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Phosphodiesterase mutations increase pneumococcus-induced NF-κB activity. (A) Relative NF-κB-mediated gene expression was measured using a mouse macrophage-like RAW264.7 cell line which had been stably transduced with a firefly luciferase transgene responsive to NF-κB. Cultures were infected 2 h with the indicated bacteria, and luciferase values were quantified using a luminometer and expressed relative to LPS positive control wells run in parallel (N = 9 experiments). (B) Relative induction of TNFα mRNA was measured in RAW264.7 cell cultures infected 2 h with the indicated bacteria. TNFα mRNA was measured and normalized to 18S rRNA using qPCR, and expressed relative to the cells infected by WT bacteria (N = 3 experiments). (C) Relative induction of IL-1β mRNA was measured in RAW264.7 cell cultures infected 2 h with the indicated bacteria. IL-1β mRNA was measured and normalized to 18S rRNA using qPCR, and expressed relative to the cells infected by WT bacteria (N = 3 experiments). Throughout panels, asterisk (*) indicates P < 0.05 compared to WT.
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Cyclic di-AMP (c-di-AMP) is an important signaling molecule for pneumococci, and as a uniquely prokaryotic product it can be recognized by mammalian cells as a danger signal that triggers innate immunity. Roles of c-di-AMP in directing host responses during pneumococcal infection are only beginning to be defined. We hypothesized that pneumococci wi...
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... In several LAB, including S. pneumoniae, Streptococcus mitis and Enterococcus faecalis, inactivation of both gdpP and dhhP genes result in significantly greater c-di-AMP levels compared to mutants with only one of these genes inactivated (Bai et al., 2013, Rørvik et al., 2020, Kundra et al., 2021. Interestingly under biofilm-like growth conditions with 5% CO 2 , a S. pneumoniae dhhP mutant, but not a gdpP mutant, contained a very high c-di-AMP level (Wooten et al., 2020). In other LAB however, inactivation of just gdpP results in very high (20-30-fold) c-di-AMP levels compared with the wild-type (Pham et al., 2018, Pham et al., 2021. ...
Cyclic-di-AMP is a nucleotide second messenger present in Gram-positive bacteria and some Gram-negative bacteria and Archaea. The intracellular concentration of cyclic-di-AMP is adjusted in response to environmental and cellular cues, primarily through the activities of synthesis and degradation enzymes. It performs its role by binding to protein and riboswitch receptors, many of which contribute to osmoregulation. Imbalances in cyclic-di-AMP can lead to pleiotropic phenotypes, affecting aspects such as growth, biofilm formation, virulence and resistance to osmotic, acid and antibiotic stressors. This review focuses on cyclic-di-AMP signalling in lactic acid bacteria (LAB) incorporating recent experimental discoveries and presenting an analysis of signalling components from a variety of LAB including those found in food, and commensal, probiotic and pathogenic species. All LAB possess enzymes for the synthesis and degradation of cyclic-di-AMP, but they are highly variable with regards to the receptors they possess. Studies in Lactococcus and Streptococcus have revealed a conserved function for c-di-AMP in inhibiting the transport of potassium and glycine betaine, either through direct binding to transporters or to a transcriptional regulator. Structural analysis of several cyclic-di-AMP receptors from LAB has also provided insights into how this nucleotide exerts its influence.
... And phosphodiesterase 1 (Pde1) and Pde2 are responsible for the degrading of c-di-AMP (237). Pneumococci deficient in Pde2 led to increased concentrations of cdi-AMP in the mutant pneumococci and resulted in the hyperactivation of STING and excessive IFN-b expression, as well as rapid cytotoxicity (238). However, another study showed that cGAS-STING pathway had no contribution to the immune response against S. pneumoniae in mice and humans, although pneumococcal DNA could be detected by cGAS and then initiated type I IFN production through STING (207). ...
The efficacious detection of pathogens and prompt induction of innate immune signaling serve as a crucial component of immune defense against infectious pathogens. Over the past decade, DNA-sensing receptor cyclic GMP-AMP synthase (cGAS) and its downstream signaling adaptor stimulator of interferon genes (STING) have emerged as key mediators of type I interferon (IFN) and nuclear factor-κB (NF-κB) responses in health and infection diseases. Moreover, both cGAS-STING pathway and pathogens have developed delicate strategies to resist each other for their survival. The mechanistic and functional comprehension of the interplay between cGAS-STING pathway and pathogens is opening the way for the development and application of pharmacological agonists and antagonists in the treatment of infectious diseases. Here, we briefly review the current knowledge of DNA sensing through the cGAS-STING pathway, and emphatically highlight the potent undertaking of cGAS-STING signaling pathway in the host against infectious pathogenic organisms.
