Sulfated Oligosaccharides Promote Hepatocyte Growth Factor Association and Govern Its Mitogenic Activity

Abstract

Hepatocyte growth factor (HGF) is a potent mitogen, motogen, and morphogen for various epithelial cell types. The pleiotropic effects of HGF are mediated by its binding to a specific high affinity receptor, c-Met. In addition, HGF binds to heparan sulfate proteoglycans on cell surfaces and within the extracellular matrix. Incubation of HGF with 0.1, 1.0, and 10 micrograms/ml of heparin, heparan sulfate, or dextran sulfate resulted in a concentration-dependent increase in mitogenic potency in a primary rat hepatocyte bioassay, whereas sodium sulfate or fucoidan did not. Although co-incubation of HGF with sulfated compounds that enhanced HGF-dependent mitogenesis did not alter the binding isotherm of HGF for the c-Met receptor in a solid phase assay, an increase in autophosphorylation of the c-Met receptor in intact A549 cells was observed upon their addition. A series of chemically sulfated malto-oligosaccharides varying in unit size and charge was tested in the bioassay in order to provide additional insights into the nature of the HGF-heparin interaction. While sulfated di-, tri-, tetra-, and pentasaccharides did not significantly potentiate HGF-dependent mitogenesis, larger oligosaccharides such as the sulfated hexa-, hepta-, or a sulfated oligosaccharide mixture containing decasaccharides resulted in an approximate 2-, 4-, and 7-fold enhancement, respectively. We observed a correlation between the sulfated oligosaccharide preparations that enhanced mitogenic potency and those that promoted HGF oligomerization in vitro, as measured by gel filtration and analytical ultracentrifugation. These findings indicate that heparin-like molecules can stabilize HGF oligomers, which may facilitate c-Met receptor dimerization and activation.

Dissociation of Heparan Sulfate and Receptor Binding Domains of Hepatocyte Growth Factor Reveals That Heparan Sulfate-c-Met Interaction Facilitates Signaling

“…In that system, added heparin facilitates HGF signaling (11), through a mechanism that was presumed to involve its demonstrated ability to enhance HGF oligomerization (12,18,19). Despite its inability to bind heparin, we found that K1 mitogenic signaling in 32D/c-Met cells was completely heparin-dependent.…”

Heparanase Induces Endothelial Cell Migration via Protein Kinase B/Akt Activation

“…The augmentation of Akt phosphorylation levels by heparin (Fig. 3, C and D) may suggest the existence of a specific heparanase receptor that, similar to several growth factor receptors, requires heparin to exert maximal activities (31,35,36). Studies aiming to identify this putative heparanase receptor are currently in progress.…”

Proteoglycans: Master regulators of molecular encounter?

“…Among these, some may be considered "HS-dependent", since in vitro manipulations that remove, or prevent the sulfation of, cellular HS result in greatly decreased growth factor binding to, and activation of, signaling receptors. The best characterized HS-dependent growth factors are FGF-1 and FGF-2, but other FGFs, HB-EGF, wingless (a member of the Wnt family) and probably several other growth factors belong to this group (Rapraeger et al, 1991;Yayon et al, 1991;Aviezer and Yayon, 1994;Tessler et al, 1994;Zioncheck et al, 1995;Reichsman et al, 1996). Models that have been put forth to explain HS-dependence include ones in which binding of HS to growth factor, receptor, or both directly alters affinity (e.g., through cross-linking, dimerization, or induction of a conformational change [Kan et al, 1993;Pantoliano et al, 1994;Spivak-Kroizman et al, 1994]).…”