A new family of neuronal survival factors comprised of glial cell line- derived neurotrophic factor (GDNF) and neurturin has recently been described (Kotzbauer, P. T., Lampe, P. A., Heuckeroth, R. O., Golden, J.P., Creedon, D. J., Johnson, E. M., Jr., and Milbrandt, J. (1997) Nature 384, 467-470). These molecules, which are related to transforming growth factor-β, are important in embryogenesis and in the survival of distinct neuronal populations. These molecules signal through a novel receptor system that includes the Ret receptor tyrosine kinase, a ligand (i.e. GDNF or neurturin), and an accessory glycosyl-phosphatidylinositol-linked molecule that is responsible for high affinity binding of the ligand. Two accessory molecules denoted GDNF family receptor 1 and 2 (GFRα-1 and GFRα-2) have been described that function in GDNF and neurturin signaling complexes. We have identified a novel co- receptor belonging to this family based on similarity to GFRα-1, which we have named GFRα-3. GFRα-3 displays 33% amino acid identity with GFRα-1 and 36% identity with GFRα-2. Despite the similarity of GFRα-3 to GFRα-1 and GFRα-2, it is unable to activate Pet in conjunction with GDNF, suggesting that there are likely additional undiscovered ligands and/or Ret-like receptors to be identified. GFRα-3 is anchored to the cell membrane by a phosphatidylinositol-specific phospholipase C-resistant glycosyl- phosphatidylinositol linkage. GFRα-3 is highly expressed by embryonic day 11 but is not appreciably expressed in the adult mouse. In situ hybridization analyses demonstrate that GFRα-3 is located in dorsal root ganglia and the superior cervical sympathetic ganglion. Comparison of the expression patterns of GFRα-3 and Ret suggests that these molecules could form a receptor pair and interact with GDNF family members to play unique roles in development.