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 7

cGAS–ZBP1–RIPK1/3 axis and GOT2/GPT2 are essential for cGAMP+AOA–induced antitumor immune response.

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cGAS–ZBP1–RIPK1/3 axis and GOT2/GPT2 are essential for cGAMP+AOA–induced...
(A) MC38 cells were transduced with control sgRNA or sgRNA targeting cGAS. Whole cell lysates (WCLs) were analyzed by immunoblotting with the indicated antibodies. (B and C) MC38 cells as described in A were subcutaneously transplanted into C57BL/6J mice and administered according to the protocol mentioned above. Tumor volume (B) and tumor photos (C) were recorded. (D and E) MC38 cells described in A were subcutaneously transplanted into C57BL/6J mice and administered according to the protocol mentioned above. The percentage of CD8+ T cells in gated CD3+ T cells (D) and IFN-γ production (E) on CD8+ T cells. (F) MC38 cells were transduced with control sgRNA or sgRNA targeting Zbp1. WCLs were analyzed by immunoblotting with the indicated antibodies. (G and H) MC38 cells as described in F were subcutaneously transplanted into C57BL/6J mice and administered according to the protocol mentioned above. Tumor volume (G) and tumor photos (H). (I and J) MC38 cells described in G were subcutaneously transplanted into C57BL/6J mice and administered according to the protocol mentioned above. The production of IFN-γ (I) and TNF-α (J) on isolated CD8+ T cells. (K) MC38 cells were transduced with control shRNA or shRNA targeting Ripk1, and the stable cells were further transduced with control shRNA or shRipk3 to generate double-knockdown (double-KD) cells. WCLs were analyzed by immunoblotting with the indicated antibodies. (L and M) MC38 cells described in K were subcutaneously transplanted into C57BL/6J mice and administered according to the protocol mentioned above. Tumor volume (L) and tumor photos (M). (N and O) The production of IFN-γ (N) and TNF-α (O) on isolated CD8+ T cells. (P) MC38 cells were transduced with control shRNA or shRNA targeting Got2, and the stable cells were further transduced with control shRNA or shGpt2 to generate double-KD cells. WCLs were analyzed by immunoblotting with the indicated antibodies. (Q and R) MC38 cells described in P were subcutaneously transplanted into C57BL/6J mice and administered according to the protocol above. Tumor volume (Q) and tumor photos (R). (S and T) The percentage of CD8+ T cells in gated CD3+ T cells (S) and IFN-γ production (T) on CD8+ T cells. Statistical analysis was performed by 2-way ANOVA and Tukey’s or Bonferroni’s test (B–T). *P < 0.05; **P < 0.01; ***P < 0.001.