Pyrimidine synthesis enzyme CTP synthetase 1 suppresses antiviral interferon induction by deamidating IRF3

Metabolism is typically contextualized in conjunction with proliferation and growth. The roles of metabolic enzymes beyond metabolism—such as in innate immune responses—are underexplored. Using a focused short hairpin RNA (shRNA)-mediated screen, we identified CTP synthetase 1 (CTPS1), a rate-limiting enzyme of pyrimidine synthesis, as a negative regulator of interferon induction. Mechanistically, CTPS1 interacts with and deamidates interferon regulatory factor 3 (IRF3). Deamidation at N85 impairs IRF3 binding to promoters containing IRF3-responsive elements, thus muting interferon (IFN) induction. Employing CTPS1 conditional deletion and IRF3 deamidated or deamidation-resistant knockin mice, we demonstrated that CTPS1-driven IRF3 deamidation restricts IFN induction in response to viral infection in vivo. However, during immune activation, IRF3 deamidation by CTPS1 is inhibited by glycogen synthase kinase 3 beta (GSK3β) to promote IFN induction. This work demonstrates how CTPS1 tames innate immunity independent of its role in pyrimidine synthesis, thus expanding the functional repertoire of metabolic enzymes into immune regulation.