title: Disrupting the cortical actin cytoskeleton points to two distinct mechanisms of yeast [PSI+] prion formation.
abstract: Mammalian and fungal prions arise de novo; however, the mechanism is poorly understood in molecular terms. One strong possibility is that oxidative damage to the non-prion form of a protein may be an important trigger influencing the formation of its heritable prion conformation. We have examined the oxidative stress-induced formation of the yeast [PSI+] prion, which is the altered conformation of the Sup35 translation termination factor. We used tandem affinity purification (TAP) and mass spectrometry to identify the proteins which associate with Sup35 in a tsa1 tsa2 antioxidant mutant to address the mechanism by which Sup35 forms the [PSI+] prion during oxidative stress conditions. This analysis identified several components of the cortical actin cytoskeleton including the Abp1 actin nucleation promoting factor, and we show that deletion of the ABP1 gene abrogates oxidant-induced [PSI+] prion formation. The frequency of spontaneous [PSI+] prion formation can be increased by overexpression of Sup35 since the excess Sup35 increases the probability of forming prion seeds. In contrast to oxidant-induced [PSI+] prion formation, overexpression-induced [PSI+] prion formation was only modestly affected in an abp1 mutant. Furthermore, treating yeast cells with latrunculin A to disrupt the formation of actin cables and patches abrogated oxidant-induced, but not overexpression-induced [PSI+] prion formation, suggesting a mechanistic difference in prion formation. [PIN+], the prion form of Rnq1, localizes to the IPOD (insoluble protein deposit) and is thought to influence the aggregation of other proteins. We show Sup35 becomes oxidized and aggregates during oxidative stress conditions, but does not co-localize with Rnq1 in an abp1 mutant which may account for the reduced frequency of [PSI+] prion formation.
authors.affiliation: University of Manchester, Faculty of Biology, Medicine and Health, The Michael Smith Building, Manchester, Unted Kindom.
authors.forename: Shaun H
authors.initials: SH
authors.lastname: Speldewinde
authors.affiliation: University of Manchester, Faculty of Biology, Medicine and Health, The Michael Smith Building, Manchester, Unted Kindom.
authors.forename: Victoria A
authors.identifier: 0000-0002-2475-3408
authors.initials: VA
authors.lastname: Doronina
authors.affiliation: Kent Fungal Group, School of Biosciences, University of Kent, Canterbury, Kent, United Kingdom.
authors.forename: Mick F
authors.initials: MF
authors.lastname: Tuite
authors.affiliation: University of Manchester, Faculty of Biology, Medicine and Health, The Michael Smith Building, Manchester, Unted Kindom.
authors.forename: Chris M
authors.identifier: 0000-0002-0616-6576
authors.initials: CM
authors.lastname: Grant
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D010454:Peptide Termination Factors / Q000235:genetics / Q000378:metabolism*; D010544:Peroxidases / Q000235:genetics / Q000378:metabolism; D011328:Prions / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D053778:Thiazolidines / Q000494:pharmacology
chemical_list: C062612:ABP1 protein, S cerevisiae; D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D008840:Microfilament Proteins; C111870:PAN1 protein, S cerevisiae; D010454:Peptide Termination Factors; D011328:Prions; C404807:RNQ1 protein, S cerevisiae; C068068:SUP35 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D053778:Thiazolidines; C072061:coronin proteins; D010544:Peroxidases; C496253:Tsa1 protein, S cerevisiae; C496254:Tsa2 protein, S cerevisiae; C037067:latrunculin A