Dietary habits, traveling and the living situation potentially influence the susceptibility to SARS-CoV-2 infection: results from healthcare workers participating in the RisCoin Study.

PURPOSE: To explore occupational and non-occupational risk and protective factors for the coronavirus disease 2019 (COVID-19) in healthcare workers (HCWs). METHODS: Serum specimens and questionnaire data were obtained between October 7 and December 16, 2021 from COVID-19-vaccinated HCWs at a quaternary care hospital in Munich, Germany, and were analyzed in the RisCoin Study. RESULTS: Of 3,696 participants evaluated, 6.6% have had COVID-19 at least once. Multivariate logistic regression analysis identified working in patient care occupations (7.3% had COVID-19, 95% CI 6.4-8.3, Pr = 0.0002), especially as nurses, to be a potential occupation-related COVID-19 risk factor. Non-occupational factors significantly associated with high rates of the disease were contacts to COVID-19 cases in the community (12.8% had COVID-19, 95% CI 10.3-15.8, Pr < 0.0001), being obese (9.9% had COVID-19, 95% CI 7.1-13.5, Pr = 0.0014), and frequent traveling abroad (9.4% had COVID-19, 95% CI 7.1-12.3, Pr = 0.0088). On the contrary, receiving the basic COVID-19 immunization early during the pandemic (5.9% had COVID-19, 95% CI 5.1-6.8, Pr < 0.0001), regular smoking (3.6% had COVID-19, 95% CI 2.1-6.0, Pr = 0.0088), living with the elderly (3.0% had COVID-19, 95% CI 1.0-8.0, Pr = 0.0475), and frequent consumption of ready-to-eat meals (2.6% had COVID-19, 95% CI 1.1-5.4, Pr = 0.0045) were non-occupational factors potentially protecting study participants against COVID-19. CONCLUSION: The newly discovered associations between the living situation, traveling as well as dietary habits and altered COVID-19 risk can potentially help refine containment measures and, furthermore, contribute to new mechanistic insights that may aid the protection of risk groups and vulnerable individuals.

Oligoadenylate synthetase 1 displays dual antiviral mechanisms in driving translational shutdown and protecting interferon production.

In response to viral infection, how cells balance translational shutdown to limit viral replication and the induction of antiviral components like interferons (IFNs) is not well understood. Moreover, how distinct isoforms of IFN-induced oligoadenylate synthetase 1 (OAS1) contribute to this antiviral response also requires further elucidation. Here, we show that human, but not mouse, OAS1 inhibits SARS-CoV-2 replication through its canonical enzyme activity via RNase L. In contrast, both mouse and human OAS1 protect against West Nile virus infection by a mechanism distinct from canonical RNase L activation. OAS1 binds AU-rich elements (AREs) of specific mRNAs, including IFNß. This binding leads to the sequestration of IFNß mRNA to the endomembrane regions, resulting in prolonged half-life and continued translation. Thus, OAS1 is an ARE-binding protein with two mechanisms of antiviral activity: driving inhibition of translation but also a broader, non-canonical function of protecting IFN expression from translational shutdown.

E-selectin-mediated rapid NLRP3 inflammasome activation regulates S100A8/S100A9 release from neutrophils via transient gasdermin D pore formation.

S100A8/S100A9 is a proinflammatory mediator released by myeloid cells during many acute and chronic inflammatory disorders. However, the precise mechanism of its release from the cytosolic compartment of neutrophils is unclear. Here, we show that E-selectin-induced rapid S100A8/S100A9 release during inflammation occurs in an NLRP3 inflammasome-dependent fashion. Mechanistically, E-selectin engagement triggers Bruton's tyrosine kinase-dependent tyrosine phosphorylation of NLRP3. Concomitant potassium efflux via the voltage-gated potassium channel KV1.3 mediates ASC oligomerization. This is followed by caspase 1 cleavage and downstream activation of pore-forming gasdermin D, enabling cytosolic release of S100A8/S100A9. Strikingly, E-selectin-mediated gasdermin D pore formation does not result in cell death but is a transient process involving activation of the ESCRT III membrane repair machinery. These data clarify molecular mechanisms of controlled S100A8/S100A9 release from neutrophils and identify the NLRP3/gasdermin D axis as a rapid and reversible activation system in neutrophils during inflammation.

