Signaling Domains of the βc Chain of the GM‐CSF/IL‐3/IL‐5 Receptor

A bstract : The granulocyte/macrophage colony‐stimulating factor (GM‐CSF)/interleukin‐3 (IL‐3)/IL‐5 receptors are a family of heterodimeric transmembrane proteins expressed by myeloid lineage cells. Each receptor has a unique ligand‐binding α chain and they share a common β chain (βc chain). Binding of GM‐CSF activates at least one receptor‐associated tyrosine kinase, JAK2, and rapidly induces tyrosine phosphorylation of the GMR βc chain (GMRβ), but not the GMR αchain (GMRα). Mutation of each of the 8 tyrosine residues in the cytoplasmic domain of the human GMRβ to phenylalanine (GMRβF 8 ) reduced tyrosine phosphorylation of GMRβ, SHP2 and SHC, but not JAK2 or STAT5. Interestingly, GMRβ‐F 8 was still capable of inducing at least short‐term proliferation and enhancing viability. The role of each individual tyrosine residue was explored by replacing each mutated phenylalanine with the wild‐type tyrosine residue. Tyrosine 577 was found to be sufficient to regenerate GM‐CSF‐dependent phosphorylation of SHC, and any of Y577, Y612, or Y695 were sufficient to regenerate GM‐CSF‐inducible phosphorylation of SHP2. Next, a series of four internal deletion mutants were generated, which deleted small sections from aa 518 to 626. One of these, deleting residues 566‐589 was profoundly defective in signaling and supporting viability, and may identify an important viability signaling domain for this receptor family. Overall, these results indicate that GMRβ tyrosine residues are not necessary for activation of the JAK/STAT pathway, or for proliferation, viability, or adhesion signaling in Ba/F3 cells, although tyrosine residues significantly affect the magnitude of the response. However, internal deletion mutant studies identify critical domains for viability and proliferation.