Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1991 Oct 2;115(2):485–493. doi: 10.1083/jcb.115.2.485

Hepatocyte adhesion to carbohydrate-derivatized surfaces. I. Surface topography of the rat hepatic lectin

PMCID: PMC2289166  PMID: 1655806

Abstract

The rat hepatic lectins, galactose- and N-acetylgalactosamine-binding proteins found on the hepatocyte cell surface, mediate adhesion of isolated primary rat hepatocytes to artificial galactose-derivatized polyacrylamide gels. Biochemical and immunohistochemical techniques were used to examine the topographical redistribution of the rat hepatic lectins in response to galactose-mediated cell adhesion. Hepatocytes isolated from rat liver by collagenase perfusion had an average of 7 x 10(5) cell surface lectin molecules per cell, representing 30-50% of the total lectin molecules per cell, the remainder residing in intracellular pools. Hepatocytes incubated on galactose-derivatized surfaces, whether at 0-4 degrees C or 37 degrees C, rapidly lost greater than 80% of their accessible cell surface lectin binding sites into an adhesive patch of characteristic morphology. The kinetics of rat hepatic lectin disappearance were used to estimate a lateral diffusion coefficient greater than 9 x 10(-9) cm2/s at 37 degrees C, suggesting rapid and unimpeded lectin diffusion in the plane of the membrane. Indirect immunofluorescence labeling of adherent cells using antihepatic lectin antibody revealed a structured ring of receptors surrounding an area of exclusion (patch) of reproducible size and shape which represented approximately 8% of the hepatocyte cell surface. Notably, adherent cells, which had lost greater than 80% of their accessible surface binding sites, still endocytosed soluble galactose-terminated radioligand at greater than 50% of the rate of nonadherent control cells. No net movement of rat hepatic lectin from intracellular pools to the cell surface was found on cells recovered after adhesion to galactose-derivatized surfaces at 37 degrees C, suggesting that the physical size and/or lectin density of the patch was restricted by kinetic or topological constraints.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Ashwell G., Harford J. Carbohydrate-specific receptors of the liver. Annu Rev Biochem. 1982;51:531–554. doi: 10.1146/annurev.bi.51.070182.002531. [DOI] [PubMed] [Google Scholar]
  2. Berg H. C., Purcell E. M. Physics of chemoreception. Biophys J. 1977 Nov;20(2):193–219. doi: 10.1016/S0006-3495(77)85544-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bischoff J., Libresco S., Shia M. A., Lodish H. F. The H1 and H2 polypeptides associate to form the asialoglycoprotein receptor in human hepatoma cells. J Cell Biol. 1988 Apr;106(4):1067–1074. doi: 10.1083/jcb.106.4.1067. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brandley B. K., Schnaar R. L. Phosphorylation of extracellular carbohydrates by intact cells. Chicken hepatocytes specifically adhere to and phosphorylate immobilized N-acetylglucosamine. J Biol Chem. 1985 Oct 15;260(23):12474–12483. [PubMed] [Google Scholar]
  5. Chao N. M., Young S. H., Poo M. M. Localization of cell membrane components by surface diffusion into a "trap". Biophys J. 1981 Oct;36(1):139–153. doi: 10.1016/S0006-3495(81)84721-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Connolly D. T., Townsend R. R., Kawaguchi K., Hobish M. K., Bell W. R., Lee Y. C. Binding and endocytosis of glycoproteins and neoglycoproteins by isolated rabbit hepatocytes. Biochem J. 1983 Aug 15;214(2):421–431. doi: 10.1042/bj2140421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. DULBECCO R., VOGT M. Plaque formation and isolation of pure lines with poliomyelitis viruses. J Exp Med. 1954 Feb;99(2):167–182. doi: 10.1084/jem.99.2.167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Drickamer K., Mamon J. F., Binns G., Leung J. O. Primary structure of the rat liver asialoglycoprotein receptor. Structural evidence for multiple polypeptide species. J Biol Chem. 1984 Jan 25;259(2):770–778. [PubMed] [Google Scholar]
  9. Drickamer K. Two distinct classes of carbohydrate-recognition domains in animal lectins. J Biol Chem. 1988 Jul 15;263(20):9557–9560. [PubMed] [Google Scholar]
  10. Dunn W. A., Connolly T. P., Hubbard A. L. Receptor-mediated endocytosis of epidermal growth factor by rat hepatocytes: receptor pathway. J Cell Biol. 1986 Jan;102(1):24–36. doi: 10.1083/jcb.102.1.24. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Guarnaccia S. P., Schnaar R. L. Hepatocyte adhesion to immobilized carbohydrates. I. Sugar recognition is followed by energy-dependent strengthening. J Biol Chem. 1982 Dec 10;257(23):14288–14292. [PubMed] [Google Scholar]
  12. Halberg D. F., Wager R. E., Farrell D. C., Hildreth J., 4th, Quesenberry M. S., Loeb J. A., Holland E. C., Drickamer K. Major and minor forms of the rat liver asialoglycoprotein receptor are independent galactose-binding proteins. Primary structure and glycosylation heterogeneity of minor receptor forms. J Biol Chem. 1987 Jul 15;262(20):9828–9838. [PubMed] [Google Scholar]
