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Mitochondrial transcription factor A serves as a danger signal by augmenting plasmacytoid dendritic cell responses to DNA.

ABSTRACT: Plasmacytoid dendritic cells (pDC) are potent APCs known to regulate immune responses to self-Ags, particularly DNA. The mitochondrial fraction of necrotic cells was found to most potently promote human pDC activation, as reflected by type I IFN release, which was dependent upon the presence of mitochondrial DNA and involved TLR9 and receptors for advanced glycation end products. Mitochondrial transcription factor A (TFAM), a highly abundant mitochondrial protein that is functionally and structurally homologous to high mobility group box protein 1, was observed to synergize with CpG-containing oligonucleotide, type A, DNA to promote human pDC activation. pDC type I IFN responses to TFAM and CpG-containing oligonucleotide, type A, DNA indicated their engagement with receptors for advanced glycation end products and TLR9, respectively, and were dependent upon endosomal processing and PI3K, ERK, and NF-?B signaling. Taken together, these results indicate that pDC contribute to sterile immune responses by recognizing the mitochondrial component of necrotic cells and further incriminate TFAM and mitochondrial DNA as likely mediators of pDC activation under these circumstances.

SUBMITTER: Julian MW

PROVIDER: S-EPMC3381894 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

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Mitochondrial transcription factor A serves as a danger signal by augmenting plasmacytoid dendritic cell responses to DNA.

Julian Mark W MW Shao Guohong G Bao Shengying S Knoell Daren L DL Papenfuss Tracey L TL VanGundy Zachary C ZC Crouser Elliott D ED

Journal of immunology (Baltimore, Md. : 1950) 20120606 1

Plasmacytoid dendritic cells (pDC) are potent APCs known to regulate immune responses to self-Ags, particularly DNA. The mitochondrial fraction of necrotic cells was found to most potently promote human pDC activation, as reflected by type I IFN release, which was dependent upon the presence of mitochondrial DNA and involved TLR9 and receptors for advanced glycation end products. Mitochondrial transcription factor A (TFAM), a highly abundant mitochondrial protein that is functionally and structu ...[more]

PMID: 22675199

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Project description:ObjectiveMitochondrial transcription factor A (TFAM) is normally bound to and remains associated with mitochondrial DNA (mtDNA) when released from damaged cells. We hypothesized that TFAM, bound to mtDNA (or equivalent CpG-enriched DNA), amplifies TNF? release from TLR9-expressing plasmacytoid dendritic cells (pDCs) by engaging RAGE.Materials and methodsMurine Flt3 ligand-expanded splenocytes obtained from C57BL/6 mice were treated with recombinant human TFAM, alone or in combination with CpG-enriched DNA with subsequent TNF? release measured by ELISA. The role of RAGE was determined by pre-treatment with soluble RAGE or heparin or by employing matching RAGE (-/-) splenocytes. TLR9 signaling was evaluated using a specific TLR9-blocking oligonucleotide and by inhibiting endosomal processing, PI3K and NF-?B. Additional studies examined whether heparin sulfate moieties or endothelin converting enzyme-1 (ECE-1)-dependent recycling of endosomal receptors were required for TFAM and CpG DNA recognition.Main resultsTFAM augmented splenocyte TNF? release in response to CpGA DNA, which was strongly dependent upon pDCs and regulated by RAGE and TLR9 receptors. Putative TLR9 signaling pathways, including endosomal acidification and signaling through PI3K and NF-?B, were essential for splenocyte TNF? release in response to TFAM+CpGA DNA. Interestingly, TNF? release depended upon endothelin converting enzyme (ECE)-1, which cleaves and presumably activates TLR9 within endosomes. Recognition of the TFAM-CpGA DNA complex was dependent upon heparin sulfate moieties, and recombinant TFAM Box 1 and Box 2 proteins were equivalent in terms of augmenting TNF? release.ConclusionsTFAM promoted TNF? release in a splenocyte culture model representing complex cell-cell interactions in vivo with pDCs playing a critical role. To our knowledge, this study is the first to incriminate ECE-1-dependent endosomal cleavage of TLR9 as a critical step in the signaling pathway leading to TNF? release. These findings, and others reported herein, significantly advance our understanding of sterile immune responses triggered by mitochondrial danger signals.

