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Binding site for p120/delta-catenin is not required for Drosophila E-cadherin function in vivo.

Pacquelet A, Lin L, Rorth P - J. Cell Biol. (2003)

Bottom Line: As expected, DE-cadherin-Delta beta did not substitute for DE-cadherin in these processes, although it retained some residual activity.Surprisingly, DE-cadherin-AAA was able to substitute for the wild-type protein in all contexts with no detectable perturbations.Thus, interaction with p120/delta-catenin does not appear to be required for DE-cadherin function in vivo.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory, 69117 Heidelberg, Germany.

ABSTRACT
Homophilic cell adhesion mediated by classical cadherins is important for many developmental processes. Proteins that interact with the cytoplasmic domain of cadherin, in particular the catenins, are thought to regulate the strength and possibly the dynamics of adhesion. beta-catenin links cadherin to the actin cytoskeleton via alpha-catenin. The role of p120/delta-catenin proteins in regulating cadherin function is less clear. Both beta-catenin and p120/delta-catenin are conserved in Drosophila. Here, we address the importance of cadherin-catenin interactions in vivo, using mutant variants of Drosophila epithelial cadherin (DE-cadherin) that are selectively defective in p120ctn (DE-cadherin-AAA) or beta-catenin-armadillo (DE-cadherin-Delta beta) interactions. We have analyzed the ability of these proteins to substitute for endogenous DE-cadherin activity in multiple cadherin-dependent processes during Drosophila development and oogenesis; epithelial integrity, follicle cell sorting, oocyte positioning, as well as the dynamic adhesion required for border cell migration. As expected, DE-cadherin-Delta beta did not substitute for DE-cadherin in these processes, although it retained some residual activity. Surprisingly, DE-cadherin-AAA was able to substitute for the wild-type protein in all contexts with no detectable perturbations. Thus, interaction with p120/delta-catenin does not appear to be required for DE-cadherin function in vivo.

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Effect of p120ctn-RNAi transgenes. (A–C) Levels of HA-p120ctn in border cells (arrow) and centripetal cells (arrowheads) overexpressing HA-p120ctn (A) and different RNAi constructs (B and C); stage 10b egg chambers. All constructs are expressed using slboGal4, which starts expressing in border cells at stage 9 and in centripetal cells at stage 10. HA-p120ctn levels are severely reduced in border cells, estimated to have been expressing RNAi constructs for 12–24 h. Bar, ∼10 μm. (D) Stage 9 egg chamber from a female of the genotype hsFLP/actin-Flipout-Gal4; UAS-RNAi/+; UAS-GFP/+. Larval heat shock was used to initiate constitutive expression of the RNAi in somatic cells (marked by GFP) one week before analysis. Border cell migration is normal. Bar, ∼15 μm. (E) Quantification of phenotypes after expression of p120ctn-RNAi transgenes in the ovary (as in D) and in embryos (maternal [mat] and zygotic [zyg] expression).
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fig5: Effect of p120ctn-RNAi transgenes. (A–C) Levels of HA-p120ctn in border cells (arrow) and centripetal cells (arrowheads) overexpressing HA-p120ctn (A) and different RNAi constructs (B and C); stage 10b egg chambers. All constructs are expressed using slboGal4, which starts expressing in border cells at stage 9 and in centripetal cells at stage 10. HA-p120ctn levels are severely reduced in border cells, estimated to have been expressing RNAi constructs for 12–24 h. Bar, ∼10 μm. (D) Stage 9 egg chamber from a female of the genotype hsFLP/actin-Flipout-Gal4; UAS-RNAi/+; UAS-GFP/+. Larval heat shock was used to initiate constitutive expression of the RNAi in somatic cells (marked by GFP) one week before analysis. Border cell migration is normal. Bar, ∼15 μm. (E) Quantification of phenotypes after expression of p120ctn-RNAi transgenes in the ovary (as in D) and in embryos (maternal [mat] and zygotic [zyg] expression).

Mentions: In an attempt to independently compromise p120ctn function, we used double-stranded RNA-mediated interference (RNAi). Two p120ctn-RNAi transgenes (RNAi-700 and RNAi-2200) were able to severely reduce the levels of co-overexpressed HA-p120ctn (Fig. 5, A–C), suggesting that levels of endogenous p120ctn would also be strongly reduced. Expression of p120-RNAi in somatic cells did not cause any detectable phenotype in the ovary (Fig. 5, D and E). This is consistent with p120ctn not being important for DE-cadherin function during oogenesis. However, the lack of phenotype may also reflect incomplete removal of p120ctn. Maternal and zygotic expression of p120ctn-RNAi transgenes did not affect embryonic viability (Fig. 5 E). Severe defects have been observed on injection of p120ctn double-stranded RNA in embryos (Magie et al., 2002). Resolving the discrepancy between these results awaits analysis of a p120ctn mutant.


