Human cytomegalovirus exploits STING signaling and counteracts IFN/ISG induction to facilitate infection of dendritic cells.

Human cytomegalovirus (HCMV) is a widespread pathogen that in immunocompromised hosts can cause life-threatening disease. Studying HCMV-exposed monocyte-derived dendritic cells by single-cell RNA sequencing, we observe that most cells are entered by the virus, whereas less than 30% of them initiate viral gene expression. Increased viral gene expression is associated with activation of the stimulator of interferon genes (STING) that usually induces anti-viral interferon responses, and with the induction of several pro- (RHOB, HSP1A1, DNAJB1) and anti-viral (RNF213, TNFSF10, IFI16) genes. Upon progression of infection, interferon-beta but not interferon-lambda transcription is inhibited. Similarly, interferon-stimulated gene expression is initially induced and then shut off, thus further promoting productive infection. Monocyte-derived dendritic cells are composed of 3 subsets, with one being especially susceptible to HCMV. In conclusion, HCMV permissiveness of monocyte-derived dendritic cells depends on complex interactions between virus sensing, regulation of the interferon response, and viral gene expression.

Secreted NS1 proteins of tick-borne encephalitis virus and West Nile virus block dendritic cell activation and effector functions.

The flavivirus non-structural protein 1 (NS1) is secreted from infected cells into the circulation and the serum levels correlate with disease severity. The effect of secreted NS1 (sNS1) on non-infected mammalian immune cells is largely unknown. Here, we expressed recombinant sNS1 proteins of tick-borne encephalitis virus (TBEV) and West Nile virus (WNV) and investigated their effects on dendritic cell (DC) effector functions. Murine bone marrow-derived DCs (BMDCs) showed reduced surface expression of co-stimulatory molecules and decreased release of pro-inflammatory cytokines when treated with sNS1 of TBEV or WNV prior to poly(I:C) stimulation. Transcriptional profiles of BMDCs that were sNS1-exposed prior to poly(I:C) stimulation showed two gene clusters that were downregulated by TBEV or WNV sNS1 and that were associated with innate and adaptive immune responses. Functionally, both sNS1 proteins modulated the capacity for BMDCs to induce specific T-cell responses as indicated by reduced IFN-γ levels in both CD4+ and CD8+ T cells after BMDC co-cultivation. In human monocyte-derived DCs, poly(I:C)-induced upregulation of co-stimulatory molecules and cytokine responses were even more strongly impaired by TBEV sNS1 or WNV sNS1 pretreatment than in the murine system. Our findings indicate that exogenous flaviviral sNS1 proteins interfere with DC-mediated stimulation of T cells, which is crucial for the initiation of cell-mediated adaptive immune responses in human flavivirus infections. Collectively, our data determine soluble flaviviral NS1 as a virulence factor responsible for a dampened immune response to flavivirus infections. IMPORTANCE The effective initiation of protective host immune responses controls the outcome of infection, and dysfunctional T-cell responses have previously been associated with symptomatic human flavivirus infections. We demonstrate that secreted flavivirus NS1 proteins modulate innate immune responses of uninfected bystander cells. In particular, sNS1 markedly reduced the capacity of dendritic cells to stimulate T-cell responses upon activation. Hence, by modulating cellular host responses that are required for effective antigen presentation and initiation of adaptive immunity, sNS1 proteins may contribute to severe outcomes of flavivirus disease.

A circular RNA derived from the insulin receptor locus protects against doxorubicin-induced cardiotoxicity.

