Defining the Roles of .BETA.-catenin and Plakoglobin in Cell-cell Adhesion: Isolation of .BETA.-catenin/plakoglobin-deficient F9 Cells

Fukunaga, Yoshitaka; Liu, Huijie; Shimizu, Masayuki; Komiya, Satoshi; Kawasuji, Michio; Nagafuchi, Akira

doi:10.1247/csf.30.25

Cited by 58 publications

(66 citation statements)

References 38 publications

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“…Thus, complex formation with ␤-catenin seemed to be a prerequisite for the exit of cadherins from the intracellular compartments. Recently, an F9 cell line lacking both ␤-catenin and plakoglobin was established (Fukunaga et al, 2005). In these cells, E-cadherin was detected in intracellular compartments, suggesting that these proteins were required not only for cell adhesion activity but also for the cell-surface expression of E-cadherin.…”

Section: Discussionmentioning

confidence: 99%

A dileucine motif in its cytoplasmic domain directs β-catenin-uncoupled E-cadherin to the lysosome

Miyashita

Ozawa

2007

Journal of Cell Science

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2+-dependent cell-cell adhesion and is localized to the basolateral membrane of polarized epithelial cells. Uncoupling ␤-catenin from E-cadherin by deletion or substitution mutations causes accumulation of these proteins in intracellular compartments, including the trans-Golgi network and early endosomes, and degradation in lysosomes. Expression of a dominant-negative dynamin did not change the pattern of the mutant E-cadherin localization, indicating that the endocytosis of the protein from the cell surface does not contribute significantly to the accumulation of the protein in the intracellular compartments. Alternatively, E-cadherin lacking its entire cytoplasmic domain (tail-less E-cadherin) was detected on the surface of cells and targeted to the basolateral membrane. We found that 20 amino acid residues within the juxtamembrane region contain the signal responsible for intracellular accumulation and the lysosomal targeting of E-cadherin. A dileucine motif within this region seems crucial, because substitution of these residues to alanines resulted in efficient surface expression of the protein. The tail-less E-cadherin construct and the dileucinesubstitution construct were detected on the basolateral membranes. Thus, the dileucine motif of E-cadherin is not required for its basolateral targeting.

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Section: Discussionmentioning

confidence: 99%

A dileucine motif in its cytoplasmic domain directs β-catenin-uncoupled E-cadherin to the lysosome

Miyashita

Ozawa

2007

Journal of Cell Science

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“…Production of autocrine hGH has also been demonstrated to stimulate the phenotypic conversion of mammary epithelial cells towards the morphological and molecular characteristics of a mesenchymal cell, including expression of fibronectin and vimentin, and loss of plakoglobin expression (Mukhina et al, 2004). Plakoglobin is a component of the adherens junctions complex, and is a multifunctional protein involved in cell-cell adhesion and transcriptional regulation (Fukunaga et al, 2005). Loss of plakoglobin expression results in dissolution of adherens junctions and has been associated with neoplastic progression and tumour invasion (Savagner, 2001;Mukhina et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

DNMT3A and DNMT3B mediate autocrine hGH repression of plakoglobin gene transcription and consequent phenotypic conversion of mammary carcinoma cells

et al. 2007

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Directed by microarray analyses, we report that autocrine human growth hormone (hGH) increased the mRNA and protein expression of DNA methyltransferase 1 (DNMT1), DNMT3A and DNMT3B in mammary carcinoma cells. Autocrine hGH stimulation of DNMT3A and DNMT3B expression was mediated by JAK2 and Src kinases, and treatment of mammary carcinoma cells with the DNMT inhibitor, 5 0 -aza-2 0 -deoxycytidine (AZA), abrogated autocrine hGH-stimulated cellular proliferation, apoptosis and anchorage-independent growth. AZA reversed the epitheliomesenchymal transition of mammary carcinoma cells induced by autocrine hGH, to an epithelioid morphology and abrogated cell migration stimulated by autocrine hGH. Autocrine hGH-stimulated hypermethylation of the first exon of the PLAKOGLOBIN gene and AZA abrogated the ability of autocrine hGH to repress plakoglobin gene transcription. Small interfering RNA (siRNA)-mediated depletion of the individual DNMT molecules did not release autocrine hGH repression of PLAKOGLOBIN promoter activity nor did individual DNMT depletion affect autocrine hGH-stimulated migration. However, concomitant siRNA-mediated depletion of both DNMT3A and DNMT3B abrogated hypermethylation of the PLAKO-GLOBIN gene stimulated by autocrine hGH and subsequent repression of plakoglobin gene transcription and increased cell migration. Thus, the autocrine hGHstimulated increases in DNMT3A and DNMT3B expression mediate repression of plakoglobin gene transcription by direct hypermethylation of its promoter and consequent phenotypic conversion of mammary carcinoma cells. Autocrine hGH, therefore, utilizes DNA methylation as a mechanism to exert its oncogenic effects in mammary carcinoma cells.

