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Drosophila p120catenin plays a supporting role in cell adhesion but is not an essential adherens junction component.

Myster SH, Cavallo R, Anderson CT, Fox DT, Peifer M - J. Cell Biol. (2003)

Bottom Line: We generated alleles of p120 and found that mutants are viable and fertile and have no substantial changes in junction structure or function.However, p120 mutations strongly enhance mutations in the genes encoding DE-cadherin or Armadillo, the beta-catenin homologue.Finally, we examined the localization of p120 during embryogenesis. p120 localizes to adherens junctions, but its localization there is less universal than that of core adherens junction proteins.

View Article: PubMed Central - PubMed

Affiliation: Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.

ABSTRACT
Cadherin-catenin complexes, localized to adherens junctions, are essential for cell-cell adhesion. One means of regulating adhesion is through the juxtamembrane domain of the cadherin cytoplasmic tail. This region is the binding site for p120, leading to the hypothesis that p120 is a key regulator of cell adhesion. p120 has also been suggested to regulate the GTPase Rho and to regulate transcription via its binding partner Kaiso. To test these hypothesized functions, we turned to Drosophila, which has only a single p120 family member. It localizes to adherens junctions and binds the juxtamembrane region of DE-cadherin (DE-cad). We generated alleles of p120 and found that mutants are viable and fertile and have no substantial changes in junction structure or function. However, p120 mutations strongly enhance mutations in the genes encoding DE-cadherin or Armadillo, the beta-catenin homologue. Finally, we examined the localization of p120 during embryogenesis. p120 localizes to adherens junctions, but its localization there is less universal than that of core adherens junction proteins. Together, these data suggest that p120 is an important positive modulator of adhesion but that it is not an essential core component of adherens junctions.

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p120 localizes to AJs and the cytoplasm. (A and B) Wild-type (WT) and homozygous p120308 (p120) embryonic extracts immunoblotted with affinity-purified rat anti-p120 (A) or rabbit anti-p120 (B). (C–O) Embryos. Embryonic stages are as in Wieschaus and Nüsslein-Volhard (1986). (C and D) Stage 11. (C) Apical section through epidermis; more basal section, cutting across the folded epithelium (D). p120 localizes to the apical cell cortex (arrows). (E–G) Stage 11. p120 (red); Arm (green). Cells are indicated in which p120 accumulation is relatively high (red arrows) or low (green arrow). (H) Stage 11 p120380 mutant. (I) Live image; p120-GFP. Higher levels are seen at the ends of cells that are stretched (arrows). (J) Stage 12, ubiquitin-myc-p120. (K and L) Stage 12, expressing myc-p120 in prd stripes. Myc (red), DE-cad (green). DE-cad is uniform across the embryo; thus, myc-p120 overexpression does not affect its localization. (M–O) Stage 17, optical cross sections. (M and N) p120 (red) and Arm (green) colocalize to apical AJs of the gut (white arrows). (O) p120-GFP (green) and DE-cad (red) colocalize at AJs of the gut (arrow) and epidermis (arrowhead). (P) Egg chamber in ovary. p120-GFP (green) and Arm (red) colocalize at follicle cell AJs (arrow). Bars, 5 μm.
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fig2: p120 localizes to AJs and the cytoplasm. (A and B) Wild-type (WT) and homozygous p120308 (p120) embryonic extracts immunoblotted with affinity-purified rat anti-p120 (A) or rabbit anti-p120 (B). (C–O) Embryos. Embryonic stages are as in Wieschaus and Nüsslein-Volhard (1986). (C and D) Stage 11. (C) Apical section through epidermis; more basal section, cutting across the folded epithelium (D). p120 localizes to the apical cell cortex (arrows). (E–G) Stage 11. p120 (red); Arm (green). Cells are indicated in which p120 accumulation is relatively high (red arrows) or low (green arrow). (H) Stage 11 p120380 mutant. (I) Live image; p120-GFP. Higher levels are seen at the ends of cells that are stretched (arrows). (J) Stage 12, ubiquitin-myc-p120. (K and L) Stage 12, expressing myc-p120 in prd stripes. Myc (red), DE-cad (green). DE-cad is uniform across the embryo; thus, myc-p120 overexpression does not affect its localization. (M–O) Stage 17, optical cross sections. (M and N) p120 (red) and Arm (green) colocalize to apical AJs of the gut (white arrows). (O) p120-GFP (green) and DE-cad (red) colocalize at AJs of the gut (arrow) and epidermis (arrowhead). (P) Egg chamber in ovary. p120-GFP (green) and Arm (red) colocalize at follicle cell AJs (arrow). Bars, 5 μm.

