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. 1986 May 1;102(5):1832–1842. doi: 10.1083/jcb.102.5.1832

Changes in membrane-microfilament interaction in intercellular adherens junctions upon removal of extracellular Ca2+ ions

PMCID: PMC2114216  PMID: 3084500

Abstract

EGTA-induced depletion of Ca2+ ions from the culture medium of Madin- Darby bovine kidney epithelial cells results in rapid splitting of adherens-type junctions and the detachment of the vinculin- and actin- containing filament bundle from the cytoplasmic faces of the plasma membrane of the zonula adhaerens. This process was monitored by phase- contrast microscopy, combined with electron microscopy and immunofluorescent localization of the two proteins. It is shown that shortly after extracellular free Ca2+ concentration is lowered to the micromolar range, the actin-containing, junction-associated belt of microfilaments, together with the vinculin-rich junctional plaque material, is irreversibly detached as one structural unit from the plasma membrane, contracts, and is displaced towards the perinuclear cytoplasm where it gradually disintegrates. Other actin- and vinculin- containing structures present in the same cells, notably the focal contacts at the substratum, are not similarly affected by the Ca2+ depletion and retain both the adhesion to the external surface and the association with the plaque and microfilament components. Electron microscopic examination has shown that the membrane domain of the zonulae adhaerentes, unlike that of desmosomes, is not endocytosed after Ca2+ removal and that the displaced actin- and vinculin- containing plaque and filament belt are not associated with a particular membrane. It is further shown that upon restoration of normal Ca2+ levels in the culture medium, new intercellular contacts are established gradually by accretion of both vinculin and actin into new belt-like plaque- and microfilament-containing structures.

