NrCAM, a neuronal system cell-adhesion molecule, is induced in papillary thyroid carcinomas

NrCAM (neuron-glia-related cell-adhesion molecule) is primarily, although not solely, expressed in the nervous system. In the present study, NrCAM expression was analysed in a series (46) of papillary thyroid carcinomas (PTCs) and paired normal tissues (NT). Quantitative reverse transcriptase (QRT)-PCR revealed that NrCAM expression was upregulated in all PTCs compared to normal thyroid, whatever the stage or size of the primary tumour. NrCAM transcript levels were 1.3- to 30.7-fold higher in PTCs than in NT. Immunohistochemistry (IHC) confirmed that the expression of NrCAM was considerably higher in tumours (score 2+/3+) than in adjacent normal paratumoural thyroid tissue. The NrCAM protein was detected in all but three (93.3%) PTC samples, and it was mainly cytoplasmic; in some cases there was additional membranous localisation – basolateral and partly apical. In the normal thyroid and tissues surrounding tumours, focal NrCAM immunolabelling was seen only in follicles containing tall cells, where staining was restricted to the apical pole of thyrocytes. Western blot analysis corroborated the QRT–PCR and IHC results, showing higher NrCAM protein levels in PTCs than in paired NT. The level of overexpression of the NrCAM mRNA in tumourous tissue appeared to be independent of the primary tumour stage (pT) or the size of the PTC. These data provide the first evidence that NrCAM is overexpressed in human PTCs at the mRNA and protein levels, whatever the tumour stage. Thus, the induction and upregulation of NrCAM expression could be implicated in the pathogenesis and behaviour of papillary thyroid cancers

Effects of small interfering RNAs targeting fascin on human esophageal squamous cell carcinoma cell lines

<p>Abstract</p> <p>Background</p> <p>Fascin induces membrane protrusions and cell motility. Fascin overexpression was associated with poor prognosis, and its downregulation reduces cell motility and invasiveness in esophageal squamous cell carcinoma (ESCC). Using a stable knockdown cell line, we revealed the effect of fascin on cell growth, cell adhesion and tumor formation.</p> <p>Methods</p> <p>We examined whether fascin is a potential target in ESCC using <it>in vitro </it>and <it>in vivo </it>studies utilizing a specific siRNA. We established a stable transfectant with downregulated fascin from KYSE170 cell line.</p> <p>Results</p> <p>The fascin downregulated cell lines showed a slower growth pattern by 40.3% (p < 0.01) and detachment from collagen-coated plates by 53.6% (p < 0.01), compared to mock cells, suggesting that fascin plays a role in cell growth by maintaining cell adhesion to the extracellular matrix. <it>In vivo</it>, the tumor size was significantly smaller in the tumor with fascin knockdown cells than in mock cells by 95% at 30 days after inoculation.</p> <p>Conclusions</p> <p>These findings suggest that fascin overexpression plays a role in tumor growth and progression in ESCC and that cell death caused by its downregulation might be induced by cell adhesion loss. This indicates that targeting fascin pathway could be a novel therapeutic strategy for the human ESCC.</p

N-Terminal 1–54 Amino Acid Sequence and Armadillo Repeat Domain Are Indispensable for P120-Catenin Isoform 1A in Regulating E-Cadherin

P120-catenin (p120ctn) exerts important roles in regulating E-cadherin and invasiveness in cancer cells. However, the mechanisms by which p120ctn isoforms 1 and 3 modulate E-cadherin expression are poorly understood. In the current study, HBE, H460, SPC and LTE cell lines were used to examine the effects of p120ctn isoforms 1A and 3A on E-cadherin expression and cell invasiveness. E-cadherin was localized on the cell membrane of HBE and H460 cells, while it was confined to the cytoplasm in SPC and LTE cells. Depletion of endogenous p120ctn resulted in reduced E-cadherin expression; however, p120ctn ablation showed opposite effects on invasiveness in the cell lines by decreasing invasiveness in SPC and LTE cells and increasing it in HBE and H460 cells. Restitution of 120ctn isoform 1A restored E-cadherin on the cell membrane and blocked cell invasiveness in H460 and HBE cells, while it restored cytoplasmic E-cadherin and enhanced cell invasiveness in SPC and LTE cells. P120ctn isoform 3A increased the invasiveness in all four cell lines despite the lack of effect on E-cadherin expression, suggesting a regulatory pathway independent of E-cadherin. Moreover, five p120ctn isoform 1A deletion mutants were constructed and expressed in H460 and SPC cells. The results showed that only the M4 mutant, which contains N-terminal 1–54 amino acids and the Armadillo repeat domain, was functional in regulating E-cadherin and cell invasiveness, as observed in p120ctn isoform 1A. In conclusion, the N-terminal 1–54 amino acid sequence and Armadillo repeat domain of p120ctn isoform 1A are indispensable for regulating E-cadherin protein. P120ctn isoform 1A exerts opposing effects on cell invasiveness, corresponding to the subcellular localization of E-cadherin

