Hypoxia-inducible factor-1α plays roles in Epstein-Barr virus’s natural life cycle and tumorigenesis by inducing lytic infection through direct binding to the immediate-early BZLF1 gene promoter

Abstract

When confronted with poor oxygenation, cells adapt by activating survival signaling pathways, including the oxygen-sensitive transcriptional regulators called hypoxia-inducible factor alphas (HIF-αs). We report here that HIF-1α also regulates the life cycle of Epstein-Barr virus (EBV). Incubation of EBV-positive gastric carcinoma AGS-Akata and SNU-719 and Burkitt lymphoma Sal and KemIII cell lines with a prolyl hydroxylase inhibitor, L-mimosine or deferoxamine, or the NEDDylation inhibitor MLN4924 promoted rapid and sustained accumulation of both HIF-1α and lytic EBV antigens. ShRNA knockdown of HIF-1α significantly reduced deferoxamine-mediated lytic reactivation. HIF-1α directly bound the promoter of the EBV primary latent-lytic switch BZLF1 gene, Zp, activating transcription via a consensus hypoxia-response element (HRE) located at nt -83 through -76 relative to the transcription initiation site. HIF-1α did not activate transcription from the other EBV immediate-early gene, BRLF1. Importantly, expression of HIF-1α induced EBV lytic-gene expression in cells harboring wild-type EBV, but not in cells infected with variants containing base-pair substitution mutations within this HRE. Human oral keratinocyte (NOK) and gingival epithelial (hGET) cells induced to differentiate by incubation with either methyl cellulose or growth in organotypic culture accumulated both HIF-1α and Blimp-1α, another cellular factor implicated in lytic reactivation. HIF-1α activity also accumulated along with Blimp-1α during B-cell differentiation into plasma cells. Furthermore, most BZLF1-expressing cells observed in lymphomas induced by EBV in NSG mice with a humanized immune system were located distal to blood vessels in hypoxic regions of the tumors. Thus, we conclude that HIF-1α plays central roles in both EBV’s natural life cycle and EBV-associated tumorigenesis. We propose that drugs that induce HIF-1α protein accumulation are good candidates for development of a lytic-induction therapy for treating some EBV-associated malignancies.

Human Cytomegalovirus Productively Replicates In Vitro in Undifferentiated Oral Epithelial Cells

“…NOKs are human telomerase reverse transcriptase (hTERT)-expressing cells from human gingival tissue (45), generously provided by Karl Münger. hGETs are hTERT-expressing normal human gingival epithelial progenitor cells (46). NOKs and hGETs were cultured as monolayers in keratinocyte serum-free medium (KSFM; Life Technologies) supplemented with 0.14 g/ml epidermal growth factor and 0.33 mg/ml bovine pituitary extract at 5% CO 2 and 37°C.…”

Herpesviral Latency—Common Themes

“…Low oxygen conditions can also facilitate EBV reactivation [196]. Hypoxia inducible factor 1α (HIF1α) can directly induce EBV reactivation by activating the expression of BZLF1, a lytic switch protein [197]. In fact, reactivation of KSHV by hypoxia and proinflammatory cytokines involves oxidative stress and reactive oxygen species (ROS).…”

Epstein–Barr Virus Infection of Oral Squamous Cells

“…We have confirmed up-regulation of KLF4 in EBV infected HSC1 cells ( Figure S3). In addition to KLF4, activation of other transcription factors that induce Zta was reported in EBV-infected epithelial cells [30][31][32]. As shown in Figure 3, the status of EBV infection in HSC1 cells and SCC25 cells was productive and abortive, respectively.…”