Characterization of the chemokine response of RAW264.7 cells to infection by murine norovirus

Waugh, Emily; Chen, Augustine; Baird, Margaret A.; Brown, Chris M.; Ward, Vernon K.

doi:10.1016/j.virusres.2013.12.025

Cited by 16 publications

(14 citation statements)

References 52 publications

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“…The TNFα and Traf1 upregulation was also observed in this study, both in MNV infected and in NS1-2 transfected cells (Table 3). The chemokines secreted during an in vitro MNV infection correlate to a Th1 phenotype, and this has been previously shown by microarray analysis for mRNA and ELISA assays to detect secreted chemokines [21]. There is also an upregulation of serum TNFα and IFNβ [22], and an increase in inflammatory dendritic cells in the intestines of mice [68], during an MNV infection that further supports a Th1 phenotype.…”

Section: Resultsmentioning

confidence: 64%

“…MNV-1 (CW1-P3) was initially generated via reverse genetics [27], and purified as previously described [21]. MNV NS1-2 (nt 1–1028) and NS1-2 dis (nt 1–431) were cloned from full-length MNV-1 cDNA using forward primers containing a T7 promoter (underlined), the MNV 5′ UTR (lowercase), and nt 6–25 at the N-terminus of the MNV genome (5′-GAAATTAATACGACTCACTATAgtgaaATGAGGATGGCAACGCCATC-3′), and reverse primers containing stop codon (bold) and unique restriction enzyme sites (italics) (NS1-2 (nt 1024–1046): 5′-AGCAAGGTCGAAGGGTTATTCGGC-3′) and (NS1-2 dis (nt 409–431): 5′-GGTGGT CTGCAG TTACTCCAAGATAGAGCCGATCACAG-3′).…”

Section: Methodsmentioning

confidence: 99%

“…These stem cells are derived from patients undergoing intestinal biopsies for a variety of pathological conditions, which would complicate transcriptomic analysis. MNV grown in monocytes has been used a model system to study norovirus pathogenesis, including intracellular signaling and effects on immune-regulatory molecules [7, 21, 22]. Since murine monocytes are well characterised in terms of both differences and similarities to its human counterpart [23, 24], and MNV infections have been well studied [25, 26], an established murine monocyte cell line was used in this study.…”

Section: Introductionmentioning

confidence: 99%

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Transcriptomic analysis of human norovirus NS1-2 protein highlights a multifunctional role in murine monocytes

et al. 2017

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BackgroundThe GII.4 Sydney 2012 strain of human norovirus (HuNoV) is a pandemic strain that is responsible for the majority of norovirus outbreaks in healthcare settings. The function of the non-structural (NS)1-2 protein from HuNoV is unknown.Results In silico analysis of human norovirus NS1-2 protein showed that it shares features with the murine NS1-2 protein, including a disordered region, a transmembrane domain and H-box and NC sequence motifs. The proteins also contain caspase cleavage and phosphorylation sites, indicating that processing and phosphorylation may be a conserved feature of norovirus NS1-2 proteins. In this study, RNA transcripts of human and murine norovirus full-length and the disordered region of NS1-2 were transfected into monocytes, and next generation sequencing was used to analyse the transcriptomic profile of cells expressing virus proteins. The profiles were then compared to the transcriptomic profile of MNV-infected cells.ConclusionsRNAseq analysis showed that NS1-2 proteins from human and murine noroviruses affect multiple immune systems (chemokine, cytokine, and Toll-like receptor signaling) and intracellular pathways (NFκB, MAPK, PI3K-Akt signaling) in murine monocytes. Comparison to the transcriptomic profile of MNV-infected cells indicated the pathways that NS1-2 may affect during norovirus infection.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3417-4) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 64%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Transcriptomic analysis of human norovirus NS1-2 protein highlights a multifunctional role in murine monocytes

et al. 2017

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“…The recent generation of an enteroid system for the growth and culture of human norovirus will enable this to be investigated in more detail (27). Notwithstanding this, we and others have observed that ISG gene induction observed following murine norovirus infection often does not correlate with the resulting levels of the induced protein (11, 28), suggesting a posttranscriptional regulatory mechanism is also involved in the control of the innate response.…”

mentioning

confidence: 55%

Norovirus-Mediated Modification of the Translational Landscape via Virus and Host-Induced Cleavage of Translation Initiation Factors

