Abstract
A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4 and ADAMTS-5 are the principal aggrecanases in mice and humans; however, mice lacking the catalytic domain of both enzymes (TS-4/5∆cat) have no skeletal phenotype, suggesting there is an alternative aggrecanase for modulating normal growth and development in these mice. We previously identified aggrecanase activity that (a) cleaved at E↓G rather than E↓A bonds in the aggrecan core protein, and (b) was upregulated by retinoic acid but not IL-1α. The present study aimed to identify the alternative aggrecanase. Femoral head cartilage explants from TS-4/5∆cat mice were stimulated with IL-1α or retinoic acid and total RNA was analysed by microarray. In addition to ADAMTS-5 and matrix metalloproteinase (MMP)-13, which are not candidates for the novel aggrecanase, the microarray analyses identified MMP-11, calpain-5 and ADAMTS-9 as candidate aggrecanases upregulated by retinoic acid. When calpain-5 and MMP-11 failed to meet subsequent criteria, ADAMTS-9 emerged as the most likely candidate for the novel aggrecanase. Immunohistochemistry revealed ADAMTS-9 expression throughout the mouse growth plate and strong expression, particularly in the proliferative zone of the TS-4/5-∆cat mice. In conclusion, ADAMTS-9 has a novel specificity for aggrecan, cleaving primarily at E↓G rather than E↓A bonds in mouse cartilage. ADAMTS-9 might have more important roles in normal skeletal development compared with ADAMTS-4 and ADAMTS-5, which have key roles in joint pathology.
Highlights
The weight-bearing properties of articular cartilage are conferred by the large and highly charged glycosaminoglycan side chains that are present on the proteoglycan aggrecan
Retinoic Acid Regulates the Expression of a Novel Aggrecanase Activity via a Transcriptional MechaWniesmpreviously identified a novel aggrecanase activity that cleaves aggrecan in the CS-rich region in cartilage explants from mice deficient in both ADAMTS-4 and ADAMTS-5 [13]
We have used immunoreactivity against the FREEE 1467 neoepitope to show that TS-4/5∆cat mice express aggrecanase activity, and we propose that this activity is likely to be the product of ADAMTS-9 (Figure 5)
Summary
The weight-bearing properties of articular cartilage are conferred by the large and highly charged glycosaminoglycan side chains that are present on the proteoglycan aggrecan. The resilience of cartilage is compromised when large aggrecan aggregates are degraded by aggrecanases that are members of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family of zinc-dependent enzymes within the metzincin family of metalloproteinases. The principal aggrecanases thought to be responsible for aggrecan degradation are ADAMTS-4 [1] and ADAMTS-5 [2]. These enzymes were originally identified by their ability to cleave the aggrecan core protein at the TEGE 373↓374 ALGS bond in the aggrecan interglobular domain (IGD) [3,4,5] and at four additional.
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