Systematic P2Y receptor survey identifies P2Y11 as modulator of immune responses and virus replication in macrophages.

The immune system is in place to assist in ensuring tissue homeostasis, which can be easily perturbed by invading pathogens or nonpathogenic stressors causing tissue damage. Extracellular nucleotides are well known to contribute to innate immune signaling specificity and strength, but how their signaling is relayed downstream of cell surface receptors and how this translates into antiviral immunity is only partially understood. Here, we systematically investigated the responses of human macrophages to extracellular nucleotides, focusing on the nucleotide-sensing GPRC receptors of the P2Y family. Time-resolved transcriptomic analysis showed that adenine- and uridine-based nucleotides induce a specific, immediate, and transient cytokine response through the MAPK signaling pathway that regulates transcriptional activation by AP-1. Using receptor trans-complementation, we identified a subset of P2Ys (P2Y1, P2Y2, P2Y6, and P2Y11) that govern inflammatory responses via cytokine induction, while others (P2Y4, P2Y11, P2Y12, P2Y13, and P2Y14) directly induce antiviral responses. Notably, P2Y11 combined both activities, and depletion or inhibition of this receptor in macrophages impaired both inflammatory and antiviral responses. Collectively, these results highlight the underappreciated functions of P2Y receptors in innate immune processes.

ß1 integrin signaling governs necroptosis via the chromatin-remodeling factor CHD4.

Fibrosis, characterized by sustained activation of myofibroblasts and excessive extracellular matrix (ECM) deposition, is known to be associated with chronic inflammation. Receptor-interacting protein kinase 3 (RIPK3), the central kinase of necroptosis signaling, is upregulated in fibrosis and contributes to tumor necrosis factor (TNF)-mediated inflammation. In bile-duct-ligation-induced liver fibrosis, we found that myofibroblasts are the major cell type expressing RIPK3. Genetic ablation of ß1 integrin, the major profibrotic ECM receptor in fibroblasts, not only abolished ECM fibrillogenesis but also blunted RIPK3 expression via a mechanism mediated by the chromatin-remodeling factor chromodomain helicase DNA-binding protein 4 (CHD4). While the function of CHD4 has been conventionally linked to the nucleosome-remodeling deacetylase (NuRD) and CHD4-ADNP-HP1(ChAHP) complexes, we found that CHD4 potently repressed a set of genes, including Ripk3, with high locus specificity but independent of either the NuRD or the ChAHP complex. Thus, our data uncover that ß1 integrin intrinsically links fibrotic signaling to RIPK3-driven inflammation via a novel mode of action of CHD4.

Hemophagocytic lymphohistiocytosis-like hyperinflammation due to a de novo mutation in DPP9.

