scholarly journals Localization of the hypothetical protein Cpn0585 in the inclusion membrane of Chlamydia pneumoniae-infected cells

2007 ◽  
Vol 42 (2-3) ◽  
pp. 111-116 ◽  
Author(s):  
Jianhua Luo ◽  
Tianjun Jia ◽  
Youmin Zhong ◽  
Ding Chen ◽  
Rhonda Flores ◽  
...  
Keyword(s):  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Inclusion Membrane ◽  
Infected Cells

scholarly journals Hypothetical Protein Cpn0308 Is Localized in the Chlamydia pneumoniae Inclusion Membrane

2006 ◽  
Vol 75 (1) ◽  
pp. 497-503 ◽  
Author(s):  
Jianhua Luo ◽  
Tianjun Jia ◽  
Rhonda Flores ◽  
Ding Chen ◽  
Guangming Zhong
Keyword(s):  
Membrane Protein ◽  
Fusion Proteins ◽  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Open Reading Frame ◽  
Inclusion Membrane ◽  
Reading Frame ◽  
Antibody Staining ◽  
Infected Cells ◽  
Antibody Labeling

ABSTRACT The hypothetical protein encoded by Chlamydia pneumoniae open reading frame cpn0308 was detected in inclusion membranes of C. pneumoniae-infected cells using antibodies raised with Cpn0308 fusion proteins. The anti-Cpn0308 antibodies did not cross-react with IncA, a known C. pneumoniae inclusion membrane protein, although the anti-Cpn0308 antibody staining overlapped with the anti-IncA antibody labeling. The labeling of the inclusion membrane by the anti-Cpn0308 antibody was specifically blocked by the Cpn0308 but not IncA fusion proteins. The Cpn0308 antigen was detectable 24 h after infection and remained in the inclusion membrane throughout the infection course.

scholarly journals Characterization of the hypothetical protein Cpn1027, a newly identified inclusion membrane protein unique to Chlamydia pneumoniae

Microbiology ◽  
2007 ◽  
Vol 153 (3) ◽  
pp. 777-786 ◽  
Author(s):  
R. Flores ◽  
J. Luo ◽  
D. Chen ◽  
G. Sturgeon ◽  
P. Shivshankar ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Inclusion Membrane

scholarly journals Characterization of Antiapoptotic Activities ofChlamydia pneumoniae in Human Cells

2001 ◽  
Vol 69 (11) ◽  
pp. 7121-7129 ◽  
Author(s):  
Silke F. Fischer ◽  
Claudia Schwarz ◽  
Juliane Vier ◽  
Georg Häcker
Keyword(s):  
Hela Cells ◽  
Chlamydia Pneumoniae ◽  
Death Receptor ◽  
Chronic Infections ◽  
Effector Caspase ◽  
Cellular Factors ◽  
Death Receptor Signaling ◽  
Infected Cells ◽  
External Stimuli ◽  
Bacterial Protein Synthesis

ABSTRACT Chlamydia pneumoniae is an obligate intracellular bacterium which frequently causes airway infection in humans and has been implicated in atherosclerosis. Here we show that infection withC. pneumoniae protects HeLa human epithelioid cells against apoptosis induced by external stimuli. In infected HeLa cells, apoptosis induced by staurosporine and CD95-death-receptor signaling was strongly reduced. Upon treatment with staurosporine, generation of effector caspase activity, processing of caspase-3 and caspase-9 and cytochrome c redistribution were all profoundly inhibited in cells infected with C. pneumoniae. Bacterial protein synthesis during early infection was required for this inhibition. Furthermore, cytochrome c-induced processing and activation of caspases were inhibited in cytosolic extracts from infected cells, suggesting that a C. pneumoniae-dependent antiapoptotic factor was generated in the cytosol upon infection. Infection with C. pneumoniaefailed to induce significant NF-κB activation in HeLa cells, indicating that no NF-κB-dependent cellular factors were involved in the protection against apoptosis. These results show that C. pneumoniae is capable of interfering with the host cell's apoptotic apparatus at probably at least two steps in signal transduction and might explain the propensity of these bacteria to cause chronic infections in humans.

