Role of GTPase activity of murine Mx1 protein in nuclear localization and anti-influenza virus activity

Toyoda, Tetsuya; Asano, Yukiyasu; Ishihama, Akira

doi:10.1099/0022-1317-76-7-1867

Cited by 7 publications

(3 citation statements)

References 10 publications

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“…If the sole purpose of Mx nuclear domains is to serve as storage depots, these domains are expected to be dispensable for antiviral activity. This is in line with previous findings showing that Mx1(C71S), a mutant form of Mx1 with a single amino acid exchange at position 71, is antiviral, although this mutant does not form the characteristic nuclear dots (Toyoda et al, 1995). However, using our high-expression vectors, this Mx1 mutant was able to accumulate in nuclear dots in a majority of cells (see Supplementary Figure in JGV Online), suggesting that dot formation most likely depends on protein expression levels, as discussed above.…”

Section: Discussionsupporting

confidence: 93%

Mx1 GTPase accumulates in distinct nuclear domains and inhibits influenza A virus in cells that lack promyelocytic leukaemia protein nuclear bodies

Engelhardt

Sirma

Pandolfi

et al. 2004

Journal of General Virology

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The interferon-induced murine Mx1 GTPase is a nuclear protein. It specifically inhibits influenza A viruses at the step of primary transcription, a process known to occur in the nucleus of infected cells. However, the exact mechanism of inhibition is still poorly understood. The Mx1 GTPase has previously been shown to accumulate in distinct nuclear dots that are spatially associated with promyelocytic leukaemia protein (PML) nuclear bodies (NBs), but the significance of this association is not known. Here it is reported that, in cells lacking PML and, as a consequence, PML NBs, Mx1 still formed nuclear dots. These dots were indistinguishable from the dots observed in wild-type cells, indicating that intact PML NBs are not required for Mx1 dot formation. Furthermore, Mx1 retained its antiviral activity against influenza A virus in these PML-deficient cells, which were fully permissive for influenza A virus. Nuclear Mx proteins from other species showed a similar subnuclear distribution. This was also the case for the human MxA GTPase when this otherwise cytoplasmic protein was translocated into the nucleus by virtue of a foreign nuclear localization signal. Human MxA and mouse Mx1 do not interact or form heterooligomers. Yet, they co-localized to a large degree when co-expressed in the nucleus. Taken together, these findings suggest that Mx1 dots represent distinct nuclear domains ('Mx nuclear domains') that are frequently associated with, but functionally independent of, PML NBs.

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Section: Discussionsupporting

confidence: 93%

Mx1 GTPase accumulates in distinct nuclear domains and inhibits influenza A virus in cells that lack promyelocytic leukaemia protein nuclear bodies

Engelhardt

Sirma

Pandolfi

et al. 2004

Journal of General Virology

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“…While we have identified an increasing rate of 2–5 OAS2 synthesis as HIV-1 infection progresses, viral infection is not diminished as demonstrated by both HIV-1p24 staining and RT activity. Mx1 and GBP (Supplemental Figure 1) both have roles in the inhibition of hepatitis C. , Mx1 also has been implicated in inhibiting influenza viral growth, and GBP is known to inhibit both vesicular stomatitis and encephalomyocarditis viruses . MX1 induction during HIV-1 infection has been identified by multiple groups .…”

Section: Resultsmentioning

confidence: 99%

Pulsed Stable Isotope Labeling of Amino Acids in Cell Culture Uncovers the Dynamic Interactions between HIV-1 and the Monocyte-Derived Macrophage

