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Expression and activity of citrullinating peptidylarginine deiminase enzymes in monocytes and macrophages
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  1. E R Vossenaar1,
  2. T R D Radstake2,
  3. A van der Heijden1,
  4. M A M van Mansum1,
  5. C Dieteren1,
  6. D-J de Rooij3,
  7. P Barrera2,
  8. A J W Zendman1,
  9. W J van Venrooij1
  1. 1Department of Biochemistry, University of Nijmegen, Nijmegen, The Netherlands
  2. 2Department of Rheumatology, University Medical Centre, St Radboud, Nijmegen, The Netherlands
  3. 3Department of Rheumatology, Sint Maartenskliniek, Nijmegen, The Netherlands
  1. Correspondence to:
    Dr E R Vossenaar
    161 Department of Biochemistry, PO Box 9101, 6500 HB Nijmegen, The Netherlands; e.vossenaarncmls.kun.nl

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 3′ UTR, and activation—the PAD proteins are only activated when sufficient Ca2+ is available. Such high Ca2+ concentrations are normally not present in living cells. In macrophages, which are abundant in the inflamed RA synovium, vimentin is specifically citrullinated after Ca2+ 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.

  • rheumatoid arthritis
  • peptidylarginine deiminase
  • citrullination
  • macrophages
  • vimentin
  • ACR, American College of Rheumatology
  • anti-MC, anti-modified citrulline antibodies
  • CCP, cyclic citrullinated peptide
  • HUGO, human genome organisation
  • NP-40, Nonidet P40
  • PAD, peptidylarginine deiminase
  • PB, peripheral blood
  • PBMCs, peripheral blood mononuclear cells
  • PMCA, plasma membrane Ca2+ pump
  • RA, rheumatoid arthritis
  • RT-PCR, reverse transcriptase-polymerase chain reaction
  • SDS-PAGE, sodium dodecyl sulphate-polyacrylamide gel electrophoresis
  • SF, synovial fluid
  • SFMCs, synovial fluid mononuclear cells
  • UTR, untranslated terminal region

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