ZO-1 and ZO-2 Independently Determine Where Claudins Are Polymerized in Tight-Junction Strand Formation

Abstract

A fundamental question in cell and developmental biology is how epithelial cells construct the diffusion barrier allowing them to separate different body compartments. Formation of tight junction (TJ) strands, which are crucial for this barrier, involves the polymerization of claudins, TJ adhesion molecules, in temporal and spatial manners. ZO-1 and ZO-2 are major PDZ-domain-containing TJ proteins and bind directly to claudins, yet their functional roles are poorly understood. We established cultured epithelial cells (1(ko)/2(kd)) in which the expression of ZO-1/ZO-2 was suppressed by homologous recombination and RNA interference, respectively. These cells were well polarized, except for a complete lack of TJs. When exogenously expressed in 1(ko)/2(kd) cells, ZO-1 and ZO-2 were recruited to junctional areas where claudins were polymerized, but truncated ZO-1 (NZO-1) containing only domains PDZ1-3 was not. When NZO-1 was forcibly recruited to lateral membranes and dimerized, claudins were dramatically polymerized. These findings indicate that ZO-1 and ZO-2 can independently determine whether and where claudins are polymerized.

Enteropathogenic Escherichia coli , Shigella flexneri , and Listeria monocytogenes Recruit a Junctional Protein, Zonula Occludens-1, to Actin Tails and Pedestals

“…We speculate that this is due to the functional redundancy between ZO-1 and ZO-2, another member of the membrane-associated guanylate kinase homologue family (15). The speculation is strongly supported by a recent report describing that in the absence of ZO-1, some physiological functions of ZO-1 are compensated for by ZO-2 in the polarized epithelial cells (56). We confirmed that ZO-2 is expressed in Caco/B7 and HeLa cells and is also recruited to the pedestals and the actin tails (our unpublished observation).…”

Tight junction-based epithelial microenvironment and cell proliferation

“…Occludin and JAM-1 were shown to be associated with ZO-1 through both non-PDZ and PDZ domains Schmidt et al, 2004). Recent studies on ZO-1/2/3-deficient cultured epithelial cells have revealed unequivocally that ZO-1/2 determines where and when the claudins are polymerized to form TJ strands with barrier functions (Umeda et al, 2006). These cells lacking in ZO-1/2/3, are normal in their basic apico-basal polarity, but are abnormal in the fine distribution of polarity proteins, such as PAR3, suggesting that tight junctions might be involved in the fine-tuning of cell polarity, although they are probably not involved in the initiation of apico-basal polarity.…”

“…At the present time, several aspects of the molecular mechanisms of TJ physiology have been implied by cellular and molecular biology studies, as described below, although the detailed molecular mechanisms still remain unclear. First, ZO-1/2-deficient cultured epithelial cells, which form epithelial cell sheets but lack TJs (Umeda et al, 2006), exhibited no paracellular barrier functions, but did exhibit the normal epithelial cell sheet formation (mediated by cell-cell adhesion through the belt-like arrangement of AJs). Thus, the proteinaceous TJ strands are dominantly responsible for the paracellular barrier functions, consistent with the dominant contribution of TJs, compared with the minor contribution of the lateral intercellular space, to paracellular resistance that was revealed by physiological measurements (Kottra et al, 1993a; Kottra and Fro¨mter, 1993b; Yap et al, 1998).…”

ZO-1- and ZO-2-Dependent Integration of Myosin-2 to Epithelial Zonula Adherens

“…ZO1(ko)/2(kd) Eph4 cells and ZO1(ko)/2(ko) F9 cells were established by Umeda et al (2006). Eph4 and F9 cells with or without ZO-1/2 were cultured in DMEM including 10% fetal calf serum.…”