Biology subjects

Brinkworth, A. J.

Publications and source records attributed to Brinkworth, A. J..

3 recordsLinked to original sources

bioRxiv2024

Chlamydia trachomatis modulates the expression of JAK-STAT signaling components to attenuate the Type II interferon response of epithelial cells

Chlamydia trachomatis has adapted to subvert signaling in epithelial cells to ensure successful intracellular development. Interferon-{gamma} (IFN{gamma}) produced by recruited lymphocytes signals through the JAK/STAT pathway to restrict chlamydial growth in the genital tract. However, during Chlamydia infection in vitro, addition of IFN{gamma} does not fully induce nuclear localization of its transcription factor STAT1 and target gene, IDO1. We hypothesize that this altered interferon response is a result of Chlamydia targeting components of the IFN{gamma}-JAK/STAT pathway. To assess the ability of replicating Chlamydia to dampen interferon signaling, HEp2 human epithelial cells were infected with C. trachomatis serovar L2 for 24 hours prior to exposure to physiologically relevant levels of IFN{gamma} (500 pg/mL). This novel approach enabled us to observe reduced phospho-activation of both STAT1 and its kinase Janus Kinase 2 (JAK2) in infected cells compared to mock-infected cells. Importantly, basal JAK2 and STAT1 transcript and protein levels were dampened by infection even in the absence of interferon, which could have implications for cytokine signaling beyond IFN{gamma}. Additionally, target genes IRF1, GBP1, APOL3, IDO1, and SOCS1 were not fully induced in response to IFN{gamma} exposure. Infection-dependent decreases in transcript, protein, and phosphoprotein were rescued when de novo bacterial protein synthesis was inhibited with chloramphenicol, restoring expression of IFN{gamma}-target genes. Similar Chlamydia-dependent dampening of STAT1 and JAK2 transcript levels were observed in infected END1 endocervical cells and in HEp2s infected with C. trachomatis serovar D, suggesting a conserved mechanism of dampening the interferon response by reducing the availability of key signaling components. ImportanceAs an obligate intracellular pathogen that has evolved to infect the genital epithelium, Chlamydia has developed strategies to prevent detection and antimicrobial signaling in its host to ensure its survival and spread. A major player in clearing Chlamydia infections is the inflammatory cytokine interferon-{gamma} (IFN{gamma}), which is produced by immune cells that are recruited to the site of infection. Reports of IFN{gamma} levels in vaginal and cervical swabs from Chlamydia-infected patients range from 1-350 pg/mL, while most in vitro studies of the effects of IFN{gamma} on chlamydial growth have used 15-85 fold higher concentrations. By using physiologically-relevant concentrations of IFN{gamma} we were able assess Chlamydias ability to modulate its signaling. We found that Chlamydia decreases the expression of multiple components that are required for inducing gene expression by IFN{gamma}, providing a possible mechanism by which C. trachomatis can attenuate the immune response in the female genital tract to cause long-term infections.

microbiology↗

bioRxiv2022

Pathogen-driven induction of a host transcriptome facilitating epithelial-to-mesenchymal transition

The obligate intracellular pathogen Chlamydia trachomatis is the causative agent of the most common bacterial sexually transmitted disease worldwide. While the host response to infection by this pathogen has been well characterized, it remains unclear to what extent host gene expression during infection is the product of Chlamydia-directed modulation of host transcription factors. In this report, we show the transcriptome of Chlamydia-infected epithelial cells exhibits gene expression consistent with activity of YAP, a transcriptional coactivator implicated in cell proliferation, organ morphogenesis, wound healing, and fibrosis. After confirming induction of YAP target genes during infection, we observed increased YAP nuclear translocation in Chlamydia-infected epithelial cells. We show that increased YAP activation is a Chlamydia-directed process occurring during midcycle infection; critically, this phenotype is sensitive to inhibition of protein synthesis by the pathogen. Infection-mediated YAP activation bypasses YAP inhibition by the Hippo kinase cascade, instead involving YAP tyrosine phospho-activation and the activity of host Src family kinases. Taken together, our results define a mechanism for Chlamydia-directed modulation of host gene expression independent of the host inflammatory response to infection, as well as introduce novel therapeutic targets for treatment of this pervasive disease.

cell biology↗

bioRxiv2017

Genome-wide transcriptional responses of iron-starved Chlamydia trachomatis reveal prioritization of metabolic precursor synthesis over protein translation

Iron is essential for growth and development of Chlamydia. Its long-term starvation in cultured mammalian cells leads to production of aberrant non-infectious chlamydial forms, also known as persistence. Immediate transcriptional responses to iron limitation have not been characterized, leaving a knowledge gap of how Chlamydia regulates its response to changes in iron availability. We used the fast-chelating agent 2,2-Bipyridyl (BPDL) to homogeneously starve Chlamydia trachomatis serovar L2 of iron, starting at 6 or 12h post-infection. Immediate transcriptional responses were monitored after only 3 or 6h of BPDL-treatment, well before formation of aberrant Chlamydia. The first genome-wide transcriptional response of C. trachomatis to iron-starvation was subsequently determined utilizing RNA-sequencing. Only 7% and 8% of the genome was differentially expressed in response to iron-starvation at early and mid-stages of development, respectively. Biological pathway analysis revealed an overarching theme. Synthesis of macromolecular precursors (deoxynucleotides, amino acids, charged tRNAs, and acetyl-coA) was up-regulated, while energy-expensive processes (ABC transport and translation) were down-regulated. A large fraction of differentially down-regulated genes are involved in translation, including ribosome assembly, initiation and termination factors, which could be analogous to the translation down-regulation triggered by stress in other prokaryotes during stringent responses. Additionally, transcriptional up-regulation of DNA repair, oxidative stress, and tryptophan salvage genes reveals a possible coordination of responses to multiple antimicrobial and immunological insults. These responses of replicative-phase Chlamydia to iron-starvation indicate a prioritization of survival over replication, enabling the pathogen to \"stock the pantry\" with ingredients needed for rapid growth once optimal iron levels are restored.\n\nIMPORTANCEBy utilizing an experimental approach that monitors the immediate global response of Chlamydia trachomatis to iron-starvation, clues to long-standing questions in Chlamydia biology are revealed, including how Chlamydia adapts to this stress. We determined that this pathogen initiates a transcriptional program that prioritizes replenishment of nutrient stores over replication, possibly in preparation for rapid growth once optimal iron levels are restored. Transcription of genes for biosynthesis of metabolic precursors was generally up-regulated, while those involved in multiple steps of translation were down-regulated. We also observed an increase in transcription of genes involved in DNA repair and neutralizing oxidative stress, indicating that Chlamydia employs an \"all-or-nothing\" strategy. Its small genome limits its ability to tailor a specific response to a particular stress. Therefore, the \"all-or-nothing\" strategy may be the most efficient way of surviving within the host, where the pathogen likely encounters multiple simultaneous immunological and nutritional insults.

microbiology↗