BACKGROUND: Genetic defects in components of inflammasomes can cause autoinflammation. Biallelic loss-of-function mutations in dipeptidyl peptidase 9 (DPP9), a negative regulator of the NLRP1 and CARD8 inflammasomes, have recently been shown to cause an inborn error of immunity characterized by pancytopenia, skin manifestations, and increased susceptibility to infections. OBJECTIVE: We sought to study the molecular basis of autoinflammation in a patient with severe infancy-onset hyperinflammation associated with signs of fulminant hemophagocytic lymphohistiocytosis. METHODS: Using heterologous cell models as well as patient cells, we performed genetic, immunologic, and molecular investigations to identify the genetic cause and to assess the impact of the identified mutation on inflammasome activation. RESULTS: The patient exhibited pancytopenia with decreased neutrophils and T, B, and natural killer cells, and markedly elevated levels of lactate dehydrogenase, ferritin, soluble IL-2 receptor, and triglycerides. In addition, serum levels of IL-1ß and IL-18 were massively increased, consistent with inflammasome activation. Genetic analysis revealed a previously undescribed de novo mutation in DPP9 (c.755G>C, p.Arg252Pro) affecting a highly conserved amino acid residue. The mutation led to destabilization of the DPP9 protein as shown in transiently transfected HEK293T cells and in patient-derived induced pluripotent stem cells. Using functional inflammasome assays in HEK293T cells, we demonstrated that mutant DPP9 failed to restrain the NLRP1 and CARD8 inflammasomes, resulting in constitutive inflammasome activation. These findings suggest that the Arg252Pro DPP9 mutation acts in a dominant-negative manner. CONCLUSIONS: A de novo mutation in DPP9 leads to severe infancy-onset autoinflammation because of unleashed inflammasome activation.

Tumor necrosis factor is a necroptosis-associated alarmin.

Necroptosis is a form of regulated cell death that can occur downstream of several immune pathways. While previous studies have shown that dysregulated necroptosis can lead to strong inflammatory responses, little is known about the identity of the endogenous molecules that trigger these responses. Using a reductionist in vitro model, we found that soluble TNF is strongly released in the context of necroptosis. On the one hand, necroptosis promotes TNF translation by inhibiting negative regulatory mechanisms acting at the post-transcriptional level. On the other hand, necroptosis markedly enhances TNF release by activating ADAM proteases. In studying TNF release at single-cell resolution, we found that TNF release triggered by necroptosis is activated in a switch-like manner that exceeds steady-state TNF processing in magnitude and speed. Although this shedding response precedes massive membrane damage, it is closely associated with lytic cell death. Further, we found that lytic cell death induction using a pore-forming toxin also triggers TNF shedding, indicating that the activation of ADAM proteases is not strictly related to the necroptotic pathway but likely associated with biophysical changes of the cell membrane upon lytic cell death. These results demonstrate that lytic cell death, particularly necroptosis, is a critical trigger for TNF release and thus qualify TNF as a necroptosis-associated alarmin.

Transcriptional licensing is required for Pyrin inflammasome activation in human macrophages and bypassed by mutations causing familial Mediterranean fever.

Pyrin is a cytosolic immune sensor that nucleates an inflammasome in response to inhibition of RhoA by bacterial virulence factors, triggering the release of inflammatory cytokines, including IL-1ß. Gain-of-function mutations in the MEFV gene encoding Pyrin cause autoinflammatory disorders, such as familial Mediterranean fever (FMF) and Pyrin-associated autoinflammation with neutrophilic dermatosis (PAAND). To precisely define the role of Pyrin in pathogen detection in human immune cells, we compared initiation and regulation of the Pyrin inflammasome response in monocyte-derived macrophages (hMDM). Unlike human monocytes and murine macrophages, we determined that hMDM failed to activate Pyrin in response to known Pyrin activators Clostridioides difficile (C. difficile) toxins A or B (TcdA or TcdB), as well as the bile acid analogue BAA-473. The Pyrin inflammasome response was enabled in hMDM by prolonged priming with either LPS or type I or II interferons and required an increase in Pyrin expression. Notably, FMF mutations lifted the requirement for prolonged priming for Pyrin activation in hMDM, enabling Pyrin activation in the absence of additional inflammatory signals. Unexpectedly, in the absence of a Pyrin response, we found that TcdB activated the NLRP3 inflammasome in hMDM. These data demonstrate that regulation of Pyrin activation in hMDM diverges from monocytes and highlights its dysregulation in FMF.

Myoglobin regulates fatty acid trafficking and lipid metabolism in mammary epithelial cells.

