MLCK-dependent exchange and actin binding region-dependent anchoring of ZO-1 regulate tight junction barrier function

Yu, Dan; Marchiando, Amanda M.; Weber, Christopher R.; Raleigh, David R.; Wang, Yingmin; Shen, Le; Turner, Jerrold R.

doi:10.1073/pnas.0908869107

Cited by 224 publications

(240 citation statements)

References 28 publications

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“…Expression of either full-length EGFP-ZO-1 or EGFP-ZO-1 ΔABR increased the peak TER, although this was 20%±1.0% greater in monolayers that expressed full-length EGFP-ZO-1 relative to those expressing EGFP-ZO-1 ΔABR . This suggests that ABR-mediated interactions play a role in initial barrier formation, consistent with their established role in barrier regulation (Yu et al, 2010). In contrast, induction of EGFP-ZO-1 ΔU5-GuK expression reduced the TER relative to that in uninduced controls (Fig.…”

Section: Zo-1 U5-guk-domain-mediated Interactions Are Necessary For Esupporting

confidence: 67%

“…On the contrary, other studies have demonstrated a regulatory function whereby the ABR stabilizes ZO-1 at the tight junction and is necessary for myosin light chain kinase (MLCK)-dependent ZO-1 trafficking and barrier regulation (Yu et al, 2010). The data here further support a model in which, without the ABR, ZO-1 scaffolding functions are diminished and ZO-1 is unable to effectively facilitate complex interactions (Shen et al, 2008;Yu et al, 2010). Nevertheless, our results indicate that a direct link with F-actin through the ABR is required for ZO-1 to regulate epithelial morphogenesis in some environments.…”

Section: δU5-gukmentioning

confidence: 97%

“…Despite the direct binding of ZO-1 and ZO-2, and therefore transmembrane tight junction proteins through ZO-1-ZO-2 interactions, to the actin cytoskeleton, most studies of the ABR in 2D cultures have identified little or no significant contributions to paracellular barrier function or cortical actin organization (Rodgers et al, 2013;Van Itallie et al, 2009). On the contrary, other studies have demonstrated a regulatory function whereby the ABR stabilizes ZO-1 at the tight junction and is necessary for myosin light chain kinase (MLCK)-dependent ZO-1 trafficking and barrier regulation (Yu et al, 2010). The data here further support a model in which, without the ABR, ZO-1 scaffolding functions are diminished and ZO-1 is unable to effectively facilitate complex interactions (Shen et al, 2008;Yu et al, 2010).…”

Section: δU5-gukmentioning

confidence: 99%

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ZO-1 interactions with F-actin and occludin direct epithelial polarization and single lumen specification in 3D culture

Odenwald

Choi

Buckley

et al. 2016

Journal of Cell Science

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103

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Epithelia within tubular organs form and expand lumens. Failure of these processes can result in serious developmental anomalies. Although tight junction assembly is crucial to epithelial polarization, the contribution of specific tight junction proteins to lumenogenesis is undefined. Here, we show that ZO-1 (also known as TJP1) is necessary for the formation of single lumens. Epithelia lacking this tight junction scaffolding protein form cysts with multiple lumens and are defective in the earliest phases of polarization, both in two and three dimensions. Expression of ZO-1 domain-deletion mutants demonstrated that the actin-binding region and U5-GuK domain are crucial to single lumen development. For actin-binding region, but not U5-GuK domain, mutants, this could be overcome by strong polarization cues from the extracellular matrix. Analysis of the U5-GuK binding partners shroom2, α-catenin and occludin showed that only occludin deletion led to multi-lumen cysts. Like ZO-1-deficiency, occludin deletion led to mitotic spindle orientation defects. Single lumen formation required the occludin OCEL domain, which binds to ZO-1. We conclude that ZO-1-occludin interactions regulate multiple phases of epithelial polarization by providing cell-intrinsic signals that are required for single lumen formation.

