RESUMO
ATP, NAD+, and nucleic acids are abundant purines that, in addition to having critical intracellular functions, have evolved extracellular roles as danger signals released in response to cell lysis, apoptosis, degranulation, or membrane pore formation. In general ATP and NAD+ have excitatory and adenosine has anti-inflammatory effects on immune cells. This review focuses on recent advances in our understanding of purine release mechanisms, ectoenzymes that metabolize purines (CD38, CD39, CD73, ENPP1, and ENPP2/autotaxin), and signaling by key P2 purinergic receptors (P2X7, P2Y2, and P2Y12). In addition to metabolizing ATP or NAD+, some purinergic ectoenzymes metabolize other inflammatory modulators, notably lysophosphatidic acid and cyclic GMP-AMP (cGAMP). Also discussed are extracellular signaling effects of NAD+ mediated by ADP-ribosylation, and epigenetic effects of intracellular adenosine mediated by modification of S-adenosylmethionine-dependent DNA methylation.
Assuntos
Inflamação/imunologia , Purinas/metabolismo , Receptores Purinérgicos/metabolismo , ADP-Ribosilação , Trifosfato de Adenosina/metabolismo , Animais , Metilação de DNA , Humanos , Inflamação/genética , Inflamação/metabolismo , Lisofosfolipídeos/metabolismo , Transdução de SinaisRESUMO
Cytoplasmic accumulation of TDP-43 is a disease hallmark for many cases of amyotrophic lateral sclerosis (ALS), associated with a neuroinflammatory cytokine profile related to upregulation of nuclear factor κB (NF-κB) and type I interferon (IFN) pathways. Here we show that this inflammation is driven by the cytoplasmic DNA sensor cyclic guanosine monophosphate (GMP)-AMP synthase (cGAS) when TDP-43 invades mitochondria and releases DNA via the permeability transition pore. Pharmacologic inhibition or genetic deletion of cGAS and its downstream signaling partner STING prevents upregulation of NF-κB and type I IFN induced by TDP-43 in induced pluripotent stem cell (iPSC)-derived motor neurons and in TDP-43 mutant mice. Finally, we document elevated levels of the specific cGAS signaling metabolite cGAMP in spinal cord samples from patients, which may be a biomarker of mtDNA release and cGAS/STING activation in ALS. Our results identify mtDNA release and cGAS/STING activation as critical determinants of TDP-43-associated pathology and demonstrate the potential for targeting this pathway in ALS.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , DNA Mitocondrial/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas de Membrana/metabolismo , Poro de Transição de Permeabilidade Mitocondrial/metabolismo , Nucleotidiltransferases/metabolismo , Alarminas/metabolismo , Esclerose Lateral Amiotrófica/patologia , Animais , Citoplasma/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Células HEK293 , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Inflamação/metabolismo , Interferon Tipo I/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , NF-kappa B/metabolismo , Degeneração Neural/patologia , Fosfotransferases (Aceptor do Grupo Álcool) , Subunidades Proteicas/metabolismo , Transdução de SinaisRESUMO
How the central innate immune protein, STING, is activated by its ligands remains unknown. Here, using structural biology and biochemistry, we report that the metazoan second messenger 2'3'-cGAMP induces closing of the human STING homodimer and release of the STING C-terminal tail, which exposes a polymerization interface on the STING dimer and leads to the formation of disulfide-linked polymers via cysteine residue 148. Disease-causing hyperactive STING mutations either flank C148 and depend on disulfide formation or reside in the C-terminal tail binding site and cause constitutive C-terminal tail release and polymerization. Finally, bacterial cyclic-di-GMP induces an alternative active STING conformation, activates STING in a cooperative manner, and acts as a partial antagonist of 2'3'-cGAMP signaling. Our insights explain the tight control of STING signaling given varying background activation signals and provide a therapeutic hypothesis for autoimmune syndrome treatment.
