Drs. Caterson and Hughes receive licensing fees from Eli Lilly for monoclonal antibody (mAb) BC-3 and from Millipore and Thermo Fisher for the sale of mAb BC-3; they also receive royalties from Abcam, Novus, Millipore, and Thermo Fisher for the sale of mAb BC-3.
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
Article first published online: 28 OCT 2013
© The Authors. Arthritis & Rheumatism is published by Wiley Periodicals, Inc. on behalf of the American College of Rheumatology.
This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
Arthritis & Rheumatism
Volume 65, Issue 11, pages 2866–2875, November 2013
How to Cite
Wainwright, S. D., Bondeson, J., Caterson, B. and Hughes, C. E. (2013), 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. Arthritis & Rheumatism, 65: 2866–2875. doi: 10.1002/art.38102
- Issue published online: 28 OCT 2013
- Article first published online: 28 OCT 2013
- Accepted manuscript online: 29 JUL 2013 09:26AM EST
- Manuscript Accepted: 18 JUL 2013
- Manuscript Received: 21 JUN 2012
- Arthritis Research UK. Grant Number: 18893
- MRC. Grant Number: G0800248
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.
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.
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 Glu373374Ala aggrecanase site.
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.