Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1987 Apr 1;104(4):905–916. doi: 10.1083/jcb.104.4.905

Formation of the apical pole of epithelial (Madin-Darby canine kidney) cells: polarity of an apical protein is independent of tight junctions while segregation of a basolateral marker requires cell-cell interactions

PMCID: PMC2114453  PMID: 3558485

Abstract

The time course of development of polarity of an apical (184-kD) and a basolateral (63-kD) plasma membrane protein of Madin-Darby canine kidney cells was followed using semiquantitative immunofluorescence on semithin (approximately 0.5-micron) frozen sections and monoclonal antibody probes. The 184-kD protein became highly polarized to the apical pole within the initial 24 h both in normal medium and in 1-5 microM Ca2+, which results in well-spread, dome-shaped cells, lacking tight junctions and other lateral membrane interactions. In contrast, the basolateral 63-kD membrane protein developed full polarity only after incubation in normal Ca2+ concentrations for greater than 72 h, a time much longer than that required for the formation of tight junctions (approximately 18 h) and failed to polarize in 1-5 microM Ca2+. These results demonstrate that intradomain restriction mechanisms independent of tight junctions, such as self-aggregation or specific interactions with the submembrane cytoskeleton, participate in the regionalization of at least some epithelial plasma membrane proteins. The full operation of these mechanisms depends on the presence of normal cell-cell interactions in the case of the basolateral 63-kD antigen but not in the case of the apical 184-kD protein.

Full Text

The Full Text of this article is available as a PDF (3.1 MB).

