Autocrine IFN-I inhibits isocitrate dehydrogenase in the TCA cycle of LPS-stimulated macrophages.

Macrophage activation in response to LPS is coupled to profound metabolic changes, typified by accumulation of the TCA cycle intermediates citrate, itaconate, and succinate. We have identified that endogenous type I IFN controls the cellular citrate/α-ketoglutarate ratio and inhibits expression and activity of isocitrate dehydrogenase (IDH); and, via 13C-labeling studies, demonstrated that autocrine type I IFN controls carbon flow through IDH in LPS-activated macrophages. We also found that type I IFN-driven IL-10 contributes to inhibition of IDH activity and itaconate synthesis in LPS-stimulated macrophages. Our findings have identified the autocrine type I IFN pathway as being responsible for the inhibition of IDH in LPS-stimulated macrophages.

Microbiota-Driven Tonic Interferon Signals in Lung Stromal Cells Protect from Influenza Virus Infection.

Type I interferon (IFNα/ß) pathways are fine-tuned to elicit antiviral protection while minimizing immunopathology; however, the initiating stimuli, target tissues, and underlying mechanisms are unclear. Using models of physiological and dysregulated IFNα/ß receptor (IFNAR1) surface expression, we show here that IFNAR1-dependent signals set the steady-state IFN signature in both hematopoietic and stromal cells. Increased IFNAR1 levels promote a lung environment refractory to early influenza virus replication by elevating the baseline interferon signature. Commensal microbiota drive the IFN signature specifically in lung stroma, as shown by antibiotic treatment and fecal transplantation. Bone marrow chimera experiments identify lung stromal cells as crucially important for early antiviral immunity and stroma-immune cell interaction for late antiviral resistance. We propose that the microbiota-driven interferon signature in lung epithelia impedes early virus replication and that IFNAR1 surface levels fine-tune this signature. Our findings highlight the interplay between bacterial and viral exposure, with important implications for antibiotic use.

An Update on Autoinflammatory Diseases: Relopathies.

PURPOSE OF REVIEW: The nuclear factor κB (NF-κB) pathway is tightly regulated through multiple posttranslational mechanisms including ubiquitination. Mutations in these regulatory pathways can cause disease and are the focus of this review. RECENT FINDINGS: The linear ubiquitin chain assembly complex (LUBAC) is a trimer made up of HOIL-1L, SHARPIN, and the catalytic subunit HOIP. Loss of function mutations in HOIL-1L and HOIP result in largely overlapping phenotypes, characterized by multi-organ autoinflammation, immunodeficiency, and amylopectinosis. Interestingly, patient fibroblasts exhibited diminished IL-1ß- and TNF-induced NF-κB activation, yet monocytes were hyper-responsive to IL-1ß, hinting at cell type or target specific roles of LUBAC-mediated ubiquitination. Ubiquitin-driven signaling is counterbalanced by deubiquitinase enzymes (DUBs), such as OTULIN and A20. Hypomorphic mutations in OTULIN result in elevated NF-κB signaling causing an autoinflammatory syndrome. Similarly, patients with high-penetrance heterozygous mutations in the gene encoding A20 (haploinsufficiency of A20 (HA20)) display excessive ubiquitination and increased activity of NF-κB and of NLRP3 inflammasome activation. HA20 patients present with Behçet-like characteristics or an autoimmune lymphoproliferative syndrome (ALPS)-like phenotype, indicating diverse protein functions. This review summarizes recent discoveries in the field of NF-kB-related autoinflammatory diseases (relopathies) within the past 3 years and points to several questions that still remain unanswered.

Natural amines inhibit activation of human plasmacytoid dendritic cells through CXCR4 engagement.

Plasmacytoid dendritic cells (pDC) are specialized in secretion of type I interferon in response to pathogens. Here we show that natural monoamines and synthetic amines inhibit pDC activation by RNA viruses. Furthermore, a synthetic analogue of histamine reduces type I interferon production in a mouse model of influenza infection. We identify CXC chemokine receptor 4 (CXCR4) as a receptor used by amines to inhibit pDC. Our study establishes a functional link between natural amines and the innate immune system and identifies CXCR4 as a potential 'on-off' switch of pDC activity with therapeutic potential.

