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Y Sommarin

Publications and source records attributed to Y Sommarin.

At least 19 recordsLinked to original sources
PubMed2000

The amino-terminal part of PRELP binds to heparin and heparan sulfate.

PRELP (proline, arginine-rich end leucine-rich repeat protein) is an extracellular matrix leucine-rich repeat protein. The amino-terminal region of PRELP differs from that of other leucine-rich repeat proteins in containing a high number of proline and arginine residues. The clustered proline and basic residues are conserved in rat, bovine, and human PRELP. Although the function of PRELP is not yet known, the clustered arginine residues suggest a heparan sulfate/heparin-binding capacity. We show here that PRELP indeed binds heparin and heparan sulfate. Truncated PRELP without the amino-terminal region does not bind heparin. The dissociation constant for the interaction of PRELP with heparin was determined by an in solution binding assay and by surface plasmon resonance analysis to be in the range of 10-30 nm. A 6-mer heparin oligosaccharide was the smallest size showing binding to PRELP. The binding increased with increasing length up to an 18-mer and depended on the degree of sulfation of heparin as well as heparan sulfate. Sulfate groups at all positions were shown to be of importance for the binding. Fibroblasts bind PRELP, and this interaction is inhibited with heparin, suggesting a function for PRELP as a linker between the matrix and cell surface proteoglycans.

Amino Acid Sequence↗
PubMed2000

Synthesis of hyaluronan by normal and wounded rabbit iris.

BACKGROUND: Endogenous hyaluronan has been found in different tissues in the normal and traumatized eye. However, the main source, the biological aspects and the full potential role of hyloronan are still unclear. METHODS: Hyaluronan production was studied both in organ culture and in vivo, using a double-label protocol with [35S]sulfate and [3H]glucosamine. RESULTS: [3H]glucosamine and [35S]sulfate were incorporated into hyaluronan and sulfated glycosaminoglycans in normal and in traumatized iris tissue in organ culture and in vivo. There was low relative hyaluronan synthesis in vivo, only 2% of total incorporated [3H]glucosamine in normal irides. Increased relative incorporation of [3H]glucosamine into hyaluronan was seen after operative trauma to iris tissue both in vivo and in vitro. CONCLUSION: Our findings demonstrate synthesis of hyaluronan by normal and traumatized iris. The iris seems to be the most important source of aqueous hyaluronan.

Animals↗
PubMed1999

The human CILP gene: exon/intron organization and chromosomal mapping.

The human cDNA for cartilage intermediate layer protein (CILP) codes for a larger precursor protein that consists of CILP and a homologue to porcine Nucleotide pyrophosphohydrolase (NTPPHase) [Lorenzo et al. 1998a. J. Biol. Chem. 273, 23469-23475]. The human gene has now been isolated and characterized. Southern blot analysis indicated a single copy of the CILP gene in the human genome. The gene spans approximately 15.3 kbp of genomic DNA, and is organized in nine exons. The 5' flanking region contains a putative promoter region with a TATA-like box localized from -29 to -23 bp upstream of the transcription start site. Analysis of the putative promoter region revealed potentially cis-regulatory eukaryotic elements such as GATA-1, MyoD, MZF1, and CdxA. The protein coding region begins in exon 2 with the putative signal peptide. CILP is encoded from exon 3 to exon 9. In addition, exon 9 also codes for the entire NTPPHase homologue and contains the 3' untranslated region of the gene. All the introns follow the 'gt-ag' rule, except the last intron, intron 8, that belongs to the minor class of pre-mRNA introns that contain 'at-ac' at their 5' and 3' ends, respectively. The CILP gene was mapped to human chromosome 15q22.

Base Sequence↗
PubMed1999

Tissue distribution of a novel cell binding protein, osteoadherin, in the rat.

