Hsp90-mediated Assembly of the 26 S Proteasome Is Involved in Major Histocompatibility Complex Class I Antigen Processing

Abstract

Heat shock protein 90 (hsp90) and the proteasome activator PA28 stimulate major histocompatibility complex (MHC) class I antigen processing. It is unknown whether hsp90 influences the proteasome activity to produce T cell epitopes, although association of PA28 with the 20 S proteasome stimulates the enzyme activity. Here, we show that hsp90 is essential in assembly of the 26 S proteasome and as a result, is involved in epitope production. Addition of recombinant hsp90␣ to cell lysate enhanced chymotrypsinlike activity of the 26 S proteasome in an ATP-dependent manner as determined by an in-gel hydrolysis assay. We successfully pulled down histidine-tagged hsp90␣-and PA28␣-induced, newly assembled 26 S proteasomes from the cell extracts for in vitro epitope production assay, and we found these structures to be sensitive to geldanamycin, an hsp90 inhibitor. We found a cleaved epitope unique to the proteasome pulled down by both hsp90␣ and PA28␣, whereas two different epitopes were identified in the hsp90␣-and PA28␣-pulldowns, respectively. Processing of these respective peptides in vivo was enhanced faithfully by the protein combinations used for the proteasome pulldowns. Inhibition of hsp90 in vivo by geldanamycin partly disrupted the 26 S proteasome structure, consistent with down-regulated MHC class I expression. Our results indicate that hsp90 facilitates MHC class I antigen processing through epitope production in a complex of the 26 S proteasome.

Heat shock protein 90 (HSP90) contributes to cytosolic translocation of extracellular antigen for cross-presentation by dendritic cells

“…Heat shock protein 90 (HSP90) has been suggested to play a role in the endogenous MHC I antigen processing pathway (12)(13)(14)(15). HSP90 associates with peptides as postproteasomal degradation products (14) or with newly synthesized polypeptides destined for degradation by the proteasome (13).…”

Hsp90 and ECM29 Are Important to Maintain the Integrity of Mammalian 26S Proteasome

“…For the first time, we ap- plied this method directly on WT and ECM29 deficient mice liver and identified Hsp90 by MS. In mammalian cells, only one study discussed the involvement of Hsp90 in the proteasome assembly in vitro [37]. In yeast, previous studies [35] [36] revealed an interaction between the proteasome and Hsp90.…”

“…Hsp90α is inducible under stress conditions, while Hsp90β is constitutively expressed. Recent studies mainly in yeast also implied that Hsp90 is involved in the maintenance of the 26S structure of the proteasome [35]- [37].…”

Alteration of the proteostasis network of plant cells promotes the post-endoplasmic reticulum trafficking of recombinant mutant (L444P) human β-glucocerebrosidase

“…Among the putative functions of HSP90, some of which are achieved along with its partners (the HSP70-HSP90 system, and/or co-chaperone p23), 51 HSP90 may participate in the "extraction" of soluble ERAD substrates from the ER lumen to the cytosol. 52 It may further contribute to the assembly and integrity of the 26S proteasome 53,54 and to directing misfolded proteins to the ubiquitin-proteasome degradation pathway. 40 One of the known effects of celastrol is an inhibition of the expression of the gene encoding this protein.…”