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Understanding the molecular basis leading the virulence of vesicular stomatitis virus in pigs

ABSTRACT: Understanding the molecular basis leading the virulence of vesicular stomatitis virus in pigs

PROVIDER: PRJNA937643 | ENA |

REPOSITORIES: ENA

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Understanding the molecular basis leading the virulence of vesicular stomatitis virus in pigs

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Next-generation sequencing of three memory OT-I cell subsets from mice infected with vesicular stomatitis virus

Project description:Purpose: RNA-seq analysis of three memory OT-I cell subsets (from a Klrg1-Cre fate reporter mouse model) isolated from the spleen of C57BL/6 mice infected with vesicular stomatitis virus. The hypothesis tested in the present study was that KLRG1+ effector CD8 T lymphocytes differentiate into KLRG1- memory CD8 T lymphocytes and provide long-lasting immunity against infectious diseases and malignancies. Methods: Total RNA was obtained from FACS-purified OT-I cell subsets isolated from spleen 70 days post infection with ovalbumin-expressing vesicular stomatitis virus (VSV-OVA) (experiment 3). Results: Using RNA-seq technology, we performed genome-wide transcriptional profiling of three memory OT-I cells (KLRG1+ Reporter+, KLRG1- Reporter+ (exKLRG1) and KLRG1- Reporter-) and identified 36 genes differentially expressed (> 1.5-fold) between exKLRG1 and KLRG1- Reporter- memory OT-I cells, and 132 differentially expressed genes between exKLRG1 and KLRG1+ Reporter+ memory OT-I cells. We then confirmed the expression of 15 genes/molecules by qRT-PCR and/or flow cytometry. Conclusions: Our study represents the first fate mapping analysis of KLRG1+ effector OT-I cells, demonstrates that KLRG1+ effector OT-I cells differentiate into all memory T cell lineages thereby promoting protective immunity. RNA-seq also identified CX3CR1 as a marker of circulating exKLRG1 early memory OT-I cells.

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Analysis of global gene expression profiles of hPAF1 deficient A549 cells during infection with H1N1 influenza A virus or vesicular stomatitis virus (VSV)

Project description:Viral infection is commonly associated with virus-driven hijacking of host proteins. We describe a novel mechanism by which influenza virus impacts host cells through the interaction of influenza NS1 protein with the infected cell epigenome. We show that the NS1 protein of influenza A H3N2 targets the transcription elongation PAF1 complex (hPAF1C). We demonstrate that binding of NS1 to hPAF1C results in suppression of hPAF1C-mediated transcriptional elongation. More importantly,in the following data sets, we show that hPAF1 plays a crucial role in the antiviral response. Loss of hPAF1C reduces antiviral gene expression and reduces inducible transcription of target genes after stimulation with viral RNA analogue poly(I:C), vesicular stomatitis virus (VSV), exogenous recombinant IFN(beta) and influenza virus (H1N1). This study underscores the importance of hPAF1C in controlling inducible antiviral gene expression. Untreated (no siRNA), control siRNA-treated and hPAF1 siRNA-treated A549 cells were stimulated with A/Puerto Rico/8/1934 influenza virus (H1N1) or vesicular stomatitis virus (VSV). Total RNA was isolated with the Qiagen RNeasy mini kit. 200ng of total RNA per sample was used to prepare biotin-labeled RNA using MessageAmp™ Premier RNA Amplification Kit (Applied Biosystems) and hybridized to HumanHT-12 v4 Expression BeadChips (Illumina). Data analysis was performed using the GeneSpring GX11.0 software (Agilent Technologies). 3 biological replicates per condition

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