The core protein of the matrix-associated heparan sulfate proteoglycan binds to fibronectin.

A cDNA clone coding for a membrane proteoglycan core protein was isolated from a neonatal rat Schwann cell cDNA library by screening with an oligonucleotide based on a conserved sequence in cDNAs coding for previously described proteoglycan core proteins. Primer extension and polymerase chain reaction amplification were used to obtain additional 5' protein coding sequences. The deduced amino acid sequence predicted a 353 amino acid polypeptide with a single membrane spanning segment and a 34 amino acid hydrophilic COOH-terminal cytoplasmic domain. The putative extracellular domain contains three potential glycosaminoglycan attachment sites, as well as a domain rich in Thr and Pro residues. Analysis of the cDNA and deduced amino acid sequences revealed a high degree of identity with the transmembrane and cytoplasmic domains of previously described proteoglycans but a unique extracellular domain sequence. On Northern blots the cDNA hybridized to a single 5.6-kb mRNA that was present in Schwann cells, neonatal rat brain, rat heart, and rat smooth muscle cells. A 16-kD protein fragment encoded by the cDNA was expressed in bacteria and used to immunize rabbits. The resulting antibodies reacted on immunoblots with the core protein of a detergent extracted heparan sulfate proteoglycan. The core protein had an apparent mass of 120 kD. When the anti-core protein antibodies were used to stain tissue sections immunoreactivity was present in peripheral nerve, newborn rat brain, heart, aorta, and other neonatal tissues. A ribonuclease protection assay was used to quantitate levels of the core protein mRNA. High levels were found in neonatal rat brain, heart, and Schwann cells. The mRNA was barely detectable in neonatal or adult liver, or adult brain.

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Selective deglycosylation of the heparan sulfate proteoglycan of bovine glomerular basement membrane and identification of the core protein.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)48328-7 ◽

1987 ◽

Vol 262 (14) ◽

pp. 6893-6898

Author(s):

A S Edge ◽

R G Spiro

Keyword(s):

Basement Membrane ◽

Heparan Sulfate ◽

Glomerular Basement Membrane ◽

Core Protein ◽

Heparan Sulfate Proteoglycan ◽

Sulfate Proteoglycan ◽

The Core

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Characterization of a novel heparan sulfate proteoglycan found in the extracellular matrix of liver sinusoids and basement membranes.

The Journal of Cell Biology ◽

10.1083/jcb.113.5.1231 ◽

1991 ◽

Vol 113 (5) ◽

pp. 1231-1241 ◽

Cited By ~ 33

Author(s):

C J Soroka ◽

M G Farquhar

Keyword(s):

Extracellular Matrix ◽

Basement Membrane ◽

Heparan Sulfate ◽

Rat Liver ◽

Core Protein ◽

Heparan Sulfate Proteoglycan ◽

Basement Membranes ◽

Sulfate Proteoglycan ◽

Sinusoidal Cell ◽

Space Of Disse

A novel heparan sulfate proteoglycan (HSPG) present in the extracellular matrix of rat liver has been partially characterized. Proteoglycans were purified from a high salt extract of total microsomes from rat liver and found to consist predominantly (approximately 90%) of HSPG. A polyclonal antiserum raised against this fraction specifically recognized HSPG by immunoprecipitation and immunoblotting. The intact, fully glycosylated HSPG migrated as a broad smear (150-300 kD) by SDS-PAGE, but after deglycosylation with trifluoromethanesulfonic acid only a single approximately 40-kD band was seen. By immunocytochemistry this HSPG was localized in the perisinusoidal space of Disse associated with irregular clumps of basement membrane-like extracellular matrix material, some of which was closely associated with the hepatocyte sinusoidal cell surface. It was also localized in biosynthetic compartments (rough ER and Golgi cisternae) of hepatocytes, suggesting that this HSPG is synthesized and deposited in the space of Disse by the hepatocyte. The anti-liver HSPG IgG also stained basement membranes of hepatic blood vessels and bile ducts as well as those of kidney and several other organs (heart, pancreas, and intestine). An antibody that recognizes the basement membrane HSPG found in the rat glomerular basement membrane did not precipitate the 150-300-kD rat liver HSPG. We conclude that the liver sinusoidal space of Disse contains a novel population of HSPG that differs in its overall size, its distribution and in the size of its core protein from other HSPG (i.e., membrane-intercalated HSPG) previously described in rat liver. It also differs in its core protein size from HSPG purified from other extracellular matrix sources. This population of HSPG appears to be a member of the basement membrane HSPG family.

