Chikungunya virus (CHIKV) is a mosquito-borne arthralgia arbovirus that is reemergent in sub-Saharan Africa and Southeast Asia. CHIKV infection has been shown to be self-limiting, but the molecular mechanisms of the innate immune response that control CHIKV replication remain undefined. Here, longitudinal transcriptional analyses of PBMCs from a cohort of CHIKV-infected patients revealed that type I IFNs controlled CHIKV infection via RSAD2 (which encodes viperin), an enigmatic multifunctional IFN-stimulated gene (ISG). Viperin was highly induced in monocytes, the major target cell of CHIKV in blood. Anti-CHIKV functions of viperin were dependent on its localization in the ER, and the N-terminal amphipathic α-helical domain was crucial for its antiviral activity in controlling CHIKV replication. Furthermore, mice lacking Rsad2 had higher viremia and severe joint inflammation compared with wild-type mice. Our data demonstrate that viperin is a critical antiviral host protein that controls CHIKV infection and provide a preclinical basis for the design of effective control strategies against CHIKV and other reemerging arthrogenic alphaviruses.
Terk-Shin Teng, Suan-Sin Foo, Diane Simamarta, Fok-Moon Lum, Teck-Hui Teo, Aleksei Lulla, Nicholas K.W. Yeo, Esther G.L. Koh, Angela Chow, Yee-Sin Leo, Andres Merits, Keh-Chuang Chin, Lisa F.P. Ng
In order to prime T cells, DCs integrate signals emanating directly from pathogens and from their noxious action on the host. DNGR-1 (CLEC9A) is a DC-restricted receptor that detects dead cells. Therefore, we investigated the possibility that DNGR-1 affects immunity to cytopathic viruses. DNGR-1 was essential for cross-presentation of dying vaccinia virus–infected (VACV-infected) cells to CD8+ T cells in vitro. Following injection of VACV or VACV-infected cells into mice, DNGR-1 detected the ligand in dying infected cells and mediated cross-priming of anti-VACV CD8+ T cells. Loss of DNGR-1 impaired the CD8+ cytotoxic response to VACV, especially against those virus strains that are most dependent on cross-presentation. The decrease in total anti-VACV CTL activity was associated with a profound increase in viral load and delayed resolution of the primary lesion. In addition, lack of DNGR-1 markedly diminished protection from infection induced by vaccination with the modified vaccinia Ankara (MVA) strain. DNGR-1 thus contributes to anti-VACV immunity, following both primary infection and vaccination. The non-redundant ability of DNGR-1 to regulate cross-presentation of viral antigens suggests that this form of regulation of antiviral immunity could be exploited for vaccination.
Salvador Iborra, Helena M. Izquierdo, María Martínez-López, Noelia Blanco-Menéndez, Caetano Reis e Sousa, David Sancho
SIVs infecting wild-living apes in west central Africa have crossed the species barrier to humans on at least four different occasions, one of which spawned the AIDS pandemic. Although the chimpanzee precursor of pandemic HIV-1 strains must have been able to infect humans, the capacity of SIVcpz strains to replicate in human lymphoid tissues (HLTs) is not known. Here, we show that SIVcpz strains from two chimpanzee subspecies are capable of replicating in human tonsillary explant cultures, albeit only at low titers. However, SIVcpz replication in HLT was significantly improved after introduction of a previously identified human-specific adaptation at position 30 in the viral Gag matrix protein. An Arg or Lys at this position significantly increased SIVcpz replication in HLT, while the same mutation reduced viral replication in chimpanzee-derived CD4+ T cells. Thus, naturally occurring SIVcpz strains are capable of infecting HLTs, the major site of HIV-1 replication in vivo. However, efficient replication requires the acquisition of a host-specific adaptation in the viral matrix protein. These results identify Gag matrix as a major determinant of SIVcpz replication fitness in humans and suggest a critical role in the emergence of HIV/AIDS.
Frederic Bibollet-Ruche, Anke Heigele, Brandon F. Keele, Juliet L. Easlick, Julie M. Decker, Jun Takehisa, Gerald Learn, Paul M. Sharp, Beatrice H. Hahn, Frank Kirchhoff
Herpes simplex viruses (HSVs) are highly prevalent neurotropic viruses. While they can replicate lytically in cells of the epithelial lineage, causing lesions on mucocutaneous surfaces, HSVs also establish latent infections in neurons, which act as reservoirs of virus for subsequent reactivation events. Immunological control of HSV involves activation of innate immune pattern-recognition receptors such as TLR3, which detects double-stranded RNA and induces type I IFN expression. Humans with defects in the TLR3/IFN pathway have an elevated susceptibility to HSV infections of the CNS. However, it is not known what cell type mediates the role of TLR3 in the immunological control of HSV, and it is not known whether TLR3 sensing occurs prior to or after CNS entry. Here, we show that in mice TLR3 provides early control of HSV-2 infection immediately after entry into the CNS by mediating type I IFN responses in astrocytes. Tlr3–/– mice were hypersusceptible to HSV-2 infection in the CNS after vaginal inoculation. HSV-2 exhibited broader neurotropism in Tlr3–/– mice than it did in WT mice, with astrocytes being most abundantly infected. Tlr3–/– mice did not exhibit a global defect in innate immune responses to HSV, but astrocytes were defective in HSV-induced type I IFN production. Thus, TLR3 acts in astrocytes to sense HSV-2 infection immediately after entry into the CNS, possibly preventing HSV from spreading beyond the neurons mediating entry into the CNS.
