Connexin45 directly binds to ZO-1 and localizes to the tight junction region in epithelial MDCK cells

Zonula occludens protein I (ZO-1) Is a cytosolic tight junction protein tha t tethers transmembrane proteins such as occludin, claudin, and junctional adhesion molecule to the actin cytoskeleton. The interaction between ZO-1 a nd claudin or junctional adhesion molecule occurs via the amino-terminal PS D95/Dlg/ZO-1 (PDZ) domains in ZO-1. A yeast two-hybrid screen to search for proteins that interact with the PDZ domains of ZO-1 identified connexin (C x) 45. Cx45 interacts with the PDZ domains of ZO-1 and ZO-3, but not ZO-2, via a short C-terminal PDZ binding motif (SVWI). In transfected epithelial Madin-Darby canine kidney cells, Cx45 co-localizes with endogenous ZO-1 at or near tight junctions and co-precipitation experiments show that Cx45 and ZO-1 directly. interact. Inactivating the C-terminal PDZ-binding motif in Cx45 affects its co-precipitation and co-localization with ZO-1. The growin g number of connexins (i.e. Cx43 and Cx45) that can associate with ZO prote ins indicate that ZO proteins may play a more general role in organizing ga p junctions and/or; in recruiting signaling molecules that regulate interce llular communication. (C) 2001 Federation, of European Biochemical Societie s. Published by Elsevier Science B.V. All rights reserved.