Interaction of Zonula Occludens-1 (ZO-1) with α-Actinin-4:  Application of Functional Proteomics for Identification of PDZ Domain-Associated Proteins

Chen, Vincent C.; Li, Xinbo; Perreault, Hélène; Nagy, J.I.

doi:10.1021/pr060216l

Cited by 43 publications

(38 citation statements)

References 55 publications

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“…Our studies indicate that F-actin localization is significantly altered in the ZO-1 knockdown cells, with an increase in actin staining at apical junctional complex and into scattered apical dots. These observations suggest that ZO-1 has very specific effects on actin dynamics at the apical junctional complex, perhaps by localizing the activity of cytoskeletal proteins [such as ␣ actinin; Chen et al, 2006), ␣ catenin (Itoh et al, 1997), or shroom2 (Etournay et al, 2007)] or signaling pathways [such as the RhoGEF TUBA (Otani et al, 2006) and G␣ 12 (Meyer et al, 2002)] that regulate actin dynamics.…”

Section: Discussionmentioning

confidence: 99%

ZO-1 Stabilizes the Tight Junction Solute Barrier through Coupling to the Perijunctional Cytoskeleton

Itallie¹,

Fanning²,

Bridges

et al. 2009

MBoC

380

415

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ZO-1 binds numerous transmembrane and cytoplasmic proteins and is required for assembly of both adherens and tight junctions, but its role in defining barrier properties of an established tight junction is unknown. We depleted ZO-1 in MDCK cells using siRNA methods and observed specific defects in the barrier for large solutes, even though flux through the small claudin pores was unaffected. This permeability increase was accompanied by morphological alterations and reorganization of apical actin and myosin. The permeability defect, and to a lesser extent morphological changes, could be rescued by reexpression of either full-length ZO-1 or an N-terminal construct containing the PDZ, SH3, and GUK domains. ZO-2 knockdown did not replicate either the permeability or morphological phenotypes seen in the ZO-1 knockdown, suggesting that ZO-1 and -2 are not functionally redundant for these functions. Wild-type and knockdown MDCK cells had differing physiological and morphological responses to pharmacologic interventions targeting myosin activity. Use of the ROCK inhibitor Y27632 or myosin inhibitor blebbistatin increased TER in wild-type cells, whereas ZO-1 knockdown monolayers were either unaffected or changed in the opposite direction; paracellular flux and myosin localization were also differentially affected. These studies are the first direct evidence that ZO-1 limits solute permeability in established tight junctions, perhaps by forming a stabilizing link between the barrier and perijunctional actomyosin. INTRODUCTIONTight junctions form size-and charge-selective barriers to the paracellular movement of solutes and ions across epithelia (Powell, 1981;Madara, 1998). More than 40 different cytoplasmic and transmembrane proteins are located at the junction (Gonzalez-Mariscal et al., 2003;Schneeberger and Lynch, 2004;Guillemot et al., 2008) yet there is limited knowledge of how most contribute to the junction's selective sealing properties. The cytoplasmic protein ZO-1 was the first tight junction protein to be identified (Stevenson et al., 1986). It is a member of the MAGUK family of membraneassociated scaffolding proteins (Funke et al., 2005) and contains protein-binding domains for all the major transmembrane barrier proteins: claudins (Furuse et al., 1998;Itoh et al., 1999), occludin (Furuse et al., 1994;Fanning et al., 1998), tricellulin (Ikenouchi et al., 2005;Riazuddin et al., 2006), JAM-A (Martin-Padura et al., 1998;Ebnet et al., 2000), and several components of the perijunctional network of actin and myosin (Tsukita et al., 2001;Hartsock and Nelson, 2008;Yamazaki et al., 2008). Although ZO-1 gene deletions are embryonic lethal in mice (Katsuno et al., 2008), the depletion of ZO-1 in cultured epithelial cell models (Umeda et al., 2004;McNeil et al., 2006) results in only a delay in barrier formation. Once tight junctions are formed in the ZO-1-depleted cells, TER and flux for large (40 kDa) solutes are reported to be essentially normal (Umeda et al., 2004). There is no direct evidence to date suggesting that ...

