Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1991 May 2;113(4):881–892. doi: 10.1083/jcb.113.4.881

Endocytosis of junctional cadherins in bovine kidney epithelial (MDBK) cells cultured in low Ca2+ ion medium

PMCID: PMC2288996  PMID: 2026652

Abstract

The release of intercellular contacts in MDBK cells, initiated by the depletion of Ca2+ ions from the culture medium, results in the endocytotic uptake of membrane vesicles containing specific membrane constituents of the zonula adhaerens (ZA). During this process the junction-derived, endocytosed vesicles remain associated with the ZA plaque components, while the plaque and its attached actin filaments retract as a whole in a ring-like fashion from the plasma membrane, often accumulating, usually in fragments, in the juxtanuclear cytoplasm. Double-label immunofluorescence microscopy with antiplakoglobin and antivinculin has indicated that both plaque proteins colocalize with the hallmark membrane glycoprotein of this junction type, E-cadherin (uvomorulin). When HRP used as a fluid phase marker is applied to the culture medium, simultaneously with the Ca2+ ion-chelator EGTA, numerous HRP-positive vesicles are found in close association with the dislocated plaque material, suggesting that the HRP is contained in the vesicles formed upon EGTA-induced junction splitting. Immunoelectron microscopy with various cadherin-specific antibodies revealed vesicle-associated labeling, confirming the derivation of these plaque-associated vesicles from the ZA. As the desmosome-specific cadherin, desmoglein, is recovered in another type of junction-derived vesicle, which is characterized by its association with a desmoplakin-plaque, we conclude that the membrane domains of both kinds of junction are endocytosed during Ca2+ depletion but stay in different vesicle populations, emphasizing the selective interaction of the specific cadherins with their respective plaque and filament partners.

Full Text

The Full Text of this article is available as a PDF (5.9 MB).

