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CEACAM engagement by human pathogens enhances cell adhesion and counteracts bacteria-induced detachment of epithelial cells.

Muenzner P, Rohde M, Kneitz S, Hauck CR - J. Cell Biol. (2005)

Bottom Line: Blockage of CD105 expression by antisense oligonucleotides abolished infection-induced cell adhesion.The CD105-mediated increase in cell adhesion was dependent on the presence and function of integrin beta1.CD105 expression did not elevate cellular integrin levels but caused a dramatic increase in the ECM-binding capacity of the cells, suggesting that CD105 affects integrin activity.

View Article: PubMed Central - PubMed

Affiliation: Zentrum für Infektionsforschung, Universität Würzburg, 97070 Würzburg, Germany.

ABSTRACT
Exfoliation, which is the detachment of infected epithelial cells, is an innate defense mechanism to prevent bacterial colonization. Indeed, infection with Neisseria gonorrhoeae induced epithelial detachment from an extracellular matrix (ECM) substrate in vitro. Surprisingly, variants of N. gonorrhoeae that bind to human carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) failed to induce detachment and, instead, promoted enhanced host cell adhesion to the ECM. Microarray analysis revealed that CEACAM engagement by several human pathogens triggers expression of CD105. Blockage of CD105 expression by antisense oligonucleotides abolished infection-induced cell adhesion. The expression of full-length CD105 promoted cell adhesion to the ECM and was sufficient to prevent infection-induced detachment. The CD105-mediated increase in cell adhesion was dependent on the presence and function of integrin beta1. CD105 expression did not elevate cellular integrin levels but caused a dramatic increase in the ECM-binding capacity of the cells, suggesting that CD105 affects integrin activity. The exploitation of CEACAMs to trigger CD105 expression and to counteract infection-induced cell detachment represents an intriguing adaptation of pathogens that are specialized to colonize the human mucosa.

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CD105 cooperates with integrin β1 to mediate enhanced cell adhesion. (A) 293T cells were transfected with control vector (pcDNA), CEACAM1, or CD105 WT; infected (black bars) or left uninfected (white bars) for 8 h with Ngo OpaCEA; and used in adhesion assays in the absence or presence of mAbs against CD105, integrin β1 (αIntβ1), CD44, or with control mouse IgG. (B) Integrin-deficient fibroblasts (GD25 cells) or GD25 cells expressing human integrin β1 were transfected with CD105-GFP or the empty vector control (pEGFP) and were used in adhesion assays. Bars represent means ± SD of five (A) or eight wells (B). Bottom panel shows Western blotting of whole cell lysates with anti-GFP antibody. (C) 293T cells transfected as in B were stained with anti–integrin β1 antibodies (clone P5D2) and phycoerythrin (PE)-conjugated secondary antibodies (black). Controls (gray lines) were stained with isotype-matched control IgG. Upon gating of GFP-positive cells, phycoerythrin-derived fluorescence was measured in 10,000 transfected cells. A representative experiment that was repeated three times is shown. FL2-H, intensity of the fluorescence signal detected in channel 2. (D) 293T cells were transfected as in C or were left untransfected (control). 2 d later, whole cell lysates as well as purified collagen type IV (Col) and fibronectin (FN) as positive controls were analyzed by Western blotting with antibodies against either collagen type IV (top left) or fibronectin (top right). After stripping, the membrane was developed with anti–β-actin antibodies demonstrating equal loading (bottom).
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fig7: CD105 cooperates with integrin β1 to mediate enhanced cell adhesion. (A) 293T cells were transfected with control vector (pcDNA), CEACAM1, or CD105 WT; infected (black bars) or left uninfected (white bars) for 8 h with Ngo OpaCEA; and used in adhesion assays in the absence or presence of mAbs against CD105, integrin β1 (αIntβ1), CD44, or with control mouse IgG. (B) Integrin-deficient fibroblasts (GD25 cells) or GD25 cells expressing human integrin β1 were transfected with CD105-GFP or the empty vector control (pEGFP) and were used in adhesion assays. Bars represent means ± SD of five (A) or eight wells (B). Bottom panel shows Western blotting of whole cell lysates with anti-GFP antibody. (C) 293T cells transfected as in B were stained with anti–integrin β1 antibodies (clone P5D2) and phycoerythrin (PE)-conjugated secondary antibodies (black). Controls (gray lines) were stained with isotype-matched control IgG. Upon gating of GFP-positive cells, phycoerythrin-derived fluorescence was measured in 10,000 transfected cells. A representative experiment that was repeated three times is shown. FL2-H, intensity of the fluorescence signal detected in channel 2. (D) 293T cells were transfected as in C or were left untransfected (control). 2 d later, whole cell lysates as well as purified collagen type IV (Col) and fibronectin (FN) as positive controls were analyzed by Western blotting with antibodies against either collagen type IV (top left) or fibronectin (top right). After stripping, the membrane was developed with anti–β-actin antibodies demonstrating equal loading (bottom).

