The Thrombospondin Motif of Aggrecanase-1 (ADAMTS-4) Is Critical for Aggrecan Substrate Recognition and Cleavage

Tortorella, Micky D.; Pratta, Michael A.; Liu, Rui‐Qin; Abbaszade, Ilgar; Ross, Harold O; Burn, Timothy C.; Arner, Elizabeth C.

doi:10.1074/jbc.m001065200

Cited by 182 publications

(152 citation statements)

References 35 publications

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“…ADAMTS-4 undergoes post-translational processing to at least 4 activation states, each with distinct substrate specificity and matrixbinding properties that could influence inhibitor potency (Gao, et al, 2002;Kashiwagi, et al, 2004;Tortorella, et al, 2000). As a result, the aggrecanase inhibitors used in this study may efficiently block aggrecanase activity in the aggrecan IGD and yet permit less destructive activity (at sites in the CS-2 domain) characteristic of lower activation states.…”

Section: Discussionmentioning

confidence: 99%

Selective and non-selective metalloproteinase inhibitors reduce IL-1-induced cartilage degradation and loss of mechanical properties

et al. 2007

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Articular cartilage undergoes matrix degradation and loss of mechanical properties when stimulated with proinflammatory cytokines such as interleukin-1 (IL-1). Aggrecanases and matrix metalloproteinases (MMPs) are thought to be principal downstream effectors of cytokine-induced matrix catabolism, and aggrecanase-or MMP-selective inhibitors reduce or block matrix destruction in several model systems. The objective of this study was to use metalloproteinase inhibitors to perturb IL-1-induced matrix catabolism in bovine cartilage explants and examine their effects on changes in tissue compression and shear properties. Explanted tissue was stimulated with IL-1 for up to 24 days in the absence or presence of inhibitors which were aggrecanase-selective, MMP-selective, or non-selective. Analysis of conditioned media and explant digests revealed that aggrecanase-mediated aggrecanolysis was delayed to varying extents with all inhibitor treatments, but that aggrecan release persisted. Collagen degradation was abrogated by MMP-and non-selective inhibitors and reduced by the aggrecanase inhibitor. The inhibitors delayed but did not reduce loss of the equilibrium compression modulus, whereas the loss of dynamic compression and shear moduli was delayed and reduced. The data suggest that non-metalloproteinase mechanisms participate in IL-1-induced matrix degradation and loss of tissue material properties.

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Section: Discussionmentioning

confidence: 99%

Selective and non-selective metalloproteinase inhibitors reduce IL-1-induced cartilage degradation and loss of mechanical properties

et al. 2007

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“…However, if ADAMTS4 cleaved at both sites in the bovine incubations, the short intervening AF-28-reactive peptide would have migrated off the gel under the electrophoretic conditions used. In other studies (46), aggrecan fragmentation was monitored by antibody BC-3 and an antibody to the chondroitin sulfate stubs on chondroitinase-treated aggrecan (monoclonal antibody 2035), neither of which would have detected the possible formation of G1-VDIPES. Most recently, a VDIPES-specific antiserum did not detect this product in an incubation of bovine aggrecan with 10 nM ADAMTS4 for 24 h (45).…”

Section: Discussionmentioning

confidence: 99%

ADAMTS4 Cleaves at the Aggrecanase Site (Glu373-Ala374) and Secondarily at the Matrix Metalloproteinase Site (Asn341-Phe342) in the Aggrecan Interglobular Domain

Westling

Fosang

Thompson

et al. 2002

Journal of Biological Chemistry

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Aggrecan is the major cartilage hyalectan (1), which, together with the collagen network, provides this tissue with its unique mechanical properties of compressibility and stiffness (2-4). Extraction of aggrecan in its native form (5) and subsequent structural analysis (6) have revealed that the molecular organization of aggrecan is perfectly suited to its central functional role in articular cartilage. The N-terminal region of aggrecan is composed of two globular domains (G1 1 and G2) separated by the interglobular domain (IGD). G1 interacts with hyaluronan and link protein, thereby keeping the aggrecan molecule anchored within the cartilage tissue. Further interactions with other matrix components such as tenascin-R and fibulin-1 and fibulin-2 (7, 8) may occur through a third globular domain (G3) at the extreme C terminus of the core protein. The extended core protein between G2 and G3 is composed of a short keratan sulfate-rich region followed by a longer chondroitin sulfate-substituted domain. The charge repulsion and hydration of the long negatively charged glycosaminoglycan (GAG) chains are thought to maintain the C-terminal portions of aggrecan in an extended conformation (9). The swelling pressure of the aggrecan-link protein complex with hyaluronan is restrained by the tension in the collagen network; and together, these components form a fiber-reinforced concentrated gel within the cartilage, which transmits forces across the articular joint.In diseases characterized by cartilage degradation such as rheumatoid arthritis and osteoarthritis, increased aggrecan release from the cartilage occurs early (10, 11) and before the bulk of the collagen network is degraded (12). Proteolytic cleavage of aggrecan within the IGD separates the GAG-rich region from the hyaluronan-anchored G1 domain, resulting in GAG release from the cartilage matrix to the synovial fluid. Biomechanical tests on cartilage discs have shown that proteolysis within the IGD of aggrecan, and not cleavages near the C terminus, is primarily responsible for the loss of compressive resistance that accompanies interleukin-1-mediated degradation of the tissue (13). Identification of the proteinases responsible for this "destructive" cleavage of aggrecan has therefore been a major focus of experimentation in arthritis-related research.In this regard, two major cleavage sites that occur in vivo have been identified in the IGD of human aggrecan. One is a matrix metalloproteinase (MMP)-sensitive site at VDIPEN 341 2F 342 FGVGG, which can be cleaved at neutral pH by any one of a range of MMPs, including MMP-1-3, -7-9, -13,

