RNAseq of 2'3' cGAMP injected Drosophila melanogaster
Ontology highlight
ABSTRACT:
ORGANISM(S): Drosophila melanogaster
PROVIDER: GSE140955 | GEO | 2020/05/31
REPOSITORIES: GEO
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Project description:The accumulation of DNA in the cytosol serves as a key immunostimulatory signal associated with infections, cancer and genomic damage. Cytosolic DNA triggers immune responses by activating the cGAS/STING pathway. The binding of DNA to the cytosolic enzyme cGAMP synthase (cGAS), activates its enzymatic activity, leading to the synthesis of a second messenger, cyclic[G(2’,5’)pA(3’,5’)] (2’3’-cGAMP). 2’3’-cGAMP, a cyclic dinucleotide (CDN), activates the protein ‘stimulator of interferon genes’ (STING), which in turn activates the transcription factors IRF3 and NF-κB promoting the transcription of genes encoding type I interferons and other cytokines and mediators that stimulate a broader immune response. Exogenous 2’3’-cGAMP produced by malignant cells and other CDNs, including CDNs produced by bacteria and synthetic CDNs used in cancer immunotherapy, must traverse the cell membrane to activate STING in target cells. How these charged CDNs pass through the lipid bilayer is unknown. Here we used a genome-wide CRISPR interference screen to identify the reduced folate carrier SLC19A1, a folate-organic phosphate antiporter, as the major transporter for CDNs. CDN uptake and functional responses are inhibited by depleting SLC19A1 from human cells and enhanced by overexpressing SLC19A1. In both human cell lines and primary cells ex vivo, CDN uptake is inhibited by folates, as well as two medications approved for treatment of inflammatory diseases, sulfasalazine and the antifolate methotrexate. The identification of SLC19A1 as the major transporter of CDNs into cells has implications for the immunotherapeutic treatment of cancer, host responsiveness to CDN-producing pathogenic microorganisms, and potentially in certain inflammatory diseases.
2019-09-01 | GSE134371 | GEO
Project description:To reconstruct the ancestry of animal innate immunity, we have developed the choanoflagellate Monosiga brevicollis, one of the closest living relatives of animals, as a model for studying mechanisms underlying pathogen recognition and immune response. We found that M. brevicollis is killed by exposure to Pseudomonas aeruginosa bacteria and selectively avoids ingesting them. Moreover, M. brevicollis expresses STING, which, in animals, activates innate immune pathways in response to cyclic dinucleotides during pathogen sensing. M. brevicollis STING increases the susceptibility of M. brevicollis to P. aeruginosa-induced cell death and is required for responding to the cyclic dinucleotide 2’3’ cGAMP. Furthermore, similar to animals, autophagic signaling in M. brevicollis is induced by 2’3’ cGAMP in a STING-dependent manner.
2021-05-13 | GSE174340 | GEO
Project description:The volume-regulated anion channel (VRAC) is a hexameric complex formed by LRRC8 proteins. VRACs are responsible for the regulatory volume decrease (RVD) after hypotonic cell swelling by mediating the efflux of chloride. Besides chloride, LRRC8 proteins transport other molecules including immunomodulatory cyclic dinucleotides (CDNs) such as 2’3’cGAMP. Here we identify LRRC8C as a critical component of VRACs in T cells, where its deletion abolishes VRAC currents and RVD. We find that LRRC8C mediates the transport of 2’3’cGAMP in T cells. T cells of Lrrc8c-/- mice have increased cell cycle progression, proliferation, survival, Ca2+ influx and cytokine production in vitro and enhanced T cell mediated immunity in vivo. We used RNA sequencing of CD4+ T cells from wild-type (WT) and Lrrc8c-/- mice to determine the effects of impaired cGAMP signaling in T cells. T cells of Lrrc8c-/- mice had impaired p53 signaling which was associated with decreased p53 expression. We found that uptake of 2’3’cGAMP and other CDNs via LRRC8C results in STING activation and p53 accumulation. Inhibition of STING in WT T cells recapitulates the phenotype of LRRC8C-deficient T cells whereas overexpression of p53 inhibits enhanced T cell function in the absence of LRRC8C. These findings establish cGAMP uptake through LRRC8C and subsequent STING-p53 signaling as a novel inhibitory signaling pathway in T cells and adaptive immunity.
2021-12-31 | GSE163679 | GEO
Project description:DNA sensing is a fundamental process in the immune system, including host defence against viruses. The DNA sensor cGAS synthesises 2’3’ cyclic GMP-AMP (cGAMP), a second messenger that activates STING, which subsequently induces innate immunity. cGAMP not only activates STING in the cell where it is produced but also transfers to other cells. Transporters, channels and pores including SLC19A1, the SLC46A family, P2X7, ABCC1 and volume-regulated anion channels (VRACs) release cGAMP into the extracellular space and/or import cGAMP into cells. Emerging evidence suggests these proteins are important in antiviral immunity. Here, we investigated whether viruses antagonise cGAMP transporters, channels and pores. We report that infection with multiple human viruses depleted cGAMP conduits from cells. This included herpes simplex virus 1 (HSV-1) that targeted the VRAC subunits LRRC8A and LRRC8C, as well as SLC46A2 and P2X7, for degradation. The HSV-1 protein UL56 was required and sufficient for these effects that were mediated at least partially by proteasomal turnover. UL56 thereby inhibited the cGAMP uptake via VRAC, SLC46A2 and P2X7. Taken together, we show that HSV-1 actively antagonises cGAMP transfer across the plasma membrane and propose this limits innate immunity by reducing cell-to-cell communication via the immunotransmitter cGAMP.
