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Regulatory mechanisms required for DE-cadherin function in cell migration and other types of adhesion.

Pacquelet A, Rørth P - J. Cell Biol. (2005)

Bottom Line: We have investigated the requirements for Drosophila melanogaster epithelial (DE) cadherin regulation in vivo.We found that (1) although linking DE-cadherin to alpha-catenin is essential, regulation of the link is not required in any of these types of adhesion; (2) beta-catenin is required only to link DE-cadherin to alpha-catenin; and (3) the cytoplasmic domain of DE-cadherin has an additional specific function for the invasive migration of border cells, which is conserved to other cadherins.The nature of this additional function is discussed.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory (EMBL), 69117 Heidelberg, Germany.

ABSTRACT
Cadherin-mediated adhesion can be regulated at many levels, as demonstrated by detailed analysis in cell lines. We have investigated the requirements for Drosophila melanogaster epithelial (DE) cadherin regulation in vivo. Investigating D. melanogaster oogenesis as a model system allowed the dissection of DE-cadherin function in several types of adhesion: cell sorting, cell positioning, epithelial integrity, and the cadherin-dependent process of border cell migration. We generated multiple fusions between DE-cadherin and alpha-catenin as well as point-mutated beta-catenin and analyzed their ability to support these types of adhesion. We found that (1) although linking DE-cadherin to alpha-catenin is essential, regulation of the link is not required in any of these types of adhesion; (2) beta-catenin is required only to link DE-cadherin to alpha-catenin; and (3) the cytoplasmic domain of DE-cadherin has an additional specific function for the invasive migration of border cells, which is conserved to other cadherins. The nature of this additional function is discussed.

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Molecular dissection of DE-cadherin cytoplasmic domain. (A) Schematic representation of proteins that were analyzed. (B) Alignment of D. melanogaster, Anopheles, human and mouse E-cadherin, and D. melanogaster N-cadherin transmembrane and cytoplasmic domains. In DE-cadherinΔCyt/α-catenin and DE-cadherinΔβ/α-catenin, α-catenin was fused at arrows 1 and 3, respectively. In DE-cadherinΔJM, the region between arrows 1 and 2 was deleted. Blue asterisks indicate p120 catenin–binding sites. Red asterisks indicate conserved tyrosines mutated in DE-cadherin–4YF. The limit between JM and COOH-terminal domain was drawn after studies on mammal E-cadherin. (C–E′) Expression of DE-cadherinΔβ/α-catenin in shg- mutant follicle (C and D) and border cells (E). (C and D) Mutant cells are above and to the right of the green line. (E) Three border cells (asterisks) are shg mutant and express DE-cadherinΔβ/α-catenin. (C and E) All DE-cadherin is detected. (D) Only surface DE-cadherin is detected. (E) Phalloidin (red) stains F-actin and DE-cadherin is in blue. (F) Border cell migration in shg mutant border cell clones expressing the indicated UAS transgenes. Bars (C and D), 20 μm; (E and E′) 10 μm.
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fig6: Molecular dissection of DE-cadherin cytoplasmic domain. (A) Schematic representation of proteins that were analyzed. (B) Alignment of D. melanogaster, Anopheles, human and mouse E-cadherin, and D. melanogaster N-cadherin transmembrane and cytoplasmic domains. In DE-cadherinΔCyt/α-catenin and DE-cadherinΔβ/α-catenin, α-catenin was fused at arrows 1 and 3, respectively. In DE-cadherinΔJM, the region between arrows 1 and 2 was deleted. Blue asterisks indicate p120 catenin–binding sites. Red asterisks indicate conserved tyrosines mutated in DE-cadherin–4YF. The limit between JM and COOH-terminal domain was drawn after studies on mammal E-cadherin. (C–E′) Expression of DE-cadherinΔβ/α-catenin in shg- mutant follicle (C and D) and border cells (E). (C and D) Mutant cells are above and to the right of the green line. (E) Three border cells (asterisks) are shg mutant and express DE-cadherinΔβ/α-catenin. (C and E) All DE-cadherin is detected. (D) Only surface DE-cadherin is detected. (E) Phalloidin (red) stains F-actin and DE-cadherin is in blue. (F) Border cell migration in shg mutant border cell clones expressing the indicated UAS transgenes. Bars (C and D), 20 μm; (E and E′) 10 μm.

Mentions: We sought to define more precisely which part of the cytoplasmic domain is specifically required for DE-cadherin function during border cell migration. We first tested whether the most COOH-terminal domain of DE-cadherin contained a contributing sequence by generating the intermediate fusion DE-cadherinΔβ/α-catenin (Fig. 6, A and B). In follicle and border cell shg mutant clones, DE-cadherinΔβ/α-catenin was expressed at high levels and accumulated inside the cells (Fig. 6, C and E) as well as at the cell surface (Fig. 6 D) in amounts similar to those observed with DE-cadherinΔCyt/CD2/α-catenin. DE-cadherinΔβ/α-catenin fully rescued shg follicle cell sorting and epithelial integrity (Fig. 6 D) as well as oocyte mispositioning (Fig. S3 A). In border cells, DE-cadherinΔβ/α-catenin partially rescued shg migration defects (Fig. 6 F). This indicates that the most COOH-terminal domain of DE-cadherin, although it is not strictly essential, contributes to efficient border cell migration. As border cells expressing DE-cadherin-FL/α-catenin in the absence of β-catenin migrate normally, this appears to be independent of binding to β-catenin.


