• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jcellbiolHomeThe Rockefeller University PressEditorsContactInstructions for AuthorsThis issue
J Cell Biol. Dec 2, 1994; 127(6): 1617–1626.
PMCID: PMC2120300

Direct association of occludin with ZO-1 and its possible involvement in the localization of occludin at tight junctions

Abstract

Occludin is an integral membrane protein localizing at tight junctions (TJ) with four transmembrane domains and a long COOH-terminal cytoplasmic domain (domain E) consisting of 255 amino acids. Immunofluorescence and laser scan microscopy revealed that chick full- length occludin introduced into human and bovine epithelial cells was correctly delivered to and incorporated into preexisting TJ. Further transfection studies with various deletion mutants showed that the domain E, especially its COOH-terminal approximately 150 amino acids (domain E358/504), was necessary for the localization of occludin at TJ. Secondly, domain E was expressed in Escherichia coli as a fusion protein with glutathione-S-transferase, and this fusion protein was shown to be specifically bound to a complex of ZO-1 (220 kD) and ZO-2 (160 kD) among various membrane peripheral proteins. In vitro binding analyses using glutathione-S-transferase fusion proteins of various deletion mutants of domain E narrowed down the sequence necessary for the ZO-1/ZO-2 association into the domain E358/504. Furthermore, this region directly associated with the recombinant ZO-1 produced in E. coli. We concluded that occludin itself can localize at TJ and directly associate with ZO-1. The coincidence of the sequence necessary for the ZO-1 association with that for the TJ localization suggests that the association with underlying cytoskeletons through ZO-1 is required for occludin to be localized at TJ.