et al. 2011

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Dynamic interactions between human immunodeficiency virus-1 (HIV-1) and the macrophage govern the tempo of viral dissemination and replication in its human host. HIV-1 affects macrophage phenotype, and the macrophage, in turn, can modulate the viral life cycle. While these processes are linked to host–cell function and survival, the precise intracellular pathways involved are incompletely understood. To elucidate such dynamic virus–cell events, we employed pulsed stable isotope labeling of amino acids in cell culture. Alterations in de novo protein synthesis of HIV-1 infected human monocyte-derived macrophages (MDM) were examined after 3, 5, and 7 days of viral infection. Synthesis rates of cellular metabolic, regulatory, and DNA packaging activities were decreased, whereas, those affecting antigen presentation (major histocompatibility complex I and II) and interferon-induced antiviral activities were increased. Interestingly, enrichment of proteins linked to chromatin assembly or disassembly, DNA packaging, and nucleosome assembly were identified that paralleled virus-induced cytopathology and replication. We conclude that HIV-1 regulates a range of host MDM proteins that affect its survival and abilities to contain infection.

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“…Previous attempts to define Mx domains important for antiviral function by mutational analyses of human or mouse Mx proteins have been hampered by the fact that most mutations resulted in a complete loss of antiviral activity (8,22,32,46,51). Therefore, we chose an alternative approach to identify functional domains in human MxA protein.…”

mentioning

confidence: 99%

Dominant-negative mutants of human MxA protein: domains in the carboxy-terminal moiety are important for oligomerization and antiviral activity

Ponten¹,

Sick²,

Weeber³

et al. 1997

J Virol

115

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Human MxA protein is an interferon-induced 76-kDa GTPase that exhibits antiviral activity against several RNA viruses. Wild-type MxA accumulates in the cytoplasm of cells. TMxA, a modified form of wild-type MxA carrying a foreign nuclear localization signal, accumulates in the cell nucleus. Here we show that MxA protein is translocated into the nucleus together with TMxA when both proteins are expressed simultaneously in the same cell, demonstrating that MxA molecules form tight complexes in living cells. To define domains important for MxA-MxA interaction and antiviral function in vivo, we expressed mutant forms of MxA together with wild-type MxA or TMxA in appropriate cells and analyzed subcellular localization and interfering effects. An MxA deletion mutant, MxA(359-572), formed heterooligomers with TMxA and was translocated to the nucleus, indicating that the region between amino acid positions 359 and 572 contains an interaction domain which is critical for oligomerization of MxA proteins. Mutant T103A with threonine at position 103 replaced by alanine had lost both GTPase and antiviral activities. T103A exhibited a dominant-interfering effect on the antiviral activity of wild-type MxA rendering MxA-expressing cells susceptible to infection with influenza A virus, Thogoto virus, and vesicular stomatitis virus. To determine which sequences are critical for the dominantnegative effect of T103A, we expressed truncated forms of T103A together with wild-type protein. A C-terminal deletion mutant lacking the last 90 amino acids had lost interfering capacity, indicating that an intact C terminus was required. Surprisingly, a truncated version of MxA representing only the C-terminal half of the molecule exerted also a dominant-negative effect on wild-type function, demonstrating that sequences in the C-terminal moiety of MxA are necessary and sufficient for interference. However, all MxA mutants formed hetero-oligomers with TMxA and were translocated to the nucleus, indicating that physical interaction alone is not sufficient for disturbing wild-type function. We propose that dominant-negative mutants directly influence wild-type activity within hetero-oligomers or else compete with wild-type MxA for a cellular or viral target.

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Role of GTPase activity of murine Mx1 protein in nuclear localization and anti-influenza virus activity

Cited by 7 publications

References 10 publications

Mx1 GTPase accumulates in distinct nuclear domains and inhibits influenza A virus in cells that lack promyelocytic leukaemia protein nuclear bodies

Mx1 GTPase accumulates in distinct nuclear domains and inhibits influenza A virus in cells that lack promyelocytic leukaemia protein nuclear bodies

Pulsed Stable Isotope Labeling of Amino Acids in Cell Culture Uncovers the Dynamic Interactions between HIV-1 and the Monocyte-Derived Macrophage

Dominant-negative mutants of human MxA protein: domains in the carboxy-terminal moiety are important for oligomerization and antiviral activity

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