Assuntos
Proteínas de Membrana/metabolismo , Sítios de Ligação , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , Dimerização , Retículo Endoplasmático/metabolismo , Células HEK293 , Humanos , Ligantes , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Modelos Moleculares , Mutagênese Sítio-Dirigida , Nucleotídeos Cíclicos/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Transdução de SinaisRESUMO
The cGAS-STING intracellular DNA-sensing pathway has emerged as a key element of innate antiviral immunity and a promising therapeutic target. The existence of an innate immune sensor that can be activated by any double-stranded DNA (dsDNA) of any origin raises fundamental questions about how cGAS is regulated and how it responds to "foreign" DNA while maintaining tolerance to ubiquitous self-DNA. In this review, we summarize recent evidence implicating important roles for cGAS in the detection of foreign and self-DNA. We describe two recent and surprising insights into cGAS-STING biology: that cGAS is tightly tethered to the nucleosome and that the cGAMP product of cGAS is an immunotransmitter acting at a distance to control innate immunity. We consider how these advances influence our understanding of the emerging roles of cGAS in the DNA damage response (DDR), senescence, aging, and cancer biology. Finally, we describe emerging approaches to harness cGAS-STING biology for therapeutic benefit.
Assuntos
Nucleotidiltransferases , Transdução de Sinais , Nucleotidiltransferases/metabolismo , Imunidade Inata , DNARESUMO
Tissues are exposed to diverse inflammatory challenges that shape future inflammatory responses. While cellular metabolism regulates immune function, how metabolism programs and stabilizes immune states within tissues and tunes susceptibility to inflammation is poorly understood. Here, we describe an innate immune metabolic switch that programs long-term intestinal tolerance. Intestinal interleukin-18 (IL-18) stimulation elicited tolerogenic macrophages by preventing their proinflammatory glycolytic polarization via metabolic reprogramming to fatty acid oxidation (FAO). FAO reprogramming was triggered by IL-18 activation of SLC12A3 (NCC), leading to sodium influx, release of mitochondrial DNA, and activation of stimulator of interferon genes (STING). FAO was maintained in macrophages by a bistable switch that encoded memory of IL-18 stimulation and by intercellular positive feedback that sustained the production of macrophage-derived 2'3'-cyclic GMP-AMP (cGAMP) and epithelial-derived IL-18. Thus, a tissue-reinforced metabolic switch encodes durable immune tolerance in the gut and may enable reconstructing compromised immune tolerance in chronic inflammation.
RESUMO
The innate immune system functions as the first line of defense against invading bacteria and viruses. In this context, the cGAS/STING [cyclic guanosine monophosphate (GMP)-adenosine monophosphate (AMP) synthase/STING] signaling axis perceives the nonself DNA associated with bacterial and viral infections, as well as the leakage of self DNA by cellular dysfunction and stresses, to elicit the host's immune responses. In this pathway, the noncanonical cyclic dinucleotide 2',3'-cyclic GMP-AMP (2',3'-cGAMP) functions as a second messenger for signal transduction: 2',3'-cGAMP is produced by the enzyme cGAS upon its recognition of double-stranded DNA, and then the 2',3'-cGAMP is recognized by the receptor STING to induce the phosphorylation of downstream factors, including TBK1 (TANK binding kinase 1) and IRF3 (interferon regulatory factor 3). Numerous crystal structures of the components of this cGAS/STING signaling axis have been reported and these clarify the structural basis for their signal transduction mechanisms. In this review, we summarize recent progress made in the structural dissection of this signaling pathway and indicate possible directions of forthcoming research.
Assuntos
DNA/imunologia , Imunidade Inata , Nucleotídeos Cíclicos/imunologia , Nucleotidiltransferases/imunologia , Sistemas do Segundo Mensageiro/imunologia , Animais , Bactérias , Cristalografia por Raios X , Citosol/química , Citosol/imunologia , DNA/química , DNA/genética , Regulação da Expressão Gênica , Humanos , Fator Regulador 3 de Interferon/química , Fator Regulador 3 de Interferon/genética , Fator Regulador 3 de Interferon/imunologia , Modelos Moleculares , Nucleotídeos Cíclicos/química , Nucleotídeos Cíclicos/genética , Nucleotidiltransferases/química , Nucleotidiltransferases/genética , Fosforilação , Conformação Proteica , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Sistemas do Segundo Mensageiro/genéticaRESUMO
Lipid metabolism has been associated with the cyclic guanosine monophosphate (GMP)-AMP synthase (cGAS) stimulator of interferon genes (STING) DNA-sensing pathway, but our understanding of how these signals are integrated into a cohesive immunometabolic program is lacking. Here, we have identified liver X receptor (LXR) agonists as potent inhibitors of STING signaling. We show that stimulation of lipid metabolism by LXR agonists specifically suppressed cyclic GMP-AMP (cGAMP)-STING signaling. Moreover, we developed cyclic dinucleotide-conjugated beads to biochemically isolate host effectors for cGAMP inhibition, and we found that LXR ligands stimulated the expression of sphingomyelin phosphodiesterase acid-like 3A (SMPDL3A), which is a 2'3'-cGAMP-degrading enzyme. Results of crystal structures suggest that cGAMP analog induces dimerization of SMPDL3A, and the dimerization is critical for cGAMP degradation. Additionally, we have provided evidence that SMPDL3A cleaves cGAMP to restrict STING signaling in cell culture and mouse models. Our results reveal SMPDL3A as a cGAMP-specific nuclease and demonstrate a mechanism for how LXR-associated lipid metabolism modulates STING-mediated innate immunity.
Assuntos
Metabolismo dos Lipídeos , Nucleotidiltransferases , Animais , Camundongos , Receptores X do Fígado/metabolismo , Nucleotidiltransferases/metabolismo , DNA , Nucleotídeos Cíclicos/metabolismo , Imunidade InataRESUMO
In mammals, the enzyme cGAS senses the presence of cytosolic DNA and synthesizes the cyclic dinucleotide (CDN) 2'3'-cGAMP, which triggers STING-dependent immunity. In Drosophila melanogaster, two cGAS-like receptors (cGLRs) produce 3'2'-cGAMP and 2'3'-cGAMP to activate STING. We explored CDN-mediated immunity in 14 Drosophila species covering 50 million years of evolution and found that 2'3'-cGAMP and 3'2'-cGAMP failed to control infection by Drosophila C virus in D. serrata and two other species. We discovered diverse CDNs produced in a cGLR-dependent manner in response to viral infection in D. melanogaster, including 2'3'-c-di-GMP. This CDN was a more potent STING agonist than cGAMP in D. melanogaster and it also activated a strong antiviral transcriptional response in D. serrata. Our results shed light on the evolution of cGLRs in flies and provide a basis for understanding the function and regulation of this emerging family of pattern recognition receptors in animal innate immunity.
Assuntos
Antivirais , Drosophila , Animais , Drosophila melanogaster , GMP Cíclico , MamíferosRESUMO
The DNA sensor cyclic GMP-AMP synthase (cGAS) is important for antiviral and anti-tumor immunity. cGAS generates cyclic GMP-AMP (cGAMP), a diffusible cyclic dinucleotide that activates the antiviral response through the adaptor protein stimulator of interferon genes (STING). cGAMP cannot passively cross cell membranes, but recent advances have established a role for extracellular cGAMP as an "immunotransmitter" that can be imported into cells. However, the mechanism by which cGAMP exits cells remains unknown. Here, we identifed ABCC1 as a direct, ATP-dependent cGAMP exporter in mouse and human cells. We show that ABCC1 overexpression enhanced cGAMP export and limited STING signaling and that loss of ABCC1 reduced cGAMP export and potentiated STING signaling. We demonstrate that ABCC1 deficiency exacerbated cGAS-dependent autoimmunity in the Trex1-/- mouse model of Aicardi-Goutières syndrome. Thus, ABCC1-mediated cGAMP export is a key regulatory mechanism that limits cell-intrinsic activation of STING and ameliorates STING-dependent autoimmune disease.
Assuntos
Proteínas de Membrana Transportadoras , Proteínas Associadas à Resistência a Múltiplos Medicamentos , Nucleotídeos Cíclicos , Trifosfato de Adenosina , Animais , DNA/metabolismo , Humanos , Interferons/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Camundongos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Nucleotídeos Cíclicos/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismoRESUMO
2',3'-cGAMP, produced by the DNA sensor cGAS, activates stimulator of interferon genes (STING) and triggers immune response during infection. Tremendous effort has been placed on unraveling the mechanism of STING activation. However, little is known about STING inhibition. Here, we found that apo-STING exhibits a bilayer with head-to-head as well as side-by-side packing, mediated by its ligand-binding domain (LBD). This type of assembly holds two endoplasmic reticulum (ER) membranes together not only to prevent STING ER exit but also to eliminate the recruitment of TBK1, representing the autoinhibited state of STING. Additionally, we obtained the filament structure of the STING/2',3'-cGAMP complex, which adopts a bent monolayer assembly mediated by LBD and transmembrane domain (TMD). The active, curved STING polymer could deform ER membrane to support its ER exit and anterograde transportation. Our data together provide a panoramic vision regarding STING autoinhibition and activation, which adds substantially to current understanding of the cGAS-STING pathway.
Assuntos
Proteínas Serina-Treonina Quinases , Transdução de Sinais , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , DNA , Imunidade InataRESUMO
The enzyme cyclic GMP-AMP synthase (cGAS) senses cytosolic DNA in infected and malignant cells and catalyzes the formation of 2'3'cGMP-AMP (cGAMP), which in turn triggers interferon (IFN) production via the STING pathway. Here, we examined the contribution of anion channels to cGAMP transfer and anti-viral defense. A candidate screen revealed that inhibition of volume-regulated anion channels (VRACs) increased propagation of the DNA virus HSV-1 but not the RNA virus VSV. Chemical blockade or genetic ablation of LRRC8A/SWELL1, a VRAC subunit, resulted in defective IFN responses to HSV-1. Biochemical and electrophysiological analyses revealed that LRRC8A/LRRC8E-containing VRACs transport cGAMP and cyclic dinucleotides across the plasma membrane. Enhancing VRAC activity by hypotonic cell swelling, cisplatin, GTPγS, or the cytokines TNF or interleukin-1 increased STING-dependent IFN response to extracellular but not intracellular cGAMP. Lrrc8e-/- mice exhibited impaired IFN responses and compromised immunity to HSV-1. Our findings suggest that cell-to-cell transmission of cGAMP via LRRC8/VRAC channels is central to effective anti-viral immunity.
Assuntos
Fibroblastos/imunologia , Interferons/imunologia , Proteínas de Membrana/imunologia , Nucleotídeos Cíclicos/imunologia , Canais de Ânion Dependentes de Voltagem/imunologia , Animais , Antivirais/imunologia , Antivirais/metabolismo , Efeito Espectador , Linhagem Celular , Células Cultivadas , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Células HeLa , Herpes Simples/imunologia , Herpes Simples/virologia , Herpesvirus Humano 1/imunologia , Herpesvirus Humano 1/fisiologia , Humanos , Interferons/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nucleotídeos Cíclicos/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/imunologia , Nucleotidiltransferases/metabolismo , Canais de Ânion Dependentes de Voltagem/metabolismoRESUMO
Interferon-inducible human oligoadenylate synthetase-like (OASL) and its mouse ortholog, Oasl2, enhance RNA-sensor RIG-I-mediated type I interferon (IFN) induction and inhibit RNA virus replication. Here, we show that OASL and Oasl2 have the opposite effect in the context of DNA virus infection. In Oasl2-/- mice and OASL-deficient human cells, DNA viruses such as vaccinia, herpes simplex, and adenovirus induced increased IFN production, which resulted in reduced virus replication and pathology. Correspondingly, ectopic expression of OASL in human cells inhibited IFN induction through the cGAS-STING DNA-sensing pathway. cGAS was necessary for the reduced DNA virus replication observed in OASL-deficient cells. OASL directly and specifically bound to cGAS independently of double-stranded DNA, resulting in a non-competitive inhibition of the second messenger cyclic GMP-AMP production. Our findings define distinct mechanisms by which OASL differentially regulates host IFN responses during RNA and DNA virus infection and identify OASL as a negative-feedback regulator of cGAS.
Assuntos
2',5'-Oligoadenilato Sintetase/metabolismo , Infecções por Vírus de DNA/imunologia , Vírus de DNA/fisiologia , Infecções por Vírus de RNA/imunologia , Vírus de RNA/imunologia , 2',5'-Oligoadenilato Sintetase/genética , Animais , AMP Cíclico/metabolismo , Humanos , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nucleotidiltransferases/metabolismo , RNA Interferente Pequeno/genética , Transdução de Sinais , Células THP-1 , Replicação ViralRESUMO
Extracellular 2'3'-cyclic-GMP-AMP (cGAMP) is an immunotransmitter exported by diseased cells and imported into host cells to activate the innate immune STING pathway. We previously identified SLC19A1 as a cGAMP importer, but its use across human cell lines is limited. Here, we identify LRRC8A heteromeric channels, better known as volume-regulated anion channels (VRAC), as widely expressed cGAMP transporters. LRRC8A forms complexes with LRRC8C and/or LRRC8E, depending on their expression levels, to transport cGAMP and other 2'3'-cyclic dinucleotides. In contrast, LRRC8D inhibits cGAMP transport. We demonstrate that cGAMP is effluxed or influxed via LRRC8 channels, as dictated by the cGAMP electrochemical gradient. Activation of LRRC8A channels, which can occur under diverse stresses, strongly potentiates cGAMP transport. We identify activator sphingosine 1-phosphate and inhibitor DCPIB as chemical tools to manipulate channel-mediated cGAMP transport. Finally, LRRC8A channels are key cGAMP transporters in resting primary human vasculature cells and universal human cGAMP transporters when activated.
Assuntos
Sistemas CRISPR-Cas , Proteínas de Membrana/metabolismo , Nucleotídeos Cíclicos/metabolismo , Transporte Biológico , Ciclopentanos/farmacologia , Humanos , Indanos/farmacologia , Lisofosfolipídeos/farmacologia , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Esfingosina/análogos & derivados , Esfingosina/farmacologia , Células U937RESUMO
Detection of cytosolic DNA by the enzyme cGAS triggers the production of cGAMP, a second messenger that binds and activates the adaptor protein STING, which leads to interferon (IFN) production. Here, we found that in vivo natural killer (NK) cell killing of tumor cells, but not of normal cells, depends on STING expression in non-tumor cells. Experiments using transplantable tumor models in STING- and cGAS-deficient mice revealed that cGAS expression by tumor cells was critical for tumor rejection by NK cells. In contrast, cGAS expression by host cells was dispensable, suggesting that tumor-derived cGAMP is transferred to non-tumor cells, where it activates STING. cGAMP administration triggered STING activation and IFN-ß production in myeloid cells and B cells but not NK cells. Our results reveal that the anti-tumor response of NK cells critically depends on the cytosolic DNA sensing pathway, similar to its role in defense against pathogens, and identify tumor-derived cGAMP as a major determinant of tumor immunogenicity with implications for cancer immunotherapy.
