A decade ago, bone morphogenetic protein 1 (BMP1) was shown to provide the activity necessary for proteolytic removal of the C-propeptides of procollagens I-III: precursors of the major fibrillar collagens. Subsequent studies have shown BMP1 to be the prototype of a small group of extracellular metalloproteinases that play manifold roles in regulating formation of the extracellular matrix (ECM). Soon after initial cloning of BMP1, genetic studies showed the related Drosophila proteinase Tolloid (TLD) to be necessary for formation of the dorsal-ventral axis in early embryogenesis. It is now clear that the BMP1/TLD-like proteinases, conserved in species ranging from Drosophila to humans, act in dorsal-ventral patterning via activation of transforming growth factor β (TGFβ)-like proteins BMP2, BMP4 (vertebrates) and decapentaplegic (arthropods). More recently, it has become apparent that the BMP1/TLD-like proteinases are key activators of a broader subset of the TGFβ superfamily of proteins, with implications that these proteinases may be key in orchestrating formation of ECM with growth factor activation and BMP signaling in morphogenetic processes.

All transforming growth factor ␤ (TGF-␤) superfamily members are synthesized as precursors with prodomain sequences that are proteolytically removed by subtilisin-like proprotein convertases (SPCs). For most superfamily members, this is believed sufficient for activation. Exceptions are TGF-␤s 1 to 3 and growth differentiation factor 8 (GDF8), also known as myostatin, which form noncovalent, latent complexes with their SPC-cleaved prodomains. Sequence similarities between TGF-␤s 1 to 3, myostatin, and superfamily member GDF11, also known as bone morphogenetic protein 11 (BMP11), prompted us to examine whether GDF11 might be capable of forming a latent complex with its cleaved prodomain. Here we demonstrate that GDF11 forms a noncovalent latent complex with its SPC-cleaved prodomain and that this latent complex is activated via cleavage at a single specific site by members of the developmentally important BMP1/Tolloid family of metalloproteinases. Evidence is provided for a molecular model whereby formation and activation of this complex may play a general role in modulating neural differentiation. In particular, mutant GDF11 prodomains impervious to cleavage by BMP1/Tolloid proteinases are shown to be potent stimulators of neurodifferentiation, with potential for therapeutic applications.

Gliomedin, which has been implicated as a major player in genesis of the nodes of Ranvier, contains two collagenous domains and an olfactomedin-like domain and belongs to the group of type II transmembrane collagens that includes collagens XIII and XVII and ectodysplasin A. One characteristic of this protein family is that constituent proteins can exist in both transmembrane and soluble forms. Recently, gliomedin expressed at the tips of Schwann cell microvilli was found to bind axonal adhesion molecules neurofascin and NrCAM in interactions essential for Na ؉ -channel clustering at the nodes of Ranvier in myelinating peripheral nerves. Interestingly, exogenously added olfactomedin domain was found to have the same effect as intact gliomedin. Here we analyze the tissue form of gliomedin and demonstrate that the molecule not only exists as full-length gliomedin but also as a soluble form shed from the cell surface in a furin-dependent manner. In addition, gliomedin can be further proteolytically processed by bone morphogenetic protein 1/Tolloid-like enzymes, resulting in release of the olfactomedin domain from the collagen domains. Interestingly, the later cleavage induces formation of higher order, insoluble molecular aggregates that may play important roles in Na ؉ -channel clustering.Collagens and other proteins with collagenous domains constitute the most abundant components of the extracellular matrix. They also play essential roles in development and provision of structural integrity to most tissues and organs and can be divided into nine subgroups (1). Type II-oriented transmembrane collagens are one such subgroup. These play roles in cellmatrix interactions both as integral membrane proteins and as shed forms cleaved from cell surfaces by limited proteolysis (2). Transmembrane collagens XIII (3) and XXV (4) and transmembrane protein ectodysplasin A, which is not formally a collagen but which contains collagenous domains (5), are shed by cleavage at the juxtamembrane consensus sequence (K/R)X n (K/R) (n ϭ 0, 2, 4, 6). Such cleavage is by furin-like proprotein convertases of the S8 family of Kex/subtilisin-related serine endopeptidases (6 -8). In contrast, collagen XVII is not directly cleaved by furin-like convertases. Rather, furin-like convertases activate ADAM (a disintegrin and metalloproteinase) family proteinases that cleave collagen XVII (2, 9).The designation colmedin (collagen repeat plus olfactomedin domain) describes a new type of transmembrane collagen (10) containing an ϳ260-amino acid olfactomedin-like domain at the C terminus. The prototype, olfactomedin, was first identified in the extracellular mucus matrix of the olfactory neuroepithelium of Rana catesbeiana (11,12). With the exception of the sea urchin protein amassin (13), all subsequently characterized olfactomedin-like domain-containing proteins are expressed in neural tissues and seem involved in neural function. Noelin-1 participates in generation of neural crest cells (14, 15), photomedins are expressed in retina (16), tiarin is re...

