Dengue virus (DENV) is a mosquito-borne flavivirus, and symptoms of infection range from asymptomatic to the severe dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). High viral loads correlate with disease severity, and both type I & II interferons (IFNs) are crucial for controlling viral replication. We have previously reported that signal transducer and activator of transcription (STAT) 1-deficient mice are resistant to DENV-induced disease, but little is known about this STAT1-independent mechanism of protection. To determine the molecular basis of the STAT1-independent pathway, mice lacking STAT1, STAT2, or both STAT1 and STAT2 were infected with a virulent mouse-adapted strain of DENV2. In the first 72 hours of infection, the single-deficient mice lacking STAT1 or STAT2 possessed 50–100 fold higher levels of viral RNA than wild type mice in the serum, spleen, and other visceral tissues, but remained resistant to DENV-induced death. In contrast, the double-deficient mice exhibited the early death phenotype previously observed in type I and II IFN receptor knockout mice (AG129), indicating that STAT2 is the mediator of the STAT1-independent host defense mechanism. Further studies demonstrated that this STAT2-dependent STAT1-independent mechanism requires the type I IFN receptor, and contributes to the autocrine amplification of type I IFN expression. Examination of gene expression in the spleen and bone marrow-derived macrophages following DENV infection revealed STAT2-dependent pathways can induce the transcription of a subset of interferon stimulated genes even in the absence of STAT1. Collectively, these results help elucidate the nature of the poorly understood STAT1-independent host defense mechanism against viruses by identifying a functional type I IFN/STAT2 signaling pathway following DENV infection in vivo.

Previous studies have shown that human cytomegalovirus (CMV) is a potent elicitor of interferon-stimulated gene (ISG) expression. Induction of the interferon pathway does not require replication-competent virus, and envelope glycoprotein B (gB) from CMV is a viral structural component that can directly induce transcription of ISGs. Here we extend these earlier findings by defining the consequences of inducing the interferon pathway. We found that cells respond to CMV or soluble gB by establishing a functional antiviral state within cell types critical in CMV biology, such as fibroblasts and endothelial cells. We have also discovered new insights into the mechanism by which the pathway is initiated. Interferon regulatory factor 3 (IRF3), a key transcriptional regulator of cellular interferon responses, is activated by CMV virions and soluble gB. Thus, IRF3 becomes activated via "outside-in" signal transduction events. This is a novel mechanism of activation of this key transcription factor by viruses. In comparison to soluble gB (gB 1-750 ), which comprises the entire ectodomain of gB, a truncation mutant encompassing only the amino-terminal region of gB (gB 1-460 ) was markedly less effective at inducing antiviral responses. This indicates that the region of gB from residues 461 to 750 is important for initiation of the antiviral response. In addition, CMV and gB establish an antiviral state in alpha/beta interferon null cells, illustrating that primary induction of ISGs by CMV and gB is sufficient to establish the antiviral response and that interferon secretion is not necessary for the antiviral effect. Taken together, our findings reveal that CMV initiates a coordinated antiviral response through contact between gB and an as-yet-unidentified cell surface receptor(s).

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. In contrast, DENV viral loads in nonlymphoid tissues were similar between Cardif ؊/؊ and wild-type mice. These results reveal that RNA helicase-mediated sensing acts as a first line of innate defense against DENV infection in vivo and functions in a tissue-dependent manner.Dengue virus (DENV) is a positive-sense, single-stranded RNA virus that belongs to the family Flaviviridae, which includes other arthropod-borne viruses such as Japanese encephalitis, yellow fever, and West Nile viruses (12). DENV is transmitted to humans by mosquitoes and causes an estimated 50 million new cases of dengue fever and 250,000 cases of dengue hemorrhagic fever/dengue shock syndrome per year worldwide (3). Despite the prevalence of DENV, the virus-host interactions that ultimately determine the disease remain largely uncharacterized.Several lines of evidence implicate a key role for type I interferon (IFN) in protection against DENV infection, as follows. (i) DENV-infected patients contain a high level of alpha interferon (IFN-␣) in the serum (11). (ii) Human peripheral blood mononuclear cells exposed to DENV-infected monocytes secrete IFN-␣ and protect other human monocytes from DENV infection (10). (iii) IFN-␣/␤ inhibits DENV infection in a variety of human and nonhuman cells (1, 2). (iv) Mice lacking receptors for both IFN-␣/␤ and IFN-␥, but not wild-type (WT) control animals, succumb to primary DENV infection (7,22). (v) Expression of DENV nonstructural proteins NS2A, NS2B, and NS4B inhibit IFN-␤-induced activation of STAT1 in a monkey cell line (15, 16). However, little is known about the mechanisms by which the anti-DENV IFN response is initiated in vivo.The retinoic acid-inducible gene I (RIG-I) protein (5), which is composed of a DExD/H-box RNA helicase domain fused to a caspase recruitment domain (CARD), mediates induction of type I IFN in response to RNA virus infection (28). Two other RIG-I-like receptors are currently known, MDA5 and LGP2, and these and RIG-I reside in the cell cytoplasm and interact with viral RNA via their helicase domains. Upon RNA binding, the CARD domain of RIG-I/ MDA5 interacts with the CARD of Cardif/MAVS/IPS-1/ VISA, ultimately leading to type I IFN production (4). DENV recognition by both RIG-I and MDA5 and downstream Cardif-dependent signaling have been recently shown in cultured fibroblasts (13), suggesting that RIG-I-like receptor signaling affects DENV replication in vivo.

Interferons, which transduce pivotal signals through signal transducer and activator of transcription (Stat)1 and Stat2, effectively suppress the replication of Legionella pneumophila in primary murine macrophages. Whereas the ability of IFN-γ to impede L. pneumophila growth is fully dependent on Stat1, IFN-α/β unexpectedly suppresses L. pneumophila growth in both Stat1 and Stat2 deficient macrophages. New studies demonstrating that the robust response to IFN-α/β is lost in Stat1-Stat2 double knockout macrophages, suggest that Stat1 and Stat2 are functionally redundant in their ability to direct an innate response towards L. pneumophila. Since the ability of IFN-α/β to signal through Stat1-dependent complexes (i.e., Stat1-Stat1 and Stat1-Stat2 dimers) has been well characterized, the current studies focus on how Stat2 is able to direct a potent response to IFN-α/β in the absence of Stat1. These studies reveal that IFN-α/β is able to drive the formation of a Stat2 and IRF9 complex that drives the expression of a subset of IFN stimulated genes (ISGs), but with substantially delayed kinetics. These observations raise the possibility that this pathway evolved in response to microbes that have devised strategies to subvert Stat1 dependent responses.