Selected References

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

  1. Beckerle M. C. Identification of a new protein localized at sites of cell-substrate adhesion. J Cell Biol. 1986 Nov;103(5):1679–1687. doi: 10.1083/jcb.103.5.1679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Behrens J., Birchmeier W., Goodman S. L., Imhof B. A. Dissociation of Madin-Darby canine kidney epithelial cells by the monoclonal antibody anti-arc-1: mechanistic aspects and identification of the antigen as a component related to uvomorulin. J Cell Biol. 1985 Oct;101(4):1307–1315. doi: 10.1083/jcb.101.4.1307. [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. Cohen S. M., Gorbsky G., Steinberg M. S. Immunochemical characterization of related families of glycoproteins in desmosomes. J Biol Chem. 1983 Feb 25;258(4):2621–2627. [PubMed] [Google Scholar]
  5. Cowin P., Garrod D. R. Antibodies to epithelial desmosomes show wide tissue and species cross-reactivity. Nature. 1983 Mar 10;302(5904):148–150. doi: 10.1038/302148a0. [DOI] [PubMed] [Google Scholar]
  6. Cowin P., Kapprell H. P., Franke W. W., Tamkun J., Hynes R. O. Plakoglobin: a protein common to different kinds of intercellular adhering junctions. Cell. 1986 Sep 26;46(7):1063–1073. doi: 10.1016/0092-8674(86)90706-3. [DOI] [PubMed] [Google Scholar]
  7. Cowin P., Kapprell H. P., Franke W. W. The complement of desmosomal plaque proteins in different cell types. J Cell Biol. 1985 Oct;101(4):1442–1454. doi: 10.1083/jcb.101.4.1442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cowin P., Mattey D., Garrod D. Distribution of desmosomal components in the tissues of vertebrates, studied by fluorescent antibody staining. J Cell Sci. 1984 Mar;66:119–132. doi: 10.1242/jcs.66.1.119. [DOI] [PubMed] [Google Scholar]
  9. Cowin P., Mattey D., Garrod D. Identification of desmosomal surface components (desmocollins) and inhibition of desmosome formation by specific Fab'. J Cell Sci. 1984 Aug;70:41–60. doi: 10.1242/jcs.70.1.41. [DOI] [PubMed] [Google Scholar]
  10. Drenckhahn D., Franz H. Identification of actin-, alpha-actinin-, and vinculin-containing plaques at the lateral membrane of epithelial cells. J Cell Biol. 1986 May;102(5):1843–1852. doi: 10.1083/jcb.102.5.1843. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Duden R., Franke W. W. Organization of desmosomal plaque proteins in cells growing at low calcium concentrations. J Cell Biol. 1988 Sep;107(3):1049–1063. doi: 10.1083/jcb.107.3.1049. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Edelman G. M. Cell adhesion molecules. Science. 1983 Feb 4;219(4584):450–457. doi: 10.1126/science.6823544. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Franke W. W., Kapprell H. P., Cowin P. Immunolocalization of plakoglobin in endothelial junctions: identification as a special type of Zonulae adhaerentes. Biol Cell. 1987;59(3):205–218. doi: 10.1111/j.1768-322x.1987.tb00532.x. [DOI] [PubMed] [Google Scholar]
  15. Franke W. W., Lüder M. R., Kartenbeck J., Zerban H., Keenan T. W. Involvement of vesicle coat material in casein secretion and surface regeneration. J Cell Biol. 1976 Apr;69(1):173–195. doi: 10.1083/jcb.69.1.173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Franke W. W., Moll R., Schiller D. L., Schmid E., Kartenbeck J., Mueller H. Desmoplakins of epithelial and myocardial desmosomes are immunologically and biochemically related. Differentiation. 1982;23(2):115–127. doi: 10.1111/j.1432-0436.1982.tb01274.x. [DOI] [PubMed] [Google Scholar]
  17. Franke W. W., Mueller H., Mittnacht S., Kapprell H. P., Jorcano J. L. Significance of two desmosome plaque-associated polypeptides of molecular weights 75 000 and 83 000. EMBO J. 1983;2(12):2211–2215. doi: 10.1002/j.1460-2075.1983.tb01725.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. 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]
  19. Gallin W. J., Sorkin B. C., Edelman G. M., Cunningham B. A. Sequence analysis of a cDNA clone encoding the liver cell adhesion molecule, L-CAM. Proc Natl Acad Sci U S A. 1987 May;84(9):2808–2812. doi: 10.1073/pnas.84.9.2808. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. 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]
  22. 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]
  23. Geiger B., Volberg T., Ginsberg D., Bitzur S., Sabanay I., Hynes R. O. Broad spectrum pan-cadherin antibodies, reactive with the C-terminal 24 amino acid residues of N-cadherin. J Cell Sci. 1990 Dec;97(Pt 4):607–614. doi: 10.1242/jcs.97.4.607. [DOI] [PubMed] [Google Scholar]
  24. 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]
  25. Giudice G. J., Cohen S. M., Patel N. H., Steinberg M. S. Immunological comparison of desmosomal components from several bovine tissues. J Cell Biochem. 1984;26(1):35–45. doi: 10.1002/jcb.240260104. [DOI] [PubMed] [Google Scholar]
  26. Gorbsky G., Steinberg M. S. Isolation of the intercellular glycoproteins of desmosomes. J Cell Biol. 1981 Jul;90(1):243–248. doi: 10.1083/jcb.90.1.243. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Graham R. C., Jr, Karnovsky M. J. The early stages of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney: ultrastructural cytochemistry by a new technique. J Histochem Cytochem. 1966 Apr;14(4):291–302. doi: 10.1177/14.4.291. [DOI] [PubMed] [Google Scholar]
  28. Green K. J., Parry D. A., Steinert P. M., Virata M. L., Wagner R. M., Angst B. D., Nilles L. A. Structure of the human desmoplakins. Implications for function in the desmosomal plaque. J Biol Chem. 1990 Feb 15;265(5):2603–2612. [PubMed] [Google Scholar]
  29. Hatta K., Nose A., Nagafuchi A., Takeichi M. Cloning and expression of cDNA encoding a neural calcium-dependent cell adhesion molecule: its identity in the cadherin gene family. J Cell Biol. 1988 Mar;106(3):873–881. doi: 10.1083/jcb.106.3.873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. 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]
