Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Dynamic expression profiling of type I and type III interferon-stimulated hepatocytes.

ABSTRACT: This dataset details the time-dependent response of human Huh7 hepatoma cells to type I and type III IFN. Despite activating similar signaling cascades, the type I and type III interferons (IFNs) differ in their ability to antagonize viral replication. However, it is not clear whether these cytokines induce unique antiviral states, particularly in the liver, where the clinically important hepatitis B and C viruses cause persistent infection. Here, microarray-based gene expression analysis is combined with mechanistic studies of signaling pathways to dynamically characterize the transcriptional responses induced by these cytokines in Huh7 hepatoma cells and primary human hepatocytes. Type I and III IFNs differed greatly in their level of interferon-stimulated gene (ISG) induction with a clearly detectable hierarchy (IFN-M-NM-2 > IFN-M-NM-1 > IFN-M-NM-;3 > IFN-M-NM-;1 > IFN-M-NM-;2). This hierarchy resulted in widely varying numbers of differentially expressed genes when quantified using common statistical thresholds, even though individual IFNs did not appear to regulate unique sets of genes. The kinetic profiles of IFN-induced gene expression were also qualitatively similar with the important exception of IFN-M-NM-1. While stimulation with either IFN-M-NM-2 or IFN-M-NM-;s resulted in a similar long-lasting ISG induction, IFN-M-NM-1 signaling peaked early after stimulation then declined due to a negative feedback mechanism. The quantitative expression hierarchy and unique kinetics of IFN-M-NM-1 suggest different roles for individual IFNs in the immune response, and help explain previously observed differences in antiviral activity. Huh7 cells were seeded into 6-well plates at the density of 3x10^5 cells/well and cultured overnight before stimulation with either 500 U/ml of IFN-M-NM-1 or IFN-M-NM-2, or 10 ng/ml of IFN-M-NM-;1, IFN-M-NM-;2, or IFN-M-NM-;3. Total RNA was harvested and isolated at 0.5, 1, 2, 4, 6, 12 and 24 hours post-incubation using RNeasy Mini Kit (Qiagen) following the Affymetrix GeneChip protocol of the Keck Affymetrix Resource facility at Yale University. IFN incubation at each time point was performed in duplicate. All subsequent processing, hybridization to the Illumina HumanHT-12 microarray, and quality control analyses were carried out by the Yale Center for Genome Analysis using standard protocols.

ORGANISM(S): Homo sapiens

SUBMITTER: Christopher Bolen

PROVIDER: E-GEOD-48400 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Dynamic expression profiling of type I and type III interferon-stimulated hepatocytes reveals a stable hierarchy of gene expression.

Bolen Christopher R CR Ding Siyuan S Robek Michael D MD Kleinstein Steven H SH

Hepatology (Baltimore, Md.) 20140218 4

Despite activating similar signaling cascades, the type I and type III interferons (IFNs) differ in their ability to antagonize virus replication. However, it is not clear whether these cytokines induce unique antiviral states, particularly in the liver, where the clinically important hepatitis B and C viruses cause persistent infection. Here, clustering and promoter analyses of microarray-based gene expression profiling were combined with mechanistic studies of signaling pathways to dynamically ...[more]

PMID: 23929627

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Project description:Interferons (IFNs) are key players in the antiviral response. IFN sensing by the cell activates transcription of IFN-stimulated genes (ISGs) able to induce an antiviral state by affecting viral replication and release. IFN also induces the expression of ISGs that function as negative regulators to limit the strength and duration of IFN response. The ISGs identified so far belong to coding genes. However, only a small proportion of the transcriptome corresponds to coding transcripts and it has been estimated that there could be as many coding as long non-coding RNAs (lncRNAs). To address whether IFN can also regulate the expression of lncRNAs, we analyzed the transcriptome of HuH7 cells treated or not with IFNα2 by expression arrays. Analysis of the arrays showed increased levels of several well-characterized coding genes that respond to IFN both at early or late times. Furthermore, we identified several IFN-stimulated or -downregulated lncRNAs (ISRs and IDRs). Further validation showed that ISR2, 8, and 12 expression mimics that of their neighboring genes GBP1, IRF1, and IL6, respectively, all related to the IFN response. These genes are induced in response to different doses of IFNα2 in different cell lines at early (ISR2 or 8) or later (ISR12) time points. IFNβ also induced the expression of these lncRNAs. ISR2 and 8 were also induced by an influenza virus unable to block the IFN response but not by other wild-type lytic viruses tested. Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV. Increased levels of ISR2 were also detected in patients chronically infected with HIV. This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response. Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response. HuH7 cells were treated with 10000 units/ml of IFN a2 and RNA was isolated 3 days post-treatment

Project description:Interferons (IFNs) are key players in the antiviral response. IFN sensing by the cell activates transcription of IFN-stimulated genes (ISGs) able to induce an antiviral state by affecting viral replication and release. IFN also induces the expression of ISGs that function as negative regulators to limit the strength and duration of IFN response. The ISGs identified so far belong to coding genes. However, only a small proportion of the transcriptome corresponds to coding transcripts and it has been estimated that there could be as many coding as long non-coding RNAs (lncRNAs). To address whether IFN can also regulate the expression of lncRNAs, we analyzed the transcriptome of HuH7 cells treated or not with IFNÎ±2 by expression arrays. Analysis of the arrays showed increased levels of several well-characterized coding genes that respond to IFN both at early or late times. Furthermore, we identified several IFN-stimulated or -downregulated lncRNAs (ISRs and IDRs). Further validation showed that ISR2, 8, and 12 expression mimics that of their neighboring genes GBP1, IRF1, and IL6, respectively, all related to the IFN response. These genes are induced in response to different doses of IFNÎ±2 in different cell lines at early (ISR2 or 8) or later (ISR12) time points. IFNÎ² also induced the expression of these lncRNAs. ISR2 and 8 were also induced by an influenza virus unable to block the IFN response but not by other wild-type lytic viruses tested. Surprisingly, both ISR2 and 8 were significantly upregulated in cultured cells and livers from patients infected with HCV. Increased levels of ISR2 were also detected in patients chronically infected with HIV. This is relevant as genome-wide guilt-by-association studies predict that ISR2, 8, and 12 may function in viral processes, in the IFN pathway and the antiviral response. Therefore, we propose that these lncRNAs could be induced by IFN to function as positive or negative regulators of the antiviral response. HuH7 cells were treated with 10000 units/ml of IFN a2 and RNA was isolated 3 days post-treatment

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Dataset Information

Dynamic expression profiling of type I and type III interferon-stimulated hepatocytes.

Publications

Dynamic expression profiling of type I and type III interferon-stimulated hepatocytes reveals a stable hierarchy of gene expression.

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