Characterization of Human DSPG3, a Small Dermatan Sulfate Proteoglycan

PG-Lb is a small dermatan sulfate proteoglycan that has been previously characterized in chicken. In the developing limb, chick PG-Lb appears to be exclusively expressed in the zone of flattened chondrocytes. We have cloned and sequenced the human homolog to chick PG-Lb from two human chondrocyte cDNA libraries and a human chondrocyte RNA sample. The human homolog has been named DSPG3, as it is the third member of the small dermatan sulfate proteoglycan family to be identified and characterized along with biglycan (PG-I) and decorin (PG-II). DSPG3 maps to chromosome 12q21 and is composed of 1515 nucleotides of cDNA that code for a 322-amino-acid protein. The protein contains three potential glycosaminoglycan attachment sites, two N-glycosylation sites, a poly- glutamic acid stretch, and six cysteines. By Northern analysis, we have demonstrated that DSPG3 is expressed in cartilage, as well as ligament and placental tissues.     

cDNA clone to chick corneal chondroitin/dermatan sulfate proteoglycan reveals identity to decorin.

A 1.6-kb cDNA clone was isolated by screening a library prepared from chick corneal mRNA with a cDNA clone to bovine decorin. The cDNA contained an open reading frame coding for a M(r) 39,683 protein. A 19-amino-acid match with sequence from the N-terminus of core protein from the corneal chondroitin/dermatan sulfate proteoglycan confirmed the clone as a corneal proteoglycan and the homology with human and bovine decorin confirmed its identity as decorin. Structural features of the deduced sequence include a 16-amino-acid signal peptide, a 14-amino-acid propeptide, cysteine residues at the N- and C-terminal regions, and a central leucine-rich region (comprising 63% of the protein) containing nine repeats of the sequence LXXLXLXXNXL/I. Chick decorin contains three variations of this sequence that are tandemly linked to form a unit and three units tandemly linked to form the leucine-rich region. The presence of beta bend amino acids flanking the units may serve to delineate the units as structural elements of the leucine-rich region. Sequence homology within the repeats and the spacing of the repeats suggest that this region arose by duplication. Chick decorin primarily differs from mammalian decorins in the 19-amino-acid sequence that starts the N-terminus of the core protein. Within this region, the serine that serves as a potential acceptor for the chondroitin/dermatan sulfate side chain is preceded by a glycine instead of being followed by a glycine as it is in the mammalian decorins and all other mammalian proteoglycans.     

Comparisons of antibody reactivity and enzyme sensitivity between small proteoglycans from bovine tendon, bone, and cartilage

Preparations of small proteoglycans from bovine tendon, bone, and cartilage have been compared for sensitivity to various enzymes and reactivity with different polyclonal antibodies. Chondroitinase ABC digestion of all proteoglycans generated a core protein preparation that migrated similarly in sodium dodecyl sulfate-polyacrylamide electrophoresis as a doublet band with Mr approximately equal to 45,000. The small proteoglycans of cartilage were divided into two populations based upon electrophoretic migration of the intact molecules (Rosenberg, L. C., Choi, H. U., Tank, L-H., Johnson, T. L., Pal, S., Webber, C., Reiner, A., and Poole, A. R. (1985) J. Biol. Chem. 260, 6304-6313). The core preparations of tendon, bone, and the faster-migrating (PG II) proteoglycans of cartilage all interacted in Western blot/enzyme-linked immunosorbent assay analysis with polyclonal antibody raised against either the tendon or bone proteoglycans. The slower-migrating (PG I) proteoglycans of cartilage did not react with these antibodies. Digestion of the tendon small proteoglycan with Staphylococcus aureus V8 protease released glycosaminoglycan chains from the molecule and generated a 40-kDa protein fragment that was resistant to further rapid degradation by the enzyme. This large digestion fragment was also prominent following V8 protease digestion of the faster-migrating (PG II) population of small cartilage proteoglycans, but not the small proteoglycan of bone. The N-terminal amino acid sequence of the tendon (PG II) proteoglycan was determined. These observations provide additional evidence for heterogeneity among the chemically similar small proteoglycans from different tissues.     

