Inactivated Wild-Type Epstein-Barr Virus (B95-8)

More than 90% of adults worldwide are infected with Epstein-Barr virus (EBV). EBV belongs to the gammaherpesvirus family and is transmitted via saliva, and new infections can lead to infectious mononucleosis if symptoms develop. Here, researchers performed objective metabolomics- and transcriptomics-based analyses and found that the activity of the kynurenine pathway and the de novo nicotinamide adenine dinucleotide (NAD) synthesis pathway are upregulated in B cells during latency following EBV infection. Researchers found that a viral protein, the EBV-encoded transactivator EBNA2, cooperates with the host B cell transcription factor EBF1 to induce the expression of indoleamine 2,3-dioxygenase 1 (IDO1), the first and rate-limiting enzyme of the kynurenine pathway. IDO1-dependent tryptophan degradation promotes de novo NAD synthesis, thereby supporting mitochondrial adenosine triphosphate generation during early stages of EBV infection. Pharmacological inhibition of IDO1 reduced the sensitivity of B cells to EBV transformation by 100-fold, while this barrier could be eliminated by re-addition of the IDO1 product kynurenine as well as supplementation with nicotinic acid mononucleotide, the immediate precursor of NAD.

To monitor metabolic and transcriptional responses associated with early EBV infection, we performed metabolomic and transcriptomic analyses at 0, 1, and 4 days post-infection. Specifically, naive B cells were isolated from healthy donors and infected with the prototype EBV strain B95-8 by spinoculation. Heat-inactivated EBV (h.i. EBV) was used as a control to detect non-infection-associated activation of B cells by pathogen-associated molecular patterns (PAMPs) and was added at the same concentration as infectious B95-8 virus. B cells were then analyzed at 0, 1, and 4 days post-infection with EBV and at 0, 1, and 4 days post-infection with h.i. EBV (Figure 1A). Researchers hypothesized that metabolic adaptations of infected B cells within 4 days post-infection are critical for cell cycle entry and initiation of hyperproliferation. Analysis of single metabolite abundance revealed that quinolinate, a metabolite formed from tryptophan in the kynurenine pathway, was the most differential metabolite, with a 32-fold increase in quinolinate in B cells at day 4 after EBV infection compared with B cells exposed to high density EBV (Figure 1B). In the kynurenine pathway, tryptophan is sequentially catabolized to L-kynurenine and quinolinate, which in some cells can be used for de novo biosynthesis of nicotinamide adenine dinucleotide (NAD). At day 4 after EBV infection, tryptophan levels were reduced in B cells, indicating enhanced activity of the kynurenine pathway (Figure 1B). The reduction in the abundance of NAD+ (the oxidized form of NAD) supports a model of early activation of the kynurenine pathway in EBV-infected B cells to replenish NAD (Figure 1B).

Figure 1. Metabolic profiling of EBV-infected B cells reveals transient up-regulation of NAD de novo biosynthesis in newly infected B cells. (Müller-Durovic B, et al., 2024)