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Distinct molecular forms of beta-catenin are targeted to adhesive or transcriptional complexes.

Gottardi CJ, Gumbiner BM - J. Cell Biol. (2004)

Bottom Line: The Wnt-stimulated, TCF-selective form is monomeric and is regulated by the COOH terminus of beta-catenin, which selectively competes cadherin binding through an intramolecular fold-back mechanism.Phosphorylation of the cadherin reverses the TCF binding selectivity, suggesting another potential layer of regulation.In contrast, the main cadherin-binding form of beta-catenin is a beta-catenin-alpha-catenin dimer, indicating that there is a distinct molecular form of beta-catenin that can interact with both the cadherin and alpha-catenin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, University of Virginia School of Medicine, Charlottesville, VA 22908, USA. c-gottardi@northwestern.edu.

ABSTRACT
Beta-catenin plays essential roles in both cell-cell adhesion and Wnt signal transduction, but what precisely controls beta-catenin targeting to cadherin adhesive complexes, or T-cell factor (TCF)-transcriptional complexes is less well understood. We show that during Wnt signaling, a form of beta-catenin is generated that binds TCF but not the cadherin cytoplasmic domain. The Wnt-stimulated, TCF-selective form is monomeric and is regulated by the COOH terminus of beta-catenin, which selectively competes cadherin binding through an intramolecular fold-back mechanism. Phosphorylation of the cadherin reverses the TCF binding selectivity, suggesting another potential layer of regulation. In contrast, the main cadherin-binding form of beta-catenin is a beta-catenin-alpha-catenin dimer, indicating that there is a distinct molecular form of beta-catenin that can interact with both the cadherin and alpha-catenin. We propose that participation of beta-catenin in adhesion or Wnt signaling is dictated by the regulation of distinct molecular forms of beta-catenin with different binding properties, rather than simple competition between cadherins and TCFs for a single constitutive form. This model explains how cells can control whether beta-catenin is used independently in cell adhesion and nuclear signaling, or competitively so that the two processes are coordinated and interrelated.

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The COOH terminus of β-catenin restricts binding to cadherin. COOH terminus of β-catenin competes cadherin, but not TCF binding. (A) Schematic shows where α-catenin, cadherin, and TCF interact with β-catenin (Huber et al., 1997; Graham et al., 2000; Pokutta and Weis, 2000; Huber and Weis, 2001). (B) The COOH terminus of β-catenin binds the arm repeat region of β-catenin in yeast-two hybrid (Cox et al., 1999) and recombinant protein assays (Piedra et al., 2001). (C) COOH-terminal region of β-catenin competes β-catenin binding to cad-GST, but not to TCF-GST fusion protein. Recombinant β-catenin (1.5 μg) purified from baculovirus (Suh and Gumbiner, 2003) was incubated with cadherin-GST (2 μg) or TCF-GST (2.4 μg) coupled agarose beads in the presence of increasing amounts of β-catenin COOH-terminal peptide (amino acids 695–781). Affinity precipitates were analyzed by SDS-PAGE and Western blotting with an antibody to β-catenin. (D) Cadherin-GST preferentially depletes the fraction of β-catenin recognized by a COOH-terminal mAb (M5.2). A cytosolic fraction from Rat1/Wnt cells was affinity precipitated (×3) with cadherin-GST (lanes 1–3). The cad-GST nonbinding pool (lanes 4 and 5) was divided in two and immunoprecipitated with either an mAb that recognizes a COOH-terminal β-catenin epitope (βC-mAb (M5.2), lane 4) or an NH2-terminal β-catenin epitope (βN-mAb (1.1), lane 5). As a control, these antibodies were used to immunoprecipitate β-catenin from the total starting material (not previously depleted with cad-GST; lanes 6 and 7).
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fig2: The COOH terminus of β-catenin restricts binding to cadherin. COOH terminus of β-catenin competes cadherin, but not TCF binding. (A) Schematic shows where α-catenin, cadherin, and TCF interact with β-catenin (Huber et al., 1997; Graham et al., 2000; Pokutta and Weis, 2000; Huber and Weis, 2001). (B) The COOH terminus of β-catenin binds the arm repeat region of β-catenin in yeast-two hybrid (Cox et al., 1999) and recombinant protein assays (Piedra et al., 2001). (C) COOH-terminal region of β-catenin competes β-catenin binding to cad-GST, but not to TCF-GST fusion protein. Recombinant β-catenin (1.5 μg) purified from baculovirus (Suh and Gumbiner, 2003) was incubated with cadherin-GST (2 μg) or TCF-GST (2.4 μg) coupled agarose beads in the presence of increasing amounts of β-catenin COOH-terminal peptide (amino acids 695–781). Affinity precipitates were analyzed by SDS-PAGE and Western blotting with an antibody to β-catenin. (D) Cadherin-GST preferentially depletes the fraction of β-catenin recognized by a COOH-terminal mAb (M5.2). A cytosolic fraction from Rat1/Wnt cells was affinity precipitated (×3) with cadherin-GST (lanes 1–3). The cad-GST nonbinding pool (lanes 4 and 5) was divided in two and immunoprecipitated with either an mAb that recognizes a COOH-terminal β-catenin epitope (βC-mAb (M5.2), lane 4) or an NH2-terminal β-catenin epitope (βN-mAb (1.1), lane 5). As a control, these antibodies were used to immunoprecipitate β-catenin from the total starting material (not previously depleted with cad-GST; lanes 6 and 7).