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Section: Zo-1 U5-guk-domain-mediated Interactions Are Necessary For Esupporting

confidence: 67%

Section: δU5-gukmentioning

confidence: 97%

Section: δU5-gukmentioning

confidence: 99%

See 1 more Smart Citation

ZO-1 interactions with F-actin and occludin direct epithelial polarization and single lumen specification in 3D culture

Odenwald

Choi

Buckley

et al. 2016

Journal of Cell Science

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103

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“…cZNA localization does not exactly mimic full length ZO-1 expression, nor does it restore the phenotypic, wavy junctions of wild type MDCK cells or dKD cells expressing full length ZO-1 (Fig. 1C) (Fanning et al, 2012), therein confirming that the C-terminal region (aa 888-1746) has a role in the orchestrating the typical architecture of the AJC in these cells (Yu et al, 2010). However, we did not investigate these differences further because they did not correlate with changes in the permeability of MDCK cells (Fig.…”

Section: Resultsmentioning

confidence: 91%

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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“…IEL migration assays into Caco-2 BBe monolayers (24,26) were performed placing flow cytometry-sorted IELs in the upper chamber of the Transwell and fixing the filters at various time points. Stable cell lines expressing pSUPER vectors containing occludin (36) or ZO-1 (26) targeting sequence resulted in the suppression of 90% of target protein expression in Caco-2 BBe cells (46).…”

Section: Methodsmentioning

confidence: 99%

Dynamic migration of γδ intraepithelial lymphocytes requires occludin

Edelblum¹,

Shen²,

Weber³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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152

191

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γδ intraepithelial lymphocytes (IELs) are located beneath or between adjacent intestinal epithelial cells and are thought to contribute to homeostasis and disease pathogenesis. Using in vivo microscopy to image jejunal mucosa of GFP γδ T-cell transgenic mice, we discovered that γδ IELs migrate actively within the intraepithelial compartment and into the lamina propria. As a result, each γδ IEL contacts multiple epithelial cells. Occludin is concentrated at sites of γδ IEL/epithelial interaction, where it forms a ring surrounding the γδ IEL. In vitro analyses showed that occludin is expressed by epithelial and γδ T cells and that occludin derived from both cell types contributes to these rings and to γδ IEL migration within epithelial monolayers. In vivo TNF administration, which results in epithelial occludin endocytosis, reduces γδ IEL migration. Further in vivo analyses demonstrated that occludin KO γδ T cells are defective in both initial accumulation and migration within the intraepithelial compartment. These data challenge the paradigm that γδ IELs are stationary in the intestinal epithelium and demonstrate that γδ IELs migrate dynamically to make extensive contacts with epithelial cells. The identification of occludin as an essential factor in γδ IEL migration provides insight into the molecular regulation of γδ IEL/epithelial interactions.intestine | tight junction T he intestine is one of the few peripheral tissues to contain a large population of intraepithelial lymphocytes (IELs), with one IEL for every 5-10 epithelial cells. Although the majority of these IELs express the γδ T cell receptor, and epidermal γδ IELs have been studied extensively (1-4), the functions of intestinal γδ IELs remain poorly understood. Some studies have shown that γδ IELs contribute to progression of immune-mediated colitis (5-7); other data suggest that γδ IELs contribute to mucosal homeostasis (8, 9) by secreting keratinocyte growth factor (10, 11) and antimicrobial peptides (12, 13), suppressing CD4 + T-cell expansion through TGF-β and IL-10 production (8, 9) and promoting barrier maintenance via poorly understood mechanisms (13-15). These observations and the small number of IELs relative to intestinal epithelial cells are difficult to reconcile with the widely held belief that γδ IELs have limited motility (1, 16).Further understanding of γδ IEL function will require definition of the molecular structures that regulate interactions between intestinal epithelial and γδ T cells. On the basis of the location of epithelial/γδ IEL contact sites along epithelial lateral membranes, it is likely that epithelial proteins targeted to these domains, including apical junction complex components, are involved in these interactions. Attractive candidates include E-cadherin, which can bind CD103 (α E β 7 integrin) expressed by IELs (17), as well as tight junction proteins. For example, γδ IELs express several epithelial tight junction proteins, including occludin and zonula occludens-1 (ZO-1) (18), that may bind directly or indirectly t...

show abstract

MLCK-dependent exchange and actin binding region-dependent anchoring of ZO-1 regulate tight junction barrier function

Cited by 224 publications

References 28 publications

ZO-1 interactions with F-actin and occludin direct epithelial polarization and single lumen specification in 3D culture

ZO-1 interactions with F-actin and occludin direct epithelial polarization and single lumen specification in 3D culture

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

Dynamic migration of γδ intraepithelial lymphocytes requires occludin

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