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The chlamydial periplasmic stress response serine protease cHtrA is secreted into host cell cytosol.

Wu X, Lei L, Gong S, Chen D, Flores R, Zhong G - BMC Microbiol. (2011)

Bottom Line: The periplasmic cHtrA protein appeared to be actively secreted into host cell cytosol since no other chlamydial periplasmic proteins were detected in the host cell cytoplasm.Most chlamydial species secreted cHtrA into host cell cytosol and the secretion was not inhibitable by a type III secretion inhibitor.Since it is hypothesized that chlamydial organisms possess a proteolysis strategy to manipulate host cell signaling pathways, secretion of the serine protease cHtrA into host cell cytosol suggests that the periplasmic cHtrA may also play an important role in chlamydial interactions with host cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology and Immunology, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX 78229, USA.

ABSTRACT

Background: The periplasmic High Temperature Requirement protein A (HtrA) plays important roles in bacterial protein folding and stress responses. However, the role of chlamydial HtrA (cHtrA) in chlamydial pathogenesis is not clear.

Results: The cHtrA was detected both inside and outside the chlamydial inclusions. The detection was specific since both polyclonal and monoclonal anti-cHtrA antibodies revealed similar intracellular labeling patterns that were only removed by absorption with cHtrA but not control fusion proteins. In a Western blot assay, the anti-cHtrA antibodies detected the endogenous cHtrA in Chlamydia-infected cells without cross-reacting with any other chlamydial or host cell antigens. Fractionation of the infected cells revealed cHtrA in the host cell cytosol fraction. The periplasmic cHtrA protein appeared to be actively secreted into host cell cytosol since no other chlamydial periplasmic proteins were detected in the host cell cytoplasm. Most chlamydial species secreted cHtrA into host cell cytosol and the secretion was not inhibitable by a type III secretion inhibitor.

Conclusion: Since it is hypothesized that chlamydial organisms possess a proteolysis strategy to manipulate host cell signaling pathways, secretion of the serine protease cHtrA into host cell cytosol suggests that the periplasmic cHtrA may also play an important role in chlamydial interactions with host cells.

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The cHtrA but not other chlamydial periplasmic proteins are secreted into host cell cytosol. HeLa cells infected with C. trachomatis organisms were processed and co-labeled with mouse antibodies against various periplasmic proteins (red) and a rabbit antibody against IncA (green) as described in Figure 1 legend. The Hoechst dye was used to visualize DNA (blue). The triple labeling was analyzed under a conventional fluorescence microscope (A) and confocal microscope (B). Under the confocal microscope, a series of four images were taken along the Z-axis by varying 1 μM between each. Note that cHtrA (red arrows) but none of the other periplasmic proteins including CT067, CT539 & CT783 was detected outside of the inclusion membrane (green arrows) by either immunofluorescence microscopy or confocal microscopy.
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Figure 3: The cHtrA but not other chlamydial periplasmic proteins are secreted into host cell cytosol. HeLa cells infected with C. trachomatis organisms were processed and co-labeled with mouse antibodies against various periplasmic proteins (red) and a rabbit antibody against IncA (green) as described in Figure 1 legend. The Hoechst dye was used to visualize DNA (blue). The triple labeling was analyzed under a conventional fluorescence microscope (A) and confocal microscope (B). Under the confocal microscope, a series of four images were taken along the Z-axis by varying 1 μM between each. Note that cHtrA (red arrows) but none of the other periplasmic proteins including CT067, CT539 & CT783 was detected outside of the inclusion membrane (green arrows) by either immunofluorescence microscopy or confocal microscopy.

Mentions: Since cHtrA is a periplasmic protein, we next tested whether localization in the host cell cytosol is a common characteristic of all chlamydial periplasmic proteins. The intracellular distributions of two periplasmic proteins involved in disulfide bond formation (CT539, TrxA or thioredoxin) and isomerization (CT783, PDI or protein disulfide bond isomerase; http://stdgen.northwestern.edu/) respectively and one periplasmic iron binding protein (CT067, YtgA, an ABC transporter system component; ref: [59,60]) were compared with that of cHtrA (Figure 3). Under a conventional fluorescence microscope (A), only cHtrA but not the other periplasmic proteins including CT067, CT539 & CT783 was detected outside of the chlamydial inclusions. This observation was confirmed under a confocal microscope (B). The Z-axis serial section images showed that cHtrA was clearly detected both inside and outside the inclusion membrane but CT067 was only detected inside the inclusion membrane.


