NF-kappaB--a potential therapeutic target for inhibition of human cytomegalovirus (re)activation?

From clinical studies the proinflammatory cytokine TNFalpha was proposed to play a key role in human cytomegalovirus (HCMV) reactivation from latency. In vitro experiments confirmed that TNFalpha stimulates the activity of the HCMV IE1/2 enhancer/promoter, which controls immediate early protein IE1 and IE2 gene expression via activation of the transcription factor NF-kappaB and its binding to putative binding sites in the IE1/2 enhancer. NF-kappaB was also proposed to be involved in IE1-mediated autostimulation of this promoter. The IE1/2 enhancer of HCMV contains four putative NF-kappaB binding sites which differ in their distance to the transcription start site as well as in their sequence. Construction and testing of a series of promoter mutants demonstrated that NF-kappaB is essential for both TNFalpha and IE1 stimulation. Furthermore, we were able to show that although all four NF-kappaB sites bind NF-kappaB with similar affinity in vitro, the contribution to TNFalpha and IE1 stimulation differs in correlation with the distance to the transcription start site and the sequence. Site 1 and 3 play the most dominant role and site 2 an intermediate, while site 4, which is conserved in sequence but far distant from the transcription start site, had no influence on NF-kappaB-mediated regulation of the IE1/2 promoter. Specific inhibition of NF-kappaB signalling by co-expression of a dominant-negative IkappaB variant reduced TNFalpha stimulation of the IE1/2 enhancer/promoter by up to 80%. From this data, inhibitors of NF-kappaB activation are suggested to be an alternative therapeutical strategy to interfere with HCMV (re)activation in undifferentiated monocyte/granulocyte progenitor cells in patients with a high risk of inflammation-related HCMV (re)activation.