Human GH (hGH) is believed to elicit its signal by promoting dimerization of the hGH receptor (hGHR). In this study, we examined a covalent linkage of receptors induced by hGH treatment of IM-9 cells. hGH induced a time- and concentration-dependent appearance of a disulfide-linked species of 215-230 kilodaltons, designated p215-230, that at 37 C was long-lived (> 1 h). p215-230 was confirmed to contain the hGHR (115-140 kilodaltons) as at least one of its constituents by two-dimensional diagonal sodium dodecyl sulfate-polyacrylamide gel electrophoresis. hGH induction of p215-230 required intact cells and was inhibitable by pretreatment of cells with N-ethylmaleimide (NEM), a sulfhydryl-reactive alkylating agent. NEM pretreatment did not, however, prevent hGH-dependent formation of a nondisulfide-linked p215-230 form, which was detected in NEM-pretreated hGH-stimulated cells by chemical cross-linking of detergent cell extracts. The disulfide-linked form of the hGHR accounted for a substantial fraction of the receptors that became tyrosine phosphorylated early into hGH treatment. However, formation of the disulfide-linked hGHR was not blocked by attenuation of tyrosine kinase activation, in that pretreatment of cells with staurosporine (1.25 μM) prevented detectable hGH-induced tyrosine phosphorylation without preventing the appearance of p215-230. These findings indicate that hGH induces its receptor to form a noncovalently associated complex, which then undergoes a rapid transition to a disulfide-linked form. These processes may have relevance to hGH signaling and/or hGHR trafficking. © 1994 by The Endocrine Society.