Ordered Assembly of Heat Shock Proteins, Hsp26, Hsp70, Hsp90, and Hsp104, on Expanded Polyglutamine Fragments Revealed by Chemical Probes

• Published in: The Journal of biological chemistry 2011-11, Vol.286 (47), p.40486-40493
• Main Authors: Walter, Gladis M., Smith, Matthew C., Wisén, Susanne, Basrur, Venkatesha, Elenitoba-Johnson, Kojo S.J., Duennwald, Martin L., Kumar, Anuj, Gestwicki, Jason E.
• Format: Article
• Language: English
• Subjects: Chaperone Chaperonin; Chemical Biology; Chromatography, Liquid; Heat Shock Protein; Heat-Shock Proteins - metabolism; HSP70 Heat-Shock Proteins - metabolism; HSP90 Heat-Shock Proteins - metabolism; Immunoprecipitation; Mass Spectrometry (MS); Molecular Bases of Disease; Molecular Probes - metabolism; Peptide Fragments - chemistry; Peptide Fragments - genetics; Peptide Fragments - metabolism; Peptides - chemistry; Peptides - genetics; Peptides - metabolism; Polyglutamine; Protein Binding; Protein Multimerization; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - metabolism; Tandem Mass Spectrometry; Time Factors; Transcriptional Activation
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.284448

In Saccharomyces cerevisae, expanded polyglutamine (polyQ) fragments are assembled into discrete cytosolic aggregates in a process regulated by the molecular chaperones Hsp26, Hsp70, Hsp90, and Hsp104. To better understand how the different chaperones might cooperate during polyQ aggregation, we used sequential immunoprecipitations and mass spectrometry to identify proteins associated with either soluble (Q25) or aggregation-prone (Q103) fragments at both early and later times after induction of their expression. We found that Hsp26, Hsp70, Hsp90, and other chaperones interact with Q103, but not Q25, within the first 2 h. Further, Hsp70 and Hsp90 appear to be partially released from Q103 prior to the maturation of the aggregates and before the recruitment of Hsp104. To test the importance of this seemingly ordered process, we used a chemical probe to artificially enhance Hsp70 binding to Q103. This treatment retained both Hsp70 and Hsp90 on the polyQ fragment and, interestingly, limited subsequent exchange for Hsp26 and Hsp104, resulting in incomplete aggregation. Together, these results suggest that partial release of Hsp70 may be an essential step in the continued processing of expanded polyQ fragments in yeast.