Full Text

The Full Text of this article is available as a PDF (2.7M).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Anderson JM, Balda MS, Fanning AS. The structure and regulation of tight junctions. Curr Opin Cell Biol. 1993 Oct;5(5):772–778. [PubMed]
  • Anderson JM, Stevenson BR, Jesaitis LA, Goodenough DA, Mooseker MS. Characterization of ZO-1, a protein component of the tight junction from mouse liver and Madin-Darby canine kidney cells. J Cell Biol. 1988 Apr;106(4):1141–1149. [PMC free article] [PubMed]
  • Bourne HR. Do GTPases direct membrane traffic in secretion? Cell. 1988 Jun 3;53(5):669–671. [PubMed]
  • Citi S. The molecular organization of tight junctions. J Cell Biol. 1993 May;121(3):485–489. [PMC free article] [PubMed]
  • Citi S, Sabanay H, Jakes R, Geiger B, Kendrick-Jones J. Cingulin, a new peripheral component of tight junctions. Nature. 1988 May 19;333(6170):272–276. [PubMed]
  • Furuse M, Hirase T, Itoh M, Nagafuchi A, Yonemura S, Tsukita S, Tsukita S. Occludin: a novel integral membrane protein localizing at tight junctions. J Cell Biol. 1993 Dec;123(6 Pt 2):1777–1788. [PMC free article] [PubMed]
  • Goud B, McCaffrey M. Small GTP-binding proteins and their role in transport. Curr Opin Cell Biol. 1991 Aug;3(4):626–633. [PubMed]
  • Gumbiner B. Structure, biochemistry, and assembly of epithelial tight junctions. Am J Physiol. 1987 Dec;253(6 Pt 1):C749–C758. [PubMed]
  • Gumbiner B, Lowenkopf T, Apatira D. Identification of a 160-kDa polypeptide that binds to the tight junction protein ZO-1. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3460–3464. [PMC free article] [PubMed]
  • Gumbiner BM. Breaking through the tight junction barrier. J Cell Biol. 1993 Dec;123(6 Pt 2):1631–1633. [PMC free article] [PubMed]
  • Howarth AG, Hughes MR, Stevenson BR. Detection of the tight junction-associated protein ZO-1 in astrocytes and other nonepithelial cell types. Am J Physiol. 1992 Feb;262(2 Pt 1):C461–C469. [PubMed]
  • Howarth AG, Singer KL, Stevenson BR. Analysis of the distribution and phosphorylation state of ZO-1 in MDCK and nonepithelial cells. J Membr Biol. 1994 Feb;137(3):261–270. [PubMed]
  • Itoh M, Nagafuchi A, Yonemura S, Kitani-Yasuda T, Tsukita S, Tsukita S. The 220-kD protein colocalizing with cadherins in non-epithelial cells is identical to ZO-1, a tight junction-associated protein in epithelial cells: cDNA cloning and immunoelectron microscopy. J Cell Biol. 1993 May;121(3):491–502. [PMC free article] [PubMed]
  • Itoh M, Yonemura S, Nagafuchi A, Tsukita S, Tsukita S. A 220-kD undercoat-constitutive protein: its specific localization at cadherin-based cell-cell adhesion sites. J Cell Biol. 1991 Dec;115(5):1449–1462. [PMC free article] [PubMed]
  • Jesaitis LA, Goodenough DA. Molecular characterization and tissue distribution of ZO-2, a tight junction protein homologous to ZO-1 and the Drosophila discs-large tumor suppressor protein. J Cell Biol. 1994 Mar;124(6):949–961. [PMC free article] [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Madara JL. Intestinal absorptive cell tight junctions are linked to cytoskeleton. Am J Physiol. 1987 Jul;253(1 Pt 1):C171–C175. [PubMed]
  • Nose A, Tsuji K, Takeichi M. Localization of specificity determining sites in cadherin cell adhesion molecules. Cell. 1990 Apr 6;61(1):147–155. [PubMed]
  • Rodriguez-Boulan E, Nelson WJ. Morphogenesis of the polarized epithelial cell phenotype. Science. 1989 Aug 18;245(4919):718–725. [PubMed]
  • Schneeberger EE, Lynch RD. Structure, function, and regulation of cellular tight junctions. Am J Physiol. 1992 Jun;262(6 Pt 1):L647–L661. [PubMed]
  • Stevenson BR, Heintzelman MB, Anderson JM, Citi S, Mooseker MS. ZO-1 and cingulin: tight junction proteins with distinct identities and localizations. Am J Physiol. 1989 Oct;257(4 Pt 1):C621–C628. [PubMed]
  • Stevenson BR, Siliciano JD, Mooseker MS, Goodenough DA. Identification of ZO-1: a high molecular weight polypeptide associated with the tight junction (zonula occludens) in a variety of epithelia. J Cell Biol. 1986 Sep;103(3):755–766. [PMC free article] [PubMed]
  • Tsukita S, Itoh M, Nagafuchi A, Yonemura S, Tsukita S. Submembranous junctional plaque proteins include potential tumor suppressor molecules. J Cell Biol. 1993 Dec;123(5):1049–1053. [PMC free article] [PubMed]
  • Tsukita S, Itoh M, Tsukita S. A new 400-kD protein from isolated adherens junctions: its localization at the undercoat of adherens junctions and at microfilament bundles such as stress fibers and circumferential bundles. J Cell Biol. 1989 Dec;109(6 Pt 1):2905–2915. [PMC free article] [PubMed]
  • Tsukita S, Tsukita S, Nagafuchi A, Yonemura S. Molecular linkage between cadherins and actin filaments in cell-cell adherens junctions. Curr Opin Cell Biol. 1992 Oct;4(5):834–839. [PubMed]
  • Weber E, Berta G, Tousson A, St John P, Green MW, Gopalokrishnan U, Jilling T, Sorscher EJ, Elton TS, Abrahamson DR, et al. Expression and polarized targeting of a rab3 isoform in epithelial cells. J Cell Biol. 1994 May;125(3):583–594. [PMC free article] [PubMed]
  • Willott E, Balda MS, Fanning AS, Jameson B, Van Itallie C, Anderson JM. The tight junction protein ZO-1 is homologous to the Drosophila discs-large tumor suppressor protein of septate junctions. Proc Natl Acad Sci U S A. 1993 Aug 15;90(16):7834–7838. [PMC free article] [PubMed]
  • Zahraoui A, Joberty G, Arpin M, Fontaine JJ, Hellio R, Tavitian A, Louvard D. A small rab GTPase is distributed in cytoplasmic vesicles in non polarized cells but colocalizes with the tight junction marker ZO-1 in polarized epithelial cells. J Cell Biol. 1994 Jan;124(1-2):101–115. [PMC free article] [PubMed]
  • Zerial M, Stenmark H. Rab GTPases in vesicular transport. Curr Opin Cell Biol. 1993 Aug;5(4):613–620. [PubMed]
  • Zhong Y, Saitoh T, Minase T, Sawada N, Enomoto K, Mori M. Monoclonal antibody 7H6 reacts with a novel tight junction-associated protein distinct from ZO-1, cingulin and ZO-2. J Cell Biol. 1993 Jan;120(2):477–483. [PMC free article] [PubMed]

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

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...