Modulation of human fetal hepatocyte survival and differentiation by interactions with a rat liver epithelial cell line

Our recent studies have shown that chick embryo epiphyseal cartilage synthesizes three distinct species of proteoglycan (PG-H, PG-Lb, and PG-Lt) which are analogous in having glycosaminoglycan side chains of the chondroitin (dermatan) sulfate type but different from one another in regard to the structure of core protein. In the present report, the expression of PG-H and PG-Lb has been studied in developing chick hind limbs (stages 19-33), using antibodies specific for these substances in indirect immunofluorescence. At the onset of cartilage morphogenesis (stage 24), PG-H became recognizable in the cartilage primordia, whereas a parallel section stained for PG-Lb showed no reaction. The first evidence of PG-Lb appearance was seen in a stage 28 cartilage (e.g., tibia) in which the cells in the middiaphysis became elongated in a direction perpendicular to the long axis of the cartilage. The PG-Lb fluorescence was confined to the zone of these flattened, disc-like cells, whereas the fluorescence for PG-H was uniformly distributed throughout the cartilage. With further development of cartilage (stage 29 approximately), the zone of flattened cells spread proximally and distally, and simultaneously large hypertrophied cells appeared at the diaphyseal region. During these zonal changes of cell morphology, the PG-Lb fluorescence remained restricted to the zone of flattened cells. Parallel sections stained for PG-H, in contrast, showed an evenly distributed pattern of the PG-H fluorescence throughout the cartilage. The results indicate that the appearance of PG-Lb is closely associated with the zonal changes of cell shape and orientation along the proximal-distal axis of the developing limb cartilage, and further suggest that the flattened chondrocytes in this particular zone have undergone additional changes in gene expression to form an extracellular matrix of still another chemical property.     

cDNA to chick lumican (corneal keratan sulfate proteoglycan) reveals homology to the small interstitial proteoglycan gene family and expression in muscle and intestine.

A 1.9-kb cDNA clone to chick lumican (keratan sulfate proteoglycan) was isolated by screening an expressing vector library made from chick corneal RNA with antiserum to chick corneal lumican. The cDNA clone contained an open reading frame coding for a 343-amino acid protein, Mr = 38,640. Structural features of the deduced sequence include: a 18-amino acid signal peptide, cysteine residues at the N- and C-terminal regions, and a central leucine-rich region (comprising 62% of the protein) containing nine repeats of the sequence LXXLXLXXNXL/I, where X represents any amino acid. Lumican contains three variations of this sequence that are tandemly linked to form a unit and three units tandemly linked to form the leucine-rich region. The sequential arrangement of these repeats and their spacing suggest that this region arose by duplication. The deduced sequence shows five potential N-linked glycosylation sites, four of which are in the leucine-rich region. These sites are also potential keratan sulfate attachment sites. The cDNA clone to lumican hybridizes to a 2.0-kb mRNA found in tissues other than cornea, predominantly muscle and intestine. Radiolabeling and immunoprecipitation studies show that lumican core protein is also synthesized by these tissues. The primary structure of lumican is similar to fibromodulin, decorin, and biglycan, which indicates it belongs to the small interstitial proteoglycan gene family. The expression of lumican in tissues other than cornea indicates a broader role for lumican besides contributing to corneal transparency.     

