In vivo monitoring of the prion replication cycle reveals a critical role for Sis1 in delivering substrates to Hsp104.
Prions in Saccharomyces cerevisiae are inherited ordered aggregates reliant upon the disaggregase Hsp104 for stable maintenance. The function of other factors in the natural prion cycle is unclear. We constructed yeast-bacterial chimeric chaperones to resolve the roles of Hsp104 domains, and by extension chaperones that interact with these domains, in prion propagation. Our results show that, as with amorphous aggregate dissolution, the Hsp70/40 system recruits prion substrates to Hsp104 via its top ring. By adapting our chimera to couple to an inactive protease "trap," we monitored the reaction products of prion propagation in vivo. We find that prion maintenance is accompanied by translocation of prion proteins through Hsp104 hexamers and that both processes critically rely upon the Hsp40 Sis1. Our data suggest that yeast prion replication is a natural extension of chaperone activity in dissolving amorphous aggregates, distinguished from its ancestral reaction by the ordered, self-propagating structure of the substrate.
Tipton
Kimberly A
KA
California Institute of Quantitative Biomedical Science, University of California, San Francisco, 1700 4th Street, San Francisco, CA 94158-2542, USA.
Verges
Katherine J
KJ
Weissman
Jonathan S
JS
D004261:DNA Replication; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D050956:HSP40 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011328:Prions / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D017434:Protein Structure, Tertiary / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013379:Substrate Specificity
D005656:Fungal Proteins; D050956:HSP40 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D011328:Prions; C070320:SIS1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins