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© The Rockefeller University Press, 0021-9525/1998//755 $5.00
The Journal of Cell Biology, Volume 141, Number 3, , 1998 755-764

Articles

Vinculin Is Part of the Cadherin–Catenin Junctional Complex: Complex Formation between {alpha}-Catenin and Vinculin



Elisabeth E. Weiss*, Martina Kroemker*, Angelika-H. Rüdiger{ddagger}, Brigitte M. Jockusch*, and Manfred Rüdiger*

* Cell Biology, Zoological Institute, Technical University, 38092 Braunschweig, Germany; and {ddagger} Division of Cell Biology and Immunology, National Research Centre for Biotechnology, 38124 Braunschweig, Germany

In epithelial cells, {alpha}-, β-, and {gamma}-catenin are involved in linking the peripheral microfilament belt to the transmembrane protein E-cadherin. {alpha}-Catenin exhibits sequence homologies over three regions to vinculin, another adherens junction protein. While vinculin is found in cell–matrix and cell–cell contacts, {alpha}-catenin is restricted to the latter. To elucidate, whether vinculin is part of the cell–cell junctional complex, we investigated complex formation and intracellular targeting of vinculin and {alpha}-catenin. We show that {alpha}-catenin colocalizes at cell–cell contacts with endogenous vinculin and also with the transfected vinculin head domain forming immunoprecipitable complexes. In vitro, the vinculin NH2-terminal head binds to {alpha}-catenin, as seen by immunoprecipitation, dot overlay, cosedimentation, and surface plasmon resonance measurements. The Kd of the complex was determined to 2–4 x 10–7 M. As seen by overlays and affinity mass spectrometry, the COOH-terminal region of {alpha}-catenin is involved in this interaction.

Complex formation of vinculin and {alpha}-catenin was challenged in transfected cells. In PtK2 cells, intact {alpha}-catenin and {alpha}-catenin1-670, harboring the β-catenin– binding site, were directed to cell–cell contacts. In contrast, {alpha}-catenin697–906 fragments were recruited to cell–cell contacts, focal adhesions, and stress fibers. Our results imply that in vivo {alpha}-catenin, like vinculin, is tightly regulated in its ligand binding activity.

Abbreviations used in this paper: DFP, diisopropylfluorophosphate; DSP, dithiobis(succinimidylproppionate); MDBK, Madin–Darby bovine kidney epithelial cells; PIP2, phosphatidylinositol-4,5-bisphosphate; PtK2, Potourus tridactylus kidney epithelial cells; RU, resonance units.

Address all correspondence to Dr. Manfred Rüdiger, Cell Biology, Zoological Institute, Technical University Braunschweig, 38092 Braunschweig, Germany. Tel.: 0531-391-3191. Fax: 0531-391-8203. E-mail: m.ruediger{at}tu-bs.de


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