Project description:BackgroundTranscription factors (TFs) control gene expression by direct binding to regulatory regions of target genes but also by impacting chromatin landscapes and modulating DNA accessibility for other TFs. In recent years several TFs have been defined that control cell fate decisions and effector functions in the immune system. Plasmacytoid dendritic cells (pDCs) are an immune cell type with the unique capacity to produce high amounts of type I interferons quickly in response to contact with viral components. Hereby, this cell type is involved in anti-infectious immune responses but also in the development of inflammatory and autoimmune diseases. To date, the global TF reservoir in pDCs early after activation remains to be fully characterized.ResultsTo fill this gap, we have performed a comprehensive analysis in naïve versus TLR9-activated murine pDCs in a time course study covering early timepoints after stimulation (2 h, 6 h, 12 h) integrating gene expression (RNA-Seq) and chromatin landscape (ATAC-Seq) studies. To unravel the biological processes underlying the changes in TF expression on a global scale gene ontology (GO) analyses were performed. We found that 70% of all genes annotated as TFs in the mouse genome (1014 out of 1636) are expressed in pDCs for at least one stimulation time point and are covering a wide range of TF classes defined by their specific DNA binding mechanisms. GO analysis revealed involvement of TLR9-induced TFs in epigenetic modulation, NFκB and JAK-STAT signaling, and protein production in the endoplasmic reticulum. pDC activation predominantly "turned on" the chromatin regions associated with TF genes. Our in silico analyses pointed at the AP-1 family of TFs as less noticed but possibly important players in these cells after activation. AP-1 family members exhibit (1) increased gene expression, (2) enhanced chromatin accessibility in their promoter region, and (3) a TF DNA binding motif that is globally enriched in genomic regions that were found more accessible in pDCs after TLR9 activation.ConclusionsIn this study we define the complete set of TLR9-regulated TFs in pDCs. Further, this study identifies the AP-1 family of TFs as potentially important but so far less well characterized regulators of pDC function.

Project description:BACKGROUND:Atopy and viral respiratory tract infections synergistically promote asthma exacerbations. IgE cross-linking inhibits critical virus-induced IFN-? responses of plasmacytoid dendritic cells (pDCs), which can be deficient in patients with allergic asthma. OBJECTIVE:We sought to determine whether reducing IgE levels in vivo with omalizumab treatment increases pDC antiviral IFN-? responses in inner-city children with asthma. METHODS:PBMCs and pDCs isolated from children with exacerbation-prone asthma before and during omalizumab treatment were stimulated ex vivo with rhinovirus and influenza in the presence or absence of IgE cross-linking. IFN-? levels were measured in supernatants, and mRNA expression of IFN-? pathway genes was determined by using quantitative RT-PCR (qRT-PCR) in cell pellets. Fc?RI? protein levels and mRNA expression were measured in unstimulated cells by using flow cytometry and qRT-PCR, respectively. Changes in these outcomes and associations with clinical outcomes were analyzed, and statistical modeling was used to identify risk factors for asthma exacerbations. RESULTS:Omalizumab treatment increased rhinovirus- and influenza-induced PBMC and rhinovirus-induced pDC IFN-? responses in the presence of IgE cross-linking and reduced pDC surface Fc?RI? expression. Omalizumab-induced reductions in pDC Fc?RI? levels were significantly associated with a lower asthma exacerbation rate during the outcome period and correlated with increases in PBMC IFN-? responses. PBMC Fc?RI? mRNA expression measured on study entry significantly improved an existing model of exacerbation prediction. CONCLUSIONS:These findings indicate that omalizumab treatment augments pDC IFN-? responses and attenuates pDC Fc?RI? protein expression and provide evidence that these effects are related. These results support a potential mechanism underlying clinical observations that allergic sensitization is associated with increased susceptibility to virus-induced asthma exacerbations.

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Mitochondrial transcription factor A serves as a danger signal by augmenting plasmacytoid dendritic cell responses to DNA.

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Mitochondrial transcription factor A serves as a danger signal by augmenting plasmacytoid dendritic cell responses to DNA.

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