Myoglobin (MB) is known to bind and deliver oxygen in striated muscles at high expression levels. MB is also expressed at much reduced levels in mammary epithelial cells, where the protein´s function is unclear. In this study, we aim to determine whether MB impacts fatty acid trafficking and facilitates aerobic fatty acid ß-oxidation in mammary epithelial cells. We utilized MB-wildtype versus MB-knockout mice and human breast cancer cells to examine the impact of MB and its oxygenation status on fatty acid metabolism in mouse milk and mammary epithelia. MB deficient cells were generated through CRISPR/Cas9 and TALEN approaches and exposed to various oxygen tensions. Fatty acid profiling of milk and cell extracts were performed along with cell labelling and immunocytochemistry. Our findings show that MB expression in mammary epithelial cells promoted fatty acid oxidation while reducing stearyl-CoA desaturase activity for lipogenesis. In cells and milk product, presence of oxygenated MB significantly elevated indices of limited fatty acid ß-oxidation, i.e., the organelle-bound removal of a C2 moiety from long-chain saturated or monounsaturated fatty acids, thus shifting the composition toward more saturated and shorter fatty acid species. Presence of the globin also increased cytoplasmic fatty acid solubility under normoxia and fatty acid deposition to lipid droplets under severe hypoxia. We conclude that MB can function in mammary epithelia as intracellular O2-dependent shuttle of oxidizable fatty acid substrates. MB's impact on limited oxidation of fatty acids could generate inflammatory mediator lipokines, such as 7-hexadecenoate. Thus, the novel functions of MB in breast epithelia described herein range from controlling fatty acid turnover and homeostasis to influencing inflammatory signalling cascade. Future work is needed to analyse to what extent these novel roles of MB also apply to myocytic cell physiology and malignant cell behaviour, respectively.

Phosphorylation of muramyl peptides by NAGK is required for NOD2 activation.

Bacterial cell wall components provide various unique molecular structures that are detected by pattern recognition receptors (PRRs) of the innate immune system as non-self. Most bacterial species form a cell wall that consists of peptidoglycan (PGN), a polymeric structure comprising alternating amino sugars that form strands cross-linked by short peptides. Muramyl dipeptide (MDP) has been well documented as a minimal immunogenic component of peptidoglycan1-3. MDP is sensed by the cytosolic nucleotide-binding oligomerization domain-containing protein 24 (NOD2). Upon engagement, it triggers pro-inflammatory gene expression, and this functionality is of critical importance in maintaining a healthy intestinal barrier function5. Here, using a forward genetic screen to identify factors required for MDP detection, we identified N-acetylglucosamine kinase (NAGK) as being essential for the immunostimulatory activity of MDP. NAGK is broadly expressed in immune cells and has previously been described to contribute to the hexosamine biosynthetic salvage pathway6. Mechanistically, NAGK functions upstream of NOD2 by directly phosphorylating the N-acetylmuramic acid moiety of MDP at the hydroxyl group of its C6 position, yielding 6-O-phospho-MDP. NAGK-phosphorylated MDP-but not unmodified MDP-constitutes an agonist for NOD2. Macrophages from mice deficient in NAGK are completely deficient in MDP sensing. These results reveal a link between amino sugar metabolism and innate immunity to bacterial cell walls.

ZAKα-driven ribotoxic stress response activates the human NLRP1 inflammasome.

Human NLRP1 (NACHT, LRR, and PYD domain-containing protein 1) is an innate immune sensor predominantly expressed in the skin and airway epithelium. Here, we report that human NLRP1 senses the ultraviolet B (UVB)- and toxin-induced ribotoxic stress response (RSR). Biochemically, RSR leads to the direct hyperphosphorylation of a human-specific disordered linker region of NLRP1 (NLRP1DR) by MAP3K20/ZAKα kinase and its downstream effector, p38. Mutating a single ZAKα phosphorylation site in NLRP1DR abrogates UVB- and ribotoxin-driven pyroptosis in human keratinocytes. Moreover, fusing NLRP1DR to CARD8, which is insensitive to RSR by itself, creates a minimal inflammasome sensor for UVB and ribotoxins. These results provide insight into UVB sensing by human skin keratinocytes, identify several ribotoxins as NLRP1 agonists, and establish inflammasome-driven pyroptosis as an integral component of the RSR.

