Wnt Meeting 1996

A meeting on Wnt genes was held at Stanford University, August 9 and 10, organized by Harold Varmus (NIH) and Roel Nusse (HHMI-Stanford). This meeting was the first formal version of a series of informal Wnt gatherings held over the past years at various locations. The previous meetings were always inspired by the importance of Wnt genes in animal development and tumorigenesis, but also frustrated by the lack of understanding of the molecular aspects of signaling by these secreted proteins. Much to the delight of the attendants, the 1996 meeting brought together a surprising number of novel findings on various aspects of Wnt signaling. Only half a year ago, very little was known about the Wnt signal transduction pathway, but recently, specific receptors and various intracellular components have been discovered and many of these novel findings were reported at the meeting. It seems that the power of Drosophila and C. elegans genetics, combined with in vitro assays for Wnt signaling and rapid biological assays in Xenopus will lead to rapid progress in understanding how Wnt molecules bring about their interesting biological effects.     

Wnt信号通路及其与疾病的关系

Wnt信号通路是参与胚胎及器官发育的主要4大类信号传导途径之一,对胚胎及器官发育起着不可替代的作用.对其在疾病产生中的作用做了简要的概述,以期在发育生物学、生物医学及交叉学科的研究上带来全新的革命.     

Interaction of dishevelled and Xenopus axin-related protein is required for wnt signal transduction

Signaling by the Wnt family of secreted proteins plays an important role in animal development and is often misregulated in carcinogenesis. Wnt signal transduction is controlled by the rate of degradation of beta-catenin by a complex of proteins including glycogen synthase kinase 3 (GSK3), adenomatous polyposis coli, and Axin. Dishevelled is required for Wnt signal transduction, and its activation results in stabilization of beta-catenin. However, the biochemical events underlying this process remain largely unclear. Here we show that Xenopus Dishevelled (Xdsh) interacts with a Xenopus Axin-related protein (XARP). This interaction depends on the presence of the Dishevelled-Axin (DIX) domains in both XARP and Xdsh. Moreover, the same domains are essential for signal transduction through Xdsh. Finally, our data point to a possible mechanism for signal transduction, in which Xdsh prevents beta-catenin degradation by displacing GSK3 from its complex with XARP.     

A WNTer wonderland in Snowbird

The Keystone Symposium on ;Wnt and beta-catenin signaling in development and disease' was held recently in Snowbird, UT, USA. Organized by Mariann Bienz and Hans Clevers, this meeting covered a wide range of topics, including Wnt protein biogenesis, Wnt receptors and signaling pathways, beta-catenin/Tcf complexes and gene expression, Wnt signaling in development, cancer, stem cell biology and regeneration, and therapeutics that target the Wnt/beta-catenin pathway.     

Wnt5a can both activate and repress Wnt/β-catenin signaling during mouse embryonic development

Embryonic development is controlled by a small set of signal transduction pathways, with vastly different phenotypic outcomes depending on the time and place of their recruitment. How the same molecular machinery can elicit such specific and distinct responses, remains one of the outstanding questions in developmental biology. Part of the answer may lie in the high inherent genetic complexity of these signaling cascades, as observed for the Wnt-pathway. The mammalian genome encodes multiple Wnt proteins and receptors, each of which show dynamic and tightly controlled expression patterns in the embryo. Yet how these components interact in the context of the whole organism remains unknown. Here we report the generation of a novel, inducible transgenic mouse model that allows spatiotemporal control over the expression of Wnt5a, a protein implicated in many developmental processes and multiple Wnt-signaling responses. We show that ectopic Wnt5a expression from E10.5 onwards results in a variety of developmental defects, including loss of hair follicles and reduced bone formation in the skull. Moreover, we find that Wnt5a can have dual signaling activities during mouse embryonic development. Specifically, Wnt5a is capable of both inducing and repressing β-catenin/TCF signaling in vivo, depending on the time and site of expression and the receptors expressed by receiving cells. These experiments show for the first time that a single mammalian Wnt protein can have multiple signaling activities in vivo, thereby furthering our understanding of how signaling specificity is achieved in a complex developmental context.     

Head inducer Dickkopf-1 is a ligand for Wnt coreceptor LRP6.