Mentions: To further examine whether fly p120 was a p120 ortholog, we examined its subcellular localization. To do so, we generated rat and rabbit polyclonal antisera to its COOH-terminal 96 aa. The work described below, unless noted, uses affinity-purified rat anti-p120. In immunoblots of embryo extracts, this primarily recognizes a single protein of ∼88 kD (Fig. 2 A; p120's predicted mol wt = 86.7 kD). This protein is absent in a p120 mutant (Fig. 2 A; see below), confirming that it is encoded by p120. This ∼88 kD protein is also the major protein recognized by rabbit anti-p120 (Fig. 2 B). Both antisera variably cross-react with other proteins, but no other protein was consistently recognized. The rat antisera specifically recognize p120 in embryos. In p120308 mutants, cell junctional staining was lost and overall staining reduced (Fig. 2, F versus H). In parallel to generating antibodies, we generated fusions of p120 with six myc epitopes at the NH2 terminus (myc-p120) or GFP at the COOH terminus (p120-GFP). These were expressed ubiquitously using the ubiquitin promoter or at specific times and places using the GAL4-UAS system. Since they were not expressed from the p120 promoter, we primarily used them to confirm p120's subcellular localization.


Drosophila p120catenin plays a supporting role in cell adhesion but is not an essential adherens junction component.

Myster SH, Cavallo R, Anderson CT, Fox DT, Peifer M - J. Cell Biol. (2003)