Novel Poxin Stable cGAMP-Derivatives Are Remarkable STING Agonists.

2',3'-cGAMP is a cyclic A- and G-containing dinucleotide second messenger, which is formed upon cellular recognition of foreign cytosolic DNA as part of the innate immune response. The molecule binds to the adaptor protein STING, which induces an immune response characterized by the production of type I interferons and cytokines. The development of STING-binding molecules with both agonistic as well as antagonistic properties is currently of tremendous interest to induce or enhance antitumor or antiviral immunity on the one hand, or to treat autoimmune diseases on the other hand. To escape the host innate immune recognition, some viruses encode poxin endonucleases that cleave 2',3'-cGAMP. Here we report that dideoxy-2',3'-cGAMP (1) and analogs thereof, which lack the secondary ribose-OH groups, form a group of poxin-stable STING agonists. Despite their reduced affinity to STING, particularly the compound constructed from two A nucleosides, dideoxy-2',3'-cAAMP (2), features an unusually high antitumor response in mice.

Molecular Signature of Astrocytes for Gene Delivery by the Synthetic Adeno-Associated Viral Vector rAAV9P1.

Astrocytes have crucial functions in the central nervous system (CNS) and are major players in many CNS diseases. Research on astrocyte-centered diseases requires efficient and well-characterized gene transfer vectors. Vectors derived from the Adeno-associated virus serotype 9 (AAV9) target astrocytes in the brains of rodents and nonhuman primates. A recombinant (r) synthetic peptide-displaying AAV9 variant, rAAV9P1, that efficiently and selectively transduces cultured human astrocytes, has been described previously. Here, it is shown that rAAV9P1 retains astrocyte-targeting properties upon intravenous injection in mice. Detailed analysis of putative receptors on human astrocytes shows that rAAV9P1 utilizes integrin subunits αv, ß8, and either ß3 or ß5 as well as the AAV receptor AAVR. This receptor pattern is distinct from that of vectors derived from wildtype AAV2 or AAV9. Furthermore, a CRISPR/Cas9 genome-wide knockout screening revealed the involvement of several astrocyte-associated intracellular signaling pathways in the transduction of human astrocytes by rAAV9P1. This study delineates the unique receptor and intracellular pathway signatures utilized by rAAV9P1 for targeting human astrocytes. These results enhance the understanding of the transduction biology of synthetic rAAV vectors for astrocytes and can promote the development of advanced astrocyte-selective gene delivery vehicles for research and clinical applications.

In-depth profiling of COVID-19 risk factors and preventive measures in healthcare workers.

PURPOSE: To determine risk factors for coronavirus disease 2019 (COVID-19) in healthcare workers (HCWs), characterize symptoms, and evaluate preventive measures against SARS-CoV-2 spread in hospitals. METHODS: In a cross-sectional study conducted between May 27 and August 12, 2020, after the first wave of the COVID-19 pandemic, we obtained serological, epidemiological, occupational as well as COVID-19-related data at a quaternary care, multicenter hospital in Munich, Germany. RESULTS: 7554 HCWs participated, 2.2% of whom tested positive for anti-SARS-CoV-2 antibodies. Multivariate analysis revealed increased COVID-19 risk for nurses (3.1% seropositivity, 95% CI 2.5-3.9%, p = 0.012), staff working on COVID-19 units (4.6% seropositivity, 95% CI 3.2-6.5%, p = 0.032), males (2.4% seropositivity, 95% CI 1.8-3.2%, p = 0.019), and HCWs reporting high-risk exposures to infected patients (5.5% seropositivity, 95% CI 4.0-7.5%, p = 0.0022) or outside of work (12.0% seropositivity, 95% CI 8.0-17.4%, p < 0.0001). Smoking was a protective factor (1.1% seropositivity, 95% CI 0.7-1.8% p = 0.00018) and the symptom taste disorder was strongly associated with COVID-19 (29.8% seropositivity, 95% CI 24.3-35.8%, p < 0.0001). An unbiased decision tree identified subgroups with different risk profiles. Working from home as a preventive measure did not protect against SARS-CoV-2 infection. A PCR-testing strategy focused on symptoms and high-risk exposures detected all larger COVID-19 outbreaks. CONCLUSION: Awareness of the identified COVID-19 risk factors and successful surveillance strategies are key to protecting HCWs against SARS-CoV-2, especially in settings with limited vaccination capacities or reduced vaccine efficacy.

