Identification of citrullinated α-enolase as a candidate autoantigen in rheumatoid arthritis

Andrew Kinloch,David Moyes,David Peston,Robin Wait,Patrick J Venables,Peter C Taylor,Verena Tatzer,Phillipe Donatien,Karin Lundberg

doi:10.1186/ar1845

Abstract

Antibodies against citrullinated proteins are highly specific for rheumatoid arthritis (RA), but little is understood about their citrullinated target antigens. We have detected a candidate citrullinated protein by immunoblotting lysates of monocytic and granulocytic HL-60 cells treated with peptidylarginine deiminase. In an initial screen of serum samples from four patients with RA and one control, a protein of molecular mass 47 kDa from monocytic HL-60s reacted with sera from the patients, but not with the serum from the control. Only the citrullinated form of the protein was recognised. The antigen was identified by tandem mass spectrometry as α-enolase, and the positions of nine citrulline residues in the sequence were determined. Serum samples from 52 patients with RA and 40 healthy controls were tested for presence of antibodies against citrullinated and non-citrullinated α-enolase by immunoblotting of the purified antigens. Twenty-four sera from patients with RA (46%) reacted with citrullinated α-enolase, of which seven (13%) also recognised the non-citrullinated protein. Six samples from the controls (15%) reacted with both forms. α-Enolase was detected in the RA joint, where it co-localised with citrullinated proteins. The presence of antibody together with expression of antigen within the joint implicates citrullinated α-enolase as a candidate autoantigen that could drive the chronic inflammatory response in RA.

Highlights

Rheumatoid arthritis (RA) is a common and disabling disease affecting about 1% of the population [1]
It was subsequently reported that binding of anti-filaggrin antibody epitopes is dependent on the presence of citrulline, an amino acid derived from arginine as a result of a post-translational modification catalysed by the enzyme peptidylarginine deiminase (PAD) [6,7]
Reactivity at 47 kDa was strongest with HL-60 cells that had been differentiated to monocytes, a similar polypeptide was seen in lysates from cells with the granulocyte phenotype

Summary

Introduction

Rheumatoid arthritis (RA) is a common and disabling disease affecting about 1% of the population [1]. Because rheumatoid factors are present in up to 75% of patients with RA, it has been suggested that immunoglobulin G is the antigen. Rheumatoid factors are present in patients with other diseases and in up to 5% of healthy individuals [2]. Other antibodies are present in sera from patients with RA, including antiperinuclear factor [3] and antikeratin antibody [4]. Because both antiperinuclear factor and antikeratin antibody react with human filaggrin and related proteins [5] they were collectively designated 'antifilaggrin antibodies'. It was subsequently reported that binding of anti-filaggrin antibody epitopes is dependent on the presence of citrulline, an amino acid derived from arginine as a result of a post-translational modification catalysed by the enzyme peptidylarginine deiminase (PAD) [6,7]

Methods

Results

Conclusion