... Higher IFN-β levels in vivo increase bacterial killing by the host (Andrade et al., 2016). Pde2-deficient S. pneumoniae with elevated c-di-AMP drive aberrant innate immune responses from macrophages involving the hyperactivation of STING, excessive IFN-β expression, and rapid cytotoxicity (Wooten et al., 2020). The effect of the Streptococcus suis SntA (hemin-binding protein) on c-di-AMP is comparable with the activity of cdnP that dampened type I IFN response (Cabezas et al., 2022). ...
Cyclic di-adenosine monophosphate (c-di-AMP) is a second messenger which is widely used in signal transduction in bacteria and archaea. c-di-AMP plays an important role in the regulation of bacterial physiological activities, such as the cell cycle, cell wall stability, environmental stress response, and biofilm formation. Moreover, c-di-AMP produced by pathogens can be recognized by host cells for the activation of innate immune responses. It can induce type I interferon (IFN) response in a stimulator of interferon genes (STING)-dependent manner, activate the nuclear factor kappa B (NF-κB) pathway, inflammasome, and host autophagy, and promote the production and secretion of cytokines. In addition, c-di-AMP is capable of triggering a host mucosal immune response as a mucosal adjuvant. Therefore, c-di-AMP is now considered to be a new pathogen-associated molecular pattern in host immunity and has become a promising target in bacterial/viral vaccine and drug research. In this review, we discussed the crosstalk between bacteria and host immunity mediated by c-di-AMP and addressed the role of c-di-AMP as a mucosal adjuvant in boosting evoked immune responses of subunit vaccines. The potential application of c-di-AMP in immunomodulation and immunotherapy was also discussed in this review.
... 2010年, Rao等人 [24] 首次在枯草 芽孢杆菌中发现, YybT蛋白具有水解c-di-AMP的能力, 后来YybT蛋白被重新命名为GdpP. 随后在一些厚壁菌 门的细菌(如金黄色葡萄球菌 [15] 、肺炎链球菌 [32] 、粪 肠球菌 [33] 等)中也发现了GdpP及其同源基因. GdpP型 PDE的N端具有两个跨膜螺旋区, 其后是PAS结构域、 GGDEF结构域及位于C端的DHH/DHHA1结构域 [24] . ...
... 肺炎链球菌中Δpde2菌株能迅速诱导巨 噬细胞中IFN-β的特异性表达, 并引起巨噬细胞病死率 增加. 进一步研究发现, cGAS-STING和IRF3/IRF7通路 通过c-di-AMP诱导IFN-β的产生是巨噬细胞对Δpde2肺 炎链球菌免疫反应的潜在机制 [32] . 与肺炎链球菌相似, 单核细胞增生李斯特菌感染骨髓来源的巨噬细胞过程 中, pdeA(GdpP同源物)和pgpH基因缺失的单核细胞增 生李斯特菌在感染骨髓来源的巨噬细胞过程中能够分 泌大量c-di-AMP到宿主胞浆, 触发了I型干扰素应答, 导致受感染宿主细胞产生和分泌大量IFN-β [13,55] . ...
Cyclic dimeric adenosine 3′,5′-monophosphate (c-di-AMP) is an emerging second messenger with multiple functions in Archaea, Gram-positive bacteria, Mycoplasma, Chlamydia, Cyanobacteria, and some other Gram-negative bacteria. C-di-AMP-specific phosphodiesterase (PDE) maintains the balance of c-di-AMP metabolism in bacteria. PDEs in bacteria have a vibrant structure and can be classified into three categories based on their functional domains. The first characterized PDE includes GdpP and DhhP homologs. GdpP has two transmembrane helices, a PAS (Per-Arnt-Sim) domain, a degenerate GGDEF domain, and the catalytic domains DHH and DHHA1. DhhP is a stand-alone DHH-DHHA1 domain protein compared with GdpP. Both DhhP and GdpP homologs degrade c-di-AMP via DHH/DHHA1 (Asp-His-His) domain. The second involves PgpH homologs, HD (Asp-His) is the main functional domain of hydrolyzing c-di-AMP. The HD domain structure remains conserved, although the amino acid sequence identity is low in certain bacteria. The newly discovered bacterial extracellular nucleosidases contain CdnP and AtaC and are sensitive to pH and metal ions. However, their catalytic domain for enzymatic activity and specific hydrolysis mechanism needs improvement. C-di-AMP-specific PDE could degrade c-di-AMP to one pApA molecule or two AMP molecules. Some PDEs hydrolyze in two steps, while others only hydrolyze in one, which could be due to conserved metal ion centers and unique substrate binding sites found in different PDEs. However, some c-di-AMP-specific PDEs, such as DhhP, can hydrolyze not only c-di-AMP and pApA but also c-di-GMP and pGpG. Most c-di-AMP-containing bacteria encode two or more PDEs simultaneously, which sense different signals or display distinct substrate preferences to perform diverse cellular functions individually or cooperatively. PDEs are important for maintaining the physiological function of bacteria and