Selected References

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

  1. Balcarova-Ständer J., Pfeiffer S. E., Fuller S. D., Simons K. Development of cell surface polarity in the epithelial Madin-Darby canine kidney (MDCK) cell line. EMBO J. 1984 Nov;3(11):2687–2694. doi: 10.1002/j.1460-2075.1984.tb02194.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bennett V. The molecular basis for membrane - cytoskeleton association in human erythrocytes. J Cell Biochem. 1982;18(1):49–65. doi: 10.1002/jcb.1982.240180106. [DOI] [PubMed] [Google Scholar]
  3. Branton D., Cohen C. M., Tyler J. Interaction of cytoskeletal proteins on the human erythrocyte membrane. Cell. 1981 Apr;24(1):24–32. doi: 10.1016/0092-8674(81)90497-9. [DOI] [PubMed] [Google Scholar]
  4. Caplan M. J., Anderson H. C., Palade G. E., Jamieson J. D. Intracellular sorting and polarized cell surface delivery of (Na+,K+)ATPase, an endogenous component of MDCK cell basolateral plasma membranes. Cell. 1986 Aug 15;46(4):623–631. doi: 10.1016/0092-8674(86)90888-3. [DOI] [PubMed] [Google Scholar]
  5. Cereijido M., Ehrenfeld J., Meza I., Martínez-Palomo A. Structural and functional membrane polarity in cultured monolayers of MDCK cells. J Membr Biol. 1980;52(2):147–159. doi: 10.1007/BF01869120. [DOI] [PubMed] [Google Scholar]
  6. Cereijido M., Robbins E. S., Dolan W. J., Rotunno C. A., Sabatini D. D. Polarized monolayers formed by epithelial cells on a permeable and translucent support. J Cell Biol. 1978 Jun;77(3):853–880. doi: 10.1083/jcb.77.3.853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chambard M., Gabrion J., Mauchamp J. Influence of collagen gel on the orientation of epithelial cell polarity: follicle formation from isolated thyroid cells and from preformed monolayers. J Cell Biol. 1981 Oct;91(1):157–166. doi: 10.1083/jcb.91.1.157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cox J. V., Moon R. T., Lazarides E. Anion transporter: highly cell-type-specific expression of distinct polypeptides and transcripts in erythroid and nonerythroid cells. J Cell Biol. 1985 May;100(5):1548–1557. doi: 10.1083/jcb.100.5.1548. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. De Camilli P., Peluchetti D., Meldolesi J. Structural difference between luminal and lateral plasmalemma in pancreatic acinar cells. Nature. 1974 Mar 15;248(445):245–247. doi: 10.1038/248245b0. [DOI] [PubMed] [Google Scholar]
  10. Diamond J. M. Twenty-first Bowditch lecture. The epithelial junction: bridge, gate, and fence. Physiologist. 1977 Feb;20(1):10–18. [PubMed] [Google Scholar]
  11. Dragsten P. R., Handler J. S., Blumenthal R. Fluorescent membrane probes and the mechanism of maintenance of cellular asymmetry in epithelia. Fed Proc. 1982 Jan;41(1):48–53. [PubMed] [Google Scholar]
  12. Drenckhahn D., Schlüter K., Allen D. P., Bennett V. Colocalization of band 3 with ankyrin and spectrin at the basal membrane of intercalated cells in the rat kidney. Science. 1985 Dec 13;230(4731):1287–1289. doi: 10.1126/science.2933809. [DOI] [PubMed] [Google Scholar]
  13. Edelman G. M. Cell adhesion molecules. Science. 1983 Feb 4;219(4584):450–457. doi: 10.1126/science.6823544. [DOI] [PubMed] [Google Scholar]
  14. Edelman G. M. Modulation of cell adhesion during induction, histogenesis, and perinatal development of the nervous system. Annu Rev Neurosci. 1984;7:339–377. doi: 10.1146/annurev.ne.07.030184.002011. [DOI] [PubMed] [Google Scholar]
  15. Ekblom P. Determination and differentiation of the nephron. Med Biol. 1981 Jun;59(3):139–160. [PubMed] [Google Scholar]
  16. FARQUHAR M. G., PALADE G. E. Junctional complexes in various epithelia. J Cell Biol. 1963 May;17:375–412. doi: 10.1083/jcb.17.2.375. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fisher K. A., Stoeckenius W. Freeze-fractured purple membrane particles: protein content. Science. 1977 Jul 1;197(4298):72–74. doi: 10.1126/science.867052. [DOI] [PubMed] [Google Scholar]