Host α-adducin is redistributed and localized to the inclusion membrane in Chlamydia- and Chlamydophila-infected cells

Microbiology ◽  
2008 ◽  
Vol 154 (12) ◽  
pp. 3848-3855 ◽  
Author(s):  
Hencelyn G. Chu ◽  
Sara K. Weeks ◽  
Diana M. Gilligan ◽  
Daniel D. Rockey
Keyword(s):  
Inclusion Membrane ◽  
Infected Cells

scholarly journals Bacteriophage T4 Self-Assembly: Localization of gp3 and Its Role in Determining Tail Length

2000 ◽  
Vol 182 (3) ◽  
pp. 680-688 ◽  
Author(s):  
A. Vianelli ◽  
G. R. Wang ◽  
M. Gingery ◽  
R. L. Duda ◽  
F. A. Eiserling ◽  
...  
Keyword(s):  
Self Assembly ◽  
High Efficiency ◽  
Bacteriophage T4 ◽  
Tail Length ◽  
Hypothetical Protein ◽  
Protein Product ◽  
Amino Acid Sequences ◽  
Tail Assembly ◽  
Infected Cells ◽  
Assembly Intermediate

ABSTRACT Gene 3 of bacteriophage T4 participates at a late stage in the T4 tail assembly pathway, but the hypothetical protein product, gp3, has never been identified in extracts of infected cells or in any tail assembly intermediate. In order to overcome this difficulty, we expressed gp3 in a high-efficiency plasmid expression vector and subsequently purified it for further analysis. The N-terminal sequence of the purified protein showed that the initial methionine had been removed. Variant C-terminal amino acid sequences were resolved by determining the cysteine content of the protein. The molecular mass of 20.6 kDa for the pure protein was confirmed by Western blotting, using a specific anti-gp3 serum for which the purified protein was the immunogen. We also demonstrated, for the first time, the physical presence of gp3 in the mature T4 phage particle and localized it to the tail tube. By finding a nonleaky, nonpermissive host for a gene 3 mutant, we could clearly demonstrate a new phenotype: the slow, aberrant elongation of the tail tube in the absence of gp3.

scholarly journals Analytical Sensitivity, Reproducibility of Results, and Clinical Performance of Five PCR Assays for Detecting Chlamydia pneumoniae DNA in Peripheral Blood Mononuclear Cells

2000 ◽  
Vol 38 (7) ◽  
pp. 2622-2627 ◽  
Author(s):  
J. B. Mahony ◽  
S. Chong ◽  
B. K. Coombes ◽  
M. Smieja ◽  
A. Petrich
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Nested Pcr ◽  
Chlamydia Pneumoniae ◽  
Mononuclear Cells ◽  
Rrna Gene ◽  
Analytical Sensitivity ◽  
Infected Cells ◽  
Pcr Assays ◽  
The 16S Rrna Gene

Chlamydia pneumoniae has been associated with atherosclerosis and coronary artery disease (CAD), and its DNA has been detected in atheromatous lesions of the aorta, carotid, and coronary arteries by a variety of PCR assays. The objective of this study was to compare the performances of five published PCR assays in the detection of C. pneumoniae in peripheral blood mononuclear cells (PBMCs) from patients with coronary artery disease. The assays included two conventional PCRs, one targeting a cloned PstI fragment and one targeting the 16S rRNA gene; two nested PCRs, one targeting the 16S rRNA gene and one targeting ompA; and a touchdown enzyme time release (TETR) PCR, targeting the 16S rRNA gene. All PCRs had similar analytical sensitivities and detected a minimum of 0.005 inclusion-forming units (IFU) of C. pneumoniae; the ompA nested PCR and the TETR PCR were slightly more sensitive and detected 0.001 IFU. Assay reproducibility was examined by testing 10 replicates of C. pneumoniae DNA by each assay. All five assays showed excellent reproducibility at high levels of DNA, with scores of 10 out of 10 for 0.01 IFU, but exhibited decreased reproducibility for smaller numbers of C. pneumoniae IFU for all tests. Pairwise comparison of test results indicated that there was a significant difference between tests (Cochran Q = 32.0, P < 0.001), with thePstI fragment (P < 0.001) and 16S rRNA (P = 0.002) assays having lower reproducibility than the nested ompA and TETR assays. To further analyze assay sensitivity, C. pneumoniae-infected U-937 mononuclear cells were added to whole blood, and extracted mononuclear-cell DNA was tested by each assay. All five assays showed similar sensitivities, detecting 15 infected cells; three assays detected 3 infected cells, while all assays were negative at the next dilution (1.5 infected cells). A striking difference in performance of the five assays was seen, however, when PBMCs from CAD patients were tested for C. pneumoniae DNA. The ompA nested PCR detected C. pneumoniae DNA in 11 of 148 (7.4%) specimens, the 16S rRNA nested PCR detected 2 positives among the 148 specimens (1.4%) (P < 0.001), and the other 3 assays detected no positive specimens (P < 0.001, compared with theompA assay). These results indicate that analytical sensitivity alone does not predict the ability of an assay to detectC. pneumoniae in whole-blood-derived PBMCs. Before standardized assays can be used in wide-scale epidemiological studies, further characterization of these assays will be required to improve our understanding of their performance in the detection of C. pneumoniae in clinical material.

