Glutamine is required for red-spotted grouper nervous necrosis virus replication via replenishing the tricarboxylic acid cycle

Glutamine, one of the most important nutrients, plays a vital role in carbon metabolic pathway and has been reported to be required for the replication of several human DNA viruses. However, whether glutamine is required for RNA virus replication and the related mechanism remains elusive. Nervous necrosis virus (NNV), a positive-stranded RNA virus, can infect a number of important aquatic species and has caused great economic losses in aquaculture industry worldwide. In this study, the effects of glutamine on red-spotted grouper nervous necrosis virus (RGNNV) replication were investigated. The results showed that lack of glutamine did not affect the cell viability, but dramatically inhibited RGNNV replication, indicating that glutamine was required for RGNNV replication. Glutamine can be converted to α-ketoglutarate (α-KG) by glutaminase (GLS) to join in the tricarboxylic acid (TCA) cycle. Inhibiting the activity of GLS by a GLS inhibitor: bis-2-5-phenylacetamido-1,3,4-thiadiazol-2-ethyl sulfide (BPTES) significantly inhibited RGNNV replication, while adding the TCA cycle intermediates: α-KG, oxaloacetic acid (OAA), or pyruvate significantly restored RGNNV replication in glutamine-free medium, indicating that the requirement of glutamine for RGNNV replication was due to replenishing the TCA cycle. Taken together, these data revealed that glutamine could regulate RGNNV replication via TCA cycle, which will pave a new way for the prevention of the RGNNV infection in the future.