Expression and activity of citrullinating peptidylarginine deiminase enzymes in monocytes and macrophages

Abstract

Background: Antibodies directed to proteins containing the non-standard amino acid citrulline, are extremely specific for rheumatoid arthritis (RA). Peptidylcitrulline can be generated by post-translational conversion of arginine residues. This process, citrullination, is catalysed by a group of calcium dependent peptidylarginine deiminase (PAD) enzymes. Objective: To investigate the expression and activity of four isotypes of PAD in peripheral blood and synovial fluid cells of patients with RA. Results: The data presented here show that citrullination of proteins by PAD enzymes is a process regulated at three levels: transcription-in peripheral blood PAD2 and PAD4 mRNAs are expressed predominantly in monocytes; PAD4 mRNA is not detectable in macrophages, translation-translation of PAD2 mRNA is subject to differentiation stage-specific regulation by its 39 UTR, and activation-the PAD proteins are only activated when sufficient Ca 2+ is available. Such high Ca 2+ concentrations are normally not present in living cells. In macrophages, which are abundant in the inflamed RA synovium, vimentin is specifically citrullinated after Ca 2+ influx. Conclusion: PAD2 and PAD4 are the most likely candidate PAD isotypes for the citrullination of synovial proteins in RA. Our results indicate that citrullinated vimentin is a candidate autoantigen in RA.

Synovial intracellular citrullinated proteins colocalizing with peptidyl arginine deiminase as pathophysiologically relevant antigenic determinants of rheumatoid arthritis–specific humoral autoimmunity

“…Of interest, the presence of not only PAD-2, but also PAD-4, was previously demonstrated in human synovial tissue, and both subtypes were also found in SF mononuclear cells (29,36). However, the association between functional haplotypes of PADI4 and RA remains controversial (40,(51)(52)(53).…”

“…However, it should be taken into account that not the mere presence, but also the activation, of PAD-2 is pathophysiologically relevant, as illustrated by the fact that not all PAD-2-positive cells contained citrullinated proteins in RA synovium and that PAD-2 was also found in some control samples. Studies are being conducted to assess whether this is related to programmed cell death, as demonstrated in vitro (25,28,29), or to other mechanisms leading to the increase of the intracellular calcium levels needed for the activation of the citrullinating PADI enzymes in human RA synovium.…”

“…Since the recent observations might indicate a pathophysiologic role in the induction of ACPAs in vivo, in this study we attempted to confirm the RA specificity of synovial intracellular citrullinated proteins, with special reference to the presence of the citrullinating enzyme peptidyl arginine deiminase (PAD) (28,29). Considering the presence of several deiminated proteins in inflamed synovium, we also sought direct evidence that these RA-specific synovial intracellular citrullinated proteins are genuine antigenic targets of the humoral autoimmune process by analyzing their influence on local and systemic ACPA levels.…”

Deiminated Epstein‐Barr virus nuclear antigen 1 is a target of anti–citrullinated protein antibodies in rheumatoid arthritis

“…In the presence of a higher EBV load, events such as apoptosis, infection, damage, or any stimulus involving calcium influx may activate PAD, which is highly expressed in RA synovia (48,49), and induce the deimination of different proteins, including viral proteins. As a result, EBNA-1 may contain posttranslationally acquired deiminated sequences.…”

Absence of citrulline‐specific autoantibodies in animal models of autoimmunity

“…In most cases autoimmune patients also have generated autoantibodies to the unmodified antigen as a result of epitope spreading. In the case of citrullination, which is known to occur during terminal differentiation or cell death (15), the antibodies are directed only to the modification. This raises the question of what mechanisms are underlying this specificity.…”