  18. Fradet Y., Cordon-Cardo C., Thomson T., Daly M. E., Whitmore W. F., Jr, Lloyd K. O., Melamed M. R., Old L. J. Cell surface antigens of human bladder cancer defined by mouse monoclonal antibodies. Proc Natl Acad Sci U S A. 1984 Jan;81(1):224–228. doi: 10.1073/pnas.81.1.224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gonzalez-Mariscal L., Chávez de Ramírez B., Cereijido M. Tight junction formation in cultured epithelial cells (MDCK). J Membr Biol. 1985;86(2):113–125. doi: 10.1007/BF01870778. [DOI] [PubMed] [Google Scholar]
  20. Goodenough D. A., Revel J. P. A fine structural analysis of intercellular junctions in the mouse liver. J Cell Biol. 1970 May;45(2):272–290. doi: 10.1083/jcb.45.2.272. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Griffiths G., Warren G., Quinn P., Mathieu-Costello O., Hoppeler H. Density of newly synthesized plasma membrane proteins in intracellular membranes. I. Stereological studies. J Cell Biol. 1984 Jun;98(6):2133–2141. doi: 10.1083/jcb.98.6.2133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Gumbiner B., Simons K. A functional assay for proteins involved in establishing an epithelial occluding barrier: identification of a uvomorulin-like polypeptide. J Cell Biol. 1986 Feb;102(2):457–468. doi: 10.1083/jcb.102.2.457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hall H. G., Farson D. A., Bissell M. J. Lumen formation by epithelial cell lines in response to collagen overlay: a morphogenetic model in culture. Proc Natl Acad Sci U S A. 1982 Aug;79(15):4672–4676. doi: 10.1073/pnas.79.15.4672. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Herzlinger D. A., Ojakian G. K. Studies on the development and maintenance of epithelial cell surface polarity with monoclonal antibodies. J Cell Biol. 1984 May;98(5):1777–1787. doi: 10.1083/jcb.98.5.1777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Hoi Sang U., Saier M. H., Jr, Ellisman M. H. Tight junction formation is closely linked to the polar redistribution of intramembranous particles in aggregating MDCK epithelia. Exp Cell Res. 1979 Sep;122(2):384–391. doi: 10.1016/0014-4827(79)90315-x. [DOI] [PubMed] [Google Scholar]
  26. Imhof B. A., Vollmers H. P., Goodman S. L., Birchmeier W. Cell-cell interaction and polarity of epithelial cells: specific perturbation using a monoclonal antibody. Cell. 1983 Dec;35(3 Pt 2):667–675. doi: 10.1016/0092-8674(83)90099-5. [DOI] [PubMed] [Google Scholar]
  27. Ingber D. E., Madri J. A., Jamieson J. D. Basement membrane as a spatial organizer of polarized epithelia. Exogenous basement membrane reorients pancreatic epithelial tumor cells in vitro. Am J Pathol. 1986 Jan;122(1):129–139. [PMC free article] [PubMed] [Google Scholar]
  28. Johnson M. H., Ziomek C. A. Induction of polarity in mouse 8-cell blastomeres: specificity, geometry, and stability. J Cell Biol. 1981 Oct;91(1):303–308. doi: 10.1083/jcb.91.1.303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Köhler G., Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975 Aug 7;256(5517):495–497. doi: 10.1038/256495a0. [DOI] [PubMed] [Google Scholar]
  30. Lamb J. F., Ogden P., Simmons N. L. Autoradiographic localisation of [3H]ouabain bound to cultured epithelial cell monolayers of MDCK cells. Biochim Biophys Acta. 1981 Jun 22;644(2):333–340. doi: 10.1016/0005-2736(81)90391-6. [DOI] [PubMed] [Google Scholar]
  31. Leighton J., Estes L. W., Mansukhani S., Brada Z. A cell line derived from normal dog kidney (MDCK) exhibiting qualities of papillary adenocarcinoma and of renal tubular epithelium. Cancer. 1970 Nov;26(5):1022–1028. doi: 10.1002/1097-0142(197011)26:5<1022::aid-cncr2820260509>3.0.co;2-m. [DOI] [PubMed] [Google Scholar]
  32. Louvard D. Apical membrane aminopeptidase appears at site of cell-cell contact in cultured kidney epithelial cells. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4132–4136. doi: 10.1073/pnas.77.7.4132. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Matlin K. S., Simons K. Sorting of an apical plasma membrane glycoprotein occurs before it reaches the cell surface in cultured epithelial cells. J Cell Biol. 1984 Dec;99(6):2131–2139. doi: 10.1083/jcb.99.6.2131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Matlin K., Bainton D. F., Pesonen M., Louvard D., Genty N., Simons K. Transepithelial transport of a viral membrane glycoprotein implanted into the apical plasma membrane of Madin-Darby canine kidney cells. I. Morphological evidence. J Cell Biol. 1983 Sep;97(3):627–637. doi: 10.1083/jcb.97.3.627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Michl J., Pieczonka M. M., Unkeless J. C., Silverstein S. C. Effects of immobilized immune complexes on Fc- and complement-receptor function in resident and thioglycollate-elicited mouse peritoneal macrophages. J Exp Med. 1979 Sep 19;150(3):607–621. doi: 10.1084/jem.150.3.607. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Misek D. E., Bard E., Rodriguez-Boulan E. Biogenesis of epithelial cell polarity: intracellular sorting and vectorial exocytosis of an apical plasma membrane glycoprotein. Cell. 1984 Dec;39(3 Pt 2):537–546. doi: 10.1016/0092-8674(84)90460-4. [DOI] [PubMed] [Google Scholar]
  37. Misfeldt D. S., Hamamoto S. T., Pitelka D. R. Transepithelial transport in cell culture. Proc Natl Acad Sci U S A. 1976 Apr;73(4):1212–1216. doi: 10.1073/pnas.73.4.1212. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Mooseker M. S. Organization, chemistry, and assembly of the cytoskeletal apparatus of the intestinal brush border. Annu Rev Cell Biol. 1985;1:209–241. doi: 10.1146/annurev.cb.01.110185.001233. [DOI] [PubMed] [Google Scholar]
  39. Moreno J. H., Diamond J. M. Cation permeation mechanisms and cation selectivity in "tight junctions" of gallbladder epithelium. Membranes. 1975;3:383–497. [PubMed] [Google Scholar]
  40. Nelson W. J., Veshnock P. J. Dynamics of membrane-skeleton (fodrin) organization during development of polarity in Madin-Darby canine kidney epithelial cells. J Cell Biol. 1986 Nov;103(5):1751–1765. doi: 10.1083/jcb.103.5.1751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Nitsch L., Wollman S. H. Ultrastructure of intermediate stages in polarity reversal of thyroid epithelium in follicles in suspension culture. J Cell Biol. 1980 Sep;86(3):875–880. doi: 10.1083/jcb.86.3.875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Pesonen M., Simons K. Transepithelial transport of a viral membrane glycoprotein implanted into the apical plasma membrane of Madin-Darby canine kidney cells. II. Immunological quantitation. J Cell Biol. 1983 Sep;97(3):638–643. doi: 10.1083/jcb.97.3.638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Pfeiffer S., Fuller S. D., Simons K. Intracellular sorting and basolateral appearance of the G protein of vesicular stomatitis virus in Madin-Darby canine kidney cells. J Cell Biol. 1985 Aug;101(2):470–476. doi: 10.1083/jcb.101.2.470. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Pisam M., Ripoche P. Redistribution of surface macromolecules in dissociated epithelial cells. J Cell Biol. 1976 Dec;71(3):907–920. doi: 10.1083/jcb.71.3.907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Pitelka D. R., Taggart B. N., Hamamoto S. T. Effects of extracellular calcium depletion on membrane topography and occluding junctions of mammary epithelial cells in culture. J Cell Biol. 1983 Mar;96(3):613–624. doi: 10.1083/jcb.96.3.613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Pitelka D. R., Taggart B. N. Mechanical tension induces lateral movement of intramembrane components of the tight junction: studies on mouse mammary cells in culture. J Cell Biol. 1983 Mar;96(3):606–612. doi: 10.1083/jcb.96.3.606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Rabito C. A., Tchao R. [3H]ouabain binding during the monolayer organization and cell cycle in MDCK cells. Am J Physiol. 1980 Jan;238(1):C43–C48. doi: 10.1152/ajpcell.1980.238.1.C43. [DOI] [PubMed] [Google Scholar]
  48. Rindler M. J., Ivanov I. E., Plesken H., Sabatini D. D. Polarized delivery of viral glycoproteins to the apical and basolateral plasma membranes of Madin-Darby canine kidney cells infected with temperature-sensitive viruses. J Cell Biol. 1985 Jan;100(1):136–151. doi: 10.1083/jcb.100.1.136. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Rodriguez Boulan E., Pendergast M. Polarized distribution of viral envelope proteins in the plasma membrane of infected epithelial cells. Cell. 1980 May;20(1):45–54. doi: 10.1016/0092-8674(80)90233-0. [DOI] [PubMed] [Google Scholar]
