Glutamine supports human cytomegalovirus induced lipidome remodeling through reductive carboxylation

Abstract Lipidome remodeling during human cytomegalovirus (HCMV) replication is a complex process that requires induction of lipogenic proteins and altered metabolite flow to support synthesis of fatty acids and lipids. HCMV infection increases the utilization of glucose and acetate to provide enough carbons to support increased demand for lipogenesis during virus replication, but other carbon contributors have not been studied. Here, we identify glutamine as a carbon source for lipogenesis during HCMV infection. Metabolic tracing with 13 C-labeled glutamine revealed carbons from glutamine are enriched in phospholipids and neutral lipids during infection, including phosphatidylcholine, phosphatidylethanolamine, diacylglycerol, and triacylglycerol. Additional metabolic tracing demonstrates that HCMV infection promotes glutamine flow to fatty acid synthesis primarily through reductive carboxylation, i.e., conversion of glutamine to citrate through isocitrate. Through the use of two different 13 C-labeled forms of glutamine, we found that ∼30% of the carbons from glutamine are delivered to fatty acid synthesis through additional metabolic means. Our current understanding of metabolite utilization during HCMV replication is based on cell culture models where there is an excess amount of glucose, suggesting that deriving carbons from glutamine might be needed when glucose levels are low. To determine if concentrations of glucose and glutamine change their contributions to fatty acid synthesis, we investigated lipogenesis when glucose and glutamine are at physiological levels (5 mM and 0.55 mM, respectively). We determined that physiological levels of glucose and glutamine are sufficient to support the increased demand for fatty acid synthesis caused by HCMV infection, despite a reduction in virus production. Using metabolic tracing with 13 C-labeled forms of glucose or glutamine, we determined that both carbon sources still contribute to fatty acid synthesis when present at physiological levels. Overall, our results identify viral activation of reductive carboxylation that increases glutamine flow to lipogenesis during infection. This work provides additional insight into metabolic reprogramming that supports HCMV-induced lipidome remodeling. Author Summary Many viruses hijack cellular metabolic processing to obtain the components needed for replication. Human cytomegalovirus (HCMV) uses several mechanisms to reprogram lipid metabolism and remodel the lipidome of infected cells. HCMV promotes synthesis of very long chain fatty acids that are found in phospholipids and triacylglycerol. Glucose and acetate contribute carbon to fatty acid synthesis and elongation following HCMV infection. In this work, we demonstrate that glutamine is an additional carbon source for fatty acid and lipid synthesis. Phospholipids and neutral lipids are enriched with carbons from glutamine during HCMV infection. Mechanistically, HCMV induces reductive carboxylation to increase glutamine flow to fatty acid synthesis and increased metabolite availability supports additional carbon flow to fatty acids. Overall, this study provides additional insight into virus-induced metabolic remodeling that supplies the molecular building blocks for virus replication.