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 2

AOA augments cGAS-STING–mediated IFN response by inducing aspartate deficiency.

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AOA augments cGAS-STING–mediated IFN response by inducing aspartate defi...
(A) Heatmap of metabolite changes in mock- and AOA-treated L929 cells followed with HT-DNA transfection or without HT-DNA transfection. The fold-change of metabolites abundances was normalized to the mock group. Each square represents the mean of 3 replicates (n = 3 independent cultures). (B) L929 cells were treated with 0.5 mM AOA for 1 hour followed by DMXAA (50 μM) stimulation for 6 hours in the absence or presence of 20 mM aspartate or asparagine, and then cells were harvested for qPCR analysis of IFN response gene expression. (C) Heatmap from RNA-seq revealing the expression of ISGs in L929 cells with indicated treatment compared with mock group [z-score–normalized log2(fold per million reads) values, n = 3 independent cultures]. (D) GSEA of interferon-beta response gene. (E) Western blot detected p-IRF3 and p-STAT1 levels in L929 cells treated as indicated. (F) Representative immunofluorescence images of IRF3 in BJ-5ta cells treated with AOA 1 hour followed by HT-DNA (0.5 μg/mL) transfection in the absence or presence of 20 mM aspartate. Scale bars, 5 μm. Data are represented as means ± SEM. Representative data are shown from 2 or 3 independent experiments. Statistical analysis was performed by 1-way ANOVA followed by Tukey’s test (B). *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.