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Impaired chromatin remodelling at STAT1-regulated promoters leads to global unresponsiveness of Toxoplasma gondii-infected macrophages to IFN-?.

ABSTRACT: Intracellular pathogens including the apicomplexan and opportunistic parasite Toxoplasma gondii profoundly modify their host cells in order to establish infection. We have shown previously that intracellular T. gondii inhibit up-regulation of regulatory and effector functions in murine macrophages (M?) stimulated with interferon (IFN)-?, which is the cytokine crucial for controlling the parasites' replication. Using genome-wide transcriptome analysis we show herein that infection with T. gondii leads to global unresponsiveness of murine macrophages to IFN-?. More than 61% and 89% of the transcripts, which were induced or repressed by IFN-? in non-infected M?, respectively, were not altered after stimulation of T. gondii-infected cells with IFN-?. These genes are involved in a variety of biological processes, which are mostly but not exclusively related to immune responses. Analyses of the underlying mechanisms revealed that IFN-?-triggered nuclear translocation of STAT1 still occurred in Toxoplasma-infected M?. However, STAT1 bound aberrantly to oligonucleotides containing the IFN-?-responsive gamma-activated site (GAS) consensus sequence. Conversely, IFN-? did not induce formation of active GAS-STAT1 complexes in nuclear extracts from infected M?. Mass spectrometry of protein complexes bound to GAS oligonucleotides showed that T. gondii-infected M? are unable to recruit non-muscle actin to IFN-?-responsive DNA sequences, which appeared to be independent of stimulation with IFN-? and of STAT1 binding. IFN-?-induced recruitment of BRG-1 and acetylation of core histones at the IFN-?-regulated CIITA promoter IV, but not ?-actin was diminished by >90% in Toxoplasma-infected M? as compared to non-infected control cells. Remarkably, treatment with histone deacetylase inhibitors restored the ability of infected macrophages to express the IFN-? regulated genes H2-A/E and CIITA. Taken together, these results indicate that Toxoplasma-infected M? are unable to respond to IFN-? due to disturbed chromatin remodelling, but can be rescued using histone deacetylase inhibitors.

SUBMITTER: Lang C

PROVIDER: S-EPMC3262016 | biostudies-literature | 2012 Jan

REPOSITORIES: biostudies-literature

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Impaired chromatin remodelling at STAT1-regulated promoters leads to global unresponsiveness of Toxoplasma gondii-infected macrophages to IFN-γ.

Lang Christine C Hildebrandt Anke A Brand Franziska F Opitz Lennart L Dihazi Hassan H Lüder Carsten G K CG

PLoS pathogens 20120119 1

Intracellular pathogens including the apicomplexan and opportunistic parasite Toxoplasma gondii profoundly modify their host cells in order to establish infection. We have shown previously that intracellular T. gondii inhibit up-regulation of regulatory and effector functions in murine macrophages (MΦ) stimulated with interferon (IFN)-γ, which is the cytokine crucial for controlling the parasites' replication. Using genome-wide transcriptome analysis we show herein that infection with T. gondii ...[more]

PMID: 22275866

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Project description:Intracellular pathogens including the apicomplexan and opportunistic parasite Toxoplasma gondii profoundly modify their host cells in order to establish infection. We have shown previously that intracellular T. gondii inhibit up-regulation of regulatory and effector functions in murine macrophages (MΦ) stimulated with interferon (IFN)-γ, which is the cytokine crucial for controlling the parasites’ replication. Using genome-wide transcriptome analysis we show herein that infection with T. gondii leads to global unresponsiveness of murine macrophages to IFN-γ. More than 61% and 89% of the transcripts, which were induced or repressed by IFN-γ in non-infected MΦ, respectively, were not altered after stimulation of T. gondii-infected cells with IFN-γ. These genes are involved in a variety of biological processes, which are mostly but not exclusively related to immune responses. Analyses of the underlying mechanisms revealed that IFN-γ-triggered nuclear translocation of STAT1 still occurred in Toxoplasma-infected MΦ. However, STAT1 bound aberrantly to oligonucleotides containing the IFN-γ-responsive gamma-activated site (GAS) consensus sequence. Conversely, IFN-γ did not induce formation of active GAS-STAT1 complexes in nuclear extracts from infected MΦ. Mass spectrometry of protein complexes bound to GAS oligonucleotides showed that T. gondii-infected MΦ are unable to recruit non-muscle actin to IFN-γ-responsive DNA sequences, which appeared to be independent of stimulation with IFN-γ and of STAT1 binding. IFN-γ-induced recruitment of BRG-1 and acetylation of core histones at the IFN-γ-regulated CIITA promoter IV, but not β-actin was diminished by >90% in Toxoplasma-infected MΦ as compared to non-infected control cells. Remarkably, treatment with histone deacetylase inhibitors restored the ability of infected macrophages to express the IFN-γ regulated genes H2-A/E and CIITA. Taken together, these results indicate that Toxoplasma-infected MΦ are unable to respond to IFN-γ due to disturbed chromatin remodelling, but can be rescued using histone deacetylase inhibitors. Comparison of 4 different RNA pools with a 2-Color-Loop Design including 10 microarrays: [1] T. gondii infected and IFN-gamma treated, [2] T. gondii infected and untreated, [3] Non-infected and IFN-gamma treated, and [4] Non-infected and untreated.

Project description:Toxoplasma gondii (T. gondii) is a parasite infecting animals and humans. In intermediate hosts, such as humans or rodents, rapidly replicating tachyzoites drive vigorous innate and adaptive immune responses resulting in bradyzoites that survive within tissue cysts. Activation of the innate immune system is critical during the early phase of infection to limit pathogen growth and to instruct parasite-specific adaptive immunity. In rodents, dendritic cells (DCs) sense T. gondii through TLR11/12, leading to IL-12 production, which activates NK cells to produce IFN-γ as an essential mechanism for early parasite control. Further, C3 can bind to T. gondii resulting in limited complement activation. Here, we determined the role of C5a/C5aR1 axis activation for the early innate immune response in a mouse model of peritoneal T. gondii infection. We found that C5ar1 -/- animals suffered from significantly higher weight loss, disease severity, mortality, and parasite burden in the brain than wild type control animals. Severe infection in C5ar1 -/- mice was associated with diminished serum concentrations of IL-12, IL-27, and IFN-γ. Importantly, the serum levels of pro-inflammatory cytokines, including IL-1α, IL-6, and TNF-α, as well as several CXC and CC chemokines, were decreased in comparison to wt animals, whereas anti-inflammatory IL-10 was elevated. The defect in IFN-γ production was associated with diminished Ifng mRNA expression in the spleen and the brain, reduced frequency of IFN-γ+ NK cells in the spleen, and decreased Nos2 expression in the brain of C5ar1 -/- mice. Mechanistically, DCs from the spleen of C5ar1 -/- mice produced significantly less IL-12 in response to soluble tachyzoite antigen (STAg) stimulation in vivo and in vitro. Our findings suggest a model in which the C5a/C5aR1 axis promotes IL-12 induction in splenic DCs that is critical for IFN-γ production from NK cells and subsequent iNOS expression in the brain as a critical mechanism to control acute T. gondii infection.

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Impaired chromatin remodelling at STAT1-regulated promoters leads to global unresponsiveness of Toxoplasma gondii-infected macrophages to IFN-?.

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Impaired chromatin remodelling at STAT1-regulated promoters leads to global unresponsiveness of Toxoplasma gondii-infected macrophages to IFN-γ.

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