The interaction of Hsp104 with yeast prion Sup35 as analyzed by fluorescence cross-correlation spectroscopy.
Prions are self-propagating amyloids. Yeast prion [PSI(+)] is a protein-based heritable element, in which amyloid aggregates of the Sup35 protein are transmitted to daughter cells. Hsp104, an ATP-dependent disaggregase, and other chaperones are essential to maintain [PSI(+)]. Although previous reports have demonstrated the physical interactions of Hsp104 and Sup35 amyloids, the mechanism how Hsp104 interacts with Sup35 amyloids remains to be elucidated. Here we investigated the interaction between Hsp104 and Sup35 in the lysates of [PSI(+)] cells using fluorescence cross-correlation spectroscopy (FCCS), which can analyze the codiffusion events of different fluorophores. FCCS analysis showed a strong interaction between Hsp104 and Sup35 in [PSI(+)] lysates, but not in [psi(-)] lysates, suggesting that Hsp104 recognizes the amyloid aggregates of Sup35. Although the interaction was retained in ATP-depleted [PSI(+)] lysates, addition of ATP or guanidine hydrochloride, which is an inhibitor of Hsp104, to [PSI(+)] lysates weakened the interaction.
Laboratory of Molecular Cell Dynamics, Advanced Life Science, Hokkaido University, Sapporo 001-0021, Japan.
Shohei
S
Ohta
Shigeko
S
Kawai-Noma
Akira
A
Kitamura
Chan-Gi
CG
Pack
Masataka
M
Kinjo
Hideki
H
Taguchi
D000682:Amyloid / Q000737:chemistry / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D010454:Peptide Termination Factors / Q000737:chemistry / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence / Q000379:methods
D000682:Amyloid; D006360:Heat-Shock Proteins; D010454:Peptide Termination Factors; C068068:SUP35 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C070972:HsP104 protein, S cerevisiae
FCCS; Fluorescence cross-correlation spectroscopy (FCCS); Hsp104; N and M domains of Sup35; RCA; Sup35; Sup35NM; Yeast prion; fluorescence cross-correlation spectroscopy; relative cross-correlation amplitude