Coexistence of nonfluorescent chromoproteins and fluorescent proteins in massive Porites spp. corals manifesting a pink pigmentation response.

Introduction: Several fluorescent proteins (FPs) and chromoproteins (CPs) are present in anthozoans and play possible roles in photoprotection. Coral tissues in massive corals often display discoloration accompanied by inflammation. Incidences of the pink pigmentation response (PPR) in massive Porites, described as inflammatory pink lesions of different shapes and sizes, has recently increased worldwide. FPs are reported to be present in PPR lesions, wherein a red fluorescent protein (RFP) appears to play a role in reducing reactive oxygen species. However, to date, the biochemical characterization and possible roles of the pigments involved are poorly understood. The present study aimed to identify and characterize the proteins responsible for pink discoloration in massive Porites colonies displaying PPRs, as well as to assess the differential distribution of pigments and the antioxidant properties of pigmented areas. Method: CPs were extracted from PPR lesions using gel-filtration chromatography and identified via genetic analysis using liquid chromatography-tandem mass spectrometry. The coexistence of CPs and RFP in coral tissues was assessed using microscopic observation. Photosynthetic antivity and hydrogen peroxide-scavenging activitiy were measured to assess coral stress conditions. Results: The present study revealed that the same CP (plut2.m8.16902.m1) isolated from massive Porites was present in both the pink spot and patch morphologies of the PPR. CPs were also found to coexist with RFP in coral tissues that manifested a PPR, with a differential distribution (coenosarc or tip of polyps' tentacles). High hydrogen peroxide-scavenging rates were found in tissues affected by PPR. Discussion and Conclusion: The coexistence of CPs and RFP suggests their possible differential role in coral immunity. CPs, which are specifically expressed in PPR lesions, may serve as an antioxidant in the affected coral tissue. Overall, this study provides new knowledge to our understanding of the role of CPs in coral immunity.

Exploring the midgut physiology of the non-haematophagous mosquito Toxorhynchites theobaldi.

Toxorhynchites mosquitoes have an exclusively phytophagous feeding habit as adults, which leads to significant differences in their morphophysiology compared with haematophagous mosquitoes. However, the molecular mechanisms of digestion in this mosquito are not well understood. In this study, RNA sequencing of the posterior midgut (PMG) of the mosquito Toxorhynchites theobaldi was undertaken, highlighting its significance in mosquito digestion. Subsequently, a comparison was made between the differential gene expression of the PMG and that of the anterior midgut. It was found that the most abundant proteases in the PMG were trypsin and chymotrypsin, and the level of gene expression for enzymes essential for digestion (such as serine protease, α-amylase and pancreatic triacylglycerol lipase) and innate immune response (including catalase, cecropin-A2 and superoxide dismutase) was like that of haematophagous mosquitoes. Peritrophin-1 was detected in the entire midgut, with an elevated expression level in the PMG. Based on our findings, it is hypothesized that a non-haematophagic habit might have been exhibited by the ancestor of Tx. theobaldi, and this trait may have been retained. This study represents a pioneering investigation at the molecular level of midgut contents in a non-haematophagous mosquito. The findings offer valuable insights into the evolutionary aspects of feeding habits in culicids.

The Fusicoccin story revisited.

In plant biology Fusicoccin (FC) is one of the most studied fungal metabolites to date. Since the structural identification in 1964, much has been learned about its effects on the physiology of plants, about the interference with the action of plant hormones, the molecular nature of the plant receptor(s) for FC and the biosynthetic pathway for FC in the fungus. The finding that the plasma membrane H+-ATPase in combination with 14-3-3 proteins acts as high-affinity receptor for FC was a breakthrough in the field. Ever since, the binding of FC to the ATPase|14-3-3 receptor has taken center stage in explaining all FC induced physiological effects. However, a more critical review shows that this is not at all evident for a number of FC induced effects. Examples of this are: the inhibition of outward rectifying K+-channels in guard cells, the phosphorylation/activation of PEP-carboxylase and malate accumulation, the antagonism with ABA induced production of H2O2 / NO and the effect on ethylene production. In addition, recently two other physiological processes were shown to be targeted by FC, viz. the activation of TORC1 and the interference of FC with the immune response to fungal elicitors. In this review, the notion will be challenged that all FC affected processes start with the binding to and activation of the PM-ATPase and the question is raised whether may be other proteins with a key role in the respective processes are directly targeted by FC. A second unresolved question is whether FC may be another example of a fungal molecule turning out to be a 'copy' of an as yet unknown plant molecule; in analogy to the fungal product and plant hormone gibberellic acid. A relevant question in this respect is whether it is a coincidence that proteins that act in a coordinated fashion during stomatal opening (the ATPases and K+-channels) are targeted by FC? Or are the sites where FC binds in the plant, conserved during evolution because they serve a physiological role, namely the accommodation of a plant produced molecule? In view of the evidence, albeit not conclusive, that plants indeed produce 'FC-like ligands', it is worthwhile to make a renewed attempt with current day improved technology to answer this question and may be upgrade FC or structural analogue(s) to a new level, the level of plant hormone.

