Differential carbohydrate processing and secretion of thyrotropin and free alpha subunit. Effects of 1-deoxynojirimycin.

In pulse-chase experiments we compared the kinetics of early carbohydrate processing and subsequent secretion of thyroid-stimulating hormone (TSH) and free alpha subunit under control conditions and after treatment with 1-deoxynojirimycin, an inhibitor of glucosidases I and II. Under control conditions TSH achieved resistance to endo-beta-N-acetylglucosaminidase H (endo H) more rapidly than free alpha (t1/2 0.3 h versus 0.9 h); however, free alpha was secreted more rapidly than TSH (t1/2 2.2 h versus 3.4 h). With 1-deoxynojirimycin, oligosaccharides co-migrating with G3Man9GlcNAc and G2Man9GlcNAc were demonstrated on TSH for the first time, suggesting that previous pulse-chase studies did not disclose these intermediates due to rapid removal of glucose residues from the common G3Man9GlcNAc2 precursor. 1-Deoxynojirimycin delayed the rate of attainment of endo H resistance for both TSH and free alpha, but there was no effect on subunit combination. With 5 mM 1-deoxynojirimycin the amount of secreted free alpha was reduced to 65% of control; secreted TSH was reduced markedly to 17% of control without intracellular accumulation, suggesting increased intracellular degradation. There was no significant toxicity from these doses of 1-deoxynojirimycin on the production or secretion of the two major nonglycosylated pituitary proteins, growth hormone and prolactin, or on at least 10 other secretory proteins. Basal differences in the relative rates of TSH and free alpha processing and secretion as well as differential sensitivity to 1-deoxynojirimycin suggest separate secretory pathways for these two closely related proteins.