Mus musculus
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PROVIDER: PRJNA350836 | ENA |
REPOSITORIES: ENA
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Project description:Glycosylation of the HA protein of H5N1 virus increases its virulence in mice by exacerbating the host immune response
| PRJNA350836 | ENA
Project description:The highly pathogenic avian influenza (HPAI) H5N1 viruses continue to circulate in nature and threaten public health. Although several viral determinants and host factors that influence the virulence of HPAI H5N1 viruses in mammals have been identified, the detailed molecular mechanism remains poorly defined and requires further clarification. In our previous studies, we characterized two naturally isolated HPAI H5N1 viruses that had similar viral genomes but differed substantially in their lethality in mice. Here, we explored the molecular determinants and potential mechanism for this difference in virulence. By using reverse genetics, we found that a single amino acid at position 158 of the hemagglutinin (HA) protein substantially affected the systemic replication and pathogenicity of these H5N1 influenza viruses in mice. We further found that the G158N mutation introduced an N-linked glycosylation at sites 158–160 of the HA protein and that this N-linked glycosylation enhanced viral productivity in infected mammalian cells and induced stronger host immune and inflammatory responses to viral infection. These findings further our understanding of the determinants of pathogenicity of H5N1 viruses in mammals.
2016-12-22 | GSE89246 | GEO
Project description:Determination of the host response (C57Bl6 mouse model) to the HA avirulent mutation in A/Vietnam/1203-CIP048_RG1/2004(H5N1) virus at 10^4PFU.
2012-04-25 | GSE37572 | GEO
Project description:Determination of the host response (C57Bl6 mouse model) to the HA avirulent mutation in A/Vietnam/1203-CIP048_RG1/2004(H5N1) virus at 10^4PFU. Groups of 20 week old C57BL6 mice were infected with the HA avirulent mutation in A/Vietnam/1203-CIP048_RG1/2004(H5N1) virus at 10^4PFU or time-matched mock infected. Time points were 1, 2, 4 and 7 d.p.i. There were 3-4 animals/dose/time point. Lung samples were collected for virus load, transcriptional and proteomics analysis. Weight loss and animal survival were also monitored.
2012-04-24 | E-GEOD-37572 | biostudies-arrayexpress
Project description:Broilers were immunized with three variants of subunit vaccines, based on the hemagglutinin (HA) DNA and Pichia-produced HA protein from H5N1 virus, in comparison to the control group, which was administered an empty vector (pCI). Gene expression changes in the spleens of chickens were investigated at 7 day post booster dose.
2018-07-12 | GSE102972 | GEO
Project description:Influenza A virus (IAV) is a human respiratory pathogen that causes yearly global epidemics, and sporadic pandemics due to human adaptation of pathogenic strains. Efficient replication of IAV in different species is, in part, dictated by its ability to exploit the genetic environment of the host cell. To investigate IAV tropism in human cells, we evaluated the replication of IAV strains in a diverse subset of epithelial cell lines. HeLa cells were refractory to growth of human H1N1 and H3N2, and low pathogenic avian influenza (LPAIs) viruses. Interestingly, a human isolate of the highly pathogenic avian influenza (HPAI) virus H5N1 successfully propagated in HeLa cells to levels comparable to a human lung cell line. Heterokaryon cells generated by fusion of HeLa and permissive cells supported H1N1 growth, suggesting the absence of a host factor(s) required for replication of H1N1, but not H5N1, in HeLa cells. The absence of this factor(s) was mapped to reduced nuclear import, replication, and translation, and deficient viral budding. Using reassortant H1N1:H5N1 viruses, we found that the combined introduction of nucleoprotein (NP) and hemagglutinin (HA) from H5N1 was necessary and sufficient to enable H1N1 growth. Overall, this study suggests the absence of one or more cellular factors in HeLa cells that results in abortive replication of H1N1, H3N2, and LPAI viruses, but can be circumvented upon introduction of H5N1 NP and HA. Further understanding of the molecular basis of this restriction will provide important insights into virus-host interactions that underlie IAV pathogenesis and tropism.
