Essential role of PKCδ in histone deacetylase inhibitor-induced Epstein–Barr virus reactivation in nasopharyngeal carcinoma cells

Abstract

Histone deactylase inhibitors (HDACi) are common chemotherapeutic agents that stimulate Epstein-Barr virus (EBV) reactivation; the detailed mechanism remains obscure. In this study, it is demonstrated that PKCd is required for induction of the EBV lytic cycle by HDACi. Inhibition of PKCd abrogates HDACi-mediated transcriptional activation of the Zta promoter and downstream lytic gene expression. Nuclear translocation of PKCd is observed following HDACi stimulation and its overexpression leads to progression of the EBV lytic cycle. Our study suggests that PKCd is a crucial mediator of EBV reactivation and provides a novel insight to study the regulation of the EBV lytic cycle.Epstein-Barr virus (EBV) is a human gammaherpesvirus with both latent and lytic states in its life cycle (Kieff & Rickinson, 2001). EBV reactivation not only produces infectious viral progeny, but also contributes to the development of EBV-related disease (Kieff & Rickinson, 2001). Although the majority of EBV infections in vivo are latent, serological studies suggest that EBV reactivation may occur months or years before the clinical diagnosis of nasopharyngeal carcinoma (NPC), Hodgkin's disease and endemic Burkitt's lymphoma, serving as a risk factor for cancer development (Kieff & Rickinson, 2001). Previous studies have shown that some physiological stimuli and pharmacological agents, including transforming growth factor beta, cross-linking of surface immunoglobulin, phorbol ester, histone deacetylase inhibitors (HDACi) and several genotoxic agents, can induce EBV from latent status into the lytic cycle (Daibata et al., 1994;Davies et al., 1991;Feng & Kenney, 2006; Feng et al., 2002;Schuster et al., 1991).The protein kinase C (PKC) family has been known for a long time to be necessary for EBV reactivation following stimulation with 12-O-tetradecanoyl phorbol 13-acetate (TPA) (Davies et al., 1991) or anti-immunoglobulin treatment (Daibata et al., 1994). The PKC family comprises 11 isozymes, which are classified into three subfamilies according to their structure and regulatory domains activated by calcium or diacylglycerol (DAG) (Newton, 1997). Conventional PKCs (PKCa, bI, bII and c) require calcium and DAG activators, and novel PKCs (PKCd, e, h and g) respond only to DAG and not to calcium (Newton, 1997). However, atypical PKC isoforms (PKCf, i/l and m) are DAG-insensitive (Newton, 1997). Until now, it has not been clear which PKC members are involved in EBV reactivation. The limited information available suggests that PKCs may play an important role during different stages of various virus infections (Constantinescu et al., 1991;Sieczkarski et al., 2003). For example, PKCf or PKCd is required for early infection or TPA-triggered lytic-cycle activation of Kaposi's sarcoma-associated herpesvirus (KSHV) (Deutsch et al., 2004;Naranatt et al., 2003).HDACi, including trichostatin A (TSA), sodium butyrate (SB) and valproic acid, are common agents used to induce the EBV lytic cycle in several EBV-harbouring epithelial and B cells (Chang & Liu, 2...

Maintenance of Epstein-Barr Virus Latent Status by a Novel Mechanism, Latent Membrane Protein 1-Induced Interleukin-32, via the Protein Kinase Cδ Pathway

“…In this work, we uncovered a novel mechanism of a cellular factor which modulates the EBV life cycle. Our previous studies indicated that PKC␦ is an important modulator of EBV lytic cycle progression and that overexpression of PKC␦ significantly stimulates Zp activity (38,54). In this study, we clearly showed that IL-32 downregulates Zp activation by interacting with PKC␦ and inhibiting the nuclear translocation of PKC␦ to maintain viral latency.…”

“…It is well documented that PKC␦ activation may be detected by its translocation from the cytoplasm to the nucleus, phosphorylation at specific sites, and cleavage to a catalytic fragment (54)(55)(56)(57). Confocal microscopy was used to investigate whether IL-32 affected the localization of PKC␦.…”

Histone Hyperacetylation Up-regulates Protein Kinase Cδ in Dopaminergic Neurons to Induce Cell Death

“…Interestingly, these findings, combined with previous evidence, indicate a cell typespecific regulation of PKC␦ gene expression in response to HDAC inhibition. For instance, in nasopharyngeal carcinoma and AA/C1 adenoma cells, HDAC inhibitors, such as TSA and NaBu, did not alter the PKC␦ expression levels but rather led to a distinct cellular sub-localization of the PKC␦ protein (92,93). By contrast, in human hepatoma Hep3B cells, TSA was found to specifically down-regulate PKC␦ levels (94).…”

Interplay between PKCδ and Sp1 on Histone Deacetylase Inhibitor-Mediated Epstein-Barr Virus Reactivation

“…In our previous studies, we found that the PKC␦ specific inhibitor, Rottlerin, alone is sufficient to block TSAinduced EBV reactivation (31). Moreover, the activation of PKC␦ can be indicated by phosphorylation at threonine 505 residues, cleavage into catalytic fragments, and protein translocation to the nucleus, and all have been observed after TSA treatment (31).…”

“…Of note, we found that both sodium butyrate (SB) and trichostatin A (TSA), two histone deacetyltransferase inhibitors, are efficient inducers of EBV reactivation in an epithelial cell model. Furthermore, PKC␦ was shown to be responsible for this specific induction (31).…”