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Section: Discussionmentioning

confidence: 99%

ZO-1 Stabilizes the Tight Junction Solute Barrier through Coupling to the Perijunctional Cytoskeleton

Itallie¹,

Fanning²,

Bridges

et al. 2009

MBoC

380

415

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show abstract

“…There is no known F-actin-binding domain within PDZ1 or PDZ3 of ZO-1 (Fanning et al, 1998;Itoh et al, 1997), therefore regulation of the contractile array is likely through the binding of ZO-1 to cytoskeletal regulatory proteins and/or other cytoskeletal components. For example, PDZ1 is known to bind to alphaactinin 4 and ARVCF (Chen et al, 2006;Kausalya et al, 2004). Future studies will address the role of PDZ1 ligands in regulating cytoskeletal structure.…”

Section: Discussionmentioning

confidence: 99%

Epithelial barrier assembly requires coordinated activity of multiple domains of the tight junction protein ZO-1

Rodgers

Beam

Anderson

et al. 2013

Journal of Cell Science

123

112

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SummaryTight junctions (TJs) regulate the paracellular movement of ions, macromolecules and immune cells across epithelia. Zonula occludens (ZO)-1 is a multi-domain polypeptide required for the assembly of TJs. MDCK II cells lacking ZO-1, and its homolog ZO-2, have three distinct phenotypes: reduced localization of occludin and some claudins to the TJs, increased epithelial permeability, and expansion of the apical actomyosin contractile array found at the apical junction complex (AJC). However, it is unclear exactly which ZO-1 binding domains are required to coordinate these activities. We addressed this question by examining the ability of ZO-1 domain-deletion transgenes to reverse the effects of ZO depletion. We found that the SH3 domain and the U5 motif are required to recruit ZO-1 to the AJC and that localization is a prerequisite for normal TJ and cytoskeletal organization. The PDZ2 domain is not required for localization of ZO-1 to the AJC, but is necessary to establish the characteristic continuous circumferential band of ZO-1, occludin and claudin-2. PDZ2 is also required to establish normal permeability, but is not required for normal cytoskeletal organization. Finally, our results demonstrate that PDZ1 is crucial for the normal organization of both the TJ and the AJC cytoskeleton. Our results establish that ZO-1 acts as a true scaffolding protein and that the coordinated activity of multiple domains is required for normal TJ structure and function.

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“…In agreement with our data, actinin-4 has been linked to cell-cell junctions. In epithelial cells, actinin-4 associates with ␤-catenin at AJs (13) and with MAGI-1 and ZO-1 at TJs (6,35). In glomerular epithelial cells (podocytes), mutations in the actinin-4 gene are shown to cause the dysfunction of slit diaphragm (SD), a junctional structure that links the interdigitating foot processes from neighboring podocytes and contains features of both AJs and TJs, and lead to focal and segmental glomerulosclerosis, which is characterized by progressive proteinuria and podocyte foot process effacement (21,24).…”

Section: Discussionmentioning

confidence: 99%

Involvement of Actinin-4 in the Recruitment of JRAB/MICAL-L2 to Cell-Cell Junctions and the Formation of Functional Tight Junctions

Nakatsuji

Nishimura²,

Yamamura³

et al. 2008

Molecular and Cellular Biology

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Interaction of Zonula Occludens-1 (ZO-1) with α-Actinin-4: Application of Functional Proteomics for Identification of PDZ Domain-Associated Proteins

Cited by 43 publications

References 55 publications

ZO-1 Stabilizes the Tight Junction Solute Barrier through Coupling to the Perijunctional Cytoskeleton

ZO-1 Stabilizes the Tight Junction Solute Barrier through Coupling to the Perijunctional Cytoskeleton

Epithelial barrier assembly requires coordinated activity of multiple domains of the tight junction protein ZO-1

Involvement of Actinin-4 in the Recruitment of JRAB/MICAL-L2 to Cell-Cell Junctions and the Formation of Functional Tight Junctions

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