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PMID: 2166051 Published · ppublish English Journal Article

Glycolipid and glycoprotein transport through the Golgi complex are similar biochemically and kinetically. Reconstitution of glycolipid transport in a cell free system.

The Journal of cell biology ·Vol. 111 ·No. 2 ·1990-08-00 ·Pages 421-8

Wattenberg BW

Abstract

Glycolipid transport between compartments of the Golgi apparatus has been reconstituted in a cell free system. Transport of lactosylceramide (galactose beta 1-4-glucose-ceramide) was followed from a donor to an acceptor Golgi population. The major glycolipid in CHO cells is GM3 (sialic acid alpha 2-3 galactose beta 1-4-glucose-ceramide). Donor membranes were derived from a Chinese hamster ovary (CHO) cell mutant (Lec2) deficient in the Golgi CMP-sialic acid transporter, and therefore contained lactosylceramide as the predominant glycolipid. Acceptor Golgi apparatus was prepared from another mutant, Lec8, which is defective in UDP-Gal transport. Thus, glucosylceramide is the major glycolipid in Lec8 cells. Transport was measured by the incorporation of labeled sialic acid into lactosylceramide (present originally in the donor) by transport to acceptor membranes, forming GM3. This incorporation was dependent on ATP, cytosolic components, intact membranes, and elevated temperature. Donor membranes were prepared from Lec2 cells infected with vesicular stomatitus virus (VSV). These membranes therefore contain the VSV membrane glycoprotein, G protein. Donor membranes derived from VSV-infected cells could then be used to monitor both glycolipid and glycoprotein transport. Transport of these two types of molecules between Golgi compartments was compared biochemically and kinetically. Glycolipid transport required the N-ethylmaleimide sensitive factor previously shown to act in glycoprotein transport (Glick, B. S., and J. E. Rothman. 1987. Nature [Lond.]. 326:309-312; Rothman, J. E. 1987. J. Biol. Chem. 262:12502-12510). GTP gamma S inhibited glycolipid and glycoprotein transport similarly. The kinetics of transport of glycolipid and glycoprotein were also compared. The kinetics of transport to the end of the pathway were similar, as were the kinetics of movement into a defined transport intermediate. It is concluded that glycolipid and glycoprotein transport through the Golgi occur by similar if not identical mechanisms.

MeSH Terms
Animals Antigens, CD Cell Line Cell Membrane/metabolism Cell Transformation, Viral Cell-Free System Cytosol/metabolism Ethylmaleimide/pharmacology G(M3) Ganglioside/metabolism Glycolipids/metabolism Glycoproteins/genetics,metabolism Glycosphingolipids/metabolism Golgi Apparatus/metabolism Guanosine 5'-O-(3-Thiotriphosphate) Guanosine Triphosphate/analogs & derivatives,pharmacology Kinetics Lactosylceramides Models, Biological N-Acetylneuraminic Acid Protein Processing, Post-Translational Sialic Acids/metabolism Thionucleotides/pharmacology Vesicular stomatitis Indiana virus/genetics
Chemicals
Antigens, CD G(M3) Ganglioside Glycolipids Glycoproteins Glycosphingolipids Lactosylceramides Sialic Acids Thionucleotides Guanosine 5'-O-(3-Thiotriphosphate) CDw17 antigen Guanosine Triphosphate N-Acetylneuraminic Acid Ethylmaleimide
Authors & Affiliations
1 authors, click to expand affiliations / ORCID
Wattenberg B W
Cell Biology Unit, Upjohn Company, Kalamazoo, Michigan 49007.
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43 references, click to expand
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.111.2.421
Published
1990-08-00
Pages
421-8
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2116202
Subset
IM
External Links
PubMed source DOI: 10.1083/jcb.111.2.421 PMC Full Text View journal details
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