Hepatitis-D Virus Infection Is Not Impaired by Innate Immunity but Increases Cytotoxic T-Cell Activity.

Approximately 70 million humans worldwide are affected by chronic hepatitis D, which rapidly leads to liver cirrhosis and hepatocellular carcinoma due to chronic inflammation. The triggers and consequences of this chronic inflammation, induced by co-infection with the hepatitis D virus (HDV) and the hepatitis B virus (HBV), are poorly understood. Using CRISPR technology, we characterized the recognition of HDV mono- and co-infection by intracellular innate immunity and determined its influence on the viral life cycle and effector T-cell responses using different HBV and HDV permissive hepatoma cell lines. We showed that HDV infection is detected by MDA5 and -after a lag phase -induces a profound type I interferon response in the infected cells. The type I interferon response, however, was not able to suppress HDV replication or spread, thus providing a persistent trigger. Using engineered T-cells directed against the envelope proteins commonly used by HBV and HDV, we found that HDV immune recognition enhanced T-cell cytotoxicity. Interestingly, the T-cell effector function was enhanced independently of antigen presentation. These findings help to explain immune mediated tissue damage in chronic hepatitis D patients and indicate that combining innate triggers with T-cell activating therapies might allow for a curative approach.

Phosphoproteome profiling uncovers a key role for CDKs in TNF signaling.

Tumor necrosis factor (TNF) is one of the few cytokines successfully targeted by therapies against inflammatory diseases. However, blocking this well studied and pleiotropic ligand can cause dramatic side-effects. Here, we reason that a systems-level proteomic analysis of TNF signaling could dissect its diverse functions and offer a base for developing more targeted therapies. Therefore, we combine phosphoproteomics time course experiments with subcellular localization and kinase inhibitor analysis to identify functional modules of protein phosphorylation. The majority of regulated phosphorylation events can be assigned to an upstream kinase by inhibiting master kinases. Spatial proteomics reveals phosphorylation-dependent translocations of hundreds of proteins upon TNF stimulation. Phosphoproteome analysis of TNF-induced apoptosis and necroptosis uncovers a key role for transcriptional cyclin-dependent kinase activity to promote cytokine production and prevent excessive cell death downstream of the TNF signaling receptor. This resource of TNF-induced pathways and sites can be explored at http://tnfviewer.biochem.mpg.de/ .

Human NLRP1 is a sensor for double-stranded RNA.

Inflammasomes function as intracellular sensors of pathogen infection or cellular perturbation and thereby play a central role in numerous diseases. Given the high abundance of NLRP1 in epithelial barrier tissues, we screened a diverse panel of viruses for inflammasome activation in keratinocytes. We identified Semliki Forest virus (SFV), a positive-strand RNA virus, as a potent activator of human but not murine NLRP1B. SFV replication and the associated formation of double-stranded (ds) RNA was required to engage the NLRP1 inflammasome. Moreover, delivery of long dsRNA was sufficient to trigger activation. Biochemical studies revealed that NLRP1 binds dsRNA through its leucine-rich repeat domain, resulting in its NACHT domain gaining adenosine triphosphatase activity. Altogether, these results establish human NLRP1 as a direct sensor for dsRNA and thus RNA virus infection.

