A Novel Naturally Occurring Tandem Promoter in Modified Vaccinia Virus Ankara Drives Very Early Gene Expression and Potent Immune Responses

Vaccinia virus (VV) infects a broad range of host cells, and while it usually causes their lysis (i.e. necrosis), the nature of the cell-death phenomenon is not well understood. In this study, we show that VV induces apoptosis of cells of the murine macrophage line J774.G8, as revealed by morphological signs, DNA ladder formation, changes of mitochondrial membrane potential and annexin-V positivity. Apoptosis occurred in both untreated and IFN-γ-pretreated macrophages, and could not be inhibited by aminoguanidine, a relatively specific inhibitor of inducible nitric oxide synthase. Inhibition of VV DNA synthesis and late gene expression by cytosine arabinoside also did not prevent apoptosis, while heat- or psoralen/UV-inactivated VV did not cause any apoptosis. Thus, VV early gene expression seems to be required for induction of apoptosis. At the cellular level, infection with VV induced a decrease in the levels of Bcl-xL, an anti-apoptotic member of the Bcl-2 family. The importance of loss of Bcl-xL was demonstrated by prevention of VV-mediated apoptosis on expression of Bcl-2, a functional homologue of Bcl-xL. Our findings provide evidence that induction of apoptosis by VV in macrophages requires virus early gene expression, does not involve nitric oxide, induces a decrease in mitochondrial membrane potential and is associated with altered levels of Bcl-xL.

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Microarray Analysis Reveals Characteristic Changes of Host Cell Gene Expression in Response to Attenuated Modified Vaccinia Virus Ankara Infection of Human HeLa Cells

Journal of Virology ◽

10.1128/jvi.78.11.5820-5834.2004 ◽

2004 ◽

Vol 78 (11) ◽

pp. 5820-5834 ◽

Cited By ~ 57

Author(s):

Susana Guerra ◽

Luis A. López-Fernández ◽

Raquel Conde ◽

Alberto Pascual-Montano ◽

Keith Harshman ◽

...

Keyword(s):

Gene Expression ◽

Immune Responses ◽

Vaccinia Virus ◽

Immune Modulation ◽

Expression Patterns ◽

Family Members ◽

Signal Transduction Pathway ◽

Gene Clusters ◽

Altered Expression ◽

Modified Vaccinia Virus Ankara

ABSTRACT The potential use of the modified vaccinia virus Ankara (MVA) strain as a live recombinant vector to deliver antigens and elicit protective immune responses against infectious diseases demands a comprehensive understanding of the effect of MVA infection on human host gene expression. We used microarrays containing more than 15,000 human cDNAs to identify gene expression changes in human HeLa cell cultures at 2, 6, and 16 h postinfection. Clustering of the 410 differentially regulated genes identified 11 discrete gene clusters with altered expression patterns after MVA infection. Clusters 1 and 2 (accounting for 16.59% [68 of 410] of the genes) contained 68 transcripts showing a robust induction pattern that was maintained during the course of infection. Changes in cellular gene transcription detected by microarrays after MVA infection were confirmed for selected genes by Northern blot analysis and by real-time reverse transcription-PCR. Upregulated transcripts in clusters 1 and 2 included 20 genes implicated in immune responses, including interleukin 1A (IL-1A), IL-6, IL-7, IL-8, and IL-15 genes. MVA infection also stimulated the expression of NF-κB and components of the NF-κB signal transduction pathway, including p50 and TRAF-interacting protein. A marked increase in the expression of histone family members was also induced during MVA infection. Expression of the Wiskott-Aldrich syndrome family members WAS, WASF1, and the small GTP-binding protein RAC-1, which are involved in actin cytoskeleton reorganization, was enhanced after MVA infection. This study demonstrates that MVA infection triggered the induction of groups of genes, some of which may be involved in host resistance and immune modulation during virus infection.

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