scholarly journals Formation of E-Cadherin/β-Catenin–based Adherens Junctions in Hepatocytes Requires Serine-10 in p27(Kip1)

2008 ◽  
Vol 19 (4) ◽  
pp. 1605-1613 ◽  
Author(s):  
Delphine Théard ◽  
Marcel A. Raspe ◽  
Dharamdajal Kalicharan ◽  
Dick Hoekstra ◽  
Sven C.D. van IJzendoorn
Keyword(s):  
Cell Adhesion ◽  
Hepg2 Cells ◽  
Molecular Mechanisms ◽  
Functional Organization ◽  
Adherens Junctions ◽  
Oncostatin M ◽  
Wild Type ◽  
P27 Kip1 ◽  
E Cadherin ◽  
Cell Cell

The adhesion between epithelial cells at adherens junctions is regulated by signaling pathways that mediate the intracellular trafficking and assembly of its core components. Insight into the molecular mechanisms of this is necessary to understand how adherens junctions contribute to the functional organization of epithelial tissues. Here, we demonstrate that in human hepatic HepG2 cells, oncostatin M-p42/44 mitogen-activated protein kinase signaling stimulates the phosphorylation of p27(Kip1) on Ser-10 and promotes cell–cell adhesion. The overexpression of wild-type p27 or a phospho-mimetic p27S10D mutant in HepG2 cells induces a hyper-adhesive phenotype. In contrast, the overexpression of a nonphosphorylatable p27S10A mutant prevents the mobilization of E-cadherin and β-catenin at the cell surface, reduces basal cell–cell adhesion strength, and prevents the stimulatory effect of oncostatin M on cell–cell adhesion. As part of the underlying molecular mechanism, it is shown that in p27S10A-expressing cells β-catenin interacts with p27 and is prevented from interacting with E-cadherin. The intracellular retention of E-cadherin and β-catenin is also observed in hepatocytes from p27S10A knockin mice that express the p27S10A mutant instead of wild-type p27. Together, these data suggest that the formation of adherens junctions in hepatocytes requires Ser-10 in p27.

scholarly journals Role of Nectin in Formation of E-Cadherin–based Adherens Junctions in Keratinocytes: Analysis with the N-Cadherin Dominant Negative Mutant

2003 ◽  
Vol 14 (4) ◽  
pp. 1597-1609 ◽  
Author(s):  
Yoshinari Tanaka ◽  
Hiroyuki Nakanishi ◽  
Shigeki Kakunaga ◽  
Noriko Okabe ◽  
Tomomi Kawakatsu ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Adhesion Molecule ◽  
Cell Adhesion Molecule ◽  
Adherens Junctions ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Adhesion Activity ◽  
Negative Mutant ◽  
E Cadherin ◽  
Cell Cell

E-Cadherin is a Ca2+-dependent cell-cell adhesion molecule at adherens junctions (AJs) of epithelial cells. A fragment of N-cadherin lacking its extracellular region serves as a dominant negative mutant (DN) and inhibits cell-cell adhesion activity of E-cadherin, but its mode of action remains to be elucidated. Nectin is a Ca2+-independent immunoglobulin-like cell-cell adhesion molecule at AJs and is associated with E-cadherin through their respective peripheral membrane proteins, afadin and catenins, which connect nectin and cadherin to the actin cytoskeleton, respectively. We showed here that overexpression of nectin capable of binding afadin, but not a mutant incapable of binding afadin, reduced the inhibitory effect of N-cadherin DN on the cell-cell adhesion activity of E-cadherin in keratinocytes. Overexpressed nectin recruited N-cadherin DN to the nectin-based cell-cell adhesion sites in an afadin-dependent manner. Moreover, overexpression of nectin enhanced the E-cadherin–based cell-cell adhesion activity. These results suggest that N-cadherin DN competitively inhibits the association of the endogenous nectin-afadin system with the endogenous E-cadherin-catenin system and thereby reduces the cell-cell adhesion activity of E-cadherin. Thus, nectin plays a role in the formation of E-cadherin–based AJs in keratinocytes.

