to form a single, novel strain after in vitro propagation in the presence of PE alone.