The Influence of Metabolic Risk Factors on the Inflammatory Response Triggered by Myocardial Infarction: Bridging Pathophysiology to Treatment.

Myocardial infarction (MI) sets off a complex inflammatory cascade that is crucial for effective cardiac healing and scar formation. Yet, if this response becomes excessive or uncontrolled, it can lead to cardiovascular complications. This review aims to provide a comprehensive overview of the tightly regulated local inflammatory response triggered in the early post-MI phase involving cardiomyocytes, (myo)fibroblasts, endothelial cells, and infiltrating immune cells. Next, we explore how the bone marrow and extramedullary hematopoiesis (such as in the spleen) contribute to sustaining immune cell supply at a cardiac level. Lastly, we discuss recent findings on how metabolic cardiovascular risk factors, including hypercholesterolemia, hypertriglyceridemia, diabetes, and hypertension, disrupt this immunological response and explore the potential modulatory effects of lifestyle habits and pharmacological interventions. Understanding how different metabolic risk factors influence the inflammatory response triggered by MI and unraveling the underlying molecular and cellular mechanisms may pave the way for developing personalized therapeutic approaches based on the patient's metabolic profile. Similarly, delving deeper into the impact of lifestyle modifications on the inflammatory response post-MI is crucial. These insights may enable the adoption of more effective strategies to manage post-MI inflammation and improve cardiovascular health outcomes in a holistic manner.

Common virulence factors between Histoplasma and Paracoccidioides: Recognition of Hsp60 and Enolase by CR3 and plasmin receptors in host cells.

Over the last two decades, the incidence of Invasive Fungal Infections (IFIs) globally has risen, posing a considerable challenge despite available antifungal therapies. Addressing this, the World Health Organization (WHO) prioritized research on specific fungi, notably Histoplasma spp. and Paracoccidioides spp. These dimorphic fungi have a mycelial life cycle in soil and a yeast phase associated with tissues of mammalian hosts. Inhalation of conidia and mycelial fragments initiates the infection, crucially transforming into the yeast form within the host, influenced by factors like temperature, host immunity, and hormonal status. Survival and multiplication within alveolar macrophages are crucial for disease progression, where innate immune responses play a pivotal role in overcoming physical barriers. The transition to pathogenic yeast, triggered by increased temperature, involves yeast phase-specific gene expression, closely linked to infection establishment and pathogenicity. Cell adhesion mechanisms during host-pathogen interactions are intricately linked to fungal virulence, which is critical for tissue colonization and disease development. Yeast replication within macrophages leads to their rupture, aiding pathogen dissemination. Immune cells, especially macrophages, dendritic cells, and neutrophils, are key players during infection control, with macrophages crucial for defense, tissue integrity, and pathogen elimination. Recognition of common virulence molecules such as heat- shock protein-60 (Hsp60) and enolase by pattern recognition receptors (PRRs), mainly via the complement receptor 3 (CR3) and plasmin receptor pathways, respectively, could be pivotal in host-pathogen interactions for Histoplasma spp. and Paracoccidioides spp., influencing adhesion, phagocytosis, and inflammatory regulation. This review provides a comprehensive overview of the dynamic of these two IFIs between host and pathogen. Further research into these fungi's virulence factors promises insights into pathogenic mechanisms, potentially guiding the development of effective treatment strategies.

IFITM3 reduces infectious bursal disease virus proliferation by regulating interferon expression.

Interferon-inducible transmembrane protein 3 (IFITM3), a member of the interferon-stimulating factor (ISG) family, has a wide range of antiviral functions. Infectious bursal disease virus (IBDV) mainly invades the bursa of Fabricius in chickens, causing a reduction in their immunity and resulting in death from secondary infections. Our previous study found that IBDV infection promotes the expression of chicken IFITM3. However, the role of chicken IFITM3 in IBDV infection remains unknown. To explore this role, the overexpression vector for IFITM3 was constructed and transfected into HD-11 and DF-1 cells. The results showed that the overexpression of IFITM3 significantly reduced IBDV proliferation. While the IBDV proliferation increased when IFITM3 was inhibited by using siRNA. To further explore the mechanism by which IFITM3 reduces IBDV proliferation, the effects of IFITM3 on interferon (IFN) were investigated. Transfecting the constructed IFITM3 vectors into HD-11 and DF-1 cells demonstrated that IFITM3 promoted the expression of IFN-α, IFN-ß, and IFN-γ. To investigate the mechanism by which IFITM3 regulates IFN expression, the effects of IFITM3 on IFN production were explored. The results showed that the IKB gene mainly affected the regulatory effects of IFITM3 on IFN. Taken together, IFITM3 may reduce viral proliferation by regulating changes in IFNs, and this process may involve a positive feedback effect of IFITM3 on IFN. IKB plays an important role in the regulation of IFN effects by IFITM3.

Cap-related modifications of RNA regulate binding to IFIT proteins.

All cells in our body are equipped with receptors to recognize pathogens and trigger a rapid defense response. As a result, foreign molecules are blocked and cells are alerted to the danger. Among the many molecules produced in response to viral infection are interferon-induced proteins with tetratricopeptide repeats (IFITs). Their role is to recognize foreign mRNA and eliminate it from the translational pool of transcripts. In the present study, we used biophysical methods to characterize the interactions between IFIT1 protein and its partners IFIT2 and IFIT3. IFIT1 interacts with IFIT3 with nanomolar binding affinity, which did not change significantly in the presence of the preformed IFIT2/3 complex. The interactions between IFIT2 and IFIT3 and IFIT1 and IFIT2 were one order of magnitude weaker. We also present kinetic data of the interactions between the IFIT protein complex and short RNA bearing various modifications at the 5' end. We show kinetic parameters for interaction between IFIT complex and RNA with m6Am modification. The results show that the cap adjacent m6Am modification is a stronger signature than cap1 alone. It blocks the formation of a complex between IFIT proteins and m7Gpppm6Am-RNA much more effectively than other cap modifications. In contrast, m6A in the 5'UTR is not recognized by IFIT proteins and does not contribute to translation repression by IFIT proteins. The data obtained are important for understanding the regulation of expression of genetic information. They indicate that 2'-O and m6Am modifications modulate the availability of mRNA molecules for proteins of innate immune response.

Association between a past history of nausea and vomiting during pregnancy and fever after the BNT162b2 COVID-19 vaccination.

PURPOSE: Nausea and vomiting during pregnancy (NVP) are common among pregnant women and can be severe enough to require hospitalization. However, the mechanism underlying NVP pathogenesis remains unclear. This study examined factors associated with adverse events after vaccination, including a past history of NVP. METHODS: A questionnaire-based survey was completed by non-pregnant women working at Nagasaki University Hospital who received two doses of the BNT162b2 coronavirus disease 2019 (COVID-19) vaccine. This study primarily examined the association between a past history of NVP and post-vaccination fever, as fever was determined to be the most objective and reliable indicator of the surveyed adverse events. RESULTS: Multivariate logistic regression analysis showed that post-vaccination fever was more strongly associated with a past history of NVP (odds ratio, 1.88; 95% confidence interval, 1.16-3.07) than either age (0.73; 0.56-0.96) or weight (0.85; 0.70-1.15), which were previously considered to be highly associated with the incidence of adverse events following COVID-19 vaccination. CONCLUSION: These results suggest an involvement of a similar pathological condition in developing NVP and post-vaccination fever.

Assessment of serum proprotein convertase subtilisin/kexin type 9 in pediatric sepsis syndrome.

Sepsis is a life-threatening condition that arises when the body's response to infection causes injury to its tissues and organs. Proprotein convertase subtilisin/kexin type 9 (PCSK9) is an enzyme released in response to the drop in cholesterol level occurring in sepsis. Our study aimed to evaluate the prognostic role of serum Proprotein convertase subtilisin/kexin type 9 (PCSK9) level in children with sepsis and severe sepsis. Sixty children were included in this study. They were divided into two groups: 30 children in the sepsis group and 30 in the severe sepsis group. Another 30 apparently healthy children were included as a control group. Blood samples were withdrawn from all included children for complete blood count (CBC), renal function tests (RFT), liver function tests (LFT), LDL-cholesterol (LDL-C), blood culture, and serum PCSK9. In this study, PCSK9 and LDL-C were higher in the two sepsis groups than in the control group (p < 0.05). They were also higher in the severe sepsis group than the sepsis group and in the non-survivors than in the survivors (p < 0.05). PCSK9 was positively correlated with length of hospital stay in surviving children (r = 0.67, p = 0.001) and had predicted significant hematological dysfunction (adjusted B = - 96.95, p = 0.03). In conclusion, the PCSK9 assay can be used as a biomarker for bad prognosis in children suffering from clinical sepsis.

Pseudomonas aeruginosa in chronic lung disease: untangling the dysregulated host immune response.

Pseudomonas aeruginosa is a highly adaptable opportunistic pathogen capable of exploiting barriers and immune defects to cause chronic lung infections in conditions such as cystic fibrosis. In these contexts, host immune responses are ineffective at clearing persistent bacterial infection, instead driving a cycle of inflammatory lung damage. This review outlines key components of the host immune response to chronic P. aeruginosa infection within the lung, beginning with initial pathogen recognition, followed by a robust yet maladaptive innate immune response, and an ineffective adaptive immune response that propagates lung damage while permitting bacterial persistence. Untangling the interplay between host immunity and chronic P. aeruginosa infection will allow for the development and refinement of strategies to modulate immune-associated lung damage and potentiate the immune system to combat chronic infection more effectively.

Comparative Analysis of Immune Gene Transcription in Sea Bream (Sparus aurata) Challenged with RGNNV or RGNNV/SJNNV Betanodaviruses.

Gilthead sea bream and European sea bass display different resistance-susceptibility patterns during infection with different nervous necrosis virus (NNV) species, which may derive from differences in the triggered immune response. Based on this premise, we analysed the transcription of several selected immune-related genes in sea bream experimentally infected with NNV isolates obtained from sea bass (DlNNV, RGNNV) or sea bream (SaNNV, RGNNV/SJNNV). Viral replication only occurred in SaNNV-inoculated fish; therefore, the differences between the immune response elicited by both viruses may be the key to understanding the mechanism behind the inhibition of DlNNV replication. Principal component analysis clustered samples according to the viral isolate from 1 day post infection onwards and evidenced differences in the immune response against both viruses, even though no mortalities or symptoms were recorded. The response against DlNNV is characterized by higher rtp3 transcription early after the infection, longer-lasting il-10 transcription and stronger induction of casp1 and hsp70. These genes should be targets for future studies in order to elucidate their role in hampering NNV replication in sea bream, which is essential for developing effective prophylactic measures.

Recent Progress in Innate Immune Responses to Enterovirus A71 and Viral Evasion Strategies.

Enterovirus A71 (EV-A71) is a major pathogen causing hand, foot, and mouth disease (HFMD) in children worldwide. It can lead to severe gastrointestinal, pulmonary, and neurological complications. The innate immune system, which rapidly detects pathogens via pathogen-associated molecular patterns or pathogen-encoded effectors, serves as the first defensive line against EV-A71 infection. Concurrently, the virus has developed various sophisticated strategies to evade host antiviral responses and establish productive infection. Thus, the virus-host interactions and conflicts, as well as the ability to govern biological events at this first line of defense, contribute significantly to the pathogenesis and outcomes of EV-A71 infection. In this review, we update recent progress on host innate immune responses to EV-A71 infection. In addition, we discuss the underlying strategies employed by EV-A71 to escape host innate immune responses. A better understanding of the interplay between EV-A71 and host innate immunity may unravel potential antiviral targets, as well as strategies that can improve patient outcomes.

SLIRP promotes autoimmune diseases by amplifying antiviral signaling via positive feedback regulation.

The abnormal innate immune response is a prominent feature underlying autoimmune diseases. One emerging factor that can trigger dysregulated immune activation is cytosolic mitochondrial double-stranded RNAs (mt-dsRNAs). However, the mechanism by which mt-dsRNAs stimulate immune responses remains poorly understood. Here, we discover SRA stem-loop interacting RNA binding protein (SLIRP) as a key amplifier of mt-dsRNA-triggered antiviral signals. In autoimmune diseases, SLIRP is commonly upregulated, and targeted knockdown of SLIRP dampens the interferon response. We find that the activation of melanoma differentiation-associated gene 5 (MDA5) by exogenous dsRNAs upregulates SLIRP, which then stabilizes mt-dsRNAs and promotes their cytosolic release to activate MDA5 further, augmenting the interferon response. Furthermore, the downregulation of SLIRP partially rescues the abnormal interferon-stimulated gene expression in autoimmune patients' primary cells and makes cells vulnerable to certain viral infections. Our study unveils SLIRP as a pivotal mediator of interferon response through positive feedback amplification of antiviral signaling.

Increased [18F]FDG uptake in the infarcted myocardial area displayed by combined PET/CMR correlates with snRNA-seq-detected inflammatory cell invasion.

Combined [18F]FDG PET-cardiac MRI imaging (PET/CMR) is a useful tool to assess myocardial viability and cardiac function in patients with acute myocardial infarction (AMI). Here, we evaluated the prognostic value of PET/CMR in a porcine closed-chest reperfused AMI (rAMI) model. Late gadolinium enhancement by PET/CMR imaging displayed tracer uptake defect at the infarction site by 3 days after the rAMI in the majority of the animals (group Match, n = 28). Increased [18F]FDG uptake at the infarcted area (metabolism/contractility mismatch) with reduced tracer uptake in the remote viable myocardium (group Mismatch, n = 12) 3 days after rAMI was observed in the animals with larger infarct size and worse left ventricular ejection fraction (LVEF) (34 ± 8.7 vs 42.0 ± 5.2%), with lower LVEF also at the 1-month follow-up (35.8 ± 9.5 vs 43.0 ± 6.3%). Transcriptome analyses by bulk and single-nuclei RNA sequencing of the infarcted myocardium and border zones (n = 3 of each group, and 3 sham-operated controls) revealed a strong inflammatory response with infiltration of monocytes and macrophages in the infarcted and border areas in Mismatch animals. Our data indicate a high prognostic relevance of combined PET/MRI in the subacute phase of rAMI for subsequent impairment of heart function and underline the adverse effects of an excessive activation of the innate immune system in the initial phase after rAMI.

Association between polymorphisms in TLR3, TICAM1 and IFNA1 genes and covid-19 severity in Southern Brazil.

BACKGROUND: A distinct phenotype in Coronavirus disease 2019 (Covid-19) was observed in severe patients, consisting of a highly impaired interferon (IFN) type I response, an exacerbated inflammatory response. OBJECTIVE: The aim of the present study was to investigate a possible association of single nucleotide polymorphisms (SNPs), in five genes related to the immune response, rs3775291 in TLR3; rs2292151 in TICAM1; rs1758566 in IFNA1; rs1800629 in TNF, and rs1800795 in IL6 with the severity of Covid-19. METHODS: A cross-sectional study was performed, with non-severe and severe/critical patients diagnosed with Covid-19, by two public hospitals in Brazil. In total, 300 patients were genotyped for the SNPs, 150 with the non-severe form of the disease and 150 with severe/critical form. RESULTS: The T/T genotype of TLR3 in recessive model shows 58% of protection against severe/critical Covid-19; as well as the genotypes G/A+A/A of TICAM1 in dominant model with 60% of protection, and in a codominant model G/A with 57% and A/A with 71% of protection against severe/critical Covid-19. Comparing severe and critical cases, the T/C genotype of IFNA1 in the codominant model and TC+C/C in the dominant model showed twice the risk of critical Covid-19. CONCLUSION: We can conclude that rs3775291, rs2292151 and rs1758566 can influence the COVID-19 severity.

Discovery and Characterization of the ddx41 Gene in Atlantic Salmon: Evolutionary Implications, Structural Functions, and Innate Immune Responses to Piscirickettsia salmonis and Renibacterium salmoninarum Infections.