Line S. Reinert, Louis Harder, Christian K. Holm, Marie B. Iversen, Kristy A. Horan, Frederik Dagnæs-Hansen, Benedicte P. Ulhøi, Thomas H. Holm, Trine H. Mogensen, Trevor Owens, Jens R. Nyengaard, Allan R. Thomsen, Søren R. Paludan
Infections by viruses are associated with approximately 12% of human cancer. Kaposi’s sarcoma-associated herpesvirus (KSHV) is causally linked to several malignancies commonly found in AIDS patients. The mechanism of KSHV-induced oncogenesis remains elusive, due in part to the lack of an adequate experimental system for cellular transformation of primary cells. Here, we report efficient infection and cellular transformation of primary rat embryonic metanephric mesenchymal precursor cells (MM cells) by KSHV. Cellular transformation occurred at as early as day 4 after infection and in nearly all infected cells. Transformed cells expressed hallmark vascular endothelial, lymphatic endothelial, and mesenchymal markers and efficiently induced tumors in nude mice. KSHV established latent infection in MM cells, and lytic induction resulted in low levels of detectable infectious virions despite robust expression of lytic genes. Most KSHV-induced tumor cells were in a latent state, although a few showed heterogeneous expression of lytic genes. This efficient system for KSHV cellular transformation of primary cells might facilitate the study of growth deregulation mechanisms resulting from KSHV infections.
Tiffany Jones, Fengchun Ye, Roble Bedolla, Yufei Huang, Jia Meng, Liwu Qian, Hongyi Pan, Fuchun Zhou, Rosalie Moody, Brent Wagner, Mazen Arar, Shou-Jiang Gao
HBV infection remains a leading cause of death worldwide. IFN-α inhibits viral replication in vitro and in vivo, and pegylated IFN-α is a commonly administered treatment for individuals infected with HBV. The HBV genome contains a typical IFN-stimulated response element (ISRE), but the molecular mechanisms by which IFN-α suppresses HBV replication have not been established in relevant experimental systems. Here, we show that IFN-α inhibits HBV replication by decreasing the transcription of pregenomic RNA (pgRNA) and subgenomic RNA from the HBV covalently closed circular DNA (cccDNA) minichromosome, both in cultured cells in which HBV is replicating and in mice whose livers have been repopulated with human hepatocytes and infected with HBV. Administration of IFN-α resulted in cccDNA-bound histone hypoacetylation as well as active recruitment to the cccDNA of transcriptional corepressors. IFN-α treatment also reduced binding of the STAT1 and STAT2 transcription factors to active cccDNA. The inhibitory activity of IFN-α was linked to the IRSE, as IRSE-mutant HBV transcribed less pgRNA and could not be repressed by IFN-α treatment. Our results identify a molecular mechanism whereby IFN-α mediates epigenetic repression of HBV cccDNA transcriptional activity, which may assist in the development of novel effective therapeutics.
Laura Belloni, Lena Allweiss, Francesca Guerrieri, Natalia Pediconi, Tassilo Volz, Teresa Pollicino, Joerg Petersen, Giovanni Raimondo, Maura Dandri, Massimo Levrero
Induction of virus-specific CD8+ T cell responses is critical for the success of vaccines against chronic viral infections. Despite the large number of potential MHC-I–restricted epitopes located in viral proteins, MHC-I–restricted epitope generation is inefficient, and factors defining the production and presentation of MHC-I–restricted viral epitopes are poorly understood. Here, we have demonstrated that the half-lives of HIV-derived peptides in cytosol from primary human cells were highly variable and sequence dependent, and significantly affected the efficiency of cell recognition by CD8+ T cells. Furthermore, multiple clinical isolates of HLA-associated HIV epitope variants displayed reduced half-lives relative to consensus sequence. This decreased cytosolic peptide stability diminished epitope presentation and CTL recognition, illustrating a mechanism of immune escape. Chaperone complexes including Hsp90 and histone deacetylase HDAC6 enhanced peptide stability by transient protection from peptidase degradation. Based on empirical results with 166 peptides, we developed a computational approach utilizing a sequence-based algorithm to estimate the cytosolic stability of antigenic peptides. Our results identify sequence motifs able to alter the amount of peptide available for loading onto MHC-I, suggesting potential new strategies to modulate epitope production from vaccine immunogens.
