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Expression of Opacity Proteins Interferes with the Transmigration of Neisseria gonorrhoeae across Polarized Epithelial Cells.

Stein DC, LeVan A, Hardy B, Wang LC, Zimmerman L, Song W - PLoS ONE (2015)

Bottom Line: When MS11Opa+, able to phase vary Opa expression, was inoculated, only those bacteria that turn off Opa expression transmigrate across the polarized epithelial monolayer.Apical inoculation of MS11Opa+, but not MS11∆Opa, induces the recruitment of the Opa host-cell receptor carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) to the apical junction and the vicinity of bacterial adherent sites.Our results suggest that Opa expression limits gonococcal ability to invade into subepithelial tissues by forming tight interactions with neighboring bacteria and by inducing CEACAM redistribution to cell junctions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology & Molecular Genetics, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
Neisseria gonorrhoeae (GC) establishes infection at the mucosal surface of the human genital tract, most of which is lined with polarized epithelial cells. GC can cause localized as well as disseminated infections, leading to various complications. GC constantly change their surface structures via phase and antigenic variation, which has been implicated as a means for GC to establish infection at various anatomic locations of male and female genital tracks. However, the exact contribution of each surface molecule to bacterial infectivity remains elusive due to their phase variation. Using a GC derivative that is genetically devoid of all opa genes (MS11∆Opa), this study shows that Opa expression interferes with GC transmigration across polarized human epithelial cells. MS11∆Opa transmigrates across polarized epithelial cells much faster and to a greater extent than MS11Opa+, while adhering at a similar level as MS11Opa+. When MS11Opa+, able to phase vary Opa expression, was inoculated, only those bacteria that turn off Opa expression transmigrate across the polarized epithelial monolayer. Similar to bacteria alone or co-cultured with non-polarized epithelial cells, MS11∆Opa fails to form large microcolonies at the apical surface of polarized epithelial cells. Apical inoculation of MS11Opa+, but not MS11∆Opa, induces the recruitment of the Opa host-cell receptor carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) to the apical junction and the vicinity of bacterial adherent sites. Our results suggest that Opa expression limits gonococcal ability to invade into subepithelial tissues by forming tight interactions with neighboring bacteria and by inducing CEACAM redistribution to cell junctions.

No MeSH data available.


Related in: MedlinePlus

Opa expression changes gonococcal interaction with polarized epithelial cells.Polarized T84 cells on transwells were incubated with or without GC MS11Opa+, MS11ΔOpa, and MS11OpaH for 4 h. Cells were fixed, permeabilized, stained for the junctional protein ZO-1 and gonococci, and analyzed using a confocal microscope. Images shown are representative images from three independent experiments. Scale bar, 5 µm.
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pone.0134342.g001: Opa expression changes gonococcal interaction with polarized epithelial cells.Polarized T84 cells on transwells were incubated with or without GC MS11Opa+, MS11ΔOpa, and MS11OpaH for 4 h. Cells were fixed, permeabilized, stained for the junctional protein ZO-1 and gonococci, and analyzed using a confocal microscope. Images shown are representative images from three independent experiments. Scale bar, 5 µm.

Mentions: We previously demonstrated that Opa expression is critical for GC-GC interactions [37]. MS11Opa+ GC (a strain that can phase vary its Opa expression) but not MS11∆Opa (a strain genetically devoid of all opa genes) forms clumps when grown in broth and co-cultured with non-polarized cells. To determine how the expression of Opa impacts gonococcal interactions on mucosal surfaces, we analyzed the distribution of GC on the apical surface of polarized T84 cells grown on transwells. Polarized epithelial cells were incubated with bacteria in the apical chamber at an MOI of ~10 for 4 h. After fixation and extensive washing, cells were stained for the apical junctional protein ZO1 and GC, and examined by confocal fluorescence microscopy. We compared the apical distribution of three GC variants of MS11: MS11Opa+ (Opa-expressing wild type), MS11ΔOpa, and MS11OpaH (a derivative of MS11ΔOpa that had been genetically modified to express OpaH constitutively). We choose to use an OpaH-expressing derivative because this Opa promotes GC invasion into a variety of epithelial cell lines [40, 41]. We found that piliated MS11Opa+ and MS11OpaH formed large microcolonies, while MS11ΔOpa spread across the apical surface of the monolayer, appearing as diplococci or small clumps (Fig 1). While each of these strain express different pilE variants, the pilin that they express all have aspartic acid at residue 140 of the hypervariable region that is associated with low adhesiveness, instead of lysine that is associated with high adhesiveness [42]. These results suggest that Opa is responsible for GC aggregation on the apical surface of polarized epithelial cells, and that GC aggregation can be mediated by the expression of a single isoform of Opa.


