Reconstitution of fibroblast growth factor receptor interactions in the yeast two hybrid system.
Fibroblast growth factors (FGF) activate their receptors through the formation of trimolecular complexes, composed of a ligand, a receptor, and a heparan sulfate oligosaccharide, all of which are members of particularly large families capable of multiple interactions in a combinatorial fashion. Understanding this large network of interactions not only presents a great challenge, but is practically beyond the capacity of most classical techniques routinely used to study ligand receptor interactions. We have used the yeast two hybrid system to study protein-protein interactions in the FGF family. Both ligand and receptor ectodomains are properly folded and functional in the yeast. Basic FGF (bFGF) expressed in the yeast dimerizes spontaneously. This self-assembly occurs at low affinity, which can be greatly enhanced by the introduction of heparin, supporting a defined role for heparin in bFGF dimerization. Screening a rat embryo cDNA library with bFGF in the yeast two hybrid system identified a short variant of FGF receptor 1, found most frequently in embryonal and tumor cells and which possesses affinity toward bFGF that is significantly greater than that of the more abundant, full-length receptor. We find the yeast two hybrid system, a most suitable alternative method for the analysis of growth factor-receptor interactions as well as for screening for novel interacting proteins and modulators of FGF and its receptors.
Aloni-Grinstein
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Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel.
Seddon
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Yayon
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D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D019281:Dimerization; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology; D051381:Rats; D017468:Receptors, Fibroblast Growth Factor / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D017468:Receptors, Fibroblast Growth Factor; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D016222:Fibroblast Growth Factor 2; D006493:Heparin