The primary structure of a bone-specific sialoprotein was deduced from cloned cDNA. One of the cDNA clones isolated from a rat osteosarcoma (ROS 17/2.8) phage Xgtll library had a 1473-base-pair-long insert that encoded a protein with 317 amino acid residues. This cDNA done appears to represent the complete coding region of sialoprotein mRNA, including a putative AUG initiation codon and a signal peptide sequence. The amino acid sequence deduced from the cDNA contains several Ser-Xaa-Glu sequences, possibly representing attachment points for O-glycosidically linked oligosaccharides and one Asn-Xaa-Ser sequence representing a likely site for the N-glycosidically linked oligosaccharide. An interesting observation is the Gly-Arg-Gly-Asp-Ser sequence, which is identical to the cell-binding sequence identified in fibronectin. The presence of this sequence prompted us to investigate the cell-binding properties of sialoprotein. The ROS 17/2.8 cells attached and attained a spread morphology on surfaces coated with sialoprotein. We could demonstrate that synthetic ArgGly-Asp-containing peptides efficiently inhibited the attachment ofcells to sialoprotein-coated substrates. The results show that the Arg-Gly-Asp sequence also confers cell-binding properties on bone-specific sialoprotein. To better reflect the potential function of bone sialoprotein-we propose the name "osteopontin" for this protein.

A key event in bone resorption is the binding of osteoclasts to the mineral matrix of bone surfaces. A candidate for mediating this binding is osteopontin, a major cell-and hydroxyapatite-binding protein synthesized by osteoblasts. In support of this hypothesis is the fact that the synthesis of osteopontin is stimulated by calcitriol (1,25-dihydroxyvitamin(D3), a substance that induces bone resorption. The present study demonstrates that osteopontin is highly enriched at regions of the bone surface where osteoclasts are anchored. Furthermore, the vitronectin receptor, which has known specificity for osteopontin, is shown preferentially localized at the corresponding area of the osteoclast plasma membrane. The results thus support the hypothesis that osteoclasts when resorbing bone are anchored by osteopontin bound both to the mineral of bone matrix and to a vitronectin receptor on the osteoclast plasma membrane.More than 100 years have passed since Kolliker gave the name "Osteoklast" to a large multinucleated cell observed along bone surfaces and suggested a role for the cell in bone resorption (1). It is now well established that osteoclasts are derived from a bone-marrow cell reaching the bone surfaces by means of blood-borne mononuclear precursor cells (2). The stimulus is provided by other bone cells, which when appropriately stimulated, produce factors that induce recruitment of osteoclast progenitor cells to select bone surfaces (3). Osteoclasts are in contact with mineralized bone matrix at two modified cell surface areas, a clear zone and a ruffled border (4, 5) ( Fig. 1). Active bone resorption is confined to the ruffled border area, where an acidic milieu is maintained. The clear zone is located immediately adjacent to the ruffled border and is thought to provide the osteoclast with a tight attachment to bone, thereby sealing off the ruffled border zone (5, 6). The mechanism for this vital attachment has remained an enigma. It has, however, been suggested that a vitronectin receptor identified on isolated osteoclasts (7) may be involved (8).Over recent years several noncollagenous matrix proteins of bone have been isolated and characterized. At least two of these proteins-i.e., osteopontin and bone sialoprotein-are acidic and bind tightly to hydroxylapatite (9, 10). A functional Arg-Gly-Asp (RGD) cell-binding sequence has been identified in both proteins by cDNA cloning and sequencing (11,12). A receptor for the proteins on cultured osteoblastic cells appears to be the vitronectin receptor (13). Both osteopontin and bone sialoproteins are products of osteoblasts, contain stretches of acidic amino acids, and are phosphorylated. Osteopontin and bone sialoprotein represent distinct gene products. The synthesis of only one of the proteins, osteopontin, is stimulated by calcitriol (14, 15), which is known to induce bone resorption. Thus, osteopontin is a candidate for mediating binding of osteoclasts to bone at the clear zone. In the present study this hypothesis was tested by using ultrastructural ...

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Collagen fibrillogenesis is finely regulated during development of tissue-specific extracellular matrices. The role(s) of a leucine-rich repeat protein subfamily in the regulation of fibrillogenesis during tendon development were defined. Lumican-, fibromodulin-, and double-deficient mice demonstrated disruptions in fibrillogenesis. With development, the amount of lumican decreases to barely detectable levels while fibromodulin increases significantly, and these changing patterns may regulate this process. Electron microscopic analysis demonstrated structural abnormalities in the fibrils and alterations in the progression through different assembly steps. In lumican-deficient tendons, alterations were observed early and the mature tendon was nearly normal. Fibromodulin-deficient tendons were comparable with the lumican-null in early developmental periods and acquired a severe phenotype by maturation. The double-deficient mice had a phenotype that was additive early and comparable with the fibromodulin-deficient mice at maturation. Therefore, lumican and fibromodulin both influence initial assembly of intermediates and the entry into fibril growth, while fibromodulin facilitates the progression through growth steps leading to mature fibrils. The observed increased ratio of fibromodulin to lumican and a competition for the same binding site could mediate these transitions. These studies indicate that lumican and fibromodulin have different developmental stage and leucine-rich repeat protein specific functions in the regulation of fibrillogenesis.