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PMID: 1747103 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Determination of the intracellular sites and topology of glucosylceramide synthesis in rat liver.

The Biochemical journal ·Vol. 280 ( Pt 2) ·1991-12-01 ·Pages 295-302

Futerman AH, Pagano RE

Abstract

We examined the intracellular site(s) and topology of glucosylceramide (GlcCer) synthesis in subcellular fractions from rat liver, using radioactive and fluorescent ceramide analogues as precursors, and compared these results with those obtained in our recent study of sphingomyelin (SM) synthesis in rat liver [Futerman, Stieger, Hubbard & Pagano (1990) J. Biol. Chem. 265, 8650-8657]. In contrast with SM synthesis, which occurs principally at the cis/medial Golgi apparatus, GlcCer synthesis was more widely distributed, with substantial amounts of synthesis detected in a heavy (cis/medial) Golgi-apparatus subfraction, a light smooth-vesicle fraction that is almost devoid of an endoplasmic-reticulum marker enzyme (glucose-6-phosphatase), and a heavy vesicle fraction. Furthermore, no GlcCer synthesis was detected in an enriched plasma-membrane fraction after accounting for contamination by Golgi-apparatus membranes. These results suggest that a significant amount of GlcCer may be synthesized in a pre- or early Golgi-apparatus compartment. Unlike SM synthesis, which occurs at the luminal surface of the Golgi apparatus, GlcCer synthesis appeared to occur at the cytosolic surface of intracellular membranes, since (i) limited proteolytic digestion of intact Golgi-apparatus vesicles almost completely inhibited GlcCer synthesis, and (ii) the extent of UDP-glucose translocation into the Golgi apparatus was insufficient to account for the amount of GlcCer synthesis measured. These findings imply that, after its synthesis, GlcCer must undergo transbilayer movement to the luminal surface to account for the known topology of higher-order glycosphingolipids within the Golgi apparatus and plasma membrane.

