Interferon-γ induces receptor dimerization in solution and on cells

The extracellular domain (ECD) of the human interferon-γ (IFNγ) receptor was stably expressed in Chinese hamster ovary cells and purified to homogeneity. Scatchard analysis of ¹²⁵I-IFNγ binding to ECD preparations revealed the formation of a ligand-receptor complex which displayed a K_a of 6.4 ± 0.9 × 10⁸ M^-1. Two types of complexes were identified by sucrose density gradient ultracentrifugation. The stoichiometry of the major ECD-ligand complex was determined by high performance liquid chromatography gel filtration. When IFNγ was incubated with a 2-fold molar excess of ECD, a 190-kDa complex was isolated that contained 2 mol of ECD per 1 mol of IFNγ. IFNγ also induced dimerization of IFNγ receptors expressed at the cell surface as detected by chemically cross-linking receptor bound ligand and analyzing cell lysates by SDS-polyacrylamide gel electrophoresis and immunoblotting. Finally, labeled forms of ECD bound to cells preincubated at 4°C with excess amounts of IFNγ indicating that the ligand could associate with more than one receptor molecule in the absence of chemical cross-linking agents. These results demonstrate that IFNγ effects dimerization of its receptor under physiologic conditions and suggest that IFNγ receptor dimerization may be an important event in inducing IFNγ-dependent biologic responses.