Cingulin Contains Globular and Coiled-Coil Domains and Interacts with Zo-1, Zo-2, Zo-3, and Myosin
- Cordenonsi, Michelangelo Department of Biology, University of Padova, 35121 Padova, Italy
- D'Atri, Fabio Department of Molecular Biology, University of Geneva, 1211 Geneva 4, Switzerland
- Hammar, Eva Department of Molecular Biology, University of Geneva, 1211 Geneva 4, Switzerland
- Parry, David A.D. Institute of Fundamental Sciences, Massey University, Palmerston North, New Zealand
- Kendrick-Jones, John Medical Research Council Laboratory of Molecular Biology, Cambridge, CB22QH UK
- Shore, David Department of Molecular Biology, University of Geneva, 1211 Geneva 4, Switzerland
- Citi, Sandra Department of Molecular Biology, University of Geneva, 1211 Geneva 4, Switzerland
- 1999-12-27
Published in:
- Journal of Cell Biology. - Rockefeller University Press. - 1999, vol. 147, no. 7, p. 1569-1582
English
We characterized the sequence and protein interactions of cingulin, an Mr 140–160-kD phosphoprotein localized on the cytoplasmic surface of epithelial tight junctions (TJ). The derived amino acid sequence of a full-length Xenopus laevis cingulin cDNA shows globular head (residues 1–439) and tail (1,326–1,368) domains and a central α-helical rod domain (440–1,325). Sequence analysis, electron microscopy, and pull-down assays indicate that the cingulin rod is responsible for the formation of coiled-coil parallel dimers, which can further aggregate through intermolecular interactions. Pull-down assays from epithelial, insect cell, and reticulocyte lysates show that an NH2-terminal fragment of cingulin (1–378) interacts in vitro with ZO-1 (Kd ∼5 nM), ZO-2, ZO-3, myosin, and AF-6, but not with symplekin, and a COOH-terminal fragment (377–1,368) interacts with myosin and ZO-3. ZO-1 and ZO-2 immunoprecipitates contain cingulin, suggesting in vivo interactions. Full-length cingulin, but not NH2-terminal and COOH-terminal fragments, colocalizes with endogenous cingulin in transfected MDCK cells, indicating that sequences within both head and rod domains are required for TJ localization. We propose that cingulin is a functionally important component of TJ, linking the submembrane plaque domain of TJ to the actomyosin cytoskeleton.
- Language
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- English
- Open access status
- bronze
- Identifiers
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- DOI 10.1083/jcb.147.7.1569
- ISSN 0021-9525
- Persistent URL
- https://sonar.ch/global/documents/149346
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