scholarly journals An association between type Iγ PI4P 5-kinase and Exo70 directs E-cadherin clustering and epithelial polarization

2012 ◽  
Vol 23 (1) ◽  
pp. 87-98 ◽  
Author(s):  
Xunhao Xiong ◽  
Qingwen Xu ◽  
Yan Huang ◽  
Raman Deep Singh ◽  
Richard Anderson ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Adherens Junctions ◽  
Direct Interaction ◽  
Phosphatidylinositol 4 ◽  
E Cadherin

E-Cadherin–mediated formation of adherens junctions (AJs) is essential for the morphogenesis of epithelial cells. However, the mechanisms underlying E-cadherin clustering and AJ maturation are not fully understood. Here we report that type Iγ phosphatidylinositol-4-phosphate 5-kinase (PIPKIγ) associates with the exocyst via a direct interaction with Exo70, the exocyst subunit that guides the polarized targeting of exocyst to the plasma membrane. By means of this interaction, PIPKIγ mediates the association between E-cadherin and Exo70 and determines the targeting of Exo70 to AJs. Further investigation revealed that Exo70 is necessary for clustering of E-cadherin on the plasma membrane and extension of nascent E-cadherin adhesions, which are critical for the maturation of cohesive AJs. In addition, we observed phosphatidylinositol-4,5-bisphosphate (PI4,5P2) accumulation at E-cadherin clusters during the assembly of E-cadherin adhesions. PIPKIγ-generated PI4,5P2 is required for recruiting Exo70 to newly formed E-cadherin junctions and facilitates the assembly and maturation of AJs. These results support a model in which PIPKIγ and PIPKIγ-generated PI4,5P2 pools at nascent E-cadherin contacts cue Exo70 targeting and orient the tethering of exocyst-associated E-cadherin. This could be an important mechanism that regulates E-cadherin clustering and AJ maturation, which is essential for the establishment of solid, polarized epithelial structures.

scholarly journals Novel Membrane Protein shrew-1 Targets to Cadherin-Mediated Junctions in Polarized Epithelial Cells

2004 ◽  
Vol 15 (1) ◽  
pp. 397-406 ◽  
Author(s):  
Sanita Bharti ◽  
Heike Handrow-Metzmacher ◽  
Silvia Zickenheiner ◽  
Andreas Zeitvogel ◽  
Rudolf Baumann ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Membrane Protein ◽  
Transmembrane Protein ◽  
Adherens Junctions ◽  
Direct Interaction ◽  
Potential Candidate ◽  
Putative Open Reading Frame ◽  
Polarized Epithelial Cells ◽  
E Cadherin

While searching for potential candidate molecules relevant for the pathogenesis of endometriosis, we discovered a 2910-base pair cDNA encoding a novel putative 411-amino acid integral membrane protein that we called shrew-1. The putative open-reading frame was confirmed with antibodies against shrew-1 peptides that labeled a protein of ∼48 kDa in extracts of shrew-1 mRNA-positive tissue and also detected ectopically expressed shrew-1. Expression of epitope-tagged shrew-1 in epithelial cells and analysis by surface biotinylation and immunoblots demonstrated that shrew-1 is indeed a transmembrane protein. Shrew-1 is able to target to E-cadherin-mediated adherens junctions and interact with the E-cadherin–catenin complex in polarized MCF7 and Madin-Darby canine kidney cells, but not with the N-cadherin–catenin complex in nonpolarized epithelial cells. Direct interaction of shrew-1 with β-catenin in in vitro pull-down assay suggests that β-catenin might be one of the proteins that targets and/or retains shrew-1 in the adherens junctions. Interestingly, shrew-1 was partially translocated in response to scatter factor (ligand of receptor tyrosine kinase c-met) from the plasma membrane to the cytoplasm where it still colocalized with endogenous E-cadherin. In summary, we introduce shrew-1 as a novel component of adherens junctions, interacting with E-cadherin–β-catenin complexes in polarized epithelial cells.

scholarly journals The Interaction of JRAB/MICAL-L2 with Rab8 and Rab13 Coordinates the Assembly of Tight Junctions and Adherens Junctions

