So Young Cha

A Wnt–Axin2–GSK3β cascade regulates Snail1 activity in breast cancer cells

Accumulating evidence indicates that hyperactive Wnt signalling occurs in association with the development and progression of human breast cancer. As a consequence of engaging the canonical Wnt pathway, a β-catenin–T-cell factor (TCF) transcriptional complex is generated, which has been postulated to trigger the epithelial–mesenchymal transition (EMT) that characterizes the tissue-invasive phenotype. However, the molecular mechanisms by which the β-catenin–TCF complex induces EMT-like programmes remain undefined. Here, we demonstrate that canonical Wnt signalling engages tumour cell dedifferentiation and tissue-invasive activity through an Axin2-dependent pathway that stabilizes the Snail1 zinc-transcription factor, a key regulator of normal and neoplastic EMT programmes. Axin2 regulates EMT by acting as a nucleocytoplasmic chaperone for GSK3β, the dominant kinase responsible for controlling Snail1 protein turnover and activity. As dysregulated Wnt signalling marks a diverse array of cancerous tissue types, the identification of a β-catenin–TCF-regulated Axin2–GSK3β–Snail1 axis provides new mechanistic insights into cancer-associated EMT programmes.

p53 and microRNA-34 are suppressors of canonical Wnt signaling.

The tumor suppressor p53 activates miRNA-34 to inhibit Wnt signaling and colorectal cancer cell invasiveness. p53 Activates MicroRNA-34 to Inhibit Wnt Signaling The tumor suppressor p53 is missing or nonfunctional in many cancers, whereas the canonical Wnt signaling pathway is frequently activated. Here, Kim et al. show that p53 restrained Wnt signaling during Xenopus development, whereas loss of p53 function led to aberrant activation of the canonical Wnt signaling pathway, with microRNA-34 (miR-34) providing the connection between the two. They found that p53 stimulated production of miR-34, which, in turn, targeted key genes in the Wnt signaling pathway. Analyses of gene expression data sets indicated that loss of p53 or miR-34 function was associated with activation of Wnt signaling in human cancers; moreover, loss of p53 function increased Wnt signaling in colon cancer cells in vitro. In p53-mutant colon cancer cells, miR-34 attenuated Wnt signaling and decreased the invasiveness of these cells in vitro. Thus, the p53–miR-34–Wnt pathway appears to be crucial not only during development but also for p53’s tumor suppressor function. Although loss of p53 function and activation of canonical Wnt signaling cascades are frequently coupled in cancer, the links between these two pathways remain unclear. We report that p53 transactivated microRNA-34 (miR-34), which consequently suppressed the transcriptional activity of β-catenin–T cell factor and lymphoid enhancer factor (TCF/LEF) complexes by targeting the untranslated regions (UTRs) of a set of conserved targets in a network of genes encoding elements of the Wnt pathway. Loss of p53 function increased canonical Wnt signaling by alleviating miR-34–specific interactions with target UTRs, and miR-34 depletion relieved p53-mediated Wnt repression. Gene expression signatures reflecting the status of β-catenin–TCF/LEF transcriptional activity in breast cancer and pediatric neuroblastoma patients were correlated with p53 and miR-34 functional status. Loss of p53 or miR-34 contributed to neoplastic progression by triggering the Wnt-dependent, tissue-invasive activity of colorectal cancer cells. Further, during development, miR-34 interactions with the β-catenin UTR affected Xenopus body axis polarity and the expression of Wnt-dependent patterning genes. These data provide insight into the mechanisms by which a p53–miR-34 network restrains canonical Wnt signaling cascades in developing organisms and human cancer.

Snail1 is stabilized by O-GlcNAc modification in hyperglycaemic condition.

Protein O‐phosphorylation often occurs reciprocally with O‐GlcNAc modification and represents a regulatory principle for proteins. O‐phosphorylation of serine by glycogen synthase kinase‐3β on Snail1, a transcriptional repressor of E‐cadherin and a key regulator of the epithelial–mesenchymal transition (EMT) programme, results in its proteasomal degradation. We show that by suppressing O‐phosphorylation‐mediated degradation, O‐GlcNAc at serine112 stabilizes Snail1 and thus increases its repressor function, which in turn attenuates E‐cadherin mRNA expression. Hyperglycaemic condition enhances O‐GlcNAc modification and initiates EMT by transcriptional suppression of E‐cadherin through Snail1. Thus, dynamic reciprocal O‐phosphorylation and O‐GlcNAc modification of Snail1 constitute a molecular link between cellular glucose metabolism and the control of EMT.

p53 regulates nuclear GSK-3 levels through miR-34-mediated Axin2 suppression in colorectal cancer cells

p53 is a bona fide tumor suppressor gene whose loss of function marks the most common genetic alteration in human malignancy. Although the causal link between loss of p53 function and tumorigenesis has been clearly demonstrated, the mechanistic links by which loss of p53 potentiates oncogenic signaling are not fully understood. Recent evidence indicates that the microRNA-34 (miR-34) family, a transcriptional target of the p53, directly suppresses a set of canonical Wnt genes and Snail, resulting in p53-mediated suppression of Wnt signaling and the EMT process. In this study, we report that p53 regulates GSK-3β nuclear localization via miR-34-mediated suppression of Axin2 in colorectal cancer. Exogenous miR-34a decreases Axin2 UTR-reporter activity through multiple binding sites within the 5′ and 3′ UTR of Axin2. Suppression of Axin2 by p53 or miR-34 increases nuclear GSK-3β abundance and leads to decreased Snail expression in colorectal cancer cells. Conversely, expression of the non-coding UTR of Axin2 causes depletion of endogenous miR-34 via the miR-sponge effect together with increased Axin2 function, supporting that the RNA-RNA interactions with Axin2 transcripts act as an endogenous decoy for miR-34. Further, RNA transcripts of miR-34 target were correlated with Axin2 in clinical data set of colorectal cancer patients. Although the biological relevance of nuclear GSK-3 level has not been fully studied, our results demonstrate that the tumor suppressor p53/miR-34 axis plays a role in regulating nuclear GSK-3 levels and Wnt signaling through the non-coding UTR of Axin2 in colorectal cancer.

