Cooperative Dimerization of Fibroblast Growth Factor 1 (FGF1) upon a Single Heparin Saccharide May Drive the Formation of 2:2:1 FGF1·FGFR2c·Heparin Ternary Complexes

Christopher J Robinson,Nicholas J Harmer,Sarah J Goodger,Tom L Blundell,John T Gallagher

doi:10.1074/jbc.m505720200

Christopher J Robinson, Nicholas J Harmer + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m505720200

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Abstract

The related glycosaminoglycans heparin and heparan sulfate are essential for the activity of the fibroblast growth factor (FGF) family as they form an integral part of the signaling complex at the cell surface. Using size-exclusion chromatography we have studied the capacities of a variety of heparin oligosaccharides to bind FGF1 and FGFR2c both separately and together in ternary complexes. In the absence of heparin, FGF1 had no detectable affinity for FGFR2c. However, 2:2:1 complexes formed spontaneously in solution between FGF1, FGFR2c, and heparin octasaccharide (dp8). The dp8 sample was the shortest chain length that bound FGFR2c, that dimerized FGF1, and that promoted a strong mitogenic response to FGF1 through FGFR2c. Heparin hexasaccharide and various selectively desulfated heparin dp12s failed to bind FGFR2c and could only interact with FGF1 monomerically. These saccharides formed 1:1:1 complexes with FGF1 and FGFR2c, which had no tendency to self-associate, suggesting that binding of two FGF1 molecules to the same saccharide chain is a prerequisite for subsequent FGFR2c dimerization. We found that FGF1 dimerization upon heparin was favored over monomeric interactions even when a large excess of saccharide was present. A cooperative mechanism of FGF1 dimerization could explain how 2:2:1 signaling complexes form at the cell surface, an environment rich in heparan sulfate.

Highlights

The FGFRs share a number of common structural features [5]
The fibroblast growth factor (FGF) are heparin-binding proteins, and it is appreciated that signal transduction requires the formation of a ternary complex at the cell surface formed between FGF, FGFR, and heparan sulfate (HS)
The heparin-bridged 2:2:1 complex [27] could form internally upon a HS chain but, because the two FGF1⁄7FGFR half-complexes do not interact directly, they are reliant on the probability that they become aligned upon opposite faces of the same short stretch of HS

Summary

Introduction

The FGFRs share a number of common structural features [5]. All have an intracellular split tyrosine kinase domain, a transmembrane sequence, and an extracellular region composed of three immunoglobulin (Ig)-like domains (D1–D3). The FGFs are heparin-binding proteins, and it is appreciated that signal transduction requires the formation of a ternary complex at the cell surface formed between FGF, FGFR, and heparan sulfate (HS). When run on a Superdex 200 column calibrated with protein standards, the dp12 sample had an apparent molecular mass of 22 kDa. Data on these and all subsequent major observed peaks are presented in TABLES ONE and TWO.

Results

Conclusion