Cellular pyrimidine imbalance triggers mitochondrial DNA-dependent innate immunity [HeLa_CAD_RNA-seq]

Cytosolic mitochondrial DNA (mtDNA) elicits a type I interferon response, but signals triggering the release of mtDNA from mitochondria remain enigmatic. Here, we show that mtDNA-dependent immune signalling via the cyclic GMP–AMP syn- thase‒stimulator of interferon genes‒TANK-binding kinase 1 (cGAS–STING–TBK1) pathway is under metabolic control and is induced by cellular pyrimidine deficiency. The mitochondrial protease YME1L preserves pyrimidine pools by supporting de novo nucleotide synthesis and by proteolysis of the pyrimidine nucleotide carrier SLC25A33. Deficiency of YME1L causes inflamma- tion in mouse retinas and in cultured cells. It drives the release of mtDNA and a cGAS–STING–TBK1-dependent inflammatory response, which requires SLC25A33 and is suppressed upon replenishment of cellular pyrimidine pools. Overexpression of SLC25A33 is sufficient to induce immune signalling by mtDNA. Similarly, depletion of cytosolic nucleotides upon inhibition of de novo pyrimidine synthesis triggers mtDNA-dependent immune responses in wild-type cells. Our results thus identify mtDNA release and innate immune signalling as a metabolic response to cellular pyrimidine deficiencies. Transcript profiles of HeLa cells treated with esiRNA targeting Gfp (control) or CAD in the presence and absence of 2',3' dideoxycytidine (ddC)

胞质线粒体DNA（mtDNA）可引发I型干扰素应答，但触发线粒体mtDNA释放的信号机制仍不明晰。本研究证实，经由环GMP-AMP合酶-干扰素基因刺激因子-TANK结合激酶1（cGAS–STING–TBK1）通路介导的mtDNA依赖性免疫信号通路受代谢调控，且可由细胞嘧啶缺乏所诱导。线粒体蛋白酶YME1L通过支持核苷酸从头合成，以及降解嘧啶核苷酸转运蛋白SLC25A33，来维持细胞内嘧啶池稳态。YME1L缺失会导致小鼠视网膜及培养细胞发生炎症反应，其可驱动mtDNA释放，并引发依赖cGAS–STING–TBK1的炎症应答；该过程依赖SLC25A33，且可通过补充细胞嘧啶池得到抑制。过表达SLC25A33即可通过mtDNA触发免疫信号通路。同样，抑制从头嘧啶合成途径导致胞内核苷酸耗竭时，也会在野生型细胞中触发mtDNA依赖性免疫应答。综上，本研究将mtDNA释放与先天免疫信号鉴定为细胞对嘧啶缺乏的代谢性应答。本数据集包含经靶向Gfp（对照）或CAD的esiRNA处理，且分别添加或不添加2',3'-二脱氧胞苷（ddC）的HeLa细胞转录组谱数据。