3 years ago

The Role of Autophagy in the Degradation of Misfolded HLA-B27 Heavy Chains

Robert A. Colbert, Keith A. Sikora, Gerlinde Layh-Schmitt, Antony Cougnoux, Fatemeh Navid
Objective To determine whether autophagy is involved in the degradation of misfolded HLA-B27 in experimental spondyloarthritis. Methods Bone marrow derived macrophages from HLA-B27 and human β2m (hβ2m) transgenic rats were examined with and without IFNγ and proteasome or autophagy inhibitors. Immunoprecipitation, western blotting, and immunofluorescence were used to measure HLA-B27 heavy chains and autophagy. Autophagy was induced using rapamycin. HLA-B7/hβ2m transgenic and wild type rat macrophages were used as controls. Results HLA-B27-expressing macrophages showed similar LC3B-II levels compared to both controls, before and after manipulating autophagy. Blocking autophagic flux with bafilomycin resulted in the accumulation of misfolded HLA-B27 dimers and oligomers as well as monomers, comparable to blocking endoplasmic reticulum-associated degradation (ERAD) with the proteasome inhibitor bortezomib. HLA-B7 monomers also accumulated after blocking each degradation pathway. The proportion of ubiquitinated heavy chains was ~3-fold lower for HLA-B27 compared to HLA-B7. Activation of autophagy with rapamycin rapidly eliminated ~50% of misfolded HLA-B27, while folded HLA-B27 or HLA-B7 monomeric heavy chains were minimally affected. Conclusion These results demonstrate for the first time that both autophagy and ERAD play a role in the elimination of excess HLA class I heavy chains expressed in transgenic rats. We found no evidence that HLA-B27 expression modulated the autophagy pathway. Our results suggest that impaired ubiquitination of HLA-B27 may play a role in the accumulation of misfolded disulfide-linked dimers, whose elimination can be enhanced by activation of autophagy. Manipulation of the autophagy pathway should be further investigated as a potential therapeutic avenue in spondyloarthritis. This article is protected by copyright. All rights reserved.

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/art.40414

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