Chlamydia trachomatis
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PROVIDER: PRJNA171054 | ENA |
REPOSITORIES: ENA
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Project description:The obligate intracellular developmental cycle of Chlamydia trachomatis presents significant challenges in defining its proteome. In this study we have applied quantitative proteomics to both the intracellular reticulate body (RB) and the extracellular elementary body (EB) from C. trachomatis. We used C. trachomatis L2 which is a model chlamydial isolate for such a study since it has a high infectivity: particle ratio and there is an excellent quality genome sequence. EBs and RBs (>99% pure) were quantified by chromosomal and plasmid copy number using PCR to determine the concentrations of chlamydial proteins per bacterial cell. RBs harvested at 15h post infection (PI) were purified by three successive rounds of gradient centrifugation. This is the earliest possible time to obtain purified RBs, free from host cell components in quantity, within the constraints of the technology, EBs were purified at 48h PI. We then used two-dimensional reverse phase UPLC to fractionate RB or EB peptides before mass spectroscopic analysis, providing absolute amount estimates of chlamydial proteins.
2016-06-28 | PXD003025 | Pride
Project description:By comprehensive quantitative proteome analysis we characterize the three growth forms elementary body (EB), reticulate body (RB) and aberrant reticulate body (ARB) of Chlamydia trachomatis genital strain D/UW-3/CX
2017-01-04 | PXD003883 | Pride
Project description:The obligate intracellular bacterium Chlamydia has a unique developmental cycle that alternates between two contrasting cell types. With a hardy envelope and highly condensed genome, the small elementary body (EB) maintains limited metabolic activities yet can survive in an extracellular environment and is infectious. After entering host cells, EBs differentiate into larger and proliferating reticulate bodies (RBs). Progeny EBs are derived from RBs in late developmental stages and eventually exit host cells. How the expression of the chlamydial genome consisting of nearly 1000 genes governs the chlamydial developmental cycle is unclear. A previous microarray study identified only 29 immediately early genes, defined as genes activated by the first hour postinoculation, in C. trachomatis. By performing RNA sequencing analysis for C. trachomatis cultures with high multiplicities of infection (i.e., MOI of 50 and 200), we observed that 730 C. trachomatis genes underwent 2- to 900-fold activation within one hour postinoculation. By conducting quantitative reverse transcription real-time PCR (qRT-PCR) analysis for 48 of the 730 genes using an MOI of 1, we confirmed the expression increases in 46 genes. Our results demonstrate that the immediate early transcriptome is tens of times more extensive than previously realized. Gene ontology analysis indicates that the activation spans across all functional categories. We conclude that a supermajority of the C. trachomatis genes are activated almost immediately after EBs are inside host cells to initiate the differentiation toward RBs and to establish an intracellular niche conducive for chlamydial development and growth. RNA-Seq analysis was performed for Chlamydia trachomatis L2
2024-01-30 | GSE248988 | GEO
Project description:The obligate intracellular human pathogen Chlamydia pneumoniae was subjected to dRNA-Seq to gain insights into the transcriptome. The two distinct life cycle forms elementary bodies (EB) and reticulate bodies (RB) were isolated from human Hep2 cell line by differential gradient centrifugation.
2011-08-18 | GSE24999 | GEO
Project description:The obligate intracellular human pathogen Chlamydia pneumoniae was subjected to dRNA-Seq to gain insights into the transcriptome. The two distinct life cycle forms elementary bodies (EB) and reticulate bodies (RB) were isolated from human Hep2 cell line by differential gradient centrifugation. Total RNA was isolated and partially treated with Terminator Exonuclease to digest RNA without 5'-PPP and thereby enrich for native 5' ends.
2011-08-18 | E-GEOD-24999 | biostudies-arrayexpress
Project description:Bacteria in the chlamydiales order are obligate intracellular parasites of eukaryotic cells. Within this order, the genus Chlamydia contains the causative agents of a number of clinically important infections of humans. Biovars of C. trachomatis are the causative agents of trachoma, the leading cause of preventable blindness worldwide, as well as sexually transmitted infections with the potential to cause pelvic inflammatory disease and infertility. Irrespective of the resulting disease, all chlamydial species share the same obligate intracellular life cycle and developmental cell forms. They are reliant on an infectious cycle consisting of at least three phenotypically distinct cell forms termed the reticulate body (RB), the intermediate body (IB) and the elementary body (EB). The EB is infectious but does not replicate. The RB replicates in the host cell but is non-infectious, while the IB is an intermediate form that transitions to the EB form. In this study, we ectopically expressed the transcriptional repressor Euo, the two nucleoid-associated proteins HctA and HctB, and the two component sensor kinase CtcB in the RB. Transcriptional analysis using RNA-seq, differential expression clustering and fluorescence in situ hybridization analysis show that the chlamydial developmental cycle is driven by three distinct regulons corresponding to the RB, IB or EB cell forms. Moreover, we show that the genes for the T3SS were cell type restricted, suggesting defined functional roles for the T3SS in specific cell forms.
