ERAP1 reduces accumulation of aberrant and disulfide-linked forms of HLA-B27 on the cell surface.
Endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) variants contribute to the risk of ankylosing spondylitis in HLA-B27 positive individuals, implying a disease-related interaction between these gene products. The aim of this study was to determine whether reduced ERAP1 expression would alter the cell surface expression of HLA-B27 and the formation of aberrant disulfide-linked forms that have been implicated in the pathogenesis of spondyloarthritis.



ERAP1 expression was knocked down in monocytic U937 cells expressing HLA-B27 and endogenous HLA class I. The effect of ERAP1 knockdown on the accumulation HLA-B alleles (B18, B51, and B27) was assessed using immunoprecipitation, isoelectric focusing, and immunoblotting, as well as flow cytometry with antibodies specific for different forms of HLA-B27. Cell surface expression of aberrant disulfide-linked HLA-B27 dimers was assessed by immunoprecipitation and electrophoresis on non-reducing polyacrylamide gels.



ERAP1 knockdown increased the accumulation of HLA-B27 on the cell surface including disulfide-linked dimers, but had no effect on levels of HLA-B18 or -B51. Antibodies with unique specificity for HLA-B27 confirmed increased cell surface expression of complexes shown previously to contain long peptides. IFN-γ treatment resulted in striking increases in the expression of disulfide-linked HLA-B27 heavy chains, even in cells with normal ERAP1 expression.



Our results suggest that normal levels of ERAP1 reduce the accumulation of aberrant and disulfide-linked forms of HLA-B27 in monocytes, and thus help to maintain the integrity of cell surface HLA-B27 complexes.
Pediatric Translational Research Branch, National Institute of Arthritis Musculoskeletal and Skin Diseases (NIAMS), National Institutes of Health (NIH), Bethesda, MD 20892, United States.
Tri M
TM
Tran
Pediatric Translational Research Branch, National Institute of Arthritis Musculoskeletal and Skin Diseases (NIAMS), National Institutes of Health (NIH), Bethesda, MD 20892, United States.
Sohee
S
Hong
Pediatric Translational Research Branch, National Institute of Arthritis Musculoskeletal and Skin Diseases (NIAMS), National Institutes of Health (NIH), Bethesda, MD 20892, United States.
Jehad H
JH
Edwan
Pediatric Translational Research Branch, National Institute of Arthritis Musculoskeletal and Skin Diseases (NIAMS), National Institutes of Health (NIH), Bethesda, MD 20892, United States. Electronic address: colbertr@mail.nih.gov.
Robert A
RA
Colbert
D000626:Aminopeptidases / Q000378:metabolism*; D005434:Flow Cytometry; D055785:Gene Knockdown Techniques; D020022:Genetic Predisposition to Disease; D015796:HLA-B27 Antigen / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D015778:Minor Histocompatibility Antigens / Q000378:metabolism*; D013167:Spondylitis, Ankylosing / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D020298:U937 Cells
D015796:HLA-B27 Antigen; D015778:Minor Histocompatibility Antigens; D000626:Aminopeptidases; C464841:ERAP1 protein, human
Ankylosing spondylitis; Epistasis; Major histocompatibility complex; Misfolding