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The NS1 Protein of the Parvovirus MVM Aids in the Localization of the Viral Genome to Cellular Sites of DNA Damage

ABSTRACT: The autonomous parvovirus Minute Virus of Mice (MVM) localizes to cellular sites of DNA damage to establish viral replication centers, visualized by focal depositions of the essential MVM phosphoprotein NS1. However, how such foci are established remains unknown. Here, we show that MVM NS1 localized to cellular sites of DNA damage independently of its known ability to covalently bind the viral genome at its known consensus DNA binding site. NS1 that retained its capacity to bind its cognate DNA binding sequence could relocate either the viral genome, or heterologous DNA, containing those sites, to cellular sites of DNA damage. These findings suggest that another way that NS1 can facilitate virus infection is by localizing the genome to cellular sites which contain replication and expression proteins.

ORGANISM(S): Mus musculus

PROVIDER: GSE152601 | GEO | 2020/10/03

REPOSITORIES: GEO

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Project description:The NS1 protein of influenza A virus (IAV) is a multifunctional virulence factor. Mouse adaptive mutations in the NS1 protein of the human isolate A/Hong Kong/1/1968(H3N2) (HK) have been previously reported to increase virulence, viral fitness, and interferon antagonism, but differ in binding to post-transcriptional processing factor CPSF30. Because nuclear trafficking is a major genetic determinant of influenza virus host adaptation, we assessed subcellular localization and host gene expression of NS1 adaptive mutations. Recombinant HK viruses with adaptive mutations in the NS1 gene were assessed for NS1 protein subcellular localization in mouse and human cells using confocal microscopy and cellular fractionation. HK-wt virus NS1 partitioned equivalently between the cytoplasm and nucleus in human cells but was defective in cytoplasmic localization in mouse cells. The adaptive mutations either increased the proportion or abundance of NS1 in the cytoplasm, and/or the nucleus. NS1 mutations that increased cytoplasmic distribution identified a putative second nuclear export signal (NES) spanning aa positions 98-106 LSEDWFMLM, (mutation sites in bold); with the strongest effect seen for mutation M106I. The putative NES in the NS3 protein was associated with cytoplasmic localization. The host gene expression profile of the adaptive mutants was determined by microarray analysis of infected mouse cells to show either high or low gene regulation (HGR or LGR) phenotypes that mapped to the amino-terminal and the carboxy-terminal regions respectively. The HGR and LGR mutations were predominantly down regulating versus up regulating respectively. The greatest effect on host gene expression in the HGR group correlated with the ability of the NS1 protein to bind CPSF30. To our knowledge this is the first report of roles of adaptive NS1 mutations that affect intracellular localization and regulation of host gene expression. biological triplicates of mock infected mouse M1 cells; cells infected with A/HK/1/1968(H3N2) wt and NS1 mtuatn with mutaions, D2N, V23A, L98S, L98S + M106I, F103L, M106V, M106V + M124I, D125G, V180A, V226I, M106I, and R227K. Cells were infected at a multiplicty of infection of 2 and cels were incubated for 8 hr at 37 C for 8 hrs before RNA extraction and analysis relative to mock PBS infected cells.

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The NS1 Protein of the Parvovirus MVM Aids in the Localization of the Viral Genome to Cellular Sites of DNA Damage

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