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“…Plakoglobin binds to the cytoplasmic domains of classical cadherins and desmosomal cadherins in adherens junctions and desmosomes, respectively (2,17,21). In adherens junctions, ␤-catenin and plakoglobin compete for binding to classical cadherins (2,10).…”

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confidence: 99%

“…TCF-4 is expressed in these cells as a main partner of ␤-catenin (our unpublished observation). We recently succeeded in disrupting the genes encoding ␤-catenin and plakoglobin in F9 cells using gene-targeting technology (10). Since almost all of the exons encoding ␤-catenin and/or plakoglobin were deleted by our targeting vectors, we obtained cells that were completely null for ␤-catenin and/or plakoglobin.…”

mentioning

confidence: 99%

Defining the Roles of β-Catenin and Plakoglobin in LEF/T-Cell Factor-Dependent Transcription Using β-Catenin/Plakoglobin-Null F9 Cells

Shimizu

Fukunaga

Ikenouchi

et al. 2008

Molecular and Cellular Biology

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␤-Catenin functions as a transcriptional regulator in Wnt signaling. Its function is regulated by a specific destruction system. Plakoglobin is a close homologue of ␤-catenin in mammalian cells and is regulated in a similar fashion. When ␤-catenin or plakoglobin is exogenously expressed in cells, endogenous ␤-catenin is stabilized, which complicates estimation of the transcriptional activities of exogenously expressed proteins. To facilitate the design of experiments aimed at investigating the transcriptional activities of ␤-catenin and plakoglobin, we utilized F9 cells in which we knocked out endogenous ␤-catenin and/or plakoglobin by gene deletion and exogenously expressed wild-type and mutant ␤-catenin and/or plakoglobin. We show that Cterminally deleted ␤-catenin, but not plakoglobin, has a strong dominant-negative effect on transcription without altering the nuclear accumulation of ␤-catenin. Moreover, we show that Wnt-3a activation of LEF/Tcell factor (TCF)-dependent transcription depends on ␤-catenin but not on plakoglobin. Using chimeras of ␤-catenin and plakoglobin, we demonstrate that plakoglobin has the potential to function in transcriptional regulation but is not responsible for Wnt-3a signaling in F9 cells. Our data show that preferential nuclear accumulation of ␤-catenin is not necessarily linked to its transcriptional activity. We also clearly demonstrate that plakoglobin is insufficient for LEF/TCF-dependent transcriptional activation by Wnt-3a in F9 cells.

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Defining the Roles of .BETA.-catenin and Plakoglobin in Cell-cell Adhesion: Isolation of .BETA.-catenin/plakoglobin-deficient F9 Cells

Cited by 58 publications

References 38 publications

A dileucine motif in its cytoplasmic domain directs β-catenin-uncoupled E-cadherin to the lysosome

A dileucine motif in its cytoplasmic domain directs β-catenin-uncoupled E-cadherin to the lysosome

DNMT3A and DNMT3B mediate autocrine hGH repression of plakoglobin gene transcription and consequent phenotypic conversion of mammary carcinoma cells

Defining the Roles of β-Catenin and Plakoglobin in LEF/T-Cell Factor-Dependent Transcription Using β-Catenin/Plakoglobin-Null F9 Cells

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