Estibaliz Lazaro, Carl Kadie, Pamela Stamegna, Shao Chong Zhang, Pauline Gourdain, Nicole Y. Lai, Mei Zhang, Sergio A. Martinez, David Heckerman, Sylvie Le Gall
Kaposi sarcoma–associated herpesvirus (KSHV) is a B-lymphotropic virus whose primary site of replication is the oropharynx. KSHV can infect both T and B cells from primary tonsillar explant cultures. However, T cells do not support lytic replication, while B cells spontaneously produce substantial amounts of infectious virus. Here, we provide evidence for a mechanism by which activated T cells may promote or stabilize latency of KSHV infection in B cells. When mixed cultures of B cells and activated T cells were exposed to KSHV, little spontaneous virus production was observed. Removing T cells from the mix or treating the mixed culture with immune suppressants enhanced virus production. Adding back activated T cells to purified infected B cells efficiently suppressed KSHV production, primarily due to CD4+ T cells. This suppressive activity required T cell activation and direct cell-cell contact, but not prior exposure to KSHV antigen. Suppression was not MHC restricted and did not result in killing of the target cell. We therefore propose that oropharyngeal T cells activated by a variety of stimuli can recognize ligands on infected target B cells, leading to signaling events that prevent spontaneous lytic activation and promote latent infection in this compartment.
Jinjong Myoung, Don Ganem
During infection with the hepatitis A virus (HAV), most patients develop mild or asymptomatic disease. However, a small number of patients develop serious, life-threatening hepatitis. We investigated this variability in disease severity by examining 30 Argentinean patients with HAV-induced acute liver failure in a case-control, cross-sectional, observational study. We found that HAV-induced severe liver disease was associated with a 6-amino-acid insertion in TIM1/HAVCR1 (157insMTTTVP), the gene encoding the HAV receptor. This polymorphism was previously shown to be associated with protection against asthma and allergic diseases and with HIV progression. In binding assays, the TIM-1 protein containing the 157insMTTTVP insertion polymorphism bound HAV more efficiently. When expressed by human natural killer T (NKT) cells, this long form resulted in greater NKT cell cytolytic activity against HAV-infected liver cells, compared with the shorter TIM-1 protein without the polymorphism. To our knowledge, the 157insMTTTVP polymorphism in TIM1 is the first genetic susceptibility factor shown to predispose to HAV-induced acute liver failure. Furthermore, these results suggest that HAV infection has driven the natural selection of shorter forms of the TIM-1 protein, which binds HAV less efficiently, thereby protecting against severe HAV-induced disease, but which may predispose toward inflammation associated with asthma and allergy.
Hye Young Kim, María Belén Eyheramonho, Muriel Pichavant, Carlos Gonzalez Cambaceres, Ponpan Matangkasombut, Guillermo Cervio, Silvina Kuperman, Rita Moreiro, Krishnamurthy Konduru, Mohanraj Manangeeswaran, Gordon J. Freeman, Gerardo G. Kaplan, Rosemarie H. DeKruyff, Dale T. Umetsu, Sergio D. Rosenzweig
Plasmacytoid DCs (pDCs) are innate immune cells that are specialized to produce IFN-α and to activate adaptive immune responses. Although IFN-α inhibits HIV-1 replication in vitro, the production of IFN-α by HIV-activated pDCs in vivo may contribute more to HIV pathogenesis than to protection. We have now shown that HIV-stimulated human pDCs allow for persistent IFN-α production upon repeated stimulation, express low levels of maturation molecules, and stimulate weak T cell responses. Persistent IFN-α production by HIV-stimulated pDCs correlated with increased levels of IRF7 and was dependent upon the autocrine IFN-α/β receptor feedback loop. Because it has been shown that early endosomal trafficking of TLR9 agonists causes strong activation of the IFN-α pathway but weak activation of the NF-κB pathway, we sought to investigate whether early endosomal trafficking of HIV, a TLR7 agonist, leads to the IFN-α–producing phenotype we observed. We demonstrated that HIV preferentially traffics to the early endosome in human pDCs and therefore skews pDCs toward a partially matured, persistently IFN-α–secreting phenotype.
Meagan O’Brien, Olivier Manches, Rachel Lubong Sabado, Sonia Jimenez Baranda, Yaming Wang, Isabelle Marie, Linda Rolnitzky, Martin Markowitz, David M. Margolis, David Levy, Nina Bhardwaj
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