C-tag TNF: a reporter system to study TNF shedding.

TNF is a highly pro-inflammatory cytokine that contributes not only to the regulation of immune responses but also to the development of severe inflammatory diseases. TNF is synthesized as a transmembrane protein, which is further matured via proteolytic cleavage by metalloproteases such as ADAM17, a process known as shedding. At present, TNF is mainly detected by measuring the precursor or the mature cytokine of bulk cell populations by techniques such as ELISA or immunoblotting. However, these methods do not provide information on the exact timing and extent of TNF cleavage at single-cell resolution and they do not allow the live visualization of shedding events. Here, we generated C-tag TNF as a genetically encoded reporter to study TNF shedding at the single-cell level. The functionality of the C-tag TNF reporter is based on the exposure of a cryptic epitope on the C terminus of the transmembrane portion of pro-TNF on cleavage. In both denatured and nondenatured samples, this epitope can be detected by a nanobody in a highly sensitive and specific manner only upon TNF shedding. As such, C-tag TNF can successfully be used for the detection of TNF cleavage in flow cytometry and live-cell imaging applications. We furthermore demonstrate its applicability in a forward genetic screen geared toward the identification of genetic regulators of TNF maturation. In summary, the C-tag TNF reporter can be employed to gain novel insights into the complex regulation of ADAM-dependent TNF shedding.

CARD8 inflammasome activation triggers pyroptosis in human T cells.

Inflammasomes execute a unique type of cell death known as pyroptosis. Mostly characterized in myeloid cells, caspase-1 activation downstream of an inflammasome sensor results in the cleavage and activation of gasdermin D (GSDMD), which then forms a lytic pore in the plasma membrane. Recently, CARD8 was identified as a novel inflammasome sensor that triggers pyroptosis in myeloid leukemia cells upon inhibition of dipeptidyl-peptidases (DPP). Here, we show that blocking DPPs using Val-boroPro triggers a lytic form of cell death in primary human CD4 and CD8 T cells, while other prototypical inflammasome stimuli were not active. This cell death displays morphological and biochemical hallmarks of pyroptosis. By genetically dissecting candidate components in primary T cells, we identify this response to be dependent on the CARD8-caspase-1-GSDMD axis. Moreover, DPP9 constitutes the relevant DPP restraining CARD8 activation. Interestingly, this CARD8-induced pyroptosis pathway can only be engaged in resting, but not in activated T cells. Altogether, these results broaden the relevance of inflammasome signaling and associated pyroptotic cell death to T cells, central players of the adaptive immune system.

Hepatitis B Virus DNA is a Substrate for the cGAS/STING Pathway but is not Sensed in Infected Hepatocytes.

Hepatitis B virus (HBV) chronic infection is a critical risk factor for hepatocellular carcinoma. The innate immune response to HBV infection is a matter of debate. In particular, viral escape mechanisms are poorly understood. Our study reveals that HBV RNAs are not immunostimulatory in immunocompetent myeloid cells. In contrast, HBV DNA from viral particles and DNA replication intermediates are immunostimulatory and sensed by cyclic GMP-AMP Synthase (cGAS) and Stimulator of Interferon Genes (STING). We show that primary human hepatocytes express DNA sensors to reduced levels compared to myeloid cells. Nevertheless, hepatocytes can respond to HBV relaxed-circular DNA (rcDNA), when transfected in sufficient amounts, but not to HBV infection. Finally, our data suggest that HBV infection does not actively inhibit the DNA-sensing pathway. In conclusion, in infected hepatocytes, HBV passively evades recognition by cellular sensors of nucleic acids by (i) producing non-immunostimulatory RNAs, (ii) avoiding sensing of its DNAs by cGAS/STING without active inhibition of the pathway.