ADAMTS: A novel family of proteases with an ADAM protease domain and thrombospondin 1 repeats

A disintegrin and metalloproteinase (ADAM) is a family of gene products with sequence homology to snake venom metalloproteinases and disintegrins [1,2]. Members of the family share several distinct protein modules, including a metalloprotease domain, a disintegrin domain, a cysteine-rich region and an EGF repeat [1,2]. These proteins, however, perform essential functions in cell adhesion and fusion in diverse systems [3]. The heterodimeric sperm protein fertilin participates in sperm-egg fusion [4], while meltrin-K is involved in myoblast fusion [5]. The Drosophila Kuzbanian (KUZ) plays a role in axonal extension and neural cell fate in £y neurogenesis [6,7]. The protease activity of some members is apparently important in ectodomain shedding of molecules such as the tumor necrosis factor K and the Drosophila signalling molecule Notch [3]. The tumor necrosis factor (TNF)-K converting enzyme (TACE, ADAM17), for example, has a role in processing multiple cell surface proteins as revealed by analysis of knockout mice [8]. The ADAM family members are generally cell surface molecules with a transmembrane domain spanning the plasma membrane. We report here a bunch of proteins with a metalloproteasedisintegrin domain and an additional distinct feature not present in other ADAM proteins, a thrombospondin type 1 (TSP1) motif. The TSP1 motifs are repeats conserved in TSP 1 and 2, which are multifunctional extracellular matrix (ECM) proteins implicated in cell adhesion, motility and growth. Thrombospondin is a major constituent of platelet K-granules. It is also secreted by a wide variety of epithelial and mesenchymal cells and the levels of thrombospondin are correlated with developmental changes in the embryo and the response to injury in the adult [9]. TSP1 motifs can also be found in several proteins in the complement cascade, properdin [10] and f-spondin, a protein involved in neural cell adhesion and neurite outgrowth [11]. A schematic representation of the domain structures of four mammalian gene products with an ADAM type metalloprotease-disintegrin domain and TSP1 repeats are depicted in Fig. 1A. The respective metalloprotease-disintegrin domain and TSP1 repeats in these proteins were revealed by the pfscan program of the ISREC bioinformatics pro¢lescan server (http://www.isrec.isb-sib.ch/software/PFSCAN_form.html). The ¢rst, ADAMTS-1, is a mouse gene cloned by di¡erential display from a cachexigenic tumor cell line [12,13]. The authors coined the name ADAM with thrombospondin motifs (ADAMTS-1) for the gene product, an informative nomenclature which we shall adhere to. Two human cDNA deposited into the database by the Kazusa DNA research institute, KIAA0688 and KIAA0366, are designated ADAMTS-2 and ADAMTS-4 respectively. KIAA0688 shares a rather high homology (44%) with ADAMTS-1, whereas KIAA0366 shares a high homology (52%) with the bovine gene for procollagen I N-proteinase [14,15]. We propose to designate this bovine gene as ADAMTS-3. Fig. 1B is a phylogenetic tree illustrating the phylogenetic distances between representatives of the ADAM family with that of ADAMTS-1^4. The ADAMTS proteins clearly belong to a subfamily. Fig. 2A shows the alignment of the metalloprotease domain of ADAMTS-1^4 with three ADAM proteins, fertilin-K (ADAM1), meltrin-K (ADAM12) and the monocyte surface antigen MS2 (ADAM8). Fig. 2B shows the alignment of the ¢rst TSP1 motif of ADAMTS-1^4 with the three TSP1 repeats found in thrombospondin 1. All the ADAMTS proteins with the exception of ADAMTS-2 have additional TSP1-like repeats (with lesser homology to that found in thrombospondin) at the carboxyl-terminal portion of the molecules. What might the functions of the ADAMTS family members be? The ADAMTS-1 gene was isolated based on its selective expression in colon 26 adenocarcinoma cachexigenic sublines under in vivo tumor bearing states. ADAMTS-1 mRNA could be induced by an in£ammatory cytokine interleukin-1 in vitro and by intravenous administration of lipopolysaccharide in vivo [12]. The expression of ADAMTS-1 is therefore associated with acute in£ammation. Procollagen I N-proteinase cleaves the amino propeptides in the processing of type I and type II procollagens into collagens. The enzyme was ¢rst puri¢ed [14] from calf skin and the bovine gene was subsequently cloned [15]. It was noted by the authors that the mRNA transcript levels in some tissues are disproportionately high relative to the apparent rate of collagen biosynthesis and the proteins may have other roles in development that are independent of their role in procollagen processing [15]. The presence of a metalloprotease and a disintegrin domain and TSP1 repeats on the same protein suggests intriguing possibilities. It should be noted that ADAMTS-1^4 all possess a Zn-protease catalytic site consensus sequence (HEXXH), which suggests an intact catalytic activity [3]. Procollagen I N-proteinase was indeed isolated as an active enzyme [14]. The other ADAMTSs could therefore be responsible for proteolysis of yet unidenti¢ed substrates at the cell surface or the ECM, if they are expressed on the cell surface or are secreted. Both ADAMTS-1 and procollagen I N-proteinase precursors have putative N-terminal signal peptides, and both are secreted when expressed in cultured cells [13,15]. ADAMTS-2 has a hydrophobic N-terminal sequence and a potential signal cleavage site, as predicted by the PSORT II program