-
Helen M Lazear,
Alissa Lancaster,
Courtney Wilkins,
Mehul S Suthar,
Albert Huang,
Sarah C Vick,
Lisa Clepper,
Larissa Thackray,
Margaret M Brassil,
Herbert W Virgin,
Janko Nikolich-Zugich,
Ashlee V Moses, Michael Gale,
Klaus Früh,
Michael S Diamond
PLoS Pathogens 05/2013; 9(5). · 9.13 Impact Factor
-
Amina A Negash,
Hilario J Ramos,
Nanette Crochet,
Daryl T Y Lau,
Brian Doehle,
Neven Papic,
Don A Delker,
Juandy Jo,
Antonio Bertoletti,
Curt H Hagedorn, Michael Gale
[show abstract]
[hide abstract]
ABSTRACT: Chronic hepatitis C virus (HCV) infection is a leading cause of liver disease. Liver inflammation underlies infection-induced fibrosis, cirrhosis and liver cancer but the processes that promote hepatic inflammation by HCV are not defined. We provide a systems biology analysis with multiple lines of evidence to indicate that interleukin-1β (IL-1β) production by intrahepatic macrophages confers liver inflammation through HCV-induced inflammasome signaling. Chronic hepatitis C patients exhibited elevated levels of serum IL-1β compared to healthy controls. Immunohistochemical analysis of healthy control and chronic hepatitis C liver sections revealed that Kupffer cells, resident hepatic macrophages, are the primary cellular source of hepatic IL-1β during HCV infection. Accordingly, we found that both blood monocyte-derived primary human macrophages, and Kupffer cells recovered from normal donor liver, produce IL-1β after HCV exposure. Using the THP-1 macrophage cell-culture model, we found that HCV drives a rapid but transient caspase-1 activation to stimulate IL-1β secretion. HCV can enter macrophages through non-CD81 mediated phagocytic uptake that is independent of productive infection. Viral RNA triggers MyD88-mediated TLR7 signaling to induce IL-1β mRNA expression. HCV uptake concomitantly induces a potassium efflux that activates the NLRP3 inflammasome for IL-1β processing and secretion. RNA sequencing analysis comparing THP1 cells and chronic hepatitis C patient liver demonstrates that viral engagement of the NLRP3 inflammasome stimulates IL-1β production to drive proinflammatory cytokine, chemokine, and immune-regulatory gene expression networks linked with HCV disease severity. These studies identify intrahepatic IL-1β production as a central feature of liver inflammation during HCV infection. Thus, strategies to suppress NLRP3 or IL-1β activity could offer therapeutic actions to reduce hepatic inflammation and mitigate disease.
PLoS Pathogens 04/2013; 9(4):e1003330. · 9.13 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Plasmacytoid Dendritic Cells (pDCs) represent a key immune cell in the defense against viruses. Through pattern recognition receptors (PRRs), these cells detect viral pathogen associated molecular patterns (PAMPs) and initiate an Interferon (IFN) response. pDCs produce the antiviral IFNs including the well-studied Type I and the more recently described Type III. Recent genome wide association studies (GWAS) have implicated Type III IFNs in HCV clearance. We examined the IFN response induced in a pDC cell line and ex vivo human pDCs by a region of the HCV genome referred to as the HCV PAMP. This RNA has been shown previously to be immunogenic in hepatocytes, whereas the conserved X-region RNA is not. We show that in response to the HCV PAMP, pDC-GEN2.2 cells upregulate and secrete Type III (in addition to Type I) IFNs and upregulate PRR genes and proteins. We also demonstrate that the recognition of this RNA is dependent on RIG-I-like Receptors (RLRs) and Toll-like Receptors (TLRs), challenging the dogma that RLRs are dispensable in pDCs. The IFNs produced by these cells in response to the HCV PAMP also control HCV replication in vitro. These data are recapitulated in ex vivo pDCs isolated from healthy donors. Together, our data shows that pDCs respond robustly to HCV RNA to make Type III Interferons that control viral replication. This may represent a novel therapeutic strategy for the treatment of HCV.
