Crosstalk between NF-kappaB and beta-catenin pathways in bacterial-colonized intestinal epithelial cells

Salmonella-epithelial cell interactions are known to activate the proinflammatory NF-kappaB signaling pathway and have recently been found to also influence the beta-catenin signaling pathway, an important regulator of epithelial cell proliferation and differentiation. Here, using polarized epithelial cell models, we demonstrate that these same bacteria-mediated effects also direct the molecular crosstalk between the NF-kappaB and beta-catenin signaling pathways. Convergence of these two pathways is a result of the direct interaction between the NF-kappaB p50 subunit and beta-catenin. We show that PhoP(c), the avirulent derivative of a wild-type Salmonella strain, attenuates NF-kappaB activity by stabilizing the association of beta-catenin with NF-kappaB. In cell lines expressing constitutively active beta-catenin, IkappaBalpha protein was indirectly stabilized and NF-kappaB activity was repressed after wild-type Salmonella colonization. Accordingly, constitutively active beta-catenin was found to inhibit the secretion of IL-8. Thus our findings strongly suggest that the crosstalk between the beta-catenin and NF-kappaB signaling pathways is an important regulator of intestinal inflammation.     

Wnt/beta-catenin signaling inhibits death receptor-mediated apoptosis and promotes invasive growth of HNSCC

The Wnt/beta-catenin signaling pathway plays a critical role in cell proliferation and oncogenesis. It has been found to be chronically activated in a variety of human cancers, including head and neck squamous cell carcinoma (HNSCC). Previously, we have found that the activation of the Wnt/beta-catenin signaling pathway inhibits mitochondria-mediated apoptosis. In this study, we extended our studies to determine whether the Wnt/beta-catenin signaling pathway inhibited death receptor-mediated apoptosis in HNSCC cells. We found that Wnt/beta-catenin inhibited not only tumor necrosis factor (TNF)/c-Myc-mediated apoptosis, but also cell detachment-mediated apoptosis (anoikis) which is dependent on the death receptor signaling pathway. Interestingly, we also observed that the Wnt/beta-catenin signaling pathway induced HNSCC cell scattering and promoted cell invasion in the Matrigel, both of which are hallmarks for the invasive growth of HNSCC. Consistently, the over-expression of beta-catenin promoted HNSCC tumor growth in nude mice. Taken together, our results suggest that the Wnt/beta-catenin signaling pathway plays dual functions in HNSCC development: promoting both cell survival and invasive growth of HNSCC cells.     

The stabilization of beta-catenin leads to impaired primordial germ cell development via aberrant cell cycle progression

Primordial germ cells (PGCs) are germ cell precursors that are committed to sperm or oocytes. Dramatic proliferation during PGC development determines the number of founder spermatogonia and oocytes. Although specified to a germ lineage, PGCs produce pluripotent embryonic germ (EG) cells in vitro and testicular teratomas in vivo. Wnt/beta-catenin signaling regulates pluripotency and differentiation in various stem cell systems, and dysregulation of this signaling causes various human cancers. Here, we examined the role of Wnt/beta-catenin signaling in PGC development. In normal PGC development, Wnt/beta-catenin signaling is suppressed by the GSK3beta-mediated active degradation of beta-catenin and the low expression of canonical Wnt molecules. The effects of aberrant activation of Wnt/beta-catenin signaling in PGCs were analyzed using mice carrying a deletion of the exon that encodes the GSK3beta phosphorylation sites in the beta-catenin locus. Despite the potential activity of Wnt/beta-catenin signaling in stem cell maintenance and carcinogenesis in various cell lineages, teratomas were not induced in the mice expressing the nuclear-localized beta-catenin in PGCs. Instead, the mutant mice showed germ cell deficiency caused by the delayed cell cycle progression of the proliferative phase PGCs. Our results show that the suppression of Wnt/beta-catenin signaling is a prerequisite for the normal development of PGCs.     

Distinct Wnt signaling pathways have opposing roles in appendage regeneration

In contrast to mammals, lower vertebrates have a remarkable capacity to regenerate complex structures damaged by injury or disease. This process, termed epimorphic regeneration, involves progenitor cells created through the reprogramming of differentiated cells or through the activation of resident stem cells. Wnt/beta-catenin signaling regulates progenitor cell fate and proliferation during embryonic development and stem cell function in adults, but its functional involvement in epimorphic regeneration has not been addressed. Using transgenic fish lines, we show that Wnt/beta-catenin signaling is activated in the regenerating zebrafish tail fin and is required for formation and subsequent proliferation of the progenitor cells of the blastema. Wnt/beta-catenin signaling appears to act upstream of FGF signaling, which has recently been found to be essential for fin regeneration. Intriguingly, increased Wnt/beta-catenin signaling is sufficient to augment regeneration, as tail fins regenerate faster in fish heterozygous for a loss-of-function mutation in axin1, a negative regulator of the pathway. Likewise, activation of Wnt/beta-catenin signaling by overexpression of wnt8 increases proliferation of progenitor cells in the regenerating fin. By contrast, overexpression of wnt5b (pipetail) reduces expression of Wnt/beta-catenin target genes, impairs proliferation of progenitors and inhibits fin regeneration. Importantly, fin regeneration is accelerated in wnt5b mutant fish. These data suggest that Wnt/beta-catenin signaling promotes regeneration, whereas a distinct pathway activated by wnt5b acts in a negative-feedback loop to limit regeneration.     