Regulatory mechanisms required for DE-cadherin function in cell migration and other types of adhesion.

Pacquelet A, Rørth P - J. Cell Biol. (2005)

Molecular dissection of DE-cadherin cytoplasmic domain. (A) Schematic representation of proteins that were analyzed. (B) Alignment of D. melanogaster, Anopheles, human and mouse E-cadherin, and D. melanogaster N-cadherin transmembrane and cytoplasmic domains. In DE-cadherinΔCyt/α-catenin and DE-cadherinΔβ/α-catenin, α-catenin was fused at arrows 1 and 3, respectively. In DE-cadherinΔJM, the region between arrows 1 and 2 was deleted. Blue asterisks indicate p120 catenin–binding sites. Red asterisks indicate conserved tyrosines mutated in DE-cadherin–4YF. The limit between JM and COOH-terminal domain was drawn after studies on mammal E-cadherin. (C–E′) Expression of DE-cadherinΔβ/α-catenin in shg- mutant follicle (C and D) and border cells (E). (C and D) Mutant cells are above and to the right of the green line. (E) Three border cells (asterisks) are shg mutant and express DE-cadherinΔβ/α-catenin. (C and E) All DE-cadherin is detected. (D) Only surface DE-cadherin is detected. (E) Phalloidin (red) stains F-actin and DE-cadherin is in blue. (F) Border cell migration in shg mutant border cell clones expressing the indicated UAS transgenes. Bars (C and D), 20 μm; (E and E′) 10 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2171345&req=5

fig6: Molecular dissection of DE-cadherin cytoplasmic domain. (A) Schematic representation of proteins that were analyzed. (B) Alignment of D. melanogaster, Anopheles, human and mouse E-cadherin, and D. melanogaster N-cadherin transmembrane and cytoplasmic domains. In DE-cadherinΔCyt/α-catenin and DE-cadherinΔβ/α-catenin, α-catenin was fused at arrows 1 and 3, respectively. In DE-cadherinΔJM, the region between arrows 1 and 2 was deleted. Blue asterisks indicate p120 catenin–binding sites. Red asterisks indicate conserved tyrosines mutated in DE-cadherin–4YF. The limit between JM and COOH-terminal domain was drawn after studies on mammal E-cadherin. (C–E′) Expression of DE-cadherinΔβ/α-catenin in shg- mutant follicle (C and D) and border cells (E). (C and D) Mutant cells are above and to the right of the green line. (E) Three border cells (asterisks) are shg mutant and express DE-cadherinΔβ/α-catenin. (C and E) All DE-cadherin is detected. (D) Only surface DE-cadherin is detected. (E) Phalloidin (red) stains F-actin and DE-cadherin is in blue. (F) Border cell migration in shg mutant border cell clones expressing the indicated UAS transgenes. Bars (C and D), 20 μm; (E and E′) 10 μm.
Mentions: We sought to define more precisely which part of the cytoplasmic domain is specifically required for DE-cadherin function during border cell migration. We first tested whether the most COOH-terminal domain of DE-cadherin contained a contributing sequence by generating the intermediate fusion DE-cadherinΔβ/α-catenin (Fig. 6, A and B). In follicle and border cell shg mutant clones, DE-cadherinΔβ/α-catenin was expressed at high levels and accumulated inside the cells (Fig. 6, C and E) as well as at the cell surface (Fig. 6 D) in amounts similar to those observed with DE-cadherinΔCyt/CD2/α-catenin. DE-cadherinΔβ/α-catenin fully rescued shg follicle cell sorting and epithelial integrity (Fig. 6 D) as well as oocyte mispositioning (Fig. S3 A). In border cells, DE-cadherinΔβ/α-catenin partially rescued shg migration defects (Fig. 6 F). This indicates that the most COOH-terminal domain of DE-cadherin, although it is not strictly essential, contributes to efficient border cell migration. As border cells expressing DE-cadherin-FL/α-catenin in the absence of β-catenin migrate normally, this appears to be independent of binding to β-catenin.

Bottom Line: We have investigated the requirements for Drosophila melanogaster epithelial (DE) cadherin regulation in vivo.We found that (1) although linking DE-cadherin to alpha-catenin is essential, regulation of the link is not required in any of these types of adhesion; (2) beta-catenin is required only to link DE-cadherin to alpha-catenin; and (3) the cytoplasmic domain of DE-cadherin has an additional specific function for the invasive migration of border cells, which is conserved to other cadherins.The nature of this additional function is discussed.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory (EMBL), 69117 Heidelberg, Germany.

ABSTRACT
Cadherin-mediated adhesion can be regulated at many levels, as demonstrated by detailed analysis in cell lines. We have investigated the requirements for Drosophila melanogaster epithelial (DE) cadherin regulation in vivo. Investigating D. melanogaster oogenesis as a model system allowed the dissection of DE-cadherin function in several types of adhesion: cell sorting, cell positioning, epithelial integrity, and the cadherin-dependent process of border cell migration. We generated multiple fusions between DE-cadherin and alpha-catenin as well as point-mutated beta-catenin and analyzed their ability to support these types of adhesion. We found that (1) although linking DE-cadherin to alpha-catenin is essential, regulation of the link is not required in any of these types of adhesion; (2) beta-catenin is required only to link DE-cadherin to alpha-catenin; and (3) the cytoplasmic domain of DE-cadherin has an additional specific function for the invasive migration of border cells, which is conserved to other cadherins. The nature of this additional function is discussed.

Show MeSH
Related in: MedlinePlus