Human Cytomegalovirus Circumvents NF-κB Dependence in Retinal Pigment Epithelial Cells

Abstract

The human CMV (HCMV) is a persistent virus that may cause severe inflammatory responses especially in immunocompromised hosts. In different cell types, HCMV infection leads to the activation of the pleiotropic transcription factor, NF-κB, which triggers virus replication but also propagates cell-mediated inflammatory mechanisms that largely depend on PG synthesis. We investigated the interactions of HCMV and the NF-κB-dependent PG synthesis pathway in cultures of retinal pigment epithelial (RPE) cells that are known to be infected in HCMV retinitis patients. Unlike in other cell types, HCMV increased neither NF-κB activity nor p65 and p105/50 mRNA levels in RPE cells. Both TNF-α and phorbol ester 12,0-tetradecanoylphorbol 13-acetate (TPA) enhanced NF-κB activity but only TPA increased HCMV replication. Cyclooxygenase-2 expression and PGE2 release was increased by TPA and TNF-α but not by HCMV infection. Stimulatory activity of TPA on HCMV replication was suppressed by protein kinase C inhibitors and inhibitors of p42/44 and p38 mitogen-activated protein kinases but not by NF-κB inhibitors. In conclusion, HCMV circumvents the NF-κB route in favor of the protein kinase C-dependent mitogen-activated protein kinase pathway in RPE cells. This virus/host cell interaction might be a mechanism that promotes HCMV persistence in immune-privileged organs such as the eye.

Phorbol Ester-Induced Human Cytomegalovirus Major Immediate-Early (MIE) Enhancer Activation through PKC-Delta, CREB, and NF-κB Desilences MIE Gene Expression in Quiescently Infected Human Pluripotent NTera2 Cells

“…Phorbol esters have been reported to enhance active HCMV infection in acutely infected human umbilical vein endothelial cells (HUVEC) (55) and retinal pigmented epithelial (RPE) cells (13). Phorbol ester treatment of HUVEC prior to infection with the laboratory-adapted HCMV Ad169 strain, which is impaired in its ability to translocate to the HUVEC nucleus (54), increases the proportion of the cell population allowing active HCMV infection in a manner that is blocked by the PKC inhibitor staurosporin or RO-31-8220 (55).…”

“…Phorbol ester treatment of HUVEC prior to infection with the laboratory-adapted HCMV Ad169 strain, which is impaired in its ability to translocate to the HUVEC nucleus (54), increases the proportion of the cell population allowing active HCMV infection in a manner that is blocked by the PKC inhibitor staurosporin or RO-31-8220 (55). The treatment of RPE cells with phorbol ester before or shortly after infection enhances HCMV activity via a PKC-dependent mitogen-activated protein kinase pathway, and this enhancement does not involve NF-B activity (13). Notably, MCMV replication in acutely infected murine fibroblasts is blocked by PKC inhibitors, which manifests as a decrease in the level of MIE gene expression (28).…”

“…Viral particle components themselves are sufficient to stimulate the signaling cascades that involve NF-B activation (60). In contrast, HCMV Ad169 infection of retinal pigmented epithelial cells does not increase NF-B DNA binding or entry into the cell nucleus (13). However, the treatment of these cells with the phorbol ester 12,0-tetradecanoylphorbol 13-acetate (TPA) greatly increases nuclear NF-B binding activity, but surprisingly, the enhanced NF-B binding activity does not account for the TPA-induced activation of HCMV Ad169 MIE gene expression in these cells (13).…”

Neutrality of the Canonical NF-κB-Dependent Pathway for Human and Murine Cytomegalovirus Transcription and Replication In Vitro

“…We further analyzed the contribution of the canonical, or classical, NF-B pathway by using dominant negative inhibitors of IB␣ and immortalized p65/RelA-deficient fibroblasts. Infection of fibroblasts by CMV has been shown to activate the translocation of transcriptionally active p50/p65 to the nucleus (10,19,27) and to induce activation of the p50 and p65 promoters by IE1 (19,54,55). Overexpression of HCMV IE1 has also recently been reported to induce transcription of the NF-B family member RelB (23).…”

“…Numerous studies have shown that human CMV infection induces activation of NF-B (9,10,45,54), and the NF-B-responsive elements present in the MIEP (Fig. 1A) bind NF-B and are responsive to NF-B-mediated transcriptional enhancement in transient-transfection reporter-based expression assays (45).…”