Expression of TNF-α by Herpes Simplex Virus-Infected Macrophages Is Regulated by a Dual Mechanism: Transcriptional Regulation by NF-κB and Activating Transcription Factor 2/Jun and Translational Regulation Through the AU-Rich Region of the 3′ Untranslated Region

Abstract

Here we have investigated the regulation of TNF-α expression in macrophages during HSV-2 infection. Despite a low basal level of TNF-α mRNA present in resting macrophages, no TNF-α protein is detectable. HSV-2 infection marginally increases the level of TNF-α mRNA and protein in resting macrophages, whereas a strong increase is observed in IFN-γ-activated cells infected with the virus. By reporter gene assay it was found that HSV infection augments TNF-α promoter activity. Moreover, treatment of the cells with actinomycin D, which totally blocked mRNA synthesis, only partially prevented accumulation of TNF-α protein, indicating that the infection lifts a block on translation of TNF-α mRNA. EMSA analysis showed that specific binding to the κB#3 site of the murine TNF-α promoter was induced within 1 h after infection and persisted beyond 5 h where TNF-α expression is down-modulated. Binding to the cAMP responsive element site was also induced but more transiently with kinetics closely following activation of the TNF-α promoter. Inhibitors against either NF-κB activation or the activating transcription factor 2 kinase p38 abrogated TNF-α expression, showing a requirement for both signals for activation of the promoter. This observation was corroborated by reporter gene assays. As to the translational regulation of TNF-α, the AU-rich sequence in the 3′ untranslated region of the mRNA was found to be responsible for this control because deletion of this region renders mRNA constitutively translationable. These results show that TNF-α production is induced by HSV-2 in macrophages through both transcriptional and translational regulation.

Induction of RANTES/CCL5 by herpes simplex virus is regulated by nuclear factor κB and interferon regulatory factor 3

“…2). NF-kBdependent HSV-induced production of antiviral substances such as IL-6, IL-12, TNF-a and NO has been established (Paludan et al, 1998;Malmgaard et al, 2000;Paludan, 2001;Paludan et al, 2001) and the present finding suggests that expression of RANTES in HSV-infected cells, also relies on NF-kB.…”

“…Previous studies from this laboratory have shown that production of IL-6 and TNF-a in HSV-infected macrophages is dependent on p38 (Paludan, 2001;Paludan et al, 2001). To investigate the role of p38 in the expression of RANTES we used SB203580, a highly specific inhibitor of p38 MAP kinase.…”

Toll-like Receptor 9 Regulates Tumor Necrosis Factor-α Expression by Different Mechanisms

“…TNF-␣ gene expression has been studied extensively and shown to be regulated on multiple levels: transcription, mRNA stability, translation, and protein stability (42)(43)(44)(45)(46). Despite a higher level of TNF-␣ release in response to CpG-ODN by cells derived from BALB/c, the induced cytokine mRNA levels were lower in these cells.…”

Post‐transcriptional regulation of gene expression by degradation of messenger RNAs

“…As to the translational regulation of TNFa, the ARE in the 3 0 UTR of the mRNA was found to be responsible for this control because deletion of this region renders mRNA constitutively translatable (Paludan et al, 2001). A feedback mechanism of protein level regulation would not be unusual in the cell.…”