p120 localizes to AJs and the cytoplasm. (A and B) Wild-type (WT) and homozygous p120308 (p120) embryonic extracts immunoblotted with affinity-purified rat anti-p120 (A) or rabbit anti-p120 (B). (C–O) Embryos. Embryonic stages are as in Wieschaus and Nüsslein-Volhard (1986). (C and D) Stage 11. (C) Apical section through epidermis; more basal section, cutting across the folded epithelium (D). p120 localizes to the apical cell cortex (arrows). (E–G) Stage 11. p120 (red); Arm (green). Cells are indicated in which p120 accumulation is relatively high (red arrows) or low (green arrow). (H) Stage 11 p120380 mutant. (I) Live image; p120-GFP. Higher levels are seen at the ends of cells that are stretched (arrows). (J) Stage 12, ubiquitin-myc-p120. (K and L) Stage 12, expressing myc-p120 in prd stripes. Myc (red), DE-cad (green). DE-cad is uniform across the embryo; thus, myc-p120 overexpression does not affect its localization. (M–O) Stage 17, optical cross sections. (M and N) p120 (red) and Arm (green) colocalize to apical AJs of the gut (white arrows). (O) p120-GFP (green) and DE-cad (red) colocalize at AJs of the gut (arrow) and epidermis (arrowhead). (P) Egg chamber in ovary. p120-GFP (green) and Arm (red) colocalize at follicle cell AJs (arrow). Bars, 5 μm.
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fig2: p120 localizes to AJs and the cytoplasm. (A and B) Wild-type (WT) and homozygous p120308 (p120) embryonic extracts immunoblotted with affinity-purified rat anti-p120 (A) or rabbit anti-p120 (B). (C–O) Embryos. Embryonic stages are as in Wieschaus and Nüsslein-Volhard (1986). (C and D) Stage 11. (C) Apical section through epidermis; more basal section, cutting across the folded epithelium (D). p120 localizes to the apical cell cortex (arrows). (E–G) Stage 11. p120 (red); Arm (green). Cells are indicated in which p120 accumulation is relatively high (red arrows) or low (green arrow). (H) Stage 11 p120380 mutant. (I) Live image; p120-GFP. Higher levels are seen at the ends of cells that are stretched (arrows). (J) Stage 12, ubiquitin-myc-p120. (K and L) Stage 12, expressing myc-p120 in prd stripes. Myc (red), DE-cad (green). DE-cad is uniform across the embryo; thus, myc-p120 overexpression does not affect its localization. (M–O) Stage 17, optical cross sections. (M and N) p120 (red) and Arm (green) colocalize to apical AJs of the gut (white arrows). (O) p120-GFP (green) and DE-cad (red) colocalize at AJs of the gut (arrow) and epidermis (arrowhead). (P) Egg chamber in ovary. p120-GFP (green) and Arm (red) colocalize at follicle cell AJs (arrow). Bars, 5 μm.
Mentions: To further examine whether fly p120 was a p120 ortholog, we examined its subcellular localization. To do so, we generated rat and rabbit polyclonal antisera to its COOH-terminal 96 aa. The work described below, unless noted, uses affinity-purified rat anti-p120. In immunoblots of embryo extracts, this primarily recognizes a single protein of ∼88 kD (Fig. 2 A; p120's predicted mol wt = 86.7 kD). This protein is absent in a p120 mutant (Fig. 2 A; see below), confirming that it is encoded by p120. This ∼88 kD protein is also the major protein recognized by rabbit anti-p120 (Fig. 2 B). Both antisera variably cross-react with other proteins, but no other protein was consistently recognized. The rat antisera specifically recognize p120 in embryos. In p120308 mutants, cell junctional staining was lost and overall staining reduced (Fig. 2, F versus H). In parallel to generating antibodies, we generated fusions of p120 with six myc epitopes at the NH2 terminus (myc-p120) or GFP at the COOH terminus (p120-GFP). These were expressed ubiquitously using the ubiquitin promoter or at specific times and places using the GAL4-UAS system. Since they were not expressed from the p120 promoter, we primarily used them to confirm p120's subcellular localization.

Bottom Line: We generated alleles of p120 and found that mutants are viable and fertile and have no substantial changes in junction structure or function.However, p120 mutations strongly enhance mutations in the genes encoding DE-cadherin or Armadillo, the beta-catenin homologue.Finally, we examined the localization of p120 during embryogenesis. p120 localizes to adherens junctions, but its localization there is less universal than that of core adherens junction proteins.

View Article: PubMed Central - PubMed

Affiliation: Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.

ABSTRACT
Cadherin-catenin complexes, localized to adherens junctions, are essential for cell-cell adhesion. One means of regulating adhesion is through the juxtamembrane domain of the cadherin cytoplasmic tail. This region is the binding site for p120, leading to the hypothesis that p120 is a key regulator of cell adhesion. p120 has also been suggested to regulate the GTPase Rho and to regulate transcription via its binding partner Kaiso. To test these hypothesized functions, we turned to Drosophila, which has only a single p120 family member. It localizes to adherens junctions and binds the juxtamembrane region of DE-cadherin (DE-cad). We generated alleles of p120 and found that mutants are viable and fertile and have no substantial changes in junction structure or function. However, p120 mutations strongly enhance mutations in the genes encoding DE-cadherin or Armadillo, the beta-catenin homologue. Finally, we examined the localization of p120 during embryogenesis. p120 localizes to adherens junctions, but its localization there is less universal than that of core adherens junction proteins. Together, these data suggest that p120 is an important positive modulator of adhesion but that it is not an essential core component of adherens junctions.

Show MeSH
Related in: MedlinePlus