BackgroundThe different mechanisms involved in p120-catenin (p120ctn) isoforms' 1/3 regulation of cell cycle progression are still not elucidated to date.Methods and FindingsWe found that both cyclin D1 and cyclin E could be effectively restored by restitution of p120ctn-1A or p120ctn-3A in p120ctn depleted lung cancer cells. When the expression of cyclin D1 was blocked by co-transfection with siRNA-cyclin D1 in p120ctn depleted cells restoring p120ctn-1A or 3A, the expression of cyclin E was slightly decreased, not increased, implying that p120ctn isoforms 1 and 3 cannot up-regulate cyclin E directly but may do so through up-regulation of cyclin D1. Interestingly, overexpression of p120ctn-1A increased β-catenin and cyclin D1 expression, while co-transfection with siRNA targeting β-catenin abolishes the effect of p120ctn-1A on up-regulation of cyclin D1, suggesting a role of β-catenin in mediating p120ctn-1A's regulatory function on cyclin D1 expression. On the other hand, overexpression of p120ctn isoform 3A reduced nuclear Kaiso localization, thus decreasing the binding of Kaiso to KBS on the cyclin D1 promoter and thereby enhancing the expression of cyclin D1 gene by relieving the repressor effect of Kaiso. Because overexpressing NLS-p120ctn-3A (p120ctn-3A nuclear target localization plasmids) or inhibiting nuclear export of p120ctn-3 by Leptomycin B (LMB) caused translocation of Kaiso to the nucleus, it is plausible that the nuclear export of Kaiso is p120ctn-3-dependent.ConclusionsOur results suggest that p120ctn isoforms 1 and 3 up-regulate cyclin D1, and thereby cyclin E, resulting in the promotion of cell proliferation and cell cycle progression in lung cancer cells probably via different protein mediators, namely, β-catenin for isoform 1 and Kaiso, a negative transcriptional factor of cyclin D1, for isoform 3.

Ezrin, one of the ezrin-radixin-moesin proteins, is involved in the formation of cell membrane processes such as lamellipodia and filopodia and acts as a membrane-cytoskeleton linker. Its aberrant expression correlates with development and progression of several human cancers. However, the expression of ezrin and its role in lung cancer are currently unknown. In this study, we performed ezrin small interfering RNA transfection in two lung cancer cell lines and examined the effects on malignant phenotypes in cancer cells by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, wound healing, and chamber transwell assays. Ezrin knockdown significantly reduced the proliferation, migration, and invasion of lung cancer cells in vitro. To address the possible mechanisms, we evaluated the expression of adhesion molecules E-cadherin and β-catenin by Western blot and reverse transcriptase-polymerase chain reaction analyses. The results demonstrated that downregulation of ezrin reduced β-catenin and increased E-cadherin at the protein level but had no effects on their mRNA levels, suggesting posttranscriptional regulation of these two adhesion molecules. Immunofluorescence assays revealed that ezrin knockdown restored membranous expression of E-cadherin and decreased cytoplasmic β-catenin in lung cancer cells. In addition, ezrin expression was immunohistochemically evaluated on 135 normal and 183 lung cancer tissues. The expression of ezrin was significantly higher in cancer samples than paired autologous normal lung tissues. In normal bronchial epithelium, ezrin was mainly localized on the apical membrane, while in lung cancers and metastatic foci, ezrin was primarily distributed in cytoplasm. Among lung cancer tissues, expression of ezrin was higher in the invasive front of primary lesions and the highest in lymphatic metastasis. Statistical analysis demonstrated that ezrin expression correlated significantly with lymphatic metastasis and advanced TNM stage. Our data suggest that ezrin may play a crucial role in governing the biological behavior of lung cancer.

Crk‐Like (CRKL) is an adapter protein that has crucial roles in multiple biological processes, including cell proliferation, adhesion, and migration. Amplification of CRKL gene was found in non‐small cell lung cancer (NSCLC). However, the expression pattern of CRKL protein and its clinical significance in human NSCLC have not been well characterized to date. In this study, expression of CRKL was evaluated in 131 NSCLC tissues by immumohistochemistry. CRKL protein was up‐regulated in the lung carcinomas compared with adjacent normal lung tissue. Overexpression of CRKL was found in 58 of 131 (44.3%) NSCLC samples and correlated with poor tumor differentiation (P = 0.0042), histological type (adenocarcinoma; P = 0.001), advanced p‐TNM stage (P = 0.0004), nodal metastasis (P = 0.0273), high proliferation index (P = 0.0062) and poor overall survival (P = 0.0084). Further univariate and multivariate analysis showed a significant association of CRKL overexpression and worse overall survival in lung cancer patients. In addition, overexpression of CRKL in HBE and H1299 cell lines promoted cell proliferation by facilitating cell cycle progression. Further analysis of cell cycle related molecules showed that CRKL induced cyclin D1, cyclin B1 expression, and increased Rb phosphorylation. In conclusion, this study demonstrated overexpression of CRKL correlated with poor prognosis and lung cancer proliferation by cell cycle regulation. © 2012 Wiley Periodicals, Inc.