PLoS Pathogens 04/2013; 9(4):e1003316. · 9.13 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Although susceptibility of neurons in the brain to microbial infection is a major determinant of clinical outcome, little is known about the molecular factors governing this vulnerability. Here we show that two types of neurons from distinct brain regions showed differential permissivity to replication of several positive-stranded RNA viruses. Granule cell neurons of the cerebellum and cortical neurons from the cerebral cortex have unique innate immune programs that confer differential susceptibility to viral infection ex vivo and in vivo. By transducing cortical neurons with genes that were expressed more highly in granule cell neurons, we identified three interferon-stimulated genes (ISGs; Ifi27, Irg1 and Rsad2 (also known as Viperin)) that mediated the antiviral effects against different neurotropic viruses. Moreover, we found that the epigenetic state and microRNA (miRNA)-mediated regulation of ISGs correlates with enhanced antiviral response in granule cell neurons. Thus, neurons from evolutionarily distinct brain regions have unique innate immune signatures, which probably contribute to their relative permissiveness to infection.
Nature medicine 03/2013; · 27.14 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The actions of the RIG-I like receptor (RLR) and type I interferon (IFN) signaling pathways are essential for a protective innate immune response against the emerging flavivirus West Nile virus (WNV). In mice lacking RLR or IFN signaling pathways, WNV exhibits enhanced tissue tropism, indicating that specific host factors of innate immune defense restrict WNV infection and dissemination in peripheral tissues. However, the immune mechanisms by which the RLR and IFN pathways coordinate and function to impart restriction of WNV infection are not well defined. Using a systems biology approach, we defined the host innate immune response signature and actions that restrict WNV tissue tropism. Transcriptional profiling and pathway modeling to compare WNV-infected permissive (spleen) and nonpermissive (liver) tissues showed high enrichment for inflammatory responses, including pattern recognition receptors and IFN signaling pathways, that define restriction of WNV replication in the liver. Assessment of infected livers from Mavs(-/-)×Ifnar(-/-) mice revealed the loss of expression of several key components within the natural killer (NK) cell signaling pathway, including genes associated with NK cell activation, inflammatory cytokine production, and NK cell receptor signaling. In vivo analysis of hepatic immune cell infiltrates from WT mice demonstrated that WNV infection leads to an increase in NK cell numbers with enhanced proliferation, maturation, and effector action. In contrast, livers from Mavs(-/-)×Ifnar(-/-) infected mice displayed reduced immune cell infiltration, including a significant reduction in NK cell numbers. Analysis of cocultures of dendritic and NK cells revealed both cell-intrinsic and -extrinsic roles for the RLR and IFN signaling pathways to regulate NK cell effector activity. Taken together, these observations reveal a complex innate immune signaling network, regulated by the RLR and IFN signaling pathways, that drives tissue-specific antiviral effector gene expression and innate immune cellular processes that control tissue tropism to WNV infection.
PLoS Pathogens 02/2013; 9(2):e1003168. · 9.13 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: West Nile virus (WNV) is an emerging neurotropic flavivirus that is transmitted to humans through the bite of an infected mosquito. WNV has disseminated broadly in the Western hemisphere and now poses a significant public health risk. The continuing spread of WNV, combined with the lack of specific therapeutics or vaccines to combat or prevent infection, imparts a pressing need to identify the viral and host processes that control the outcome of and immunity to WNV infection. Here, we provide an overview of recent research that has revealed the virus-host interface controlling WNV infection and immunity.
Nature Reviews Microbiology 02/2013; 11(2):115-28. · 21.18 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: In this issue of Immunity, studies by Blanc et al. (2013) and Liu et al. (2013) reveal how interferon induction of cholesterol-25-hydroxylase mediates innate immunity against multiple diverse viruses.