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 Regulation of cell–cell adhesion in prostate cancer cells by microRNA-96 through upregulation of E-Cadherin and EpCAM

2019 ◽  
Vol 41 (7) ◽  
pp. 865-874
Author(s):  
Gjendine Voss ◽  
Benedikta S Haflidadóttir ◽  
Helena Järemo ◽  
Margareta Persson ◽  
Tina Catela Ivkovic ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Adhesion ◽  
Bone Metastasis ◽  
Cancer Cells ◽  
Molecular Mechanisms ◽  
Cell Interaction ◽  
Metastatic Progression ◽  
Prostate Cancer Cells ◽  
E Cadherin ◽  
Cell Cell

Abstract Prostate cancer is one of the most common cancers in men, yet the biology behind lethal disease progression and bone metastasis is poorly understood. In this study, we found elevated levels of microRNA-96 (miR-96) in prostate cancer bone metastasis samples. To determine the molecular mechanisms by which miR-96 deregulation contributes to metastatic progression, we performed an Argonaute2-immunoprecipitation assay, in which mRNAs associated with cell–cell interaction were enriched. The expression of two cell adhesion molecules, E-Cadherin and EpCAM, was upregulated by miR-96, and potential targets sites were identified in the coding sequences of their mRNAs. We further showed that miR-96 enhanced cell–cell adhesion between prostate cancer cells as well as their ability to bind to osteoblasts. Our findings suggest that increased levels of miR-96 give prostate cancer cells an advantage at forming metastases in the bone microenvironment due to increased cell–cell interaction. We propose that miR-96 promotes bone metastasis in prostate cancer patients by facilitating the outgrowth of macroscopic tumours in the bone.

scholarly journals Different effects of dominant negative mutants of desmocollin and desmoglein on the cell-cell adhesion of keratinocytes

2000 ◽  
Vol 113 (10) ◽  
pp. 1803-1811
Author(s):  
Y. Hanakawa ◽  
M. Amagai ◽  
Y. Shirakata ◽  
K. Sayama ◽  
K. Hashimoto
Keyword(s):  
Cell Adhesion ◽  
Adherens Junctions ◽  
Dominant Negative ◽  
Cell Contact ◽  
Beta Catenin ◽  
Hacat Cells ◽  
Subsequent Formation ◽  
Contact Sites ◽  
E Cadherin ◽  
Cell Cell

Desmosomes contain two types of cadherin: desmocollin (Dsc) and desmoglein (Dsg). In this study, we examined the different roles that Dsc and Dsg play in the formation of desmosomes, by using dominant-negative mutants. We constructed recombinant adenoviruses (Ad) containing truncated mutants of E-cadherin, desmocollin 3a, and desmoglein 3 lacking a large part of their extracellular domains (EcaddeltaEC, Dsc3adeltaEC, Dsg3deltaEC), using the Cre-loxP Ad system to circumvent the problem of the toxicity of the mutants to virus-producing cells. When Dsc3adeltaEC Ad-infected HaCaT cells were cultured with high levels of calcium, E-cadherin and beta-catenin, which are marker molecules for the adherens junction, disappeared from the cell-cell contact sites, and cell-cell adhesion was disrupted. This also occurred in the cells infected with EcaddeltaEC Ad. With Dsg3deltaEC Ad infection, keratin insertion at the cell-cell contact sites was inhibited and desmoplakin, a marker of desmosomes, was stained in perinuclear dots while the adherens junctions remained intact. Dsc3adeltaEC Ad inhibited the induction of adherens junctions and the subsequent formation of desmosomes with the calcium shift, while Dsg3deltaEC Ad only inhibited the formation of desmosomes. To further determine whether Dsc3adeltaEC directly affected adherens junctions, mouse fibroblast L cells transfected with E-cadherin (LEC5) were infected with these mutant Ads. Both Dsc3adeltaEC and EcaddeltaEC inhibited the cell-cell adhesion of LEC5 cells, as determined by the cell aggregation assay, while Dsg3deltaEC did not. These results indicate that the dominant negative effects of Dsg3deltaEC were restricted to desmosomes, while those of Dsc3adeltaEC were observed in both desmosomes and adherens junctions. Furthermore, the cytoplasmic domain of Dsc3adeltaEC coprecipitated both plakoglobin and beta-catenin in HaCaT cells. In addition, beta-catenin was found to bind the endogenous Dsc in HaCaT cells. These findings lead us to speculate that Dsc interacts with components of the adherens junctions through beta-catenin, and plays a role in nucleating desmosomes after the adherens junctions have been established.

scholarly journals A Novel Role of Nectins in Inhibition of the E-Cadherin–induced Activation of Rac and Formation of Cell-Cell Adherens Junctions

2004 ◽  
Vol 15 (3) ◽  
pp. 1077-1088 ◽  
Author(s):  
Takashi Hoshino ◽  
Kazuya Shimizu ◽  
Tomoyuki Honda ◽  
Tomomi Kawakatsu ◽  
Taihei Fukuyama ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
G Protein ◽  
Adherens Junctions ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Inhibitory Effect ◽  
Small G Protein ◽  
E Cadherin ◽  
Cell Cell