Mentions: Several findings influenced our investigation of a mechanism that could generate a form of β-catenin that binds selectively to TCF. The COOH terminus of β-catenin can interact with the armadillo repeat region of β-catenin (Cox et al., 1999; Piedra et al., 2001) and compete with β-catenin binding to the cadherin cytoplasmic domain in vitro (Castano et al., 2002). These observations raised the possibility that conformational changes in the COOH terminus of β-catenin, and in particular, a “closed” conformation, might be incompatible with cadherin binding. Because Wnt signaling alters β-catenin binding to the cadherin and TCF differently (Fig. 1), we sought to determine whether the COOH terminus of β-catenin might contribute to this binding selectivity. Indeed, although the COOH terminus of β-catenin can compete β-catenin binding to the cadherin, as demonstrated previously (Castano et al., 2002), its interaction with TCF is not competed (Fig. 2 C). Thus, if a COOH-terminal conformational change giving rise to a closed form of β-catenin were to occur during Wnt signaling, it could alter β-catenin ligand interactions selectively.


Distinct molecular forms of beta-catenin are targeted to adhesive or transcriptional complexes.

Gottardi CJ, Gumbiner BM - J. Cell Biol. (2004)

The COOH terminus of β-catenin restricts binding to cadherin. COOH terminus of β-catenin competes cadherin, but not TCF binding. (A) Schematic shows where α-catenin, cadherin, and TCF interact with β-catenin (Huber et al., 1997; Graham et al., 2000; Pokutta and Weis, 2000; Huber and Weis, 2001). (B) The COOH terminus of β-catenin binds the arm repeat region of β-catenin in yeast-two hybrid (Cox et al., 1999) and recombinant protein assays (Piedra et al., 2001). (C) COOH-terminal region of β-catenin competes β-catenin binding to cad-GST, but not to TCF-GST fusion protein. Recombinant β-catenin (1.5 μg) purified from baculovirus (Suh and Gumbiner, 2003) was incubated with cadherin-GST (2 μg) or TCF-GST (2.4 μg) coupled agarose beads in the presence of increasing amounts of β-catenin COOH-terminal peptide (amino acids 695–781). Affinity precipitates were analyzed by SDS-PAGE and Western blotting with an antibody to β-catenin. (D) Cadherin-GST preferentially depletes the fraction of β-catenin recognized by a COOH-terminal mAb (M5.2). A cytosolic fraction from Rat1/Wnt cells was affinity precipitated (×3) with cadherin-GST (lanes 1–3). The cad-GST nonbinding pool (lanes 4 and 5) was divided in two and immunoprecipitated with either an mAb that recognizes a COOH-terminal β-catenin epitope (βC-mAb (M5.2), lane 4) or an NH2-terminal β-catenin epitope (βN-mAb (1.1), lane 5). As a control, these antibodies were used to immunoprecipitate β-catenin from the total starting material (not previously depleted with cad-GST; lanes 6 and 7).
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fig2: The COOH terminus of β-catenin restricts binding to cadherin. COOH terminus of β-catenin competes cadherin, but not TCF binding. (A) Schematic shows where α-catenin, cadherin, and TCF interact with β-catenin (Huber et al., 1997; Graham et al., 2000; Pokutta and Weis, 2000; Huber and Weis, 2001). (B) The COOH terminus of β-catenin binds the arm repeat region of β-catenin in yeast-two hybrid (Cox et al., 1999) and recombinant protein assays (Piedra et al., 2001). (C) COOH-terminal region of β-catenin competes β-catenin binding to cad-GST, but not to TCF-GST fusion protein. Recombinant β-catenin (1.5 μg) purified from baculovirus (Suh and Gumbiner, 2003) was incubated with cadherin-GST (2 μg) or TCF-GST (2.4 μg) coupled agarose beads in the presence of increasing amounts of β-catenin COOH-terminal peptide (amino acids 695–781). Affinity precipitates were analyzed by SDS-PAGE and Western blotting with an antibody to β-catenin. (D) Cadherin-GST preferentially depletes the fraction of β-catenin recognized by a COOH-terminal mAb (M5.2). A cytosolic fraction from Rat1/Wnt cells was affinity precipitated (×3) with cadherin-GST (lanes 1–3). The cad-GST nonbinding pool (lanes 4 and 5) was divided in two and immunoprecipitated with either an mAb that recognizes a COOH-terminal β-catenin epitope (βC-mAb (M5.2), lane 4) or an NH2-terminal β-catenin epitope (βN-mAb (1.1), lane 5). As a control, these antibodies were used to immunoprecipitate β-catenin from the total starting material (not previously depleted with cad-GST; lanes 6 and 7).