Immunity 01/2013; 38(1):3-5. · 21.64 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: West Nile Virus (WNV) is a RNA virus of the family Flaviviridae and the leading cause of mosquito-borne encephalitis in the United States. Humoral immunity is essential for protection against WNV infection; however, the requirements for initiating effective antibody responses against WNV infection are still unclear. CD22 (Siglec-2) is expressed on B cells and regulates B cell receptor signaling, cell survival, proliferation and antibody production. Here we investigated how CD22 contributes to protection against WNV infection, and found that CD22 knockout (Cd22(-/-)) mice were highly susceptible to WNV infection and had increased viral loads in the serum and central nervous system (CNS) compared to wild type (WT) mice. This was not due to a defect in humoral immunity as Cd22(-/-) mice had normal WNV-specific antibody responses. However, Cd22(-/-) mice had decreased WNV-specific CD8(+) T cell responses compared to WT mice. These defects were not simply due to reduced cytotoxic activity or increased cell death, but rather were associated with decreased lymphocyte migration into the draining lymph nodes (dLNs) of infected Cd22(-/-) mice. Cd22(-/-) mice had reduced production of the chemokine CCL3 in the dLNs after infection, suggesting that CD22 affects chemotaxis via controlling chemokine production. CD22 was not restricted to B cells but was also expressed on a subset of splenic DCIR2(+) dendritic cells that rapidly expand early after WNV infection. Thus, CD22 plays an essential role in controlling WNV infection by governing cell migration and CD8(+) T cell responses.
Journal of Virology 01/2013; · 5.40 Impact Factor
-
Helen M Lazear,
Alissa Lancaster,
Courtney Wilkins,
Mehul S Suthar,
Albert Huang,
Sarah C Vick,
Lisa Clepper,
Larissa Thackray,
Margaret M Brassil,
Herbert W Virgin,
Janko Nikolich-Zugich,
Ashlee V Moses, Michael Gale,
Klaus Früh,
Michael S Diamond
[show abstract]
[hide abstract]
ABSTRACT: Although the transcription factors IRF-3 and IRF-7 are considered master regulators of type I interferon (IFN) induction and IFN stimulated gene (ISG) expression, Irf3(-/-)×Irf7(-/-) double knockout (DKO) myeloid dendritic cells (mDC) produce relatively normal levels of IFN-β after viral infection. We generated Irf3(-/-)×Irf5(-/-)×Irf7(-/-) triple knockout (TKO) mice to test whether IRF-5 was the source of the residual induction of IFN-β and ISGs in mDCs. In pathogenesis studies with two unrelated positive-sense RNA viruses (West Nile virus (WNV) and murine norovirus), TKO mice succumbed at rates greater than DKO mice and equal to or approaching those of mice lacking the type I IFN receptor (Ifnar(-/-)). In ex vivo studies, after WNV infection or exposure to Toll-like receptor agonists, TKO mDCs failed to produce IFN-β or express ISGs. In contrast, this response was sustained in TKO macrophages following WNV infection. To define IRF-regulated gene signatures, we performed microarray analysis on WNV-infected mDC from wild type (WT), DKO, TKO, or Ifnar(-/-) mice, as well as from mice lacking the RIG-I like receptor adaptor protein MAVS. Whereas the gene induction pattern in DKO mDC was similar to WT cells, remarkably, almost no ISG induction was detected in TKO or Mavs(-/-) mDC. The relative equivalence of TKO and Mavs(-/-) responses suggested that MAVS dominantly regulates ISG induction in mDC. Moreover, we showed that MAVS-dependent induction of ISGs can occur through an IRF-5-dependent yet IRF-3 and IRF-7-independent pathway. Our results establish IRF-3, -5, and -7 as the key transcription factors responsible for mediating the type I IFN and ISG response in mDC during WNV infection and suggest a novel signaling link between MAVS and IRF-5.