Nuclear beta-catenin localization is not sufficient for canonical Wnt signaling activation in human meianoma cell lines

In most cases, advanced stages of melanoma are practically incurable due to high metastatic potential of tumor cells. Multiple observations support the idea that aberrations in Wnt signaling pathway play a significant role in melanoma development and progression. Canonical Wnt signaling activation results in stabilization and accumulation of the major effector molecule called beta-catenin. Mutations promoting beta-catenin stabilization and, thereby, activation of canonical Wnt signaling pathway are frequently found in different cancers, but rarely observed in melanomas. Nevertheless, beta-catenin nuclear and cytoplasmic accumulation is the feature of many human melanoma cell lines and original tumors. That is why, the aim of the investigation was to elucidate the relation between beta-catenin intracellular localization and activity status of Wnt signaling pathway in human melanoma cell lines. Ten human melanoma cell lines were characterized on the basis of the following parameters: canonical Wnt ligand expression, intracellular beta-catenin localization, and activity status of canonical Wnt signaling pathway. Here, it has been demonstrated that nuclear localization of beta-catenin does not always correspond to active status canonical Wnt signaling pathway. Moreover, in the majority of cell lines with nuclear beta-catenin canonical Wnt signaling can't be activated by exogenous expression of an appropriate ligand. Human melanoma cell lines differ in activity of canonical Wnt signaling pathway as well as in mechanisms of its regulation. Therefore, the pathway-targeted potential antineoplastic therapy requires the formation of a "molecular pattern of cancer" for localization of the defect in Wnt signaling cascade in the each case.     

Rac1 activation controls nuclear localization of beta-catenin during canonical Wnt signaling

Canonical Wnt signaling critically regulates cell fate and proliferation in development and disease. Nuclear localization of beta-catenin is indispensable for canonical Wnt signaling; however, the mechanisms governing beta-catenin nuclear localization are not well understood. Here we demonstrate that nuclear accumulation of beta-catenin in response to Wnt requires Rac1 activation. The role of Rac1 depends on phosphorylation of beta-catenin at Ser191 and Ser605, which is mediated by JNK2 kinase. Mutations of these residues significantly affect Wnt-induced beta-catenin nuclear accumulation. Genetic ablation of Rac1 in the mouse embryonic limb bud ectoderm disrupts canonical Wnt signaling and phenocopies deletion of beta-catenin in causing severe truncations of the limb. Finally, Rac1 interacts genetically with beta-catenin and Dkk1 in controlling limb outgrowth. Together these results uncover Rac1 activation and subsequent beta-catenin phosphorylation as a hitherto uncharacterized mechanism controlling canonical Wnt signaling and may provide additional targets for therapeutic intervention of this important pathway.     

The Wnt connection to tumorigenesis

Wnt signaling has been identified as one of the key signaling pathways in cancer, regulating cell growth, motility and differentiation. Because of its widespread activation in diverse human tumor diseases, the Wnt pathway has gained considerable and growing interest in tumor research over recent years. Evidence that altered Wnt signaling is important for human tumor development came from three major findings: (i) the tumor suppressor adenomatous polyposis coli (APC) binds to the Wnt pathway component beta-catenin and is involved in its degradation, (ii) mutations of APC in colon tumors lead to stabilization of the beta-catenin protein and (iii) tumor-associated mutations of beta-catenin in colorectal cancer as well as in other tumor types lead to its stabilisation, qualifying beta-catenin as a proto-oncogene. Here we will describe the biochemical interactions which shape the Wnt pathway and focus on its role in tumorigenesis.     