eliciting the immune response of host cells by regulating the bacterial c-di-AMP level. Recent studies suggest that PDE deletion or mutation affects multiple physiological functions including bacterial growth, virulence, and sensitivity to the external environment. GdpP leads to a dramatic increase of c-di-AMP in Staphylococcus aureus, thus reducing bacterial metabolic activity and growth. PDE absence in Lactococcus lactis improves bacterial heat tolerance and increases susceptibility to osmotic stress. PDE deletion could inhibit the overall virulence of particle pathogenic bacteria such as Streptococcus. Thus, PDE can be concluded as a potential virulence factor of pathogenic bacteria. Meanwhile, PDE also mediates bacterial biofilm formation via modulating c-di-AMP, which may be a key element in microbial drug resistance. Furthermore, c-di-AMP can effectively activate key molecules of the host cell immune response during pathogen infection; however, its levels are strictly regulated by PDE. This indicates the PDE’s importance in bacterial physiological function and host immune response. Therefore, c-di-AMP-specific PDEs have great significance in the field of antibacterial drugs as a target for preventing and treating pathogenic infections. This paper reviews the types, structures, and biological functions of PDE, aiming to provide references for related research.
... It enters mature phagolysosomes and releases nucleic acids and c-di-AMP into the host cytosol via a secretion system or following partial lysis. S. pneumoniae c-di-AMP initiates host innate immune responses by activating STING directly and increasing the expression of IFN-b (99). IFN-b production is also induced by pneumolysin (Ply), a pore-forming protein and a major virulence factor of S. pneumoniae. ...
Cyclic guanosine monophosphate (GMP)-adenosine monophosphate (AMP) (cGAMP) synthase (cGAS), along with the adaptor stimulator of interferon genes (STING), are crucial components of the innate immune system, and their study has become a research hotspot in recent years. Many biochemical and structural studies that have collectively elucidated the mechanism of activation of the cGAS-STING pathway with atomic resolution have provided insights into the roles of the cGAS-STING pathway in innate immunity and clues to the origin and evolution of the modern cGAS-STING signaling pathway. The cGAS-STING pathway has been identified to protect the host against viral infection. After detecting viral dsDNA, cGAS synthesizes a second messenger to activate STING, eliciting antiviral immune responses by promoting the expression of interferons (IFNs) and hundreds of IFN-stimulated genes (ISGs). Recently, the cGAS-STING pathway has also been found to be involved in response to bacterial infections, including bacterial pneumonia, melioidosis, tuberculosis, and sepsis. However, compared with its functions in viral infection, the cGAS-STING signaling pathway in bacterial infection is more complex and diverse since the protective and detrimental effects of type I IFN (IFN-I) on the host depend on the bacterial species and infection mode. Besides, STING activation can also affect infection prognosis through other mechanisms in different bacterial infections, independent of the IFN-I response. Interestingly, the core protein components of the mammalian cGAS-STING signaling pathway have been found in the bacterial defense system, suggesting that this widespread signaling pathway may have originated in bacteria. Here, we review recent findings related to the structures of major molecules involved in the cGAS-STING pathway and the effects of the cGAS-STING pathway in various bacterial infections and bacterial immunity, which may pave the way for the development of new antibacterial drugs that specifically kill bacteria without harmful effects on the host.
... In particular, dhhP deletion or simultaneous deletions of both PDEs significantly increase the intracellular c-di-AMP pool and cause severe growth defects in rich media (Bowman et al., 2016;Konno et al., 2018;Wooten et al., 2020;Ye et al., 2014). As an interesting exception, dhhP is essential in the gram-negative pathogen Borrelia burgdorferi (Ye et al., 2014). ...
