Glucose removal from N-linked oligosaccharides is required for efficient maturation of certain secretory glycoproteins from the rough endoplasmic reticulum to the Golgi complex.

Abstract

ABSTRACT1-Deoxynojirimycin is a specific inhibitor of glucosidases I and II, the first enzymes that process N-linked oligosaccharides after their transfer to polypeptides in the rough endoplasmic reticulum . In a pulse-chase experiment, 1-deoxynojirimycin greatly reduced the rate of secretion of a1-antitrypsin and al -antichymotrypsin by human hepatoma HepG2 cells, but had marginal effects on secretion of the glycoproteins C3 and transferrin, or of albumin . As judged by equilibrium gradient centrifugation, 1-deoxynojirimycin caused al-antitrypsin and al-antichymotrypsin to accumulate in the rough endoplasmic reticulum . The oligosaccharides on cell-associated al-antitrypsin and al-antichymotrypsin synthesized in the presence of 1-deoxynojirimycin, remained sensitive to Endoglycosidase H and most likely had the structure Glu 1 -3 Man 9 GIcNAC 2 . Tunicamycin, an antibiotic that inhibits addition of N-linked oligosaccharide units to glycoproteins, had a similar differential effect on secretion of these proteins . Swainsonine, an inhibitor of the Golgi enzyme a-mannosidase II, had no effect on the rates of protein secretion, although the proteins were in this case secreted with an abnormal N-linked, partially complex, oligosaccharide . We conclude that the movement of al-antitrypsin and al-antichymotrypsin from the rough endoplasmic reticulum to the Golgi requires that the N-linked oligosaccharides be processed to at least the Man9 GlcNAc 2 form; possibly this oligosaccharide forms part of the recognition site of a transport receptor for certain secretory proteins .Secretory proteins and cell surface glycoproteins are synthesized on ribosomes attached to the rough endoplasmic reticulum membrane . Cotranslationally, the polypeptides are transported across or inserted into the rough endoplasmic reticulum membrane (1-3), and one or more oligosaccharides, of structure G1c 3 Man9GlcNAc2 , are transferred to specific asn residues (4) . By a series of membrane vesicles these proteins are transported first to the Golgi complex of membranes, then to the cell surface (1-3, 5, 6).How proteins move from the rough endoplasmic reticulum to the Golgi is not known . A common hypothesis is that small vesicles bud off from a specialized region of the rough endoplasmic reticulum, containing a sample of the luminal contents and membrane proteins. These vesicles subsequently fuse with vesicles of the cis-(or proximal) region of the Golgi cisternae (2, 3). In hepatic parenchymal cells albumin and lipoprotein particles are localized to different terminal dilations of the rough endoplasmic reticulum and cis-Golgi (7) . Thus it is possible that different vesicles transport different secretory products to the Golgi .It is generally believed that all secretory and plasma membrane glycoprotein proteins migrate through the same Golgi vesicles. A combination of cell-fractionation and immunoelectronmicroscopic approaches has shown that albumin, transferrin, and the vesicular stomatitis viral glycoprotein and other glycoproteins are found in the...

Hepatitis B virus (HBV) envelope glycoproteins vary drastically in their sensitivity to glycan processing: Evidence that alteration of a single N-linked glycosylation site can regulate HBV secretion

“…Inhibition of early glycan processing events leads to the retention and misfolding of some glycoproteins within the ER (15). This may be the result of triglucosylated glycoproteins being unable to interact with ER chaperones such as calnexin, which only recognize monoglucosylated glycoproteins (16,17).…”

“…3, while the secretion of the L and S proteins within subviral particles was reduced by only 20-30%, the secreted subviral particles were virtually devoid of the M protein. That is, while subviral particles containing mostly the L and S glycoproteins are secreted near normally, as are many host glycoproteins from ␣-glucosidase-inhibited cells (15,19,20), there was a 16-fold reduction the secretion of Mcontaining subviral particles. Preliminary evidence suggest that the subviral particles containing the M protein carry hyperglucosylated glycan structures and are targeted to lysosomal compartments (14).…”

“…Inhibitors of ␣-glucosidase are known to have different effects on different glycoproteins (15,20). However, the differential effect of the ␣-glucosidase inhibitor on the three HBV envelope proteins (L, M, and S) proved quite surprising.…”

Secretion of high‐mannose‐type α1‐proteinase inhibitor and α1‐acid glycoprotein by primary cultures of rat hepatocytes in the presence of the mannosidase I inhibitor 1‐deoxymannojirimycin

“…Lodish and Kong [23] localized the unsecreted al-proteinase inhibitor within the rough endoplasmic reticulum. Inhibition of mannosidase I1 by swainsonine led to the formation of hybrid-type al-proteinase inhibitor without affecting its secretion [23,381. After these studies the question remained open whether al-proteinase inhibitor, carrying deglucosylated oligosaccharide side-chains of the high-mannose type, could be secreted at a normal rate.…”

“…In the presence of the glucosidase inhibitor I-deoxynojirimycin glucosylated high-mannosetype a,-proteinase inhibitor accumulated intracellularly [20]. Lodish and Kong [23] localized the unsecreted al-proteinase inhibitor within the rough endoplasmic reticulum. Inhibition of mannosidase I1 by swainsonine led to the formation of hybrid-type al-proteinase inhibitor without affecting its secretion [23,381.…”