Abstract
IntroductionBRAF (v raf murine sarcoma viral oncogene homologue B1) is a serine-threonine kinase involved in the mitogen-activated protein kinase (MAPK) signalling pathway, known to be implicated in the production of pro-inflammatory cytokines.We have observed that sera from rheumatoid arthritis (RA) patients recognize the BRAF's catalytic domain, which encompasses amino acids 416 to 766. Here, we identify peptide targets of anti-BRAF autoantibodies and test whether anti-BRAF autoantibodies may interfere with BRAF kinase activity.MethodsAnti-BRAF autoantibodies were detected by ELISA (enzyme-linked immunosorbent assay) in the serum of RA patients and controls, using 40 overlapping 20mer peptides encompassing the catalytic domain of BRAF as immunosorbents. To test whether autoantibodies to BRAF influence BRAF kinase activity, we developed an in vitro phosphorylation assay of MEK1 (mitogen extracellular regulated kinase), a major BRAF substrate. MEK1 phosphorylation by BRAF was tested in the presence of purified anti-BRAF autoantibodies from RA patients or control antibody.ResultsWe found that one BRAF peptide, P25 (656 to 675), is specifically recognized by autoantibodies from RA patients. Of interest, anti-P25 autoantibodies are detected in 21% of anti-CCP (cyclic citrullinated peptides) negative RA patients. Anti-BRAF autoantibodies activate the in vitro phosphorylation of MEK1 mediated by BRAF.ConclusionsAnti-BRAF autoantibodies from RA patients preferentially recognize one BRAF peptide: P25. Autoantibody responses to P25 are detected in 21% of anti-CCP negative RA patients. Most anti-BRAF autoantibodies activate BRAF kinase activity.
Highlights
BRAF (v raf murine sarcoma viral oncogene homologue B1) is a serine-threonine kinase involved in the mitogen-activated protein kinase (MAPK) signalling pathway, known to be implicated in the production of proinflammatory cytokines
Autoantibody responses to P25 are detected in 21% of anti-cyclic citrullinated peptides (CCP) negative rheumatoid arthritis (RA) patients
We found that one BRAF peptide, P25 (656 to 675), is recognized by autoantibodies from RA patients
Summary
BRAF (v raf murine sarcoma viral oncogene homologue B1) is a serine-threonine kinase involved in the mitogen-activated protein kinase (MAPK) signalling pathway, known to be implicated in the production of proinflammatory cytokines. We have observed that sera from rheumatoid arthritis (RA) patients recognize the BRAF’s catalytic domain, which encompasses amino acids 416 to 766. We identify peptide targets of anti-BRAF autoantibodies and test whether anti-BRAF autoantibodies may interfere with BRAF kinase activity. Rheumatoid arthritis (RA) is a chronic inflammatory joint disease with a prevalence of 0.5% worldwide [1]. The sera of RA patients contain many autoantibodies. The most characteristic are directed at citrullinated proteins (ACPA) [2]. ACPA recognize citrulline (a posttranslationally modified form of arginin) containing epitopes on various proteins, such as filaggrin, vimentin, and fibrinogen [3,4,5,6]. ACPAs can be detected by commercially available enzyme-linked immunoabsorbent assays using synthetic cyclic citrullinated peptides (CCP).
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