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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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Time course of cHtrA expression during C. trachomatis infection. The C. trachomatis-infected culture samples were processed at various times after infection (as indicated on the top) for immunofluorescence staining as described in Figure 1 legend. The mouse anti-cHtrA (a to h) and anti-CPAF (mAb 100a; i to p) were visualized with a goat anti-mouse IgG conjugated with Cy3 (red) while the chlamydial organisms were visualized with a rabbit anti-chlamydia antibody plus a goat anti-rabbit IgG-Cy2 conjugate (green). Note that cHtrA was first detected inside the chlamydial inclusions at 12 hours after infection [panel d, yellow (overlapping with organisms) & red (free of chlamydial organisms) arrowheads], similar to the detection of CPAF. However, cHtrA secretion into host cell cytosol was only detected 24 h after infection while secretion of CPAF was already obvious by 16 h post infection.
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Figure 5: Time course of cHtrA expression during C. trachomatis infection. The C. trachomatis-infected culture samples were processed at various times after infection (as indicated on the top) for immunofluorescence staining as described in Figure 1 legend. The mouse anti-cHtrA (a to h) and anti-CPAF (mAb 100a; i to p) were visualized with a goat anti-mouse IgG conjugated with Cy3 (red) while the chlamydial organisms were visualized with a rabbit anti-chlamydia antibody plus a goat anti-rabbit IgG-Cy2 conjugate (green). Note that cHtrA was first detected inside the chlamydial inclusions at 12 hours after infection [panel d, yellow (overlapping with organisms) & red (free of chlamydial organisms) arrowheads], similar to the detection of CPAF. However, cHtrA secretion into host cell cytosol was only detected 24 h after infection while secretion of CPAF was already obvious by 16 h post infection.

Mentions: We further used the specific anti-cHtrA antibodies to characterize the endogenous cHtrA. As shown in Figure 5, cHtrA protein was detected inside the inclusions as early as 12 h after infection and secretion of cHtrA into host cell cytosol became apparent by 24 h post infection. Although CPAF was also detectable at 12 h, the secretion of CPAF was more robust and became very obvious as early as 16 h after infection. The cHtrA protein was detected both within the chlamydial inclusions and in the host cell cytosol while CPAF mainly accumulated in the host cell cytosol as infection progressed. Although both CPAF and cHtrA are serine proteases secreted by C. trachomatis organisms, their distinct secretion kinetics and intracellular distribution patterns suggest that they may fulfill different functions during chlamydial infection. To further evaluate whether cHtrA secretion is common to all chlamydial organisms, we monitored the cHtrA protein distribution in cells infected with various serovars and strains from different chlamydial species, including 13 C. trachomatis serovars and also isolates representing species of C. muridarum, C. caviae, C. pneumoniae and C. psittaci (Figure 6). The cHtrA protein was consistently detected in both the lumen of chlamydial inclusion and cytosol of host cells infected with all serovars of C. trachomatis organisms and isolates of C. muridarum, C. caviae and C. pneumoniae but not C. psittaci. Although secretion of cHtrA into the inclusion lumen and further into the cytosol of the infected cells seems to be a common feature of most chlamydial organisms tested, it is not known at this moment why the species C. psittaci, which primarily infect birds, failed to secrete cHtrA into host cytosol.


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)

Time course of cHtrA expression during C. trachomatis infection. The C. trachomatis-infected culture samples were processed at various times after infection (as indicated on the top) for immunofluorescence staining as described in Figure 1 legend. The mouse anti-cHtrA (a to h) and anti-CPAF (mAb 100a; i to p) were visualized with a goat anti-mouse IgG conjugated with Cy3 (red) while the chlamydial organisms were visualized with a rabbit anti-chlamydia antibody plus a goat anti-rabbit IgG-Cy2 conjugate (green). Note that cHtrA was first detected inside the chlamydial inclusions at 12 hours after infection [panel d, yellow (overlapping with organisms) & red (free of chlamydial organisms) arrowheads], similar to the detection of CPAF. However, cHtrA secretion into host cell cytosol was only detected 24 h after infection while secretion of CPAF was already obvious by 16 h post infection.
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Related In: Results  -  Collection

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Figure 5: Time course of cHtrA expression during C. trachomatis infection. The C. trachomatis-infected culture samples were processed at various times after infection (as indicated on the top) for immunofluorescence staining as described in Figure 1 legend. The mouse anti-cHtrA (a to h) and anti-CPAF (mAb 100a; i to p) were visualized with a goat anti-mouse IgG conjugated with Cy3 (red) while the chlamydial organisms were visualized with a rabbit anti-chlamydia antibody plus a goat anti-rabbit IgG-Cy2 conjugate (green). Note that cHtrA was first detected inside the chlamydial inclusions at 12 hours after infection [panel d, yellow (overlapping with organisms) & red (free of chlamydial organisms) arrowheads], similar to the detection of CPAF. However, cHtrA secretion into host cell cytosol was only detected 24 h after infection while secretion of CPAF was already obvious by 16 h post infection.
Mentions: We further used the specific anti-cHtrA antibodies to characterize the endogenous cHtrA. As shown in Figure 5, cHtrA protein was detected inside the inclusions as early as 12 h after infection and secretion of cHtrA into host cell cytosol became apparent by 24 h post infection. Although CPAF was also detectable at 12 h, the secretion of CPAF was more robust and became very obvious as early as 16 h after infection. The cHtrA protein was detected both within the chlamydial inclusions and in the host cell cytosol while CPAF mainly accumulated in the host cell cytosol as infection progressed. Although both CPAF and cHtrA are serine proteases secreted by C. trachomatis organisms, their distinct secretion kinetics and intracellular distribution patterns suggest that they may fulfill different functions during chlamydial infection. To further evaluate whether cHtrA secretion is common to all chlamydial organisms, we monitored the cHtrA protein distribution in cells infected with various serovars and strains from different chlamydial species, including 13 C. trachomatis serovars and also isolates representing species of C. muridarum, C. caviae, C. pneumoniae and C. psittaci (Figure 6). The cHtrA protein was consistently detected in both the lumen of chlamydial inclusion and cytosol of host cells infected with all serovars of C. trachomatis organisms and isolates of C. muridarum, C. caviae and C. pneumoniae but not C. psittaci. Although secretion of cHtrA into the inclusion lumen and further into the cytosol of the infected cells seems to be a common feature of most chlamydial organisms tested, it is not known at this moment why the species C. psittaci, which primarily infect birds, failed to secrete cHtrA into host cytosol.

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