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Conformational variations in an infectious protein determine prion strain differences

Abstract

A remarkable feature of prion biology is the strain phenomenon wherein prion particles apparently composed of the same protein lead to phenotypically distinct transmissible states1,2,3,4. To reconcile the existence of strains with the ‘protein-only’ hypothesis of prion transmission, it has been proposed that a single protein can misfold into multiple distinct infectious forms, one for each different strain1,2,3,5. Several studies have found correlations between strain phenotypes and conformations of prion particles6,7,8,9,10; however, whether such differences cause or are simply a secondary manifestation of prion strains remains unclear, largely due to the difficulty of creating infectious material from pure protein3,5. Here we report a high-efficiency protocol for infecting yeast with the [PSI+] prion using amyloids composed of a recombinant Sup35 fragment (Sup-NM). Using thermal stability and electron paramagnetic resonance spectroscopy, we demonstrate that Sup-NM amyloids formed at different temperatures adopt distinct, stably propagating conformations. Infection of yeast with these different amyloid conformations leads to different [PSI+] strains. These results establish that Sup-NM adopts an infectious conformation before entering the cell—fulfilling a key prediction of the prion hypothesis5—and directly demonstrate that differences in the conformation of the infectious protein determine prion strain variation.

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Figure 1: Induction of the [PSI+] prion by in-vitro-converted Sup-NM amyloid fibres.
Figure 2: Generation of multiple [PSI+] strains by Sup-NM amyloid fibres converted in vitro.
Figure 3: Different conformations of Sup-NM amyloid fibres are formed at different temperatures.
Figure 4: Induction of distinct [PSI+] strains by Sup-NM amyloid fibres formed at different temperatures.

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Acknowledgements

We thank S. Collins, J. Newman, L. Osherovich, K. Tipton and members of the Weissman laboratory for discussion and reading of the manuscript. M.T. was supported by JSPS postdoctoral fellowships for research abroad. P.C. was supported by National Science Foundation Graduate Fellowships and the ARCS foundation. Funding was also provided by Howard Hughes Medical Instititute, The David and Lucile Packard Foundation and the National Institutes of Health.

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Correspondence to Jonathan S. Weissman.

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Supplementary information

Supplementary Information

Preparation of lyticase (DOC 5 kb)

Supplementary Figure 1

Effects of proteinase K and deoxyribonuclease I on prion infectivity. (JPG 95 kb)

Supplementary Figure 2

Non-Mendelian inheritance of a [PSI+] strain generated by protein infection. (JPG 78 kb)

Supplementary Figure 3

Sedimentation analysis of strong and weak [PSI+] strains generated by protein infection. (JPG 55 kb)

Supplementary Figure 4

Induction of [PSI+] by over-expression of Sup-NM-GFP in [psi-][PIN+] strain. (JPG 44 kb)

Supplementary Figure 5

Induction of prion state by partially purified prion particles derived from strong and weak [PSI+] strains. (JPG 75 kb)

Supplementary Figure 6

Partial proteolysis of Sup-NM fibres formed at 4, 23, or 37°C. (JPG 51 kb)

Supplementary Figure 7

Curing efficiency of strong and weak [PSI+] strains generated by protein infection, by Hsp104 over-expression. (JPG 45 kb)

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Tanaka, M., Chien, P., Naber, N. et al. Conformational variations in an infectious protein determine prion strain differences. Nature 428, 323–328 (2004). https://doi.org/10.1038/nature02392

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