  31. Kapprell H. P., Cowin P., Franke W. W. Biochemical characterization of the soluble form of the junctional plaque protein, plakoglobin, from different cell types. Eur J Biochem. 1987 Aug 3;166(3):505–517. doi: 10.1111/j.1432-1033.1987.tb13543.x. [DOI] [PubMed] [Google Scholar]
  32. Kapprell H. P., Cowin P., Franke W. W., Ponstingl H., Opferkuch H. J. Biochemical characterization of desmosomal proteins isolated from bovine muzzle epidermis: amino acid and carbohydrate composition. Eur J Cell Biol. 1985 Mar;36(2):217–229. [PubMed] [Google Scholar]
  33. Kartenbeck J., Franke W. W., Moser J. G., Stoffels U. Specific attachment of desmin filaments to desmosomal plaques in cardiac myocytes. EMBO J. 1983;2(5):735–742. doi: 10.1002/j.1460-2075.1983.tb01493.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. 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]
  35. Koch P. J., Walsh M. J., Schmelz M., Goldschmidt M. D., Zimbelmann R., Franke W. W. Identification of desmoglein, a constitutive desmosomal glycoprotein, as a member of the cadherin family of cell adhesion molecules. Eur J Cell Biol. 1990 Oct;53(1):1–12. [PubMed] [Google Scholar]
  36. MADIN S. H., DARBY N. B., Jr Established kidney cell lines of normal adult bovine and ovine origin. Proc Soc Exp Biol Med. 1958 Jul;98(3):574–576. doi: 10.3181/00379727-98-24111. [DOI] [PubMed] [Google Scholar]
  37. Mattey D. L., Garrod D. R. Splitting and internalization of the desmosomes of cultured kidney epithelial cells by reduction in calcium concentration. J Cell Sci. 1986 Sep;85:113–124. doi: 10.1242/jcs.85.1.113. [DOI] [PubMed] [Google Scholar]
  38. Mueller H., Franke W. W. Biochemical and immunological characterization of desmoplakins I and II, the major polypeptides of the desmosomal plaque. J Mol Biol. 1983 Feb 5;163(4):647–671. doi: 10.1016/0022-2836(83)90116-x. [DOI] [PubMed] [Google Scholar]
  39. Nagafuchi A., Takeichi M. Cell binding function of E-cadherin is regulated by the cytoplasmic domain. EMBO J. 1988 Dec 1;7(12):3679–3684. doi: 10.1002/j.1460-2075.1988.tb03249.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Nagafuchi A., Takeichi M. Transmembrane control of cadherin-mediated cell adhesion: a 94 kDa protein functionally associated with a specific region of the cytoplasmic domain of E-cadherin. Cell Regul. 1989 Nov;1(1):37–44. doi: 10.1091/mbc.1.1.37. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Ringwald M., Schuh R., Vestweber D., Eistetter H., Lottspeich F., Engel J., Dölz R., Jähnig F., Epplen J., Mayer S. The structure of cell adhesion molecule uvomorulin. Insights into the molecular mechanism of Ca2+-dependent cell adhesion. EMBO J. 1987 Dec 1;6(12):3647–3653. doi: 10.1002/j.1460-2075.1987.tb02697.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Schmelz M., Duden R., Cowin P., Franke W. W. A constitutive transmembrane glycoprotein of Mr 165,000 (desmoglein) in epidermal and non-epidermal desmosomes. I. Biochemical identification of the polypeptide. Eur J Cell Biol. 1986 Dec;42(2):177–183. [PubMed] [Google Scholar]
  43. Schmelz M., Duden R., Cowin P., Franke W. W. A constitutive transmembrane glycoprotein of Mr 165,000 (desmoglein) in epidermal and non-epidermal desmosomes. II. Immunolocalization and microinjection studies. Eur J Cell Biol. 1986 Dec;42(2):184–199. [PubMed] [Google Scholar]
  44. Schwarz M. A., Owaribe K., Kartenbeck J., Franke W. W. Desmosomes and hemidesmosomes: constitutive molecular components. Annu Rev Cell Biol. 1990;6:461–491. doi: 10.1146/annurev.cb.06.110190.002333. [DOI] [PubMed] [Google Scholar]
  45. 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]
  46. Steinberg M. S., Shida H., Giudice G. J., Shida M., Patel N. H., Blaschuk O. W. On the molecular organization, diversity and functions of desmosomal proteins. Ciba Found Symp. 1987;125:3–25. doi: 10.1002/9780470513408.ch2. [DOI] [PubMed] [Google Scholar]
  47. Takeichi M. Functional correlation between cell adhesive properties and some cell surface proteins. J Cell Biol. 1977 Nov;75(2 Pt 1):464–474. doi: 10.1083/jcb.75.2.464. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Takeichi M. The cadherins: cell-cell adhesion molecules controlling animal morphogenesis. Development. 1988 Apr;102(4):639–655. doi: 10.1242/dev.102.4.639. [DOI] [PubMed] [Google Scholar]
  49. Tsukita S., Tsukita S., Nagafuchi A. The undercoat of adherens junctions: a key specialized structure in organogenesis and carcinogenesis. Cell Struct Funct. 1990 Feb;15(1):7–12. doi: 10.1247/csf.15.7. [DOI] [PubMed] [Google Scholar]
  50. Vestweber D., Gossler A., Boller K., Kemler R. Expression and distribution of cell adhesion molecule uvomorulin in mouse preimplantation embryos. Dev Biol. 1987 Dec;124(2):451–456. doi: 10.1016/0012-1606(87)90498-2. [DOI] [PubMed] [Google Scholar]
  51. Vestweber D., Kemler R. Some structural and functional aspects of the cell adhesion molecule uvomorulin. Cell Differ. 1984 Dec;15(2-4):269–273. doi: 10.1016/0045-6039(84)90084-8. [DOI] [PubMed] [Google Scholar]
  52. Volberg T., Geiger B., Kartenbeck J., Franke W. W. Changes in membrane-microfilament interaction in intercellular adherens junctions upon removal of extracellular Ca2+ ions. J Cell Biol. 1986 May;102(5):1832–1842. doi: 10.1083/jcb.102.5.1832. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Yoshida C., Takeichi M. Teratocarcinoma cell adhesion: identification of a cell-surface protein involved in calcium-dependent cell aggregation. Cell. 1982 Feb;28(2):217–224. doi: 10.1016/0092-8674(82)90339-7. [DOI] [PubMed] [Google Scholar]

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

RESOURCES