Changes in the Turnover of the Cellular Proteome during Metabolic Reprogramming: A Role for mtROS in Proteostasis

The
role played by protein turnover in metabolic reprogramming
is unknown. Herein, using a SILAC approach, we have studied the changes
in the half-life of 266 proteins of energy metabolism and of translation
during the metabolic switch induced by the prolyl hydroxylases inhibitor
dimethyloxalylglycine (DMOG). DMOG induces HIF-1α expression
and triggers the activation of glycolysis and the concurrent inhibition
of mitochondrial respiration in colon cancer cells. Changes in the
activity of energy provision pathways correlated with increased turnover
rates of glycolytic enzymes and the stabilization of mitochondrial
proteins. Moreover, reprogramming also stabilized the proteins of
translation. The partial DMOG-mediated arrest of the synthesis of
mitochondrial and translation proteins results from the inhibition
of the mTORC1/p70SK/S6 signaling pathway. In contrast, DMOG stimulated
the synthesis of glycolytic enzymes, emphasizing the opposite and
differential regulation of the two pathways of energy provision. Addition
of MitoQ, a mitochondrial reactive oxygen species (mtROS) scavenger,
stabilized the turnover of cellular proteins similarly as when protein
degradation is inhibited with leupeptin, a serine-protease inhibitor.
Overall, the results show that the higher the activity of a pathway
the lower is the half-life of the proteins involved and suggest a
role for mtROS in cellular proteostasis. Data are available via ProteomeXchange
with identifier PXD013482.

蛋白质周转（protein turnover）在代谢重编程中所发挥的作用尚未明确。本研究采用稳定同位素细胞培养氨基酸标记（SILAC）技术，探究了脯氨酰羟化酶抑制剂二甲基草酰甘氨酸（DMOG）诱导的代谢转换过程中，266种能量代谢与翻译相关蛋白质的半衰期变化。DMOG可诱导结肠癌细胞内缺氧诱导因子-1α（HIF-1α）的表达，激活糖酵解通路并同时抑制线粒体呼吸。能量供给通路的活性变化与糖酵解酶周转速率升高及线粒体蛋白质稳定性提升呈相关性。此外，代谢重编程还可提升翻译相关蛋白质的稳定性。DMOG介导的线粒体与翻译相关蛋白质合成部分阻滞，源于mTORC1/p70SK/S6信号通路的抑制。与之相反，DMOG可促进糖酵解酶的合成，凸显了两条能量供给通路的反向差异化调控。加入线粒体活性氧（mitochondrial reactive oxygen species, mtROS）清除剂MitoQ后，细胞蛋白质的周转稳定性得到提升，其效果与使用丝氨酸蛋白酶抑制剂亮肽素（leupeptin）抑制蛋白质降解时一致。综上，本研究结果显示，某一通路的活性越高，其所涉及的蛋白质半衰期越短，同时提示mtROS在细胞蛋白质稳态（cellular proteostasis）中发挥作用。相关研究数据可通过标识符为PXD013482的蛋白质组交换数据库（ProteomeXchange）获取。