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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 expression results in enhanced integrin activity. (A) 293T cells were transfected as indicated and seeded on glass coverslips. The cells were either incubated with collagen- (col), α–integrin β1- (α-int), or BSA-coated microspheres for 2 h at 37°C. Nonadherent beads were removed by washing with PBS, and the cells were fixed and analyzed by microscopy. Arrowheads indicate transfected cells. (B) Quantification of cell-associated beads from samples shown in A. Bars represent the mean number of beads per transfected cell ± SD observed in three independent experiments in which 60 transfected cells for each sample were counted. (C) 293T cells expressing either RFP (red arrows) or CD105-GFP (green arrowheads) were mixed and plated together on glass coverslips. Cells were incubated with collagen- or α–integrin β1 antibody–coated microspheres for 2 h, washed, fixed, and analyzed by microscopy. Shown are representative fields of view.
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fig8: CD105 expression results in enhanced integrin activity. (A) 293T cells were transfected as indicated and seeded on glass coverslips. The cells were either incubated with collagen- (col), α–integrin β1- (α-int), or BSA-coated microspheres for 2 h at 37°C. Nonadherent beads were removed by washing with PBS, and the cells were fixed and analyzed by microscopy. Arrowheads indicate transfected cells. (B) Quantification of cell-associated beads from samples shown in A. Bars represent the mean number of beads per transfected cell ± SD observed in three independent experiments in which 60 transfected cells for each sample were counted. (C) 293T cells expressing either RFP (red arrows) or CD105-GFP (green arrowheads) were mixed and plated together on glass coverslips. Cells were incubated with collagen- or α–integrin β1 antibody–coated microspheres for 2 h, washed, fixed, and analyzed by microscopy. Shown are representative fields of view.

Mentions: A characteristic property of integrins is their ability to switch from a low to a high affinity state with respect to ligand binding in a process termed integrin activation (Hynes, 2002). Integrin activation can be promoted by signals from within the cell (inside-out signaling) by long-range conformational changes that reposition the ligand-binding site of the heterodimeric receptor and allow substrate binding (Xiao et al., 2004). To investigate whether CD105 expression has an influence on integrin activation, we incubated cells expressing GFP, CD105 WT, or CD105 ΔCT with microsphere beads coated with either BSA as a control or integrin β1 ligand collagen. After gentle washing, GFP-positive cells as well as cell-bound beads were visualized to allow an estimate of integrin activation on the single cell level (Fig. 8 A). Strikingly, there was a strong and selective binding of collagen-coated beads to CD105 WT–transfected cells. Untransfected, GFP-transfected, or CD105 ΔCT–expressing cells exhibited very little binding of collagen-coated beads that was indistinguishable from the low background binding of BSA control beads (Fig. 8 A). In addition, beads that were coated with an antibody directed against integrin β1 bound equally to all transfected as well as untransfected cells, demonstrating that integrin levels were not altered by CD105 expression and further indicating that CD105 influences integrin activity (Fig. 8 A). Quantification of the mean number of beads that were bound per cell corroborated the view that CD105 WT–expressing cells have a higher collagen-binding capacity as a result of increased integrin activity and not as a result of changes in integrin expression (Fig. 8 B). To finally demonstrate the cis-acting properties of CD105 on cell adhesion, cells expressing either RFP or CD105 WT–GFP were mixed, plated, and incubated with collagen-coated or anti–integrin β1-coated microspheres (Fig. 8 C). Whereas beads coated with the integrin β1 antibody bound equally well to both cells, the collagen-coated beads bound with high affinity almost exclusively to the CD105 WT–GFP-expressing cells (Fig. 8 C). Together, these results support the idea that CD105 positively influences integrin activation via its cytoplasmic domain, thereby increasing integrin affinity for ECM proteins and leading to enhanced cell adhesion.


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 expression results in enhanced integrin activity. (A) 293T cells were transfected as indicated and seeded on glass coverslips. The cells were either incubated with collagen- (col), α–integrin β1- (α-int), or BSA-coated microspheres for 2 h at 37°C. Nonadherent beads were removed by washing with PBS, and the cells were fixed and analyzed by microscopy. Arrowheads indicate transfected cells. (B) Quantification of cell-associated beads from samples shown in A. Bars represent the mean number of beads per transfected cell ± SD observed in three independent experiments in which 60 transfected cells for each sample were counted. (C) 293T cells expressing either RFP (red arrows) or CD105-GFP (green arrowheads) were mixed and plated together on glass coverslips. Cells were incubated with collagen- or α–integrin β1 antibody–coated microspheres for 2 h, washed, fixed, and analyzed by microscopy. Shown are representative fields of view.
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Related In: Results  -  Collection

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fig8: CD105 expression results in enhanced integrin activity. (A) 293T cells were transfected as indicated and seeded on glass coverslips. The cells were either incubated with collagen- (col), α–integrin β1- (α-int), or BSA-coated microspheres for 2 h at 37°C. Nonadherent beads were removed by washing with PBS, and the cells were fixed and analyzed by microscopy. Arrowheads indicate transfected cells. (B) Quantification of cell-associated beads from samples shown in A. Bars represent the mean number of beads per transfected cell ± SD observed in three independent experiments in which 60 transfected cells for each sample were counted. (C) 293T cells expressing either RFP (red arrows) or CD105-GFP (green arrowheads) were mixed and plated together on glass coverslips. Cells were incubated with collagen- or α–integrin β1 antibody–coated microspheres for 2 h, washed, fixed, and analyzed by microscopy. Shown are representative fields of view.
Mentions: A characteristic property of integrins is their ability to switch from a low to a high affinity state with respect to ligand binding in a process termed integrin activation (Hynes, 2002). Integrin activation can be promoted by signals from within the cell (inside-out signaling) by long-range conformational changes that reposition the ligand-binding site of the heterodimeric receptor and allow substrate binding (Xiao et al., 2004). To investigate whether CD105 expression has an influence on integrin activation, we incubated cells expressing GFP, CD105 WT, or CD105 ΔCT with microsphere beads coated with either BSA as a control or integrin β1 ligand collagen. After gentle washing, GFP-positive cells as well as cell-bound beads were visualized to allow an estimate of integrin activation on the single cell level (Fig. 8 A). Strikingly, there was a strong and selective binding of collagen-coated beads to CD105 WT–transfected cells. Untransfected, GFP-transfected, or CD105 ΔCT–expressing cells exhibited very little binding of collagen-coated beads that was indistinguishable from the low background binding of BSA control beads (Fig. 8 A). In addition, beads that were coated with an antibody directed against integrin β1 bound equally to all transfected as well as untransfected cells, demonstrating that integrin levels were not altered by CD105 expression and further indicating that CD105 influences integrin activity (Fig. 8 A). Quantification of the mean number of beads that were bound per cell corroborated the view that CD105 WT–expressing cells have a higher collagen-binding capacity as a result of increased integrin activity and not as a result of changes in integrin expression (Fig. 8 B). To finally demonstrate the cis-acting properties of CD105 on cell adhesion, cells expressing either RFP or CD105 WT–GFP were mixed, plated, and incubated with collagen-coated or anti–integrin β1-coated microspheres (Fig. 8 C). Whereas beads coated with the integrin β1 antibody bound equally well to both cells, the collagen-coated beads bound with high affinity almost exclusively to the CD105 WT–GFP-expressing cells (Fig. 8 C). Together, these results support the idea that CD105 positively influences integrin activation via its cytoplasmic domain, thereby increasing integrin affinity for ECM proteins and leading to enhanced cell adhesion.

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