GAPDH was used as an interior control. that may mark the activation of glutaminolysis. Cell proliferation and viability of latently EBV-infected cells were notably inhibited by KGA/GAC, as well as GLUD1 inhibitors. Taken together, our results suggest that c-Myc-dependent regulation of KGA and GAC enhances mitochondrial functions to support the rapid proliferation of the EBV-infected cells, and these metabolic processes could be therapeutically exploited by targeting KGA/GAC and GLUD1 to prevent EBV-associated cancers. Keywords: EBV, KGA, GAC, glutaminolysis, mitochondrial metabolism, cell proliferation 1. Introduction EpsteinCBarr computer virus (EBV) or human herpesvirus 4 (HHV-4) is an oncogenic computer virus that infects and establishes latency in more than 90% of the worlds human population. EBV is usually associated with a variety of B-cell lymphomas, such as Burkitts lymphoma, Rabbit Polyclonal to NSF Hodgkins lymphoma, and post-transplantation lymphoproliferative disorders, and two epithelial cancers, nasopharyngeal carcinoma and gastric carcinoma [1,2,3]. Depending on the expression of EBV-latent proteins, distinct latency patterns are associated with specific EBV-associated cancers. Burkitts lymphoma is usually associated with the expression of type I latency genes, EBV-nuclear antigen 1 (EBNA1) and EBV-encoded RNAs (EBERs). Hodgkins lymphoma and nasopharyngeal carcinoma are characterized by the expression of type II latency genes, EBNA1, latent membrane protein 1 (LMP1), LMP2A, LMP2B, and EBERs. In post-transplantation lymphoproliferative diseases and in vitro immortalized lymphoblastoid cell lines (LCLs), EBV has the most composite expression profile, latency III, which involves the expression of five EBNAs (EBNA1, 2, 3A, 3B, 3C), two LMPs (LMP1, LMP2A), and two EBERs [4,5,6]. Although the pattern of latent gene expression varies among EBV-associated cancers, they share several cellular metabolic adaptations that influence the uptake of nutrients to support rapid cell proliferation, cell growth, Tenacissoside H and survival [7]. c-Myc is an important cellular oncogene which co-ordinates viral gene expression and metabolic reprograming in EBV-associated cancers [8]. Notably, in Burkitts lymphoma, chromosomal translocation of c-Myc to the IgG locus leads to overexpression of c-Myc, resulting in increased cell proliferation and malignant transformation [9]. c-Myc upregulation has also been reported in about 90% of EBV-associated nasopharyngeal carcinomas [10,11,12]. Additionally, the activation of c-Myc transcription by EBNA2 was reported to play an instrumental role in EBV-infected LCL proliferation and survival [13]. c-Myc is usually a transcription factor which regulates the expression of many genes associated with Tenacissoside H the cellular metabolic processes to meet the high metabolic and bioenergetics demands of cancer cells [14]. Aerobic glycolysis and glutaminolysis are the major metabolic pathways used by cancer cells to fuel their bioenergetic and biosynthetic needs [15]. As the energy derived from aerobic glycolysis is not sufficient to meet the energy requirements of actively dividing cancer cells, they also depend on increased glutamine uptake and glutaminolysis to sustain a functional TCA cycle for the production of energy, reductive equivalents, and the biosynthesis of various macromolecules supporting tumor growth and proliferation [16]. Glutaminolysis involves the deamination of glutamine to glutamate catalyzed by GLS. Glutamate is usually subsequently converted to a TCA cycle intermediate, alpha-ketoglutarate, Tenacissoside H mediated by glutamate dehydrogenases or aminotransferases that replenish the TCA cycle. The mitochondrial enzyme GLS1 plays a crucial role in maintaining metabolism and homeostasis. In mammalian cells, GLS1 encodes two isoforms: kidney (K-type) glutaminase (KGA) and glutaminase C (GAC) [17]. Despite distinct tissue distribution patterns, the functionality of both isoforms remains the same. GAC is found to be the more catalytically active and predominant isoform, with implications in tumor metabolism [18]. However, the involvement of KGA and GAC in oncogenic cellular energy metabolism and cell proliferation, as well as its connection with the aberrantly expressed c-Myc, is not completely comprehended in EBV-associated cancers. We hypothesized that this expression of KGA and GAC may be an adaptation of rapidly proliferating EBV-infected cells to upgrade the efficiency of glutaminolysis for the sustenance of the increased energy demands of tumor metabolism. Thus, understanding the interplay of KGA and GAC isoforms in the oncogenic cellular energy metabolism of EBV-infected cancers could unravel the bioenergetics in EBVs pathobiology. In the present study, we demonstrate that EBV contamination upregulates the expression of GLS1 isoforms KGA and GAC, which in turn are regulated by c-Myc. Both KGA and GAC localize to the mitochondria, where they are involved in the conversion of glutamine.