Mentions: Because the enhanced cell adhesion that was triggered by CD105 was most pronounced on surfaces coated with ligands for β1 integrins, we wondered whether this process involved integrins. Cell adhesion assays that were performed in the presence of monoclonal antibodies directed against either CD105, integrin β1, or the hyaluronate receptor CD44 demonstrated that interference with both CD105 or integrin β1 abolished enhanced cell adhesion after bacterial infection or CD105 expression (Fig. 7 A). In contrast, anti-CD44 antibodies or control mouse IgG had no influence on cell adhesion to collagen (Fig. 7 A). Importantly, CD105 expression did not lead to enhanced cell adhesion in cells that were genetically deficient for integrin β1, whereas reexpression of human integrin β1 restored the ability of these cells to display enhanced cell adhesion upon CD105 expression, demonstrating that integrin β1 is essential for this process (Fig. 7 B). Flow cytometry revealed that the levels of integrin β1 are not modulated by the presence of CD105 (Fig. 7 C). Furthermore, CD105 expression did not alter the production of ECM proteins, such as collagen or fibronectin, by 293T cells (Fig. 7 D), suggesting that CD105 exerts its effect by altering the ligand-binding properties of integrin β1.


CEACAM engagement by human pathogens enhances cell adhesion and counteracts bacteria-induced detachment of epithelial cells.

Muenzner P, Rohde M, Kneitz S, Hauck CR - J. Cell Biol. (2005)

CD105 cooperates with integrin β1 to mediate enhanced cell adhesion. (A) 293T cells were transfected with control vector (pcDNA), CEACAM1, or CD105 WT; infected (black bars) or left uninfected (white bars) for 8 h with Ngo OpaCEA; and used in adhesion assays in the absence or presence of mAbs against CD105, integrin β1 (αIntβ1), CD44, or with control mouse IgG. (B) Integrin-deficient fibroblasts (GD25 cells) or GD25 cells expressing human integrin β1 were transfected with CD105-GFP or the empty vector control (pEGFP) and were used in adhesion assays. Bars represent means ± SD of five (A) or eight wells (B). Bottom panel shows Western blotting of whole cell lysates with anti-GFP antibody. (C) 293T cells transfected as in B were stained with anti–integrin β1 antibodies (clone P5D2) and phycoerythrin (PE)-conjugated secondary antibodies (black). Controls (gray lines) were stained with isotype-matched control IgG. Upon gating of GFP-positive cells, phycoerythrin-derived fluorescence was measured in 10,000 transfected cells. A representative experiment that was repeated three times is shown. FL2-H, intensity of the fluorescence signal detected in channel 2. (D) 293T cells were transfected as in C or were left untransfected (control). 2 d later, whole cell lysates as well as purified collagen type IV (Col) and fibronectin (FN) as positive controls were analyzed by Western blotting with antibodies against either collagen type IV (top left) or fibronectin (top right). After stripping, the membrane was developed with anti–β-actin antibodies demonstrating equal loading (bottom).
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Related In: Results  -  Collection

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fig7: CD105 cooperates with integrin β1 to mediate enhanced cell adhesion. (A) 293T cells were transfected with control vector (pcDNA), CEACAM1, or CD105 WT; infected (black bars) or left uninfected (white bars) for 8 h with Ngo OpaCEA; and used in adhesion assays in the absence or presence of mAbs against CD105, integrin β1 (αIntβ1), CD44, or with control mouse IgG. (B) Integrin-deficient fibroblasts (GD25 cells) or GD25 cells expressing human integrin β1 were transfected with CD105-GFP or the empty vector control (pEGFP) and were used in adhesion assays. Bars represent means ± SD of five (A) or eight wells (B). Bottom panel shows Western blotting of whole cell lysates with anti-GFP antibody. (C) 293T cells transfected as in B were stained with anti–integrin β1 antibodies (clone P5D2) and phycoerythrin (PE)-conjugated secondary antibodies (black). Controls (gray lines) were stained with isotype-matched control IgG. Upon gating of GFP-positive cells, phycoerythrin-derived fluorescence was measured in 10,000 transfected cells. A representative experiment that was repeated three times is shown. FL2-H, intensity of the fluorescence signal detected in channel 2. (D) 293T cells were transfected as in C or were left untransfected (control). 2 d later, whole cell lysates as well as purified collagen type IV (Col) and fibronectin (FN) as positive controls were analyzed by Western blotting with antibodies against either collagen type IV (top left) or fibronectin (top right). After stripping, the membrane was developed with anti–β-actin antibodies demonstrating equal loading (bottom).
Mentions: Because the enhanced cell adhesion that was triggered by CD105 was most pronounced on surfaces coated with ligands for β1 integrins, we wondered whether this process involved integrins. Cell adhesion assays that were performed in the presence of monoclonal antibodies directed against either CD105, integrin β1, or the hyaluronate receptor CD44 demonstrated that interference with both CD105 or integrin β1 abolished enhanced cell adhesion after bacterial infection or CD105 expression (Fig. 7 A). In contrast, anti-CD44 antibodies or control mouse IgG had no influence on cell adhesion to collagen (Fig. 7 A). Importantly, CD105 expression did not lead to enhanced cell adhesion in cells that were genetically deficient for integrin β1, whereas reexpression of human integrin β1 restored the ability of these cells to display enhanced cell adhesion upon CD105 expression, demonstrating that integrin β1 is essential for this process (Fig. 7 B). Flow cytometry revealed that the levels of integrin β1 are not modulated by the presence of CD105 (Fig. 7 C). Furthermore, CD105 expression did not alter the production of ECM proteins, such as collagen or fibronectin, by 293T cells (Fig. 7 D), suggesting that CD105 exerts its effect by altering the ligand-binding properties of integrin β1.

Bottom Line: Blockage of CD105 expression by antisense oligonucleotides abolished infection-induced cell adhesion.The CD105-mediated increase in cell adhesion was dependent on the presence and function of integrin beta1.CD105 expression did not elevate cellular integrin levels but caused a dramatic increase in the ECM-binding capacity of the cells, suggesting that CD105 affects integrin activity.

View Article: PubMed Central - PubMed

Affiliation: Zentrum für Infektionsforschung, Universität Würzburg, 97070 Würzburg, Germany.

ABSTRACT
Exfoliation, which is the detachment of infected epithelial cells, is an innate defense mechanism to prevent bacterial colonization. Indeed, infection with Neisseria gonorrhoeae induced epithelial detachment from an extracellular matrix (ECM) substrate in vitro. Surprisingly, variants of N. gonorrhoeae that bind to human carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) failed to induce detachment and, instead, promoted enhanced host cell adhesion to the ECM. Microarray analysis revealed that CEACAM engagement by several human pathogens triggers expression of CD105. Blockage of CD105 expression by antisense oligonucleotides abolished infection-induced cell adhesion. The expression of full-length CD105 promoted cell adhesion to the ECM and was sufficient to prevent infection-induced detachment. The CD105-mediated increase in cell adhesion was dependent on the presence and function of integrin beta1. CD105 expression did not elevate cellular integrin levels but caused a dramatic increase in the ECM-binding capacity of the cells, suggesting that CD105 affects integrin activity. The exploitation of CEACAMs to trigger CD105 expression and to counteract infection-induced cell detachment represents an intriguing adaptation of pathogens that are specialized to colonize the human mucosa.

Show MeSH
Related in: MedlinePlus