title: The low binding affinity of ADAMTS4 for citrullinated fibronectin may contribute to the destruction of joint cartilage in rheumatoid arthritis.
abstract: OBJECTIVE
  Rapid cartilage degradation in the joints is observed in rheumatoid arthritis (RA). ADAMTS4 (a disintegrin and metalloproteinase with thrombospondin motifs-4) degrades aggrecan, the primary component of cartilage, therefore contributing to joint erosion in RA. The proteolytic activity of ADAMTS4 is inhibited by fibronectin (FN). FN is abundantly expressed in the synovia in RA and is modified by citrullination, the conversion of peptidylarginine to citrulline. This study aims to investigate the binding ability of citrullinated FN (cFN) to ADAMTS4 and the effect of cFN on aggrecanase activity.
  
  
  METHODS
  The full-length recombinant ADAMTS4 was purified from HEK293 cells that were transiently transfected with a full-length cDNA coding for human ADAMTS4. A 40-kDa FN fragment exhibiting heparin binding was citrullinised with rabbit peptidylarginine deaminase. The binding activity of the full-length recombinant ADAMTS4 to cFN was investigated in an in vitro binding assay. The proteolytic activity of ADAMTS4 after incubation with cFN was determined using an aggrecanase activity kit, in which the ARGSVIL peptide is produced by digestion with aggrecanase.
  
  
  RESULTS
  cFN displayed significantly decreased binding activity with ADAMTS4 compared with FN. The full-length ADAMTS4 produced large amounts of the ARGSVIL peptide, but the amount was markedly decreased in the presence of FN. The production of this peptide approached the normal level when the full-length ADAMTS4 was incubated with cFN.
  
  
  CONCLUSIONS
  FN following citrullination is less effective in inhibiting the proteolytic activity of ADAMTS4. It is known that PADI4, an enzyme active in citrullination, is highly expressed in the synovial tissue in RA. Our results suggest that PADI4 in the RA synovium may contribute to cartilage destruction via the citrullination of FN.
authors.affiliation: Orthopaedic Surgery Center, Shandong University, Shandong, People's Republic of China. yanxinfengra@163.com
authors.forename: Xinfeng
authors.initials: X
authors.lastname: Yan
authors.forename: Luxu
authors.initials: L
authors.lastname: Yin
authors.forename: Yuejian
authors.initials: Y
authors.lastname: Wang
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhao
authors.forename: Xiaotian
authors.initials: X
authors.lastname: Chang
mesh_terms: D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism*; D000071121:ADAMTS4 Protein; D053674:Aggrecans / Q000378:metabolism; D001172:Arthritis, Rheumatoid / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D002358:Cartilage, Articular / Q000201:enzymology* / Q000473:pathology; D002956:Citrulline / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D006867:Hydrolases / Q000378:metabolism; D007596:Joints / Q000201:enzymology* / Q000473:pathology; D011348:Procollagen N-Endopeptidase / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection
chemical_list: D053674:Aggrecans; D005353:Fibronectins; D011994:Recombinant Proteins; D002956:Citrulline; D006867:Hydrolases; D051722:ADAM Proteins; D011348:Procollagen N-Endopeptidase; D000071121:ADAMTS4 Protein; C000603918:ADAMTS4 protein, human; C000629235:PADI4 protein, human; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases