Distinct structural elements in GDNF mediate binding to GFRalpha1 and activation of the GFRalpha1-c-Ret receptor complex.
Ligand-induced receptor oligomerization is a widely accepted mechanism for activation of cell-surface receptors. We investigated ligand-receptor interactions in the glial cell-line derived neurotrophic factor (GDNF) receptor complex, formed by the c-Ret receptor tyrosine kinase and the glycosylphosphatidylinositol (GPI)-anchored subunit GDNF family receptor alpha-1 (GFRalpha1). As only GFRalpha1 can bind GDNF directly, receptor complex formation is thought to be initiated by GDNF binding to this receptor. Here we identify an interface in GDNF formed by exposed acidic and hydrophobic residues that is critical for binding to GFRalpha1. Unexpectedly, several GDNF mutants deficient in GFRalpha1 binding retained the ability to bind and activate c-Ret at normal levels. Although impaired in binding GFRalpha1 efficiently, these mutants still required GFRalpha1 for c-Ret activation. These findings support a role for c-Ret in ligand binding and indicate that GDNF does not initiate receptor complex formation, but rather interacts with a pre-assembled GFRalpha1- c-Ret complex.
Eketjäll
S
S
Division of Molecular Neurobiology, Department of Neuroscience, Karolinska Institute, 17177 Stockholm, Sweden.
Fainzilber
M
M
Murray-Rust
J
J
Ibáñez
C F
CF
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D003432:Cross-Linking Reagents; D019281:Dimerization; D029721:Drosophila Proteins; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D051379:Mice; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D029721:Drosophila Proteins; C495040:Gdnf protein, mouse; C494988:Gfra1 protein, mouse; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; D011518:Proto-Oncogene Proteins; D019000:Phosphotyrosine; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases; C416649:Ret protein, Drosophila; C494999:Ret protein, mouse