Nectins are Ca2+-independent immunoglobulin (Ig)-like cell-cell adhesion molecules. The trans-interactions of nectins recruit cadherins to the nectin-based cell-cell adhesion, resulting in formation of cell-cell adherens junctions (AJs) in epithelial cells and fibroblasts. The trans-interaction of E-cadherin induces activation of Rac small G protein, whereas the trans-interactions of nectins induce activation of not only Rac but also Cdc42 small G protein. We showed by the fluorescent resonance energy transfer (FRET) imaging that the trans-interaction of E-cadherin induced dynamic activation and inactivation of Rac, which led to dynamic formation and retraction of lamellipodia. Moreover, we found here that the nectins, which did not trans-interact with other nectins (non–trans-interacting nectins), inhibited the E-cadherin–induced activation of Rac and reduced the velocity of the formation of the E-cadherin-based cell-cell AJs. The inhibitory effect of non–trans-interacting nectins was suppressed by the activation of Cdc42 induced by the trans-interactions of nectins. These results indicate a novel role of nectins in regulation of the E-cadherin–induced activation of Rac and formation of cell-cell AJs.

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.

Rescuing the N-cadherin knockout by cardiac-specific expression of N- or E-cadherin

Development ◽  
2001 ◽  
Vol 128 (4) ◽  
pp. 459-469
Author(s):  
Y. Luo ◽  
M. Ferreira-Cornwell ◽  
H. Baldwin ◽  
I. Kostetskii ◽  
J. Lenox ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Heart Development ◽  
Critical Role ◽  
Embryonic Cells ◽  
Wild Type ◽  
Specific Expression ◽  
Developmental Defects ◽  
E Cadherin ◽  
Cell Cell

Cell-cell adhesion mediated by some members of the cadherin family is essential for embryonic survival. The N-cadherin-null embryo dies during mid-gestation, with multiple developmental defects. We show that N-cadherin-null embryos expressing cadherins using muscle-specific promoters, alpha- or beta-myosin heavy chain, are partially rescued. Somewhat surprisingly, either N-cadherin or E-cadherin was effective in rescuing the embryos. The rescued embryos exhibited an increased number of somites, branchial arches and the presence of forelimb buds; however, in contrast, brain development was severely impaired. In rescued animals, the aberrant yolk sac morphology seen in N-cadherin-null embryos was corrected, demonstrating that this phenotype was secondary to the cardiac defect. Dye injection studies and analysis of chimeric animals that have both wild-type and N-cadherin-null cells support the conclusion that obstruction of the cardiac outflow tract represents a major defect that is likely to be the primary cause of pericardial swelling seen in null embryos. Although rescued embryos were more developed than null embryos, they were smaller than wild-type embryos, even though the integrity of the cardiovascular system appeared normal. The smaller size of rescued embryos may be due, at least in part, to increased apoptosis observed in tissues not rescued by transgene expression, indicating that N-cadherin-mediated cell adhesion provides an essential survival signal for embryonic cells. Our data provide in vivo evidence that cadherin adhesion is essential for cell survival and for normal heart development. Our data also show that E-cadherin can functionally substitute for N-cadherin during cardiogenesis, suggesting a critical role for cadherin-mediated cell-cell adhesion, but not cadherin family member-specific signaling, at the looping stage of heart development.

scholarly journals Enhanced RhoA signaling stabilizes E-cadherin in migrating epithelial monolayers.

2021 ◽  
Author(s):  
Shafali Gupta ◽  
Kinga Duszyc ◽  
Suzie Verma ◽  
Srikanth Budnar ◽  
Xuan Liang ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Mechanical Stress ◽  
Adherens Junctions ◽  
Mechanical Tension ◽  
Epithelial Migration ◽  
Epithelial Monolayers ◽  
Rhoa Signaling ◽  
Mcf 7 ◽  
E Cadherin ◽  
Cell Cell

Epithelia migrate as physically coherent populations of cells. Earlier studies revealed that mechanical stress accumulates in these cellular layers as they move. These stresses are characteristically tensile in nature and have often been inferred to arise when moving cells pull upon the cell-cell adhesions that hold them together. We now report that epithelial tension at adherens junctions between migrating cells also reflects an increase in RhoAmediated junctional contractility. We find that active RhoA levels were stimulated by p114 RhoGEF at the junctions between migrating MCF-7 monolayers, and this is accompanied by increased levels of actomyosin and mechanical tension. By applying a strategy to restore active RhoA specifically at adherens junctions by manipulating its scaffold, anillin, we found that this junctional RhoA signal was necessary to stabilize junctional E-cadherin during epithelial migration. We suggest that stabilization of E-cadherin by RhoA serves to increase cell-cell adhesion against the mechanical stresses of migration.

scholarly journals Enhanced RhoA signaling stabilizes E-cadherin in migrating epithelial monolayers

2021 ◽  
Author(s):  
Shafali Gupta ◽  
Kinga Duszyc ◽  
Suzie Verma ◽  
Srikanth Budnar ◽  
Xuan Liang ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Mechanical Stress ◽  
Adherens Junctions ◽  
Mechanical Tension ◽  
Epithelial Migration ◽  
Epithelial Monolayers ◽  
Rhoa Signaling ◽  
Mcf 7 ◽  
E Cadherin ◽  
Cell Cell

Epithelia migrate as physically coherent populations of cells. Earlier studies revealed that mechanical stress accumulates in these cellular layers as they move. These stresses are characteristically tensile in nature and have often been inferred to arise when moving cells pull upon the cell-cell adhesions that hold them together. We now report that epithelial tension at adherens junctions between migrating cells also increases due to an increase in RhoA-mediated junctional contractility. We find that active RhoA levels were stimulated by p114 RhoGEF at the junctions between migrating MCF-7 monolayers, and this was accompanied by increased levels of actomyosin and mechanical tension. Applying a strategy to restore active RhoA specifically at adherens junctions by manipulating its scaffold, anillin, we found that this junctional RhoA signal was necessary to stabilize junctional E-cadherin during epithelial migration and promoted orderly collective movement. We suggest that stabilization of E-cadherin by RhoA serves to increase cell-cell adhesion against the mechanical stresses of migration.

scholarly journals The kinesin-8 member Kif19 alters microtubule dynamics, suppresses cell adhesion, and promotes cancer cell invasion

2020 ◽  
Author(s):  
Samuel C. Eisenberg ◽  
Abhinav Dey ◽  
Rayna Birnbaum ◽  
David J. Sharp
Keyword(s):  
Cell Adhesion ◽  
Cancer Cell ◽  
Cell Invasion ◽  
Adherens Junctions ◽  
Focal Adhesions ◽  
Microtubule Dynamics ◽  
Surrounding Tissue ◽  
Cancer Cell Invasion ◽  
E Cadherin ◽  
Cell Cell

AbstractMetastasis is one of the deadliest aspects of cancer. Initial Metastatic spread is dependent on the detachment and dissemination of cells from a parent tumor, and invasion into the surrounding tissue. In this study, we characterize the kinesin-8 member Kif19 as a promoter of cancer cell invasion that suppresses cell-cell adherens junctions and cell-matrix focal adhesions. Initial analysis of publicly available cancer patient data sets demonstrated that Kif19 expression correlates with worse overall survival probability in several cancers and that Kif19 expression is increased in metastases of colorectal and breast carcinoma compared to the primary tumor. Depletion of Kif19 from two human cancer cell lines (DMS53 and MDA-MB-231) did not alter viability, but decreased the cells’ ability to invade a Matrigel matrix by half and impaired the invasion of spheroids into a primary cell monolayer. Ectopically expressed Kif19 localized to, and partially depolymerized, microtubules in the cell periphery. However, Kif19 depletion increased microtubule dynamicity and sensitivity to pharmacological depolymerization without altering total microtubule polymer levels. These data indicate that Kif19 can both depolymerize and stabilize microtubules. Given this activity, we then studied Kif19’s effect on focal adhesions and adherens junctions, which are both regulated by microtubule dynamics. Kif19 knockdown increased the proportion of cell surface area covered by Vinculin focal adhesions. Further, Kif19 depletion increased whole cell E-cadherin expression and the accumulation of E-cadherin at cell-cell adherens junctions. Conversely, ectopic overexpression of full-length Kif19 led to proportionally smaller focal adhesions and impaired E-cadherin accumulation at cell-cell junctions. Our current hypothesis is that aberrant Kif19 expression in cancer alters focal adhesion dynamics and suppresses E-cadherin expression, which enhance cell invasiveness. Further, we propose that these changes in cell adhesion are due to modification of peripheral microtubule dynamics by Kif19, potentially through disruption of local rho GTPase activity.

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