PLoS Pathogens 01/2013; 9(1):e1003118. · 9.13 Impact Factor
-
Hilario J Ramos,
Marion C Lanteri,
Gabriele Blahnik,
Amina Negash,
Mehul S Suthar,
Margaret M Brassil,
Khushbu Sodhi,
Piper M Treuting,
Michael P Busch,
Philip J Norris, Michael Gale
[show abstract]
[hide abstract]
ABSTRACT: West Nile virus (WNV) is an emerging flavivirus capable of infecting the central nervous system (CNS) and mediating neuronal cell death and tissue destruction. The processes that promote inflammation and encephalitis within the CNS are important for control of WNV disease but, how inflammatory signaling pathways operate to control CNS infection is not defined. Here, we identify IL-1β signaling and the NLRP3 inflammasome as key host restriction factors involved in viral control and CNS disease associated with WNV infection. Individuals presenting with acute WNV infection displayed elevated levels of IL-1β in their plasma over the course of infection, suggesting a role for IL-1β in WNV immunity. Indeed, we found that in a mouse model of infection, WNV induced the acute production of IL-1β in vivo, and that animals lacking the IL-1 receptor or components involved in inflammasome signaling complex exhibited increased susceptibility to WNV pathogenesis. This outcome associated with increased accumulation of virus within the CNS but not peripheral tissues and was further associated with altered kinetics and magnitude of inflammation, reduced quality of the effector CD8(+) T cell response and reduced anti-viral activity within the CNS. Importantly, we found that WNV infection triggers production of IL-1β from cortical neurons. Furthermore, we found that IL-1β signaling synergizes with type I IFN to suppress WNV replication in neurons, thus implicating antiviral activity of IL-1β within neurons and control of virus replication within the CNS. Our studies thus define the NLRP3 inflammasome pathway and IL-1β signaling as key features controlling WNV infection and immunity in the CNS, and reveal a novel role for IL-1β in antiviral action that restricts virus replication in neurons.
PLoS Pathogens 11/2012; 8(11):e1003039. · 9.13 Impact Factor
-
Daryl T-Y Lau,
Amina Negash,
Jie Chen,
Nanette Crochet,
Mala Sinha,
Yuhong Zhang,
Jeremie Guedj,
Sharon Holder,
Takeshi Saito,
Stanley M Lemon,
Bruce A Luxon,
Alan S Perelson, Michael Gale
[show abstract]
[hide abstract]
ABSTRACT: BACKGROUND & AIMS: In patients with hepatitis C virus (HCV) infection, interferon (IFN)-α alters expression of IFN-stimulated genes (ISGs), but little is understood about factors that determine outcomes of therapy. We used a systems biology approach to evaluate the acute response of patients with chronic hepatitis C to IFN-α therapy. METHODS: We collected liver biopsy samples from 8 treatment-naïve patients with chronic HCV genotype 1 infection at baseline and 24 hours after they received IFN-α2a (10 MUsc). Blood samples were collected before, and up to 48 hours after, IFN-α2a administration, to measure HCV RNA and for gene expression analysis. Patients then received pegylated IFN-α2a and ribavirin on day 5 of the study; therapy continued for up to 48 weeks. RESULTS: Based on kinetics of HCV RNA during the first 12 weeks of therapy, 2 patients were rapid virologic responders, 4 were early virologic responders, and 2 did not respond to therapy (nonresponders). Nonresponders had high pretreatment levels of ISG expression in liver, but not in peripheral blood monocyte cells. In responders, after administration of IFN-α, intrahepatic ISG expression increased significantly from baseline and was associated with rapid phase-1 decrease in HCV. We identified distinct hepatic expression and tissue distribution patterns of ISGs that segregated with treatment outcome. Importantly, Kupffer cells were a local source of IFN that promoted basal expression of ISG in hepatocytes of nonresponders. This finding was validated in cultured THP1 human macrophage that expressed IFN-β after exposure to viable HCV2a. When Huh7 K2040 and Huh7 L2198S hepatoma cells were incubated with IFN-α2a, expression of ISGs peaked by 4 hours and decreased by 72 hours, associated with an increase in level of HCV RNA. This indicates that constitutive exposure to IFN causes hepatoma cells to become tolerant to ISG function. CONCLUSIONS: In patients with chronic HCV infection, IFN production by Kupffer cells might promote innate immune tolerance, characterized by a lack of response to IFN therapy. Strategies to disrupt the virus-host interactions that induce innate immune tolerance should improve therapy.