The primary structure of the core protein of the small, leucine-rich proteoglycan (PG I) from bovine articular cartilage

Two forms of small, interstitial proteoglycans have been isolated from bovine articular cartilage and have different core proteins, based on NH2-terminal analysis and peptide mapping (Choi, H. U., Johnson, T. L., Pal, S., Tang, L-H., Rosenberg, L. C., and Neame, P. J. (1989) J. Biol. Chem. 264, 2876-2884). These proteoglycans have been called PG I and PG II. Since they were first described, they have also been called "biglycan" (PG I), "decorin," and "DS-PG" (PG II). This report describes the primary structure of PG I from bovine articular cartilage. The protein core consists of 331 amino acids with a molecular mass of 37,280 Da. The amino acid sequence shows 55% identity to the cDNA-derived sequence of PG II from bovine bone. There are four discrete domains in the amino acid sequence. Domain 1, at the NH2 terminus (approximately 23 amino acids), contains two sites of attachment of dermatan sulfate, both of which match the consensus sequence of Asp/Glu-X-X-Ser-Gly-hydrophobic. Neither of these sites is substituted to 100% with glycosaminoglycan in native PG I. Domain 2, near the NH2 terminus and containing approximately 28 amino acids, has a cysteine pattern similar to a domain near the COOH terminus of mouse metallothionein and contains at least one disulfide bond (between the first and fourth cysteine residues). The majority of the core protein of PG I (domain 3) is a leucine-rich domain containing ten repeating units (approximately 231 amino acids). Patthy [1987) J. Mol. Biol. 198, 567-577) has shown that for PG II, the majority of domain 3 shows considerable similarity to leucine-rich alpha 2-glycoprotein (LRG) from serum. Domain 2 of PG I or PG II also has an analog in LRG, in that it has two cysteines in a similar place. The major motif in the PG I described here, in PG II and in LRG, is a series of leucine-rich repeats. PG I and PG II both contain 10 leucine-rich repeats which are 14 amino acids long and which are somewhat irregularly spaced, while LRG contains 9 leucine-rich repeats spaced 10 amino acids apart. Other proteins which contain leucine repeats are the platelet glycoprotein Ib, which is involved in platelet adherence to subendothelium (eight repeats in the alpha chain and two in the beta chain), the protein encoded by the Toll gene (involved in lateral and ventral spatial organization in Drosophila) and chaoptin (a protein involved in Drosophila photoreceptor morphogenesis).(ABSTRACT TRUNCATED AT 400 WORDS)     

Molecular cloning and sequence analysis of the cDNA for small proteoglycan II of bovine bone.

The cDNA for the full-length core protein of the small chondroitin sulphate proteoglycan II of bovine bone was cloned and sequenced. A 1.3 kb clone (lambda Pg28) was identified by plaque hybridization with a previously isolated 1.0 kb proteoglycan cDNA clone (lambda Pg20), positively identified previously by polyclonal and monoclonal antibody reactivity and by hybrid-selected translation in vitro [Day, Ramis, Fisher, Gehron Robey, Termine & Young (1986) Nucleic Acids Res. 14, 9861-9876]. The cDNA sequences of both clones were identical in areas of overlap. The 360-amino-acid-residue protein contains a 30-residue propeptide of which the first 15 residues are highly hydrophobic. The mature protein consists of 330 amino acid residues corresponding to an Mr of 36,383. The core protein contains three potential glycosaminoglycan-attachment sites (Ser-Gly), only one of which is within a ten-amino-acid-residue homologous sequence seen at the known attachment sites of related small proteoglycans. Comparisons of the published 24-residue N-terminal protein sequence of bovine skin proteoglycan II core protein with the corresponding region in the deduced sequence of the bovine core protein reveals complete homology. Comparison of the cDNA-derived sequences of bovine bone and human embryonic fibroblast proteoglycans shows a hypervariable region near the N-terminus. Nucleotide homology between bone and fibroblast core proteins was 87% and amino acid homology was 90%.     