scholarly journals Inclusion Fluorescent-Antibody Test as a Screening Assay for Detection of Antibodies to Chlamydia pneumoniae

2002 ◽  
Vol 9 (3) ◽  
pp. 562-567
Author(s):  
Olga Tapia ◽  
Anatoly Slepenkin ◽  
Evgueni Sevrioukov ◽  
Kathi Hamor ◽  
Luis M. de la Maza ◽  
...  
Keyword(s):  
Screening Test ◽  
Chlamydia Pneumoniae ◽  
Fluorescent Antibody ◽  
Acute Infection ◽  
Serum Samples ◽  
Neutralization Titer ◽  
Screening Assay ◽  
Specific Test ◽  
Infected Cells

ABSTRACT A study was conducted to determine the ability of the inclusion immunofluorescence assay (inclusion IFA) to act as a screening test to detect samples with antibodies to Chlamydia pneumoniae; microimmunofluorescence (MIF) was used as the “gold standard.” In addition, the inclusion IFA was compared using HEp-2 cells infected with either C. pneumoniae CM-1 or Chlamydia trachomatis serovar E. A total of 331 serum samples representing a range of MIF titers were evaluated. The sensitivities of the inclusion IFA for detecting samples with C. pneumoniae MIF titers of ≥16 were 96.9 and 74.8% with C. pneumoniae- and C. trachomatis-infected cells, respectively. For samples with an elevated C. pneumoniae MIF titer of ≥512, the sensitivities of the C. pneumoniae- and C. trachomatis-based inclusion IFA were 97.0 and 8.8%, respectively. These results suggest that the inclusion IFA is not a genus-specific test, as evidenced by the failure of the C. trachomatis-infected cells to detect a significant number of samples with C. pneumoniae antibodies. Samples that had elevated C. pneumoniae inclusion IFA and MIF titers but that were found negative (titer, <16) by the C. trachomatis inclusion IFA were further tested by an in vitro neutralization assay for functional antibodies that might not have been detected by the serological assays. The in vitro neutralization results corroborated the serological results in that all seven sera tested had a neutralization titer for C. pneumoniae (range, 20 to 225), while all but one failed to have any effect on the infectivity of C. trachomatis serovar E. While the C. pneumoniae inclusion IFA had a high sensitivity for detecting chlamydial antibodies, depending on whether it was used as a screening test for detecting samples with low (≥16) or elevated (≥512) MIF titers, its specificity ranged from 53.4 to 77.1%. In conclusion, the inclusion IFA with C. pneumoniae-infected cells was best suited as a sensitive screening test for identifying specimens with elevated MIF titers (those associated with a possible acute infection with C. pneumoniae).

scholarly journals Inhibition of Fusion of Chlamydia trachomatis Inclusions at 32°C Correlates with Restricted Export of IncA

2002 ◽  
Vol 70 (7) ◽  
pp. 3816-3823 ◽  
Author(s):  
K. A. Fields ◽  
E. Fischer ◽  
T. Hackstadt
Keyword(s):  
Chlamydia Trachomatis ◽  
Hela Cells ◽  
Immunofluorescence Microscopy ◽  
Parasitophorous Vacuole ◽  
Obligate Intracellular Bacterium ◽  
Inclusion Membrane ◽  
Obligate Intracellular ◽  
Transmission Electron ◽  
Infected Cells ◽  
Exocytic Pathway

