HLA-B27 in Transgenic Rats Forms Disulfide-Linked Heavy Chain Oligomers and Multimers That Bind to the Chaperone BiP

Abstract

To test the hypothesis that HLA-B27 predisposes to disease by forming disulfide-linked homodimers, we examined rats transgenic for HLA-B27, mutant Cys67Ser HLA-B27, or HLA-B7. In splenic Con A blasts from high transgene copy B27 lines that develop inflammatory disease, the anti-H chain mAb HC10 precipitated four bands of molecular mass 78–105 kDa and additional higher molecular mass material, seen by nonreducing SDS-PAGE. Upon reduction, all except one 78-kDa band resolved to 44 kDa, the size of the H chain monomer. The 78-kDa band was found to be BiP/Grp78, and the other high molecular mass material was identified as B27 H chain. Analysis of a disease-resistant low copy B27 line showed qualitatively similar high molecular mass bands that were less abundant relative to H chain monomer. Disease-prone rats with a Cys67Ser B27 mutant showed B27 H chain bands at 95 and 115 kDa and a BiP band at 78 kDa, whereas only scant high molecular mass bands were found in cells from control HLA-B7 rats. 125I-surface labeled B27 oligomers were immunoprecipitated with HC10, but not with a mAb to folded B27-β2-microglobulin-peptide complexes. Immunoprecipitation of BiP with anti-BiP Abs coprecipitated B27 H chain multimers. Folding and maturation of B27 were slow compared with B7. These data indicate that disulfide-linked intracellular H chain complexes are more prone to form and bind BiP in disease-prone wild-type B27 and B27-C67S rats than in disease-resistant HLA-B7 rats. The data support the hypothesis that accumulation of misfolded B27 participates in the pathogenesis of B27-associated disease.

Endoplasmic reticulum stress and the unfolded protein response are linked to synergistic IFN‐β induction via X‐box binding protein 1

“…We also examined rat macrophages overexpressing HLA-B7, an allele that does not misfold or activate the UPR [26,28], even with up-regulation [30]. XBP-1 splicing and IFN-b induction in response to LPS were not different between HLA-B7-Tg cells and WT controls (Fig.…”

“…HLA-B27 is an MHC-encoded class I protein that is linked to the development of spondyloarthritis (SpA) including ankylosing spondylitis [22,23], and when expressed in rats along with human b 2 m (HLA-B27-Tg) results in the development of an inflammatory disease that recapitulates many features of human SpA [24]. A proportion of HLA-B27 misfolds in the ER, resulting in increased degradation by ERAD [25], and formation of disulfide-linked and BiP-bound complexes [26][27][28]. Up-regulation of HLA-B27 exacerbates misfolding and activates the UPR in macrophages from Tg rats [29,30].…”

“…Up-regulation of HLA-B27 exacerbates misfolding and activates the UPR in macrophages from Tg rats [29,30]. HLA-B27 misfolding has been correlated with disease in these animals [28], but links between protein misfolding and the development of the immune dysregulation have not been established.…”

“…Up-regulation of HLA-B27 exacerbates misfolding and activates the UPR in macrophages from Tg rats [29,30]. HLA-B27 misfolding has been correlated with disease in these animals [28], but links between protein misfolding and the development of the immune dysregulation have not been established.Although much is known about innate immune responses to external threats, and in parallel, ER signaling pathways that orchestrate the response to internal stress, little is known about how these pathways intersect. Here, we show that pharmacologic agents and proteins that misfold and activate the UPR synergize strongly with certain PRR agonists in the induction of IFN-b, and that XBP-1 is required for this effect.…”

Additional human β2‐microglobulin curbs HLA–B27 misfolding and promotes arthritis and spondylitis without colitis in male HLA–B27–transgenic rats

“…Cell-surface-expressed MHC class I molecules are thought normally to be formed of folded noncovalent complexes consisting of the heavy chain, ␤ 2 m, and peptide that are assembled in the endoplasmic reticulum through association with a series of chaperones (19). Both in cells from the B27-transgenic rats and in human and rat cell lines, B27 heavy chain has been shown to be unusually inefficient in its assembly with ␤ 2 m and peptide, readily forming heavy chain homodimers (20)(21)(22)(23). Based on these observations, the interaction of B27 heavy chain homodimers with receptors of innate immunity (24) and the triggering of the unfolded protein response (UPR) and downstream inflammation by B27 heavy chain misfolding (25) have been proposed as potential mechanisms to explain the role of B27 in disease.…”