Transport of free polymannose-type oligosaccharides from the endoplasmic reticulum into the cytosol is inhibited by mannosides and requires a thapsigargin-sensitive calcium store |
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Authors: | Moore SE |
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Institution: | Unite de Neuroendocrinologie et Cellulaire Digestives, Institut National de la Sante et de la Recherche Medicale, U410, Faculte de Medecine Xavier Bichat, 16 Rue Henri Huchard, 75018 Paris, France. |
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Abstract: | 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; alpha-methyl mannoside (IC50, 2.0
mM), mannoheptulose (IC50, 1.6 mM), and alpha-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.
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