Phosphorylation of LCRMP-1 by GSK3β Promotes Filopoda Formation, Migration and Invasion Abilities in Lung Cancer Cells

LCRMP-1, a novel isoform of CRMP-1, can promote cancer cell migration, invasion and associate with poor clinical outcome in patients with non-small-cell lung cancer (NSCLC). However, the underlying regulatory mechanisms of LCRMP-1 in cancer cell invasiveness still remain obscure. Here, we report that GSK3β can phosphorylate LCRMP-1 at Thr-628 in consensus sequences and this phosphorylation is crucial for function of LCRMP-1 to promote filopodia formation, migration and invasion in cancer cells. Impediment of Thr-628 phosphorylation attenuates the stimulatory effects of LCRMP-1 on filopodia forming, migration and invasion abilities in cancer cells; simultaneously, kinase-dead GSK3β diminishes regulation of LCRMP-1 on cancer cell invasion. Furthermore, we also found that patients with low-level Ser-9-phosphorylated GSK3β expression and high-level LCRMP-1 expression have worse overall survival than those with high-level inactive GSK3β expressions and low-level LCRMP-1 expressions (P<0.0001). Collectively, these results demonstrate that GSK3β-dependent phosphorylation of LCRMP-1 provides an important mechanism for regulation of LCRMP-1 on cancer cell invasiveness and clinical outcome

Upregulation of calcium-sensing receptor and mitogen-activated protein kinase signalling in the regulation of growth and differentiation in colon carcinoma

In the present study, we demonstrate that Ca2+-induced growth inhibition and induction of differentiation in a line of human colon carcinoma cells (CBS) is dependent on mitogen-activated protein (MAP) kinase signaling and is associated with upregulation of extracellular calcium-sensing receptor (CaSR) expression. When CBS cells were grown in Ca2+-free medium and then switched to medium supplemented with 1.4 mM Ca2+, proliferation was reduced and morphologic features of differentiation were expressed. E-cadherin, which was minimally expressed in nonsupplemented medium, was rapidly induced in response to Ca2+ stimulation. Sustained activation of the extracellular signal-regulated kinase (ERK) occured in Ca2+-supplemented medium. When an inhibitor of ERK activation (10 μM U0126) was included in the Ca2+-supplemented culture medium, ERK-activation did not occur. Concomitantly, E-cadherin was not induced, cell proliferation remained high and differentiation was not observed. The same level of Ca2+ supplementation that induced MAP kinase activation also stimulated CaSR upregulation in CBS cells. A clonal isolate of the CBS line that did not upregulate CaSR expression in response to extracellular Ca2+ was isolated from the parent cells. This isolate failed to produce E-cadherin or undergo growth inhibition/induction of differentiation when exposed to Ca2+ in the culture medium. However, ERK-activation occurred as efficiently in this isolate as in parent CBS cells or in a cloned isolate that underwent growth reduction and differentiation in response to Ca2+ stimulation. Together, these data indicate that CaSR upregulation and MAP kinase signalling are both intermediates in the control of colon carcinoma cell growth and differentiation. They appear to function, at least in part, independently of one another

β-Catenin is involved in alterations in mitochondrial activity in non-transformed intestinal epithelial and colon cancer cells

