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Molecular and Cellular Biology, December 2006, p. 9003-9015, Vol. 26, No. 23
0270-7306/06/$08.00+0     doi:10.1128/MCB.01811-05
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Normal Establishment of Epithelial Tight Junctions in Mice and Cultured Cells Lacking Expression of ZO-3, a Tight-Junction MAGUK Protein{triangledown} ,{ddagger} ,§

Makoto Adachi,1,{dagger}* Akihito Inoko,1,{dagger} Masaki Hata,2 Kyoko Furuse,2 Kazuaki Umeda,1 Masahiko Itoh,1 and Shoichiro Tsukita1

Department of Cell Biology, Faculty of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan,1 KAN Research Institute, Inc., Kyoto Research Park, Chudoji, Shimogyo-ku, Kyoto 600-8815, Japan2

Received 15 September 2005/ Returned for modification 19 November 2005/ Accepted 1 September 2006

ZO-1, ZO-2, and ZO-3 are closely related MAGUK family proteins that localize at the cytoplasmic surface of tight junctions (TJs). ZO-1 and ZO-2 are expressed in both epithelia and endothelia, whereas ZO-3 is exclusively expressed in epithelia. In spite of intensive studies of these TJ MAGUKs, our knowledge of their functions in vivo, especially those of ZO-3, is still fragmentary. Here, we have generated mice, as well as F9 teratocarcinoma cell lines, that do not express ZO-3 by homologous recombination. Unexpectedly, ZO-3–/– mice were viable and fertile, and rigorous phenotypic analyses identified no significant abnormalities. Moreover, ZO-3-deficient F9 teratocarcinoma cells differentiated normally into visceral endoderm epithelium-like cells in the presence of retinoic acid. These cells had a normal epithelial appearance, and the molecular architecture of their TJs did not appear to be affected, except that TJ localization of ZO-2 was upregulated. Suppression of ZO-2 expression by RNA interference in ZO-3–/– cells, however, did not affect the architecture of TJs. Furthermore, the speed with which TJs formed after a Ca2+ switch was indistinguishable between wild-type and ZO-3–/– cells. These findings indicate that ZO-3 is dispensable in vivo in terms of individual viability, epithelial differentiation, and the establishment of TJs, at least in the laboratory environment.

* Corresponding author. Mailing address: Department of Cell Biology, Faculty of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan. Phone: 81-75-753-4375. Fax: 81-75-753-4660. E-mail: ada{at}mfour.med.kyoto-u.ac.jp.

{triangledown} Published ahead of print on 25 September 2006.

{ddagger} Supplemental material for this article may be found at http://mcb.asm.org/.

§ Dedicated to the memory of coauthor Shoichiro Tsukita, now deceased.

{dagger} M.A. and A.I. contributed equally to this study.

Molecular and Cellular Biology, December 2006, p. 9003-9015, Vol. 26, No. 23
0270-7306/06/$08.00+0     doi:10.1128/MCB.01811-05
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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