IFNλ is a potent anti-influenza therapeutic without the inflammatory side effects of IFNα treatment.

Influenza A virus (IAV)-induced severe disease is characterized by infected lung epithelia, robust inflammatory responses and acute lung injury. Since type I interferon (IFNαß) and type III interferon (IFNλ) are potent antiviral cytokines with immunomodulatory potential, we assessed their efficacy as IAV treatments. IFNλ treatment of IAV-infected Mx1-positive mice lowered viral load and protected from disease. IFNα treatment also restricted IAV replication but exacerbated disease. IFNα treatment increased pulmonary proinflammatory cytokine secretion, innate cell recruitment and epithelial cell death, unlike IFNλ-treatment. IFNλ lacked the direct stimulatory activity of IFNα on immune cells. In epithelia, both IFNs induced antiviral genes but no inflammatory cytokines. Similarly, human airway epithelia responded to both IFNα and IFNλ by induction of antiviral genes but not of cytokines, while hPBMCs responded only to IFNα. The restriction of both IFNλ responsiveness and productive IAV replication to pulmonary epithelia allows IFNλ to limit IAV spread through antiviral gene induction in relevant cells without overstimulating the immune system and driving immunopathology. We propose IFNλ as a non-inflammatory and hence superior treatment option for human IAV infection.

TRAIL+ monocytes and monocyte-related cells cause lung damage and thereby increase susceptibility to influenza-Streptococcus pneumoniae coinfection.

Streptococcus pneumoniae coinfection is a major cause of influenza-associated mortality; however, the mechanisms underlying pathogenesis or protection remain unclear. Using a clinically relevant mouse model, we identify immune-mediated damage early during coinfection as a new mechanism causing susceptibility. Coinfected CCR2(-/-) mice lacking monocytes and monocyte-derived cells control bacterial invasion better, show reduced epithelial damage and are overall more resistant than wild-type controls. In influenza-infected wild-type lungs, monocytes and monocyte-derived cells are the major cell populations expressing the apoptosis-inducing ligand TRAIL. Accordingly, anti-TRAIL treatment reduces bacterial load and protects against coinfection if administered during viral infection, but not following bacterial exposure. Post-influenza bacterial outgrowth induces a strong proinflammatory cytokine response and massive inflammatory cell infiltrate. Depletion of neutrophils or blockade of TNF-α facilitate bacterial outgrowth, leading to increased mortality, demonstrating that these factors aid bacterial control. We conclude that inflammatory monocytes recruited early, during the viral phase of coinfection, induce TRAIL-mediated lung damage, which facilitates bacterial invasion, while TNF-α and neutrophil responses help control subsequent bacterial outgrowth. We thus identify novel determinants of protection versus pathology in influenza-Streptococcus pneumoniae coinfection.

Themis2 is not required for B cell development, activation, and antibody responses.

Themis1 is a protein implicated in transducing signals from the TCR. Mice deficient in Themis1 show a strong impairment in T cell selection in the thymus and defective T cell activation. The related Themis2 protein is expressed in B cells where it associates with signaling proteins Grb2 and Vav1, and is tyrosine phosphorylated after BCR stimulation. Thus, it has been proposed that Themis2 may transduce BCR signals, and hence play important roles in B cell development and activation. In this article, we show that Themis2 is expressed in all developing subsets of B cells, in mature follicular and marginal zone B cells, and in activated B cells, including germinal center B cells and plasma cells. In contrast, B lineage cells express no other Themis-family genes. Activation of B cells leads to reduced Themis2 expression, although it remains the only Themis-family protein expressed. To analyze the physiological function of Themis2, we generated a Themis2-deficient mouse strain. Surprisingly, we found that Themis2 is not required for B cell development, for activation, or for Ab responses either to model Ags or to influenza viral infection.

Pathogenic potential of interferon αß in acute influenza infection.

