Hsp31 is a stress-response chaperone that prevents alpha-synuclein aggregation

Chia-Jui Tsai, Purdue University

Abstract

Saccharomyces cerevisiae heat shock protein Hsp31 is a stress-inducible homodimeric protein belonging to DJ-1/ThiJ/PfpI superfamily. Our studies demonstrate that substoichiometric concentrations of Hsp31 can abrogate the aggregation of a broad array of substrates in vitro and that Hsp31 shares a conserved target substrate with human DJ-1, α-synuclein (αSyn). Hsp31 suppresses the in vitro fibrillization of αSyn or aggregation of citrate synthase and insulin. Chaperone activity was also observed in vivo because constitutive overexpression of Hsp31 reduced the incidence of αSyn accumulations and yeast cells are rescued from αSyn generated death upon Hsp31 overexpression. Moreover, we confirmed observations from previous studies that Hsp31 expression is induced during the diauxic phase in basal growth conditions and in cells under αSyn mediated stress. The protective role of Hsp31 against cellular stress has been observed in many studies and our results suggest that the protective effect is achieved by chaperone activity against different kinds of misfolded proteins and in particular, αSyn. We next verified the interaction between αSyn and Hsp31 and identified the nature of the oligomeric species of αSyn capable of interacting with Hsp31. Results from native PAGE gel indicated that Hsp31 interacts with intermediately sized oligomeric species that have a smaller molecular weight than αSyn protofibrils. In contrast, it appears that DJ-1 interacted preferentially with αSyn protofibrils. Furthermore, SDS-PAGE gel result showed that in presence of Hsp31, there was an increased amount of αSyn self-proteolytic products. Based on the observations from the native PAGE gel and SDS-PAGE, we conclude that Hsp31 suppresses oligomerization of αSyn monomer. As a result the higher oligomeric αSyn are reduced and αSyn proteolytic products were increased. There is a conserved cysteine, which was shown to affect the cellular function of DJ-1 upon oxidation, in the DJ-1 superfamily. The only cysteine, which is a key amino acid residue in a putative catalytic triad, in Hsp31 was engineered to examine its functional role in αSyn aggregation. We provide evidence showing that the cysteine mutants of Hsp31 do not greatly affect the in vitro fibrilization of αSyn. Overall, our data have revealed the chaperone activity of Hsp31 with different misfolded proteins. Specifically, Hsp31 was shown to interact with αSyn monomer providing an important clue as to its mode of chaperone activity. The future direction of these studies includes delineating the mechanism by which Hsp31 has chaperone activity, identifying key regulators of Hsp31 chaperone function and the natural substrates of Hsp31.

Degree

Ph.D.

Advisors

Hazbun, Purdue University.

Subject Area

Neurosciences|Biochemistry

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