The DNAJB6 and DNAJB8 Protein Chaperones Prevent Intracellular Aggregation of Polyglutamine Peptides

Judith Gillis,Sabine Schipper-Krom,Katrin Juenemann,Anna Gruber,Silvia Coolen,Rian Van Den Nieuwendijk,Henk Van Veen,Hermen Overkleeft,Joachim Goedhart,Harm H Kampinga,Eric A Reits

doi:10.1074/jbc.m112.421685

Judith Gillis, Sabine Schipper-Krom + Show 9 more

Open Access

https://doi.org/10.1074/jbc.m112.421685

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Abstract

Fragments of proteins containing an expanded polyglutamine (polyQ) tract are thought to initiate aggregation and toxicity in at least nine neurodegenerative diseases, including Huntington's disease. Because proteasomes appear unable to digest the polyQ tract, which can initiate intracellular protein aggregation, preventing polyQ peptide aggregation by chaperones should greatly improve polyQ clearance and prevent aggregate formation. Here we expressed polyQ peptides in cells and show that their intracellular aggregation is prevented by DNAJB6 and DNAJB8, members of the DNAJ (Hsp40) chaperone family. In contrast, HSPA/Hsp70 and DNAJB1, also members of the DNAJ chaperone family, did not prevent peptide-initiated aggregation. Intriguingly, DNAJB6 and DNAJB8 also affected the soluble levels of polyQ peptides, indicating that DNAJB6 and DNAJB8 inhibit polyQ peptide aggregation directly. Together with recent data showing that purified DNAJB6 can suppress fibrillation of polyQ peptides far more efficiently than polyQ expanded protein fragments in vitro, we conclude that the mechanism of DNAJB6 and DNAJB8 is suppression of polyQ protein aggregation by directly binding the polyQ tract.

Highlights

Intracellular aggregation of polyglutamine proteins can be prevented by the chaperones DNAJB6 and DNAJB8
Together with recent data showing that purified DNAJB6 can suppress fibrillation of polyQ peptides far more efficiently than polyQ expanded protein fragments in vitro, we conclude that the mechanism of DNAJB6 and DNAJB8 is suppression of polyQ protein aggregation by directly binding the polyQ tract
DNAJB6b and DNAJB8 Reduce Aggregation of PolyQ Peptides—To determine the effect of chaperones on polyQ peptide aggregation in living cells, we used GFP-ubiquitin-polyQ (GFPUb-polyQ) constructs that are immediately cleaved upon expression into GFP-Ub and polyQ peptides by Ub C-terminal hydrolases (Fig. 1A) [39]

Summary

Background

Intracellular aggregation of polyglutamine (polyQ) proteins can be prevented by the chaperones DNAJB6 and DNAJB8. Fragments of proteins containing an expanded polyglutamine (polyQ) tract are thought to initiate aggregation and toxicity in at least nine neurodegenerative diseases, including Huntington’s disease. We expressed polyQ peptides in cells and show that their intracellular aggregation is prevented by DNAJB6 and DNAJB8, members of the DNAJ (Hsp40) chaperone family. In a recent comparative screen of all members of the HSPA, HSPH (Hsp110), and DNAJ (sub)families, DNAJB6 and DNAJB8 were identified as the two most potent suppressors of aggregation and related toxicity of expanded polyQ proteins [33]. When examining the characteristics of these two chaperones in more detail, we find that DNAJB6 and DNAJB8 can directly suppress aggregation of polyQ peptides generated in living cells This action is largely dependent on the serine-rich region (SSF-SST) within the C terminus and less dependent on interaction with Hsp. The ability of DNAJB6 and DNAJB8 to reduce aggregation of expanded polyQ peptides prevents these polyQ peptides from acting as inducers of aggregation and improves their clearance

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION