Translocation of cytidine 5'-monophosphosialic acid across Golgi apparatus membranes.
Golgi apparatus, isolated from rat liver, incorporate [14C]sialic acid from CMP[14C]sialic acid into endogenous glycolipid and glycoprotein acceptors. Incorporation of [14C]sialic acid into endogenous glycoprotein acceptors was stimulated an average of 3-fold by Triton X-100 at an optimal concentration of 0.05% and was inhibited at higher concentrations. Incorporation of [14C]sialic acid into endogenous glycolipid acceptors was not stimulated by detergent. The major glycolipid product was identified by thin-layer chromatography as the ganglioside GD3. SDS-polyacrylamide gel electrophoresis on the glycoprotein products demonstrated incorporation of [14C]sialic acid into 6--7 major bands. Neuraminidase studies determined that approximately 60% of the [14C]sialic acid incorporated into endogenous acceptors in the absence of detergent had a luminal orientation. Furthermore, electron microscopy studies showed that the isolated Golgi apparatus fraction consisted of intact membrane cisternae. Our results demonstrate that sialylation of cisternal acceptors located on the inside of the membrane occurs in the absence of detergent. They are consistent with carrier-mediated transport as a mechanism to allow CMPsialic acid to traverse the Golgi apparatus membrane and to be used to glycosylate endogenous glycoprotein and glycolipid acceptors.
D000818:Animals; D001692:Biological Transport; D002250:Carbon Radioisotopes; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid / Q000378:metabolism*; D003902:Detergents / Q000494:pharmacology; D006023:Glycoproteins / Q000096:biosynthesis; D006056:Golgi Apparatus / Q000378:metabolism* / Q000648:ultrastructure; D007425:Intracellular Membranes / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008854:Microscopy, Electron; D008970:Molecular Weight; D009439:Neuraminidase; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D051381:Rats; D012794:Sialic Acids / Q000378:metabolism*; D012799:Sialyltransferases / Q000378:metabolism; D014531:Uridine Diphosphate Galactose / Q000378:metabolism
D002250:Carbon Radioisotopes; D003902:Detergents; D006023:Glycoproteins; D012794:Sialic Acids; D014531:Uridine Diphosphate Galactose; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid; D011092:Polyethylene Glycols; D017830:Octoxynol; D012799:Sialyltransferases; D009439:Neuraminidase