2008 ◽  
Vol 19 (3) ◽  
pp. 971-983 ◽  
Author(s):  
Rie Yamamura ◽  
Noriyuki Nishimura ◽  
Hiroyoshi Nakatsuji ◽  
Seiji Arase ◽  
Takuya Sasaki
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Tight Junctions ◽  
Binding Protein ◽  
Adherens Junctions ◽  
Binding Domain ◽  
Endocytic Recycling ◽  
E Cadherin

The assembly of tight junctions (TJs) and adherens junctions (AJs) is regulated by the transport of integral TJ and AJ proteins to and/or from the plasma membrane (PM) and it is tightly coordinated in epithelial cells. We previously reported that Rab13 and a junctional Rab13-binding protein (JRAB)/molecule interacting with CasL-like 2 (MICAL-L2) mediated the endocytic recycling of an integral TJ protein occludin and the formation of functional TJs. Here, we investigated the role of Rab13 and JRAB/MICAL-L2 in the transport of other integral TJ and AJ proteins claudin-1 and E-cadherin to the PM by using a Ca2+-switch model. Although knockdown of Rab13 specifically suppressed claudin-1 and occludin but not E-cadherin transport, knockdown of JRAB/MICAL-L2 and expression of its Rab13-binding domain (JRAB/MICAL-L2-C) inhibited claudin-1, occludin, and E-cadherin transport. We then identified Rab8 as another JRAB/MICAL-L2-C-binding protein. Knockdown of Rab8 inhibited the Rab13-independent transport of E-cadherin to the PM. Rab8 and Rab13 competed with each other for the binding to JRAB/MICAL-L2 and functionally associated with JRAB/MICAL-L2 at the perinuclear recycling/storage compartments and PM, respectively. These results suggest that the interaction of JRAB/MICAL-L2 with Rab8 and Rab13 coordinates the assembly of AJs and TJs.

scholarly journals Severe neonatal anemia increases intestinal permeability by disrupting epithelial adherens junctions

2020 ◽  
Vol 318 (4) ◽  
pp. G705-G716 ◽  
Author(s):  
Krishnan MohanKumar ◽  
Kopperuncholan Namachivayam ◽  
Nithya Sivakumar ◽  
Natascha G. Alves ◽  
Venkataramana Sidhaye ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intestinal Permeability ◽  
Adherens Junctions ◽  
Severe Anemia ◽  
Intestinal Epithelial ◽  
Mucosal Permeability ◽  
Chain Reactions ◽  
Age Related ◽  
Epithelial Junctions ◽  
E Cadherin

Anemia is a frequent diagnosis in critically ill infants, but the clinical implications of severe anemia in these patients remain unclear. In this study, we examined preweaned mice to investigate the effects of severe anemia during early infancy on gut mucosal permeability. C57BL/6 mice were subjected to timed phlebotomy between postnatal days (P) 2–10 to induce severe anemia (hematocrits 20%–24%), and intestinal permeability was tracked longitudinally between P10 and P20 as intestine-to-plasma translocation of enteral macromolecules and bacterial translocation. Epithelial junctions were evaluated by electron microscopy, polymerase chain reactions, immunohistochemistry, and/or enzyme immunoassays on intestinal tissues, Caco-2 intestinal epithelial-like cells, and colonic organoids. Preweaned mouse pups showed an age-related susceptibility to severe anemia, with increased intestinal permeability to enteral macromolecules (dextran, ovalbumin, β-lactoglobulin) and luminal bacteria. Electron micrographs showed increased paracellular permeability and ultrastructural abnormalities of the adherens junctions. These findings were explained by the loss of E-cadherin in epithelial cells, which was caused by destabilization of the E-cadherin ( Cdh1) mRNA because of microRNA let-7e-5p binding to the 3′-untranslated region. Severe anemia resulted in a disproportionate and persistent increase in intestinal permeability in preweaned mice because of the disruption of epithelial adherens junctions. These changes are mediated via microRNA let-7e-mediated depletion of Cdh1 mRNA. NEW & NOTEWORTHY This research article shows that newborn infants with severe anemia show an age-related susceptibility to developing increased intestinal permeability to ingested macromolecules. This abnormal permeability develops because of abnormalities in intestinal epithelial junctions caused by a deficiency of the molecule E-cadherin in epithelial cells. The deficiency of E-cadherin is caused by destabilization of its mRNA precursor because of increased expression and binding of another molecule, the microRNA let-7e-5p, to the E-cadherin mRNA.