The innate immune response in Salmo salar, mediated by pattern recognition receptors (PRRs), is crucial for defending against pathogens. This study examined DDX41 protein functions as a cytosolic/nuclear sensor for cyclic dinucleotides, RNA, and DNA from invasive intracellular bacteria. The investigation determined the existence, conservation, and functional expression of the ddx41 gene in S. salar. In silico predictions and experimental validations identified a single ddx41 gene on chromosome 5 in S. salar, showing 83.92% homology with its human counterpart. Transcriptomic analysis in salmon head kidney confirmed gene transcriptional integrity. Proteomic identification through mass spectrometry characterized three unique peptides with 99.99% statistical confidence. Phylogenetic analysis demonstrated significant evolutionary conservation across species. Functional gene expression analysis in SHK-1 cells infected by Piscirickettsia salmonis and Renibacterium salmoninarum indicated significant upregulation of DDX41, correlated with increased proinflammatory cytokine levels and activation of irf3 and interferon signaling pathways. In vivo studies corroborated DDX41 activation in immune responses, particularly when S. salar was challenged with P. salmonis, underscoring its potential in enhancing disease resistance. This is the first study to identify the DDX41 pathway as a key component in S. salar innate immune response to invading pathogens, establishing a basis for future research in salmonid disease resistance.

Mutations in nonstructural proteins essential for pathogenicity in SARS-CoV-2-infected mice.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) has resulted in substantial morbidity and mortality. The basis of severe disease in humans is difficult to determine without the use of experimental animal models. Mice are resistant to infection with ancestral strains of SARS-CoV-2, although many variants that arose later in the pandemic were able to directly infect mice. In almost all cases, viruses that naturally infected mice or were engineered to enable mouse infection required mouse passage to become virulent. In most cases, changes in structural and nonstructural changes occurred during mouse adaptation. However, the mechanism of increased virulence in mice is not understood. Here, using a recently described strain of mouse-adapted SARS-CoV-2 (rSARS2-MA30N501Y), we engineered a series of recombinant viruses that expressed a subset of the mutations present in rSARS2-MA30N501Y. Mutations were detected in the spike protein and in three nonstructural proteins (nsp4, nsp8, and nsp9). We found that infection of mice with recombinant SARS-CoV-2 expressing only the S protein mutations caused very mild infection. Addition of the mutations in nsp4 and nsp8 was required for complete virulence. Of note, all these recombinant viruses replicated equivalently in cultured cells. The innate immune response was delayed after infection with virulent compared to attenuated viruses. Further, using a lineage tracking system, we found that attenuated virus was highly inhibited in the ability to infect the parenchyma, but not the airway after infection. Together, these results indicate that mutations in both the S protein and nonstructural proteins are required for maximal virulence during mouse adaptation.IMPORTANCEUnderstanding the pathogenesis of coronavirus disease 2019 (COVID-19) requires the study of experimental animals after infection with severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2). For this purpose, several mouse-adapted SARS-CoV-2 strains have been developed. Here, using a strain of mouse-adapted virus that causes a range of diseases ranging from mild to severe, we show that mutations in both a structural protein [spike (S) protein] and nonstructural proteins are required for maximal virulence. Thus, changes in the S protein, the most widely studied viral protein, while required for mouse adaptation, are not sufficient to result in a virulent virus.

Nervous necrosis virus capsid protein and Protein A dynamically modulate the fish cGAS-mediated IFN signal pathway to facilitate viral evasion.

