Functional Interaction of the Ankylosing Spondylitis-Associated Endoplasmic Reticulum Aminopeptidase 2 With the HLA-B*27 Peptidome in Human Cells.
To determine the influence of endoplasmic reticulum aminopeptidase 2 (ERAP-2) expression on the HLA-B*27 peptidome in live cells.



Using immunoaffinity chromatography and acid extraction, HLA-B*27:05-bound peptides were isolated from 2 ERAP-2-negative lymphoblastoid cell lines and 1 ERAP-2-positive lymphoblastoid cell line expressing functionally indistinguishable ERAP-1 variants. More than 2,000-4,000 B*27:05 ligands were identified from each cell line, and their relative abundance was established by quantitative tandem mass spectrometry and MaxQuant-based peptide analyses. Pairwise comparisons were used to determine the structural features of peptides whose relative abundance was dependent on the presence of ERAP-2. Synthetic peptide digestions were performed with recombinant ERAP-1 and ERAP-2. Peptide affinity was estimated with standard algorithms.



The B*27:05 peptidome from ERAP-2-positive cells showed 3-4% fewer peptides with N-terminal basic residues than did the peptidome from ERAP-2-negative cells. Among the shared peptides, those most abundant in the presence of ERAP-2 included more nonamers, fewer decamers, and fewer N-terminal basic residues than the peptides predominant in ERAP-2-negative cells. These ERAP-2-dependent changes did not alter the global affinity of the B*27:05 peptidome.



ERAP-2 significantly influences the B*27:05-bound peptidome by destroying some ligands and decreasing the abundance of many more ligands with N-terminal basic residues, while increasing the abundance of nonamers. The former effects are best explained by direct ERAP-2 trimming. The effects on peptide length might be attributed to ERAP-2-induced activation of ERAP-1 trimming. These data support the notion of a peptide-mediated mechanism as the basis for the association of ERAP-2 with ankylosing spondylitis. Analogous effects on other major histocompatibility complex class I peptidomes might explain the involvement of ERAP-2 in HLA-B27-negative spondyloarthritis.
Centro de Biología Molecular Severo Ochoa (Consejo Superior de Investigaciones Científicas and Universidad Autónoma), Madrid, Spain.
Adrian
A
Martín-Esteban
Centro de Biología Molecular Severo Ochoa (Consejo Superior de Investigaciones Científicas and Universidad Autónoma), Madrid, Spain.
Pablo
P
Guasp
Technion-Israel Institute of Technology, Haifa, Israel.
Eilon
E
Barnea
Technion-Israel Institute of Technology, Haifa, Israel.
Arie
A
Admon
Centro de Biología Molecular Severo Ochoa (Consejo Superior de Investigaciones Científicas and Universidad Autónoma), Madrid, Spain. aldecastro@cbm.csic.es.
José A
JA
López de Castro
D000626:Aminopeptidases / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015153:Blotting, Western; D002460:Cell Line; D005838:Genotype; D015796:HLA-B27 Antigen / Q000378:metabolism*; D006801:Humans; D008214:Lymphocytes / Q000378:metabolism*; D015778:Minor Histocompatibility Antigens / Q000235:genetics / Q000494:pharmacology; D010455:Peptides / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins; D013167:Spondylitis, Ankylosing / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
C464936:HLA-B*27:05 antigen; D015796:HLA-B27 Antigen; D015778:Minor Histocompatibility Antigens; D010455:Peptides; D011994:Recombinant Proteins; D000626:Aminopeptidases; C464841:ERAP1 protein, human; C503944:ERAP2 protein, human