Citrulline depletion by ASS1 is required for proinflammatory macrophage activation and immune responses

Citrulline can be converted into argininosuccinate by argininosuccinate synthetase (ASS1) in the urea cycle and the citrulline-nitric oxide cycle. However, the regulation and biological function of citrulline metabolism remain obscure in the immune system. Unexpectedly, we found that macrophage citrulline declines rapidly after interferon gamma (IFN-γ) and/or lipopolysaccharide (LPS) stimulation, which is required for efficient proinflammatory signaling activation. Mechanistically, IFN-γ and/or LPS stimulation promotes signal transducers and activators of transcription 1 (STAT1)-mediated ASS1 transcription and Janus kinase2 (JAK2)-mediated phosphorylation of ASS1 at tyrosine 87, thereby leading to citrulline depletion. Reciprocally, increased citrulline directly binds to JAK2 and inhibits JAK2-STAT1 signaling. Blockage of ASS1-mediated citrulline depletion suppresses the host defense against bacterial infection in vivo . We therefore define a central role for ASS1 in controlling inflammatory macrophage activation and antibacterial defense through depletion of cellular citrulline and, further, identify citrulline as an innate immune-signaling metabolite that engages a metabolic checkpoint for proinflammatory responses. • Proinflammatory macrophage activation reprograms citrulline metabolism • Transcriptional and post-translational regulation of ASS1 by JAK2-STAT1 signaling • JAK2 signals citrulline depletion to proinflammatory macrophage activation • Citrulline depletion by ASS1 is vital for macrophage innate immune response Mao et al. showed that phosphorylation and transcriptional upregulation of ASS1 mediated by JAK2 and STAT1, respectively, to deplete citrulline is essential for proinflammatory macrophage activation. High levels of intracellular citrulline resulting from ASS1 inhibition can bind and inhibit JAK2, leading to a blocked innate immune response in macrophages.