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Prior infection with Type A Francisella tularensis antagonizes the pulmonary transcriptional response to an aerosolized Toll-like receptor 4 agonist.

ABSTRACT: Francisella infection attenuates immune cell infiltration and expression of selected pro-inflammatory cytokines in response to endogenous LPS, suggesting the bacteria is actively antagonizing at least some part of the response to Toll-like receptor 4 (TLR4) engagement. The ability of different Francisella strains to inhibit the ability of E. coli LPS to induce a pulmonary inflammatory response, as measured by gene expression profiling, was examined to define the scope of modulation and identify of inflammatory genes/pathways that are specifically antagonized by a virulent F. tularensis infection.Prior aerosol exposure to F. tularensis subsp. tularensis, but not the live attenuated strain (LVS) of F. tularensis subsp. holarctica or F. novicida, significantly antagonized the transcriptional response in the lungs of infected mice exposed to aerosolized E. coli LPS. The response to E. coli LPS was not completely inhibited, suggesting that the bacteria is targeting further downstream of the TLR4 molecule. Analysis of the promotors of LPS-responsive genes that were perturbed by Type A Francisella infection identified candidate transcription factors that were potentially modulated by the bacteria, including multiple members of the forkhead transcription factor family (FoxA1, Foxa2, FoxD1, Foxd3, Foxf2, FoxI1, Fox03, Foxq1), IRF1, CEBPA, and Mef2. The annotated functional roles of the affected genes suggested that virulent Francisella infection suppressed cellular processes including mRNA processing, antiviral responses, intracellular trafficking, and regulation of the actin cytoskeleton. Surprisingly, despite the broad overall suppression of LPS-induced genes by virulent Francisella, and contrary to what was anticipated from prior studies, Type A Francisella did not inhibit the expression of the majority of LPS-induced cytokines, nor the expression of many classic annotated inflammatory genes.Collectively, this analysis demonstrates clear differences in the ability of different Francisella strains to modulate TLR4 signaling and identifies genes/pathways that are specifically targeted by virulent Type A Francisella.

SUBMITTER: Walters KA

PROVIDER: S-EPMC4625460 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Prior infection with Type A Francisella tularensis antagonizes the pulmonary transcriptional response to an aerosolized Toll-like receptor 4 agonist.

Walters Kathie-Anne KA Olsufka Rachael R Kuestner Rolf E RE Wu Xiagang X Wang Kai K Skerrett Shawn J SJ Ozinsky Adrian A

BMC genomics 20151028

<h4>Background</h4>Francisella infection attenuates immune cell infiltration and expression of selected pro-inflammatory cytokines in response to endogenous LPS, suggesting the bacteria is actively antagonizing at least some part of the response to Toll-like receptor 4 (TLR4) engagement. The ability of different Francisella strains to inhibit the ability of E. coli LPS to induce a pulmonary inflammatory response, as measured by gene expression profiling, was examined to define the scope of modul ...[more]

PMID: 26510639

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Project description:The alarmone ppGpp is a critical regulator of virulence gene expression in Francisella tularensis In this intracellular pathogen, ppGpp is thought to work in concert with the putative DNA-binding protein PigR and the SspA protein family members MglA and SspA to control a common set of genes. MglA and SspA form a complex that interacts with RNA polymerase (RNAP), and PigR functions by interacting with the RNAP-associated MglA-SspA complex. Prior work suggested that ppGpp indirectly exerts its regulatory effects in F. tularensis by promoting the accumulation of polyphosphate in the cell, which in turn was required for formation of the MglA-SspA complex. Here we show that in Escherichia coli, neither polyphosphate nor ppGpp is required for formation of the MglA-SspA complex but that ppGpp promotes the interaction between PigR and the MglA-SspA complex. Moreover, we show that polyphosphate kinase, the enzyme responsible for the synthesis of polyphosphate, antagonizes virulence gene expression in F. tularensis, a finding that is inconsistent with the notion that polyphosphate accumulation promotes virulence gene expression in this organism. Our findings identify polyphosphate kinase as a novel negative regulator of virulence gene expression in F. tularensis and support a model in which ppGpp exerts its positive regulatory effects by promoting the interaction between PigR and the MglA-SspA complex.IMPORTANCE In Francisella tularensis, MglA and SspA form a complex that associates with RNA polymerase to positively control the expression of key virulence genes. The MglA-SspA complex works together with the putative DNA-binding protein PigR and the alarmone ppGpp. PigR functions by interacting directly with the MglA-SspA complex, but how ppGpp exerts its effects was unclear. Prior work indicated that ppGpp acts by promoting the accumulation of polyphosphate, which is required for MglA and SspA to interact. Here we show that formation of the MglA-SspA complex does not require polyphosphate. Furthermore, we find that polyphosphate antagonizes the expression of virulence genes in F. tularensis Thus, ppGpp does not promote virulence gene expression in this organism through an effect on polyphosphate.

Project description:Francisella tularensis is composed of a number of subspecies with varied geographic distribution, host ranges, and virulence. In view of these marked differences, comparative functional genomics may elucidate some of the molecular mechanism(s) behind these differences. In this study a shared probe microarray was designed that could be used to compare the transcriptomes of Francisella tularensis subsp. tularensis Schu S4 (Ftt), Francisella tularensis subsp. holarctica OR960246 (Fth), Francisella tularensis subsp. holarctica LVS (LVS), and Francisella novicida U112 (Fn). To gain insight into expression differences that may be related to the differences in virulence of these subspecies, transcriptomes were measured from each strain grown in vitro under identical conditions, utilizing a shared probe microarray. The human avirulent Fn strain exhibited high levels of transcription of genes involved in general metabolism, which are pseudogenes in the human virulent Ftt and Fth strains, consistent with the process of genome decay in the virulent strains. Genes encoding an efflux system (emrA2 cluster of genes), siderophore (fsl operon), acid phosphatase, LPS synthesis, polyamine synthesis, and citrulline ureidase were all highly expressed in Ftt when compared to Fn, suggesting that some of these may contribute to the relative high virulence of Ftt. Genes expressed at a higher level in Ftt when compared to the relatively less virulent Fth included genes encoding isochorismatases, cholylglycine hydrolase, polyamine synthesis, citrulline ureidase, Type IV pilus subunit, and the Francisella Pathogenicity Island protein PdpD. Fth and LVS had very few expression differences, consistent with the derivation of LVS from Fth. This study demonstrated that a shared probe microarray designed to detect transcripts in multiple species/subspecies of Francisella enabled comparative transcriptional analyses that may highlight critical differences that underlie the relative pathogenesis of these strains for humans. This strategy could be extended to other closely-related bacterial species for inter-strain and inter-species analyses.

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Prior infection with Type A Francisella tularensis antagonizes the pulmonary transcriptional response to an aerosolized Toll-like receptor 4 agonist.

Publications

Prior infection with Type A Francisella tularensis antagonizes the pulmonary transcriptional response to an aerosolized Toll-like receptor 4 agonist.

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