Summary auto-generated
This study reveals an unexpected mechanism of HSV-1 immediate early (IE) gene regulation. Using HSV-1 mutants deficient in three major transactivators (VP16, ICP0, and ICP4), researchers found that inhibition of protein synthesis with cycloheximide paradoxically stimulates IE gene expression from the ICP0 and ICP27 promoters, similar to how cellular IE genes respond to protein synthesis inhibitors. This activation occurs through increased mRNA production rather than mRNA stabilization. The effect was confirmed using alternative protein synthesis inhibitors (anisomycin, puromycin, emetine), with cycloheximide and anisomycin showing stronger activation correlating with their ability to activate cellular stress-response pathways. The human cytomegalovirus major IE promoter, when integrated into HSV-1, also showed this response. These findings suggest that cellular signal transduction pathways activated by translation inhibition can influence viral IE gene expression, potentially important during viral latency and reactivation when VP16 may be inactive.
Key findings
- HSV-1 IE promoters (ICP0 and ICP27) are stimulated by protein synthesis inhibitors through a mechanism independent of the three major viral transactivators (VP16, ICP0, ICP4)
- Activation results from increased mRNA production rather than mRNA stabilization
- The response correlates with activation of cellular stress-response pathways, particularly SAPK/c-Jun N-terminal kinase signaling
- Human cytomegalovirus IE promoter shows similar activation when integrated into HSV-1 genome
- This regulatory mechanism may be functionally important during viral latency and reactivation when VP16 is unavailable
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Abstract
Herpes simplex virus type 1 (HSV-1) transcription can be arrested at the immediate early (IE) stage by continuous treatment of cells with inhibitors of protein synthesis, usually cycloheximide, from the time of infection. We have analysed the effect of cycloheximide on IE gene expression with HSV-1 mutants deficient in the production of functional levels of the three major transactivators, the virion protein (VP16) and two IE proteins (ICP0 and ICP4). Expression from the HSV-1 IE promoters that control synthesis of ICP0 and ICP27 was, unexpectedly, stimulated by inhibition of protein synthesis. The effect was observed for the ICP0 promoter in its normal genome location and also when cloned upstream of the Escherichia coli lacZ coding sequences and inserted into the viral thymidine kinase locus. Expression from the human cytomegalovirus major IE promoter, when cloned into the genome of HSV-1 mutants, was also increased by inhibition of protein synthesis. Cycloheximide did not affect the intracellular stability of lacZ- specific RNA, suggesting that the response represented an increase in mRNA production. Activation of the ICP0 promoter was observed when protein synthesis was blocked by alternative agents. Since inhibitors of protein synthesis are known to activate cellular signal transduction pathways, our findings demonstrate new mechanisms for the regulation of HSV-1 IE gene expression which may be important during latency and reactivation. The results also highlight previously unrecognized difficulties in analysing the intrinsic activities of promoters when cloned into the HSV-1 genome.