The mammalian bone morphogenetic protein-1 (BMP-1)/Tolloid-related metalloproteinases play key roles in regulating formation of the extracellular matrix (ECM) via biosynthetic processing of various precursor proteins into mature functional enzymes, structural proteins, and proteins involved in initiating the mineralization of hard tissue ECMs. They also have been shown to activate several members of the transforming growth factor-␤ superfamily, and may serve to coordinate such activation with formation of the ECM in morphogenetic events. Osteoglycin (OGN), a small leucine-rich proteoglycan with unclear functions, is found in cornea, bone, and other tissues, and appears to undergo proteolytic processing in vivo. Here we have successfully generated recombinant OGN and have employed it to demonstrate that a pro-form of OGN is processed to varying extents by all four mammalian BMP-1/Tolloid-like proteinases, to generate a 27-kDa species that corresponds to the major form of OGN found in cornea. Moreover, whereas wild-type mouse embryo fibroblasts (MEFs) produce primarily the processed, mature form of OGN, MEFs homozygous null for genes encoding three of the four mammalian BMP-1/Tolloid-related proteinases produce only unprocessed pro-OGN. Thus, all detectable pro-OGN processing activity in MEFs is accounted for by products of these genes. We also demonstrate that both proand mature OGN can regulate type I collagen fibrillogenesis, and that processing of the prodomain by BMP-1 potentiates the ability of OGN to modulate the formation of collagen fibrils.Bone morphogenetic protein-1 (BMP-1) 1 is the prototype of a class of structurally similar metalloproteinases that play various morphogenetic roles in a broad spectrum of species (1, 2).There are four mammalian members of this family: BMP-1, mammalian Tolloid (mTLD), and mammalian Tolloid-like 1 and 2 (mTLL-1 and mTLL-2) (3-5). BMP-1 and mTLD are encoded by alternatively spliced mRNAs of the same gene (3), whereas mTLL-1 and -2 are genetically distinct (4, 5). These proteinases play key roles in regulating formation of mammalian extracellular matrix (ECM), via biosynthetic processing of precursor proteins to form mature, functional matrix components. In the case of collagen fibers, this includes processing of the C-propeptides of procollagens I-III to yield the major fibrous components of ECM (5-9); proteolytic activation of the enzyme lysyl oxidase (10), which is necessary to the formation of covalent cross-links in collagen and elastic fibers (11); and processing of NH 2 -terminal globular domains, or in some cases C-propeptides, of minor fibrillar procollagen V and XI chains (12-14) to yield type V and XI monomers. Such monomers are incorporated into collagen types I and II fibrils, respectively, and appear to control the shapes and diameters of the resultant heterotypic collagen fibrils (15-18). Members of the same small group of proteinases also process precursors for laminin 5 (19, 20) and type VII collagen (21), both of which are involved in securing epithelia...

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