ADAMTS-5 deficiency does not block aggrecanolysis at preferred cleavage sites in the chondroitin sulfate-rich region of aggrecan

Abstract

In the mouse, proteolysis in the aggrecan interglobular domain is driven by ADAMTS-5, and mice deficient in ADAMTS-5 catalytic activity are protected against aggrecan loss and cartilage damage in experimental models of arthritis. Here we show that despite ablation of ADAMTS-5 activity, aggrecanolysis can still occur at two preferred sites in the chondroitin sulfate-rich region. Retinoic acid was more effective than interleukin-1α (IL) in promoting cleavage at these sites in ADAMTS-5-deficient cartilage. These results suggest that cleavage at preferred sites in the chondroitin sulfate-rich region is mediated by ADAMTS-4 or an aggrecanase other than ADAMTS-5. Following retinoic acid or IL-1α stimulation of cartilage explants, aggrecan fragments in medium and extracts contained SELE1279 or FREEE1467 C-terminal sequences. Some SELE1279 and FREEE1467 fragments were retained in the cartilage, with intact G1 domains. Other SELE1279 fragments were released into the medium and co-migrated with the 374ALGS neoepitope, indicating they were aggrecanase-derived fragments. In contrast none of the FREEE1467 fragments released into the medium co-migrated with the 374ALGS neoepitope, suggesting that, despite their size, these fragments were not products of aggrecanase cleavage in the interglobular domain. ADAMTS-5, but not ADAMTS-1, -4, or -9, was up-regulated 8-fold by retinoic acid and 17-fold by IL-1α treatment. The data show that whereas ADAMTS-5 is entirely responsible for cleavage in the interglobular domain, cleavage in the chondroitin sulfate-rich region is driven either by ADAMTS-4, which compensates for loss of ADAMTS-5 in this experimental system, or possibly by another aggrecanase. The data show that there are differential aggrecanase activities with preferences for separate regions of the core protein. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc.