News and Publications

Role of Ectodomain Shedding in Regulation of Fibroblast Growth Factor 2 Activity

A. Hanneken, M. Kanemitsu, M. Duanmu, M. Mercado

A fundamental question in growth factor biology is how the activities of extracellular growth factors are so tightly regulated. We study the regulation of fibroblast growth factor 2 (FGF-2), a highly conserved protein that is widely distributed throughout cells and tissues in vivo. FGF-2 plays multiple roles in growth and development, including initiation of cell growth and differentiation, induction of mesodermal development, and enhancement of angiogenesis. We are focusing on the hypothesis that shedding of the extracellular domain of the high-affinity FGF receptor 1 (FGFR-1) regulates the biological activity of FGF-2.

Ectodomain shedding by cell-surface growth factors, receptors, and cell adhesion molecules is an essential process for normal cellular growth and differentiation. Ectodomain shedding occurs via proteolytic cleavage of the extracellular domain of a cell-surface protein from the full-length molecule. We identified soluble FGF receptors in blood and in the extracellular matrix of endothelial cells and showed that these proteins consist of multiple isoforms of the extracellular domain of the high-affinity receptor FGFR-1 and are primarily generated by proteolytic cleavage from full-length FGFR-1 molecules. We hypothesized that the soluble FGF receptors modulate the biological activity of FGF-2 in vivo by 2 mechanisms: downregulation of cell-surface FGF receptors and competition with cell-surface FGF receptors for ligand binding.

A paradigm is unfolding in our laboratory that supports the role of the soluble FGF receptors as inhibitors of the biological activity of the FGF family of growth factors. This model is based on the results of 3 primary research areas: structural characterization of the soluble FGF receptors found in blood, identification of an enzyme that cleaves the soluble FGF receptors from the cell surface, and description of the biological activities of the soluble receptors.

We recently showed that shedding of the FGFR-1 ectodomain can be induced by a member of the ADAM family of proteins. These multifunctional enzymes contain both a disntegrin domain and a metalloprotease domain. We found that ectodomain shedding by FGFR-1 could be enhanced by the metalloprotease domain of ADAM-12, a protein important for normal muscle differentiation. The substrate for ADAM-12 was previously unknown. We showed that FGFR-1 ectodomain shedding is inhibited by a dominant negative ADAM-12 construct containing a mutation in the zinc-binding site of the catalytic domain. We also found that the release of FGFR-1 ectodomains leads to downregulation of the full-length FGF receptor on the cell surface.

These findings support the proposal that the critical roles of ADAM-12 in muscle differentiation are to induce the downregulation of FGFR-1 and to release soluble inhibitors of FGF that sequester FGF-2 and inhibit proliferation of myoblasts. These results provide the first evidence that FGFR-1 is a substrate for ADAM-12 and are consistent with the paradigm that shedding of FGFR-1 ectodomains inhibits the biological activity of FGF-2 during muscle cell differentiation.

PUBLICATIONS

Hanneken, A. Structural characterization of the soluble FGF receptors reveals multiple isoforms generated by secretion and ectodomain shedding. FEBS Lett. 489:176, 2001.