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Wingless signalling alters the levels, subcellular distribution and dynamics of Armadillo and E-cadherin in third instar larval wing imaginal discs.

Somorjai IM, Martinez-Arias A - PLoS ONE (2008)

Bottom Line: Surprisingly, DeltaNArm(1-155) caused displacement of both Armadillo and E-Cadherin, results supported by our novel method of quantification.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of Cambridge, Cambridge, United Kingdom. ildiko.somorjai@obs-banyuls.fr

ABSTRACT

Background: 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.

Methodology/principal findings: 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.

Conclusions/significance: 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.

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Related in: MedlinePlus

ArmS10 induced signalling correlates with clearing of endogenous Armadillo from the AJ and dynamic regulation of subcellular distribution subapically.(A) Similarly to Wingless ligand-dependent signalling, ArmS10 does not affect the distribution of E-Cadherin-GFP (E-CadGFP) in the AJ. (B) In contrast, ArmS10 entirely displaces endogenous Armadillo from the AJ, which can often be identified as puncta immediately subapically (red channel, inset). (C) E-Cadherin-GFP (E-CadGFP) accumulates in puncta upon ArmS10 overpexression similarly to ligand-dependent signalling. (D) In contrast, ArmS10 causes an accumulation of endogenous Armadillo to high levels in the centre of the dppGAL4 overexpression domain, but results in complete loss from the cytoplasm and basolateral membrane at the edges (red arrows).
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pone-0002893-g007: ArmS10 induced signalling correlates with clearing of endogenous Armadillo from the AJ and dynamic regulation of subcellular distribution subapically.(A) Similarly to Wingless ligand-dependent signalling, ArmS10 does not affect the distribution of E-Cadherin-GFP (E-CadGFP) in the AJ. (B) In contrast, ArmS10 entirely displaces endogenous Armadillo from the AJ, which can often be identified as puncta immediately subapically (red channel, inset). (C) E-Cadherin-GFP (E-CadGFP) accumulates in puncta upon ArmS10 overpexression similarly to ligand-dependent signalling. (D) In contrast, ArmS10 causes an accumulation of endogenous Armadillo to high levels in the centre of the dppGAL4 overexpression domain, but results in complete loss from the cytoplasm and basolateral membrane at the edges (red arrows).

Mentions: As with Wingless signalling, little change was seen in apicobasal polarity markers at the adherens and septate junctions, with E-Cadherin expression at 0% identical in ArmS10 and Wg overexpression experiments (compare Figures 6A and 7A; and data not shown). In contrast, ArmS10 was found to be more stable than endogenous Armadillo in the AJs (Figure 7B), the latter being completely excluded except for a punctate distribution immediately basal to the AJ (Figure 7B, red channel, inset).


Wingless signalling alters the levels, subcellular distribution and dynamics of Armadillo and E-cadherin in third instar larval wing imaginal discs.

Somorjai IM, Martinez-Arias A - PLoS ONE (2008)

ArmS10 induced signalling correlates with clearing of endogenous Armadillo from the AJ and dynamic regulation of subcellular distribution subapically.(A) Similarly to Wingless ligand-dependent signalling, ArmS10 does not affect the distribution of E-Cadherin-GFP (E-CadGFP) in the AJ. (B) In contrast, ArmS10 entirely displaces endogenous Armadillo from the AJ, which can often be identified as puncta immediately subapically (red channel, inset). (C) E-Cadherin-GFP (E-CadGFP) accumulates in puncta upon ArmS10 overpexression similarly to ligand-dependent signalling. (D) In contrast, ArmS10 causes an accumulation of endogenous Armadillo to high levels in the centre of the dppGAL4 overexpression domain, but results in complete loss from the cytoplasm and basolateral membrane at the edges (red arrows).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2483348&req=5

pone-0002893-g007: ArmS10 induced signalling correlates with clearing of endogenous Armadillo from the AJ and dynamic regulation of subcellular distribution subapically.(A) Similarly to Wingless ligand-dependent signalling, ArmS10 does not affect the distribution of E-Cadherin-GFP (E-CadGFP) in the AJ. (B) In contrast, ArmS10 entirely displaces endogenous Armadillo from the AJ, which can often be identified as puncta immediately subapically (red channel, inset). (C) E-Cadherin-GFP (E-CadGFP) accumulates in puncta upon ArmS10 overpexression similarly to ligand-dependent signalling. (D) In contrast, ArmS10 causes an accumulation of endogenous Armadillo to high levels in the centre of the dppGAL4 overexpression domain, but results in complete loss from the cytoplasm and basolateral membrane at the edges (red arrows).
Mentions: As with Wingless signalling, little change was seen in apicobasal polarity markers at the adherens and septate junctions, with E-Cadherin expression at 0% identical in ArmS10 and Wg overexpression experiments (compare Figures 6A and 7A; and data not shown). In contrast, ArmS10 was found to be more stable than endogenous Armadillo in the AJs (Figure 7B), the latter being completely excluded except for a punctate distribution immediately basal to the AJ (Figure 7B, red channel, inset).

Bottom Line: Surprisingly, DeltaNArm(1-155) caused displacement of both Armadillo and E-Cadherin, results supported by our novel method of quantification.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of Cambridge, Cambridge, United Kingdom. ildiko.somorjai@obs-banyuls.fr

ABSTRACT

Background: 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.

Methodology/principal findings: 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.

Conclusions/significance: 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.

Show MeSH
Related in: MedlinePlus