Growth hormone (GH) plays a significant role in normal growth and development. Signaling to the cell is believed to require growth hormone receptor (GHR) dimerization, which occurs following binding of a single growth hormone molecule to each of two receptors. We have developed human growth hormone receptor-specific monoclonal antibodies, one of which was used here to characterize hormone/receptor interactions. This antibody, GHR05, is directed against the hinge spanning subdomains I and II of the receptor's extracellular region. Antibody binding to the cell surface receptor increases upon receptor binding to growth hormone, but not when it binds a mutant form, hGHG120R, which does not trigger receptor activation. Growth hormone binding thus appears to lead to a conformational change in the receptor epitope recognized by GHR05, giving rise to the active dimer configuration, necessary for signal transduction. Using a chimeric receptor-expressing, growth hormone-dependent murine cell line, we find that GHR05 binds to the receptor in the absence of human GH and delivers a signal leading to cell proliferation. Finally, GHR05 treatment of IM-9 cells, a human cell line expressing a functional human GHR, leads to cell proliferation mediated by the generation of GHspecific signals, including phosphorylation of the JAK2 tyrosine kinase and activation of STAT5.The 22-kDa polypeptide human growth hormone (hGH), 1 essential for normal growth and development, induces a variety of biological effects including linear growth, lactation, nitrogen retention, diabetogenic and insulin-like effects, and macrophage activation (1-4). Each of these effects is initiated by hGH interaction with specific cell receptors. The hGH receptor (hGHR) belongs to a large cytokine receptor family (5), which includes receptors for prolactin, erythropoietin, interleukins (IL)-2, -4, and -6, granulocyte/macrophage colony-stimulating factor, and granulocyte colony-stimulating factor (G-CSF) (6 -10). The three-domain organization of these receptors comprises a heavily N-glycosylated extracellular ligand binding domain, a single transmembrane segment and an intracellular domain, the last of which shares little sequence identity within the family (11). The hGHR extracellular region has two subdomains, one implicated mainly in hGH binding (subdomain I) and the other in hGHR dimerization (subdomain II) (7,8).The extracellular hGHR domain is found in serum in the form of a hormone binding protein (hGHBP), which binds hGH with approximately the same affinity and specificity as the intact receptor (12). Nonglycosylated recombinant bacterial hGHBP (13) has the same binding affinity (K d ϭ 0.4 nM) (14) and specificity for hGH as the mammalian binding protein.Crystallization studies of hGH and hGHBP show that a single hGH molecule binds two hGHBP molecules (15). Each hGH molecule is bivalent, containing two separate hGHBP binding sites; site I is a high affinity site and site II, a low affinity site. In contrast, the hGHBP is univalent, as it uses the same amino ...