Direct Evidence of Abortive Lytic Infection-Mediated Establishment of Epstein-Barr Virus Latency During B-Cell Infection

Abstract

Viral infection induces dynamic changes in transcriptional profiles. Virus-induced and antiviral responses are intertwined during the infection. Epstein-Barr virus (EBV) is a human gammaherpesvirus that provides a model of herpesvirus latency. To measure the transcriptome changes during the establishment of EBV latency, we infected EBV-negative Akata cells with EBV-EGFP and performed transcriptome sequencing (RNA-seq) at 0, 2, 4, 7, 10, and 14 days after infection. We found transient downregulation of mitotic division-related genes, reflecting reprogramming of cell growth by EBV, and a burst of viral lytic gene expression in the early phase of infection. Experimental and mathematical investigations demonstrate that infectious virions were not produced in the pre-latent phase, suggesting the presence of an abortive lytic infection. Fate mapping using recombinant EBV provided direct evidence that the abortive lytic infection in the pre-latent phase converges to latent infection during EBV infection of B-cells, shedding light on novel roles of viral lytic gene(s) in establishing latency. Furthermore, we find that the BZLF1 protein, which is a key regulator of reactivation, was dispensable for abortive lytic infection in the pre-latent phase, suggesting the divergent regulation of viral gene expressions from a productive lytic infection.

Reversal of splicing infidelity is a pre-activation step in B cell differentiation

“…Our analysis of the fully EBV-immortalized LCL GM12878 indicated that unannotated splicing in this cell line was comparable to that of activated, differentiated B cells in vivo (Figure 1). To investigate the stage at which the unique splicing pattern of blood NBCs is reversed, we obtained RNA-Seq datasets from timecourse experiments of EBV infection leading to LCL establishment (17)(18)(19)(20). Consistent with previously observed similarities between in vitro EBV-associated activation and in vivo antigen-mediated activation (16,17), fully EBV-immortalized LCLs from two different laboratories showed strong enrichment in the expression of genes associated with PCs relative to their parental blood B cells (Figure 2A).…”

Replication Compartments—The Great Survival Strategy for Epstein–Barr Virus Lytic Replication

“…DDR is also temporarily induced and attenuated until latency is established [ 106 ]. However, unlike lytic replication, cellular DNA synthesis and cell division occur several days after primary infection, progeny viruses are not produced, and BZLF1 is not necessary for infection during the pre-latent phase [ 107 , 108 ]. Thus, because cellular aspects during the pre-latent phase differ from those of the lytic phase, little is known about how viral DNAs are synthesized, including whether RCs are formed, although viral DNA is abundantly amplified during the pre-latent phase.…”

“…Moreover, recently it has been suggested that cells during the “abortive lytic phase” exist [ 109 ]. The “abortive lytic phase” is, so to speak, an “incomplete lytic cycle”, wherein several early lytic genes are expressed without progeny viral production like the pre-latent phase [ 108 , 110 ]. Studying these “incomplete lytic phases”, including whether RCs are assembled or not, might unveil the novel significance of RC formation.…”

Roles of Lytic Viral Replication and Co-Infections in the Oncogenesis and Immune Control of the Epstein–Barr Virus

“…Although it has been commonly acknowledged that EBV + tumors are largely composed of latently infected cells, accumulating evidence has shown a remarkable role of EBV lytic gene products in EBV-driven oncogenesis [ 1 , 89 , 90 ]. While a number of EBV lytic genes have been found to be expressed in a “leaky” fashion during the early stage of EBV infection, a very limited number of infectious virions were detected shortly after EBV entry into B cells, with reactivation usually not occurring for two weeks thereafter [ 91 , 92 , 93 ]. An abortive (incomplete) lytic replication cycle in the pre-latent stage is suggested as a first phase during the development of the EBV infection within the host [ 1 , 91 , 94 , 95 ].…”

“…While a number of EBV lytic genes have been found to be expressed in a “leaky” fashion during the early stage of EBV infection, a very limited number of infectious virions were detected shortly after EBV entry into B cells, with reactivation usually not occurring for two weeks thereafter [ 91 , 92 , 93 ]. An abortive (incomplete) lytic replication cycle in the pre-latent stage is suggested as a first phase during the development of the EBV infection within the host [ 1 , 91 , 94 , 95 ]. This implies that the transient lytic gene expression may play a supportive role during latency development, tumor formation, and LCL generation.…”