Emmott

Sorgeloos

Caddy

et al. 2017

Molecular & Cellular Proteomics

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Noroviruses produce viral RNAs lacking a 5′ cap structure and instead use a virus-encoded viral protein genome-linked (VPg) protein covalently linked to viral RNA to interact with translation initiation factors and drive viral protein synthesis. Norovirus infection results in the induction of the innate response leading to interferon stimulated gene (ISG) transcription. However, the translation of the induced ISG mRNAs is suppressed. A SILAC-based mass spectrometry approach was employed to analyze changes to protein abundance in both whole cell and m7GTP-enriched samples to demonstrate that diminished host mRNA translation correlates with changes to the composition of the eukaryotic initiation factor complex. The suppression of host ISG translation correlates with the activity of the viral protease (NS6) and the activation of cellular caspases leading to the establishment of an apoptotic environment. These results indicate that noroviruses exploit the differences between viral VPg-dependent and cellular cap-dependent translation in order to diminish the host response to infection.

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“…To address the specificity of VSV-induced transcriptome changes with other antiviral response, we compared our 6 h transcriptome data with the transcriptome of RAW264.7 cells infected with Murine norovirus (MNV1) for 9 h. 28 There was a good correlation (Pearson correlation=0.53) between the differentially expressed VSV-induced transcriptome at 6 h with MNV1-induced transcriptome at 9 h. The transcript levels of type-I IFNs, IFN-stimulated genes (ISGs), and proinflammatory cytokines were upregulated in both VSV and MNV1 transcriptome (Supplementary Figure S1H). To elucidate the specificity of VSV-induced phosphoproteome change, we compared our data set with LPS-induced phosphoproteome changes in RAW264.7 cells.…”

Section: Resultsmentioning

confidence: 99%

A time-resolved molecular map of the macrophage response to VSV infection

et al. 2016

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Studying the relationship between virus infection and cellular response is paradigmatic for our understanding of how perturbation changes biological systems. Immune response, in this context is a complex yet evolutionarily adapted and robust cellular change, and is experimentally amenable to molecular analysis. To visualize the full cellular response to virus infection, we performed temporal transcriptomics, proteomics, and phosphoproteomics analysis of vesicular stomatitis virus (VSV)-infected mouse macrophages. This enabled the understanding of how infection-induced changes in host gene and protein expression are coordinated with post-translational modifications by cells in time to best measure and control the infection process. The vast and complex molecular changes measured could be decomposed in a limited number of clusters within each category (transcripts, proteins, and protein phosphorylation) each with own kinetic parameter and characteristic pathways/processes, suggesting multiple regulatory options in the overall sensing and homeostatic program. Altogether, the data underscored a prevalent executive function to phosphorylation. Resolution of the molecular events affecting the RIG-I pathway, central to viral recognition, reveals that phosphorylation of the key innate immunity adaptor mitochondrial antiviral-signaling protein (MAVS) on S328/S330 is necessary for activation of type-I interferon and nuclear factor κ B (NFκB) pathways. To further understand the hierarchical relationships, we analyzed kinase–substrate relationships and found RAF1 and, to a lesser extent, ARAF to be inhibiting VSV replication and necessary for NFκB activation, and AKT2, but not AKT1, to be supporting VSV replication. Integrated analysis using the omics data revealed co-regulation of transmembrane transporters including SLC7A11, which was subsequently validated as a host factor in the VSV replication. The data sets are predicted to greatly empower future studies on the functional organization of the response of macrophages to viral challenges.

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Characterization of the chemokine response of RAW264.7 cells to infection by murine norovirus

Cited by 16 publications

References 52 publications

Transcriptomic analysis of human norovirus NS1-2 protein highlights a multifunctional role in murine monocytes

Transcriptomic analysis of human norovirus NS1-2 protein highlights a multifunctional role in murine monocytes

Norovirus-Mediated Modification of the Translational Landscape via Virus and Host-Induced Cleavage of Translation Initiation Factors

A time-resolved molecular map of the macrophage response to VSV infection

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