BACKGROUND: Dickkopf-1 (Dkk-1) is a head inducer secreted from the vertebrate head organizer and induces anterior development by antagonizing Wnt signaling. Although several families of secreted antagonists have been shown to inhibit Wnt signal transduction by binding to Wnt, the molecular mechanism of Dkk-1 action is unknown. The Wnt family of secreted growth factors initiates signaling via the Frizzled (Fz) receptor and its candidate coreceptor, LDL receptor-related protein 6 (LRP6), presumably through Fz-LRP6 complex formation induced by Wnt. The significance of the Fz-LRP6 complex in signal transduction remains to be established. RESULTS: We report that Dkk-1 is a high-affinity ligand for LRP6 and inhibits Wnt signaling by preventing Fz-LRP6 complex formation induced by Wnt. Dkk-1 binds neither Wnt nor Fz, nor does it affect Wnt-Fz interaction. Dkk-1 function in head induction and Wnt signaling inhibition strictly correlates with its ability to bind LRP6 and to disrupt the Fz-LRP6 association. LRP6 function and Dkk-1 inhibition appear to be specific for the Wnt/Fz beta-catenin pathway. CONCLUSIONS: Our results demonstrate that Dkk-1 is an LRP6 ligand and inhibits Wnt signaling by blocking Wnt-induced Fz-LRP6 complex formation. Our findings thus reveal a novel mechanism for Wnt signal modulation. LRP6 is a Wnt coreceptor that appears to specify Wnt/Fz signaling to the beta-catenin pathway, and Dkk-1, distinct from Wnt binding antagonists, may be a specific inhibitor for Wnt/beta-catenin signaling. Our findings suggest that Wnt-Fz-LRP6 complex formation, but not Wnt-Fz interaction, triggers Wnt/beta-catenin signaling.     

A positive role for the PP2A catalytic subunit in Wnt signal transduction

Protein phosphatase-2A (PP2A) is a multisubunit serine/threonine phosphatase involved in intracellular signaling, gene regulation, and cell cycle progression. Different subunits of PP2A bind to Axin and Adenomatous Polyposis Coli, components of the Wnt signal transduction pathway. Using early Xenopus embryos, we studied how PP2A functions in Wnt signal transduction. The catalytic subunit of PP2A (PP2A-C) potentiated secondary axis induction and Siamois reporter gene activation by Dishevelled, a component of the Wnt pathway, indicating a positive regulatory role of this enzyme in Wnt signaling. In contrast, small t antigen, an antagonist of PP2A-C, inhibited Dishevelled-mediated signal transduction, as did the regulatory PP2A-B'epsilon subunit, consistent with the requirement of PP2A function in this pathway. Although Wnt signaling is thought to occur via regulation of beta-catenin degradation, PP2A-C did not significantly affect beta-catenin stability. Moreover, the pathway activated by a stabilized form of beta-catenin was sensitive to PP2A-C and its inhibitors, suggesting that PP2A-C acts downstream of beta-catenin. Because previous work has suggested that PP2A can act upstream of beta-catenin, we propose that PP2A regulates the Wnt pathway at multiple levels.     

Multi-lineage differentiation of mesenchymal stem cells – To Wnt,or not Wnt

Mesenchymal stem cells (MSCs) are multipotent precursor cells originating from several adult connective tissues. MSCs possess the ability to self-renew and differentiate into several lineages, and are recognized by the expression of unique cell surface markers. Several lines of evidence suggest that various signal transduction pathways and their interplay regulate MSC differentiation. To that end, a critical player in regulating MSC differentiation is a group of proteins encoded by the Wnt gene family, which was previously known for influencing various stages of embryonic development and cell fate determination. As MSCs have gained significant clinical attention for their potential applications in regenerative medicine, it is imperative to unravel the mechanisms by which molecular regulators control differentiation of MSCs for designing cell-based therapeutics. It is rather coincidental that the functional outcome(s) of Wnt-induced signals share similarities with cellular redox-mediated networks from the standpoint of MSC biology. Furthermore, there is evidence for a crosstalk between Wnt and redox signalling, which begs the question whether Wnt-mediated differentiation signals involve the intermediary role of reactive oxygen species. In this review, we summarize the impact of Wnt signalling on multi-lineage differentiation of MSCs, and attempt to unravel the intricate interplay between Wnt and redox signals.     

Meeting report: Signal transduction meets systems biology

ABSTRACT: In the 21st century, systems-wide analyses of biological processes are getting more and more realistic. Especially for the in depth analysis of signal transduction pathways and networks, various approaches of systems biology are now successfully used. The EU FP7 large integrated project SYBILLA (Systems Biology of T-cell Activation in Health and Disease) coordinates such an endeavor. By using a combination of experimental data sets and computational modelling, the consortium strives for gaining a detailed and mechanistic understanding of signal transduction processes that govern T-cell activation. In order to foster the interaction between systems biologists and experimentally working groups, SYBILLA co-organized the 15th meeting "Signal Transduction: Receptors, Mediators and Genes" together with the Signal Transduction Society (STS). Thus, the annual STS conference, held from November 7 to 9, 2011 in Weimar, Germany, provided an interdisciplinary forum for research on signal transduction with a major focus on systems biology addressing signalling events in T-cells. Here we report on a selection of ongoing projects of SYBILLA and how they were discussed at this interdisciplinary conference.     

Wnt/beta-catenin signaling in development and disease

A remarkable interdisciplinary effort has unraveled the WNT (Wingless and INT-1) signal transduction cascade over the last two decades. Wnt genes encode small secreted proteins that are found in all animal genomes. Wnt signaling is involved in virtually every aspect of embryonic development and also controls homeostatic self-renewal in a number of adult tissues. Germline mutations in the Wnt pathway cause several hereditary diseases, and somatic mutations are associated with cancer of the intestine and a variety of other tissues.     