Binding site for p120/delta-catenin is not required for Drosophila E-cadherin function in vivo.

Pacquelet A, Lin L, Rorth P - J. Cell Biol. (2003)

Effect of p120ctn-RNAi transgenes. (A–C) Levels of HA-p120ctn in border cells (arrow) and centripetal cells (arrowheads) overexpressing HA-p120ctn (A) and different RNAi constructs (B and C); stage 10b egg chambers. All constructs are expressed using slboGal4, which starts expressing in border cells at stage 9 and in centripetal cells at stage 10. HA-p120ctn levels are severely reduced in border cells, estimated to have been expressing RNAi constructs for 12–24 h. Bar, ∼10 μm. (D) Stage 9 egg chamber from a female of the genotype hsFLP/actin-Flipout-Gal4; UAS-RNAi/+; UAS-GFP/+. Larval heat shock was used to initiate constitutive expression of the RNAi in somatic cells (marked by GFP) one week before analysis. Border cell migration is normal. Bar, ∼15 μm. (E) Quantification of phenotypes after expression of p120ctn-RNAi transgenes in the ovary (as in D) and in embryos (maternal [mat] and zygotic [zyg] expression).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172659&req=5

fig5: Effect of p120ctn-RNAi transgenes. (A–C) Levels of HA-p120ctn in border cells (arrow) and centripetal cells (arrowheads) overexpressing HA-p120ctn (A) and different RNAi constructs (B and C); stage 10b egg chambers. All constructs are expressed using slboGal4, which starts expressing in border cells at stage 9 and in centripetal cells at stage 10. HA-p120ctn levels are severely reduced in border cells, estimated to have been expressing RNAi constructs for 12–24 h. Bar, ∼10 μm. (D) Stage 9 egg chamber from a female of the genotype hsFLP/actin-Flipout-Gal4; UAS-RNAi/+; UAS-GFP/+. Larval heat shock was used to initiate constitutive expression of the RNAi in somatic cells (marked by GFP) one week before analysis. Border cell migration is normal. Bar, ∼15 μm. (E) Quantification of phenotypes after expression of p120ctn-RNAi transgenes in the ovary (as in D) and in embryos (maternal [mat] and zygotic [zyg] expression).
Mentions: In an attempt to independently compromise p120ctn function, we used double-stranded RNA-mediated interference (RNAi). Two p120ctn-RNAi transgenes (RNAi-700 and RNAi-2200) were able to severely reduce the levels of co-overexpressed HA-p120ctn (Fig. 5, A–C), suggesting that levels of endogenous p120ctn would also be strongly reduced. Expression of p120-RNAi in somatic cells did not cause any detectable phenotype in the ovary (Fig. 5, D and E). This is consistent with p120ctn not being important for DE-cadherin function during oogenesis. However, the lack of phenotype may also reflect incomplete removal of p120ctn. Maternal and zygotic expression of p120ctn-RNAi transgenes did not affect embryonic viability (Fig. 5 E). Severe defects have been observed on injection of p120ctn double-stranded RNA in embryos (Magie et al., 2002). Resolving the discrepancy between these results awaits analysis of a p120ctn mutant.

Bottom Line: As expected, DE-cadherin-Delta beta did not substitute for DE-cadherin in these processes, although it retained some residual activity.Surprisingly, DE-cadherin-AAA was able to substitute for the wild-type protein in all contexts with no detectable perturbations.Thus, interaction with p120/delta-catenin does not appear to be required for DE-cadherin function in vivo.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory, 69117 Heidelberg, Germany.

ABSTRACT
Homophilic cell adhesion mediated by classical cadherins is important for many developmental processes. Proteins that interact with the cytoplasmic domain of cadherin, in particular the catenins, are thought to regulate the strength and possibly the dynamics of adhesion. beta-catenin links cadherin to the actin cytoskeleton via alpha-catenin. The role of p120/delta-catenin proteins in regulating cadherin function is less clear. Both beta-catenin and p120/delta-catenin are conserved in Drosophila. Here, we address the importance of cadherin-catenin interactions in vivo, using mutant variants of Drosophila epithelial cadherin (DE-cadherin) that are selectively defective in p120ctn (DE-cadherin-AAA) or beta-catenin-armadillo (DE-cadherin-Delta beta) interactions. We have analyzed the ability of these proteins to substitute for endogenous DE-cadherin activity in multiple cadherin-dependent processes during Drosophila development and oogenesis; epithelial integrity, follicle cell sorting, oocyte positioning, as well as the dynamic adhesion required for border cell migration. As expected, DE-cadherin-Delta beta did not substitute for DE-cadherin in these processes, although it retained some residual activity. Surprisingly, DE-cadherin-AAA was able to substitute for the wild-type protein in all contexts with no detectable perturbations. Thus, interaction with p120/delta-catenin does not appear to be required for DE-cadherin function in vivo.

Show MeSH
Related in: MedlinePlus