Helicobacter pylori CagA promotes Snail-mediated epithelial-mesenchymal transition by reducing GSK-3 activity.

Cytotoxin-associated gene A (CagA) is an oncoprotein and a major virulence factor of H. pylori. CagA is delivered into gastric epithelial cells via a type IV secretion system and causes cellular transformation. The loss of epithelial adhesion that accompanies the epithelial-mesenchymal transition (EMT) is a hallmark of gastric cancer. Although CagA is a causal factor in gastric cancer, the link between CagA and the associated EMT has not been elucidated. Here, we show that CagA induces the EMT by stabilizing Snail, a transcriptional repressor of E-cadherin expression. Mechanistically we show that CagA binds GSK-3 in a manner similar to Axin and causes it to shift to an insoluble fraction, resulting in reduced GSK-3 activity. We also find that the level of Snail protein is increased in H. pylori infected epithelium in clinical samples. These results suggest that H. pylori CagA acts as a pathogenic scaffold protein that induces a Snail-mediated EMT via the depletion of GSK-3.

Snail reprograms glucose metabolism by repressing phosphofructokinase PFKP allowing cancer cell survival under metabolic stress

Dynamic regulation of glucose flux between aerobic glycolysis and the pentose phosphate pathway (PPP) during epithelial–mesenchymal transition (EMT) is not well-understood. Here we show that Snail (SNAI1), a key transcriptional repressor of EMT, regulates glucose flux toward PPP, allowing cancer cell survival under metabolic stress. Mechanistically, Snail regulates glycolytic activity via repression of phosphofructokinase, platelet (PFKP), a major isoform of cancer-specific phosphofructokinase-1 (PFK-1), an enzyme involving the first rate-limiting step of glycolysis. The suppression of PFKP switches the glucose flux towards PPP, generating NADPH with increased metabolites of oxidative PPP. Functionally, dynamic regulation of PFKP significantly potentiates cancer cell survival under metabolic stress and increases metastatic capacities in vivo. Further, knockdown of PFKP rescues metabolic reprogramming and cell death induced by loss of Snail. Thus, the Snail-PFKP axis plays an important role in cancer cell survival via regulation of glucose flux between glycolysis and PPP.

Niclosamide is a potential therapeutic for familial adenomatosis polyposis by disrupting Axin-GSK3 interaction

The epithelial-mesenchymal transition (EMT) is implicated in tumorigenesis and cancer progression, and canonical Wnt signaling tightly controls Snail, a key transcriptional repressor of EMT. While the suppression of canonical Wnt signaling and EMT comprises an attractive therapeutic strategy, molecular targets for small molecules reverting Wnt and EMT have not been widely studied. Meanwhile, the anti-helminthic niclosamide has been identified as a potent inhibitor of many oncogenic signaling pathways although its molecular targets have not yet been clearly identified. In this study, we show that niclosamide directly targets Axin-GSK3 interaction, at least in part, resulting in suppression of Wnt/Snail-mediated EMT. In vitro and in vivo, disruption of Axin-GSK3 complex by niclosamide induces mesenchymal to epithelial reversion at nM concentrations, accompanied with suppression of the tumorigenic potential of colon cancer. Niclosamide treatment successfully attenuates Snail abundance while increasing E-cadherin abundance in xenograft tumor. Notably, oral administration of niclosamide significantly suppressed adenoma formation in an APC-MIN mice model, indicating that niclosamide is an effective therapeutic for familial adenomatosis polyposis (FAP) patients. In this study, we identified a novel target to control the canonical Wnt pathway and Snail-mediated EMT program, and discovered a repositioned therapeutics for FAP patients.

Dishevelled has a YAP nuclear export function in a tumor suppressor context-dependent manner

Phosphorylation-dependent YAP translocation is a well-known intracellular mechanism of the Hippo pathway; however, the molecular effectors governing YAP cytoplasmic translocation remains undefined. Recent findings indicate that oncogenic YAP paradoxically suppresses Wnt activity. Here, we show that Wnt scaffolding protein Dishevelled (DVL) is responsible for cytosolic translocation of phosphorylated YAP. Mutational inactivation of the nuclear export signal embedded in DVL leads to nuclear YAP retention, with an increase in TEAD transcriptional activity. DVL is also required for YAP subcellular localization induced by E-cadherin, α-catenin, or AMPK activation. Importantly, the nuclear-cytoplasmic trafficking is dependent on the p53-Lats2 or LKB1-AMPK tumor suppressor axes, which determine YAP phosphorylation status. In vivo and clinical data support that the loss of p53 or LKB1 relieves DVL-linked reciprocal inhibition between the Wnt and nuclear YAP activity. Our observations provide mechanistic insights into controlled proliferation coupled with epithelial polarity during development and human cancer.Hippo and Wnt pathways are important for cancer development, and they can cross talk; however, the mechanisms behind this connection are unknown. Here the authors show that DVL (a scaffold protein in the Wnt pathway) regulates the shuttling of YAP (a key component of the Hippo pathway) between cytoplasm and nucleus in specific tumor suppressor contexts.