  50. Rodriguez Boulan E., Sabatini D. D. Asymmetric budding of viruses in epithelial monlayers: a model system for study of epithelial polarity. Proc Natl Acad Sci U S A. 1978 Oct;75(10):5071–5075. doi: 10.1073/pnas.75.10.5071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Rodriguez-Boulan E., Paskiet K. T., Sabatini D. D. Assembly of enveloped viruses in Madin-Darby canine kidney cells: polarized budding from single attached cells and from clusters of cells in suspension. J Cell Biol. 1983 Mar;96(3):866–874. doi: 10.1083/jcb.96.3.866. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Rodriguez-Boulan E. Polarized assembly of enveloped viruses from cultured epithelial cells. Methods Enzymol. 1983;98:486–501. doi: 10.1016/0076-6879(83)98176-4. [DOI] [PubMed] [Google Scholar]
  53. Salas P. J., Misek D. E., Vega-Salas D. E., Gundersen D., Cereijido M., Rodriguez-Boulan E. Microtubules and actin filaments are not critically involved in the biogenesis of epithelial cell surface polarity. J Cell Biol. 1986 May;102(5):1853–1867. doi: 10.1083/jcb.102.5.1853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Segrest J. P., Gulik-Krzywicki T., Sardet C. Association of the membrane-penetrating polypeptide segment of the human erythrocyte MN-glycoprotein with phospholipid bilayers. I. Formation of freeze-etch intramembranous particles. Proc Natl Acad Sci U S A. 1974 Aug;71(8):3294–3298. doi: 10.1073/pnas.71.8.3294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Simons K., Fuller S. D. Cell surface polarity in epithelia. Annu Rev Cell Biol. 1985;1:243–288. doi: 10.1146/annurev.cb.01.110185.001331. [DOI] [PubMed] [Google Scholar]
  56. Simons K., Garoff H. The budding mechanisms of enveloped animal viruses. J Gen Virol. 1980 Sep;50(1):1–21. doi: 10.1099/0022-1317-50-1-1. [DOI] [PubMed] [Google Scholar]
  57. Sugrue S. P., Hay E. D. Response of basal epithelial cell surface and Cytoskeleton to solubilized extracellular matrix molecules. J Cell Biol. 1981 Oct;91(1):45–54. doi: 10.1083/jcb.91.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Tillack T. W., Scott R. E., Marchesi V. T. The structure of erythrocyte membranes studied by freeze-etching. II. Localization of receptors for phytohemagglutinin and influenza virus to the intramembranous particles. J Exp Med. 1972 Jun 1;135(6):1209–1227. doi: 10.1084/jem.135.6.1209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. 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]
  60. Tsien R. Y. New calcium indicators and buffers with high selectivity against magnesium and protons: design, synthesis, and properties of prototype structures. Biochemistry. 1980 May 27;19(11):2396–2404. doi: 10.1021/bi00552a018. [DOI] [PubMed] [Google Scholar]
  61. Weibel E. R. Stereological principles for morphometry in electron microscopic cytology. Int Rev Cytol. 1969;26:235–302. doi: 10.1016/s0074-7696(08)61637-x. [DOI] [PubMed] [Google Scholar]
  62. White J., Kielian M., Helenius A. Membrane fusion proteins of enveloped animal viruses. Q Rev Biophys. 1983 May;16(2):151–195. doi: 10.1017/s0033583500005072. [DOI] [PubMed] [Google Scholar]
  63. Whittembury G., Rawlins F. A. Evidence of a paracellular pathway for ion flow in the kidney proximal tubule. Electromicroscopic demonstration of lanthanum precipitate in the tight junction. Pflugers Arch. 1971;330(4):302–309. doi: 10.1007/BF00588582. [DOI] [PubMed] [Google Scholar]
  64. Yelton D. E., Scharff M. D. Monoclonal antibodies: a powerful new tool in biology and medicine. Annu Rev Biochem. 1981;50:657–680. doi: 10.1146/annurev.bi.50.070181.003301. [DOI] [PubMed] [Google Scholar]
  65. Ziomek C. A., Schulman S., Edidin M. Redistribution of membrane proteins in isolated mouse intestinal epithelial cells. J Cell Biol. 1980 Sep;86(3):849–857. doi: 10.1083/jcb.86.3.849. [DOI] [PMC free article] [PubMed] [Google Scholar]
  66. van Meer G., Simons K. The function of tight junctions in maintaining differences in lipid composition between the apical and the basolateral cell surface domains of MDCK cells. EMBO J. 1986 Jul;5(7):1455–1464. doi: 10.1002/j.1460-2075.1986.tb04382.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES