Phorbol ester-induced human cytomegalovirus major immediate-early (MIE) enhancer activation through PKC-delta, CREB, and NF-kappaB desilences MIE gene expression in quiescently infected human pluripotent NTera2 cells.

The ways in which human cytomegalovirus (HCMV) major immediate-early (MIE) gene expression breaks silence from latency to initiate the viral replicative cycle are poorly understood. A delineation of the signaling cascades that desilence the HCMV MIE genes during viral quiescence in the human pluripotent N-Tera2 (NT2) cell model provides insight into the molecular mechanisms underlying HCMV reactivation. In this model, we show that phorbol 12-myristate 13-acetate (PMA) immediately activates the expression of HCMV MIE RNA and protein and greatly increases the MIE-positive (MIE(+)) NT2 cell population density; levels of Oct4 (pluripotent cell marker) and HCMV genome penetration are unchanged.

Breaking human cytomegalovirus major immediate-early gene silence by vasoactive intestinal peptide stimulation of the protein kinase A-CREB-TORC2 signaling cascade in human pluripotent embryonal NTera2 cells.

The triggering mechanisms underlying reactivation of human cytomegalovirus (HCMV) in latently infected persons are unclear. During latency, HCMV major immediate-early (MIE) gene expression breaks silence to initiate viral reactivation. Using quiescently HCMV-infected human pluripotent embryonal NTera2 cells (NT2) to model HCMV reactivation, we show that vasoactive intestinal peptide (VIP), an immunomodulatory neuropeptide, immediately and dose-dependently (1 to 500 nM) activates HCMV MIE gene expression.

Reversal of human cytomegalovirus major immediate-early enhancer/promoter silencing in quiescently infected cells via the cyclic AMP signaling pathway.

The human cytomegalovirus (HCMV) major immediate-early (MIE) enhancer contains five functional cyclic AMP (cAMP) response elements (CRE). Because the CRE in their native context do not contribute appreciably to MIE enhancer/promoter activity in lytically infected human fibroblasts and NTera2 (NT2)-derived neurons, we postulated that they might have a role in MIE enhancer/promoter reactivation in quiescently infected cells. Here, we show that stimulation of the cAMP signaling pathway by treatment with forskolin (FSK), an adenylyl cyclase activator, greatly alleviates MIE enhancer/promoter silencing in quiescently infected NT2 neuronal precursors.