Gastroenterology 11/2012; · 11.68 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Type 1 interferon (IFN) continues to be the foundation for the current standard of care combination therapy for chronic hepatitis C virus (HCV) infection, yet the component interferon-stimulated genes (ISGs) that mediate the antiviral actions of IFN are not fully defined. Interferon-induced transmembrane protein 1 (IFITM1) is an ISG product that suppresses early stage infection by a number of viruses through an as yet unknown mechanism of action. Moreover, the actions of IFITM1 on HCV infection are not fully elucidated. Here we identify IFITM1 as a hepatocyte tight junction protein and a potent anti-HCV effector molecule. IFITM1 expression is induced early during IFN treatment of hepatocytes and accumulates at hepatic tight junctions in HCV-infected human patient liver during IFN therapy. Additionally, we found that IFITM1 interacts with HCV co-receptors including CD81 and occludin to disrupt the process of viral entry. Thus, IFITM1 is an anti-HCV ISG whose actions impart control of HCV infection through interruption of viral coreceptor function. Conclusion: This study defines IFITM1 as an ISG effector with action against HCV entry. Design of therapy regimens to enhance IFITM1 expression should improve the virologic response among HCV patients undergoing treatment with type I IFN. (HEPATOLOGY 2012.).
Hepatology 09/2012; · 11.66 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: While 170 million people worldwide are chronically infected with HCV, the response rate to the current treatment regimens of pegylated IFN-α (IFN) in combination with ribavirin is only approximately 55 % of all HCV patients undergoing therapy. This IFN-based therapy is now slated to serve as the backbone for future combination therapeutics involving direct-acting antiviral compounds, including HCV protease inhibitors, viral polymerase inhibitors, and other small molecules. It is essential that the application of IFN be improved for overall enhancement of therapy outcome to effectively cure HCV infection. Systems approaches, including genomics and network modeling, are particularly powerful tools that are now being used to dissect the underlying mechanisms of successful or failed treatment response in an effort to design improved IFN-based therapeutic regimens. Furthermore, systems applications can be used to define virus-host interactions and map their variation within viral and host genomes, leading to identification of targets for novel therapy strategies. Using these approaches, we have defined distinct hepatic expression and tissue distribution of innate immune signaling molecules and gene networks that associate with IFN-based treatment outcome for HCV infection. This chapter will focus on using systems approaches to understand the host response to both HCV infection and therapy to drive the development of improved HCV therapeutics.
Current topics in microbiology and immunology 08/2012; · 4.93 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Viral infection of mammalian cells triggers the innate immune response through non-self recognition of pathogen associated molecular patterns (PAMPs) in viral nucleic acid. Accurate PAMP discrimination is essential to avoid self recognition that can generate autoimmunity, and therefore should be facilitated by the presence of multiple motifs in a PAMP that mark it as non-self. Hepatitis C virus (HCV) RNA is recognized as non-self by RIG-I through the presence of a 5'-triphosphate (5'-ppp) on the viral RNA in association with a 3' poly-U/UC tract. Here we define the HCV PAMP and the criteria for RIG-I non-self discrimination of HCV by examining the RNA structure-function attributes that impart PAMP function to the poly-U/UC tract. We found that the 34 nucleotide poly-uridine "core" of this sequence tract was essential for RIG-I activation, and that interspersed ribocytosine nucleotides between poly-U sequences in the RNA were required to achieve optimal RIG-I signal induction. 5'-ppp poly-U/UC RNA variants that stimulated strong RIG-I activation efficiently bound purified RIG-I protein in vitro, and RNA interaction with both the repressor domain and helicase domain of RIG-I was required to activate signaling. When appended to 5'-ppp RNA that lacks PAMP activity, the poly-U/UC U-core sequence conferred non-self recognition of the RNA and innate immune signaling by RIG-I. Importantly, HCV poly-U/UC RNA variants that strongly activated RIG-I signaling triggered potent anti-HCV responses in vitro and hepatic innate immune responses in vivo using a mouse model of PAMP signaling. These studies define a multi-motif PAMP signature of non-self recognition by RIG-I that incorporates a 5'-ppp with poly-uridine sequence composition and length. This HCV PAMP motif drives potent RIG-I signaling to induce the innate immune response to infection. Our studies define a basis of non-self discrimination by RIG-I and offer insights into the antiviral therapeutic potential of targeted RIG-I signaling activation.