2019-11-21 | GSE140759 | GEO
Project description:The underlying molecular mechanisms of pathogenesis and outcome of disease to different pathotypes of H5N1 influenza infection in ducks remain unclear. For that, we studied genome wide host gene expression of lung tissues infected with A/duck/India/02CA10/2011(AD2011) H5N1 virus and A/duck/Tripura/103597/2008 (AD2008) H5N1 virus in ducks using custom designed microarray. AD2011 is highly pathogenic whereas AD2008 is low pathogenic to ducks. Comparative analysis of differentially expressed genes revealed that 688 genes were commonly expressed, 877 and 1556 genes are uniquely expressed to infection with AD2011 and AD2008 virus isolate, respectively. The up-regulation of cytokines genes OAS, IL1B, IL17, IFITM2, CCL4, CXCR4, STAT3, TGFB1 and TGFB2 in the lungs tissues may cause high mortality in ducks infected with AD2011 virus. The expression of important antiviral immune genes IFIT5, IFITM5, RSAD2, EIF2AK2 (PKR), Mx, β-defensins, TRIM23 and SLC16A3 to AD2008 infection, but not in AD2011 infection, cause the host may fine-tune their innate immune responses and prevent from cytokines storms and tissue damage. Several immune related Gene ontology (GO) terms and immune pathways activated were qualitatively similar but quantitatively different to both virus infections. Based on these findings, we conclude that subtle differences in host immune responses may determine the different outcome of H5N1 infection in ducks. Agilent Custom Duck Gene Expression 8X60k (AMADID: G4102A_059612) designed by Genotypic Technology Private Limited , Labeling kit: Agilent Quick-Amp labeling Kit (p/n5190-0442)
2016-07-04 | E-GEOD-65230 | biostudies-arrayexpress
Project description:Influenza A virus (IAV) mutates rapidly, preventing a single seasonal vaccine from targeting more than one strain, and year-to-year vaccine effectiveness is low. One challenge in designing effective vaccines is that genetic mutations frequently cause amino acid variations in IAV envelope protein hemagglutinin (HA) that create new N-glycosylation sequons; resulting N-glycans cause antigenic shielding, allowing viral escape from adaptive immune responses. Vaccine candidate strain selection currently involves correlating antigenicity with HA protein sequence among circulating strains, but quantitative comparison of site-specific glycosylation information is likely to improve the ability to design vaccines with broader effectiveness against evolved strains. However, there is poor understanding of the influence of glycosylation on immunodominance, antigenicity, and immunogenicity of HA, and there are no well-tested methods for comparing glycosylation similarity among virus samples. Here, we present a method for statistically rigorous quantification of similarity between two related virus strains that considers the presence and abundance of glycopeptide glycoforms. We demonstrate the strength of our approach by determining that there was a quantifiable difference in glycosylation at the protein level between wild-type IAV HA from A/Switzerland/9715293/2013 (SWZ13) and a mutant strain of SWZ13, even though no N-glycosylation sequons were changed. We determined site-specifically that WT and mutant HA have varying similarity at the glycosylation sites of the head domain, reflecting competing pressures to evade host immune response while retaining viral fitness. To our knowledge, our results are the first to quantify changes in glycosylation state that occur in related proteins of considerable glycan heterogeneity. Our results provide a method for understanding how changes in glycosylation state are correlated with variations in protein sequence, which is necessary for improving IAV vaccine strain selection. Understanding glycosylation will be especially important as we find new expression vectors for vaccine production, as glycosylation state depends greatly on the host species.
