title: [PSI(+)] aggregate enlargement in rnq1 nonprion domain mutants, leading to a loss of prion in yeast.
abstract: [PIN(+)] is the prion form of the Rnq1 protein of unknown function in Saccharomyces cerevisiae. A glutamine/asparagine (Q/N)-rich C-terminal domain is necessary for the propagation of [PIN(+)], whereas the N-terminal region is non-Q/N-rich and considered the nonprion domain. Here, we isolated numerous single-amino-acid mutations in Rnq1, phenotypically similar to Rnq1Δ100, which inhibit [PSI(+)] propagation in the [PIN(+)] state, but not in the [pin(-)] state, when overproduced. The dynamics of the prion aggregates was analyzed by semi-denaturing detergent-agarose gel electrophoresis and fluorescence correlation spectroscopy. The results indicated that [PSI(+)] aggregates were enlarged in mother cells and, instead, not apparently transmitted into daughter cells. Under these conditions, the activity of Hsp104, a known prion disaggregase, was not affected when monitored for the thermotolerance of the rnq1 mutants. These [PSI(+)]-inhibitory rnq1 mutations did not affect [PIN(+)] propagation itself when over-expressed from a strong promoter, but instead destabilized [PIN(+)] when expressed from the weak authentic RNQ1 promoter. The majority of these mutated residues are mapped to the surface, and on one side, of contiguous α-helices of the nonprion domain of Rnq1, suggesting its involvement in interactions with a prion or a factor necessary for prion development.
authors: Department of Basic Medical Sciences, Institute of Medical Science, University of Tokyo, Shirokanedai, Minato-ku, Japan.
authors: Hiroshi
authors: H
authors: Kurahashi
authors: Chan-Gi
authors: CG
authors: Pack
authors: Shoichiro
authors: S
authors: Shibata
authors: Keita
authors: K
authors: Oishi
authors: Yasushi
authors: Y
authors: Sako
authors: Yoshikazu
authors: Y
authors: Nakamura
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D015966:Gene Expression Regulation, Fungal; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011328:Prions / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence / Q000379:methods; D013696:Temperature
chemical_list: D006360:Heat-Shock Proteins; D011328:Prions; C404807:RNQ1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C070972:HsP104 protein, S cerevisiae; D049452:Green Fluorescent Proteins