Signal Transduction and Transcription Factor Modification during Reactivation of Epstein-Barr Virus from Latency

Abstract

Reactivation of Epstein-Barr virus (EBV) from latency involves activation of theLatency and reactivation are defining features of herpesviruses, and several common human diseases are caused by their reactivation. Epstein-Barr virus (EBV) is particularly favorable for the study of reactivation because efficient reactivation can be induced in some Burkitt's lymphoma cell lines in response to physiologically relevant stimuli such as B-cell receptor (BCR) stimulation (38, 39) and cytokine treatment (3, 12). The key step in reactivation of the lytic cycle is induction of the BZLF1 transcription factor through its promoter, Zp (4, 9). The mechanism of induction of Zp in response to cross-linking the BCR with anti-immunoglobulin (anti-Ig) is the subject of this study.Initial studies of signaling to Zp were carried out with various signal transduction inhibitors to inhibit reactivation. Genistein (a tyrosine kinase inhibitor) inhibited reactivation induced by BCR cross-linking (10), and a protein kinase C antagonist, staurosporine, completely blocked the appearance of BZLF1 after anti-Ig treatment (27). Treatment with the calcium ionophore A23187 increased Zp activity. When the calcium ionophore was used in conjunction with tetradecanoyl phorbol acetate, a protein kinase C activator, Zp induction was synergistically enhanced. 1-(5-Isoquinolinylsulfonyl)-2-methylpiperazine, an inhibitor of protein kinase C, inhibited the anti-Ig inducibility of Zp. Two calmodulin antagonists, compound R24571 and trifluoperazine, blocked Zp activation with anti-Ig. These findings suggested that Zp responds directly to changes in the activity of both protein kinase C and calcium-or calmodulin-dependent protein kinase.A requirement for tyrosine kinase activation for anti-Igmediated Zp activation was also demonstrated through the use of the tyrosine kinase inhibitor herbimycin (11). Anti-IgG induced rapid phosphorylation of mitogen-activated protein kinase in Akata cells. The phosphorylation was inhibited by the mitogen-activated protein kinase/ERK kinase-specific inhibitor PD98059. Expression of the EBV immediate-early BZLF1 mRNA and its protein product, ZEBRA, and early antigen was also prevented by the inhibitor. These results indicated that mitogen-activated protein kinase may be involved in the pathways of EBV activation (34). In EBV-immortalized B-cell lines, the LMP1 and LMP2A proteins prevent reactivation of the lytic cycle (1, 28). The LMP1 mechanism is not yet known, but LMP2A protein prevents reactivation by interfering with BCR signaling. In EBVinfected cells in which LMP2A is expressed, cross-linking of secretory immunoglobulin fails to activate Lyn, Syk, phosphatidylinositol 3-kinase, phospholipase C gamma 2, Vav, Shc, and mitogen-activated protein kinase. In contrast, cross-linking of secretory immunoglobulin on cells infected with EBV recombinants with null mutations in LMP2A resulted in transient tyrosine phosphorylation of Lyn, Syk, phospholipase C gamma 2, and phosphatidylinositol 3-kinase, a transient increase in intracel...

Valproic Acid Enhances the Efficacy of Chemotherapy in EBV-Positive Tumors by Increasing Lytic Viral Gene Expression

“…Consistent with this, our results indicate that VPA significantly enhanced the luciferase activity derived from the Zp-luciferase construct even in the absence of a chromatinized template. Interestingly, one of the most important regulators of Zp transcription is the MEF-2D transcription factor, which is constitutively bound to the BZLF1 promoter but can serve either as a positive or negative regulator of BZLF1 transcription (47). When MEF-2D is complexed with HDAC proteins, it acts as negative regulator of transcription, and thus the ability of VPA to inhibit HDAC activity would be expected to convert MEF-2D bound to the BZLF1 promoter from a negative to a positive regulator of Zp transcription.…”

Virally targeted therapies for EBV-associated malignancies

“…The ability of chemotherapy to induce lytic EBV reactivation appears to require the simultaneous engagement of several different signal transduction pathways, including cellular stress MAP kinase p38, PI3 kinase, PKC delta, and ERK 1/2, but not cellular apoptosis per se (Feng et al, 2002a, and unpublished data). Interestingly, activation of these same cellular signal transduction pathways is also required for lytic EBV infection following engagement of the B-cell receptor (Adamson et al, 2000;Darr et al, 2001;Bryant and Farrell, 2002), suggesting that similar transcription factors may be involved. In addition, BRLF1-induced activation of the BZLF1 promoter requires the stress MAP kinases, p38 and c-jun Nterminal kinase, as well as PI3 kinase (Adamson et al, 2000;Darr et al, 2001).…”

“…In the case of the BZLF1 promoter (Zp), full activation Toxin expression specific to EBV-containing cells Figure 2 Latent viral promoters that require EBV-encoded gene products in trans, or the oriP replication origin that requires EBNA-1 in trans, can be used to produce cellular toxins specifically in EBV-positive cells requires both a 'ZII' motif (which binds CREB, ATF1, and a c-jun/ATF-2 heterodimer) and several different 'ZI' binding sites, which bind MEF2D as well as Sp1 and Sp3 (Flemington and Speck, 1990;Daibata et al, 1994;Speck et al, 1997;Wang et al, 1997;Adamson et al, 2000;Binne et al, 2002;Bryant and Farrell, 2002;Gruffat et al, 2002). Calcium-induced activation of MEF2D (mediated by calcineurin) and CREB (mediated by CaMKIV/GR) appears to be important for BZLF1 promoter activation following ligation of the B-cell receptor Liu et al, 1997).…”

Impact of Human Cytomegalovirus Latent Infection on Myeloid Progenitor Cell Gene Expression

“…HDACs have been implicated in the regulation of latency of other herpesviruses, including Epstein Barr virus and human herpesvirus 8 (7,16). In addition, a recent study by Murphy et al (37) suggested that HDACs mediate repression of the CMV MIE promoter (MIEP) in nonpermissive cells because inhibition of HDACs induced viral permissiveness and increased MIEP activity.…”