Detection of a LAG phase in gonadotropin-releasing hormone stimulated synthesis of lutropin peptide chains in cultured rat anterior pituitary cells

Abstract

We have studied the time course (0-5h) of the stimulatory effect of the hypothalamic gonadotropin-releasing hormone (GnRH) on the biosynthesis of lutropin (LH) polypeptide chains, as measured by the incorporation of [35S] methionine into proteins synthesized in cultured rat anterior pituitary cells in the absence or presence of 10nM GnRH. Labeled polypeptides, immunologically related to LH subunits alpha and beta, were isolated by specific immunoprecipitation, analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, then revealed by fluorography and quantified by counting the excised bands. This methodology allowed us to detect the radioactivity incorporated into LH subunits after less than 15 min of incubation. During first 1h of the time-course the quantity of [35S]Met incorporated into both alpha and LH beta subunits was not increased by the presence of GnRH in the incubation medium. A significant increase in the incorporation of radioactivity into LH subunits was observed after 2h of GnRH treatment. However, the increase in LH release into the medium in response to GnRH, as measured by RIA, was immediate. These data demonstrate that GnRH-stimulated synthesis of LH polypeptide chains occurs after a lag of approximately 1h and involves mechanisms different from those governing the stimulation of LH release.