The heparin-binding domain of HB-EGF mediates localization to sites of cell-cell contact and prevents HB-EGF proteolytic release

Abstract

SummaryHeparin-binding EGF-like growth factor (HB-EGF) is a ligand for EGF receptor (EGFR) and possesses the ability to signal in juxtacrine, autocrine and/or paracrine mode, with these alternatives being governed by the degree of proteolytic release of the ligand. Although the spatial range of diffusion of released HB-EGF is restricted by binding heparan-sulfate proteoglycans (HSPGs) in the extracellular matrix and/or cellular glycocalyx, ascertaining mechanisms governing non-released HB-EGF localization is also important for understanding its effects. We have employed a new method for independently tracking the localization of the extracellular EGFlike domain of HB-EGF and the cytoplasmic C-terminus. A striking observation was the absence of the HB-EGF transmembrane proform from the leading edge of COS-7 cells in a wound-closure assay; instead, this protein localized in regions of cell-cell contact. A battery of detailed experiments found that this localization derives from a trans interaction between extracellular HSPGs and the HB-EGF heparin-binding domain, and that disruption of this interaction leads to increased release of soluble ligand and a switch in cell phenotype from juxtacrine-induced growth inhibition to autocrine-induced proliferation. Our results indicate that extracellular HSPGs serve to sequester the transmembrane pro-form of HB-EGF at the point of cell-cell contact, and that this plays a role in governing the balance between juxtacrine versus autocrine and paracrine signaling.

ETOILERegulates Developmental Patterning in the Filamentous Brown AlgaEctocarpus siliculosus

“…Sulfated polysaccharides present within the ECM play a determining role in cell fate by regulating ligand binding to transmembrane receptors. For example, heparan sulfate, a sulfated ECM glycosaminoglycan, regulates the diffusion of the ligand of the epidermal growth factor (EGF) receptor through the ECM by mediating its sequestration by proteoglycans (Prince et al, 2010). This sequestration engages the cell in growth inhibition instead of cell proliferation.…”

Pharmacology of Heparin and Related Drugs

“…In animal models, application of heparin-binding epidermal growth factor-like growth factor (HB-EGF)-which is known to be upregulated both in human burn tissue and during healing of partial-thickness burn injuries specifically through potentiation of the expression of transforming growth factor-a, another member of the EGF family of growth factors involved in wound repair (Cribbs et al, 1998)-can promote healing of ileal tissue 120 after experimental reanastomosis surgery (Radulescu et al, 2011). Localization of HB-EGF through binding to HSPG facilitates juxtacrine inhibition of cell proliferation by this growth factor, and disruption of this binding allows the released HB-EGF to function as an autocrine mitogen (Prince et al, 2010). Therefore, it is possible that soluble heparin at the site of injury could act competitively to release this growth factor from HS binding sites and promote participation in tissue repair.…”

Trafficking of the EGFR ligand Spitz regulates its signaling activity in polarized tissues

“…Studies in vitro showed that a TGF-a proprotein carrying mutations in the metalloproteinase cleavage site could signal, but in vivo, mutation of the metalloproteinase itself appeared to recapitulate loss of TGF-a (Brachmann et al, 1989;Peschon et al, 1998;Sanderson et al, 2006;Wong et al, 1989). In recent years, mounting evidence has been provided in support of pro-protein signaling, both by TGF-a and by HB-EGF, which has opposing effects on cell survival in its membrane-bound and secreted forms (Pan et al, 2002;Prince et al, 2010;Shi et al, 2000;Singh et al, 2007;Yang et al, 2000). Some pro-proteins, such as amphiregulin, might be released on exosomes, allowing them to act at a distance (Higginbotham et al, 2011).…”

“…Mammalian EGFR ligands have also been observed to induce juxtacrine pathway activation by binding to their receptors in their transmembrane pro-protein forms (Anklesaria et al, 1990;Brachmann et al, 1989;Wong et al, 1989). Juxtacrine and paracrine activation of the receptor can result in different outcomes, suggesting that regulation of ligand cleavage can determine the consequences of pathway activity (Iwamoto et al, 1999;Pan et al, 2002;Prince et al, 2010;Singh et al, 2004;Takemura et al, 1997).…”