Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1994 Nov 8;91(23):10913–10917. doi: 10.1073/pnas.91.23.10913

Localization of the long form of beta-1,4-galactosyltransferase to the plasma membrane and Golgi complex of 3T3 and F9 cells by immunofluorescence confocal microscopy.

A Youakim 1, D H Dubois 1, B D Shur 1
PMCID: PMC45136  PMID: 7971983

Abstract

beta-1,4-Galactosyltransferase (GalTase) is localized to two subcellular compartments, the Golgi complex, where it participates in cellular glycosylation, and the plasma membrane, where it functions as a receptor for oligosaccharide ligands on opposing cells or in the extracellular matrix. The gene for GalTase encodes two nearly identical proteins that differ only in their N-terminal cytoplasmic domains: both short and long GalTases share an 11-aa cytoplasmic tail, but long GalTase has an additional 13-aa sequence on its cytoplasmic domain. In this study, we investigated the subcellular distribution of endogenous long GalTase in untransfected F9 and 3T3 cells by using confocal microscopy and antibodies specific for the 13-aa sequence unique to long GalTase. Long GalTase was found in the Golgi complex as expected; long GalTase was also found on the plasma membrane in cell-type-specific distributions. In 3T3 cells, long GalTase was evident on the basal surface of cells possessing a migratory phenotype, being concentrated at the leading and trailing edges; nonmigratory cells had little detectable surface immunoreactivity. In F9 cells, long GalTase was localized on the plasma membrane, being concentrated at the apical aspect of intercellular junctions. These results demonstrate that in 3T3 and F9 cells, long GalTase is present on the cell surface in addition to the Golgi complex. The pattern of surface expression shows cell-type specificity that is consistent with GalTase function in cellular interactions.

Full text

PDF
10913

Images in this article

10914Image
on p.10914
10915Image
on p.10915
10915Image
on p.10915
10916Image
on p.10916
10916Image
on p.10916

