Metabolic Remodeling of Human B-Cells During Latent Epstein-Barr Virus (EBV) Infection
Wang, Liang Wei
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CitationWang, Liang Wei. 2019. Metabolic Remodeling of Human B-Cells During Latent Epstein-Barr Virus (EBV) Infection. Doctoral dissertation, Harvard University, Graduate School of Arts & Sciences.
AbstractEpstein Barr-virus (EBV) is a ubiquitous herpesvirus that causes infectious mononucleosis and is associated with a variety of human cancers. Ordinarily, EBV persists in healthy individuals by establishing latent infection in the B-cell compartment. Immunosuppression can result in unchecked outgrowth of latently infected cells, culminating in lymphoproliferative disorders and lymphomas. How EBV induces and supports the physiological shift from quiescence to long-term growth and proliferation as modeled by in vitro lymphoblastic transformation is unknown. Multiplexed proteomics on de novo infected primary B-cells was performed to uncover metabolic and signaling pathways that facilitate outgrowth and survival of B-cells latently infected with EBV. By combining chemical inhibitor studies and clustered regularly interspaced short palindromic repeats (CRISPR) genetic perturbation experiments, we discovered viral upregulation of mitochondrial one-carbon (1C) metabolism and de novo synthesis of serine, cholesterol and fatty acids, to be crucial for EBV-positive B-cell proliferation and viability. We demonstrated that mitochondrial 1C metabolism broke down serine to generate formate for nucleotide synthesis, glycine for glutathione production and cellular reducing power in the form of NADPH. We also showed that intramitochondrial NADPH was important for efficient LCL proliferation. De novo serine synthesis (DNSS) may augment 1C metabolism and/or buffer the infected cells from external fluctuations in serine availability. Both mitochondrial 1C metabolism and DNSS depended on EBV-encoded nuclear antigen 2 (EBNA2) and its target MYC for activation. EBV-infected cells also utilized cholesterol and fatty acid synthetic pathways to produce geranylgeranyl pyrophosphate (GGPP) and palmitate, respectively, to reorganize the signaling milieu of the transforming cell. Rab geranylgeranyltransferase was specifically required for EBV-positive B-cell outgrowth and proliferation. We also identified RAB13 as a potent driver of latently infected cell growth and survival, whose expression was driven by EBNA3C and likely functioned as a chaperone for viral latent membrane proteins (LMPs) to maintain the latter’s oncogenic signaling capacities. Taken altogether, our results provide an expansive investigation into the pathways that underlie the transformative change of a B lymphocyte from quiescence to activation and growth and identify vulnerabilities in EBV-driven lymphoblastic disorders that may represent new therapeutic targets.
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