HLA B-27 rats peptidome - The HLA-B27 peptidome in vivo in spondyloarthritis-susceptible HLA-B27 transgenic rats and the effect of ERAP1 deletion

HLA-B27 is a class I major histocompatibility (MHC-I) allele that confers susceptibility to the rheumatic disease ankylosing spondylitis (AS) by an unknown mechanism. ERAP1 is an aminopeptidase that trims peptides in the endoplasmic reticulum for binding to MHC-I molecules. ERAP1 shows genetic epistasis with HLA-B27 in conferring susceptibility to AS. Male HLA-B27 transgenic rats develop arthritis and serve as an animal model of AS, whereas female B27 transgenic rats remain healthy. We used large-scale quantitative mass-spectrometry to identify over 15,000 unique HLA-B27 peptide ligands, isolated after immunoaffinity purification of the B27 molecules from the spleens of HLA-B27 transgenic rats. Heterozygous deletion of ERAP1, which reduced ERAP1 level to less than half, had no qualitative or quantitative effects on the B27 peptidome. Homozygous deletion of ERAP1 affected approximately one third of the B27 peptidome, but left most of the B27 peptidome unchanged, suggesting the possibility that some of the HLA-B27 immunopeptidome is not processed in the presence of ERAP1. Deletion on ERAP1 was permissive for the AS-like phenotype. Deletion of ERAP1 increased mean peptide length, and increased the frequency of C-terminal hydrophobic residues and of N-terminal Ala, Ser or Lys. The presence of ERAP1 increased the frequency of C-terminal Lys and Arg, of Glu and Asp at intermediate residues, and of N-terminal Gly. Several peptides of potential interest in AS pathogenesis, previously identified in human cell lines, were isolated. However, rats susceptible to arthritis had B27 peptidomes similar to those of non-susceptible rats, and no peptides were found to be uniquely associated with arthritis. Whether specific B27-bound peptides are required for AS pathogenesis remains to be determined.

HLA-B27是一类I类主要组织相容性复合体（major histocompatibility complex class I, MHC-I）等位基因，可通过尚不明确的机制增加风湿性疾病强直性脊柱炎（ankylosing spondylitis, AS）的易感风险。ERAP1是一种氨肽酶（aminopeptidase），能够在内质网（endoplasmic reticulum）中对肽段进行修剪，使其得以结合MHC-I分子。ERAP1与HLA-B27在增加强直性脊柱炎易感风险的过程中存在遗传上位性效应。携带HLA-B27转基因的雄性大鼠会诱发关节炎，可作为强直性脊柱炎的动物模型，而同类转基因雌性大鼠则保持健康。本研究采用大规模定量质谱技术，从携带HLA-B27转基因大鼠的脾脏中免疫亲和纯化B27分子后，分离鉴定出超过15000种独特的HLA-B27肽配体。ERAP1的杂合缺失可使ERAP1表达水平降至原水平的一半以下，该操作对B27肽组（peptidome）的定性与定量特征均无显著影响。ERAP1的纯合缺失则影响了约三分之一的B27肽组，但绝大多数B27肽组未发生改变，这提示部分HLA-B27免疫肽组（immunopeptidome）在ERAP1存在的情况下并未被加工处理。ERAP1缺失可允许强直性脊柱炎样表型的产生。ERAP1缺失会提升肽段的平均长度，并增加C端疏水氨基酸残基以及N端丙氨酸（alanine, Ala）、丝氨酸（serine, Ser）或赖氨酸（lysine, Lys）的出现频率。而ERAP1的存在则会提升C端赖氨酸（Lys）、精氨酸（arginine, Arg）的出现频率，以及肽段中间位置谷氨酸（glutamic acid, Glu）、天冬氨酸（aspartic acid, Asp）的出现频率，同时增加N端甘氨酸（glycine, Gly）的占比。本研究分离得到了若干此前在人类细胞系中被鉴定出的、与强直性脊柱炎发病机制潜在相关的肽段。然而，易患关节炎的大鼠其B27肽组与不易感大鼠的B27肽组并无显著差异，且未发现任何仅与关节炎相关的特异性肽段。强直性脊柱炎的发病是否依赖特定的B27结合肽段，仍有待进一步研究阐明。