ADAMTS‐4_v1 Is a Splice Variant of ADAMTS‐4 That Is Expressed as a Protein in Human Synovium and Cleaves Aggrecan at the Interglobular Domain. Issue 11 (28th October 2013)

• Record Type: Journal Article
• Authors: Wainwright, Shane D.; Bondeson, Jan; Caterson, Bruce; Hughes, Clare E.
• Is Part Of: Arthritis and rheumatism. Volume 65:Issue 11(2013:Nov.)
• Page Start: 2866
• Page End: 2875
• Publication Date: 2013-10-28
• Subjects: Arthritis -- Periodicals; Rheumatism -- Periodicals; Arthritis -- Periodicals; Rheumatic Diseases -- Periodicals; Rhumatisme -- Périodiques; Arthrite -- Périodiques; 616.72
• DOI: 10.1002/art.38102
• Languages: English
• ISSNs: 0004-3591

Abstract

Abstract : Objective: We previously described a messenger RNA variant of ADAMTS4 ( ADAMTS4_v1 ) in human synovial cell cocultures obtained from patients with osteoarthritis (OA). This RNA message has been found only in OA synovium and, if translated, would result in a protein identical to ADAMTS‐4, except that the C‐terminal spacer domain would be different. The purpose of this study was to determine whether ADAMTS4_v1 is translated into a protein, is expressed in vivo, and acts as a functional aggrecanase. Methods: Polyclonal antibodies were raised against unique C‐terminal sequences of ADAMTS‐4_v1. An immunohistochemical study of human OA synovium was performed. A mammalian expression vector coding for FLAG‐tagged human ADAMTS4 was mutated to contain the different sequences of ADAMTS4_v1, and the resultant plasmid was used to transfect HEK 293 cells. ADAMTS‐4_v1 produced by these cells was purified via the FLAG epitope, and the ability of this recombinant protein to cleave aggrecan, biglycan, and decorin was investigated. Results: An antibody specific for ADAMTS‐4_v1 was found to bind to the synovial membrane surface on cryosections, and the protein was detected in cell lysates from synovium obtained from OA patients. The recombinant ADAMTS‐4_v1 demonstrated enzyme activity toward the target substrate in a commercial aggrecanase 1 enzyme‐linked immunosorbent assay and was also found to cleave aggrecan at the pathologically important Glu 373↓374 Ala aggrecanase site. Conclusion: ADAMTS‐4_v1 is expressed as a protein in vivo in human OA synovium, functions as an aggrecanase, and cleaves other proteoglycan substrates. This splice variant may be a major contributor to loss of aggrecan from the superficial zone of OA cartilage.