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“…The C-terminal TSP1 domains of ADAMTS1 are reported to bind endothelial cells (29) and to have anti-angiogenic activity, which is attributed to binding and sequestration of vascular endothelial growth factor (30). The TSP1 motif of ADAMTS4 is required for aggrecan cleavage (31), and its Cys-rich and spacer domains bind sulfated glycosaminoglycans (32). ADAMTS12 also employs C-terminal TSP1 domains to bind extracellular matrix (33).…”

Section: Fig 5 Adamts13 Variants Expressed In Cos-7 Cellsmentioning

confidence: 99%

Cleavage of von Willebrand Factor Requires the Spacer Domain of the Metalloprotease ADAMTS13

Zheng

Nishio

Majerus

et al. 2003

Journal of Biological Chemistry

180

236

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ADAMTS13 consists of a reprolysin-type metalloprotease domain followed by a disintegrin domain, a thrombospondin type 1 motif (TSP1), Cys-rich and spacer domains, seven more TSP1 motifs, and two CUB domains. ADAMTS13 limits platelet accumulation in microvascular thrombi by cleaving the Tyr 1605 -Met 1606 bond in von Willebrand factor, and ADAMTS13 deficiency causes a lethal syndrome, thrombotic thrombocytopenic purpura. ADAMTS13 domains required for substrate recognition were localized by the characterization of recombinant deletion mutants. Constructs with C-terminal His 6 and V5 epitopes were expressed by transient transfection of COS-7 cells or in a baculovirus system. No association with extracellular matrix or cell surface was detected for any ADAMTS13 variant by immunofluorescence microscopy or chemical modification. Both plasma and recombinant full-length ADAMTS13 cleaved von Willebrand factor subunits into two fragments of 176 kDa and 140 kDa. Recombinant ADAMTS13 was divalent metal ion-dependent and was inhibited by IgG from a patient with idiopathic thrombotic thrombocytopenic purpura. ADAMTS13 that was truncated after the metalloprotease domain, the disintegrin domain, the first TSP1 repeat, or the Cys-rich domain was not able to cleave von Willebrand factor, whereas addition of the spacer region restored protease activity. Therefore, the spacer region is necessary for normal ADAMTS13 activity toward von Willebrand factor, and the more Cterminal TSP1 and CUB domains are dispensable in vitro.Thrombotic thrombocytopenic purpura (TTP) 1 is a syndrome characterized by microangiopathic hemolytic anemia and thrombocytopenia, and it may be accompanied by neurological dysfunction, renal failure, and fever (1-3). If untreated, the mortality can exceed 90%, but plasma-exchange therapy has reduced the mortality to less than 20% (4). Although the pathophysiology of TTP is not fully understood, a plausible model has been proposed in which the proteolysis of von Willebrand factor (VWF) plays a central role (5). VWF is a multimeric protein that binds receptors on the surface of platelets and in connective tissue, thereby mediating the adhesion of platelets to sites of vascular injury. If unchecked, the process can lead to microvascular thrombosis. A plasma VWF-cleaving protease has been described that acts on the Tyr 1605 -Met 1606 bond in the central A2 domain of the VWF subunit, and cleavage is stimulated by shear forces like those occurring at sites of thrombosis or by low concentrations of urea or guanidine (6, 7). This proteolytic reaction limits VWF-dependent platelet adhesion, and most adults with idiopathic TTP have an acquired autoantibody that inhibits the VWF-cleaving protease (8, 9). Therefore, therapeutic plasma exchange may ameliorate TTP by replacing the missing protease and removing the inhibitory antibody.The VWF-cleaving protease was recently purified and identified as a new member of the ADAMTS family of metalloproteases (10, 11), so named for the combination of a disintegrin-like and metalloprote...

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The Thrombospondin Motif of Aggrecanase-1 (ADAMTS-4) Is Critical for Aggrecan Substrate Recognition and Cleavage

Cited by 182 publications

References 35 publications

Selective and non-selective metalloproteinase inhibitors reduce IL-1-induced cartilage degradation and loss of mechanical properties

Selective and non-selective metalloproteinase inhibitors reduce IL-1-induced cartilage degradation and loss of mechanical properties

ADAMTS4 Cleaves at the Aggrecanase Site (Glu373-Ala374) and Secondarily at the Matrix Metalloproteinase Site (Asn341-Phe342) in the Aggrecan Interglobular Domain

Cleavage of von Willebrand Factor Requires the Spacer Domain of the Metalloprotease ADAMTS13

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