Abstract
Radical S-adenosylmethionine (SAM) domain-containing protein 2 (RSAD2; viperin) is a key enzyme in innate immune responses that is highly expressed in response to viral infection and inflammatory stimuli in many cell types. Recently, it was found that RSAD2 catalyses transformation of cytidine triphosphate (CTP) to its analogue 3'-deoxy-3',4'-didehydro-CTP (ddhCTP). The cellular function of this metabolite is unknown. Here, we analysed the extra- and intracellular metabolite levels in human induced pluripotent stem cell (hiPSC)-derived macrophages using high-resolution LC-MS/MS. The results together with biochemical assays and molecular docking simulations revealed that ddhCTP inhibits the NAD+ -dependent activity of enzymes including that of the housekeeping enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH). We propose that ddhCTP regulates cellular metabolism in response to inflammatory stimuli such as viral infection, pointing to a broader function of RSAD2 than previously thought.
Highlights
Radical S-adenosylmethionine domain-containing protein 2 (RSAD2), known as viperin, is highly conserved from fungi to human
Further analysis of data revealed that the cellular activity of RSAD2 abolished extracellular levels of products of phenylalanine and tyrosine metabolism, namely phenylacetic acid and 4-hydroxyphenyllactic acid (Figure 2)
The results revealed that the mixture of ddhCTP and 5 ́-deoxyadenosine (5 ́-dA) reduces NAD+-dependent activity of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) by approximately 3-fold (Figure 5A)
Summary
Radical S-adenosylmethionine domain-containing protein 2 (RSAD2), known as viperin, is highly conserved from fungi to human. Based on cell biological and biochemical assays, it is suggested that ddhCTP is a chain terminator of the RNA-dependent RNA polymerases (RdRps) of a number of flaviviruses [21] This conclusion, was challenged after a reevaluation of the data [20]. It is demonstrated that the reported data are consistent with the ddhCTP product of the radical-SAM activity of RSAD2 abolishing the cellular level of nucleotides and inhibiting mitochondrial activity [20]. These data suggest that ddhCTP modulates metabolism to restrict viral replication. It is not known how ddhCTP affects metabolism (Figure 1)
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