scholarly journals E–N-cadherin heterodimers define novel adherens junctions connecting endoderm-derived cells

2011 ◽  
Vol 195 (5) ◽  
pp. 873-887 ◽  
Author(s):  
Beate K. Straub ◽  
Steffen Rickelt ◽  
Ralf Zimbelmann ◽  
Christine Grund ◽  
Caecilia Kuhn ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Adherens Junctions ◽  
Tumor Biology ◽  
Intercellular Junctions ◽  
Pivotal Role ◽  
Major Portion ◽  
Tissue Development ◽  
Cadherin Switch ◽  
And Function ◽  
E Cadherin

Intercellular junctions play a pivotal role in tissue development and function and also in tumorigenesis. In epithelial cells, decrease or loss of E-cadherin, the hallmark molecule of adherens junctions (AJs), and increase of N-cadherin are widely thought to promote carcinoma progression and metastasis. In this paper, we show that this “cadherin switch” hypothesis does not hold for diverse endoderm-derived cells and cells of tumors derived from them. We show that the cadherins in a major portion of AJs in these cells can be chemically cross-linked in E–N heterodimers. We also show that cells possessing E–N heterodimer AJs can form semistable hemihomotypic AJs with purely N-cadherin–based AJs of mesenchymally derived cells, including stroma cells. We conclude that these heterodimers are the major AJ constituents of several endoderm-derived tissues and tumors and that the prevailing concept of antagonistic roles of these two cadherins in developmental and tumor biology has to be reconsidered.

scholarly journals Cross-Talk between Adherens Junctions and Desmosomes Depends on Plakoglobin

1997 ◽  
Vol 136 (4) ◽  
pp. 919-934 ◽  
Author(s):  
Jani E. Lewis ◽  
James K. Wahl ◽  
Kristin M. Sass ◽  
Pamela J. Jensen ◽  
Keith R. Johnson ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cell Line ◽  
Adherens Junctions ◽  
Adherens Junction ◽  
Epithelial Cell Line ◽  
Cell Contact ◽  
Common Component ◽  
Classical Cadherins ◽  
E Cadherin

Squamous epithelial cells have both adherens junctions and desmosomes. The ability of these cells to organize the desmosomal proteins into a functional structure depends upon their ability first to organize an adherens junction. Since the adherens junction and the desmosome are separate structures with different molecular make up, it is not immediately obvious why formation of an adherens junction is a prerequisite for the formation of a desmosome. The adherens junction is composed of a transmembrane classical cadherin (E-cadherin and/or P-cadherin in squamous epithelial cells) linked to either β-catenin or plakoglobin, which is linked to α-catenin, which is linked to the actin cytoskeleton. The desmosome is composed of transmembrane proteins of the broad cadherin family (desmogleins and desmocollins) that are linked to the intermediate filament cytoskeleton, presumably through plakoglobin and desmoplakin. To begin to study the role of adherens junctions in the assembly of desmosomes, we produced an epithelial cell line that does not express classical cadherins and hence is unable to organize desmosomes, even though it retains the requisite desmosomal components. Transfection of E-cadherin and/or P-cadherin into this cell line did not restore the ability to organize desmosomes; however, overexpression of plakoglobin, along with E-cadherin, did permit desmosome organization. These data suggest that plakoglobin, which is the only known common component to both adherens junctions and desmosomes, must be linked to E-cadherin in the adherens junction before the cell can begin to assemble desmosomal components at regions of cell–cell contact. Although adherens junctions can form in the absence of plakoglobin, making use only of β-catenin, such junctions cannot support the formation of desmosomes. Thus, we speculate that plakoglobin plays a signaling role in desmosome organization.

scholarly journals Phosphatidylinositol-4-phosphate 5-kinase 1α Mediates Extracellular Calcium-induced Keratinocyte Differentiation

2009 ◽  
Vol 20 (6) ◽  
pp. 1695-1704 ◽  
Author(s):  
Zhongjian Xie ◽  
Sandra M. Chang ◽  
Sally D. Pennypacker ◽  
Er-Yuan Liao ◽  
Daniel D. Bikle
Keyword(s):  
Plasma Membrane ◽  
Second Messengers ◽  
Human Keratinocytes ◽  
Extracellular Calcium ◽  
Keratinocyte Differentiation ◽  
Activity Levels ◽  
Differentiation Markers ◽  
Resultant Increase ◽  
Phosphatidylinositol 4 ◽  
E Cadherin

Extracellular calcium (Cao) is a major regulator of keratinocyte differentiation, but the mechanism is unclear. Phosphatidylinositol-4-phosphate 5-kinase 1α (PIP5K1α) is critical in synthesizing phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. In this study, we sought to determine whether PIP5K1α plays a role in mediating the ability of Cao to induce keratinocyte differentiation. We found that treatment of human keratinocytes in culture with Cao resulted in increased PIP5K1α level and activity, as well as PI(4,5)P2 level, binding of phosphatidylinositol 3,4,5-triphosphate [PI(3,4,5)P3] to and activation of phospholipase C-γ1 (PLC-γ1), with the resultant increase in inositol 1,4,5-trisphosphate (IP3) and intracellular calcium (Cai). Knockdown of PIP5K1α in human keratinocytes blocked Cao-induced increases in the binding of PI(3,4,5)P3 to PLC-γ1; PLC-γ1 activity; levels of PI(4,5)P2, IP3, and Cai; and induction of keratinocyte differentiation markers. Coimmunoprecipitation and confocal studies revealed that Cao stimulated PIP5K1α recruitment to the E-cadherin–catenin complex in the plasma membrane. Knockdown of E-cadherin or β-catenin blocked Cao-induced activation of PIP5K1α. These results indicate that after Cao stimulation PIP5K1α is recruited by the E-cadherin–catenin complex to the plasma membrane where it provides the substrate PI(4,5)P2 for both PI3K and PLC-γ1. This signaling pathway is critical for Cao-induced generation of the second messengers IP3 and Cai and keratinocyte differentiation.

scholarly journals E-cadherin redistribution mediated by Clostridioides difficile toxins increases spore-association to adherens junctions

2021 ◽  
Author(s):  
Pablo Castro-Cordova ◽  
Macarena Otto-Medina ◽  
Borden Lacy ◽  
Daniel Paredes-Sabja
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Adherens Junctions ◽  
Immunogold Labelling ◽  
Intestinal Epithelial ◽  
Clostridioides Difficile ◽  
Spore Adherence ◽  
E Cadherin

Nearly ~20% of patients with C. difficile infection (CDI) manifest recurrence of CDI (R-CDI). During CDI, C. difficile forms spores essential for R-CDI. Interactions of C. difficile spores with intestinal epithelial cells (IECs) contribute to R-CDI. However, this interaction remains poorly understood. Here, we provide evidence that C. difficile spores interact with E-cadherin, contributing to spore-adherence and internalization into IECs. C. difficile toxins TcdA/TcdB lead to adherens junctions opening and increase spore-adherence to IECs. Confocal micrographs demonstrate that C. difficile spores associate with accessible E-cadherin; spore-E-cadherin association increases upon TcdA/TcdB intoxication. The presence of anti-E-cadherin antibodies decreased spore adherence and entry into IECs. By ELISA, immunofluorescence, and immunogold labelling, we observed that E-cadherin binds to C. difficile spores, specifically to the hair-like projections of the spore. Overall, these results expand our knowledge of how C. difficile spores bind to IECs and how toxin-mediated damage affects spore interactions with IECs.

scholarly journals Tyrosine phosphorylation regulates the adhesions of ras-transformed breast epithelia.