Lebewesen nutzen nukleotid-basierte sekundäre Botenstoffe um extra- und intrazelluläre Signale zur Induktion einer entsprechenden Zellantwort weiterzuleiten. Das zyklische Dinukleotid c-di-AMP steuert verschiedene physiologische Prozesse und ist für viele Bakterien unter bestimmten Bedingungen essentiell. Dieses Signalmolekül muss präzise reguliert werden, da seine Akkumulation oft toxisch ist. Diadenylatzyklasen mit einer DAC-Domäne synthetisieren c-di-AMP, welches von Phosphodiesterasen (PDE) mit DHH/DHHA1- oder HD-Domänen abgebaut wird. Streptomyceten sind im Boden lebende, Gram-positive Actinobakterien mit einem komplexen Lebenszyklus, während welchem sie vielzählige sekundäre Metabolite, inklusive Antibiotika, produzieren. Die Regulierung des zellulären c-di-AMP und seine Bedeutung in der Streptomyceten-Biologie waren zu Beginn dieser Studie weitgehend unbekannt. Zur c-di-AMP-Synthese nutzen Streptomyces die DAC DisA, besitzen aber keine der typischen PDEs sowie die meisten der bekannten c-di-AMP-bindenden Effektoren. Diese Arbeit zeigt, dass DisA die wichtigste c-di-AMP-Synthetase in Streptomyces venezuelae ist. AtaC wurde als eine PDE identifiziert, welche eine neue Klasse von c-di-AMP PDEs begründet. Während eine ataC-Deletion zu Störungen in Differenzierung und Wachstum in S. Venezuelae führt, führt die Inaktivierung von disA zur Sensitivität gegenüber erhöhten Konzentrationen von monovalenten Kationen im Medium. CpeA und CpeD wurden als erste c-di-AMP-bindende Proteine im Streptomyces Signalnetzwerk charakterisiert. Die entsprechenden Gene sind in Operons mit putativen Kation/Proton-Antiportern cpeB bzw. cpeE kodiert und die jeweiligen Genprodukte interagieren c-di-AMP-abhängig in vivo. Obwohl Deletion von cpe und Überexpression von cpeABC in S. venezuelae keine Phänotypen zeigten, verbesserte die CpeABC-Expression in Escherichia coli das Wachstum in Kalium-supplementierten Medien, was auf eine Funktion von cpe in der Regulation von Kalium hindeutet.
Cyclic dimeric adenosine monophosphate (c‐di‐AMP) has been well studied in bacteria, including those of the genus Streptococcus , since the first recognition of this dinucleotide in 2008. Streptococci possess a sole diadenylate cyclase, CdaA, and distinct c‐di‐AMP phosphodiesterases. Interestingly, cdaA is required for viability of some streptococcal species but not all when streptococci are grown in standard laboratory media. Bacteria of this genus also have distinct c‐di‐AMP effector proteins, diverse c‐di‐AMP‐signaling pathways, and subsequent biological outcomes. In streptococci, c‐di‐AMP may influence bacterial growth, morphology, biofilm formation, competence program, drug resistance, and bacterial pathogenesis. c‐di‐AMP secreted by streptococci has also been shown to interact with the mammalian host and induces immune responses including type I interferon production. In this review, we summarize the reported c‐di‐AMP networks in seven species of the genus Streptococcus , which cause diverse clinical manifestations, and propose future perspectives to investigate the signaling molecule in these streptococcal pathogens.
Streptococcus pneumoniae is a major human pathogen and rising resistance to β-lactam antibiotics, such as penicillin, is a significant threat to global public health. Mutations occurring in the penicillin-binding proteins (PBPs) can confer high-level penicillin resistance but other poorly understood genetic factors are also important. Here, we combined strictly controlled laboratory experiments and population analyses to identify a new penicillin resistance pathway that is independent of PBP modification. Initial laboratory selection experiments identified high-frequency pde1 mutations conferring S. pneumoniae penicillin resistance. The importance of variation at the pde1 locus was confirmed in natural and clinical populations in an analysis of >7,200 S. pneumoniae genomes. The pde1 mutations identified by these approaches reduce the hydrolytic activity of the Pde1 enzyme in bacterial cells and thereby elevate levels of cyclic-di-adenosine monophosphate and penicillin resistance. Our results reveal rapid de novo loss of function mutations in pde1 as an evolutionary gateway conferring low-level penicillin resistance. This relatively simple genomic change allows cells to persist in populations on an adaptive evolutionary pathway to acquire further genetic changes and high-level penicillin resistance.
Recent studies have illustrated the functional significance of DNA recognition in the activation of innate immune responses among a variety of diseases. The cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway has been found to be modulated by post-translational modifications and can regulate the immune response via type I IFNs. Accumulating evidence indicates a pivotal role of cGAS-STING signaling, being protective or pathogenic, in the development of diseases. Thus, a comprehensive understanding of the post-translational modifications of cGAS-STING pathway and their role in disease development will provide insights in predicting individual disease outcomes and developing appropriate therapies. In this review, we will discuss the regulation of the cGAS-STING pathway and its implications in disease pathologies, as well as pharmacologic strategies to target the cGAS-STING pathway for therapeutic intervention.