MeSH Terms
Animals Cell Fractionation Glucosylceramides/biosynthesis Golgi Apparatus/metabolism Hydrogen-Ion Concentration Intracellular Membranes/metabolism Liver/drug effects,metabolism Magnesium/pharmacology Manganese/pharmacology Microscopy, Electron Microsomes, Liver/metabolism,ultrastructure Rats Sphingosine/analogs & derivatives,metabolism
Chemicals
Glucosylceramides hexanoyl-GlcCer Manganese Magnesium Sphingosine
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Futerman A H
Department of Embryology, Carnegie Institution of Washington, Baltimore, MD 21210-3301.
Pagano R E
References (43)
43 references, click to expand
  1. A rapid method of total lipid extraction and purification.
    Can J Biochem Physiol. 1959 Aug;37(8):911-7 PMID: 13671378
  2. The lysosomal localization of sphingolipid hydrolases.
    Biochim Biophys Acta. 1968 Apr 24;159(1):141-6 PMID: 5650428
  3. Glucosyceramide synthase of mouse kidney: further characterization with an improved assay method.
    Arch Biochem Biophys. 1990 Dec;283(2):372-8 PMID: 2148864
  4. Metabolism and intracellular transport of glycosphingolipids.
    Biochemistry. 1990 Dec 11;29(49):10865-71 PMID: 2271686
  5. Lipid traffic in eukaryotic cells: mechanisms for intracellular transport and organelle-specific enrichment of lipids.
    Curr Opin Cell Biol. 1990 Aug;2(4):652-63 PMID: 2252591
  6. Topology of glucosylceramide synthesis in Golgi membranes from porcine submaxillary glands.
    Biochim Biophys Acta. 1986 Jun 13;858(1):6-12 PMID: 2939881
  7. Lysogangliosides: synthesis and use in preparing labeled gangliosides.
    Methods Enzymol. 1987;138:319-41 PMID: 3600331
  8. Glycosyltransferases. Structure, localization, and control of cell type-specific glycosylation.
    J Biol Chem. 1989 Oct 25;264(30):17615-8 PMID: 2681181
  9. Two glycosphingolipid sialyltransferases are localized in different sub-Golgi compartments in rat liver.
    J Biol Chem. 1989 Sep 25;264(27):15766-9 PMID: 2777762
  10. UDPglucose-ceramide glucosyltransferase from porcine submaxillary glands is associated with the Golgi apparatus.
    Biochim Biophys Acta. 1985 Mar 28;814(1):1-7 PMID: 3156635
  11. Topography of glycosylation reactions in the rough endoplasmic reticulum membrane.
    J Biol Chem. 1986 May 25;261(15):6822-30 PMID: 2422171
  12. Free sphingosine formation from endogenous substrates by a liver plasma membrane system with a divalent cation dependence and a neutral pH optimum.
    J Biol Chem. 1989 Jun 25;264(18):10371-7 PMID: 2499578
  13. Modulation of ganglioside biosynthesis in primary cultured neurons.
    J Neurochem. 1989 Jan;52(1):207-14 PMID: 2908888
  14. Mechanism of galactosylation in the Golgi apparatus. A Chinese hamster ovary cell mutant deficient in translocation of UDP-galactose across Golgi vesicle membranes.
    J Biol Chem. 1986 Jan 5;261(1):96-100 PMID: 3510203
  15. Biosynthesis and modification of Golgi mannosidase II in HeLa and 3T3 cells.
    J Biol Chem. 1985 Jun 10;260(11):6654-62 PMID: 3922977
  16. Translocation of UDP-N-acetylglucosamine into vesicles derived from rat liver rough endoplasmic reticulum and Golgi apparatus.
    J Biol Chem. 1985 Apr 25;260(8):4671-8 PMID: 3988731
  17. Intracellular translocation of fluorescent sphingolipids in cultured fibroblasts: endogenously synthesized sphingomyelin and glucocerebroside analogues pass through the Golgi apparatus en route to the plasma membrane.
    J Cell Biol. 1985 Jan;100(1):27-34 PMID: 3965473
  18. Glycolipid and glycoprotein transport through the Golgi complex are similar biochemically and kinetically. Reconstitution of glycolipid transport in a cell free system.
    J Cell Biol. 1990 Aug;111(2):421-8 PMID: 2166051
  19. Solubilization of UDPglucose-ceramide glucosyltransferase from the Golgi apparatus.
    Biochim Biophys Acta. 1990 May 24;1024(2):263-6 PMID: 2141281
  20. Isolation of a functional vesicular intermediate that mediates ER to Golgi transport in yeast.
    J Cell Biol. 1990 Jul;111(1):45-53 PMID: 2195039
  21. Transmembrane movements of lipids.
    Experientia. 1990 Jun 15;46(6):644-56 PMID: 2193828
  22. Glycolipid transfer protein and intracellular traffic of glucosylceramide.
    Experientia. 1990 Jun 15;46(6):611-6 PMID: 2193825