Assuntos
Interferons/imunologia , Células Matadoras Naturais/imunologia , Proteínas de Membrana/imunologia , Neoplasias/imunologia , Nucleotídeos Cíclicos/imunologia , Animais , Linhagem Celular , Linhagem Celular Tumoral , Regulação da Expressão Gênica/imunologia , Humanos , Interferons/metabolismo , Células Matadoras Naturais/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neoplasias/genética , Neoplasias/metabolismo , Nucleotídeos Cíclicos/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/imunologia , Nucleotidiltransferases/metabolismo , Transdução de Sinais/imunologiaRESUMO
2'3'-cyclic-GMP-AMP (cGAMP) is a second messenger that activates the antiviral stimulator of interferon genes (STING) pathway. We recently identified a novel role for cGAMP as a soluble, extracellular immunotransmitter that is produced and secreted by cancer cells. Secreted cGAMP is then sensed by host cells, eliciting an antitumoral immune response. Due to the antitumoral effects of cGAMP, other CDN-based STING agonists are currently under investigation in clinical trials for metastatic solid tumors. However, it is unknown how cGAMP and other CDNs cross the cell membrane to activate intracellular STING. Using a genome-wide CRISPR screen, we identified SLC19A1 as the first known importer of cGAMP and other CDNs, including the investigational new drug 2'3'-bisphosphosphothioate-cyclic-di-AMP (2'3'-CDAS). These discoveries will provide insight into cGAMP's role as an immunotransmitter and aid in the development of more targeted CDN-based cancer therapeutics.
Assuntos
Imunidade Inata/genética , Neoplasias/genética , Nucleotídeos Cíclicos/genética , Proteína Carregadora de Folato Reduzido/genética , Sistemas CRISPR-Cas/genética , Linhagem Celular Tumoral , Membrana Celular/genética , Genoma Humano/genética , Humanos , Proteínas de Membrana/genética , Neoplasias/imunologia , Nucleotídeos Cíclicos/imunologia , Transdução de Sinais/genéticaRESUMO
Upon activation by double-stranded DNA (dsDNA), the cytosolic dsDNA sensor cyclic GMP-AMP synthase (cGAS) synthesizes the diffusible cyclic dinucleotide 2'3'-cGAMP (cyclic GMP-AMP), which subsequently binds to the adaptor STING, triggering a cascade of events leading to an inflammatory response. Recent studies have highlighted the role of 2'3'-cGAMP as an 'immunotransmitter' between cells, a process facilitated by gap junctions as well as by specialized membrane-spanning importer and exporter channels. This review highlights recent advances from a structural perspective of intercellular trafficking of 2'3'-cGAMP, with particular emphasis on the binding of importer SLC19A1 to 2'3'-cGAMP, as well as the significance of associated folate nutrients and antifolate therapeutics. This provides a path forward for structure-guided understanding of the transport cycle in immunology, as well as for candidate targeting approaches towards therapeutic intervention in inflammation.
Assuntos
Proteínas de Membrana , Nucleotidiltransferases , Humanos , Inflamação , Proteínas de Membrana/metabolismo , Nucleotídeos Cíclicos/metabolismo , Nucleotidiltransferases/metabolismoRESUMO
Ectonucleotide pyrophosphatase/phosphodiesterase 1 (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 stimulator of interferon genes (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, we show that ENPP1 in both cancer and normal tissues drives primary breast tumor growth and metastasis by dampening extracellular 2'3'-cyclic-GMP-AMP (cGAMP)-STING-mediated antitumoral immunity. ENPP1 loss-of-function in both cancer cells and normal tissues slowed primary tumor growth and abolished metastasis. Selectively abolishing the cGAMP hydrolysis activity of ENPP1 phenocopied ENPP1 knockout in a STING-dependent manner, demonstrating that restoration of paracrine cGAMP-STING signaling is the dominant anti-cancer mechanism of ENPP1 inhibition. Finally, ENPP1 expression in breast tumors deterministically predicated whether patients would remain free of distant metastasis after pembrolizumab (anti-PD-1) treatment followed by surgery. Altogether, ENPP1 blockade represents a strategy to exploit cancer-produced extracellular cGAMP for controlled local activation of STING and is therefore a promising therapeutic approach against breast cancer.