Nervous necrosis virus (NNV), an aquatic RNA virus belonging to Betanodavirus, infects a variety of marine and freshwater fishes, leading to massive mortality of cultured larvae and juveniles and substantial economic losses. The enzyme cyclic guanosine monophosphate-adenosine monophosphate synthase (cGAS) is widely recognized as a central component in the innate immune response to cytosolic DNA derived from different pathogens. However, little is known about the response of cGAS to aquatic RNA viruses. This study found that Epinephelus coioides cGAS (EccGAS) overexpression inhibited NNV replication, whereas EccGAS silencing promoted NNV replication. The anti-NNV activity of EccGAS was involved in interferon (IFN) signaling activation including tumor necrosis factor receptor-associated factor family member-associated NF-kappa-B activator-binding kinase 1 (TBK1) phosphorylation, interferon regulatory factor 3 (IRF3) nuclear translocation, and the subsequent induction of IFNc and ISGs. Interestingly, NNV employed its capsid protein (CP) or Protein A (ProA) to negatively or positively modulate EccGAS-mediated IFN signaling by simultaneously targeting EccGAS. CP interacted with EccGAS via the arm-P, S-P, and SD structural domains and promoted its polyubiquitination with K48 and K63 linkages in an EcUBE3C (the ubiquitin ligase)-dependent manner, ultimately leading to EccGAS degradation. Conversely, ProA bound to EccGAS and inhibited its ubiquitination and degradation. In regulating EccGAS protein content, CP's inhibitory action was more pronounced than ProA's protective effect, allowing successful NNV replication. These novel findings suggest that NNV CP and ProA dynamically modulate the EccGAS-mediated IFN signaling pathway to facilitate the immune escape of NNV. Our findings shed light on a novel mechanism of virus-host interaction and provide a theoretical basis for the prevention and control of NNV.IMPORTANCEAs a well-known DNA sensor, cGAS is a pivotal component in innate anti-viral immunity to anti-DNA viruses. Although there is growing evidence regarding the function of cGAS in the resistance to RNA viruses, the mechanisms by which cGAS participates in RNA virus-induced immune responses in fish and how aquatic viruses evade cGAS-mediated immune surveillance remain elusive. Here, we investigated the detailed mechanism by which EccGAS positively regulates the anti-NNV response. Furthermore, NNV CP and ProA interacted with EccGAS, regulating its protein levels through ubiquitin-proteasome pathways, to dynamically modulate the EccGAS-mediated IFN signaling pathway and facilitate viral evasion. Notably, NNV CP was identified to promote the ubiquitination of EccGAS via ubiquitin ligase EcUBE3C. These findings unveil a novel strategy for aquatic RNA viruses to evade cGAS-mediated innate immunity, enhancing our understanding of virus-host interactions.

Micronuclei induced by radiation, replication stress, or chromosome segregation errors do not activate cGAS-STING.

The cyclic guanosine monophosphate (GMP)-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway plays a pivotal role in innate immune responses to viral infection and inhibition of autoimmunity. Recent studies have suggested that micronuclei formed by genotoxic stress can activate innate immune signaling via the cGAS-STING pathway. Here, we investigated cGAS localization, activation, and downstream signaling from micronuclei induced by ionizing radiation, replication stress, and chromosome segregation errors. Although cGAS localized to ruptured micronuclei via binding to self-DNA, we failed to observe cGAS activation; cGAMP production; downstream phosphorylation of STING, TBK1, or IRF3; nuclear accumulation of IRF3; or expression of interferon-stimulated genes. Failure to activate the cGAS-STING pathway was observed across primary and immortalized cell lines, which retained the ability to activate the cGAS-STING pathway in response to dsDNA or modified vaccinia virus infection. We provide evidence that micronuclei formed by genotoxic insults contain histone-bound self-DNA, which we show is inhibitory to cGAS activation in cells.

Multifactoral immune modulation potentiates durable remission in multiple models of aggressive malignancy.

Tumors typically lack canonical danger signals required to activate adaptive immunity and also frequently employ substantial immunomodulatory mechanisms that downregulate adaptive responses and contribute to escape from immune surveillance. Given the variety of mechanisms involved in shielding tumors from immune recognition, it is not surprising that single-agent immunomodulatory approaches have been largely unsuccessful in generating durable antitumor responses. Here we report a unique combination of immunomodulatory and cytostatic agents that recondition the tumor microenvironment and eliminate complex and/or poor-prognosis tumor types including the non-immunogenic 4T-1 model of TNBC, the aggressive MOC-2 model of HNSCC, and the high-risk MYCN-amplified model of neuroblastoma. A course of therapy optimized for TNBC cured a majority of tumors in both ectopic and orthotopic settings and eliminated metastatic spread in all animals tested at the highest doses. Immune responses were transferable between therapeutic donor and naïve recipient through adoptive transfer, and a sizeable abscopal effect on distant, untreated lesions could be demonstrated experimentally. Similar results were observed in HNSCC and neuroblastoma models, with characteristic remodeling of the tumor microenvironment documented in all model systems. scRNA-seq analysis implicated upregulation of innate immune responses and antigen presentation in tumor cells and the myeloid cell compartment as critical early events. This analysis also highlighted the potential importance of the autonomic nervous system in the governance of inflammatory processes. The data indicate that the targeting of multiple pathways and mechanisms of action can result in substantial synergistic antitumor effects and suggest follow-up in the neoadjuvant setting may be warranted.