ADAMTS4_v1 is a splice variant of ADAMTS4 that is expressed as a protein in human synovium and cleaves aggrecan at the interglobular domain

Wainwright, Shane Daniel, Bondeson, Jan, Caterson, Bruce ORCID: https://orcid.org/0000-0001-6016-0661 and Hughes, Clare Elizabeth ORCID: https://orcid.org/0000-0003-4726-5877 2013. ADAMTS4_v1 is a splice variant of ADAMTS4 that is expressed as a protein in human synovium and cleaves aggrecan at the interglobular domain. Arthritis & Rheumatism 65 (11) , pp. 2866-2875. 10.1002/art.38102

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Abstract

Objective. We have previously reported a mRNA variant of ADAMTS4 (ADAMTS4_v1) in human synovial cell co-cultures obtained from osteoarthritic patients. This RNA message has only been found in OA synovium and if translated would result in a protein identical to ADAMTS4 except that the C-terminal spacer domain would be different. In this study, we set out to determine if ADAMTS4_v1 is translated into a protein, expressed in vivo and acts as a functional aggrecanase. Methods. Polyclonal antibodies were raised against unique C-terminus sequences of ADAMTS4_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 HEK293 cells. ADAMTS4_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 to ADAMTS4_v1 was found to bind to the synovial membrane surface on cryosections and the protein was detected in synovial cell lysates from human OA patient synovium. The recombinant ADAMTS4_v1 demonstrated enzyme activity towards the target substrate in a commercial aggrecanase-1 ELISA assay and was also found to cleave aggrecan at the pathologically important Glu373↓374Ala aggrecanase site. Conclusion. ADAMTS4_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 superficial zone loss of aggrecan in OA cartilage.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Biosciences
Subjects:	Q Science > QH Natural history > QH301 Biology Q Science > QP Physiology
Additional Information:	Online publication date: 29 July 2013.
Publisher:	John Wiley & Sons
ISSN:	0004-3591
Date of First Compliant Deposit:	30 March 2016
Last Modified:	10 Apr 2024 06:55
URI:	https://orca.cardiff.ac.uk/id/eprint/49639

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