AIMS: Cardiotoxicity leading to heart failure (HF) is a growing problem in many cancer survivors. As specific treatment strategies are not available, RNA discovery pipelines were employed and a new and powerful circular RNA (circRNA)-based therapy was developed for the treatment of doxorubicin-induced HF. METHODS AND RESULTS: The circRNA sequencing was applied and the highly species-conserved circRNA insulin receptor (Circ-INSR) was identified, which participates in HF processes, including those provoked by cardiotoxic anti-cancer treatments. Chemotherapy-provoked cardiotoxicity leads to the down-regulation of Circ-INSR in rodents and patients, which mechanistically contributes to cardiomyocyte cell death, cardiac dysfunction, and mitochondrial damage. In contrast, Circ-INSR overexpression prevented doxorubicin-mediated cardiotoxicity in both rodent and human cardiomyocytes in vitro and in a mouse model of chronic doxorubicin cardiotoxicity. Breast cancer type 1 susceptibility protein (Brca1) was identified as a regulator of Circ-INSR expression. Detailed transcriptomic and proteomic analyses revealed that Circ-INSR regulates apoptotic and metabolic pathways in cardiomyocytes. Circ-INSR physically interacts with the single-stranded DNA-binding protein (SSBP1) mediating its cardioprotective effects under doxorubicin stress. Importantly, in vitro transcribed and circularized Circ-INSR mimics also protected against doxorubicin-induced cardiotoxicity. CONCLUSION: Circ-INSR is a highly conserved non-coding RNA which is down-regulated during cardiotoxicity and cardiac remodelling. Adeno-associated virus and circRNA mimics-based Circ-INSR overexpression prevent and reverse doxorubicin-mediated cardiomyocyte death and improve cardiac function. The results of this study highlight a novel and translationally important Circ-INSR-based therapeutic approach for doxorubicin-induced cardiac dysfunction.

Toll-like Receptors in Viral Encephalitis.

Viral encephalitis is a rare but serious syndrome. In addition to DNA-encoded herpes viruses, such as herpes simplex virus and varicella zoster virus, RNA-encoded viruses from the families of Flaviviridae, Rhabdoviridae and Paramyxoviridae are important neurotropic viruses. Whereas in the periphery, the role of Toll-like receptors (TLR) during immune stimulation is well understood, TLR functions within the CNS are less clear. On one hand, TLRs can affect the physiology of neurons during neuronal progenitor cell differentiation and neurite outgrowth, whereas under conditions of infection, the complex interplay between TLR stimulated neurons, astrocytes and microglia is just on the verge of being understood. In this review, we summarize the current knowledge about which TLRs are expressed by cell subsets of the CNS. Furthermore, we specifically highlight functional implications of TLR stimulation in neurons, astrocytes and microglia. After briefly illuminating some examples of viral evasion strategies from TLR signaling, we report on the current knowledge of primary immunodeficiencies in TLR signaling and their consequences for viral encephalitis. Finally, we provide an outlook with examples of TLR agonist mediated intervention strategies and potentiation of vaccine responses against neurotropic virus infections.

Sequential MAVS and MyD88/TRIF signaling triggers anti-viral responses of tick-borne encephalitis virus-infected murine astrocytes.

Tick-borne encephalitis virus (TBEV), a member of the Flaviviridae family, is typically transmitted upon tick bite and can cause meningitis and encephalitis in humans. In TBEV-infected mice, mitochondrial antiviral-signaling protein (MAVS), the downstream adaptor of retinoic acid-inducible gene-I (RIG-I)-like receptor (RLR) signaling, is needed to induce early type I interferon (IFN) responses and to confer protection. To characterize the brain-resident cell subset that produces protective IFN-ß in TBEV-infected mice, we isolated neurons, astrocytes, and microglia from mice and exposed these cell types to TBEV in vitro. Under such conditions, neurons showed the highest percentage of infected cells, whereas astrocytes and microglia were infected to a lesser extent. In the supernatant (SN) of infected neurons, IFN-ß was not detectable, while infected astrocytes showed high and microglia low IFN-ß expression. Transcriptome analyses of astrocytes implied that MAVS signaling was needed early after TBEV infection. Accordingly, MAVS-deficient astrocytes showed enhanced TBEV infection and significantly reduced early IFN-ß responses. Nevertheless, at later time points, moderate amounts of IFN-ß were detected in the SN of infected MAVS-deficient astrocytes. Transcriptome analyses indicated that MAVS deficiency negatively affected the induction of early anti-viral responses, which resulted in significantly increased TBEV replication. Treatment with MyD88 and TRIF inhibiting peptides reduced only late IFN-ß responses of TBEV-infected WT astrocytes and blocked entirely IFN-ß responses of infected MAVS-deficient astrocytes. Thus, upon TBEV exposure of brain-resident cells, astrocytes are important IFN-ß producers showing biphasic IFN-ß induction that initially depends on MAVS and later on MyD88/TRIF signaling.