Ciliary margin transdifferentiation from neural retina is controlled by canonical Wnt signaling

The epithelial layers of the ciliary body (CB) and iris are non-neural structures that differentiate from the anterior region of the eyecup, the ciliary margin (CM). We show here that activation of the canonical Wnt signaling pathway is sufficient and necessary for the normal development of anterior eye structures. Pharmacological activation of beta-catenin signaling with lithium (Li(+)) treatment in retinal explants in vitro induced the ectopic expression of the CM markers Otx1 and Msx1. Cre-mediated stabilization of beta-catenin expression in the peripheral retina in vivo induced a cell autonomous upregulation of CM markers at the expense of neural retina (NR) markers and inhibited neurogenesis. Consistent with a cell autonomous conversion to peripheral eye fates, the proliferation index in the region of the retina that expressed stabilized beta-catenin was identical to the wild-type CM and there was an expansion of CB-like structures at later stages. Conversely, Cre-mediated inactivation of beta-catenin reduced CM marker expression as well as the size of the CM and CB/iris. Aberrant CB development in both mouse models was also associated with a reduction in the number of retinal stem cells in vitro. In summary, activation of canonical Wnt signaling is sufficient to promote the development of peripheral eyecup fates at the expense of the NR and is also required for the normal development of anterior eyecup structures.     

Siah-1 mediates a novel beta-catenin degradation pathway linking p53 to the adenomatous polyposis coli protein

The adenomatous polyposis coli (APC) tumor-suppressor protein, together with Axin and GSK3beta, forms a Wnt-regulated signaling complex that mediates phosphorylation-dependent degradation of beta-catenin by the proteasome. Siah-1, the human homolog of Drosophila seven in absentia, is a p53-inducible mediator of cell cycle arrest, tumor suppression, and apoptosis. We have now found that Siah-1 interacts with the carboxyl terminus of APC and promotes degradation of beta-catenin in mammalian cells. The ability of Siah-1 to downregulate beta-catenin signaling was also demonstrated by hypodorsalization of Xenopus embryos. Unexpectedly, degradation of beta-catenin by Siah-1 was independent of GSK3beta-mediated phosphorylation and did not require the F box protein beta-TrCP. These results indicate that APC and Siah-1 mediate a novel beta-catenin degradation pathway linking p53 activation to cell cycle control.     

Endogenous protein kinase CK2 participates in Wnt signaling in mammary epithelial cells

Protein kinase CK2 (formerly casein kinase II) is a serine/threonine kinase overexpressed in many human tumors, transformed cell lines, and rapidly proliferating tissues. Recent data have shown that many cancers involve inappropriate reactivation of Wnt signaling through ectopic expression of Wnts themselves, as has been seen in a number of human breast cancers, or through mutation of intermediates in the Wnt pathway, such as adenomatous polyposis coli or beta-catenin, as described in colon and other cancers. Wnts are secreted factors that are important in embryonic development, but overexpression of certain Wnts, such as Wnt-1, leads to proliferation and transformation of cells. We report that upon stable transfection of Wnt-1 into the mouse mammary epithelial cell line C57MG, morphological changes and increased proliferation are accompanied by increased levels of CK2, as well as of beta-catenin. CK2 and beta-catenin co-precipitate with the Dvl proteins, which are Wnt signaling intermediates. A major phosphoprotein of the size of beta-catenin appears in in vitro kinase reactions performed on the Dvl immunoprecipitates. In vitro translated beta-catenin, Dvl-2, and Dvl-3 are phosphorylated by CK2. The selective CK2 inhibitor apigenin blocks proliferation of Wnt-1-transfected cells, abrogates phosphorylation of beta-catenin, and reduces beta-catenin and Dvl protein levels. These results demonstrate that endogenous CK2 is a positive regulator of Wnt signaling and growth of mammary epithelial cells.     

Interactions Between the Canonical WNT/Beta-Catenin Pathway and PPAR Gamma on Neuroinflammation,Demyelination, and Remyelination in Multiple Sclerosis

Multiple sclerosis (MS) is marked by neuroinflammation and demyelination with loss of oligodendrocytes in the central nervous system. The immune response is regulated by WNT/beta-catenin pathway in MS. Activated NF-kappaB, a major effector of neuroinflammation, and upregulated canonical WNT/beta-catenin pathway positively regulate each other. Demyelinating events present an upregulation of WNT/beta-catenin pathway, whereas proper myelinating phases show a downregulation of WNT/beta-catenin pathway essential for the promotion of oligodendrocytes precursors cells proliferation and differentiation. The activation of WNT/beta-catenin pathway results in differentiation failure and impairment in remyelination. However, PI3K/Akt pathway and TCF7L2, two downstream targets of WNT/beta-catenin pathway, are upregulated and promote proper remyelination. The interactions of these signaling pathways remain unclear. PPAR gamma activation can inhibit NF-kappaB, and can also downregulate the WNT/beta-catenin pathway. PPAR gamma and canonical WNT/beta-catenin pathway act in an opposite manner. PPAR gamma agonists appear as a promising treatment for the inhibition of demyelination and the promotion of proper remyelination through the control of both NF-kappaB activity and canonical WNT/beta-catenin pathway.