Mentions: Several findings influenced our investigation of a mechanism that could generate a form of β-catenin that binds selectively to TCF. The COOH terminus of β-catenin can interact with the armadillo repeat region of β-catenin (Cox et al., 1999; Piedra et al., 2001) and compete with β-catenin binding to the cadherin cytoplasmic domain in vitro (Castano et al., 2002). These observations raised the possibility that conformational changes in the COOH terminus of β-catenin, and in particular, a “closed” conformation, might be incompatible with cadherin binding. Because Wnt signaling alters β-catenin binding to the cadherin and TCF differently (Fig. 1), we sought to determine whether the COOH terminus of β-catenin might contribute to this binding selectivity. Indeed, although the COOH terminus of β-catenin can compete β-catenin binding to the cadherin, as demonstrated previously (Castano et al., 2002), its interaction with TCF is not competed (Fig. 2 C). Thus, if a COOH-terminal conformational change giving rise to a closed form of β-catenin were to occur during Wnt signaling, it could alter β-catenin ligand interactions selectively.

Bottom Line: The Wnt-stimulated, TCF-selective form is monomeric and is regulated by the COOH terminus of beta-catenin, which selectively competes cadherin binding through an intramolecular fold-back mechanism.Phosphorylation of the cadherin reverses the TCF binding selectivity, suggesting another potential layer of regulation.In contrast, the main cadherin-binding form of beta-catenin is a beta-catenin-alpha-catenin dimer, indicating that there is a distinct molecular form of beta-catenin that can interact with both the cadherin and alpha-catenin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, University of Virginia School of Medicine, Charlottesville, VA 22908, USA. c-gottardi@northwestern.edu.

ABSTRACT
Beta-catenin plays essential roles in both cell-cell adhesion and Wnt signal transduction, but what precisely controls beta-catenin targeting to cadherin adhesive complexes, or T-cell factor (TCF)-transcriptional complexes is less well understood. We show that during Wnt signaling, a form of beta-catenin is generated that binds TCF but not the cadherin cytoplasmic domain. The Wnt-stimulated, TCF-selective form is monomeric and is regulated by the COOH terminus of beta-catenin, which selectively competes cadherin binding through an intramolecular fold-back mechanism. Phosphorylation of the cadherin reverses the TCF binding selectivity, suggesting another potential layer of regulation. In contrast, the main cadherin-binding form of beta-catenin is a beta-catenin-alpha-catenin dimer, indicating that there is a distinct molecular form of beta-catenin that can interact with both the cadherin and alpha-catenin. We propose that participation of beta-catenin in adhesion or Wnt signaling is dictated by the regulation of distinct molecular forms of beta-catenin with different binding properties, rather than simple competition between cadherins and TCFs for a single constitutive form. This model explains how cells can control whether beta-catenin is used independently in cell adhesion and nuclear signaling, or competitively so that the two processes are coordinated and interrelated.

Show MeSH
Related in: MedlinePlus