The chlamydial periplasmic stress response serine protease cHtrA is secreted into host cell cytosol.

Wu X, Lei L, Gong S, Chen D, Flores R, Zhong G - BMC Microbiol. (2011)

The cHtrA but not other chlamydial periplasmic proteins are secreted into host cell cytosol. HeLa cells infected with C. trachomatis organisms were processed and co-labeled with mouse antibodies against various periplasmic proteins (red) and a rabbit antibody against IncA (green) as described in Figure 1 legend. The Hoechst dye was used to visualize DNA (blue). The triple labeling was analyzed under a conventional fluorescence microscope (A) and confocal microscope (B). Under the confocal microscope, a series of four images were taken along the Z-axis by varying 1 μM between each. Note that cHtrA (red arrows) but none of the other periplasmic proteins including CT067, CT539 & CT783 was detected outside of the inclusion membrane (green arrows) by either immunofluorescence microscopy or confocal microscopy.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3107777&req=5

Figure 3: The cHtrA but not other chlamydial periplasmic proteins are secreted into host cell cytosol. HeLa cells infected with C. trachomatis organisms were processed and co-labeled with mouse antibodies against various periplasmic proteins (red) and a rabbit antibody against IncA (green) as described in Figure 1 legend. The Hoechst dye was used to visualize DNA (blue). The triple labeling was analyzed under a conventional fluorescence microscope (A) and confocal microscope (B). Under the confocal microscope, a series of four images were taken along the Z-axis by varying 1 μM between each. Note that cHtrA (red arrows) but none of the other periplasmic proteins including CT067, CT539 & CT783 was detected outside of the inclusion membrane (green arrows) by either immunofluorescence microscopy or confocal microscopy.
Mentions: Since cHtrA is a periplasmic protein, we next tested whether localization in the host cell cytosol is a common characteristic of all chlamydial periplasmic proteins. The intracellular distributions of two periplasmic proteins involved in disulfide bond formation (CT539, TrxA or thioredoxin) and isomerization (CT783, PDI or protein disulfide bond isomerase; http://stdgen.northwestern.edu/) respectively and one periplasmic iron binding protein (CT067, YtgA, an ABC transporter system component; ref: [59,60]) were compared with that of cHtrA (Figure 3). Under a conventional fluorescence microscope (A), only cHtrA but not the other periplasmic proteins including CT067, CT539 & CT783 was detected outside of the chlamydial inclusions. This observation was confirmed under a confocal microscope (B). The Z-axis serial section images showed that cHtrA was clearly detected both inside and outside the inclusion membrane but CT067 was only detected inside the inclusion membrane.

Bottom Line: The periplasmic cHtrA protein appeared to be actively secreted into host cell cytosol since no other chlamydial periplasmic proteins were detected in the host cell cytoplasm.Most chlamydial species secreted cHtrA into host cell cytosol and the secretion was not inhibitable by a type III secretion inhibitor.Since it is hypothesized that chlamydial organisms possess a proteolysis strategy to manipulate host cell signaling pathways, secretion of the serine protease cHtrA into host cell cytosol suggests that the periplasmic cHtrA may also play an important role in chlamydial interactions with host cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology and Immunology, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX 78229, USA.

ABSTRACT

Background: The periplasmic High Temperature Requirement protein A (HtrA) plays important roles in bacterial protein folding and stress responses. However, the role of chlamydial HtrA (cHtrA) in chlamydial pathogenesis is not clear.

Results: The cHtrA was detected both inside and outside the chlamydial inclusions. The detection was specific since both polyclonal and monoclonal anti-cHtrA antibodies revealed similar intracellular labeling patterns that were only removed by absorption with cHtrA but not control fusion proteins. In a Western blot assay, the anti-cHtrA antibodies detected the endogenous cHtrA in Chlamydia-infected cells without cross-reacting with any other chlamydial or host cell antigens. Fractionation of the infected cells revealed cHtrA in the host cell cytosol fraction. The periplasmic cHtrA protein appeared to be actively secreted into host cell cytosol since no other chlamydial periplasmic proteins were detected in the host cell cytoplasm. Most chlamydial species secreted cHtrA into host cell cytosol and the secretion was not inhibitable by a type III secretion inhibitor.

Conclusion: Since it is hypothesized that chlamydial organisms possess a proteolysis strategy to manipulate host cell signaling pathways, secretion of the serine protease cHtrA into host cell cytosol suggests that the periplasmic cHtrA may also play an important role in chlamydial interactions with host cells.

Show MeSH
Related in: MedlinePlus