Limits...
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

Only Opa- GC can transmigrate across the polarized epithelial monolayer.Polarized T84 cells on transwells were incubated with GC apically at a MOI of 10 for 6 h. The basolateral media were collected. T84 cells were washed and lysed to release cell-associated GC. The inoculum (A), cell lysates (B, adhered) and the basolateral media (C, transmigrated) were culture on GCK plates. CFU were determined. Opa expression of the GC colonies were determined using light microscopy (A and B) or colony blotting based on their binding activity to Mab 4B12. Shown were the average data from three independent experiments each with triplicate samples. **p<0.005, ***p< 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4526573&req=5

pone.0134342.g004: Only Opa- GC can transmigrate across the polarized epithelial monolayer.Polarized T84 cells on transwells were incubated with GC apically at a MOI of 10 for 6 h. The basolateral media were collected. T84 cells were washed and lysed to release cell-associated GC. The inoculum (A), cell lysates (B, adhered) and the basolateral media (C, transmigrated) were culture on GCK plates. CFU were determined. Opa expression of the GC colonies were determined using light microscopy (A and B) or colony blotting based on their binding activity to Mab 4B12. Shown were the average data from three independent experiments each with triplicate samples. **p<0.005, ***p< 0.001.

Mentions: Our finding that Opa expression decreases GC transmigration across polarized epithelial cells was surprising. To further confirm this result, we determined the number of GC and the Opa expression state of the inoculated (Fig 4A), attached (Fig 4B), and transmigrated GC (Fig 4C) after apical incubation for 6 h with MS11Opa+ (phase variable), MS11Opa- (phenotypically Opa- and phase variable), and MS11ΔOpa. The Opa expression phenotype was determined based on the morphology of resultant bacterial colonies. We found that the Opa phenotype of the attaching GC reflected the phenotype of the inoculum, with ~90% of attached MS11Opa+ strain expressing Opa and ~90% attached MS11Opa-lacking Opa expression (Fig 4A and 4B). In sharp contrast, all transmigrated bacteria were found to be Opa negative even when Opa-expressing MS11 were used to initiate the infection (Fig 4C). These results further confirm that GC that do not express Opa have an advantage over Opa-expressing GC in transmigrating across an epithelial barrier.


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)

Only Opa- GC can transmigrate across the polarized epithelial monolayer.Polarized T84 cells on transwells were incubated with GC apically at a MOI of 10 for 6 h. The basolateral media were collected. T84 cells were washed and lysed to release cell-associated GC. The inoculum (A), cell lysates (B, adhered) and the basolateral media (C, transmigrated) were culture on GCK plates. CFU were determined. Opa expression of the GC colonies were determined using light microscopy (A and B) or colony blotting based on their binding activity to Mab 4B12. Shown were the average data from three independent experiments each with triplicate samples. **p<0.005, ***p< 0.001.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0134342.g004: Only Opa- GC can transmigrate across the polarized epithelial monolayer.Polarized T84 cells on transwells were incubated with GC apically at a MOI of 10 for 6 h. The basolateral media were collected. T84 cells were washed and lysed to release cell-associated GC. The inoculum (A), cell lysates (B, adhered) and the basolateral media (C, transmigrated) were culture on GCK plates. CFU were determined. Opa expression of the GC colonies were determined using light microscopy (A and B) or colony blotting based on their binding activity to Mab 4B12. Shown were the average data from three independent experiments each with triplicate samples. **p<0.005, ***p< 0.001.
Mentions: Our finding that Opa expression decreases GC transmigration across polarized epithelial cells was surprising. To further confirm this result, we determined the number of GC and the Opa expression state of the inoculated (Fig 4A), attached (Fig 4B), and transmigrated GC (Fig 4C) after apical incubation for 6 h with MS11Opa+ (phase variable), MS11Opa- (phenotypically Opa- and phase variable), and MS11ΔOpa. The Opa expression phenotype was determined based on the morphology of resultant bacterial colonies. We found that the Opa phenotype of the attaching GC reflected the phenotype of the inoculum, with ~90% of attached MS11Opa+ strain expressing Opa and ~90% attached MS11Opa-lacking Opa expression (Fig 4A and 4B). In sharp contrast, all transmigrated bacteria were found to be Opa negative even when Opa-expressing MS11 were used to initiate the infection (Fig 4C). These results further confirm that GC that do not express Opa have an advantage over Opa-expressing GC in transmigrating across an epithelial barrier.

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