ABSTRACT Chlamydia trachomatis is an obligate intracellular bacterium that develops within a parasitophorous vacuole termed an inclusion. The inclusion is nonfusogenic with lysosomes but intercepts lipids from a host cell exocytic pathway. Initiation of chlamydial development is concurrent with modification of the inclusion membrane by a set of C. trachomatis-encoded proteins collectively designated Incs. One of these Incs, IncA, is functionally associated with the homotypic fusion of inclusions. Inclusions also do not fuse when cultures are multiply infected with C. trachomatis and cultivated at 32°C. We obtained evidence linking these experimental observations by characterizing IncA localization in 32°C cultures. Analysis of inclusions by light and transmission electron microscopy confirmed that HeLa cells infected with multiple C. trachomatis elementary bodies and cultivated at 32°C for 24 h contained multiple, independent inclusions. Reverse transcriptase PCR and immunoblot analyses of C. trachomatis-infected HeLa cells demonstrated the presence of IncA at 24 h in 32°C cultures. When parallel cultures were probed with IncA-specific antibodies in indirect immunofluorescence assays, IncA was detectable in intracellular chlamydiae but not within the inclusion membrane. In addition, analysis of purified reticulate bodies from 37 and 32°C cultures showed that bacterium-associated pools of IncA are enriched in cultures grown at 32°C. Microscopic observation of infected cells revealed that some vacuoles had fused by 48 h postinfection, and this finding was correlated with the detection of IncA in inclusion membranes by immunofluorescence microscopy. The data are consistent with a requirement for IncA in fusions of C. trachomatis inclusions and suggest that the effect of incubation at 32°C is manifested by restricted export of IncA to the inclusion membrane.

scholarly journals Characterization of hypothetical proteins Cpn0146, 0147, 0284 & 0285 that are predicted to be in the Chlamydia pneumoniae inclusion membrane

2007 ◽  
Vol 7 (1) ◽  
pp. 38 ◽  
Author(s):  
Jianhua Luo ◽  
Guangchao Liu ◽  
Youmin Zhong ◽  
Tianjun Jia ◽  
Kaiyang Liu ◽  
...  
Keyword(s):  
Chlamydia Pneumoniae ◽  
Hypothetical Proteins ◽  
Inclusion Membrane

scholarly journals Characterization of Fifty Putative Inclusion Membrane Proteins Encoded in the Chlamydia trachomatis Genome

2008 ◽  
Vol 76 (6) ◽  
pp. 2746-2757 ◽  
Author(s):  
Zhongyu Li ◽  
Chaoqun Chen ◽  
Ding Chen ◽  
Yimou Wu ◽  
Youmin Zhong ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Membrane Proteins ◽  
Chlamydial Infection ◽  
Distribution Patterns ◽  
Western Blot Assay ◽  
Inclusion Membrane ◽  
Urogenital Infection ◽  
Infected Cells ◽  
Inclusion Membrane Proteins

ABSTRACT Although the Chlamydia trachomatis genome is predicted to encode 50 inclusion membrane proteins, only 18 have been experimentally localized in the inclusion membrane of C. trachomatis-infected cells. Using fusion proteins and anti-fusion protein antibodies, we have systematically evaluated all 50 putative inclusion membrane proteins for their localization in the infected cells, distribution patterns, and effects on subsequent chlamydial infection when expressed ectopically, as well as their immunogenicity during chlamydial infection in humans. Twenty-two of the 50 proteins were localized in the inclusion membrane, and 7 were detected inside the inclusions, while the location of the remaining 21 was not defined. Four (CT225, CT228, CT358, and CT440) of the 22 inclusion membrane-localized proteins were visualized in the inclusion membrane of Chlamydia-infected cells for the first time in the current study. The seven intra-inclusion-localized proteins were confirmed to be chlamydial organism proteins in a Western blot assay. Further characterization of the 50 proteins revealed that neither colocalization with host cell endoplasmic reticulum nor inhibition of subsequent chlamydial infection by ectopically expressed proteins correlated with the inclusion membrane localization. Interestingly, antibodies from women with C. trachomatis urogenital infection preferentially recognized proteins localized in the inclusion membrane, and the immunodominant regions were further mapped to the region predicted to be on the cytoplasmic side of the inclusion membrane. These observations suggest that most of the inclusion membrane-localized proteins are both expressed and immunogenic during C. trachomatis infection in humans and that the cytoplasmic exposure may enhance the immunogenicity.

Sign in / Sign up

Export Citation Format

Share Document