BACKGROUND: Alteration in respiratory activity and mitochondrial DNA (mtDNA) transcription seems to be an important feature of cancer cells. Leukotriene D(4) (LTD(4)) is a proinflammatory mediator implicated in the pathology of chronic inflammation and cancer. We have shown earlier that LTD(4) causes translocation of beta-catenin both to the mitochondria, in which it associates with the survival protein Bcl-2 identifying a novel role for beta-catenin in cell survival, and to the nucleus in which it activates the TCF/LEF transcription machinery. METHODS: Here we have used non-transformed intestinal epithelial Int 407 cells and Caco-2 colon cancer cells, transfected or not with wild type and mutated (S33Y) beta-catenin to analyse its effect on mitochondria activity. We have measured the ATP/ADP ratio, and transcription of the mtDNA genes ND2, ND6 and 16 s in these cells stimulated or not with LTD(4). RESULTS: We have shown for the first time that LTD(4) triggers a cellular increase in NADPH dehydrogenase activity and ATP/ADP ratio. In addition, LTD(4) significantly increased the transcription of mtDNA genes. Overexpression of wild-type beta-catenin or a constitutively active beta-catenin mutant mimicked the effect of LTD(4) on ATP/ADP ratio and mtDNA transcription. These elevations in mitochondrial activity resulted in increased reactive oxygen species levels and subsequent activations of the p65 subunit of NF-kappaB. CONCLUSIONS: The present novel data show that LTD(4), presumably through beta-catenin accumulation in the mitochondria, affects mitochondrial activity, lending further credence to the idea that inflammatory signalling pathways are intrinsically linked with potential oncogenic signals

γ-Catenin is overexpressed in acute myeloid leukemia and promotes the stabilization and nuclear localization of β-catenin

Canonical Wnt signaling regulates the transcription of T-cell factor (TCF)-responsive genes through the stabilization and nuclear translocation of the transcriptional co-activator, β-catenin. Overexpression of β-catenin features prominently in acute myeloid leukemia (AML) and has previously been associated with poor clinical outcome. Overexpression of γ-catenin mRNA (a close homologue of β-catenin) has also been reported in AML and has been linked to the pathogenesis of this disease, however, the relative roles of these catenins in leukemia remains unclear. Here we report that overexpression and aberrant nuclear localization of γ-catenin is frequent in AML. Significantly, γ-catenin expression was associated with β-catenin stabilization and nuclear localization. Consistent with this, we found that ectopic γ-catenin expression promoted the stabilization and nuclear translocation of β-catenin in leukemia cells. β-Catenin knockdown demonstrated that both γ- and β-catenin contribute to TCF-dependent transcription in leukemia cells. These data indicate that γ-catenin expression is a significant factor in the stabilization of β-catenin in AML. We also show that although normal cells exclude nuclear translocation of both γ- and β-catenin, this level of regulation is lost in the majority of AML patients and cell lines, which allow nuclear accumulation of these catenins and inappropriate TCF-dependent transcription

NEDD9 Is a Positive Regulator of Epithelial-Mesenchymal Transition and Promotes Invasion in Aggressive Breast Cancer

Epithelial to mesenchymal transition (EMT) plays an important role in many biological processes. The latest studies revealed that aggressive breast cancer, especially the triple-negative breast cancer (TNBC) subtype was frequently associated with apparent EMT, but the mechanisms are still unclear. NEDD9/HEF1/Cas-L is a member of the Cas protein family and was identified as a metastasis marker in multiple cancer types. In this study, we wished to discern the role of NEDD9 in breast cancer progression and to investigate the molecular mechanism by which NEDD9 regulates EMT and promotes invasion in triple-negative breast cancer. We showed that expression of NEDD9 was frequently upregulated in TNBC cell lines, and in aggressive breast tumors, especially in TNBC subtype. Knockdown of endogenous NEDD9 reduced the migration, invasion and proliferation of TNBC cells. Moreover, ectopic overexpression of NEDD9 in mammary epithelial cells led to a string of events including the trigger of EMT, activation of ERK signaling, increase of several EMT-inducing transcription factors and promotion of their interactions with the E-cadherin promoter. Data presented in this report contribute to the understanding of the mechanisms by which NEDD9 promotes EMT, and provide useful clues to the evaluation of the potential of NEDD9 as a responsive molecular target for TNBC chemotherapy