Shared and distinct structural features of interstitial proteoglycans from different bovine tissues revealed by electron microscopy

Large and small interstitial proteoglycans were purified from different bovine tissues, i.e. cartilage, sclera, tendon, aorta, cornea, and bone. The structure of the molecules was compared using the glycerol spraying/rotary shadowing technique for electron microscopy. Large proteoglycans from sclera and tendon have a core protein with a domain structure similar to that previously reported for cartilage proteoglycans (Paulsson, M., M?rgelin, M., Wiedemann, H., Beardmore-Gray, M., Dunham, D., Hardingham, T., Heineg?rd, D., Timpl, R., and Engel, J. (1987) Biochem. J. 245, 763-772). It is comprised of a pair of globules at one end of the molecule, connected by a short extended segment, followed by a long extended domain which is terminated by a third globular domain. Large aorta proteoglycans show a somewhat different structure, with only one globular domain at each end of a long extended segment. Large sclera and aorta proteoglycans form aggregates with hyaluronate and cartilage link protein in a manner similar to that of large cartilage proteoglycans. The large proteoglycans show considerable tissue variability with regard to number, length, and spacing of glycosaminoglycan side chains. The small proteoglycans reveal a small globular core protein to which one or two glycosaminoglycans are attached. Although the main structural features do not differ, proteoglycans of the S1 class have an average glycosylation close to two glycosaminoglycans/molecule, while that of the S2 class is close to one. Differences in glycosaminoglycan length were observed between tissues and between the S1 and S2 class of proteoglycan derived from a single tissue.     

Immunological evidence for two distinct chondroitin sulfate proteoglycan core proteins: Differential expression in cartilage matrix deficient mice

The expression and core protein structure of two proteoglycans, the major cartilage proteoglycan isolated from a rat chondrosarcoma and a small molecular weight chondroitin sulfate proteoglycan isolated from a rat yolk sac tumor, have been compared. The cartilage proteoglycan was not detectable in the cartilage tissue of cartilage matrix deficient ($\text{cmd}\text{cmd}$) neonatal mice by immunofluorescence, but the cmd cartilage did react with antibodies against the core protein of the yolk sac tumor proteoglycan. Radioimmunoassays showed that the core proteins of these proteoglycans are not cross-reactive with each other. Analysis of the core proteins by sodium dodecyl sulfate/polyacrylamide gel electrophoresis after chondroitinase ABC treatment of the proteoglycan revealed a large difference in their sizes. The cartilage proteoglycan core protein had a molecular weight of about 200,000 while the yolk sac tumor proteoglycan core protein migrated with an apparent molecular weight of about 20,000. In addition, the cultured yolk sac tumor cells that make the small proteoglycan did not react with antiserum against the cartilage proteoglycan. These results indicate that the proteoglycan isolated from the yolk sac tumor is similar to the small chondroitin sulfate proteoglycan species found in cartilage and support the existence of at least two dissimilar and genetically independent chondroitin sulfate proteoglycan core proteins.     

Electron microscopic characterization of chick embryonic skeletal muscle proteoglycans

In this article, proteoglycans from embryonic chick leg muscle are quantitatively and qualitatively compared with day 8 high density cell culture cartilage proteoglycans by electron microscopy of proteoglycan-cytochrome c monolayers. The visualized proteoglycan profiles were separated into four categories according to shape, size, and complexity. The two major categories were further characterized by lengths of core proteins, lengths of side projections, and distance between side projections. Two large proteoglycans are identifiable in spread leg muscle preparations. One group has a core protein (mean length of 205 nm) from which extend long thin side projections that we interpret to be groups of chondroitin sulfate glycosaminoglycans with a mean length of 79 nm. This large chondroitin sulfate proteoglycan is the only type found in muscle cultures as determined both biochemically in the past and now by electron microscopy and is referred to as muscle proteoglycan. The second large proteoglycan has a mean core protein length of 250 nm and side projections that are visibly shorter (mean length of 38 nm) and thicker than those of the muscle proteoglycan. This group is referred to as the mesenchymal proteoglycan since its biosynthetic origin is still uncertain. We compare these two profiles with the chick cartilage chondroitin sulfate proteoglycan that has a mean core protein length of 202 nm and side projections with a mean length of 50 nm. The data presented here substantiate the earlier biochemical characterization of these noncartilage proteoglycans and establish the unique structural features of the muscle proteoglycan as compared with the similar profiles of the cartilage and mesenchymal proteoglycans.     