2025-01-26 | GSE287626 | GEO
Project description:Chlamydia is an obligate intracellular bacteria that undergoes a complex biphasic developmental cycle, alternating between the smaller infectious non-dividing elementary body (EB) and the larger non-infectious but dividing reticulate body (RB). Due to the differences between these functionally and morphologically distinct forms, we hypothesize protein degradation is essential to chlamydial differentiation. The bacterial Clp system, consisting of an ATPase unfoldase (e.g., ClpX or ClpC) and a proteolytic component (e.g., ClpP), is critical for the physiology of bacteria through its recognition, and usually degradation, of specific substrates. We observed by transmission electron microscopy that overexpression of wild-type ClpC, but not an ATPase mutant isoform, in Chlamydia increased glycogen accumulation within the vacuolar niche of the bacteria earlier in the developmental cycle than typically observed. This suggested ClpC activity may increase expression of EB-associated genes. Consistent with this, targeted RT-qPCR analyses demonstrated a significant increase in several EB-associated gene transcripts earlier in development. These effects were not observed with overexpression of the ATPase mutant of ClpC, providing strong evidence that the activity of ClpC drives secondary differentiation. By analyzing the global transcriptional response to ClpC overexpression using RNA sequencing, we observed a global shift to earlier expression of canonical late developmental cycle genes and other EB-associated genes. Finally, we directly linked overexpression of ClpC with earlier production of EBs. Conversely, disrupting normal ClpC function with an ATPase mutant caused a delay in secondary differentiation. Overall, these findings provide the first mechanistic insight for initiation of secondary differentiation in Chlamydia.
2024-11-12 | GSE255503 | GEO
Project description:Chlamydia trachomatis is the most common sexually transmitted infection and the bacterial agent of trachoma globally. C. trachomatis undergoes a biphasic developmental cycle involving an infectious elementary body and a replicative reticulate body. Little is currently known about the expression of host cell mRNAs, lncRNAs, and miRNAs at different stages of C. trachomatis development. Here, we performed RNA-seq and miR-seq on HeLa cells infected with C. trachomatis serovar E at 20 hpi and 44 hpi with or without IFN-γ treatment. Our study identified and validated differentially expressed host cell mRNAs, lncRNAs, and miRNAs during infection. Infection at 20 hpi showed the most differential upregulation of both coding and non-coding genes while infection at 44 hpi in the presence of IFN-γ resulted in a dramatic downregulation of a large proportion of genes. Using RT-qPCR, we validated the top 5 stage-specific upregulated mRNAs and miRNAs. One of the commonly expressed miRNAs at all three stages, miR-193b-5p, showed significant expression in clinical serum samples of C. trachomatis-infected patients as compared to sera from healthy controls and HIV-1-infected patients. Furthermore, at 20 hpi we observed significant upregulation of antigen processing and presentation, and T helper cell differentiation pathways whereas T cell receptor, mTOR, and Rap1 pathways were modulated at 44 hpi. Treatment with IFN-γ at 44 hpi showed the regulation of cytokine-cytokine receptor interaction, FoxO signaling, and Ras signaling pathways. Our study documents a role for the stage-specific transcriptional manipulation of the host cell genome and important signaling pathways that are necessary for the survival of pathogen and could serve as potential biomarkers in the diagnosis and management of the disease.
2021-01-13 | GSE158814 | GEO
Project description:Chlamydia trachomatis is a significant human pathogen yet their obligate intracellular nature severe restrictions upon research. Chlamydiae undergo a complex developmental cycle characterized by an infectious cell type known as the elementary body (EB) and an intracellular active replicative form called the reticulate body (RB). EBs have historically been described as metabolically dormant. A cell-free (axenic) culture system was developed which showed high levels of metabolic and biosynthetic activity from both EBs and RBs. EBs preferentially utilized glucose-6-phosphate as an energy source whereas RBs required ATP. Both developmental forms showed improved activity when incubated under microaerobic conditions. Incorporation of isotopically-labeled amino acids into proteins from both developmental forms indicated unique expression profiles which were confirmed by genome-wide transcriptional analysis. The described axenic culture system will greatly enhance biochemical and physiological analyses of chlamydiae. Chlamydia axenic metabolic activity
2012-11-20 | E-GEOD-39530 | biostudies-arrayexpress
Project description:The pathogenic bacterium Chlamydia reproduces via two specialized forms inside a eukaryotic host cell. The dividing form called the reticulate body (RB) must convert at late times into the infectious elementary body (EB) for spread to a new host cell. Late genes are a temporal class of chlamydial genes believed to be responsible for RB-to-EB conversion, but late gene regulation is incompletely understood. In this study, we used chromatin immunoprecipitation (ChIP) to investigate two alternative sigma factors, σ28 and σ54, that alter the promoter specificity of C. trachomatis RNA-polymerase. σ28 ChIP-seq identified hctB and tsp as the only promoters bound by σ28, and binding only occurred late, around the time of RB-to-EB conversion. A σ28 overexpression strain confirmed that these genes are transcribed in a σ28-dependent manner. σ54 ChIP-seq showed that σ54 only bound ctl0021 and ctl0052, and only at late times. This σ54 regulon appears to be conserved as in silico analysis identified σ54 promoter sequences upstream of ctl0021 and ctl0052 homologs in all Chlamydia spp. The genes encoding σ28 and σ54 were only transcribed at late times, but ChIP analysis with the late regulator Euo showed that Euo only controls σ28 expression, and late transcription of σ54 is regulated in an Euo-dependent manner. Thus, multiple mechanisms regulate late genes, including Euo and different forms of RNA polymerase. The dedicated use of two alternative RNA polymerases to control just four late genes suggests that these genes and the independent control of their temporal expression are important for RB-to-EB conversion.
2025-04-30 | GSE294424 | GEO