Abstract
Platelet-derived growth factor (PDGF) is a dimeric protein that exerts its effects through tyrosine kinase alpha- and beta-receptors. The extracellular part of each receptor is composed of five Ig-like domains. Recombinant forms of alpha-receptor domains 1-4 (alphaRD1-4), 1-3 (alphaRD1-3), and 1 and 2 (alphaRD1-2) were prepared after expression in Chinese hamster ovary cells and were used to study the assembly of soluble ligand-receptor complexes. When incubated with micromolar concentrations of PDGF, both alphaRD1-3 and alphaRD1-4 formed complexes of 1:2 molar composition, i.e. one dimeric PDGF molecule bound two soluble receptors. alphaRD1-3, in contrast to alphaRD1-4, formed detectable 1:1 complexes under conditions of ligand excess. alphaRD1-4 displayed an increased ability to form 1:2 complexes as compared with alphaRD1-3 under conditions of limiting concentrations of ligand. We thus conclude that Ig-like domain 4-mediated receptor-receptor interactions contribute to 1:2 PDGF.alphaRD1-4 complex formation. Since alphaRD1-4 and alphaRD1-3 were equipotent in blocking binding of subnanomolar concentrations of PDGF to cell-surface receptors, we also conclude that this effect is predominantly achieved through formation of Ig-like domain 4-independent 1:1 ligand-receptor complexes. Finally, since alphaRD1-2 bound PDGF-BB with high affinity, whereas PDGF-AA was bound only with low affinity, we conclude that Ig-like domain 3 of the PDGF alpha-receptor contains epitopes of particular importance for PDGF-AA binding and that most of the PDGF-BB-binding epitopes reside in Ig-like domains 1 and 2.
Highlights
Platelet-derived growth factors (PDGFs)1 are a family of disulfide-bonded dimeric isoforms of A- and B-chains with potent mitogenic activity on connective tissue cells, glia cells, and endothelial cells
We show that this property is a consequence of Ig-like domain 4-mediated receptor-receptor interactions. We show that these receptor-receptor interactions do not contribute significantly to the inhibitory effect of soluble receptors on binding of subnanomolar concentrations of PDGF to cell-surface receptors
␣RD1–2 Binds PDGF-BB, but Not PDGF-AA—To further localize the region(s) within ␣RD1–3 that mediates ligand binding, we investigated the properties of ␣RD1–2 in complex forming assays and in cell-surface receptor binding inhibition assays (Figs. 8 and 9)
Summary
Platelet-derived growth factors (PDGFs) are a family of disulfide-bonded dimeric isoforms of A- and B-chains with potent mitogenic activity on connective tissue cells, glia cells, and endothelial cells (reviewed in Ref. 1). To further study the structural basis for PDGF-induced receptor dimerization, the properties of CHO cell-derived recombinant proteins consisting of PDGF ␣-receptor Ig-like domains 1– 4 (␣RD1– 4), 1–3 (␣RD1–3), and 1 and 2 (␣RD1–2) were compared. Using these receptor fragments, we demonstrate that ␣RD1– 4 at micromolar concentrations, in contrast to ␣RD1–3, forms a 1:2 ligand-receptor complex under conditions of ligand excess. We demonstrate that ␣RD1– 4 at micromolar concentrations, in contrast to ␣RD1–3, forms a 1:2 ligand-receptor complex under conditions of ligand excess We show that this property is a consequence of Ig-like domain 4-mediated receptor-receptor interactions. Characterization of the PDGF binding properties of ␣RD1–2 demonstrates that Ig-like domain 3 of the PDGF ␣-receptor contains epitopes of particular importance for PDGF-AA binding and that most of the PDGF-BB-binding epitopes are localized within the ␣RD1–2 fragment
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