Localization of human platelet proteoglycan gene to chromosome 10, band q22.1, by in situ hybridization

Summary A cDNA probe of 527 base pairs coding for the human platelet proteoglycan (PPG) protein core demonstrated that the PPG gene lies on the long arm of chromosome 10, band q22.1. This result and other available data concerning proteoglycans containing serine-glycine repeats indicate that this gene is involved in the expression of a proteoglycan in various blood cell types.     

Identification of the keratan sulfate attachment sites on bovine fibromodulin

The small keratan sulfate-substituted proteoglycan (fibromodulin) from articular cartilage was shown to contain keratan sulfate linked to the core protein through N-glycosidic linkages to residues Asn-109, Asn-147, Asn-182, and Asn-272. Biosynthetic experiments with articular chondrocytes in the presence of tunicamycin, an inhibitor of N-linked oligosaccharide synthesis, demonstrated a specific inhibition of [35S]SO4 incorporation into fibromodulin. Under the same conditions no effect on the addition of keratan sulfate to the large aggregating proteoglycan was detected. Fibromodulin substituted with keratan sulfate was purified from bovine articular cartilage extracts by density gradient centrifugation, ion-exchange chromatography, and gel-permeation chromatography. Isolation of glycosylated peptides from tryptic digests of fibromodulin by ion-exchange chromatography and reversed-phase high performance liquid chromatography revealed four separate hexosamine-rich species, that were also immunoreactive with monoclonal antibody 5D4. Sequence analysis of these glycopeptides gave blank cycles at positions which corresponded to Asn followed by X-Ser/Thr in the sequence derived from cDNA (Oldberg, A., Antonsson, P., Lindblom, K., and Heinegard, D. (1989) EMBO J. 8, 2601-2604). Hence, all four Asn residues in the leucine-rich region of the fibromodulin core protein can serve as acceptor sites for keratan sulfate addition.     

Proteoglycans of developing bone

We purified and characterized the bone proteoglycans from fetal calves, growing rats, and human fetuses. The major proteoglycan is part of the mineralized tissue matrix and only 10-20% can be extracted prior to demineralization. This bone proteoglycan is a small glycoconjugate (Mr = 80,000-120,000) containing approximately 20-30% protein and either one or two chondroitin sulfate chains (Mr = 40,000) attached to a relatively monodisperse protein core (Mr = 38,000). "O"-linked and "N"-linked oligosaccharide units are also present. Antibodies directed against the protein core of calf bone proteoglycan do not cross-react with cartilage, skin, corneal, or basement membrane proteoglycans in immunoassays and have minimal cross-reactivity with scleral proteoglycans. Quantitative immunoassays and indirect immunofluorescence were used to show that the molecule is localized to forming bone trabeculae and dentin, but not to any other tissue. Osteoblasts and osteoprogenitor cells adjacent to areas undergoing rapid osteogenesis also contain this small proteoglycan. A second proteoglycan (Mr approximately equal to 1,000,000) was extracted from newly forming bone prior to demineralization. This large proteoglycan, which was isolated from the cartilage-free areas of developing intramembranous bone, has a protein core similar to that of the cartilage aggregating proteoglycan and cross-reacts with antisera raised against these cartilage proteoglycans but not with the small mineral-entrapped proteoglycan. It contains larger (Mr = 40,000) and fewer chondroitin sulfate chains than its cartilage-derived analogue, and is localized to the soft connective tissue mesenchyme lying between growing bone trabeculae. More fully formed compact bone did not contain detectable quantities of this proteoglycan.     

