GNRH-STIMULATED GLYCOSYLATION (PROXIMAL AND DISTAL) OF LUTEINIZING-HORMONE BY CULTURED RAT PITUITARY-CELLS

In the present work, the effects of GnRH on the translation (by [C-14] leucine incorporation; [C-14]Leu-LH) and the glycosylation of LH by ra t pituitary cells in primary culture were established, The use of spec ific markers as radioactive precursors made it possible to discriminat e the action of the neurohormone on proximal glycosylation (by [H-3]ma nnose incorporation; [H-3]Man-LH) as well as distal glycosylation (by [H-3]galactose incorporation; [H-3]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 containin g [C-14]Leu plus [H-3]Man or [C-14]Leu plus [H-3]Gal with or without 1 0 nM GnRH. GnRH increased synthesis and release of newly synthesized L H. The magnitude of the stimulatory effect on the kinetics of [C-14]Le u (slope =63.58; 158% of control) and [H-3]Man (slope=75.15; 161%) inc orporation to LH was similar, The action of the neurohormone appears t o be exerted on translation, the increased [H-3]Man incorporation bein g a secondary phenomenon arising from the greater amount of available polypeptide chains as accep ters of the polymannose core, However, a d irect effect of GnRH on proximal glycosylation cannot be excluded, GnR H also stimulated the kinetics of release of [C-14]Leu-LH (slope=6.14; 236% of control) and [H-3]Man-LH (slope=8.06; 191%), Comparatively, t he effect of GnRH on [H-3]Gal-LH was detected earlier than that on LH labeled with the other precursors; increases in rates of production (s lope=71.57; 278% of control) and release (slope=32.08; 494%) were high er than those in [C-14]Leu- and [H-3]Man-LH kinetics, indicating that GnRH acts specifically on this distal step of LH glycosylation. GnRH e nhanced the relative terminal glycosylation ([H-3]Gal/[C-14]Leu ratio) of total and released LH without modifying the relative proximal glyc osylation ([H-3]Man/[C-14]Leu ratio) of the hormone. We conclude that GnRH can induce not only changes in the quantity (greater number of mo lecules) but also in the quality (molecules more glycosylated) of the secreted LH by acting directly at translation and distal glycosylation level.