  13. Herzig M. C., Weigel P. H. Surface and internal galactosyl receptors are heterooligomers and retain this structure after ligand internalization or receptor modulation. Biochemistry. 1990 Jul 10;29(27):6437–6447. doi: 10.1021/bi00479a015. [DOI] [PubMed] [Google Scholar]
  14. Kreamer B. L., Staecker J. L., Sawada N., Sattler G. L., Hsia M. T., Pitot H. C. Use of a low-speed, iso-density percoll centrifugation method to increase the viability of isolated rat hepatocyte preparations. In Vitro Cell Dev Biol. 1986 Apr;22(4):201–211. doi: 10.1007/BF02623304. [DOI] [PubMed] [Google Scholar]
  15. Lee Y. C., Stowell C. P., Krantz M. J. 2-Imino-2-methoxyethyl 1-thioglycosides: new reagents for attaching sugars to proteins. Biochemistry. 1976 Sep 7;15(18):3956–3963. doi: 10.1021/bi00663a008. [DOI] [PubMed] [Google Scholar]
  16. McLean I. W., Nakane P. K. Periodate-lysine-paraformaldehyde fixative. A new fixation for immunoelectron microscopy. J Histochem Cytochem. 1974 Dec;22(12):1077–1083. doi: 10.1177/22.12.1077. [DOI] [PubMed] [Google Scholar]
  17. Mueller S. C., Hubbard A. L. Receptor-mediated endocytosis of asialoglycoproteins by rat hepatocytes: receptor-positive and receptor-negative endosomes. J Cell Biol. 1986 Mar;102(3):932–942. doi: 10.1083/jcb.102.3.932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Obrink B., Kuhlenschmidt M. S., Roseman S. Adhesive specificity of juvenile rat and chicken liver cells and membranes. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1077–1081. doi: 10.1073/pnas.74.3.1077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Oka J. A., Weigel P. H. Binding and spreading of hepatocytes on synthetic galactose culture surfaces occur as distinct and separable threshold responses. J Cell Biol. 1986 Sep;103(3):1055–1060. doi: 10.1083/jcb.103.3.1055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pless D. D., Lee Y. C., Roseman S., Schnaar R. L. Specific cell adhesion to immobilized glycoproteins demonstrated using new reagents for protein and glycoprotein immobilization. J Biol Chem. 1983 Feb 25;258(4):2340–2349. [PubMed] [Google Scholar]
  21. Rice K. G., Weisz O. A., Barthel T., Lee R. T., Lee Y. C. Defined geometry of binding between triantennary glycopeptide and the asialoglycoprotein receptor of rat heptocytes. J Biol Chem. 1990 Oct 25;265(30):18429–18434. [PubMed] [Google Scholar]
  22. Sawyer J. T., Sanford J. P., Doyle D. Identification of a complex of the three forms of the rat liver asialoglycoprotein receptor. J Biol Chem. 1988 Jul 25;263(21):10534–10538. [PubMed] [Google Scholar]
  23. Schnaar R. L., Langer B. G., Brandley B. K. Reversible covalent immobilization of ligands and proteins on polyacrylamide gels. Anal Biochem. 1985 Dec;151(2):268–281. doi: 10.1016/0003-2697(85)90175-7. [DOI] [PubMed] [Google Scholar]
  24. Schnaar R. L., Weigel P. H., Kuhlenschmidt M. S., Lee Y. C., Roseman S. Adhesion of chicken hepatocytes to polyacrylamide gels derivatized with N-acetylglucosamine. J Biol Chem. 1978 Nov 10;253(21):7940–7951. [PubMed] [Google Scholar]
  25. Schwartz A. L., Marshak-Rothstein A., Rup D., Lodish H. F. Identification and quantification of the rat hepatocyte asialoglycoprotein receptor. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3348–3352. doi: 10.1073/pnas.78.6.3348. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Seglen P. O. Preparation of isolated rat liver cells. Methods Cell Biol. 1976;13:29–83. doi: 10.1016/s0091-679x(08)61797-5. [DOI] [PubMed] [Google Scholar]
  27. Spiess M. The asialoglycoprotein receptor: a model for endocytic transport receptors. Biochemistry. 1990 Oct 30;29(43):10009–10018. doi: 10.1021/bi00495a001. [DOI] [PubMed] [Google Scholar]
  28. Weaver D. L. Diffusion-mediated localization on membrane surfaces. Biophys J. 1983 Jan;41(1):81–86. doi: 10.1016/S0006-3495(83)84407-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Weigel P. H. Characterization of the asialoglycoprotein receptor on isolated rat hepatocytes. J Biol Chem. 1980 Jul 10;255(13):6111–6120. [PubMed] [Google Scholar]
  30. Weigel P. H. Rat hepatocytes bind to synthetic galactoside surfaces via a patch of asialoglycoprotein receptors. J Cell Biol. 1980 Dec;87(3 Pt 1):855–861. doi: 10.1083/jcb.87.3.855. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Weigel P. H., Schmell E., Lee Y. C., Roseman S. Specific adhesion of rat hepatocytes to beta-galactosides linked to polyacrylamide gels. J Biol Chem. 1978 Jan 25;253(2):330–333. [PubMed] [Google Scholar]
  32. Weigel P. H., Schnaar R. L., Kuhlenschmidt M. S., Schmell E., Lee R. T., Lee Y. C., Roseman S. Adhesion of hepatocytes to immobilized sugars. A threshold phenomenon. J Biol Chem. 1979 Nov 10;254(21):10830–10838. [PubMed] [Google Scholar]
  33. Weisz O. A., Schnaar R. L. Hepatocyte adhesion to carbohydrate-derivatized surfaces. II. Regulation of cytoskeletal organization and cell morphology. J Cell Biol. 1991 Oct;115(2):495–504. doi: 10.1083/jcb.115.2.495. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES