Intracellular compartmentalization and degradation of free polymannose oligosaccharides released during glycoprotein biosynthesis.
The intracellular site for the degradation of free polymannose oligosaccharides released during glycoprotein biosynthesis has been studied by permeabilizing the plasma membrane of metabolically radiolabeled HepG2 cells with streptolysin O. This pore-forming agent permitted us to examine the breakdown in both the cytosolic and vesicular compartments of the previously recognized (Anumula, K. R., and Spiro, R. G. (1983) J. Biol. Chem. 258, 15274-15282) polymannose components terminating in a di-N-acetylchitobiose sequence (OS-Glc-NAc2) or a single N-acetylglucosamine residue (OS-Glc-NAc1) residue. Pulse-chase studies indicated that although the OS-GlcNAc2 saccharides were about equally distributed between vesicles and cytosol and rapidly disappeared after reaching the Man8 stage, the OS-GlcNAc1 species were found predominantly in the extravesicular compartment and there underwent a distinctive demannosylation sequence resulting in the formation of a Man5GlcNAc isomer (Man alpha 1-->2Man alpha 1-->2Man alpha 1-->3(Man alpha 1-->6)Man beta 1-->4GlcNAc) which was different from the product of Golgi processing enzymes. Further trimming of this cytosolic limit product required its translocation into a vesicular compartment, believed to be lysosomes, in which Man2-4GlcNAc components appeared as the metabolic chase progressed. The accumulation of Glc1Man5GlcNAc in the cytosol during the chase suggested that glucose interferes with the cytosolic-vesicular transfer and this became even more evident by the pronounced pile-up of extravesicular Glc3Man5GlcNAc when the cells were incubated in the presence of castanospermine. Although the biological significance and mechanism of free polymannose oligosaccharide entry into the cytosol is not yet known, the possibility that it may reflect an endoplasmic reticulum-situated degradative process of glycoproteins merits consideration.
Moore
S E
SE
Department of Biological Chemistry, Harvard Medical School, Boston, Massachusetts 02215.
Spiro
R G
RG
D001426:Bacterial Proteins; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002451:Cell Compartmentation; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D003600:Cytosol / Q000378:metabolism; D006023:Glycoproteins / Q000096:biosynthesis*; D006801:Humans; D007700:Kinetics; D008358:Mannose / Q000378:metabolism*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D013301:Streptolysins
D001426:Bacterial Proteins; D006023:Glycoproteins; D009844:Oligosaccharides; D013301:Streptolysins; C025964:streptolysin O; D008358:Mannose