Abstract
The class I protein HLA-B27 confers susceptibility to inflammatory arthritis in humans and when overexpressed in rodents for reasons that remain unclear. We demonstrated previously that HLA-B27 heavy chains (HC) undergo endoplasmic reticulum (ER)-associated degradation. We report here that HLA-B27 HC also forms two types of aberrant disulfide-linked complexes (dimers) during the folding and assembly process that can be distinguished by conformation-sensitive antibodies W6/32 and HC10. HC10-reactive dimers form immediately after HC synthesis in the ER and constitute at least 25% of the HC pool, whereas W6/32-reactive dimers appear several hours later and represent less than 10% of the folded HC. HC10-reactive dimers accumulate in the absence of tapasin or beta(2)-microglobulin, whereas W6/32-reactive dimers are not detected. Efficient formation of W6/32-reactive dimers appears to depend on the transporter associated with antigen processing, tapasin, and beta(2)-microglobulin. The unpaired Cys(67) and residues at the base of the B pocket that dramatically impair HLA-B27 HC folding are critical for the formation of HC10-reactive ER dimers. Although certain other alleles also form dimers late in the assembly pathway, ER dimerization of HLA-B27 may be unique. These results demonstrate that residues comprising the HLA-B27 B pocket result in aberrant HC folding and disulfide bond formation, and thus confer unusual properties on this molecule that are unrelated to peptide selection per se, yet may be important in disease pathogenesis.
Highlights
The class I protein HLA-B27 confers susceptibility to inflammatory arthritis in humans and when overexpressed in rodents for reasons that remain unclear
Further retention of partially folded heavy chains (HC)/2m heterodimers is mediated by tapasin, an major histocompatibility complex (MHC) class I-specific chaperone which forms a bridge between HC and TAP in the peptide loading complex
Our results indicate that HLA-B27 HC form two types of aberrant disulfide-linked complexes that are distinguishable by several criteria, including kinetics and site of formation, migration on nonreducing SDS-PAGE, dependence on 2m, TAP, and tapasin, and recognition by monoclonal antibodies HC10 and W6/32. Sialylated forms (W6/32)
Summary
HLA-B27 is highly associated with susceptibility to ankylosing spondylitis and other human spondyloarthropathies [20], and can cause spondyloarthropathy-like inflammatory conditions when overexpressed in rodents [21, 22]. The same spontaneous arthritis phenotype has been reported in the absence of HLA-B27, in either 2m or TAP-deficient mice with a mixed genetic background [26] These studies raise the possibility that class I HC misfolding in general, and intracellular sequelae might be a critical component of the disease mechanism, perhaps through the generation of ER stress [27]. Along these lines, effects of HLAB27 expression in monocytes and epithelial cells that cannot be explained by its function as an antigen presenting molecule have been described (28 –30). Our studies emphasize abnormalities in the folding of HLA-B27 that may be unique to this allele, and could be involved in the pathogenesis of spondyloarthropathies
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