Aspartate deficiency amplifies cGAS-STING signaling in antitumor immunity
Yuheng Liao, Hanze Wang, Hengxin Liu, Xi Chen, Renqiang Sun, Xie Li, Zhen Yang, Chenying Liu, Wei Wu, Ziqian He, Yuzheng Zhao, Ying Mao, Dan Ye, Hui Yang
Yuheng Liao, Hanze Wang, Hengxin Liu, Xi Chen, Renqiang Sun, Xie Li, Zhen Yang, Chenying Liu, Wei Wu, Ziqian He, Yuzheng Zhao, Ying Mao, Dan Ye, Hui Yang
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Research Article Metabolism Oncology

Aspartate deficiency amplifies cGAS-STING signaling in antitumor immunity

Abstract

Metabolic signals critically shape innate immune responses. Through pharmacological screening of metabolic pathways, we identified aspartate metabolism as a key regulator of cyclic GMP-AMP synthase (cGAS)–stimulator of interferon genes (STING) signaling. Genetically or aminooxyacetic acid–mediated (AOA-mediated) pharmacologically reducing aspartate levels markedly potentiated the cGAS-STING pathway, leading to stronger upregulation of type I interferons and interferon-stimulated genes. Mechanistically, disruption of de novo pyrimidine synthesis, a major downstream pathway of aspartate, induced mtDNA replication stress and increased mtDNA double-strand breaks, promoting mtDNA release into the cytosol. Cytosolic mtDNA synergized with cGAS-STING agonists to upregulate Z-DNA binding protein 1 (ZBP1), which recruits RIPK1/3 to sustain IRF3 phosphorylation, forming a positive feedback loop that amplifies innate immune signaling. In immunocompetent mouse models, AOA enhanced the antitumor efficacy of STING agonists, chemotherapy, or radiotherapy, whereas aspartate supplementation abrogated these effects. Consistently, aspartate levels negatively correlated with antitumor immunity in colorectal cancer patient samples. Together, our study identifies aspartate–pyrimidine metabolism as a critical metabolic checkpoint that licenses STING signaling by enabling mtDNA stress to cooperate with agonist stimulation, driving type I interferon–dependent ZBP1 induction and feed-forward amplification of STING signaling, thus offering a promising strategy to enhance antitumor immunity.

Authors

Yuheng Liao, Hanze Wang, Hengxin Liu, Xi Chen, Renqiang Sun, Xie Li, Zhen Yang, Chenying Liu, Wei Wu, Ziqian He, Yuzheng Zhao, Ying Mao, Dan Ye, Hui Yang

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Figure 1

The aminotransferase inhibitor AOA amplifies cGAS-STING signaling.

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The aminotransferase inhibitor AOA amplifies cGAS-STING signaling. (A) O...
(A) Outline of metabolic inhibitors for screening. (B) Human hTERT-immortalized foreskin fibroblast (BJ-5ta, left) and mouse fibroblast (L929, right) cells were treated with inhibitors from A followed by 0.5 μg/mL HT-DNA transfection for 5 hours. Cells were harvested for qPCR analysis of human IFNB or mouse Cxcl10. (C) BJ-5ta cells were treated with 1 mM AOA for 1 hour followed by HT-DNA (0.5 μg/mL) transfection for the indicated time. Cells were harvested for qPCR analysis of IFN response gene expression. (D and E) L929 (D) and MC38 (E) were treated with 0.5 mM AOA for 1 hour followed by DMXAA (50 μM) or HT-DNA (0.5 μg/mL) stimulation for the indicated time. Cells were harvested for qPCR analysis of IFN response gene expression. (F–H) Western blot detected phosphorylated (p-) p-IRF3 and p-STAT1 levels in BJ-5ta, L929, and MC38 cells treated as in C–E. (I) Representative immunofluorescence images of IRF3 in BJ-5ta cells treated with AOA 1 hour followed by HT-DNA (0.5 μg/mL) transfection or not. Scale bars, 5 μm. (J) Schematic of the main targets of AOA. (K) Western blot detected p-IRF3, p-STAT1, GOT2, and GPT2 levels in BJ-5ta cells treated as indicated. (L) The relative IFNB and CXCL10 mRNA expression in the control BJ-5ta cells versus GOT2 and GPT2 siRNA-silenced BJ-5ta cells after indicated treatment. Data are represented as means ± SEM. Representative data are shown from 2 or 3 independent experiments. Statistical analysis was performed by unpaired t test (C–E, I, and L). *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.