title: Characterization of Autoantigens Targeted by Anti-Citrullinated Protein Antibodies In Vivo: Prominent Role for Epitopes Derived from Histone 4 Proteins.
abstract: Anti-citrullinated protein antibodies (ACPA) have become an integral part of the clinical definition of rheumatoid arthritis, and are hypothesized to be important in the immunopathogenesis of this autoimmune disease. Several citrullinated proteins have been demonstrated to serve as candidate autoantigens for the ACPA, based on in vitro immune reactions between citrullinated peptides/proteins and RA sera. Yet it remains unclear whether the autoantigens identified in vitro are indeed directly and specifically targeted by the ACPA in vivo. Moreover, it is unclear whether ACPA present in RA sera are directed towards the same spectrum of autoantigens as the ACPA present within the synovial compartment. In this study, we isolated ACPA immune complexes from RA synovial fluids (SF) and sera by using immobilized cyclic citrullinated peptides (CCP3) based immune affinity, and characterized the proteins that are directly and specifically associated with them by mass spectrometry. The results demonstrate that four histone proteins are prominent ACPA autoantigens, with the frequency of detection being histone H4 (89%), H2B (63%), H3 (63%), and H2A (58%) in ACPA positive RA SF. We further demonstrate that a histone 4 peptide containing citrulline at position Cit39 was recognized by 100% of ACPA positive RA SF. An adjacent citrulline residue at Cit40 was recognized by 34% of ACPA positive RA SF. An H4 peptide containing Cit39-40 was recognized in the serum of 94% ACPA positive RA, 77% ACPA positive first-degree relatives (FDR) of RA patients, and 2.5% of healthy controls. The Cit39-40 peptide substantially blocked the ACPA reactivity in both SF and serum. Although the spectrum of ACPA we identified was limited to those isolated using immobilized CCP3 peptides, the findings indicate that H4 is a widely recognized RA autoantigen in both the synovial and serum compartments. The identification of this immunodominant ACPA epitope may be valuable in designing approaches to immune tolerance induction in RA.
authors: Arthritis Centre, Department of Internal Medicine, University of Manitoba, RR149, 800 Sherbrook St. Winnipeg, MB, R3A 1M4, Canada.|Manitoba Centre for Proteomics and Systems Biology, University of Manitoba, 799 JBRC, 715 McDermot Ave, Winnipeg, MB, R3E 3P4, Canada.
authors: Xiaobo
authors: X
authors: Meng
authors: Manitoba Centre for Proteomics and Systems Biology, University of Manitoba, 799 JBRC, 715 McDermot Ave, Winnipeg, MB, R3E 3P4, Canada.
authors: Peyman
authors: P
authors: Ezzati
authors: Arthritis Centre, Department of Internal Medicine, University of Manitoba, RR149, 800 Sherbrook St. Winnipeg, MB, R3A 1M4, Canada.
authors: Irene
authors: I
authors: Smolik
authors: Department of Internal Medicine, University of Manitoba, Winnipeg, Manitoba, Canada.
authors: Charles N
authors: CN
authors: Bernstein
authors: Arthritis Centre, Department of Internal Medicine, University of Manitoba, RR149, 800 Sherbrook St. Winnipeg, MB, R3A 1M4, Canada.
authors: Carol Ann
authors: CA
authors: Hitchon
authors: Arthritis Centre, Department of Internal Medicine, University of Manitoba, RR149, 800 Sherbrook St. Winnipeg, MB, R3A 1M4, Canada.|Manitoba Centre for Proteomics and Systems Biology, University of Manitoba, 799 JBRC, 715 McDermot Ave, Winnipeg, MB, R3E 3P4, Canada.
authors: Hani S
authors: HS
authors: El-Gabalawy
mesh_terms: D000595:Amino Acid Sequence; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000276:immunology; D001323:Autoantibodies / Q000097:blood / Q000737:chemistry / Q000276:immunology*; D001324:Autoantigens / Q000276:immunology*; D002956:Citrulline / Q000378:metabolism*; D000939:Epitopes / Q000276:immunology*; D006657:Histones / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D010456:Peptides, Cyclic / Q000276:immunology; D013582:Synovial Fluid / Q000276:immunology
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; D000939:Epitopes; D006657:Histones; D010456:Peptides, Cyclic; C487763:cyclic citrullinated peptide; D002956:Citrulline