Members of the Tcf family of HMG box-containing transcriptional regulators mediate Wnt signalling in the nucleus. Current models suggest that in the absence of Wnt signalling, Tcf interacts with the repressor protein Groucho and suppresses the expression of Wnt targets. Wnt signalling leads to increases in the level of cytoplasmic beta catenin, which enters the nucleus, displaces Tcf from Groucho and leads to transcriptional activation. In order to test this model we have studied the effects of Drosophila Tcf (dTcf) on signalling by Wingless, a Drosophila member of the Wnt family. We show that overexpression of wild-type dTcf during the development and patterning of the wing antagonises Wingless signalling. Furthermore, increases in the concentration of Armadillo, the Drosophila homologue of beta catenin, do not appear to be sufficient to trigger the change from antagonism to activation. This leads us to suggest that the inactivation of the repressive activity of dTcf requires the activity of Wingless in a manner that is independent of Armadillo. We observe that a Groucho molecule devoid of the WD40 repeats can interact with dTcf and acts as a dominant repressor of Wingless signalling in vivo and in vitro. Coexpression of this molecule with dTcf however, does not lead to enhancement of the repressive effects of dTcf alone. This observation suggests that repression by dTcf might not simply be mediated by an interaction with Groucho but that dTcf may have an intrinsic repressive activity that has to be antagonised by Wingless signalling.
"Genetic or chemical inhibition of GSK3 results in an increase in the cytosolic levels of β-catenin, which can enter the nucleus and, through an interaction with members of the Tcf family of transcription factors, can promote gene expression. However, the levels of β-catenin might not be diagnostic of its transcriptional activity (Fagotto et al., 1997; Guger and Gumbiner, 2000; Lawrence et al., 2000; Staal et al., 2002; Tolwinski et al., 2003; Fodde and Brabletz, 2007; Hendriksen et al., 2008) and this has led to the suggestion that there is a transcriptionally competent form of β-catenin that represents a small fraction of the total pool (Staal et al., 2002; Maher et al., 2010; Muñoz Descalzo et al., 2011). "
[Show abstract][Hide abstract] ABSTRACT: The maintenance of pluripotency in mouse embryonic stem cells (mESCs) relies on the activity of a transcriptional network that is fuelled by the activity of three transcription factors (Nanog, Oct4 and Sox2) and balanced by the repressive activity of Tcf3. Extracellular signals modulate the activity of the network and regulate the differentiation capacity of the cells. Wnt/β-catenin signaling has emerged as a significant potentiator of pluripotency: increases in the levels of β-catenin regulate the activity of Oct4 and Nanog, and enhance pluripotency. A recent report shows that β-catenin achieves some of these effects by modulating the activity of Tcf3, and that this effect does not require its transcriptional activation domain. Here, we show that during self-renewal there is negligible transcriptional activity of β-catenin and that this is due to its tight association with membranes, where we find it in a complex with Oct4 and E-cadherin. Differentiation triggers a burst of Wnt/β-catenin transcriptional activity that coincides with the disassembly of the complex. Our results establish that β-catenin, but not its transcriptional activity, is central to pluripotency acting through a β-catenin/Oct4 complex.
Development 03/2013; 140(6):1171-83. DOI:10.1242/dev.085654 · 6.46 Impact Factor
"Inactivation of the Axin/APC based scaffold through the adaptor protein Dishevelled results in a rise of cytosolic levels of ß-catenin and its entry into the nucleus where it modulates transcription through an interaction with members of the Tcf family of DNA binding proteins  . Although the mantra of Wnt signalling links directly the levels of cytosolic ß-catenin to its transcriptional function, there is abundant evidence that this association is weak       . These studies indicate that there are at least two pools of ß-catenin, a cytosolic and a transcriptionally competent, which are represented by different phosphoisoforms and might have different subcellular locations; the transcriptionally competent pool  represents less than 1% of the total ß-catenin . "
[Show abstract][Hide abstract] ABSTRACT: During development, the emergence of different cell fates and their patterning into tissues and organs requires spatio-temporal coordination that controls the relative number of different cell types. Genetic analyses in different systems have revealed that interactions between Wnt and Notch signalling play pervasive roles in these processes. While many of these interactions can be explained in terms of transcriptional cross-talk between the effectors of these pathways, some of them require a different explanation. Experiments in Drosophila, Xenopus and mouse have revealed that Notch plays an important role in the modulation of the transcriptional activity of β-catenin (the main effector of Wnt signalling pathway, independently of its well characterized function as a membrane tethered transcription factor. These studies suggest that rather than two separate pathways, elements of Wnt and Notch signalling configure a single functional module, Wntch, that plays a key role in the resolution of cell fate decisions. Here we review the evidence for Wntch and present a current circuit view of the system, its control and its role in development with a special focus on stem cell populations.
Seminars in Cell and Developmental Biology 02/2012; 23(4):443-9. DOI:10.1016/j.semcdb.2012.01.012 · 6.27 Impact Factor
"It has been broadly assumed that the key component of Wnt signalling is the stabilisation and subsequent rise in concentration of a cytoplasmic pool of ß-catenin (reviewed in ). However, there is evidence that the concentration of β-catenin alone is not a determinant of Wnt signalling , –, and it is becoming increasingly clear that other factors, like nuclear shuttling and cytoplasmic tethering, affect the nuclear availability of ß-catenin, and thus its ability to interact with the transcriptional machinery , . It is possible that the association of ß-catenin with E-Cadherin also influences its activity under steady state conditions, not only in pathological or overexpression situations. "
[Show abstract][Hide abstract] ABSTRACT: Armadillo, the Drosophila orthologue of vertebrate ss-catenin, plays a dual role as the key effector of Wingless/Wnt1 signalling, and as a bridge between E-Cadherin and the actin cytoskeleton. In the absence of ligand, Armadillo is phosphorylated and targeted to the proteasome. Upon binding of Wg to its receptors, the "degradation complex" is inhibited; Armadillo is stabilised and enters the nucleus to transcribe targets.
Although the relationship between signalling and adhesion has been extensively studied, few in vivo data exist concerning how the "transcriptional" and "adhesive" pools of Armadillo are regulated to orchestrate development. We have therefore addressed how the subcellular distribution of Armadillo and its association with E-Cadherin change in larval wing imaginal discs, under wild type conditions and upon signalling. Using confocal microscopy, we show that Armadillo and E-Cadherin are spatio-temporally regulated during development, and that a punctate species becomes concentrated in a subapical compartment in response to Wingless. In order to further dissect this phenomenon, we overexpressed Armadillo mutants exhibiting different levels of activity and stability, but retaining E-Cadherin binding. Arm(S10) displaces endogenous Armadillo from the AJ and the basolateral membrane, while leaving E-Cadherin relatively undisturbed. Surprisingly, DeltaNArm(1-155) caused displacement of both Armadillo and E-Cadherin, results supported by our novel method of quantification. However, only membrane-targeted Myr-DeltaNArm(1-155) produced comparable nuclear accumulation of Armadillo and signalling to Arm(S10). These experiments also highlighted a row of cells at the A/P boundary depleted of E-Cadherin at the AJ, but containing actin.
Taken together, our results provide in vivo evidence for a complex non-linear relationship between Armadillo levels, subcellular distribution and Wingless signalling. Moreover, this study highlights the importance of Armadillo in regulating the subcellular distribution of E-Cadherin.
PLoS ONE 02/2008; 3(8):e2893. DOI:10.1371/journal.pone.0002893 · 3.23 Impact Factor
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