Subcellular Localization of MxB Determines Its Antiviral Potential against Influenza A Virus

ABSTRACT Mx proteins are interferon (IFN) type I (α/β)- and type III (λ)-induced effector proteins with intrinsic antiviral activity. Mammalian Mx proteins show different subcellular localizations and distinct yet partially overlapping viral specificities. However, the precise mechanism(s) of antiviral action are still unresolved. Human MxA accumulates in the cytoplasm and inhibits a wide variety of RNA and DNA viruses, among them influenza A virus (IAV). In contrast, MxB, the second human Mx protein, localizes via its amino (N) terminus to the outer nuclear membrane at or near nuclear pores and inhibits the nuclear import of incoming human immunodeficiency viruses (HIV) and herpesviruses, but not that of IAV. Here, we evaluated whether the antiviral specificity of MxB is determined by its subcellular localization. For this purpose, we redirected MxB to the nucleus or cytoplasm by either attaching a nuclear localization signal to its N terminus or by exchanging the N terminus of MxB with that of MxA. Interestingly, ectopic expression of these MxB variants in the nucleus or in the cytoplasm rendered the host cells resistant to IAV, revealing that the capacity of MxB to block IAV replication critically depends on the site where the protein accumulates in the infected cell. Furthermore, coimmunoprecipitation (co-IP) assays demonstrated that MxB physically interacted with the nucleoprotein (NP) of IAV. Taken together, the data indicate that the subcellular localization of the MxB protein plays a pivotal role in determining its antiviral specificity. IMPORTANCE The interferon system plays a pivotal role in the defense against viral infections. The dynamin-related Mx proteins form a small family of interferon-induced effector proteins with distinct antiviral specificities and subcellular localizations. So far, it is not clear whether the different virus specificities of Mx proteins are the result of distinct mechanisms of action or are due rather to their different subcellular localization. We show here that the human MxB protein, normally localized to the outer membrane of the cell nucleus, acquires antiviral activity against IAV when redirected to the nucleus or cytoplasm, subcellular sites where other members of the Mx protein family efficiently interfere with IAV replication. Our findings thus strongly suggest that Mx proteins act primarily through a common mechanism and that their viral specificity is at least in part determined by their individual subcellular localization.

Interferon-Stimulated Genes—Mediators of the Innate Immune Response during Canine Distemper Virus Infection

The demyelinating canine distemper virus (CDV)-leukoencephalitis represents a translational animal model for multiple sclerosis. The present study investigated the expression of type I interferon (IFN-I) pathway members in CDV-induced cerebellar lesions to gain an insight into their role in lesion development. Gene expression of 110 manually selected genes in acute, subacute and chronic lesions was analyzed using pre-existing microarray data. Interferon regulatory factor (IRF) 3, IRF7, signal transducer and activator of transcription (STAT) 1, STAT2, MX protein, protein kinase R (PKR), 2′-5′-oligoadenylate synthetase (OAS) 1 and interferon-stimulated gene (ISG) 15 expression were also evaluated using immunohistochemistry. Cellular origin of STAT1, STAT2, MX and PKR were determined using immunofluorescence. CDV infection caused an increased expression of the antiviral effector proteins MX, PKR, OAS1 and ISG15, which probably contributed to a restricted viral replication, particularly in neurons and oligodendrocytes. This increase might be partly mediated by IRF-dependent pathways due to the lack of changes in IFN-I levels and absence of STAT2 in astrocytes. Nevertheless, activated microglia/macrophages showed a strong expression of STAT1, STAT2 and MX proteins in later stages of the disease, indicating a strong activation of the IFN-I signaling cascade, which might be involved in the aggravation of bystander demyelination.

Chicken Mx gene polymorphisms in Indian native chicken breeds and White Leghorn by real time multiplex allele specific PCR

Chicken Mx protein confers resistance against viral infections including avian influenza virus. Mx protein is one of the strong antiviral proteins induced by interferon system. Different alleles of chicken Mx gene have been shown to vary in their antiviral activity against influenza virus. A non-synonymous mutation at position 2032 of chicken Mx gene responsible for variation of amino acid (Ser to Asn) in Mx protein is responsible for negative to positive antiviral activity of Mx protein. In the present study it was analyzed that SNP at G2032A in chicken Mx gene in two Indian native chicken breeds, Aseel, Kadaknath and compared with White Leghorn layer. A total of 90 samples, including 50 of White Leghorn and 20 each from native chicken breed were screened for SNP at position 2032 of Mx gene by using real time multiplex allele specific PCR. The resistant allele (A) frequency is higher than susceptible allele in all three chicken breeds investigated. Among the breeds, the resistant allele frequency (A) is higher in White Leghorn (0.95) followed by Aseel (0.75) and Kadaknath (0.625). Homozygous to resistant allele (AA) is the major genotype in White Leghorn whereas heterozygous to both alleles is the major genotype in Kadaknath. Variations in the gene frequency might have emerged from varying degree of selection pressure the different breeds had undergone. In future, the resistant genotype could be selected in modern breeding programs to improve the genetic status for disease resistance against viral infections