GnRH-stimulated glycosylation (proximal and distal) of luteinizing hormone by cultured rat pituitary cells.

Abstract

In the present work, the effects of GnRH on the translation (by [14C]leucine incorporation; [14C]Leu-LH) and the glycosylation of LH by rat pituitary cells in primary culture were established. The use of specific markers as radioactive precursors made it possible to discriminate the action of the neurohormone on proximal glycosylation (by[3H]mannose incorporation; [3H]Man-LH) as well as distal glycosylation (by [3H]galactose incorporation; [3H]Gal-LH) in the course of synthesis and release of LH. Pituitary cells from ovariectomized adult rats were incubated for different periods between 0 and 5 h in medium containing [14C]Leu plus [3H]Man or [14C]Leu plus [3H]Gal with or without 10 nM GnRH. GnRH increased synthesis and release of newly synthesized LH. The magnitude of the stimulatory effect on the kinetics of [14C]Leu (slope = 63.58; 158% of control) and [3H]Man (slope = 75.15; 161%) incorporation to LH was similar. The action of the neurohormone appears to be exerted on translation, the increased [3H]Man incorporation being a secondary phenomenon arising from the greater amount of available polypeptide chains as acceptors of the polymannose core. However, a direct effect of GnRH on proximal glycosylation cannot be excluded. GnRH also stimulated the kinetics of release of [14C]Leu-LH (slope = 6.14; 236% of control) and [3H]Man-LH (slope = 8.06; 191%). Comparatively, the effect of GnRH on [3H]Gal-LH was detected earlier than that on LH labeled with the other precursors; increases in rates of production (slope = 71.57; 278% of control) and release (slope = 32.08; 494%) were higher than those in [14C]Leu- and [3H]Man-LH kinetics, indicating that GnRH acts specifically on this distal step of LH glycosylation. GnRH enhanced the relative terminal glycosylation ([3H]Gal/[14C]Leu ratio) of total and release LH without modifying the relative proximal glycosylation ([3H]Man/[14C]Leu ration) of the hormone. We conclude that GnRH can induce not only changes in the quantity (greater number of molecules) but also in the quality (molecules more glycosylated) of the secreted LH by acting directly at translation and distal glycosylation level.