  23. Sphingomyelin synthesis in rat liver occurs predominantly at the cis and medial cisternae of the Golgi apparatus.
    J Biol Chem. 1990 May 25;265(15):8650-7 PMID: 2187869
  24. Subcellular biosynthesis and transport of gangliosides formed from exogenous lactosylceramide in rat liver.
    Biochem J. 1990 Mar 1;266(2):363-9 PMID: 2317192
  25. Dissection of the Golgi complex. I. Monensin inhibits the transport of viral membrane proteins from medial to trans Golgi cisternae in baby hamster kidney cells infected with Semliki Forest virus.
    J Cell Biol. 1983 Mar;96(3):835-50 PMID: 6682112
  26. Compartmentation of asparagine-linked oligosaccharide processing in the Golgi apparatus.
    J Cell Biol. 1983 Jul;97(1):270-5 PMID: 6223041
  27. Evidence for coupling between transport of UDP-glucose and its synthesis by membrane-bound pyrophosphorylase in Golgi apparatus of cat liver.
    Biochim Biophys Acta. 1984 Jan 25;769(2):377-80 PMID: 6320876
  28. Inhibition of the action of pyrophosphatase and phosphatase on sugar nucleotides.
    J Biol Chem. 1981 Jul 25;256(14):7139-41 PMID: 6114094
  29. Isolation of rat hepatocyte plasma membranes. I. Presence of the three major domains.
    J Cell Biol. 1983 Jan;96(1):217-29 PMID: 6298249
  30. Enzymatic glucosylation of dolichol monophosphate and transfer of glucose from isolated dolichyl-D-glucosyl phosphate to ceramides by BHK-21 cell microsomes.
    Eur J Biochem. 1984 Sep 3;143(2):447-53 PMID: 6236085
  31. A simple and fast method for the isolation of basolateral plasma membranes from rat small-intestinal epithelial cells.
    Biochem J. 1980 Jan 15;186(1):177-81 PMID: 6245639
  32. Tunicamycin inhibits ganglioside biosynthesis in rat liver Golgi apparatus by blocking sugar nucleotide transport across the membrane vesicles.
    Proc Natl Acad Sci U S A. 1983 Dec;80(23):7075-9 PMID: 6417662
  33. Dissection of the Golgi complex. II. Density separation of specific Golgi functions in virally infected cells treated with monensin.
    J Cell Biol. 1983 Mar;96(3):851-6 PMID: 6403555
  34. Action of monensin, a monovalent cationophore, on cultured human fibroblasts: evidence that it induces high cellular accumulation of glucosyl- and lactosylceramide (gluco- and lactocerebroside).
    Biochemistry. 1984 Mar 13;23(6):1043-6 PMID: 6712936
  35. Galactose transfer to endogenous acceptors within Golgi fractions of rat liver.
    J Cell Biol. 1982 Jan;92(1):139-46 PMID: 6799523
  36. Glycosphingolipids in cellular interaction, differentiation, and oncogenesis.
    Annu Rev Biochem. 1981;50:733-64 PMID: 7023369
  37. Distribution of terminal glycosyltransferases in hepatic Golgi fractions.
    J Cell Biol. 1980 Jan;84(1):87-101 PMID: 7350172
  38. The subcellular localization of neutral sphingomyelinase in rat liver.
    J Lipid Res. 1979 May;20(4):456-63 PMID: 37279
  39. Chemical and enzymic degradations of nucleoside mono- and diphosphate sugars. I. Determination of the degradation rate during the glycosyltransferase assays.
    Biochim Biophys Acta. 1979 May 1;584(2):203-15 PMID: 435509
  40. Passage of serum-destined proteins through the Golgi apparatus of rat liver. An examination of heavy and light Golgi fractions.
    J Cell Biol. 1978 Jan;76(1):87-97 PMID: 618900
  41. Golgi fractions prepared from rat liver homogenates. I. Isolation procedure and morphological characterization.
    J Cell Biol. 1973 Oct;59(1):45-72 PMID: 4356571
  42. An improved cell fractionation procedure for the preparation of rat liver membrane-bound ribosomes.
    J Cell Biol. 1973 Jan;56(1):191-205 PMID: 4345164
  43. Sub-Golgi distribution in rat liver of CMP-NeuAc GM3- and CMP-NeuAc:GT1b alpha 2----8sialyltransferases and comparison with the distribution of the other glycosyltransferase activities involved in ganglioside biosynthesis.
    J Biol Chem. 1990 Oct 25;265(30):18242-7 PMID: 2120225
Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2800295
Published
1991-12-01
Pages
295-302
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1130545
Subset
IM
Grants
NIGMS NIH HHS · 2 R37 GM-22942 · United States
External Links
PubMed source DOI: 10.1042/bj2800295 PMC Full Text View journal details
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