Transport of free polymannose-type oligosaccharides from the endoplasmic reticulum into the cytosol is inhibited by mannosides and requires a thapsigargin-sensitive calcium store

• Published in: Glycobiology (Oxford) 1998-04, Vol.8 (4), p.373-381
• Main Author: Moore, S E
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - metabolism; Biological Transport, Active - drug effects; calcium; Calcium - metabolism; Calcium-Transporting ATPases - antagonists & inhibitors; Carbohydrate Sequence; Cell Line; Cytosol - metabolism; endoplasmic reticulum; Endoplasmic Reticulum - metabolism; Enzyme Inhibitors - pharmacology; free oligosaccharides; Humans; lectin; Mannans - chemistry; Mannans - metabolism; Mannosides - chemistry; Mannosides - pharmacology; Molecular Sequence Data; Oligosaccharides - chemistry; Oligosaccharides - metabolism; Structure-Activity Relationship; Thapsigargin - pharmacology; transport
• ISSN: 0959-6658; 1460-2423
• DOI: 10.1093/glycob/8.4.373

The transport of free polymannose-type oligosaccharides from the lumen of the endoplasmic reticulum into the cytosol has been recently demonstrated (Moore,S.E.H., et al., 1995, EMBO J., 14, 6034–6042), but at present little is known of the characteristics of this process. Here, it is shown that inhibition of the transport of endogenously synthesized metabolically radiolabeled free oligosaccharides out of the endoplasmic reticulum into the cytosol of permeabilized HepG2 cells occurs when assays are conducted in the presence of mannose (IC50, 4.9 mM), or its derivatives modified at the first carbon (C1) of the sugar ring; α-methyl mannoside (IC50, 2.0 mM), mannoheptulose (IC50, 1.6 mM), and α-benzyl mannoside (IC50, 0.8 mM), whereas other monosaccharides (50 mM), differing from mannose at position; C2 (glucose), C3 (altrose), C4 (talose), C5 (L-rhamnose), and C6 (mannoheptose), have little effect. N-Acetylglucosamine does not inhibit oligosaccharide transport and, furthermore, although mannobioses and a mannotriose inhibit free oligosaccharide transport, di-N-acetylchitobiose is without effect. It is also shown that if the transport assay buffer is either depleted of calcium ions, or supplemented with the Ca2+/Mg2+ ATPase inhibitor, thapsigargin, or with calcium ionophores, free oligosaccharide transport out of the endoplasmic reticulum is inhibited. These results demonstrate that the terminal nonreducing mannosyl residues of free polymannose-type oligosaccharides and not their N-acetylglucosamine-containing reducing termini, play an important role in the interaction of the free oligosaccharide with the transport machinery, and that this transport process requires the presence of calcium sequestered in the lumen of the endoplasmic reticulum.