The elimination of the yeast [PSI+] prion by guanidine hydrochloride is the result of Hsp104 inactivation.
In the yeast Saccharomyces cerevisiae, Sup35p (eRF3), a subunit of the translation termination complex, can take up a prion-like, self-propagating conformation giving rise to the non-Mendelian [PSI+] determinant. The replication of [PSI+] prion seeds can be readily blocked by growth in the presence of low concentrations of guanidine hydrochloride (GdnHCl), leading to the generation of prion-free [psi-] cells. Here, we provide evidence that GdnHCl blocks seed replication in vivo by inactivation of the molecular chaperone Hsp104. Although growth in the presence of GdnHCl causes a modest increase in HSP104 expression (20-90%), this is not sufficient to explain prion curing. Rather, we show that GdnHCl inhibits two different Hsp104-dependent cellular processes, namely the acquisition of thermotolerance and the refolding of thermally denatured luciferase. The inhibitory effects of GdnHCl protein refolding are partially suppressed by elevating the endogenous cellular levels of Hsp104 using a constitutive promoter. The kinetics of GdnHCl-induced [PSI+] curing could be mimicked by co-expression of an ATPase-negative dominant HSP104 mutant in an otherwise wild-type [PSI+] strain. We suggest that GdnHCl inactivates the ATPase activity of Hsp104, leading to a block in the replication of [PSI+] seeds.
Ferreira
P C
PC
Department of Biosciences, University of Kent, Canterbury, Kent CT2 7NJ, UK.
Ness
F
F
Edwards
S R
SR
Cox
B S
BS
Tuite
M F
MF
D005656:Fungal Proteins / Q000187:drug effects* / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects; D019791:Guanidine / Q000494:pharmacology*; D006360:Heat-Shock Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D006358:Hot Temperature; D007700:Kinetics; D008156:Luciferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010454:Peptide Termination Factors; D011328:Prions / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins
D005656:Fungal Proteins; D006360:Heat-Shock Proteins; D010454:Peptide Termination Factors; D011328:Prions; D011994:Recombinant Proteins; C068068:SUP35 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C070972:HsP104 protein, S cerevisiae; D008156:Luciferases; D019791:Guanidine