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Protection of armadillo/β-Catenin by armless, a novel positive regulator of wingless signaling.

Reim G, Hruzova M, Goetze S, Basler K - PLoS Biol. (2014)

Bottom Line: We found by genetic and biochemical analyses that Als functions downstream of the destruction complex, at the level of the SCF/Slimb/βTRCP E3 Ub ligase.In the absence of Als, Arm levels are severely reduced.We suggest that Als antagonizes Ter94's positive effect on E3 ligase function and propose that Als promotes Wg signaling by rescuing Arm from proteolytic degradation, spotlighting an unexpected step where the Wg pathway signal is modulated.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland.

ABSTRACT
The Wingless (Wg/Wnt) signaling pathway is essential for metazoan development, where it is central to tissue growth and cellular differentiation. Deregulated Wg pathway activation underlies severe developmental abnormalities, as well as carcinogenesis. Armadillo/β-Catenin plays a key role in the Wg transduction cascade; its cytoplasmic and nuclear levels directly determine the output activity of Wg signaling and are thus tightly controlled. In all current models, once Arm is targeted for degradation by the Arm/β-Catenin destruction complex, its fate is viewed as set. We identified a novel Wg/Wnt pathway component, Armless (Als), which is required for Wg target gene expression in a cell-autonomous manner. We found by genetic and biochemical analyses that Als functions downstream of the destruction complex, at the level of the SCF/Slimb/βTRCP E3 Ub ligase. In the absence of Als, Arm levels are severely reduced. We show by biochemical and in vivo studies that Als interacts directly with Ter94, an AAA ATPase known to associate with E3 ligases and to drive protein turnover. We suggest that Als antagonizes Ter94's positive effect on E3 ligase function and propose that Als promotes Wg signaling by rescuing Arm from proteolytic degradation, spotlighting an unexpected step where the Wg pathway signal is modulated.

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Related in: MedlinePlus

als functions downstream of the Arm/β-Catenin destruction complex and upstream of activated Armadillo.Overexpression of the positive components Arrow (A), Dishevelled (L), ArmS10 (F and Q), or Arm (G and R) and depletion of negative components such as wingful (B), shaggy (C, H, and I), apc (D and J), slimb (E, O, and P), and axin (K) cause phenotypes that reflect ectopic Wg signaling: ectopic sensory bristles (A–G), ectopic veins (A–C), and tissue overgrowth (A, H–K, O, and P), but also reduced organ size upon strong pathway activation (B–F, L, M, and Q) and caused ectopic head cuticle (H–J, L, and M). Co-expression of alsRNAi could suppress these phenotypes in the wing (A′–D′) and the eye (H′–M′). Phenotypes based on slimbRNAi and ArmS10 expression could not be suppressed upon als depletion (E′, O′, P′, F′, and Q′). Arm overexpession could not rescue alsRNAi phenotypes (G′ and R′). Heads were photographed from dorsal views (left pictures of H–K′) or lateral views, anterior to the left (right pictures of H–K′, L–R′). alsRNAi lines: oligo310UAS (A′–E′) and for the eye analysis; oligo2_5′utr (G′).
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pbio-1001988-g005: als functions downstream of the Arm/β-Catenin destruction complex and upstream of activated Armadillo.Overexpression of the positive components Arrow (A), Dishevelled (L), ArmS10 (F and Q), or Arm (G and R) and depletion of negative components such as wingful (B), shaggy (C, H, and I), apc (D and J), slimb (E, O, and P), and axin (K) cause phenotypes that reflect ectopic Wg signaling: ectopic sensory bristles (A–G), ectopic veins (A–C), and tissue overgrowth (A, H–K, O, and P), but also reduced organ size upon strong pathway activation (B–F, L, M, and Q) and caused ectopic head cuticle (H–J, L, and M). Co-expression of alsRNAi could suppress these phenotypes in the wing (A′–D′) and the eye (H′–M′). Phenotypes based on slimbRNAi and ArmS10 expression could not be suppressed upon als depletion (E′, O′, P′, F′, and Q′). Arm overexpession could not rescue alsRNAi phenotypes (G′ and R′). Heads were photographed from dorsal views (left pictures of H–K′) or lateral views, anterior to the left (right pictures of H–K′, L–R′). alsRNAi lines: oligo310UAS (A′–E′) and for the eye analysis; oligo2_5′utr (G′).