PLoS Pathogens 08/2012; 8(8):e1002839. · 9.13 Impact Factor
-
Mehul S Suthar,
Hilario J Ramos,
Margaret M Brassil,
Jason Netland,
Craig P Chappell,
Gabriele Blahnik,
Aimee McMillan,
Michael S Diamond,
Edward A Clark,
Michael J Bevan, Michael Gale
[show abstract]
[hide abstract]
ABSTRACT: The RIG-I-like receptors (RLRs) signal innate immune defenses upon RNA virus infection, but their roles in adaptive immunity have not been clearly defined. Here, we showed that the RLR LGP2 was not essential for induction of innate immune defenses, but rather was required for controlling antigen-specific CD8(+) T cell survival and fitness during peripheral T cell-number expansion in response to virus infection. Adoptive transfer and biochemical studies demonstrated that T cell-receptor signaling induced LGP2 expression wherein LGP2 operated to regulate death-receptor signaling and imparted sensitivity to CD95-mediated cell death. Thus, LGP2 promotes an essential prosurvival signal in response to antigen stimulation to confer CD8(+) T cell-number expansion and effector functions against divergent RNA viruses, including West Nile virus and lymphocytic choriomeningitis virus.
Immunity 07/2012; 37(2):235-48. · 21.64 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Interferon regulatory factor-3 (IRF-3) is a master transcription factor that drives the host intracellular innate immune response to virus infection. The importance of IRF-3 in innate immune responses is highlighted by the fact that pathogenic viruses have developed strategies for antagonism of IRF-3. Several tools exist for evaluation of viral regulation of IRF-3 activation and function, but high-quality monoclonal antibodies that mark the differential activation states of human IRF-3 are lacking. To study IRF-3 activation, turnover, and depletion in a high-throughput manner in the context of virus infection, we have developed two new monoclonal antibodies to human IRF-3. These antibodies detect IRF-3 in virus-infected cells in a wide variety of assays and provide a new tool to study virus-host interactions and innate immune signaling.
Methods 06/2012; · 4.01 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Acute virus infection induces a cell-intrinsic innate immune response comprising our first line of immunity to limit virus replication and spread, but viruses have developed strategies to overcome these defenses. HIV-1 is a major public health problem; however, the virus-host interactions that regulate innate immune defenses against HIV-1 are not fully defined. We have recently identified the viral protein Vpu to be a key determinant responsible for HIV-1 targeting and degradation of interferon regulatory factor 3 (IRF3), a central transcription factor driving host cell innate immunity. IRF3 plays a major role in pathogen recognition receptor (PRR) signaling of innate immunity to drive the expression of type I interferon (IFN) and interferon-stimulated genes (ISGs), including a variety of HIV restriction factors, that serve to limit viral replication directly and/or program adaptive immunity. Here we interrogate the cellular responses to target cell infection with Vpu-deficient HIV-1 strains. Remarkably, in the absence of Vpu, HIV-1 triggers a potent intracellular innate immune response that suppresses infection. Thus, HIV-1 can be recognized by PRRs within the host cell to trigger an innate immune response, and this response is unmasked only in the absence of Vpu. Vpu modulation of IRF3 therefore prevents virus induction of specific innate defense programs that could otherwise limit infection. These observations show that HIV-1 can indeed be recognized as a pathogen in infected cells and provide a novel and effective platform for defining the native innate immune programs of target cells of HIV-1 infection.