Assuntos
Neoplasias da Mama , Feminino , Humanos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Imunidade Inata , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Diester Fosfórico Hidrolases/genética , Diester Fosfórico Hidrolases/metabolismo , Pirofosfatases/metabolismoRESUMO
Viruses represent a major threat to all animals, which defend themselves through induction of a large set of virus-stimulated genes that collectively control the infection. In vertebrates, these genes include interferons that play a critical role in the amplification of the response to infection. Virus- and interferon-stimulated genes include restriction factors targeting the different steps of the viral replication cycle, in addition to molecules associated with inflammation and adaptive immunity. Predictably, antiviral genes evolve dynamically in response to viral pressure. As a result, each animal has a unique arsenal of antiviral genes. Here, we exploit the capacity to experimentally activate the evolutionarily conserved stimulator of IFN genes (STING) signaling pathway by injection of the cyclic dinucleotide 2'3'-cyclic guanosine monophosphate-adenosine monophosphate into flies to define the repertoire of STING-regulated genes in 10 Drosophila species, spanning 40 million years of evolution. Our data reveal a set of conserved STING-regulated factors, including STING itself, a cGAS-like-receptor, the restriction factor pastel, and the antiviral protein Vago, but also 2 key components of the antiviral RNA interference pathway, Dicer-2, and Argonaute2. In addition, we identify unknown species- or lineage-specific genes that have not been previously associated with resistance to viruses. Our data provide insight into the core antiviral response in Drosophila flies and pave the way for the characterization of previously unknown antiviral effectors.
Assuntos
Drosophila , Imunidade Inata , Animais , Nucleotídeos Cíclicos , Antivirais/farmacologiaRESUMO
The cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway is central for the initiation of anti-tumoural immune responses. Enormous effort has been made to optimise the design and administration of STING agonists to stimulate tumour immunogenicity. However, in certain contexts the cGAS-STING axis fuels tumourigenesis. Here, we review recent findings on the regulation of cGAS expression and activity. We particularly focus our attention on the DNA-dependent protein kinase (DNA-PK) complex, that recently emerged as an activator of inflammatory responses in tumour cells. We propose that stratification analyses on cGAS and DNA-PK expression/activation status should be carried out to predict treatment efficacy. We herein also provide insights into non-canonical functions borne by cGAS and cGAMP, highlighting how they may influence tumourigenesis. All these parameters should be taken into consideration concertedly to choose strategies aiming to effectively boost tumour immunogenicity.
Assuntos
Neoplasias , Proteínas Quinases , Humanos , Carcinogênese , DNA , Neoplasias/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , AnimaisRESUMO
The metazoan innate immune second messenger 2'3'-cGAMP is present both inside and outside cells. However, only extracellular cGAMP can be negatively regulated by the extracellular hydrolase ENPP1. Here, we determine whether ENPP1's regulation of extracellular cGAMP is a ubiquitous mechanism of attenuating stimulator of interferon genes (STING) signaling. We identified ENPP1H362A, a point mutation that cannot degrade the 2'-5' linkage in cGAMP while maintaining otherwise normal function. The selectivity of this histidine is conserved down to bacterial nucleotide pyrophosphatase/phosphodiesterase (NPP), allowing structural analysis and suggesting an unexplored ancient history of 2'-5' cyclic dinucleotides. Enpp1H362A mice demonstrated that extracellular cGAMP is not responsible for the devastating phenotype in ENPP1-null humans and mice but is responsible for antiviral immunity and systemic inflammation. Our data define extracellular cGAMP as a pivotal STING activator, identify an evolutionarily critical role for ENPP1 in regulating inflammation, and suggest a therapeutic strategy for viral and inflammatory conditions by manipulating ENPP1 activity.