Mentions: We manipulated the Wg signaling cascade in the wing and the eye at successively more downstream positions and assessed the requirement of Als for the manifestation of the respective Wg gain-of-signaling phenotypes (see legend of Figure 5 for a detailed description of phenotypes). First, we found that Als depletion suppressed Wg pathway activation caused by RNAi of notum/wingful (encoding an extracellular inhibitor of Wg [28],[29]) or by overexpression of Arrow (co-receptor of Wg, [17]) or Dishevelled (Figure 5A–5B′, 5L, and 5L′). Next, we tested knock-downs of components of the destruction complex, both in the wing and the eye, and found that aberrant Wg output was suppressed by alsRNAi for Axin, APC, and Shaggy/GSK3β (Figure 5C, 5C′, 5D, 5D′, 5H–5K′, 5M, and 5M′). Only the phenotypes of RNAi against the SCF/Slimb/βTRCP E3 ubiquitin ligase [30] and expression of ArmS10, a constitutively active form of Arm/β-Catenin [31], could not be ameliorated by als depletion (Figure 5E–5F′ and 5O–5Q′). Interestingly, wild-type Arm could not overcome the alsRNAi phenotypes in the wing and the eye, and did not lead to the emergence of ectopic sensory bristles (Figure 5G, 5G′, 5R, and 5R′). We interpret these genetic findings to indicate that Als functions upstream of Arm, but downstream of the Arm/β-Catenin destruction complex, possibly at the level of the SCF/Slimb/βTRCP E3 Ub ligase.


Protection of armadillo/β-Catenin by armless, a novel positive regulator of wingless signaling.

Reim G, Hruzova M, Goetze S, Basler K - PLoS Biol. (2014)

als functions downstream of the Arm/β-Catenin destruction complex and upstream of activated Armadillo.Overexpression of the positive components Arrow (A), Dishevelled (L), ArmS10 (F and Q), or Arm (G and R) and depletion of negative components such as wingful (B), shaggy (C, H, and I), apc (D and J), slimb (E, O, and P), and axin (K) cause phenotypes that reflect ectopic Wg signaling: ectopic sensory bristles (A–G), ectopic veins (A–C), and tissue overgrowth (A, H–K, O, and P), but also reduced organ size upon strong pathway activation (B–F, L, M, and Q) and caused ectopic head cuticle (H–J, L, and M). Co-expression of alsRNAi could suppress these phenotypes in the wing (A′–D′) and the eye (H′–M′). Phenotypes based on slimbRNAi and ArmS10 expression could not be suppressed upon als depletion (E′, O′, P′, F′, and Q′). Arm overexpession could not rescue alsRNAi phenotypes (G′ and R′). Heads were photographed from dorsal views (left pictures of H–K′) or lateral views, anterior to the left (right pictures of H–K′, L–R′). alsRNAi lines: oligo310UAS (A′–E′) and for the eye analysis; oligo2_5′utr (G′).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4219662&req=5