Journal of Virology 05/2012; 86(16):8499-506. · 5.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: RIG-I is a cytosolic pathogen recognition receptor that initiates immune responses against RNA viruses. Upon viral RNA recognition, antiviral signaling requires RIG-I redistribution from the cytosol to membranes where it binds the adaptor protein, MAVS. Here we identify the mitochondrial targeting chaperone protein, 14-3-3ε, as a RIG-I-binding partner and essential component of a translocation complex or "translocon" containing RIG-I, 14-3-3ε, and the TRIM25 ubiquitin ligase. The RIG-I translocon directs RIG-I redistribution from the cytosol to membranes where it mediates MAVS-dependent innate immune signaling during acute RNA virus infection. 14-3-3ε is essential for the stable interaction of RIG-I with TRIM25, which facilitates RIG-I ubiquitination and initiation of innate immunity against hepatitis C virus and other pathogenic RNA viruses. Our results define 14-3-3ε as a key component of a RIG-I translocon required for innate antiviral immunity.
Cell host & microbe 05/2012; 11(5):528-37. · 13.02 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: HIV has evolved sophisticated mechanisms to avoid restriction by intracellular innate immune defenses that otherwise serve to control acute viral infection and virus dissemination. Innate defenses are triggered when pattern recognition receptor (PRR) proteins of the host cell engage pathogen-associated molecule patterns (PAMPs) present in viral products. Interferon regulatory factor 3 (IRF3) plays a central role in PRR signaling of innate immunity to drive the expression of type I interferon (IFN) and interferon-stimulated genes (ISGs), including a variety of HIV restriction factors, that serve to limit viral replication directly and/or program adaptive immunity. Productive infection of T cells by HIV is dependent upon the targeted proteolysis of IRF3 that occurs through a virus-directed mechanism that results in suppression of innate immune defenses. However, the mechanisms by which HIV controls innate immune signaling and IRF3 function are not defined. Here, we examined the innate immune response induced by HIV strains identified through their differential control of PRR signaling. We identified viruses that, unlike typical circulating HIV strains, lack the ability to degrade IRF3. Our studies show that IRF3 regulation maps specifically to the HIV accessory protein Vpu. We define a molecular interaction between Vpu and IRF3 that redirects IRF3 to the endolysosome for proteolytic degradation, thus allowing HIV to avoid the innate antiviral immune response. Our studies reveal that Vpu is an important IRF3 regulator that supports acute HIV infection through innate immune suppression. These observations define the Vpu-IRF3 interface as a novel target for therapeutic strategies aimed at enhancing the immune response to HIV.
Journal of Virology 05/2012; 86(16):8367-74. · 5.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Prior studies have shown that 2'-O methyltransferase activity of flaviviruses, coronaviruses, and poxviruses promotes viral evasion of Ifit1, an interferon-stimulated innate immune effector protein. Viruses lacking 2'-O methyltransferase activity exhibited attenuation in primary macrophages that was rescued in cells lacking Ifit1 gene expression. Here, we examined the role of Ifit1 in restricting pathogenesis in vivo of wild type WNV (WNV-WT) and a mutant in the NS5 gene (WNV-E218A) lacking 2'-O methylation of the 5' viral RNA cap. While deletion of Ifit1 had marginal effects on WNV-WT pathogenesis, WNV-E218A showed increased replication in peripheral tissues of Ifit1⁻/⁻ mice after subcutaneous infection, yet this failed to correlate with enhanced infection in the brain or lethality. In comparison, WNV-E218A was virulent after intracranial infection as judged by increased infection in different regions of the central nervous system (CNS) and a greater than 16,000-fold decrease in LD(50) values in Ifit1⁻/⁻ compared to wild type mice. Ex vivo infection experiments revealed cell-type specific differences in the ability of an Ifit1 deficiency to complement the replication defect of WNV-E218A. In particular, WNV-E218A infection was impaired in both wild type and Ifit1⁻/⁻ brain microvascular endothelial cells, which are believed to participate in blood-brain barrier (BBB) regulation of virus entry into the CNS. A deficiency of Ifit1 also was associated with increased neuronal death in vivo, which was both cell-intrinsic and mediated by immunopathogenic CD8⁺ T cells. Our results suggest that virulent strains of WNV have largely evaded the antiviral effects of Ifit1, and viral mutants lacking 2'-O methylation are controlled in vivo by Ifit1-dependent and -independent mechanisms in different cell types.
PLoS Pathogens 05/2012; 8(5):e1002698. · 9.13 Impact Factor
