Memory T cells specific to citrullinated α-enolase are enriched in the rheumatic joint

J Autoimmun. 2018 Aug;92:47-56. doi: 10.1016/j.jaut.2018.04.004. Epub 2018 May 28.


ACPA-positive rheumatoid arthritis (RA) is associated with distinct HLA-DR alleles and immune responses to many citrullinated self-antigens. Herein we investigated the T cell epitope confined within α-enolase326-340 in the context of HLA-DRB1*04:01 and assessed the corresponding CD4+ T cells in both the circulation and in the rheumatic joint. Comparative crystallographic analyses were performed for the native and citrullinated α-enolase326-340 peptides in complex with HLA-DRB1*04:01. HLA-tetramers assembled with either the native or citrullinated peptide were used for ex vivo and in vitro assessment of α-enolase-specific T cells in peripheral blood, synovial fluid and synovial tissue by flow cytometry. The native and modified peptides take a completely conserved structural conformation within the peptide-binding cleft of HLA-DRB1*04:01. The citrulline residue-327 was located N-terminally, protruding towards TCRs. The frequencies of T cells recognizing native eno326-340 were similar in synovial fluid and peripheral blood, while in contrast, the frequency of T cells recognizing cit-eno326-340 was significantly elevated in synovial fluid compared to peripheral blood (3.6-fold, p = 0.0150). Additionally, citrulline-specific T cells with a memory phenotype were also significantly increased (1.6-fold, p = 0.0052) in synovial fluid compared to peripheral blood. The native T cell epitope confined within α-enolase326-340 does not appear to lead to complete negative selection of cognate CD4+ T cells. In RA patient samples, only T cells recognizing the citrullinated version of α-enolase326-340 were found at elevated frequencies implicating that neo-antigen formation is critical for breach of tolerance.

Keywords: Autoimmunity; Citrullination; Crystal structure; HLA class II tetramers.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Arthritis, Rheumatoid / immunology*
  • CD4-Positive T-Lymphocytes / immunology*
  • Cells, Cultured
  • Citrullination
  • Epitopes, T-Lymphocyte / chemistry
  • Epitopes, T-Lymphocyte / immunology
  • Epitopes, T-Lymphocyte / metabolism*
  • Female
  • Flow Cytometry
  • Humans
  • Immune Tolerance
  • Immunologic Memory
  • Joints / immunology*
  • Male
  • Middle Aged
  • Peptides / chemistry
  • Peptides / immunology
  • Peptides / metabolism*
  • Phosphopyruvate Hydratase / chemistry
  • Phosphopyruvate Hydratase / immunology
  • Phosphopyruvate Hydratase / metabolism*
  • Young Adult


  • Epitopes, T-Lymphocyte
  • Peptides
  • Phosphopyruvate Hydratase