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Basement membrane proteoglycan in various tissues: characterization using monoclonal antibodies to the Engelbreth-Holm-Swarm mouse tumor low density heparan sulfate proteoglycan.

The Journal of Cell Biology ◽

10.1083/jcb.106.6.2203 ◽

1988 ◽

Vol 106 (6) ◽

pp. 2203-2210 ◽

Cited By ~ 46

Author(s):

M Kato ◽

Y Koike ◽

S Suzuki ◽

K Kimata

Keyword(s):

Monoclonal Antibodies ◽

Heparan Sulfate ◽

Heparan Sulfate Proteoglycan ◽

Basement Membranes ◽

Immunofluorescent Staining ◽

Mouse Tumor ◽

Low Density ◽

Sulfate Proteoglycan ◽

The Core ◽

Core Proteins

The Engelbreth-Holm-Swarm mouse tumor has been found to produce at least two molecular species of heparan sulfate proteoglycan, a low density one (LD) and a high density one, which differ not only in core proteins but also in glycosaminoglycan structures (Kato, M., Y. Koike, Y. Ito, S. Suzuki, and K. Kimata. 1987. J. Biol. Chem. 262:7180-7188). With aim at investigating their distribution and possible functions in tissues, monoclonal antibodies were produced. Hybridomas obtained by fusion of NS-1 mouse myeloma cells with spleen cells from the rat immunized with a mixture of these proteoglycans were selected by their ability to react with the antigen. Two of them secreted monoclonal antibodies (IgG2a), designated HK-84 and HK-102, that recognize specifically the core protein moiety of LD. Immunofluorescent staining of various tissues (skeletal muscle, cardiac muscle, lung, brain, and kidney) with these monoclonal antibodies has demonstrated that the antigen molecules were present in all basement membranes of these tissues. SDS-PAGE of heparitinase-treated proteoglycan fractions prepared from these tissues and subsequent immunoblotting using these monoclonal antibodies have confirmed that the antigen molecule was LD, and further suggested that there was a tissue-specific variation in the core molecular size. Based on these results, we propose that LD may be an essential component in all basement membranes.

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Matrix-associated heparan sulfate proteoglycan: core protein-specific monoclonal antibodies decorate the pericellular matrix of connective tissue cells and the stromal side of basement membranes.

The Journal of Cell Biology ◽

10.1083/jcb.109.6.3199 ◽

1989 ◽

Vol 109 (6) ◽

pp. 3199-3211 ◽

Cited By ~ 42

Author(s):

A Heremans ◽

B van der Schueren ◽

B de Cock ◽

M Paulsson ◽

J J Cassiman ◽

...

Keyword(s):

Extracellular Matrix ◽

Monoclonal Antibodies ◽

Epithelial Cell ◽

Heparan Sulfate ◽

Core Protein ◽

Structural Features ◽

Heparan Sulfate Proteoglycan ◽

Basement Membranes ◽

Lung Fibroblasts ◽

Sulfate Proteoglycan

Cultured human lung fibroblasts produce a large, nonhydrophobic heparan sulfate proteoglycan that accumulates in the extracellular matrix of the monolayer (Heremans, A., J. J. Cassiman, H. Van den Berghe, and G. David. 1988. J. Biol. Chem. 263: 4731-4739). A panel of four monoclonal antibodies, specific for four distinct epitopes on the 400-kD core protein of this extracellular matrix heparan sulfate proteoglycan, detects similar proteoglycans in human epithelial cell cultures. Immunohistochemistry of human tissues with the monoclonal antibodies reveals that these proteoglycans are concentrated at cell-matrix interfaces. Immunogold labeling of ultracryosections of human skin indicates that the proteoglycan epitopes are nonhomogeneously distributed over the width of the basement membrane. Immunochemical investigations and amino acid sequence analysis indicate that the proteoglycan from the fibroblast matrix shares several structural features with the large, low density heparan sulfate proteoglycan isolated from the Engelbreth-Holm-Swarm sarcoma. Thus, both epithelial cell sheets and individual mesenchymal cells accumulate a large heparan sulfate proteoglycan(s) at the interface with the interstitial matrix, where the proteoglycan may adopt a specific topological orientation with respect to this matrix.

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