Prion strains are different self-propagating conformers of the same infectious protein. Three strains of the [PSI] prion, infectious forms of the yeast Sup35 protein, have been previously characterized in our laboratory. Here we report the discovery of a new [PSI] strain, named W8. We demonstrate its robust cellular propagation as well as the protein-only transmission. To reveal strain-specific sequence requirement, mutations that interfered with the propagation of W8 were identified by consecutive substitution of residues 5-55 of Sup35 by proline and insertion of glycine at alternate sites in this segment. Interestingly, propagating W8 with single mutations at residues 5-7 and around residue 43 caused the strain to transmute. In contrast to the assertion that [PSI] existed as a dynamic cloud of sub-structures, no random drift in transmission characteristics was detected in mitotically propagated W8 populations. Electron diffraction and mass-per-length measurements indicate that, similar to the 3 previously characterized strains, W8 fibers are composed of about 1 prion molecule per 4.7-Å cross-β repeat period. Thus differently folded single Sup35 molecules, not dimeric and trimeric assemblies, form the basic repeating units to build the 4 [PSI] strains.
Keywords: 5-FOA, 5-fluoroorotic acid; Aβ, amyloid β-protein; GFP, green fluorescent protein; PrP, prion protein; SC, synthetic complete; STEM, scanning transmission electron microscopy; YPD, yeast extract, peptone, dextrose; amyloid, prion strain, [PSI+], SUP35, yeast; mpl, mass per length.