Expression of Opacity Proteins Interferes with the Transmigration of Neisseria gonorrhoeae across Polarized Epithelial Cells.

Stein DC, LeVan A, Hardy B, Wang LC, Zimmerman L, Song W - PLoS ONE (2015)

Opa expression changes gonococcal interaction with polarized epithelial cells.Polarized T84 cells on transwells were incubated with or without GC MS11Opa+, MS11ΔOpa, and MS11OpaH for 4 h. Cells were fixed, permeabilized, stained for the junctional protein ZO-1 and gonococci, and analyzed using a confocal microscope. Images shown are representative images from three independent experiments. Scale bar, 5 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526573&req=5

pone.0134342.g001: Opa expression changes gonococcal interaction with polarized epithelial cells.Polarized T84 cells on transwells were incubated with or without GC MS11Opa+, MS11ΔOpa, and MS11OpaH for 4 h. Cells were fixed, permeabilized, stained for the junctional protein ZO-1 and gonococci, and analyzed using a confocal microscope. Images shown are representative images from three independent experiments. Scale bar, 5 µm.
Mentions: We previously demonstrated that Opa expression is critical for GC-GC interactions [37]. MS11Opa+ GC (a strain that can phase vary its Opa expression) but not MS11∆Opa (a strain genetically devoid of all opa genes) forms clumps when grown in broth and co-cultured with non-polarized cells. To determine how the expression of Opa impacts gonococcal interactions on mucosal surfaces, we analyzed the distribution of GC on the apical surface of polarized T84 cells grown on transwells. Polarized epithelial cells were incubated with bacteria in the apical chamber at an MOI of ~10 for 4 h. After fixation and extensive washing, cells were stained for the apical junctional protein ZO1 and GC, and examined by confocal fluorescence microscopy. We compared the apical distribution of three GC variants of MS11: MS11Opa+ (Opa-expressing wild type), MS11ΔOpa, and MS11OpaH (a derivative of MS11ΔOpa that had been genetically modified to express OpaH constitutively). We choose to use an OpaH-expressing derivative because this Opa promotes GC invasion into a variety of epithelial cell lines [40, 41]. We found that piliated MS11Opa+ and MS11OpaH formed large microcolonies, while MS11ΔOpa spread across the apical surface of the monolayer, appearing as diplococci or small clumps (Fig 1). While each of these strain express different pilE variants, the pilin that they express all have aspartic acid at residue 140 of the hypervariable region that is associated with low adhesiveness, instead of lysine that is associated with high adhesiveness [42]. These results suggest that Opa is responsible for GC aggregation on the apical surface of polarized epithelial cells, and that GC aggregation can be mediated by the expression of a single isoform of Opa.

Bottom Line: When MS11Opa+, able to phase vary Opa expression, was inoculated, only those bacteria that turn off Opa expression transmigrate across the polarized epithelial monolayer.Apical inoculation of MS11Opa+, but not MS11∆Opa, induces the recruitment of the Opa host-cell receptor carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) to the apical junction and the vicinity of bacterial adherent sites.Our results suggest that Opa expression limits gonococcal ability to invade into subepithelial tissues by forming tight interactions with neighboring bacteria and by inducing CEACAM redistribution to cell junctions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology & Molecular Genetics, University of Maryland, College Park, Maryland, United States of America.

ABSTRACT
Neisseria gonorrhoeae (GC) establishes infection at the mucosal surface of the human genital tract, most of which is lined with polarized epithelial cells. GC can cause localized as well as disseminated infections, leading to various complications. GC constantly change their surface structures via phase and antigenic variation, which has been implicated as a means for GC to establish infection at various anatomic locations of male and female genital tracks. However, the exact contribution of each surface molecule to bacterial infectivity remains elusive due to their phase variation. Using a GC derivative that is genetically devoid of all opa genes (MS11∆Opa), this study shows that Opa expression interferes with GC transmigration across polarized human epithelial cells. MS11∆Opa transmigrates across polarized epithelial cells much faster and to a greater extent than MS11Opa+, while adhering at a similar level as MS11Opa+. When MS11Opa+, able to phase vary Opa expression, was inoculated, only those bacteria that turn off Opa expression transmigrate across the polarized epithelial monolayer. Similar to bacteria alone or co-cultured with non-polarized epithelial cells, MS11∆Opa fails to form large microcolonies at the apical surface of polarized epithelial cells. Apical inoculation of MS11Opa+, but not MS11∆Opa, induces the recruitment of the Opa host-cell receptor carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) to the apical junction and the vicinity of bacterial adherent sites. Our results suggest that Opa expression limits gonococcal ability to invade into subepithelial tissues by forming tight interactions with neighboring bacteria and by inducing CEACAM redistribution to cell junctions.

No MeSH data available.


Related in: MedlinePlus