Influenza symptoms vary from mild disease to death; however, determinants of severity are unclear. Type I interferons (IFNαß) are recognized as key antiviral cytokines. Here we show that, surprisingly, influenza-infected 129 mice have increased lung damage, morbidity and mortality, yet higher levels of IFNαß, than C57BL/6 mice. Consistently, IFNα treatment of influenza-infected C57BL/6 mice increases morbidity. IFNαß receptor deficiency in 129 mice decreases morbidity, lung damage, proinflammatory cytokines and lung-infiltrating inflammatory cells, and reduces expression of the death-inducing receptor DR5 on lung epithelia and its ligand TRAIL on inflammatory monocytes. Depletion of PDCA-1+ cells or interruption of TRAIL-DR5 interaction protects infected 129 mice. Selective lack of IFNαß signalling in stromal cells abolishes epithelial DR5 upregulation and apoptosis, reducing host susceptibility. Hence, excessive IFNαß signalling in response to acute influenza infection can result in uncontrolled inflammation and TRAIL-DR5-mediated epithelial cell death, which may explain morbidity and has important implications for treatment of severe disease.

The transcription factor E4BP4 is not required for extramedullary pathways of NK cell development.

NK cells contribute to antitumor and antiviral immunosurveillance. Their development in the bone marrow (BM) requires the transcription factor E4BP4/NFIL3, but requirements in other organs are less well defined. In this study, we show that CD3(-)NK1.1(+)NKp46(+)CD122(+) NK cells of immature phenotype and expressing low eomesodermin levels are found in thymus, spleen, and liver of E4BP4-deficient mice, whereas numbers of mature, eomesodermin(high) conventional NK cells are drastically reduced. E4BP4-deficient CD44(+)CD25(-) double-negative 1 thymocytes efficiently develop in vitro into NK cells with kinetics, phenotype, and functionality similar to wild-type controls, whereas no NK cells develop from E4BP4-deficient BM precursors. In E4BP4/Rag-1 double-deficient (DKO) mice, NK cells resembling those in Rag-1-deficient controls are found in similar numbers in the thymus and liver. However, NK precursors are reduced in DKO BM, and no NK cells develop from DKO BM progenitors in vitro. DKO thymocyte precursors readily develop into NK cells, but DKO BM transfers into nude recipients and NK cells in E4BP4/Rag-1/IL-7 triple-KO mice indicated thymus-independent NK cell development. In the presence of T cells or E4BP4-sufficient NK cells, DKO NK cells have a selective disadvantage, and thymic and hepatic DKO NK cells show reduced survival when adoptively transferred into lymphopenic hosts. This correlates with higher apoptosis rates and lower responsiveness to IL-15 in vitro. In conclusion, we demonstrate E4BP4-independent development of NK cells of immature phenotype, reduced fitness, short t1/2, and potential extramedullary origin. Our data identify E4BP4-independent NK cell developmental pathways and a role for E4BP4 in NK cell homeostasis.

Type I and type III interferons drive redundant amplification loops to induce a transcriptional signature in influenza-infected airway epithelia.

Interferons (IFNs) are a group of cytokines with a well-established antiviral function. They can be induced by viral infection, are secreted and bind to specific receptors on the same or neighbouring cells to activate the expression of hundreds of IFN stimulated genes (ISGs) with antiviral function. Type I IFN has been known for more than half a century. However, more recently, type III IFN (IFNλ, IL-28/29) was shown to play a similar role and to be particularly important at epithelial surfaces. Here we show that airway epithelia, the primary target of influenza A virus, produce both IFN I and III upon infection, and that induction of both depends on the RIG-I/MAVS pathway. While IRF3 is generally regarded as the transcription factor required for initiation of IFN transcription and the so-called "priming loop", we find that IRF3 deficiency has little impact on IFN expression. In contrast, lack of IRF7 reduced IFN production significantly, and only IRF3(-/-)IRF7(-/-) double deficiency completely abolished it. The transcriptional response to influenza infection was largely dependent on IFNs, as it was reduced to a few upregulated genes in epithelia lacking receptors for both type I and III IFN (IFNAR1(-/-)IL-28Rα(-/-)). Wild-type epithelia and epithelia deficient in either the type I IFN receptor or the type III IFN receptor exhibit similar transcriptional profiles in response to virus, indicating that none of the induced genes depends selectively on only one IFN system. In chimeric mice, the lack of both IFN I and III signalling in the stromal compartment alone significantly increased the susceptibility to influenza infection. In conclusion, virus infection of airway epithelia induces, via a RIG-I/MAVS/IRF7 dependent pathway, both type I and III IFNs which drive two completely overlapping and redundant amplification loops to upregulate ISGs and protect from influenza infection.