Interaction of endothelial cell growth factor with heparin: characterization by receptor and antibody recognition.

Schreiber, Alain B.; Kenney, John S.; Kowalski, William J.; Friesel, Robert; Mehlman, Tevie; Maciag, Thomas

doi:10.1073/pnas.82.18.6138

Cited by 272 publications

(172 citation statements)

References 26 publications

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“…This affinity is considered to be involved in binding to cell-surface matrixes which is distinguishable from receptor binding, so-called high-affinity binding [29]. The physiological role of this affinity for heparin is thought to be to retain the molecule in basement membranes [30,311, potentiating and stabilizing it [32]. This affinity appears to protect FGF from inactivation [33] or proteolytic degradation [34].…”

Section: Discussionmentioning

confidence: 99%

Carboxyl‐terminal structure of basic fibroblast growth factor significantly contributes to its affinity for heparin

Seno

Sasada

Kurokawa

et al. 1990

European Journal of Biochemistry

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The carboxyl-terminal sequence of basic fibroblast growth factor (bFGF) is rich in basic amino acid residues, a common characteristic amongst fibroblast growth factors, and is considered to contribute greatly to the binding to negatively charged extracellular matrixes such as heparin. To study the relationship between the affinity for heparin and the carboxyl-terminal structure of bFGF, amino-or carboxyl-terminal truncated molecules were produced in Escherichia coli using recombinant DNA techniques. These terminally truncated bFGFs were applied to a heparin-affinity HPLC column. Truncation of more than six amino acid residues from the carboxyl-terminal made the bFGF produced in E. coli markedly difficult to solubilize and weakened its affinity for heparin, though bFGF having up to 46 amino acids removed showed significant stimulation of the DNA synthesis of BALB/c3T3 cells. This stimulation of the DNA synthesis was also recognized by the bFGF having 40 amino acids removed from its amino-terminal, while the affinity of this peptide for heparin has been shown to be equal to that of the mature bFGF (146 amino acids). These results show that the affinity of bFGF for heparin depends significantly on its carboxyl-terminal structure and that the essential part for receptor binding is present between Asp4' and SerlOO. Moreover, it suggests that the Phe139Leu'40Pro141, present in all members of the FGF family, contributes greatly to the stable structure of the intact molecule. . Extraordinarily high affinity for heparin is one of the important characteristics of this family. In fact, this characteristic has been applied to the isolation and characterization of these proteins, especially basic FGF (bFGF) and acidic FGF (aFGF), from various tissues [15] and tumors [16]. The identification of specific fundamental domains of bFGF for receptor and heparin binding has been reported using synthetic polypeptides; these have suggested the existence of two domains for heparin binding and one for receptor binding [I 5 , 171. We isolated the cDNA for human bFGF and studied the expression of this cDNA in mammalian cells [18, 191 and in bacterial cells (201. We also purified and characterized the recombinant bFGF produced in Escherichia coli [20] and studied the contribution of the four cysteine residues to the protein conformation of bFGF through a site-directed mutagenesis procedure [21]. For further characterization, we constructed several bFGF molecules whose amino-or carboxyl-terminus was deleted. Here we report the structural characterization for heparin binding using these terminally truncated molecules.Correspondence to M . Seno, Biotechnology Research Laboratories, Research and Development Div., Takeda Chemical Industries Ltd, 2-17-85 Jusohonmachi, Yodogawa-ku, Osaka 532, JapanAbbreviations. F G F , fibroblast growth factor; bFGF, basic fibroblast growth factor; aFGF, acidic fibroblast growth factor.Enzymes. DNA polymerase I (EC 2.7.7.7); exonucleasc I11 (EC 3.1.11.2); lysozyme (EC 3.2.1.17); peroxidase (EC 1.11.1.7). MA...

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Section: Discussionmentioning

confidence: 99%

Carboxyl‐terminal structure of basic fibroblast growth factor significantly contributes to its affinity for heparin

Seno

Sasada

Kurokawa

et al. 1990

European Journal of Biochemistry

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“…From the data discussed here, it is proposable that heparin, byselectively binding in the proximity of certain areas of the cell surface, might interfere with growth factor/receptor coupling, thus disrupting the flow of mitogenic signalling from the surface to the nucleus. An interaction between heparin and growth factors has been proposed to explain some effects of heparin on endothelial cell growth [22,23]. The matter, however, is far from being clear, since heparin enhances the effect of ECGF and a-FGF [9], but inhibits proliferation of exponentially growing HOME cells [I I].…”

Section: Discussionmentioning

confidence: 99%

Effect of heparin on proliferation and signalling in BC3H‐1 muscle cells

Vannucchi¹,

Pasquali²,

Fiorelli

et al. 1990

FEBS Letters

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We studied binding and growth inhibitory properties of different glycosaminoglycans in growing and differentiated BC3H-I muscle cells. Heparin (10 pg/ml) and heparan sulfate (10 ,ug/ml) significantly inhibited DNA synthesis in growing and differentiated cells, as monitored by PH]thymidine incorporation. Binding of heparin to BC3H-1 cells was specific and time-dependent. Heparan sulfate was the only glycosaminoglycan able to displace PH]heparin (I&,, 3.2 x lo-' M), although it was lo-fold less effective than heparin itself (IC,,, 3.6 x lo-* M). Scatchard analysis revealed the existence of high-alBnity heparin binding sites (K,,, 5 x lo-* M). Furthermore, heparin inhibited serum-induced stimulation of inositol lipid turnover. Taken together, these results indicate that heparin inhibits BC3H-1 cell growth by interacting with the cell surface, possibly disrupting the flow of growth factor-related mitogenic signalling.

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“…Other factors that exert a phenotypic effect on PC12 cells include FGF, IL-6, and PMA. FGF (50 ng/ml), in the presence of heparin [which stabilizes and potentiates the effect of acidic FGF (29) but has no effect by itself] (Fig. 2B, lane 4), induced an increase of PC3 mRNA levels =1/15th of those induced by NGF, whereas PMA (0.1 ,uM) and IL-6 (1000 units) were even less effective (=1/50th) (Fig.…”

mentioning

confidence: 99%

Molecular cloning of PC3, a putatively secreted protein whose mRNA is induced by nerve growth factor and depolarization.

Bradbury

Possenti

Shooter

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

173

145

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PC3 is an immediate early gene induced by nerve growth factor in PC12 cells, a cell line derived from a tumor of the adrenal medulla that undergoes neuronal differentiation in the presence of nerve growth factor. This induction is independent of new protein synthesis as it can occur in the presence of cycloheximide. PC3 is also induced with similar kinetics, but at lower levels, by membrane depolarization (both in vivo and in vitro) and epidermal growth factor. It is induced at much lower levels by fibroblast growth factor and interleukin 6. In vivo it is found expressed in tissues, such as brain at embryonic day 13.5, placenta, amnion, and spleen, which are proliferating and/or differentiating. The deduced protein sequence from the cDNA indicates the presence of a signal peptide, suggesting that PC3 is secreted.

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Interaction of endothelial cell growth factor with heparin: characterization by receptor and antibody recognition.

Abstract: ABSTRACT

Cited by 272 publications

References 26 publications

Carboxyl‐terminal structure of basic fibroblast growth factor significantly contributes to its affinity for heparin

Carboxyl‐terminal structure of basic fibroblast growth factor significantly contributes to its affinity for heparin

Effect of heparin on proliferation and signalling in BC3H‐1 muscle cells

Molecular cloning of PC3, a putatively secreted protein whose mRNA is induced by nerve growth factor and depolarization.

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