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. 1985 Jan 1;100(1):118–125. doi: 10.1083/jcb.100.1.118

Immunocytochemical demonstration of ecto-galactosyltransferase in absorptive intestinal cells

PMCID: PMC2113462  PMID: 3917437

Abstract

Galactosyltransferase immunoreactive sites were localized in human duodenal enterocytes by the protein A-gold technique on thin sections from low temperature Lowicryl K4M embedded biopsy specimens. Antigenic sites detected with affinity-purified, monospecific antibodies were found at the plasma membrane of absorptive enterocytes with the most intense labeling appearing along the brush border membrane. The lateral plasma membrane exhibited a lower degree of labeling at the level of the junctional complexes but the membrane interdigitations were intensely labeled. The labeling intensity decreased progressively towards the basal part of the enterocytes and reached the lowest degree along the basal plasma membrane. Quantitative evaluation of the distribution of gold-particle label proved its preferential orientation to the outer surface of the plasma membrane. In addition to this membrane-associated labeling, the glycocalyx extending from the microvillus tips was heavily labeled. Occasionally, cells without plasma membrane labeling were found adjacent to positive cells. The demonstration of ecto-galactosyltransferase on membranes other than Golgi membranes precludes its general use as a marker for Golgi membrane fractions. The possible function of galactosyltransferase on a luminal plasma membrane is unclear at present, but a role in adhesion appears possible on the basolateral plasma membrane.

