Misfolding of Proteins with a Polyglutamine Expansion Is Facilitated by Proteasomal Chaperones

Abstract

Deposition of misfolded proteins with a polyglutamine expansion is a hallmark of Huntington disease and other neurodegenerative disorders. Impairment of the proteolytic function of the proteasome has been reported to be both a cause and a consequence of polyglutamine accumulation. Here we found that the proteasomal chaperones that unfold proteins to be degraded by the proteasome but also have non-proteolytic functions colocalized with huntingtin inclusions both in primary neurons and in Huntington disease patients and formed a complex independently of the proteolytic particle. Overexpression of Rpt4 or Rpt6 facilitated aggregation of mutant huntingtin and ataxin-3 without affecting proteasomal degradation. Conversely, reducing Rpt6 or Rpt4 levels decreased the number of inclusions in primary neurons, indicating that endogenous Rpt4 and Rpt6 facilitate inclusion formation. In vitro reconstitution experiments revealed that purified 19S particles promote mutant huntingtin aggregation. When fused to the ornithine decarboxylase destabilizing sequence, proteins with expanded polyglutamine were efficiently degraded and did not aggregate. We propose that aggregation of proteins with expanded polyglutamine is not a consequence of a proteolytic failure of the 20S proteasome. Rather, aggregation is elicited by chaperone subunits of the 19S particle independently of proteolysis.

Spatial control of Shoc2 scaffold-mediated ERK1/2 signaling requires remodeling activity of the ATPase PSMC5

“…Furthermore, PSMC5 unfolds proteins in a proteasomeindependent manner [e.g. transcription complexes or poly(Q) protein aggregates of Huntingtin] (Rousseau et al, 2009;Sun et al, 2002). Here, we provide compelling evidence that PSMC5 is an integral partner in the Shoc2 scaffolding complex.…”

“…For studies in living cells we utilized the tagRFP (tRFP)-fusion protein of Shoc2 described in our earlier studies (Galperin et al, 2012). Given that overexpression of PSMC5 does not impair the function of the proteasome (Rousseau et al, 2009), we utilized a previously characterized fluorescently tagged PSMC5 (CFP-PSMC5) (Laporte et al, 2008). Fluorescence microscopy revealed that Shoc2-tRFP and CFP-PSMC5 are distributed in the cytosol ( Fig.…”

“…For instance, several biochemical and genetic studies have indicated that PSMC5 plays a distinct proteasome-independent role in regulating transcription activation and elongation, DNA repair and chromatin remodeling (Ferdous et al, 2002;Ferry et al, 2009;Gonzalez et al, 2002;Sulahian et al, 2006;Swaffield et al, 1992). PSMC5 is also involved in facilitating misfolding and aggregation of proteins with a poly(Q) expansion in Huntington's disease (Rousseau et al, 2009). These activities rely on the non-proteolytic function of PSMC5 as a remodeling chaperone.…”

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

“…Transfections were done with Lipofectamine 2000 (Invitrogen) and Dharmacon SMARTpool siRNA siGenome DNAJB6 (M-013020-00-0005) and non-targeting siRNA control siGENOME (D-001206- [13][14][15][16][17][18][19][20].…”

Molecular chaperones antagonize proteotoxicity by differentially modulating protein aggregation pathways

“…The proteasomal chaperones, Rpt4 and Rpt6 (AAA ATPase subunits of the 19S proteasomal particle), facilitated the aggregation of different expanded-polyQ disease proteins, huntingtin and ataxin-3, without affecting proteasomal degradation. 90 Furthermore, in vitro reconstitution experiments showed that purified 19S proteasomal particles enhanced aggregation of expanded-polyQ huntingtin protein. 90 Thus, a number of molecular chaperones can promote the assembly of disease proteins into amyloid-like aggregates.…”