Cancer Innovation Laboratory, NCI-Frederick, Frederick, MD, 21702, USA.
Protein Characterization Laboratory, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD, 21702, USA.
Nat Commun. 2023 Aug 22;14(1):5114. doi: 10.1038/s41467-023-40738-4.
M1 macrophages enter a glycolytic state when endogenous nitric oxide (NO) reprograms mitochondrial metabolism by limiting aconitase 2 and pyruvate dehydrogenase (PDH) activity. Here, we provide evidence that NO targets the PDH complex by using lipoate to generate nitroxyl (HNO). PDH E2-associated lipoate is modified in NO-rich macrophages while the PDH E3 enzyme, also known as dihydrolipoamide dehydrogenase (DLD), is irreversibly inhibited. Mechanistically, we show that lipoate facilitates NO-mediated production of HNO, which interacts with thiols forming irreversible modifications including sulfinamide. In addition, we reveal a macrophage signature of proteins with reduction-resistant modifications, including in DLD, and identify potential HNO targets. Consistently, DLD enzyme is modified in an HNO-dependent manner at Cys and Cys, and molecular modeling and mutagenesis show these modifications impair the formation of DLD homodimers. In conclusion, our work demonstrates that HNO is produced physiologically. Moreover, the production of HNO is dependent on the lipoate-rich PDH complex facilitating irreversible modifications that are critical to NO-dependent metabolic rewiring.
M1 巨噬细胞进入糖酵解状态，此时内源性一氧化氮（NO）通过限制柠檬酸合酶 2 和丙酮酸脱氢酶（PDH）的活性来重新编程线粒体代谢。在这里，我们提供的证据表明，NO 通过使用硫辛酸生成亚硝酰（HNO）来靶向 PDH 复合物。富含 NO 的巨噬细胞中 PDH E2 相关的硫辛酸发生修饰，而 PDH E3 酶，也称为二氢硫辛酰胺脱氢酶（DLD），则不可逆地被抑制。从机制上讲，我们表明硫辛酸促进了 NO 介导的 HNO 的产生，该物质与巯基相互作用形成不可逆修饰，包括亚磺酰胺。此外，我们揭示了巨噬细胞中具有还原抗性修饰的蛋白质特征，包括 DLD，并确定了潜在的 HNO 靶标。一致地，DLD 酶以依赖 HNO 的方式在 Cys 和 Cys 处发生修饰，分子建模和突变分析表明这些修饰会损害 DLD 同源二聚体的形成。总之，我们的工作表明 HNO 是生理性产生的。此外，HNO 的产生依赖于富含硫辛酸的 PDH 复合物，该复合物促进了对 NO 依赖性代谢重编程至关重要的不可逆修饰。