2024-04-15 | PXD043229 | Pride
Project description:In mammals, the cGAS-cGAMP-STING pathway is crucial for sensing viral infection and initiating an anti-viral type I interferon response. cGAS and STING are highly conserved genes that originated in bacteria and are present in most animals. By contrast, interferons only emerged in vertebrates; thus, the function of STING in invertebrates is unclear. Here, we use the STING ligand 2'3'-cGAMP to activate immune responses in a model cnidarian invertebrate, the starlet sea anemone Nematostella vectensis. Using RNA-Seq, we found that 2'3'-cGAMP induces robust transcription of both anti-viral and anti-bacterial genes, including the conserved transcription factor NF-κB. Knockdown experiments identified a role for NF-κB in specifically inducing anti-bacterial genes downstream of 2'3'-cGAMP, and some of these genes were also found to be induced during Pseudomonas aeruginosa infection. Furthermore, we characterized the protein product of one of the putative anti-bacterial genes, the N. vectensis homolog of Dae4, and found that it has conserved anti-bacterial activity. This work describes an unexpected role of a cGAMP sensing pathway in anti-bacterial immunity and suggests that a broad transcriptional response is an evolutionarily ancestral output of 2'3'-cGAMP signaling in animals.
2021-06-03 | GSE175984 | GEO
Project description:We report positional cloning and characterization of a novel Sting allele that fails to activate IFN production in the wild-derived mice of MOLF/Ei strain in response to HSV (Herpes Simplex Virus) and Listeria monocytogenes both in vitro and in vivo. We show that previously uncharacterized mutations in the N-terminal of STING are responsible for low levels of IFN due to failure of MOLF STING to respond to cytosolic STING agonists such as 2’3’cGAMP, 5,6-Dimethylxanthenone-4-acetic acid (DMXAA) or poly(deoxyadenylic-deoxythymidylic) acid (dAdT). Using Next-Generation Sequencing (NGS) for the analysis of DNA-responses in congenic C57BL6.StingMOLF/MOLF (MOLF STING) mouse macrophages, we show that these mutations in MOLF/Ei discriminate in responses between different STING agonists. Macrophages from C57BL6.StingB6/B6¬ (B6 STING), or B6 STING expressing littermates were stimulated as a positive control for STING activation, and STING -/- (STING KO) macrophages served as a negative control.
2016-01-03 | GSE68140 | GEO
Project description:We report positional cloning and characterization of a novel Sting allele that fails to activate IFN production in the wild-derived mice of MOLF/Ei strain in response to HSV (Herpes Simplex Virus) and Listeria monocytogenes both in vitro and in vivo. We show that previously uncharacterized mutations in the N-terminal of STING are responsible for low levels of IFN due to failure of MOLF STING to respond to cytosolic STING agonists such as 2â3âcGAMP, 5,6-Dimethylxanthenone-4-acetic acid (DMXAA) or poly(deoxyadenylic-deoxythymidylic) acid (dAdT). Using Next-Generation Sequencing (NGS) for the analysis of DNA-responses in congenic C57BL6.StingMOLF/MOLF (MOLF STING) mouse macrophages, we show that these mutations in MOLF/Ei discriminate in responses between different STING agonists. Macrophages from C57BL6.StingB6/B6¬ (B6 STING), or B6 STING expressing littermates were stimulated as a positive control for STING activation, and STING -/- (STING KO) macrophages served as a negative control. To examine differential gene regulation downstream of murine Tmem173 (STING) allelic variants. We stimulated macrophages from B6 mice congenic for MOLF STING; macrophages from B6 STING expressing littermates were stimulated as a positive control for STING activation, and STING -/- (STING KO) macrophages served as a negative control. One replicate of RNA-seq reads after each stimulation provided. Conditions are peritoneal macrophages from B6 congenic mice expressing: B6 STING, MOLF STING, or STING KO stimulated with 2â3âcGAMP, 5,6-Dimethylxanthenone-4-acetic acid (DMXAA), poly(deoxyadenylic-deoxythymidylic) acid (dAdT), or no treatment. This constitutes a total of 12 reads analyzed in this data set.
2016-07-05 | E-GEOD-68140 | biostudies-arrayexpress
Project description:RNAseq of 2'3' cGAMP injected Drosophila melanogaster
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Project description:ENPP1 expression correlates with poor prognosis in many cancers, and we previously discovered that ENPP1 is the dominant hydrolase of extracellular cGAMP: a cancer-cell-produced immunotransmitter that activates the anticancer STING pathway. However, ENPP1 has other catalytic activities and the molecular and cellular mechanisms contributing to its tumorigenic effects remain unclear. Here, using single cell RNA-seq (scRNA-seq), we show that ENPP1 overexpression drives primary tumor growth and metastasis by synergistically dampening extracellular cGAMP-STING mediated antitumoral immunity and activating immunosuppressive extracellular adenosine (eADO) signaling. In addition to cancer cells, stromal and immune cells in the tumor microenvironment (TME) also express ENPP1 which functions as a gate keeper to block cGAMP sensing in these cells. Enpp1 knockout in both cancer cells and normal tissues slowed primary tumor growth and prevented metastasis in an extracellular cGAMP- and STING-dependent manner. Lastly, an ENPP1 knock-in mutation that selectively abolishes ENPP1’s cGAMP hydrolysis activity phenocopied total ENPP1 knockout, demonstrating that restoration of paracrine cGAMP-STING signaling is the dominant anti-cancer mechanism of ENPP1 inhibition. In summary, selectively blocking ENPP1’s cGAMP hydrolysis activity is a promising therapeutic approach for treating cancer.
2023-08-07 | GSE233659 | GEO
Project description:STING R284S variant is constitutively active and induces inflammatory genes even in absence of STING ligand cGAMP.
2019-02-27 | GSE111196 | GEO