Analysis of intimal proteoglycans in atherosclerosis-prone and atherosclerosis-resistant human arteries by mass spectrometry

The propensity to develop atherosclerosis varies markedly among different sites in the human vasculature. To determine a possible cause for such differences in atherosclerosis susceptibility, a proteomics-based approach was used to assess the extracellular proteoglycan core protein composition of intimal hyperplasia from both the atherosclerosis-prone internal carotid artery and the atherosclerosis-resistant internal thoracic artery. The intimal proteoglycan composition in these preatherosclerotic lesions was found to be more complex than previously appreciated with up to eight distinct core proteins present, including the large extracellular proteoglycans versican and aggrecan, the basement membrane proteoglycan perlecan, the class I small leucine-rich proteoglycans biglycan and decorin, and the class II small leucine-rich proteoglycans lumican, fibromodulin, and prolargin/PRELP (proline arginine-rich end leucine-rich repeat protein). Although most of these proteoglycans seem to be present in similar amounts at the two locations, there was a selective enhanced deposition of lumican in the intima of the atherosclerosis-prone internal carotid artery compared with the intima of the atherosclerosis-resistant internal thoracic artery. The enhanced deposition of lumican in the intima of an atherosclerosis prone artery has important implications for the pathogenesis of atherosclerosis.     

The partial amino acid sequence of bovine cartilage proteoglycan, deduced from a cDNA clone, contains numerous Ser-Gly sequences arranged in homologous repeats.

We have determined the sequence of a partial cDNA clone encoding the C-terminal region of bovine cartilage aggregating proteoglycan core protein. The deduced amino acid sequence contains a cysteine-rich region which is homologous with chicken hepatic lectin. This lectin-homologous region has previously been identified in rat and chicken cartilage proteoglycan. The bovine sequence presented here is highly homologous with the rat and chicken amino acid sequences in this apparently globular region. A region containing clusters of Ser-Gly sequences is located N-terminal to the lectin homology domain. These Ser-Gly-rich segments are arranged in tandemly repeated, approx. 100-residue-long, homology domains. Each homology domain consists of an approx. 75-residue-long Ser-Gly-rich region separated by an approx. 25-residue-long segment lacking Ser-Gly dipeptides. These dipeptides are arranged in 10-residue-long segments in the 100-residue-long homology domains. The shorter homologous segments are tandemly repeated some six times in each 100-residue-long homology domain. Serine residues in these repeats are potential attachment sites for chondroitin sulphate chains.     

Ultrastructural characterization of embryonic chick cartilage proteoglycan core protein and the mapping of a monoclonal antibody epitope.

The ultrastructure of embryonic chick cartilage proteoglycan core protein was investigated by electron microscopy of specimens prepared by low angle shadowing. The molecular images demonstrated a morphological substructural arrangement of three globular and two linear regions within each core protein. The internal globular region (G2) was separated from two terminally located globular regions (G1 and G3) by two elongated strands with lengths of 21 +/- 3 nm (E1) and 105 +/- 22 nm (E2). The two N-terminal globular regions, separated by the 21-nm segment, were consistently visualized in well spread molecules and showed little variation in the length of the linear segment connecting them. The E2 segment, however, was quite variable in length, and the C-terminal globular region (G3) was detected in only 53% of the molecules. The G1, G2, and G3 regions in chick core protein were 10.1 +/- 1.7 nm, 9.7 +/- 1.3 nm, and 8.3 +/- 1.3 nm in diameter, respectively. These results are similar to those described previously for proteoglycan core proteins isolated from rat chondrosarcoma, bovine nasal cartilage, and pig laryngeal cartilage (Paulsson, M., Morgelin, M., Wiedemann, H., Beardmore-Gray, M., Dunham, D., Hardingham, T., Heinegard, D., Timpl, R., and Engel, J. (1987) Biochem. J. 245, 763-772). However, a significant difference was detected between the length of the elongated strand (E2) of core proteins isolated from chick cartilage, E2 length = 105 +/- 22 nm, compared to bovine nasal cartilage, E2 length = 260 +/- 39 nm. The epitope of the proteoglycan core protein-specific monoclonal antibody, S103L, was visualized by electron microscopy, and the distance from the core protein N terminus to the S103L binding site was measured. The S103L binding site was localized to the E2 region, 111 +/- 20 nm from the G1 (N terminus) domain and 34 nm from the G3 (C terminus) domain. cDNA clones selected from an expression vector library of chicken cartilage mRNA also show this epitope to be located near the C-terminal region (R. C. Krueger, T. A. Fields, J. Mensch, and B. Schwartz (1990) J. Biol. Chem. 265, 12088-12097).     