2020-07-03 | PXD018920 | Pride
Project description:Gene transcription effects of mutations in the infuenza virus A/Hong Kong/1/1968(H3N2) nonstructural 1 NS1 gene in infected human A549 (lung epithilium) cells Influenza A/Hong Kong/156/1997(H5N1) virus NS1 gene mutations F103L and M106I both increase IFN antagonism, virulence and cytoplasmic localization but differ in binding to RIG-I and CPSF30 (manuscript submitted to Virology Journal). Human cells were infected with influenza viruses mutants with specific gain of function mutations in the NS1 gene in order to assess the affects of each mutation on host gene expression. Human (A549) and mouse (M1) cells were infected at a multiplicity of infection of 2 (infectious viruses/cell) and incubated for 8 hr before collection of total RNA and microarray anlaysis using the Affymetrix platforms. Samples were compared in triplicate to mock PBS infected (uninfected) cells to detecte dysregulated genes for A/HK/1/1968(H3N2) wt, and the following NS1 gene mutants: F103L, M106I, M106V, and F103L + M106I. Background: The genetic basis for avian to mammalian host switching in influenza A virus is largely unknown. The human A/HK/156/1997(H5N1) virus that transmitted from poultry possesses NS1 gene mutations F103L + M106I that are virulence determinants in the mouse model of pneumonia; however their individual roles have not been determined. The emergent A/Shanghai/1/2013(H7N9)-like viruses also possess these mutations which may contribute to their virulence and ability to switch species. Methods: NS1 mutant viruses were constructed by reverse genetics and site directed mutagenesis on human and mouse-adapted backbones. Mouse infections assessed virulence, virus yield, tissue infection, and IFN induction. NS1 protein proprieties were assessed for subcellular distribution, IFN antagonism (mouse and human), CPSF30 and RIG-I domain binding, effect on host gene transcription (microarray); and the natural prevalence of 103L and 106I mutants was assessed. Results: Each of the F103L and M106I mutations contributes additively to virulence to reduce the lethal dose by >800 and >3,200 fold respectively by mediating alveolar tissue infection with >100 fold increased infectious yields. The 106I NS1 mutant lost CPSF binding but the 103L mutant maintained binding that correlated with an increased general decrease in host gene expression in human but not mouse cells. Each mutation positively modulated the inhibition of IFN induction in mouse cells and activation of the IFN-M-NM-2 promoter in human cells but not in combination in human cells indicating negative epistasis. Each of the F103L and M106I mutations restored a defect in cytoplasmic localization of H5N1 NS1 in mouse cells. Human H1N1 and H3N2 NS1 proteins bound to the CARD, helicase and RD RIG-I domains, whereas the H5N1 NS1 with the same consensus 103F and 106M mutations did not bind these domains, which was partially or totally restored by the F103L or M106I mutations respectively. Conclusions: The F103L and M106I mutations in the H5N1 NS1 protein each increased IFN antagonism and mediated interstitial pneumonia in mice that was associated with increased cytoplasmic localization and altered host factor binding. These mutations may contribute to the ability of previous HPAI H5N1 and recent LPAI H7N9 viruses to switch hosts and cause severe disease in mammals. Triplicate biological replicates of mock PBS treated (uninfected) cells to detecte dysregulated genes for A/HK/1/1968(H3N2) wt, and the following NS1 gene mutants: F103L, M106I, M106V, and F103L + M106I. Cells were infected at a multiplicty of infection of 2 and cells were incubated for 8 hr at 37 C for 8 hrs before RNA extraction and analysis of 3 biological replicates relative to mock PBS infected cells.
2013-06-22 | E-GEOD-48200 | biostudies-arrayexpress
Project description:This study revealed important similarities but also critical differences between the H5N1 and 1918-reassortant viruses, highlighting aspects of the host–pathogen interface caused by highly virulent influenza viruses. Animals assigned to 4 experimental groups (n = 8) matched for age, weight, and sex, were inoculated by intratracheal, intranasal, tonsillar, and conjunctival routes with a total of 107 pfu of A/Vietnam/1203/2004 (H5N1) virus, A/Texas/36/91 (H1N1) virus or reassortants of this virus containing either 2 (HA, NA) or 3 (HA, NA, NS) genes from the 1918 virus. Two animals per group were scheduled for sacrifice on days 1, 2, 4, and 7. Two additional animals were used as uninfected control animals and killed on day 7. Oligoarray analyses consisted of comparing array profiles of individual lung samples (with representative pathology and degree of infection) from infected animals to pooled equal masses of mRNA from all lung lobes of 7 reference cynomolgus macaques obtained through the tissue program of the University of Washington National Primate Research Center. There are 2 technical array replicates fro each sample.
2011-11-01 | E-GEOD-33351 | biostudies-arrayexpress