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Selected References

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  1. Andrews P. Purification of lactose synthetase a protein from human milk and demonstration of its interaction with alpha-lactalbumin. FEBS Lett. 1970 Aug 31;9(5):297–300. doi: 10.1016/0014-5793(70)80382-9. [DOI] [PubMed] [Google Scholar]
  2. Bennett G., Leblond C. P., Haddad A. Migration of glycoprotein from the Golgi apparatus to the surface of various cell types as shown by radioautography after labelled fucose injection into rats. J Cell Biol. 1974 Jan;60(1):258–284. doi: 10.1083/jcb.60.1.258. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berger E. G., Mandel T., Schilt U. Immunohistochemical localization of galactosyltransferase in human fibroblasts and HeLa cells. J Histochem Cytochem. 1981 Mar;29(3):364–370. doi: 10.1177/29.3.6787115. [DOI] [PubMed] [Google Scholar]
  4. Berger E. G., Verdon B., Mandel T., Fey H., Strous G. Electrophoretic comparison of cellular and soluble galactosyltransferase (lactose synthetase A protein) using specific antibodies. Enzyme. 1983;29(3):175–182. doi: 10.1159/000469630. [DOI] [PubMed] [Google Scholar]
  5. Bretz R., Bretz H., Palade G. E. Distribution of terminal glycosyltransferases in hepatic Golgi fractions. J Cell Biol. 1980 Jan;84(1):87–101. doi: 10.1083/jcb.84.1.87. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cummings R. D., Cebula T. A., Roth S. Characterization of a galactosyltransferase in plasma membrane-enriched fractions from Balb/c 3T12 cells. J Biol Chem. 1979 Feb 25;254(4):1233–1240. [PubMed] [Google Scholar]
  7. Davis B. W., Berger E. G., Locher G. W., Zeller M., Goldhirsch A. Immunohistochemical localization of galactosyltransferase in the normal human ovary and fallopian tube. J Histochem Cytochem. 1984 Jan;32(1):92–96. doi: 10.1177/32.1.6418792. [DOI] [PubMed] [Google Scholar]
  8. Deppert W., Werchau H., Walter G. Differentiation between intracellular and cell surface glycosyl transferases: galactosyl transferase activity in intact cells and in cell homogenate. Proc Natl Acad Sci U S A. 1974 Aug;71(8):3068–3072. doi: 10.1073/pnas.71.8.3068. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Farquhar M. G., Palade G. E. The Golgi apparatus (complex)-(1954-1981)-from artifact to center stage. J Cell Biol. 1981 Dec;91(3 Pt 2):77s–103s. doi: 10.1083/jcb.91.3.77s. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gerber A. C., Kozdrowski I., Wyss S. R., Berger E. G. The charge heterogeneity of soluble human galactosyltransferases isolated from milk, amniotic fluid and malignant ascites. Eur J Biochem. 1979 Feb 1;93(3):453–460. doi: 10.1111/j.1432-1033.1979.tb12843.x. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Huggins J. W., Trenbeath T. P., Chesnut R. W., Carothers C. A., Carraway K. L. Purification of plasma membranes of rat mammary gland. Comparisons of subfractions with rat milk fat globule membrane. Exp Cell Res. 1980 Apr;126(2):279–288. doi: 10.1016/0014-4827(80)90266-9. [DOI] [PubMed] [Google Scholar]
  13. Keenan T. W., Morré D. J. Glycosyltransferases: do they exist on the surface membrane of mammalian cells? FEBS Lett. 1975 Jul 15;55(1):8–13. doi: 10.1016/0014-5793(75)80944-6. [DOI] [PubMed] [Google Scholar]
  14. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  15. Lau J. T., Carlson D. M. Galactosyltransferase activities in rat intestinal mucosa. Inhibition of nucleotide pyrophosphatase. J Biol Chem. 1981 Jul 25;256(14):7142–7145. [PubMed] [Google Scholar]
  16. Patt L. M., Grimes W. J. Cell surface glycolipid and glycoprotein glycosyltransferases of normal and transformed cells. J Biol Chem. 1974 Jul 10;249(13):4157–4165. [PubMed] [Google Scholar]
  17. Pestalozzi D. M., Hess M., Berger E. G. Immunohistochemical evidence for cell surface and Golgi localization of galactosyltransferase in human stomach, jejunum, liver and pancreas. J Histochem Cytochem. 1982 Nov;30(11):1146–1152. doi: 10.1177/30.11.6815262. [DOI] [PubMed] [Google Scholar]
  18. Pierce M., Turley E. A., Roth S. Cell surface glycosyltransferase activities. Int Rev Cytol. 1980;65:1–47. doi: 10.1016/s0074-7696(08)61958-0. [DOI] [PubMed] [Google Scholar]
  19. Podolsky D. K., Weiser M. M. Galactosyltransferase activities in human sera: detection of a cancer-associated isoenzyme. Biochem Biophys Res Commun. 1975 Jul 22;65(2):545–551. doi: 10.1016/s0006-291x(75)80181-1. [DOI] [PubMed] [Google Scholar]
  20. Porter C. W., Bernacki R. J. Ultrastructural evidence for ectoglycosyltransferase systems. Nature. 1975 Aug 21;256(5519):648–650. doi: 10.1038/256648a0. [DOI] [PubMed] [Google Scholar]
  21. Quaroni A., Kirsch K., Weiser M. M. Synthesis of membrane glycoproteins in rat small-intestinal villus cells. Effect of colchicine on the redistribution of L-[1,5,6-3H]fucose-labelled membrane glycoproteins among Golgi, lateral basal and microvillus membranes. Biochem J. 1979 Jul 15;182(1):213–221. doi: 10.1042/bj1820213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rauvala H., Prieels J. P., Finne J. Cell adhesion mediated by a purified fucosyltransferase. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3991–3995. doi: 10.1073/pnas.80.13.3991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Rindler M. J., Ivanov I. E., Plesken H., Rodriguez-Boulan E., Sabatini D. D. Viral glycoproteins destined for apical or basolateral plasma membrane domains traverse the same Golgi apparatus during their intracellular transport in doubly infected Madin-Darby canine kidney cells. J Cell Biol. 1984 Apr;98(4):1304–1319. doi: 10.1083/jcb.98.4.1304. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Roth J. Application of lectin--gold complexes for electron microscopic localization of glycoconjugates on thin sections. J Histochem Cytochem. 1983 Aug;31(8):987–999. doi: 10.1177/31.8.6190857. [DOI] [PubMed] [Google Scholar]
  26. Roth J., Bendayan M., Orci L. Ultrastructural localization of intracellular antigens by the use of protein A-gold complex. J Histochem Cytochem. 1978 Dec;26(12):1074–1081. doi: 10.1177/26.12.366014. [DOI] [PubMed] [Google Scholar]
  27. Roth J., Berger E. G. Immunocytochemical localization of galactosyltransferase in HeLa cells: codistribution with thiamine pyrophosphatase in trans-Golgi cisternae. J Cell Biol. 1982 Apr;93(1):223–229. doi: 10.1083/jcb.93.1.223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sharon N., Eshdat Y., Silverblatt F. J., Ofek I. Bacterial adherence to cell surface sugars. Ciba Found Symp. 1981;80:119–141. doi: 10.1002/9780470720639.ch9. [DOI] [PubMed] [Google Scholar]
  29. Shur B. D. Evidence that galactosyltransferase is a surface receptor for poly(N)-acetyllactosamine glycoconjugates on embryonal carcinoma cells. J Biol Chem. 1982 Jun 25;257(12):6871–6878. [PubMed] [Google Scholar]
  30. Shur B. D., Roth S. Cell surface glycosyltransferases. Biochim Biophys Acta. 1975 Dec 29;415(4):473–512. doi: 10.1016/0304-4157(75)90007-6. [DOI] [PubMed] [Google Scholar]
  31. Strous G. J., Berger E. G. Biosynthesis, intracellular transport, and release of the Golgi enzyme galactosyltransferase (lactose synthetase A protein) in HeLa cells. J Biol Chem. 1982 Jul 10;257(13):7623–7628. [PubMed] [Google Scholar]
  32. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vegt G. B., Di Bon-De Ruijter M., Hekkens W. T. Studies on the determination of extracellular galactosyltransferase in human intestinal tissue. Clin Chim Acta. 1981 Jan 22;109(2):145–150. doi: 10.1016/0009-8981(81)90328-4. [DOI] [PubMed] [Google Scholar]
  34. Verbert A., Cacan R., Montreuil J. Ectogalactosyltransferase. Presence of enzyme and acceptors on the rat lymphocyte cell surface. Eur J Biochem. 1976 Nov 1;70(1):49–53. doi: 10.1111/j.1432-1033.1976.tb10954.x. [DOI] [PubMed] [Google Scholar]
  35. Weiser M. M. Intestinal epithelial cell surface membrane glycoprotein synthesis. II. Glycosyltransferases and endogenous acceptors of the undifferentiated cell surface membrane. J Biol Chem. 1973 Apr 10;248(7):2542–2548. [PubMed] [Google Scholar]
  36. Weiser M. M., Neumeier M. M., Quaroni A., Kirsch K. Synthesis of plasmalemmal glycoproteins in intestinal epithelial cells. Separation of Golgi membranes from villus and crypt cell surface membranes; glycosyltransferase activity of surface membrane. J Cell Biol. 1978 Jun;77(3):722–734. doi: 10.1083/jcb.77.3.722. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wilson J. R., Weiser M. M., Albini B., Schenck J. R., Rittenhouse H. G., Hirata A. A., Berger E. G. Co-purification of soluble human galactosyltransferase and immunoglobulins. Biochem Biophys Res Commun. 1982 Mar 30;105(2):737–744. doi: 10.1016/0006-291x(82)91496-6. [DOI] [PubMed] [Google Scholar]

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