Epstein-Barr virus inactivates the transcriptome and disrupts the chromatin architecture of its host cell in the first phase of lytic reactivation

Abstract

Epstein-Barr virus (EBV), a herpes virus also termed HHV 4 and the first identified human tumor virus, establishes a stable, long-term latent infection in human B cells, its preferred host. Upon induction of EBV’s lytic phase, the latently infected cells turn into a virus factory, a process that is governed by EBV. In the lytic, productive phase, all herpes viruses ensure the efficient induction of all lytic viral genes to produce progeny, but certain of these genes also repress the ensuing antiviral responses of the virally infected host cells, regulate their apoptotic death or control the cellular transcriptome. We now find that EBV causes previously unknown massive and global alterations in the chromatin of its host cell upon induction of the viral lytic phase and prior to the onset of viral DNA replication. The viral initiator protein of the lytic cycle, BZLF1, binds to >105 binding sites with different sequence motifs in cellular chromatin in a concentration dependent manner implementing a binary molar switch probably to prevent noise-induced erroneous induction of EBV’s lytic phase. Concomitant with DNA binding of BZLF1, silent chromatin opens locally as shown by ATAC-seq experiments, while previously wide-open cellular chromatin becomes inaccessible on a global scale within hours. While viral transcripts increase drastically, the induction of the lytic phase results in a massive reduction of cellular transcripts and a loss of chromatin-chromatin interactions of cellular promoters with their distal regulatory elements as shown in Capture-C experiments. Our data document that EBV’s lytic cycle induces discrete early processes that disrupt the architecture of host cellular chromatin and repress the cellular epigenome and transcriptome likely supporting the efficient de novo synthesis of this herpes virus.

Systematic analysis of Epstein-Barr virus genes and their individual contribution to virus production and composition

“…The mix was agitated on a mixing roller at 4 °C for 3 h. The cells were pelleted at 500 g at 4 °C for 10 min and washed with 1 ml ice-cold staining buffer (1% FBS and 2 mM EDTA in PBS). The anti-gp350 antibody (6G4) (Buschle et al, 2021) coupled to Alexa647 in 50 μl staining buffer (1:250 dilution) was added and the Elijah cells were incubated at 4 °C for 20 min. 1 ml staining buffer was added and the cells were washed and resuspended in 300 μl staining buffer and analyzed by flow cytometry (BD FACSCanto).…”

“…BZLF1 is a homodimeric bZIP transcription and pioneer factor (Schaeffner et al, 2019) and binds two classes of BZLF1 (or ZEBRA) response elements (ZREs): CpG-free motifs resembling the consensus AP-1 site recognized by cellular bZIP proteins such as c-Jun/c-Fos (Farrell et al, 1989), and CpG-containing motifs that are selectively bound by BZLF1 upon cytosine methylation (Bhende et al, 2004; Dickerson et al, 2009; Bergbauer et al, 2010; Flower et al, 2011; Woellmer et al, 2012; Tillo et al, 2017; Buschle et al, 2021; Bernaudat et al, 2021). EBV’s virion DNA, which is unmethylated upon infection (Kalla et al, 2010), precludes the binding of BZLF1 to certain promoters of essential lytic viral genes in the pre-latent phase of infection.…”

“…EBV’s virion DNA, which is unmethylated upon infection (Kalla et al, 2010), precludes the binding of BZLF1 to certain promoters of essential lytic viral genes in the pre-latent phase of infection. The requirement of CpG methylated viral DNA to support EBV’s lytic phase also means that EBV cannot turn a newly infected cell into a viral factory (Kalla et al, 2010; Woellmer et al, 2012; Buschle et al, 2021). Instead, EBV establishes a strictly latent infection initially and escapes from it later when viral DNA has reached a certain level of CpG methylation (Kalla et al, 2010, 2012).…”

Interaction sites of the Epstein–Barr virus Zta transcription factor with the host genome in epithelial cells

“…In addition, Zta binds to and activates the expression of some cellular genes [35, 38–42]. Binding sites for Zta on the cellular cell genome in several B-cell lines have been identified [43, 44]. It was recently revealed that Zta binding to the human genome results in global changes to the architecture of cellular chromatin, opening some areas and making other areas inaccessible [44].…”

“…Binding sites for Zta on the cellular cell genome in several B-cell lines have been identified [43, 44]. It was recently revealed that Zta binding to the human genome results in global changes to the architecture of cellular chromatin, opening some areas and making other areas inaccessible [44]. The high number of Zta-binding sites has been postulated to contribute a buffer zone for Zta function; to prevent low levels of Zta initiating virus lytic cycle erroneously [44].…”