NCAM-induced focal adhesion assembly: a functional switch upon loss of E-cadherin.

Loss of expression of the cell-cell adhesion molecule E-cadherin is a hallmark of epithelial-mesenchymal transition (EMT) in development and in the progression from epithelial tumours to invasive and metastatic cancers. Here, we demonstrate that the loss of E-cadherin function upregulates expression of the neuronal cell adhesion molecule (NCAM). Subsequently, a subset of NCAM translocates from fibroblast growth factor receptor (FGFR) complexes outside lipid rafts into lipid rafts where it stimulates the non-receptor tyrosine kinase p59(Fyn) leading to the phosphorylation and activation of focal adhesion kinase and the assembly of integrin-mediated focal adhesions. Ablation of NCAM expression during EMT inhibits focal adhesion assembly, cell spreading and EMT. Conversely, forced expression of NCAM induces epithelial cell delamination and migration, and high NCAM expression correlates with tumour invasion. These results establish a mechanistic link between the loss of E-cadherin expression, NCAM function, focal adhesion assembly and cell migration and invasion.

A genetic screen identifies mutations in the yeast WAR1 gene, linking transcription factor phosphorylation to weak-acid stress adaptation.

Exposure of the yeast Saccharomyces cerevisiae to weak organic acids such as the food preservatives sorbate, benzoate and propionate leads to the pronounced induction of the plasma membrane ATP-binding cassette (ABC) transporter, Pdr12p. This protein mediates efflux of weak acid anions, which is essential for stress adaptation. Recently, we identified War1p as the dedicated transcriptional regulator required for PDR12 stress induction. Here, we report the results from a genetic screen that led to the isolation of two war1 alleles encoding mutant variants, War1-28p and War1-42p, which are unable to support cell growth in the presence of sorbate. DNA sequencing revealed that War1-28 encodes a truncated form of the transcriptional regulator, and War1-42 carries three clustered mutations near the C-terminal activation domain. Although War1-42 is expressed and properly localized in the nucleus, the War1-42p variant fails to bind the weak-acid-response elements in the PDR12 promoter, as shown by in vivo footprinting. Importantly, in contrast with wild-type War1p, War1-42p is also no longer phosphorylated upon weak-acid challenge, demonstrating that phosphorylation of War1p, its activation and DNA binding are tightly linked processes that are essential for adaptation to weak-acid stress.

War1p, a novel transcription factor controlling weak acid stress response in yeast.

The Saccharomyces cerevisiae ATP-binding cassette (ABC) transporter Pdr12p effluxes weak acids such as sorbate and benzoate, thus mediating stress adaptation. In this study, we identify a novel transcription factor, War1p, as the regulator of this stress adaptation through transcriptional induction of PDR12. Cells lacking War1p are weak acid hypersensitive, since they fail to induce Pdr12p. The nuclear Zn2Cys6 transcriptional regulator War1p forms homodimers and is rapidly phosphorylated upon sorbate stress. The appearance of phosphorylated War1p isoforms coincides with transcriptional activation of PDR12. Promoter deletion analysis identified a novel cis-acting weak acid response element (WARE) in the PDR12 promoter required for PDR12 induction. War1p recognizes and decorates the WARE both in vitro and in vivo, as demonstrated by band shift assays and in vivo footprinting. Importantly, War1p occupies the WARE in the presence and absence of stress, demonstrating constitutive DNA binding in vivo. Our results suggest that weak acid stress triggers phosphorylation and perhaps activation of War1p. In turn, War1p activation is necessary for the induction of PDR12 through a novel signal transduction event that elicits weak organic acid stress adaptation.

Neural cell adhesion molecule regulates the cellular response to fibroblast growth factor.

Neural cell adhesion molecule (NCAM) mediates cell-cell adhesion and signaling in the nervous system, yet NCAM is also expressed in non-neural tissues, in which its function has in most parts remained elusive. We have previously reported that NCAM stimulates cell-matrix adhesion and neurite outgrowth by activating fibroblast growth factor receptor (FGFR) signaling. Here, we investigated whether the interplay between NCAM and FGFR has any impact on the response of FGFR to its classical ligands, FGFs. To this end, we employed two fibroblast cell lines, NCAM-negative L cells and NCAM-positive NIH-3T3 cells, in which the expression of NCAM was manipulated by means of transfection or RNAi technologies, respectively. The results demonstrate that NCAM expression reduces FGF-stimulated ERK1/2 activation, cell proliferation and cell-matrix adhesion, in both L and NIH-3T3 cells. Furthermore, our data show that NCAM inhibits the binding of FGF to its high-affinity receptor in a competitive manner, providing the mechanisms for the NCAM-mediated suppression of FGF function. In this context, a small peptide that mimics the binding of NCAM to FGFR was sufficient to block FGF-dependent cell proliferation. These findings point to NCAM as being a major regulator of FGF-FGFR interaction, thus introducing a novel type of control mechanism for FGFR activity and opening new therapeutic perspectives for those diseases characterized by aberrant FGFR function.

Increased tumor cell dissemination and cellular senescence in the absence of beta1-integrin function.

Integrins are transmembrane receptors that bind extracellular matrix proteins and enable cell adhesion and cytoskeletal organization, as well as transduction of signals into cells, to promote various aspects of cellular behavior, such as proliferation or survival. Integrins participate in many aspects of tumor biology. Here, we have employed the Rip1Tag2 transgenic mouse model of pancreatic beta cell carcinogenesis to investigate the role of beta(1)-integrin in tumor progression. Specific ablation of beta(1)-integrin function in pancreatic beta cells resulted in a defect in sorting between insulin-expressing beta cells and glucagon-expressing alpha cells in islets of Langerhans. Ablation of beta(1)-integrin in beta tumor cells of Rip1Tag2 mice led to the dissemination of tumor cell emboli into lymphatic blood vessels in the absence of ongoing lymphangiogenesis. Yet, disseminating beta(1)-integrin-deficient beta tumor cells did not elicit metastasis. Rather, primary tumor growth was significantly impaired by reduced tumor cell proliferation and the acquisition of cellular senescence by beta(1)-integrin-deficient beta tumor cells. The results indicate a critical role of beta(1)-integrin function in mediating metastatic dissemination and preventing tumor cell senescence.