Search

ß-Catenin is an essential element for the transcriptional activation of target genes in the Wnt signaling cascade and is also a cell adhesion molecule that couples with cadherins. Although plakoglobin (?-catenin), a closely related homologue of ß-catenin, is also known to be a cell adhesion molecule, its function as a transcriptional factor has not been revealed in detail. Using a human malignant mesothelioma cell line, NCI-H28, in which we have identified a homozygous deletion of the ß-catenin gene, we studied whether plakoglobin has a T-cell factor/lymphocyte enhancer factor (TCF/LEF) family-dependent transcriptional activity. Transfection with the wild-type plakoglobin expression vector induced accumulation of plakoglobin in the nucleus. Immunoprecipitation assay with cotransfection of plakoglobin and either TCF-4 or LEF-1 detected binding of plakoglobin to TCF-4 or LEF-1. Luciferase reporter assay demonstrated transcriptional activity of the wild-type plakoglobin when transfected with TCF/LEF, although plakoglobin showed less activity than ß-catenin. Exogenous plakoglobin was also shown to promote entrance of exogenous ß-catenin into the nuclei. Furthermore, small interfering RNA directed against plakoglobin suppressed expression of endogenous plakoglobin and its transcriptional activity, suggesting that endogenous plakoglobin has a weak transcriptional activity. These results suggest that plakoglobin can activate the Wnt signaling cascade directly without interaction of ß-catenin, and that plakoglobin has multiple functions as a transcriptional activator and a cell adhesion molecule like ß-catenin.Oncogene (2004) 23, 964-972. doi:10.1038/sj.onc.1207254 Published online 8 December 2003 [ABSTRACT FROM AUTHOR]

We have found that a malignant mesothelioma cell line, NCI-H28, had a chromosome 3p21.3 homozygous deletion containing the ß-catenin gene (CTNNB1), which suggested that the deletion of ß-catenin might have a growth advantage in the development of this tumor. To determine whether ß-catenin has a growth-inhibitory activity, we transfected wild-type ß-catenin, Ser37Cys mutant ß-catenin as an activated type, and C-terminus deletion mutant ß-catenin that lacks the transcription activity, into the NCI-H28 cells. A non-small cell lung cancer cell line, NCI-H1299, which expressed endogenous ß-catenin, was also studied. We tested the localization of exogenous ß-catenin in the NCI-H28 cells with immunofluorescence, and found that the wild-type ß-catenin and the C-terminus deletion mutant were more strongly expressed in the plasma membrane and cytoplasm than in the nucleus, while the Ser37Cys mutant was more in the nucleus than in the cytoplasm. By using luciferase-reporter assay, the ß-catenin/T-cell factor 4-mediated transactivity of the Ser37Cys mutant was shown to be higher than that of the wild-type ß-catenin in both cell lines. However, the transactivity of the C-terminus deletion mutant was strongly reduced in both. Colony formation of the NCI-H28 cells was reduced by 50% after transfection with the wild-type ß-catenin, and 60% with the Ser37Cys mutant, but only 20% with the C-terminus deletion mutant compared to the vector control. Inhibition of colony formation in NCI-H28 cells was because of apoptosis, manifested by positive staining of Annexin V and TUNEL assays in transfected cells. In contrast, when transfected with the wild-type ß-catenin, no significant reduction in colony formation was seen in ß-catenin wild-type NCI-H1299 cells. In conclusion, our data indicate that inactivation of ß-catenin by a 3p21.3 homozygous deletion might be a crucial event in the development of the mesothelioma NCI-H28 cells. Thus, while ß-catenin is well known to be a positive growth-stimulating factor for many human cancers, it can also act as a potential growth suppressor in some types of human cancer cells.Oncogene (2003) 22, 5978-5986. doi:10.1038/sj.onc.1206533 [ABSTRACT FROM AUTHOR]

Glial cell line-derived neurotrophic factor (GDNF) signals through a unique receptor system that includes Ret receptor tyrosine kinase and a glycosyl-phosphatidylinositol-linked cell surface protein. In the present study, we have identified several proteins in neuroblastoma cells that are phosphorylated on tyrosine in response to GDNF. The phosphorylated proteins include focal adhesion kinase (FAK), paxillin and Crk-associated substrate, p130Cas, all of which are known to be associated with focal adhesions. Of these, paxillin and p130Cas interacted with Crk proteins in GDNF-treated neuroblastoma cells. GDNF also induced reorganization of the actin cytoskelton. Tyrosine phosphorylation of FAK, paxillin and p130Cas was inhibited by cytochalasin D or two specific inhibitors of phosphatidylinositol-3′ kinase (PI-3′ kinase), wortmannin and LY294002, indicating that their tyrosine phosphorylation depends on the formation of actin stress fiber and activation of PI-3′ kinase. In addition, phosphorylation of FAK but not of paxillin and p130Cas was markedly impaired by the Clostridium botulinum C3 exoenzyme that specifically ADP-ribosylates and inactivates Rho. These results suggested the presence of Rho-dependent and -independent signaling pathways downstream of PI-3′ kinase that mediate tyrosine phosphorylation of FAK, paxillin and p130Cas through Ret kinase. [ABSTRACT FROM AUTHOR]

We have found that a malignant mesothelioma cell line, NCI-H28, had a chromosome 3p21.3 homozygous deletion containing the beta-catenin gene (CTNNB1), which suggested that the deletion of beta-catenin might have a growth advantage in the development of this tumor. To determine whether beta-catenin has a growth-inhibitory activity, we transfected wild-type beta-catenin, Ser37Cys mutant beta-catenin as an activated type, and C-terminus deletion mutant beta-catenin that lacks the transcription activity, into the NCI-H28 cells. A non-small cell lung cancer cell line, NCI-H1299, which expressed endogenous beta-catenin, was also studied. We tested the localization of exogenous beta-catenin in the NCI-H28 cells with immunofluorescence, and found that the wild-type beta-catenin and the C-terminus deletion mutant were more strongly expressed in the plasma membrane and cytoplasm than in the nucleus, while the Ser37Cys mutant was more in the nucleus than in the cytoplasm. By using luciferase-reporter assay, the beta-catenin/T-cell factor 4-mediated transactivity of the Ser37Cys mutant was shown to be higher than that of the wild-type beta-catenin in both cell lines. However, the transactivity of the C-terminus deletion mutant was strongly reduced in both. Colony formation of the NCI-H28 cells was reduced by 50% after transfection with the wild-type beta-catenin, and 60% with the Ser37Cys mutant, but only 20% with the C-terminus deletion mutant compared to the vector control. Inhibition of colony formation in NCI-H28 cells was because of apoptosis, manifested by positive staining of Annexin V and TUNEL assays in transfected cells. In contrast, when transfected with the wild-type beta-catenin, no significant reduction in colony formation was seen in beta-catenin wild-type NCI-H1299 cells. In conclusion, our data indicate that inactivation of beta-catenin by a 3p21.3 homozygous deletion might be a crucial event in the development of the mesothelioma NCI-H28 cells. Thus, while beta-catenin is well known to be a positive growth-stimulating factor for many human cancers, it can also act as a potential growth suppressor in some types of human cancer cells.