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Selected References

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  1. Abercrombie M., Dunn G. A. Adhesions of fibroblasts to substratum during contact inhibition observed by interference reflection microscopy. Exp Cell Res. 1975 Apr;92(1):57–62. doi: 10.1016/0014-4827(75)90636-9. [DOI] [PubMed] [Google Scholar]
  2. Avnur Z., Small J. V., Geiger B. Actin-independent association of vinculin with the cytoplasmic aspect of the plasma membrane in cell-contact areas. J Cell Biol. 1983 Jun;96(6):1622–1630. doi: 10.1083/jcb.96.6.1622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boller K., Vestweber D., Kemler R. Cell-adhesion molecule uvomorulin is localized in the intermediate junctions of adult intestinal epithelial cells. J Cell Biol. 1985 Jan;100(1):327–332. doi: 10.1083/jcb.100.1.327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Burridge K., Connell L. A new protein of adhesion plaques and ruffling membranes. J Cell Biol. 1983 Aug;97(2):359–367. doi: 10.1083/jcb.97.2.359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Burridge K., Connell L. Talin: a cytoskeletal component concentrated in adhesion plaques and other sites of actin-membrane interaction. Cell Motil. 1983;3(5-6):405–417. doi: 10.1002/cm.970030509. [DOI] [PubMed] [Google Scholar]
  6. Burridge K., Feramisco J. R. Microinjection and localization of a 130K protein in living fibroblasts: a relationship to actin and fibronectin. Cell. 1980 Mar;19(3):587–595. doi: 10.1016/s0092-8674(80)80035-3. [DOI] [PubMed] [Google Scholar]
  7. CURTIS A. S. THE MECHANISM OF ADHESION OF CELLS TO GLASS. A STUDY BY INTERFERENCE REFLECTION MICROSCOPY. J Cell Biol. 1964 Feb;20:199–215. doi: 10.1083/jcb.20.2.199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Damsky C. H., Richa J., Solter D., Knudsen K., Buck C. A. Identification and purification of a cell surface glycoprotein mediating intercellular adhesion in embryonic and adult tissue. Cell. 1983 Sep;34(2):455–466. doi: 10.1016/0092-8674(83)90379-3. [DOI] [PubMed] [Google Scholar]
  9. 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]
  10. Galli P., Brenna A., Camilli de P., Meldolesi J. Extracellular calcium and the organization of tight junctions in pancreatic acinar cells. Exp Cell Res. 1976 Apr;99(1):178–183. doi: 10.1016/0014-4827(76)90694-7. [DOI] [PubMed] [Google Scholar]
  11. Gallin W. J., Edelman G. M., Cunningham B. A. Characterization of L-CAM, a major cell adhesion molecule from embryonic liver cells. Proc Natl Acad Sci U S A. 1983 Feb;80(4):1038–1042. doi: 10.1073/pnas.80.4.1038. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Geiger B. A 130K protein from chicken gizzard: its localization at the termini of microfilament bundles in cultured chicken cells. Cell. 1979 Sep;18(1):193–205. doi: 10.1016/0092-8674(79)90368-4. [DOI] [PubMed] [Google Scholar]
  13. Geiger B., Avnur Z., Kreis T. E., Schlessinger J. The dynamics of cytoskeletal organization in areas of cell contact. Cell Muscle Motil. 1984;5:195–234. doi: 10.1007/978-1-4684-4592-3_5. [DOI] [PubMed] [Google Scholar]
  14. Geiger B., Avnur Z., Rinnerthaler G., Hinssen H., Small V. J. Microfilament-organizing centers in areas of cell contact: cytoskeletal interactions during cell attachment and locomotion. J Cell Biol. 1984 Jul;99(1 Pt 2):83s–91s. doi: 10.1083/jcb.99.1.83s. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Geiger B., Dutton A. H., Tokuyasu K. T., Singer S. J. Immunoelectron microscope studies of membrane-microfilament interactions: distributions of alpha-actinin, tropomyosin, and vinculin in intestinal epithelial brush border and chicken gizzard smooth muscle cells. J Cell Biol. 1981 Dec;91(3 Pt 1):614–628. doi: 10.1083/jcb.91.3.614. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Geiger B. Involvement of vinculin in contact-induced cytoskeletal interactions. Cold Spring Harb Symp Quant Biol. 1982;46(Pt 2):671–682. doi: 10.1101/sqb.1982.046.01.063. [DOI] [PubMed] [Google Scholar]
  17. Geiger B. Membrane-cytoskeleton interaction. Biochim Biophys Acta. 1983 Aug 11;737(3-4):305–341. doi: 10.1016/0304-4157(83)90005-9. [DOI] [PubMed] [Google Scholar]
  18. Geiger B., Schmid E., Franke W. W. Spatial distribution of proteins specific for desmosomes and adhaerens junctions in epithelial cells demonstrated by double immunofluorescence microscopy. Differentiation. 1983;23(3):189–205. doi: 10.1111/j.1432-0436.1982.tb01283.x. [DOI] [PubMed] [Google Scholar]
  19. Geiger B., Tokuyasu K. T., Dutton A. H., Singer S. J. Vinculin, an intracellular protein localized at specialized sites where microfilament bundles terminate at cell membranes. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4127–4131. doi: 10.1073/pnas.77.7.4127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Geiger B., Volk T., Volberg T. Molecular heterogeneity of adherens junctions. J Cell Biol. 1985 Oct;101(4):1523–1531. doi: 10.1083/jcb.101.4.1523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Heaysman J. E., Pegrum S. M. Early contacts between fibroblasts. An ultrastructural study. Exp Cell Res. 1973 Mar 30;78(1):71–78. doi: 10.1016/0014-4827(73)90039-6. [DOI] [PubMed] [Google Scholar]
  22. Hyafil F., Babinet C., Jacob F. Cell-cell interactions in early embryogenesis: a molecular approach to the role of calcium. Cell. 1981 Nov;26(3 Pt 1):447–454. doi: 10.1016/0092-8674(81)90214-2. [DOI] [PubMed] [Google Scholar]
  23. Izzard C. S., Lochner L. R. Cell-to-substrate contacts in living fibroblasts: an interference reflexion study with an evaluation of the technique. J Cell Sci. 1976 Jun;21(1):129–159. doi: 10.1242/jcs.21.1.129. [DOI] [PubMed] [Google Scholar]
  24. Jones J. C., Goldman A. E., Steinert P. M., Yuspa S., Goldman R. D. Dynamic aspects of the supramolecular organization of intermediate filament networks in cultured epidermal cells. Cell Motil. 1982;2(3):197–213. doi: 10.1002/cm.970020302. [DOI] [PubMed] [Google Scholar]
  25. Kartenbeck J., Schmid E., Franke W. W., Geiger B. Different modes of internalization of proteins associated with adhaerens junctions and desmosomes: experimental separation of lateral contacts induces endocytosis of desmosomal plaque material. EMBO J. 1982;1(6):725–732. doi: 10.1002/j.1460-2075.1982.tb01237.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lazarides E., Burridge K. Alpha-actinin: immunofluorescent localization of a muscle structural protein in nonmuscle cells. Cell. 1975 Nov;6(3):289–298. doi: 10.1016/0092-8674(75)90180-4. [DOI] [PubMed] [Google Scholar]
  27. Lehto V. P., Virtanen I. Vinculin in cultured bovine lens-forming cells. Cell Differ. 1985 May;16(3):153–160. doi: 10.1016/0045-6039(85)90512-3. [DOI] [PubMed] [Google Scholar]
  28. Palant C. E., Duffey M. E., Mookerjee B. K., Ho S., Bentzel C. J. Ca2+ regulation of tight-junction permeability and structure in Necturus gallbladder. Am J Physiol. 1983 Sep;245(3):C203–C212. doi: 10.1152/ajpcell.1983.245.3.C203. [DOI] [PubMed] [Google Scholar]
  29. Philp N. J., Nachmias V. T. Components of the cytoskeleton in the retinal pigmented epithelium of the chick. J Cell Biol. 1985 Aug;101(2):358–362. doi: 10.1083/jcb.101.2.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. 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]
  31. Staehelin L. A. Structure and function of intercellular junctions. Int Rev Cytol. 1974;39:191–283. doi: 10.1016/s0074-7696(08)60940-7. [DOI] [PubMed] [Google Scholar]
  32. Tokuyasu K. T., Dutton A. H., Geiger B., Singer S. J. Ultrastructure of chicken cardiac muscle as studied by double immunolabeling in electron microscopy. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7619–7623. doi: 10.1073/pnas.78.12.7619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Volk T., Geiger B. A 135-kd membrane protein of intercellular adherens junctions. EMBO J. 1984 Oct;3(10):2249–2260. doi: 10.1002/j.1460-2075.1984.tb02123.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Yoshida-Noro C., Suzuki N., Takeichi M. Molecular nature of the calcium-dependent cell-cell adhesion system in mouse teratocarcinoma and embryonic cells studied with a monoclonal antibody. Dev Biol. 1984 Jan;101(1):19–27. doi: 10.1016/0012-1606(84)90112-x. [DOI] [PubMed] [Google Scholar]

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