Osteoadherin is a cell binding keratan sulfate proteoglycan which was recently isolated from mineralized bovine bone and subsequently cloned and sequenced. For studies of osteoadherin expression in rat tissues we isolated and sequenced a 1.3-kbp partial cDNA covering most of the coding region using a rat calvaria cDNA library. The most 5' end of the cDNA was obtained by reverse transcription PCR from the bone total RNA preparation. The deduced, translated protein sequence containing 423 amino acid residues shows high sequence identity to mouse, bovine and human osteoadherin except in the very acidic C-terminal region. However, the rat counterpart showed a similarly high content of acidic amino acid residues. Ribonuclease protection assay showed osteoadherin mRNA to be expressed in femoral bone and calvaria tissues, while no expression was detected in cartilage, tendon or liver. Using very sensitive nested RT-PCR, however, message was detected in femoral head, rib, tendon and bone marrow total RNA preparations. An antiserum specific for the rat C-terminal region of osteoadherin was generated and used for studies of protein distribution by immunohistochemistry during femoral head development. Osteoadherin was primarily present in bone trabeculae and no staining was seen in cartilage. In situ hybridization showed the strongest expression in osteoblasts close to the cartilage/bone interface of the growth plate and lower expression in diaphyseal osteoblasts. On maturation of the femoral head on day 60 some expression was detected immediately below the forming articular cartilage. Our data indicated that osteoadherin is primarily expressed by osteoblasts and might have a role in regulation of mineralization.

Amino Acid Sequence↗
PubMed1998

Cloning and deduced amino acid sequence of a novel cartilage protein (CILP) identifies a proform including a nucleotide pyrophosphohydrolase.

The cDNA cloning and expression in vitro and in eukaryotic cells of a novel protein isolated from human articular cartilage, cartilage intermediate layer protein (CILP) is described. A single 4. 2-kilobase mRNA detected in human articular cartilage encodes a polypeptide of 1184 amino acids with a calculated molecular mass of 132.5 kDa. The protein has a putative signal peptide of 21 amino acids, and is a proform of two polypeptides. The amino-terminal half corresponds to CILP (molecular mass of 78.5 kDa, not including post-translational modifications) and the carboxyl-terminal half corresponds to a protein homologous to a porcine nucleotide pyrophosphohydrolase, NTPPHase (molecular mass of 51.8 kDa, not including post-translational modifications). CILP has 30 cysteines and six putative N-glycosylation sites. The human homolog of porcine NTPPHase described here contains 10 cysteine residues and two putative N-glycosylation sites. In the precursor protein the NTPPHase region is immediately preceded by a tetrapeptide conforming to a furin proteinase cleavage consensus sequence. Expression of the full-length cDNA in a cell-free translation system and in COS-7 or EBNA cells indicates that the precursor protein is synthesized as a single polypeptide chain that is processed, possibly by a furin-like protease, into two polypeptides upon or preceding secretion.

Amino Acid Sequence↗
PubMed1998

Osteoadherin, a cell-binding keratan sulfate proteoglycan in bone, belongs to the family of leucine-rich repeat proteins of the extracellular matrix.

Osteoadherin is a recently described bone proteoglycan containing keratan sulfate. It promotes integrin (alphav beta3)-mediated cell binding (Wendel, M., Sommarin, Y., and Heinegârd, D. (1998) J. Cell Biol. 141, 839-847). The primary structure of bovine osteoadherin has now been determined by nucleotide sequencing of a cDNA clone from a primary bovine osteoblast expression library. The entire translated primary sequence corresponds to a 49,116-Da protein with a calculated isoelectric point for the mature protein of 5.2. The dominating feature is a central region consisting of 11 B-type, leucine-rich repeats ranging in length from 20 to 30 residues. The full, primary sequence contains four putative sites for tyrosine sulfation, three of which are at the N-terminal end of the molecule. There are six potential sites for N-linked glycosylation present. Osteoadherin shows highest sequence identity, 42%, to bovine keratocan and 37-38% identity to bovine fibromodulin, lumican, and human PRELP. Unique to osteoadherin is the presence of a large and very acidic C-terminal domain. The distribution of cysteine residues resembles that of other leucine-rich repeat proteins except for two centrally located cysteines. Northern blot analysis of RNA samples from various bovine tissues showed a 4.5-kilobase pair message for osteoadherin to be expressed in bone only. Osteoadherin mRNA was detected by in situ hybridization in mature osteoblasts located superficially on trabecular bone.

Amino Acid Sequence↗
PubMed1998

Bone matrix proteins: isolation and characterization of a novel cell-binding keratan sulfate proteoglycan (osteoadherin) from bovine bone.

A small cell-binding proteoglycan for which we propose the name osteoadherin was extracted from bovine bone with guanidine hydrochloride-containing EDTA. It was purified to homogeneity using a combination of ion-exchange chromatography, hydroxyapatite chromatography, and gel filtration. The Mof the proteoglycan was 85, 000 as determined by SDS-PAGE. The protein is rich in aspartic acid, glutamic acid, and leucine. Two internal octapeptides from the proteoglycan contained the sequences Glu-Ile-Asn-Leu-Ser-His-Asn-Lys and Arg-Asp-Leu-Tyr-Phe-Asn-Lys-Ile. These sequences are not previously described, and support the notion that osteoadherin belongs to the family of leucine-rich repeat proteins. A monospecific antiserum was raised in rabbits. An enzyme-linked immunosorbent assay was developed, and showed the osteoadherin content of bone extracts to be 0.4 mg/g of tissue wet weight, whereas none was found in extracts of various other bovine tissues. Metabolic labeling of primary bovine osteoblasts followed by immunoprecipitation showed the cells to synthesize and secrete the proteoglycan. Digesting the immunoprecipitated osteoadherin with N-glycosidase reduced its apparent size to 47 kD, thus showing the presence of several N-linked oligosaccharides. Digestion with keratanase indicated some of the oligosaccharides to be extended to keratan sulfate chains. In immunohistochemical studies of the bovine fetal rib growth plate, osteoadherin was exclusively identified in the primary bone spongiosa. Osteoadherin binds to hydroxyapatite. A potential function of this proteoglycan is to bind cells, since we showed it to be as efficient as fibronectin in promoting osteoblast attachment in vitro. The binding appears to be mediated by the integrin alphavbeta3, since this was the only integrin isolated by osteoadherin affinity chromatography of surface-iodinated osteoblast extracts.

Amino Acid Sequence↗
PubMed1998

Chondroadherin expression changes in skeletal development.

Chondroadherin is a cartilage protein with cell binding properties. The expression of chondroadherin was studied in rat tissues and during postnatal femoral head development. For design of oligonucleotide probes and primers a 1664 bp, full length, rat chondroadherin cDNA was isolated from a rat chondrosarcoma library and sequenced. Northern blot analysis showed chondroadherin mRNA to be present in femoral head and rib cartilage, as well as in tendon. More sensitive reverse-transcriptase PCR additionally identified the mRNA in calvaria, long bone and bone marrow. Localization of chondroadherin by immunocytochemistry in the developing femoral head from postnatal day 14 to day 60 showed presence of the protein in cartilaginous regions. With increasing age a very distinct localization of chondroadherin was seen in the territorial matrix around late proliferative cells in the growth plate as well as in the developing articular cartilage in the maturing femoral head. Localization of chondroadherin mRNA by in situ hybridization was in agreement with immunocytochemistry with strong hybridization signals in late proliferative cells in the growth plate. In the articular cartilage the expression was restricted to cells in the lower regions. A three-fold increase of cartilage chondroadherin content in the growing femoral head was demonstrated by Western blot analysis. The high expression of this cell binding protein in a dynamic region of cartilage suggests an important role for chondroadherin in the regulation of chondrocyte growth and proliferation.

Amino Acid Sequence↗
PubMed1998

The mouse chondroadherin gene: characterization and chromosomal localization.

The mouse chondroadherin gene was isolated from a cosmid genomic library by the use of a rat chondroadherin cDNA probe. Southern blot analysis of mouse genomic DNA revealed a simple pattern of hybridization indicating a single copy gene for chondroadherin. The mouse chondroadherin gene encompasses 4.1 kb and consists of four exons separated by one large intron of 1929 bp followed by two smaller introns of 247 and 225 bp, respectively. Most of the translated region, including the start codon and the main part of a leucine-rich region, is contained within the first exon. Two small exons of 164 and 146 bp encode the rest of the protein. Interestingly, 4 bases from the stop codon, in the 3'-UTR, a third intron is located. A putative promoter region of 669 bp was sequenced and shown to contain a potential TATAA-box signal 29 bp upstream of the transcription start site and several recognition sites for transcription factors. The exon/intron organization of the chondroadherin gene differs from those of the other known genes of the leucine-rich repeat (LRR) family in the extracellular matrix. Taken together with comparison of protein sequences of other members of the LRR family in the extracellular matrix, the data suggest that chondroadherin has evolved along a different pathway. The chondroadherin gene was mapped to mouse chromosome 11, near D11Mit14, by single-strand conformation polymorphism linkage analysis.

Amino Acid Sequence↗
PubMed1995

The primary structure of a basic leucine-rich repeat protein, PRELP, found in connective tissues.

We have determined the primary structure of a connective tissue matrix protein from the nucleotide sequence of a clone isolated from a human articular chondrocyte cDNA library. The major part of the amino acid sequence has also been determined by direct protein sequencing. The translated primary sequence corresponds to 382 amino acid residues, including a 20-residue signal peptide. The molecular mass of the mature protein is 41,646 Da. The main part of the protein consists of 10 leucine-rich repeats ranging in length from 20 to 26 residues, with asparagine at position 10 (B-type). The N-terminal part is unusual in that it is basic and rich in arginine and proline. There are four potential N-linked glycosylation sites present. In three of these sites, post-translational modifications are likely to be present since Asn was not found by direct protein sequencing. The amino- and carboxyl-terminal parts contain four and two cysteine residues, respectively, probably forming disulfide bonds by analogy with the other members of this family. The protein shows highest identity (36%) to fibromodulin and 33% to bovine lumican, two other leucine-rich repeat connective tissue proteins. Northern blot analysis showed the presence of an approximately 3.8-kilobase mRNA in different types of bovine cartilage and cultured osteoblasts, whereas RNAs isolated from bovine kidney, skin, spleen, thymus, and trabecular bone and rat calvaria were negative. Human articular chondrocyte and rat chondrosarcoma cell RNAs contained an additional mRNA of approximately 1.6 and 1.8 kilobases, respectively.

Amino Acid Sequence↗
PubMed1995

Isolation, characterization, and primary structure of a calcium-binding 63-kDa bone protein.

A novel noncollagenous protein of the mineralized matrix of bovine bone was isolated by ion exchange and gel permeation chromatography. The apparent M(r) of the protein is 63,000 as determined by SDS-polyacrylamide gel electrophoresis. The protein is a rather minor constituent in bone and could not be detected in other connective tissues by enzyme-linked immunosorbent assay of guanidine HCl extracts. The 63-kDa protein was detected in the osteoid and around the osteocytes upon immuno-histochemical staining of bovine compact bone. The sequence of the 63-kDa protein was deduced from cDNA clones isolated from a rat calvaria lambda gt11 expression library. The protein contains two centrally located EF-hand Ca(2+)-binding domains. Seven heptad repeats are present indicating the ability of the protein for coiled-coil interactions. Ability to bind calcium was confirmed by 45Ca2+ binding to protein blotted onto nitrocellulose membrane. The protein was synthesized in calvaria explants as detected by immunoprecipitation of radiolabeled protein from the culture medium. Although the protein can be detected in biochemical amounts in bone only, varying amounts of mRNA for this protein were detected in several rat tissues by RNase protection assay with highest levels in rat calvaria. This extracellular protein corresponds to a mouse protein called nucleobindin.

Amino Acid Sequence↗
PubMed1995

Distribution and expression of cartilage oligomeric matrix protein and bone sialoprotein show marked changes during rat femoral head development.

Distribution and sites of synthesis of a cartilage extracellular matrix protein, cartilage oligomeric matrix protein (COMP), and of a bone extracellular matrix protein, bone sialoprotein (BSP), were studied in the femoral head of growing Wistar rats from day 14 to day 60 by immunocytochemistry and in situ hybridization. This period includes formation of the secondary ossification center and differentiation of articular cartilage. At early stages, immunoreactivity for COMP was pronounced throughout the cartilage. The localization of COMP was predominantly territorial in the center of the immature femoral head and in the growth plate at all ages studied. In the superficial parts, a shift from a uniform extracellular matrix staining at day 14 to an interterritorial localization at day 33 to day 60 was seen, apparently concurrent with formation of articular cartilage. COMP staining, representing cartilage remnants, also extended into the center of the trabecular bone in the primary spongiosa. In the secondary ossification center, the staining for COMP decreased at the onset of calcification. The protein was only synthesized by chondrocytes, as shown by in situ hybridization. The highest level of COMP mRNA was detected in chondrocytes in the central region of the growth plate. In the layer corresponding to the articular cartilage of the femoral head, mRNA levels for COMP were low from day 14 to day 33 but were increased on day 60. This shows substantial synthesis in the developing articular cartilage. Immunoreactivity for BSP was detected in bone trabeculae of primary spongiosa. In situ hybridization showed the highest levels of BSP mRNA in regions of newly formed bone. BSP mRNA was detected in hypertrophic chondrocytes in the secondary ossification center as early as day 18, well before the appearance of immunochemically detectable BSP. Interestingly, simultaneous expression of COMP and BSP mRNA was seen after day 18 in hypertrophic chondrocytes of the growth plate and later also in hypertrophic chondrocytes close to the mineralization zone of the articular cartilage.

Animals↗
PubMed1994

The structure of a 38-kDa leucine-rich protein (chondroadherin) isolated from bovine cartilage.

A leucine-rich protein, chondroadherin, has been isolated from dissociative extracts of articular cartilage, and its primary structure has been determined by both direct protein sequencing and DNA sequence analysis of polymerase chain reaction products and cDNA clones. This protein is identical to the 36-kDa protein which was isolated by Larsson et al. (Larsson, T., Sommarin, Y., Paulsson, M., Antonsson, P., Hedbom, E., Wendel, M., and Heinegård, D. (1991) J. Biol. Chem. 266, 20428-20433). It has 337 amino acids and exists in several isoforms. The two major isoforms are a form with a calculated molecular weight of 38,353 and a pI of 9.76 and a smaller form with a calculated molecular weight of 37,304 and a pI of 9.5. The two isoforms result from a cleavage near the C terminus. A further level of heterogeneity is found in that an extra alanine can be found prior to the N-terminal cysteine. There are 9 cysteines; disulfide bonds have been directly identified between Cys282-Cys324 and Cys284-Cys304. The principal feature of the protein is a series of 10 leucine-rich repeats. The most N-terminal of these repeats contains a cysteine (Cys63) which is not disulfide-bonded and which is difficult to derivatize. It is likely that this free cysteine is involved in structure-stabilizing hydrogen bonding. The mRNA is approximately 1.6 kilobases, of which 511 base pairs is a 3'-untranslated region between the stop codon and the polyadenylation signal. Based on anchored polymerase chain reaction analysis of the mRNA, there is some minor heterogeneity in the position of the 5' end of the message.

Amino Acid Sequence↗
PubMed1992

Cartilage matrix proteins. An acidic oligomeric protein (COMP) detected only in cartilage.

An Mr = 524,000 oligomeric protein was isolated from bovine cartilage and designated COMP (Cartilage Oligomeric Matrix Protein). The protein is composed of disulfide-bonded subunits with an apparent Mr of 100,000 each. It is markedly anionic, probably due to its high contents of aspartic acid and glutamic acid, as well as to its substitution with negatively charged carbohydrates. COMP was found in all cartilages analyzed, but could not be detected in other tissues by enzyme-linked immunosorbent assay of guanidine HCl extracts. Within a given cartilage, COMP shows a preferential localization to the territorial matrix surrounding the chondrocytes.

Amino Acids↗
PubMed1991

Cartilage matrix proteins. A basic 36-kDa protein with a restricted distribution to cartilage and bone.

A non-collagenous quantitatively prominent protein was purified from guanidine hydrochloride extracts of bovine tracheal cartilage. Purification was achieved by cesium chloride density gradient centrifugation and chromatography on DEAE-cellulose at pH 7.0 followed by CM-cellulose at pH 5.0. The protein has a marked tendency to form aggregates in denaturing solutions of high ionic strength, e.g. 6 M guanidine hydrochloride. The purified protein contains a single, Mr 36,000 polypeptide chain, with a particularly high content of leucine. It contains about 1% carbohydrate with a remarkable absence of hexosamines and sialic acid, whereas xylose, galactose, mannose, and fucose were identified in the preparation. The protein was identified in extracts of cartilage and bone and could be shown to be primarily extracellular. Tendon may contain trace amounts of the protein, whereas extracts of several other tissues showed no immunoreactivity in enzyme-linked immunosorbent assay.

Animals↗
PubMed1989

Chondrocyte-matrix interactions. Attachment to proteins isolated from cartilage.

Interactions between bovine chondrocytes and cartilage extracellular matrix proteins and proteoglycans were demonstrated by cell attachment to plastic surfaces coated with the macromolecules. Chondrocytes, which had been maintained in suspension culture, attached to fibronectin, bone sialoprotein, collagen II, and two novel 36- and 58-kDa proteins isolated from cartilage. Attachment to fibronectin and bone sialoprotein was inhibited by competition with an Arg-Gly-Asp containing peptide, whereas attachment to the 36- and the 58-kDa proteins was not affected. The chondrocytes attached but did not spread on 36- and 58-kDa proteins. Other matrix proteins tested gave little or no binding of the chondrocytes. These include a 59-kDa protein, a 100-kDa subunit protein, and a 148-kDa protein. Freshly isolated chondrocytes also attached to proteoglycans, while attachment to the other matrix constituents was the same as for cells maintained in spinner culture prior to assay.

Animals↗