title: Critical role of endoplasmic reticulum aminopeptidase 1 in determining the length and sequence of peptides bound and presented by HLA-B27.
abstract: OBJECTIVE
  HLA-B27 and endoplasmic reticulum aminopeptidase 1 (ERAP1) are the two strongest genetic factors predisposing to ankylosing spondylitis (AS). A key aminopeptidase in class I major histocompatibility complex presentation, ERAP1 potentially contributes to the pathogenesis of AS by altering HLA-B27 peptide presentation. The aim of this study was to analyze the effects of ERAP1 on the HLA-B27 peptide repertoire and peptide presentation to cytotoxic T lymphocytes (CTLs).
  
  
  METHODS
  ERAP1-silenced and -competent HeLa.B27 and C1R.B27 cells were isotope-labeled, mixed, lysed, and then immunoprecipitated using W6/32 or ME1 antibodies. Peptides bound to HLA-B27 were eluted and analyzed by tandem mass spectrometry. Selected peptides were synthesized and tested for HLA-B27 binding ability. The effect of ERAP1 silencing/mutation on presentation of an immunodominant viral HLA-B27 epitope, KK10, to CTLs was also studied.
  
  
  RESULTS
  In both HeLa.B27 and C1R.B27 cells, the proportion of 9-mer HLA-B27-bound peptides was decreased by ERAP1 silencing, whereas the percentages of longer peptides (11-13 mer) were increased. Surprisingly, following ERAP1 silencing, C-terminally extended peptides were readily identified. These were better able to bind to HLA-B27 than were N-terminally extended peptides lacking an arginine at position 2. In both HeLa.B27 cells and mouse fibroblasts expressing HLA-B27, the absence of ERAP1 reduced peptide recognition by HLA-B27-restricted KK10-specific CTLs following infection with recombinant vaccinia virus or transfection with minigenes expressing KK10 precursors. Presence of an AS-protective variant of ERAP1, K528R, as compared to wild-type ERAP1, reduced the peptide recognition by KK10 CTLs following transfection with extended KK10 minigenes.
  
  
  CONCLUSIONS
  These results show that ERAP1 directly alters peptide binding and presentation by HLA-B27, thus demonstrating a potential pathogenic mechanism in AS. Inhibition of ERAP1 could potentially be used for treatment of AS and other ERAP1-associated diseases.
authors: University of Oxford, Oxford, UK.
authors: Liye
authors: L
authors: Chen
authors: Roman
authors: R
authors: Fischer
authors: Yanchun
authors: Y
authors: Peng
authors: Emma
authors: E
authors: Reeves
authors: Kirsty
authors: K
authors: McHugh
authors: Nicola
authors: N
authors: Ternette
authors: Tomas
authors: T
authors: Hanke
authors: Tao
authors: T
authors: Dong
authors: Tim
authors: T
authors: Elliott
authors: Nilabh
authors: N
authors: Shastri
authors: Simon
authors: S
authors: Kollnberger
authors: Edward
authors: E
authors: James
authors: Benedikt
authors: B
authors: Kessler
authors: Paul
authors: P
authors: Bowness
mesh_terms: D000626:Aminopeptidases / Q000172:deficiency / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001402:B-Lymphocytes; D002460:Cell Line; D002478:Cells, Cultured; D004195:Disease Models, Animal; D000939:Epitopes / Q000235:genetics; D020868:Gene Silencing / Q000187:drug effects; D015796:HLA-B27 Antigen / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D018345:Mice, Knockout; D015778:Minor Histocompatibility Antigens; D009154:Mutation / Q000235:genetics; D010455:Peptides / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D013167:Spondylitis, Ankylosing / Q000378:metabolism* / Q000473:pathology; D013602:T-Lymphocytes, Cytotoxic / Q000378:metabolism / Q000473:pathology
chemical_list: D000939:Epitopes; D015796:HLA-B27 Antigen; D015778:Minor Histocompatibility Antigens; D010455:Peptides; D034741:RNA, Small Interfering; D000626:Aminopeptidases; C464841:ERAP1 protein, human; C509944:ERAP1 protein, mouse