A monoclonal antibody that specifically recognizes a glucuronic acid 2-sulfate-containing determinant in intact chondroitin sulfate chain

Monoclonal antibodies produced against chick embryo limb bud proteoglycan (PG-M) were selected for their ability to recognize determinants on intact chondroitin sulfate chains. One of these monoclonal antibodies (IgM; designated MO-225) reacts with PG-M, chick embryo cartilage proteoglycans (PG-H, PG-Lb, and PG-Lt), and bovine nasal cartilage proteoglycan, but not with Swarm rat chondrosarcoma proteoglycan. The reactivity of PG-H to MO-225 is not affected by keratanase digestion but is completely abolished after chondroitinase digestion. Competitive binding analyses with various glycosaminoglycan samples indicate that the determinant recognized by MO-225 resides in a D-glucuronic acid 2-sulfate(beta 1----3)N-acetylgalactosamine 6-sulfate disaccharide unit (D-unit) common to antigenic chondroitin sulfates. A tetrasaccharide trisulfate containing D-unit at the reducing end is the smallest chondroitin sulfate fragment that can inhibit the binding of the antibody to PG-H. Decreasing the size of a D-unit-rich chondroitin sulfate by hyaluronidase digestion results in progressive reduction in its inhibitory activity. The results suggest that the epitope has a requirement for a long stretch of a disaccharide-repeating structure for a better fit to the antibody.     

Deduced protein sequence of bone small proteoglycan I (biglycan) shows homology with proteoglycan II (decorin) and several nonconnective tissue proteins in a variety of species

The small proteoglycans (PG) of bone consist of two different molecular species: one containing one chondroitin sulfate chain (PG II) and the other, two chains (PG I). These two proteoglycans are found in many connective tissues and have Mr = 45,000 core proteins with clear differences in their NH2-terminal sequences. Using antisera produced against synthetic peptides derived from the human PG I and PG II NH2 termini, we have isolated several cDNA clones from a lambda gt11 expression library made against mRNA isolated from human bone-derived cells. The clones, which reacted with antisera to the PG II peptide, were sequenced and found to be identical with the PG II class of proteoglycan from human fibroblasts known as PG-40 or decorin. The clones reacting to the PG I antisera, however, had a unique sequence. The derived protein sequence of PG I showed sufficient homology with the PG II sequence (55% of the amino acids are identical, with most others involving chemically similar amino acid substitutions) to strongly suggest that the two proteins were the result of a gene duplication. PG II (decorin) contains one attached glycosaminoglycan chain, while PG I probably contains two chains. For this reason, we suggest that PG I be called biglycan. The biglycan protein sequence contains 368 residues (Mr = 42,510 for the complete sequence and Mr = 37,983 for the secreted form) that appears to consist predominantly of a series of 12 tandem repeats of 24 residues. The repeats are recognized by their conserved leucines (and leucine-like amino acids) in positions previously reported for a diverse collection of proteins (none of which is thought to be proteoglycans) including: two morphogenic proteins (toll and chaoptin) in the fruit fly; a yeast adenylate cyclase; and two human proteins, the von Willebrand Factor-binding platelet membrane protein, GPIb, and a rare serum protein, leucine-rich glycoprotein.