Human interleukin-3 (IL-3) induces disulfide-linked IL-3 receptor α- and β-chain heterodimerization, which is required for receptor activation but not high-affinity binding

The human interleukin-3 receptor (IL-3R) is a heterodimer that comprises an IL-3-specific α chain (IL-3Rα) and a common β chain (β_c) that is shared with the receptors for granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-5. These receptors belong to the cytokine receptor superfamily, but they are structurally and functionally more related to each other and thus make up a distinct subfamily. Although activation of the normal receptor occurs only in the presence of ligand, the underlying mechanisms are not known. We show here that human IL-3 induces heterodimerization of IL-3Rα and β_c and that disulfide linkage of these chains is involved in receptor activation but not high-affinity binding. Monoclonal antibodies (MAb) to IL-3Rα and β_c were developed which immunoprecipitated, in the absence of IL-3, the respective chains from cells labelled with ¹²⁵I on the cell surface. However, in the presence of IL-3, each MAb immunoprecipitated both IL-3Rα and β_c. IL-3-induced receptor dimers were disulfide and nondisulfide linked and were dependent on IL-3 interacting with both IL-3Rα and β_c. In the presence of IL-3 and under nonreducing conditions, MAb to either IL-3 Rα or β_c immunoprecipitated complexes with apparent molecular weights of 215,000 and 245,000 and IL-3Rct and β_c monomers. Preincubation with iodoacetamide prevented the formation of the two high-molecular-weight complexes without affecting noncovalent dimer formation or high-affinity IL-3 binding. Two-dimensional gel electrophoresis and Western blotting (immunoblotting) demonstrated the presence of both IL-3Rα and β_c in the disulfide-linked complexes. IL-3 could also be coimmunoprecipitated with anti-IL-3Rα or anti-β_c MAb, but it was not covalently attached to the receptor. Following IL-3 stimulation, only the disulfide-linked heterodimers exhibited reactivity with antiphosphotyrosine antibodies, with β_c but not IL-3Rα being the phosphorylated species. A model of IL-3R activation is proposed which may be also applicable to the related GM-CSF and IL-5 receptors.