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J. Cell Biol.,
Volume 139, Number 4, November 17, 1997 1047-1059
* Department of Molecular Biology and Biochemistry, Osaka University Medical School, Suita 565, Japan; and The Rho small G protein family, consisting
of the Rho, Rac, and Cdc42 subfamilies, regulates various cell functions, such as cell shape change, cell motility, and cytokinesis, through reorganization of the actin
cytoskeleton. We show here that the Rac and Rho subfamilies furthermore regulate cell-cell adhesion. We
prepared MDCK cell lines stably expressing each of
dominant active mutants of RhoA (sMDCK-RhoDA),
Rac1 (sMDCK-RacDA), and Cdc42 (sMDCK-Cdc42DA) and dominant negative mutants of Rac1
(sMDCK-RacDN) and Cdc42 (sMDCK-Cdc42DN)
and analyzed cell adhesion in these cell lines. The actin
filaments at the cell-cell adhesion sites markedly increased in sMDCK-RacDA cells, whereas they apparently decreased in sMDCK-RacDN cells, compared
with those in wild-type MDCK cells. Both E-cadherin
and Takai
Biotimer Project, ERATO, Japan Science and Technology Corporation, JCR Pharmaceuticals Co., Ltd., Nishi-ku, Kobe
651-22, Japan
-catenin, adherens junctional proteins, at the
cell-cell adhesion sites also increased in sMDCK-RacDA cells, whereas both of them decreased in
sMDCK-RacDN cells. The detergent solubility assay
indicated that the amount of detergent-insoluble E-cadherin increased in sMDCK-RacDA cells,
whereas it slightly decreased in sMDCK-RacDN cells,
compared with that in wild-type MDCK cells. In
sMDCK-RhoDA, -Cdc42DA, and -Cdc42DN cells,
neither of these proteins at the cell-cell adhesion sites
was apparently affected. ZO-1, a tight junctional protein, was not apparently affected in any of the transformant cell lines. Electron microscopic analysis revealed
that sMDCK-RacDA cells tightly made contact with
each other throughout the lateral membranes, whereas
wild-type MDCK and sMDCK-RacDN cells tightly and
linearly made contact at the apical area of the lateral
membranes. These results suggest that the Rac subfamily regulates the formation of the cadherin-based cell-
cell adhesion. Microinjection of C3 into wild-type
MDCK cells inhibited the formation of both the cadherin-based cell-cell adhesion and the tight junction,
but microinjection of C3 into sMDCK-RacDA cells
showed little effect on the localization of the actin filaments and E-cadherin at the cell-cell adhesion sites. These results suggest that the Rho subfamily is necessary for the formation of both the cadherin-based cell-
cell adhesion and the tight junction, but not essential
for the Rac subfamily-regulated, cadherin-based cell-
cell adhesion.
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