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aoki D., Lee N., Yamaguchi N., Dubois C., Fukuda M. N. Golgi retention of a trans-Golgi membrane protein, galactosyltransferase, requires cysteine and histidine residues within the membrane-anchoring domain. Proc Natl Acad Sci U S A. 1992 May 15;89(10):4319–4323. doi: 10.1073/pnas.89.10.4319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Appeddu P. A., Shur B. D. Molecular analysis of cell surface beta-1,4-galactosyltransferase function during cell migration. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2095–2099. doi: 10.1073/pnas.91.6.2095. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bayna E. M., Shaper J. H., Shur B. D. Temporally specific involvement of cell surface beta-1,4 galactosyltransferase during mouse embryo morula compaction. Cell. 1988 Apr 8;53(1):145–157. doi: 10.1016/0092-8674(88)90496-5. [DOI] [PubMed] [Google Scholar]
  4. Burke J., Pettitt J. M., Schachter H., Sarkar M., Gleeson P. A. The transmembrane and flanking sequences of beta 1,2-N-acetylglucosaminyltransferase I specify medial-Golgi localization. J Biol Chem. 1992 Dec 5;267(34):24433–24440. [PubMed] [Google Scholar]
  5. Childs R. A., Berger E. G., Thorpe S. J., Aegerter E., Feizi T. Blood-group-related carbohydrate antigens are expressed on human milk galactosyltransferase and are immunogenic in rabbits. Biochem J. 1986 Sep 1;238(2):605–611. doi: 10.1042/bj2380605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Colley K. J., Lee E. U., Paulson J. C. The signal anchor and stem regions of the beta-galactoside alpha 2,6-sialyltransferase may each act to localize the enzyme to the Golgi apparatus. J Biol Chem. 1992 Apr 15;267(11):7784–7793. [PubMed] [Google Scholar]
  7. Eckstein D. J., Shur B. D. Laminin induces the stable expression of surface galactosyltransferase on lamellipodia of migrating cells. J Cell Biol. 1989 Jun;108(6):2507–2517. doi: 10.1083/jcb.108.6.2507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Evans S. C., Lopez L. C., Shur B. D. Dominant negative mutation in cell surface beta 1,4-galactosyltransferase inhibits cell-cell and cell-matrix interactions. J Cell Biol. 1993 Feb;120(4):1045–1057. doi: 10.1083/jcb.120.4.1045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hagopian A., Bosmann H. B., Eylar E. H. Glycoprotein biosynthesis: the localization of polypeptidyl: N-acetylgalactosaminyl, collagen: glucosyl, and glycoprotein:galactosyl transferases in HeLa cell membrane fractions. Arch Biochem Biophys. 1968 Nov;128(2):387–396. doi: 10.1016/0003-9861(68)90045-3. [DOI] [PubMed] [Google Scholar]
  10. Harduin-Lepers A., Shaper J. H., Shaper N. L. Characterization of two cis-regulatory regions in the murine beta 1,4-galactosyltransferase gene. Evidence for a negative regulatory element that controls initiation at the proximal site. J Biol Chem. 1993 Jul 5;268(19):14348–14359. [PubMed] [Google Scholar]
  11. Hathaway H. J., Runyan R. B., Khounlo S., Shur B. D. Purification and characterization of avian beta 1,4 galactosyltransferase: comparison with the mammalian enzyme. Glycobiology. 1991 Mar;1(2):211–221. doi: 10.1093/glycob/1.2.211. [DOI] [PubMed] [Google Scholar]
  12. Joziasse D. H. Mammalian glycosyltransferases: genomic organization and protein structure. Glycobiology. 1992 Aug;2(4):271–277. doi: 10.1093/glycob/2.4.271. [DOI] [PubMed] [Google Scholar]
  13. Lopez L. C., Maillet C. M., Oleszkowicz K., Shur B. D. Cell surface and Golgi pools of beta-1,4-galactosyltransferase are differentially regulated during embryonal carcinoma cell differentiation. Mol Cell Biol. 1989 Jun;9(6):2370–2377. doi: 10.1128/mcb.9.6.2370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lopez L. C., Youakim A., Evans S. C., Shur B. D. Evidence for a molecular distinction between Golgi and cell surface forms of beta 1,4-galactosyltransferase. J Biol Chem. 1991 Aug 25;266(24):15984–15991. [PubMed] [Google Scholar]
  15. Masibay A. S., Balaji P. V., Boeggeman E. E., Qasba P. K. Mutational analysis of the Golgi retention signal of bovine beta-1,4-galactosyltransferase. J Biol Chem. 1993 May 5;268(13):9908–9916. [PubMed] [Google Scholar]
  16. Mengle-Gaw L., McCoy-Haman M. F., Tiemeier D. C. Genomic structure and expression of human beta-1,4-galactosyltransferase. Biochem Biophys Res Commun. 1991 May 15;176(3):1269–1276. doi: 10.1016/0006-291x(91)90423-5. [DOI] [PubMed] [Google Scholar]
  17. Munro S. Sequences within and adjacent to the transmembrane segment of alpha-2,6-sialyltransferase specify Golgi retention. EMBO J. 1991 Dec;10(12):3577–3588. doi: 10.1002/j.1460-2075.1991.tb04924.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nilsson T., Lucocq J. M., Mackay D., Warren G. The membrane spanning domain of beta-1,4-galactosyltransferase specifies trans Golgi localization. EMBO J. 1991 Dec;10(12):3567–3575. doi: 10.1002/j.1460-2075.1991.tb04923.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Roseman S. The synthesis of complex carbohydrates by multiglycosyltransferase systems and their potential function in intercellular adhesion. Chem Phys Lipids. 1970 Oct;5(1):270–297. doi: 10.1016/0009-3084(70)90024-1. [DOI] [PubMed] [Google Scholar]
  20. Roth S., McGuire E. J., Roseman S. Evidence for cell-surface glycosyltransferases. Their potential role in cellular recognition. J Cell Biol. 1971 Nov;51(21):536–547. doi: 10.1083/jcb.51.2.536. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Russo R. N., Shaper N. L., Shaper J. H. Bovine beta 1----4-galactosyltransferase: two sets of mRNA transcripts encode two forms of the protein with different amino-terminal domains. In vitro translation experiments demonstrate that both the short and the long forms of the enzyme are type II membrane-bound glycoproteins. J Biol Chem. 1990 Feb 25;265(6):3324–3331. [PubMed] [Google Scholar]
  22. Russo R. N., Shaper N. L., Taatjes D. J., Shaper J. H. Beta 1,4-galactosyltransferase: a short NH2-terminal fragment that includes the cytoplasmic and transmembrane domain is sufficient for Golgi retention. J Biol Chem. 1992 May 5;267(13):9241–9247. [PubMed] [Google Scholar]
  23. Shur B. D. Glycosyltransferases as cell adhesion molecules. Curr Opin Cell Biol. 1993 Oct;5(5):854–863. doi: 10.1016/0955-0674(93)90035-o. [DOI] [PubMed] [Google Scholar]
  24. Strous G. J. Golgi and secreted galactosyltransferase. CRC Crit Rev Biochem. 1986;21(2):119–151. doi: 10.3109/10409238609113610. [DOI] [PubMed] [Google Scholar]
  25. Teasdale R. D., D'Agostaro G., Gleeson P. A. The signal for Golgi retention of bovine beta 1,4-galactosyltransferase is in the transmembrane domain. J Biol Chem. 1992 Feb 25;267(6):4084–4096. [PubMed] [Google Scholar]
  26. Youakim A., Hathaway H. J., Miller D. J., Gong X., Shur B. D. Overexpressing sperm surface beta 1,4-galactosyltransferase in transgenic mice affects multiple aspects of sperm-egg interactions. J Cell Biol. 1994 Sep;126(6):1573–1583. doi: 10.1083/jcb.126.6.1573. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Youakim A., Shur B. D. Effects of overexpression of beta 1,4-galactosyltransferase on glycoprotein biosynthesis in F9 embryonal carcinoma cells. Glycobiology. 1993 Apr;3(2):155–163. doi: 10.1093/glycob/3.2.155. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES