scholarly journals Microtubule Disruption in Keratinocytes Induces Cell-Cell Adhesion through Activation of Endogenous E-Cadherin

2001 ◽  
Vol 12 (7) ◽  
pp. 1983-1993 ◽  
Author(s):  
Sun-Ho Kee ◽  
Peter M. Steinert
Keyword(s):  
Cell Adhesion ◽  
Cell Lines ◽  
Epidermal Keratinocytes ◽  
Cell Periphery ◽  
Microtubule Disruption ◽  
Low Calcium ◽  
High Calcium ◽  
Long Term Treatment ◽  
E Cadherin ◽  
Cell Cell

The association of the cytoskeleton with the cadherin–catenin complex is essential for strong cell-cell adhesion in epithelial cells. In this study, we have investigated the effect of microtubule organization on cell-cell adhesion in differentiating keratinocytes. When microtubules of normal human epidermal keratinocytes (NHEKs) grown in low calcium media (0.05 mM) were disrupted with nocodazole or colcemid, cell-cell adhesion was induced through relocalization of the E-cadherin–catenin–actin complex to the cell periphery. This was accompanied by actin polymerization. Also, it was found that microtubule disruption-induced cell-cell adhesion was significantly reduced in more advanced differentiated keratinocytes. For example, when NHEK cells cultured under high calcium (1.2 mM) for 8 d and then in low calcium for 1 d were treated with nocodazole, there was no induction of cell-cell adhesion. Also long-term treatment of a phorbol ester for 48 h inhibited nocodazole-induced cell-cell adhesion of NHEK. Furthermore, this nocodazole-induced cell-cell adhesion could be observed in squamous cancer cell lines (A431 and SCC-5, -9, and -25) under low calcium condition, but not in the keratinocyte cell lines derived from normal epidermis (HaCaT, RHEK). On the other hand, HaCaT cells continuously cultivated in low calcium media regained a less differentiated phenotype such as decreased expression of cytokeratin 10, and increased K5; these changes were accompanied with inducibility of cell-cell adhesion by nocodazole. Together, our results suggest that microtubule disruption can induce the cell-cell adhesion via activation of endogenous E-cadherin in non- or early differentiating keratinocytes. However, this is no longer possible in advanced terminally differentiating keratinocytes, possibly due to irreversible changes effected by cell envelope barrier formation.

scholarly journals Changes in E-cadherin rigidity sensing regulate cell adhesion

2017 ◽  
Vol 114 (29) ◽  
pp. E5835-E5844 ◽  
Author(s):  
Caitlin Collins ◽  
Aleksandra K. Denisin ◽  
Beth L. Pruitt ◽  
W. James Nelson
Keyword(s):  
Cell Adhesion ◽  
Signaling Molecules ◽  
Membrane Dynamics ◽  
Cell Contact ◽  
Adhesion Assay ◽  
Cell Periphery ◽  
Actin Organization ◽  
Rigidity Sensing ◽  
E Cadherin ◽  
Cell Cell

Mechanical cues are sensed and transduced by cell adhesion complexes to regulate diverse cell behaviors. Extracellular matrix (ECM) rigidity sensing by integrin adhesions has been well studied, but rigidity sensing by cadherins during cell adhesion is largely unexplored. Using mechanically tunable polyacrylamide (PA) gels functionalized with the extracellular domain of E-cadherin (Ecad-Fc), we showed that E-cadherin–dependent epithelial cell adhesion was sensitive to changes in PA gel elastic modulus that produced striking differences in cell morphology, actin organization, and membrane dynamics. Traction force microscopy (TFM) revealed that cells produced the greatest tractions at the cell periphery, where distinct types of actin-based membrane protrusions formed. Cells responded to substrate rigidity by reorganizing the distribution and size of high-traction-stress regions at the cell periphery. Differences in adhesion and protrusion dynamics were mediated by balancing the activities of specific signaling molecules. Cell adhesion to a 30-kPa Ecad-Fc PA gel required Cdc42- and formin-dependent filopodia formation, whereas adhesion to a 60-kPa Ecad-Fc PA gel induced Arp2/3-dependent lamellipodial protrusions. A quantitative 3D cell–cell adhesion assay and live cell imaging of cell–cell contact formation revealed that inhibition of Cdc42, formin, and Arp2/3 activities blocked the initiation, but not the maintenance of established cell–cell adhesions. These results indicate that the same signaling molecules activated by E-cadherin rigidity sensing on PA gels contribute to actin organization and membrane dynamics during cell–cell adhesion. We hypothesize that a transition in the stiffness of E-cadherin homotypic interactions regulates actin and membrane dynamics during initial stages of cell–cell adhesion.

scholarly journals A role for the E-cadherin cell-cell adhesion molecule during tumor progression of mouse epidermal carcinogenesis.

1991 ◽  
Vol 115 (2) ◽  
pp. 517-533 ◽  
Author(s):  
P Navarro ◽  
M Gómez ◽  
A Pizarro ◽  
C Gamallo ◽  
M Quintanilla ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Cell Lines ◽  
Spindle Cell ◽  
Inverse Correlation ◽  
Malignant Cell ◽  
Mouse Tumors ◽  
Keratinocyte Cell ◽  
Well Differentiated ◽  
E Cadherin ◽  
Cell Cell

The expression of the cell-cell adhesion molecules E- and P-cadherin has been analyzed in seven mouse epidermal keratinocyte cell lines representative of different stages of epidermal carcinogenesis. An inverse correlation between the amount of E-cadherin protein and tumorigenicity of the cell lines has been found, together with a complete absence of E-cadherin protein and mRNA expression in three carcinoma cell lines (the epithelioid HaCa4 and the fibroblastoid CarB and CarC cells). A similar result has been detected in tumors induced in nude mice by the cell lines, where induction of E-cadherin expression takes place in moderately differentiated squamous cell carcinomas induced by HaCa4 cells, although at much lower levels than in well-differentiated tumors induced by the epithelial PDV or PDVC57 cell lines. Complete absence of E-cadherin expression has been observed in spindle cell carcinomas induced by CarB or CarC cells. P-cadherin protein was detected in all cell lines that exhibit an epithelial (MCA3D, AT5, PDV, and PDVC57) or epithelioid (HaCa4) morphology, as well as in nude mouse tumors, independent of their tumorigenic capabilities. However, complete absence of P-cadherin was observed in the fibroblast-like cells (CarB and CarC) and in spindle cell carcinomas. The introduction of an exogenous E-cadherin cDNA into HaCa4 cells, or reactivation of the endogenous E-cadherin gene, leads to a partial suppression of the tumorigenicity of this highly malignant cell line. These results suggest a role for E-cadherin in the progression to malignancy of mouse epidermal carcinogenesis. They also suggest that the loss of both E- and P-cadherin could be associated to the final stage of carcinogenesis, the development of spindle cell carcinomas.

scholarly journals RAC1 Regulates Adherens Junctions through Endocytosis of E-Cadherin

2001 ◽  
Vol 12 (4) ◽  
pp. 847-862 ◽  
Author(s):  
Nasreen Akhtar ◽  
Neil A. Hotchin
Keyword(s):  
Cell Adhesion ◽  
Fluorescent Protein ◽  
Adherens Junctions ◽  
Expression Vectors ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Cell Contacts ◽  
Green Fluorescent ◽  
E Cadherin ◽  
Cell Cell

The establishment of cadherin-dependent cell–cell contacts in human epidermal keratinocytes are known to be regulated by the Rac1 small GTP-binding protein, although the mechanisms by which Rac1 participates in the assembly or disruption of cell–cell adhesion are not well understood. In this study we utilized green fluorescent protein (GFP)-tagged Rac1 expression vectors to examine the subcellular distribution of Rac1 and its effects on E-cadherin–mediated cell–cell adhesion. Microinjection of keratinocytes with constitutively active Rac1 resulted in cell spreading and disruption of cell–cell contacts. The ability of Rac1 to disrupt cell–cell adhesion was dependent on colony size, with large established colonies being resistant to the effects of active Rac1. Disruption of cell–cell contacts in small preconfluent colonies was achieved through the selective recruitment of E-cadherin–catenin complexes to the perimeter of multiple large intracellular vesicles, which were bounded by GFP-tagged L61Rac1. Similar vesicles were observed in noninjected keratinocytes when cell–cell adhesion was disrupted by removal of extracellular calcium or with the use of an E-cadherin blocking antibody. Moreover, formation of these structures in noninjected keratinocytes was dependent on endogenous Rac1 activity. Expression of GFP-tagged effector mutants of Rac1 in keratinocytes demonstrated that reorganization of the actin cytoskeleton was important for vesicle formation. Characterization of these Rac1-induced vesicles revealed that they were endosomal in nature and tightly colocalized with the transferrin receptor, a marker for recycling endosomes. Expression of GFP-L61Rac1 inhibited uptake of transferrin-biotin, suggesting that the endocytosis of E-cadherin was a clathrin-independent mechanism. This was supported by the observation that caveolin, but not clathrin, localized around these structures. Furthermore, an inhibitory form of dynamin, known to inhibit internalization of caveolae, inhibited formation of cadherin vesicles. Our data suggest that Rac1 regulates adherens junctions via clathrin independent endocytosis of E-cadherin.

scholarly journals Two Cell Adhesion Molecules, Nectin and Cadherin, Interact through Their Cytoplasmic Domain–Associated Proteins

2000 ◽  
Vol 150 (5) ◽  
pp. 1161-1176 ◽  
Author(s):  
Kouichi Tachibana ◽  
Hiroyuki Nakanishi ◽  
Kenji Mandai ◽  
Kumi Ozaki ◽  
Wataru Ikeda ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Cell Lines ◽  
Cell Adhesion Molecules ◽  
Cytoplasmic Domain ◽  
Adhesion Sites ◽  
Adhesion System ◽  
Associated Proteins ◽  
Mechanism Of Interaction ◽  
E Cadherin ◽  
Cell Cell

We have found a new cell–cell adhesion system at cadherin-based cell–cell adherens junctions (AJs) consisting of at least nectin and l-afadin. Nectin is a Ca2+-independent homophilic immunoglobulin-like adhesion molecule, and l-afadin is an actin filament-binding protein that connects the cytoplasmic region of nectin to the actin cytoskeleton. Both the trans-interaction of nectin and the interaction of nectin with l-afadin are necessary for their colocalization with E-cadherin and catenins at AJs. Here, we examined the mechanism of interaction between these two cell–cell adhesion systems at AJs by the use of α-catenin–deficient F9 cell lines and cadherin-deficient L cell lines stably expressing their various components. We showed here that nectin and E-cadherin were colocalized through l-afadin and the COOH-terminal half of α-catenin at AJs. Nectin trans-interacted independently of E-cadherin, and the complex of E-cadherin and α- and β-catenins was recruited to nectin-based cell–cell adhesion sites through l-afadin without the trans-interaction of E-cadherin. Our results indicate that nectin and cadherin interact through their cytoplasmic domain–associated proteins and suggest that these two cell–cell adhesion systems cooperatively organize cell–cell AJs.

scholarly journals A Novel Role for Integrin-linked Kinase in Epithelial Sheet Morphogenesis

2005 ◽  
Vol 16 (9) ◽  
pp. 4084-4095 ◽  
Author(s):  
Alisa Vespa ◽  
Sudhir J.A. D'Souza ◽  
Lina Dagnino
Keyword(s):  
Cell Membrane ◽  
Adherens Junction ◽  
Focal Adhesions ◽  
Cell Junctions ◽  
Keratinocyte Differentiation ◽  
Epidermal Keratinocytes ◽  
Cell Periphery ◽  
Integrin Linked Kinase ◽  
E Cadherin ◽  
Cell Cell

Integrin-linked kinase (ILK) is a multidomain protein involved in cell motility and cell-extracellular matrix interactions. ILK is found in integrin-containing focal adhesions in undifferentiated primary epidermal keratinocytes. Induction of keratinocyte differentiation by treatment with Ca2+ triggers formation of cell–cell junctions, loss of focal adhesions, and ILK distribution to cell borders. We now show that Ca2+ treatment of keratinocytes induces rapid (≤1 h) translocation to the cell membrane of the adherens junction (AJ) proteins E-cadherin and β-catenin. This is followed by slower (>6 h) localization of tight junction (TJ) proteins. The kinetics of ILK movement toward the cell periphery mimics that of AJ components, suggesting that ILK plays a role in the early formation of cell–cell contacts. Whereas the N terminus in ILK mediates localization to cell borders, expression of an ILK deletion mutant incapable of localizing to the cell membrane (ILK 191-452) interferes with translocation of E-cadherin/β-catenin to cell borders, precluding Ca2+-induced AJ formation. Cells expressing ILK 191-452 also fail to form TJ and sealed cell–cell borders and do not form epithelial sheets. Thus, we have uncovered a novel role for ILK in epithelial cell–cell adhesion, independent of its well-established role in integrin-mediated adhesion and migration.

scholarly journals Regulation of Cell–Cell Adhesion by Rac and Rho Small G Proteins in MDCK Cells

1997 ◽  
Vol 139 (4) ◽  
pp. 1047-1059 ◽  
Author(s):  
Kenji Takaishi ◽  
Takuya Sasaki ◽  
Hirokazu Kotani ◽  
Hideo Nishioka ◽  
Yoshimi Takai
Keyword(s):  
Cell Adhesion ◽  
Tight Junction ◽  
Cell Lines ◽  
Actin Filaments ◽  
Mdck Cells ◽  
Wild Type ◽  
Cell Functions ◽  
Adhesion Sites ◽  
E Cadherin ◽  
Cell Cell