Wnt activates the Tak1/Nemo-like kinase pathway

Genetic studies on endoderm-mesoderm specification in Caenorhabditis elegans have demonstrated a role for several Wnt cascade components as well as for a MAPK-like pathway in this process. The latter pathway includes the MAPK kinase kinase-like MOM-4/Tak1, its adaptor TAP-1/Tab1, and the MAPK-like LIT-1/Nemo-like kinase. A model has been proposed in which the Tak1 kinase cascade counteracts the Wnt cascade at the level of beta-catenin/TCF phosphorylation. In this model, the signal that activates the Tak1 kinase cascade is unknown. As an alternative explanation of these genetic data, we have explored whether Tak1 is directly activated by Wnt. We find that Wnt1 stimulation results in autophosphorylation and activation of MOM-4/Tak1 in a TAP-1/Tab1-dependent fashion. Wnt1-induced Tak1 stimulation activates Nemo-like kinase, resulting in the phosphorylation of TCF. Our results combined with the genetic data from C. elegans imply a mechanism whereby Wnt directly activates the MOM-4/Tak1 kinase signaling pathway. Thus, Wnt signal transduction through the canonical pathway activates beta-catenin/TCF, whereas Wnt signal transduction through the Tak1 pathway phosphorylates and inhibits TCF, which might function as a feedback mechanism.     

Wnt signaling: an embarrassment of receptors

Recent genetic studies in Drosophila and mouse have uncovered a new aspect of the Wnt signal transduction machinery. Mutations disrupting LDL-receptor related proteins produce loss-of-function Wnt phenotypes, suggesting that these cell surface molecules may represent essential co-receptors for Wnt ligands.     

Polyethylenimine-cationized β-catenin protein transduction activates the Wnt canonical signaling pathway more effectively than cationic lipid-based transduction

The Wnt canonical signaling pathway is essential for the early development of eukaryotic organisms and plays a key role in cell proliferation, differentiation, and oncogenesis. Moreover, the Wnt canonical signaling pathway contributes to the self-renewal of mouse hematopoietic stem cells (HSCs). Here, we demonstrate artificial activation of the Wnt canonical signaling pathway by β-catenin protein transduction. Constitutively active β-catenin protein was introduced into human embryonic kidney HEK-293 cells using a polyethylenimine (PEI) cationization method, or with the BioPORTER protein transduction reagent. We have previously shown that modification with PEI effectively causes proteins to be internalized by living mammalian cells. PEI-cationized, constitutively active β-catenin protein was added to HEK-293 cells, and induction of several Wnt/β-catenin target genes was detected by real-time PCR. However, using BioPORTER to introduce active β-catenin did not activate the Wnt canonical signaling pathway. Introduction of eGFPNuc (enhanced green fluorescent protein variant containing a nuclear localization signal) into HEK-293 cells using the BioPORTER reagent caused significant cell death, as determined by propidium iodide staining. In contrast, the PEI-modified eGFPNuc did not impair survival of HEK-293 cells. These results indicate that the Wnt canonical signaling pathway could be successfully activated by transduction of PEI-cationized active β-catenin, and the PEI-cationization method is an effective and safe technology for protein transduction into mammalian cells.     

Wnt signaling: multivesicular bodies hold GSK3 captive

Two key events in Wnt signal transduction, receptor endocytosis and inactivation of Glycogen Synthase Kinase 3 (GSK3), remain incompletely understood. Taelman et?al. (2010) discover that Wnt signaling inactivates GSK3 by sequestering the enzyme in multivesicular bodies, thus linking these two events and providing a new framework for understanding Wnt signaling.     

Knockout mouse models to study Wnt signal transduction

Wnt signal transduction is crucial for maintaining the balance between proliferation and differentiation throughout embryogenesis and postnatal life. Here, we provide a comprehensive overview of the conventional knockout mouse studies of both the canonical and the noncanonical Wnt pathways during mammalian development. Many of these knockout mice display early embryonic lethality, underscoring the fundamental importance of Wnt signal transduction, but precluding functional analyses at later stages. Use of conditional or inducible mouse models will enable us to study the role of Wnt signaling during later stages of development and adult life. Furthermore, genomic-scale approaches and advanced imaging techniques could provide a means to start dissecting the mechanism behind the observed phenotypes.     

WNT secretion and signalling in human disease

Wnt signalling, a key pathway involved in various aspects of embryonic development, also underlies many human diseases, in particular, cancer. Research focused on signal transduction within signal-receiving cells led to the discovery of many Wnt pathway components, but study of the secretion of Wnt ligands themselves was neglected until recently. Attention was drawn to this highly regulated process by the association of aberrant Wnt levels with an increasing number of diseases. Studying the biogenesis and processing of active Wnt ligands will open new avenues for generating therapeutics to specifically target aberrant Wnt signalling. Here we review the proteins required for Wnt secretion and signalling at the plasma membrane, ending with a discussion on potential therapeutic approaches to treat Wnt-induced diseases.