1995 ◽  
Vol 130 (2) ◽  
pp. 461-471 ◽  
Author(s):  
M S Kinch ◽  
G J Clark ◽  
C J Der ◽  
K Burridge
Keyword(s):  
Epithelial Cells ◽  
Actin Cytoskeleton ◽  
Tyrosine Phosphorylation ◽  
Tyrosine Kinases ◽  
Adherens Junctions ◽  
Focal Adhesions ◽  
Transformed Cells ◽  
Beta Catenin ◽  
Altered Cell ◽  
E Cadherin

Transformed epithelial cells often are characterized by a fibroblastic or mesenchymal morphology. These cells exhibit altered cell-cell and cell-substrate interactions. Here we have identified changes in the adhesions and cytoskeletal interactions of transformed epithelial cells that contribute to their altered morphology. Using MCF-10A human breast epithelial cells as a model system, we have found that transformation by an activated form of ras is characterized by less developed adherens-type junctions between cells but increased focal adhesions. Contributing to the modified adherens junctions of the transformed cells are decreased interactions among beta-catenin, E-cadherin, and the actin cytoskeleton. The ras-transformed cells reveal elevated phosphotyrosine in many proteins, including beta-catenin and p120 Cas. Whereas in the normal cells beta-catenin is found in association with E-cadherin, p120 Cas is not. In the ras-transformed cells, the situation is reversed; tyrosine-phosphorylated p120 Cas, but not tyrosine-phosphorylated beta-catenin, now is detected in E-cadherin complexes. The tyrosine-phosphorylated beta-catenin also shows increased detergent solubility, suggesting a decreased association with the actin cytoskeleton. p120 Cas, whether tyrosine phosphorylated or not, partitions into the detergent soluble fraction, suggesting that it is not tightly bound to the actin cytoskeleton in either the normal or ras-transformed cells. Inhibitors of tyrosine kinases decrease the level of tyrosine phosphorylation and restore a normal epithelial morphology to the ras-transformed cells. In particular, decreased tyrosine phosphorylation of beta-catenin is accompanied by increased interaction with both E-cadherin and the detergent insoluble cytoskeletal fraction. These results suggest that elevated tyrosine phosphorylation of proteins such as beta-catenin and p120 Cas contribute to the altered adherens junctions of ras-transformed epithelia.

scholarly journals Dual pulse-chase microscopy reveals early divergence in the biosynthetic trafficking of the Na,K-ATPase and E-cadherin

2015 ◽  
Vol 26 (24) ◽  
pp. 4401-4411 ◽  
Author(s):  
Glen A. Farr ◽  
Michael Hull ◽  
Emily H. Stoops ◽  
Rosalie Bateson ◽  
Michael J. Caplan
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Cell Surface ◽  
Golgi Complex ◽  
Plasma Membranes ◽  
Secretory Pathway ◽  
Trans Golgi Network ◽  
Polarized Epithelial Cells ◽  
Golgi Network ◽  
E Cadherin

Recent evidence indicates that newly synthesized membrane proteins that share the same distributions in the plasma membranes of polarized epithelial cells can pursue a variety of distinct trafficking routes as they travel from the Golgi complex to their common destination at the cell surface. In most polarized epithelial cells, both the Na,K-ATPase and E-cadherin are localized to the basolateral domains of the plasma membrane. To examine the itineraries pursued by newly synthesized Na,K-ATPase and E-cadherin in polarized MDCK epithelial cells, we used the SNAP and CLIP labeling systems to fluorescently tag temporally defined cohorts of these proteins and observe their behaviors simultaneously as they traverse the secretory pathway. These experiments reveal that E-cadherin is delivered to the cell surface substantially faster than is the Na,K-ATPase. Furthermore, the surface delivery of newly synthesized E-cadherin to the plasma membrane was not prevented by the 19°C temperature block that inhibits the trafficking of most proteins, including the Na,K-ATPase, out of the trans-Golgi network. Consistent with these distinct behaviors, populations of newly synthesized E-cadherin and Na,K-ATPase become separated from one another within the trans-Golgi network, suggesting that they are sorted into different carrier vesicles that mediate their post-Golgi trafficking.

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