Induction of alpha/beta interferon by myxoma virus is selectively abrogated when primary mouse embryo fibroblasts become immortalized.
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ABSTRACT:
SUBMITTER: Wang F
PROVIDER: S-EPMC2681981 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The purpose of this experiment was to obtain samples for microRNA analysis in IHH cells infected with human interferon alpha. Immortalized human hepatocyte (IHH) cells were seeded at 1 x 10^6 cells per well on 6 well plates 3 days before infection. Treatments were initiated by replacing culture medium (containing 10% FBS) with fresh medium containing 500 U/ml of human interferon α (PBL Biosciences). Mock-treated cells were given normal culture medium only. Mock and treated cells were incubated at 37°C until sample collection time points. Infected samples were collected in triplet; time-matched mocks were collected in triplet in parallel with infected samples. Time points: 6, 12, and 18 h post-infection
2015-08-06 | E-GEOD-71733 | biostudies-arrayexpress
Project description:The purpose of this experiment was to obtain samples for mRNA analysis in IHH cells infected with human interferon alpha. Immortalized human hepatocyte (IHH) cells were seeded at 1 x 10^6 cells per well on 6 well plates 3 days before infection. Treatments were initiated by replacing culture medium (containing 10% FBS) with fresh medium containing 500 U/ml of human interferon α (PBL Biosciences). Mock-treated cells were given normal culture medium only. Mock and treated cells were incubated at 37°C until sample collection time points. Infected samples were collected in triplet; time-matched mocks were collected in triplet in parallel with infected samples. Time points: 6, 12, and 18 h post-infection
2015-08-06 | E-GEOD-71732 | biostudies-arrayexpress
Project description:Bats are considered to be natural reservoirs for several viruses of clinical importance, including rabies virus, Nipah virus, and Hendra virus. Type I interferons (IFNs) is an important part of the immune system in the defense against viral infection. To investigate the function of type I IFNs upon viral infection in bats, the nucleic acid, and amino acid sequences of Egyptian Rousette (Rousettus aegyptiacus) IFN-alpha and -beta were characterized. Sequence data indicated that bat IFN-alpha consists of 562-bp encoded 187-aa, and IFN-beta consisted of 558-bp encoded 186-aa. Phylogenetic analysis of the overall identity of IFN-beta shared the highest sequence homology with pig IFN-beta in both nucleotide and amino acid level. Stimulation of bat primary kidney cells (BPKCs) and bat lung cell lines, Tb-1 Lu, with polyinosinic-polycytidylic acid (poly(I:C)) or exogenous bat type I IFNs resulted in increased type I IFNs mRNA expression in BPKCs, but not in Tb-1 Lu. Characterization of the bat IFN-alpha and -beta genes allows understanding of the immune responses upon stimulation in different tissues, thus providing practical strategies for control and treatment of clinically important diseases. These results are important especially for the virus infection, and suggest that future molecular studies on virus infection experiment of bats in vitro will require careful consideration of the differences of type I IFN expression patterns in different cell types.
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Project description:We previously identified a subset of interferon-stimulated genes (ISGs) upregulated by West Nile virus (WNV) infection in wild-type mouse embryo fibroblasts (MEFs) after viral proteins had inhibited type I interferon (IFN)-mediated JAK-STAT signaling and also in WNV-infected RIG-I-/-, MDA5-/-, STAT1-/-, STAT2-/-, IFNAR-/-, IRF3-/-, IRF7-/-, and IRF3/7-/- MEFs. In this study, ISG upregulation by WNV infection in IFNAR-/- MEFs was confirmed by transcriptome sequencing (RNA-seq). ISG upregulation by WNV infection was inhibited in RIG-I/MDA5-/- MEFs. ISGs were upregulated in IRF1-/- and IRF5-/- MEFs but only minimally upregulated in IRF3/5/7-/- MEFs, suggesting redundant IRF involvement. We previously showed that a single proximal interferon-stimulated response element (ISRE) in the Oas1a and Oas1b promoters bound the ISGF3 complex after type I IFN treatment. In this study, we used wild-type and mutant promoter luciferase reporter constructs to identify critical regions in the Oas1b and Ifit1 promoters for gene activation in infected IFNAR-/- MEFs. Two ISREs were required in both promoters. Mutation of these ISREs in an Ifit1 promoter DNA probe reduced in vitro complex formation with infected nuclear extracts. An NF-κB inhibitor decreased Ifit1 promoter activity in cells and in vitro complex formation. IRF3 and p50 promoter binding was detected by chromatin immunoprecipitation (ChIP) for upregulated ISGs with two proximal ISREs. The data indicate that ISREs function cooperatively to upregulate the expression of some ISGs when type I IFN signaling is absent, with the binding complex consisting of IRF3, IRF5, and/or IRF7 and an NF-κB component(s) as well as other, as-yet-unknown factors. IMPORTANCE Type I IFN signaling in mammalian cells induces formation of the ISGF3 transcription factor complex, which binds to interferon stimulated response elements (ISREs) in the promoters of interferon-stimulated genes (ISGs) in the cell nucleus. Flavivirus proteins counteract type I IFN signaling by preventing either the formation or nuclear localization of ISGF3. A subset of ISRE-regulated ISGs was still induced in West Nile virus (WNV)-infected mouse embryo fibroblasts (MEFs), indicating that cells have an alternative mechanism for activating these ISGs. In this study, cellular components involved in this ISG upregulation mechanism were identified using gene knockout MEFs and ChIP, and critical promoter regions for gene activation were mapped using reporter assays. The data indicate a cooperative function between two ISREs and required binding of IRF3, IRF5, and/or IRF7 and an NF-κB component(s). Moreover, type I IFN signaling-independent ISG activation requires different additional promoter activation regions than type I IFN-dependent activation.
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Project description:The purpose of this experiment was to obtain samples for microRNA analysis in IHH cells infected with human interferon alpha.
2015-08-06 | GSE71733 | GEO
Project description:The purpose of this experiment was to obtain samples for mRNA analysis in IHH cells infected with human interferon alpha.
2015-08-06 | GSE71732 | GEO