The Rho small G protein family, consisting of the Rho, Rac, and Cdc42 subfamilies, regulates various cell functions, such as cell shape change, cell motility, and cytokinesis, through reorganization of the actin cytoskeleton. We show here that the Rac and Rho subfamilies furthermore regulate cell–cell adhesion. We prepared MDCK cell lines stably expressing each of dominant active mutants of RhoA (sMDCK-RhoDA), Rac1 (sMDCK-RacDA), and Cdc42 (sMDCK-Cdc42DA) and dominant negative mutants of Rac1 (sMDCK-RacDN) and Cdc42 (sMDCK-Cdc42DN) and analyzed cell adhesion in these cell lines. The actin filaments at the cell–cell adhesion sites markedly increased in sMDCK-RacDA cells, whereas they apparently decreased in sMDCK-RacDN cells, compared with those in wild-type MDCK cells. Both E-cadherin and β-catenin, adherens junctional proteins, at the cell–cell adhesion sites also increased in sMDCK-RacDA cells, whereas both of them decreased in sMDCK-RacDN cells. The detergent solubility assay indicated that the amount of detergent-insoluble E-cadherin increased in sMDCK-RacDA cells, whereas it slightly decreased in sMDCK-RacDN cells, compared with that in wild-type MDCK cells. In sMDCK-RhoDA, -Cdc42DA, and -Cdc42DN cells, neither of these proteins at the cell–cell adhesion sites was apparently affected. ZO-1, a tight junctional protein, was not apparently affected in any of the transformant cell lines. Electron microscopic analysis revealed that sMDCK-RacDA cells tightly made contact with each other throughout the lateral membranes, whereas wild-type MDCK and sMDCK-RacDN cells tightly and linearly made contact at the apical area of the lateral membranes. These results suggest that the Rac subfamily regulates the formation of the cadherin-based cell– cell adhesion. Microinjection of C3 into wild-type MDCK cells inhibited the formation of both the cadherin-based cell–cell adhesion and the tight junction, but microinjection of C3 into sMDCK-RacDA cells showed little effect on the localization of the actin filaments and E-cadherin at the cell–cell adhesion sites. These results suggest that the Rho subfamily is necessary for the formation of both the cadherin-based cell– cell adhesion and the tight junction, but not essential for the Rac subfamily-regulated, cadherin-based cell– cell adhesion.

scholarly journals Phorbol ester-induced scattering of HT-29 human intestinal cancer cells is associated with down-modulation of E-cadherin

1993 ◽  
Vol 106 (2) ◽  
pp. 513-521
Author(s):  
M. Fabre ◽  
A. Garcia de Herreros
Keyword(s):  
Cell Adhesion ◽  
Phorbol Ester ◽  
Cell Attachment ◽  
Cell Aggregation ◽  
Tumor Promoter ◽  
High Concentration ◽  
High Calcium ◽  
Ht 29 ◽  
E Cadherin ◽  
Cell Cell

The effects of tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) on the growth characteristics of the colon cancer cell line HT-29 M6 were studied. TPA induced the scattering of proliferative HT-29 M6 cells: in the presence of the phorbol ester, HT-29 M6 colonies scattered and the cells acquired a flatter aspect with diminished cell-cell contacts. This effect of TPA required a persistent activation of PK-C and was accompanied by a slight decrease (30%) in the growth rate. Modifications by TPA of two scattering associated properties of these cells were also detected: TPA decreased cell-to-cell aggregation and enhanced the cellular attachment to matrix substrata (collagen, laminin). The decrease in cell-to-cell adhesion was correlated with a loss of cellular E-cadherin as evidenced by immunofluorescence or immunoblotting with a specific monoclonal antibody. Cell scattering was dependent on the extracellular concentration of Ca2+; an increase from 1.6 to 10 mM in the concentration of this ion completely blocked the morphological effects of TPA as well as its action on cell aggregation. This high concentration of Ca2+ also prevented the down modulation of E-cadherin as determined by immunofluorescence. However, the TPA-induced increase in cell attachment to the matrix was not affected by high calcium. These findings support the importance of altered cell-cell adhesion in the process of scattering and provide a good system for the study of down modulation of E-cadherin, a protein involved in the control of cell growth, differentiation and invasion of epithelial cells.

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