pbio-1001988-g005: als functions downstream of the Arm/β-Catenin destruction complex and upstream of activated Armadillo.Overexpression of the positive components Arrow (A), Dishevelled (L), ArmS10 (F and Q), or Arm (G and R) and depletion of negative components such as wingful (B), shaggy (C, H, and I), apc (D and J), slimb (E, O, and P), and axin (K) cause phenotypes that reflect ectopic Wg signaling: ectopic sensory bristles (A–G), ectopic veins (A–C), and tissue overgrowth (A, H–K, O, and P), but also reduced organ size upon strong pathway activation (B–F, L, M, and Q) and caused ectopic head cuticle (H–J, L, and M). Co-expression of alsRNAi could suppress these phenotypes in the wing (A′–D′) and the eye (H′–M′). Phenotypes based on slimbRNAi and ArmS10 expression could not be suppressed upon als depletion (E′, O′, P′, F′, and Q′). Arm overexpession could not rescue alsRNAi phenotypes (G′ and R′). Heads were photographed from dorsal views (left pictures of H–K′) or lateral views, anterior to the left (right pictures of H–K′, L–R′). alsRNAi lines: oligo310UAS (A′–E′) and for the eye analysis; oligo2_5′utr (G′).
Mentions: We manipulated the Wg signaling cascade in the wing and the eye at successively more downstream positions and assessed the requirement of Als for the manifestation of the respective Wg gain-of-signaling phenotypes (see legend of Figure 5 for a detailed description of phenotypes). First, we found that Als depletion suppressed Wg pathway activation caused by RNAi of notum/wingful (encoding an extracellular inhibitor of Wg [28],[29]) or by overexpression of Arrow (co-receptor of Wg, [17]) or Dishevelled (Figure 5A–5B′, 5L, and 5L′). Next, we tested knock-downs of components of the destruction complex, both in the wing and the eye, and found that aberrant Wg output was suppressed by alsRNAi for Axin, APC, and Shaggy/GSK3β (Figure 5C, 5C′, 5D, 5D′, 5H–5K′, 5M, and 5M′). Only the phenotypes of RNAi against the SCF/Slimb/βTRCP E3 ubiquitin ligase [30] and expression of ArmS10, a constitutively active form of Arm/β-Catenin [31], could not be ameliorated by als depletion (Figure 5E–5F′ and 5O–5Q′). Interestingly, wild-type Arm could not overcome the alsRNAi phenotypes in the wing and the eye, and did not lead to the emergence of ectopic sensory bristles (Figure 5G, 5G′, 5R, and 5R′). We interpret these genetic findings to indicate that Als functions upstream of Arm, but downstream of the Arm/β-Catenin destruction complex, possibly at the level of the SCF/Slimb/βTRCP E3 Ub ligase.

Bottom Line: We found by genetic and biochemical analyses that Als functions downstream of the destruction complex, at the level of the SCF/Slimb/βTRCP E3 Ub ligase.In the absence of Als, Arm levels are severely reduced.We suggest that Als antagonizes Ter94's positive effect on E3 ligase function and propose that Als promotes Wg signaling by rescuing Arm from proteolytic degradation, spotlighting an unexpected step where the Wg pathway signal is modulated.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland.

ABSTRACT
The Wingless (Wg/Wnt) signaling pathway is essential for metazoan development, where it is central to tissue growth and cellular differentiation. Deregulated Wg pathway activation underlies severe developmental abnormalities, as well as carcinogenesis. Armadillo/β-Catenin plays a key role in the Wg transduction cascade; its cytoplasmic and nuclear levels directly determine the output activity of Wg signaling and are thus tightly controlled. In all current models, once Arm is targeted for degradation by the Arm/β-Catenin destruction complex, its fate is viewed as set. We identified a novel Wg/Wnt pathway component, Armless (Als), which is required for Wg target gene expression in a cell-autonomous manner. We found by genetic and biochemical analyses that Als functions downstream of the destruction complex, at the level of the SCF/Slimb/βTRCP E3 Ub ligase. In the absence of Als, Arm levels are severely reduced. We show by biochemical and in vivo studies that Als interacts directly with Ter94, an AAA ATPase known to associate with E3 ligases and to drive protein turnover. We suggest that Als antagonizes Ter94's positive effect on E3 ligase function and propose that Als promotes Wg signaling by rescuing Arm from proteolytic degradation, spotlighting an unexpected step where the Wg pathway signal is modulated.

Show MeSH
Related in: MedlinePlus