Substantial Influence of ERAP2 on the HLA-B40:02 Peptidome: Implications for HLA-B27-Negative Ankylosing Spondylitis

ERAP2 and HLA-B*40:02 are associated with ankylosing spondylitis independently of HLA-B*27. ERAP2 process MHC-I ligands, preferentially trimming N-terminal basic residues. The B*40:02 peptidomes from wild-type and ERAP2-KO cells were compared, which demonstrated a substantial role of ERAP2 on the generation/destruction balance of HLA-B*40:02 ligands. The major effect was on N-terminal residues, although other peptide positions were also affected. We propose that the non-epistatic association of ERAP2 with spondyloarthropathy might be related to processing of peptides with N-terminal basic residues. [Display omitted] Highlights •HLA-B*40:02 and ERAP2 are risk factors for ankylosing spondylitis.•The effects of ERAP2 on the B*40:02 peptidome are defined.•ERAP2 has a major influence mainly due to alterations of N-terminal residues.•These effects provide a basis for the association of ERAP2 with disease. HLA-B*40:02 is one of a few major histocompatibility complex class I (MHC-I) molecules associated with ankylosing spondylitis (AS) independently of HLA-B*27. The endoplasmic reticulum aminopeptidase 2 (ERAP2), an enzyme that process MHC-I ligands and preferentially trims N-terminal basic residues, is also a risk factor for this disease. Like HLA-B*27 and other AS-associated MHC-I molecules, HLA-B*40:02 binds a relatively high percentage of peptides with ERAP2-susceptible residues. In this study, the effects of ERAP2 depletion on the HLA-B*40:02 peptidome were analyzed. ERAP2 protein expression was knocked out by CRISPR in the transfectant cell line C1R-B*40:02, and the differences between the peptidomes from the wild-type and ERAP2-KO cells were determined by label-free quantitative comparisons. The qualitative changes dependent on ERAP2 affected about 5% of the peptidome, but quantitative changes in peptide amounts were much more substantial, reflecting a significant influence of this enzyme on the generation/destruction balance of HLA-B*40:02 ligands. As in HLA-B*27, a major effect was on the frequencies of N-terminal residues. In this position, basic and small residues were increased, and aliphatic/aromatic ones decreased in the ERAP2 knockout. Other peptide positions were also affected. Because most of the non-B*27 MHC-I molecules associated with AS risk bind a relatively high percentage of peptides with N-terminal basic residues, we hypothesize that the non-epistatic association of ERAP2 with AS might be related to the processing of peptides with these res