Human DnaJB6 Antiamyloid Chaperone Protects Yeast from Polyglutamine Toxicity Separately from Spatial Segregation of Aggregates

ABSTRACT

Polyglutamine (polyQ) aggregates are associated with pathology in protein-folding diseases and with toxicity in the yeast Saccharomyces cerevisiae. Protection from polyQ toxicity in yeast by human DnaJB6 coincides with sequestration of aggregates. Gathering of misfolded proteins into deposition sites by protein quality control (PQC) factors has led to the view that PQC processes protect cells by spatially segregating toxic aggregates. Whether DnaJB6 depends on this machinery to sequester polyQ aggregates, if this sequestration is needed for DnaJB6 to protect cells, and the identity of the deposition site are unknown. Here, we found DnaJB6-driven deposits share characteristics with perivacuolar insoluble protein deposition sites (IPODs). Binding of DnaJB6 to aggregates was necessary, but not enough, for detoxification. Focal formation required a DnaJB6-Hsp70 interaction and actin, polyQ could be detoxified without sequestration, and segregation of aggregates alone was not protective. Our findings suggest DnaJB6 binds to smaller polyQ aggregates to block their toxicity. Assembly and segregation of detoxified aggregates are driven by an Hsp70- and actin-dependent process. Our findings show sequestration of aggregates is not the primary mechanism by which DnaJB6 suppresses toxicity and raise questions regarding how and when misfolded proteins are detoxified during spatial segregation.

KEYWORDS:

• DnaJB6
• Huntington's disease
• J-protein
• molecular chaperone
• heat shock protein (HSP)
• amyloid

ACKNOWLEDGMENTS

We thank our NIH and NIDDK colleagues for helpful discussions, Ruchika Sharma for plasmids, and Mike Reidy for valuable technical and intellectual support. Plasmid pCUP1-DuDre-Atg8-404 was a gift from Zhiping Xie (69201; Addgene).

This work was supported by the Intramural Program of the National Institutes of Health, National Institute of Diabetes, and Digestive and Kidney Diseases.

N.L.K. initiated the project and, with J.K. and D.C.M., conceived ideas for experimental approaches, analyzed data, and interpreted results. D.C.M. constructed strains and some plasmids; J.K. and N.L.K. conducted experiments and wrote the paper with D.C.M.

We have no conflicts of interest to declare.