Proteomics Reveals that Methylmalonyl-CoA Mutase Modulates Cell Architecture and Increases Susceptibility to Stress
Proteomics Reveals that Methylmalonyl-CoA Mutase Modulates Cell Architecture and Increases Susceptibility to Stress
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DOI：
10.3390/ijms21144998
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发表时间：
2020-07-01
影响因子：
5.6
通讯作者：
Ruoppolo, Margherita
中科院分区：
文献类型：
--
作者：
Costanzo, Michele;Caterino, Marianna;Ruoppolo, Margherita
甲基丙二酸血症（MMA）是一种罕见的先天性代谢缺陷引起的缺乏甲基丙二酰辅酶A（MUT）酶。下游MUT缺陷，甲基丙二酸积累与毒性代谢产物一起从丙酰辅酶A和其他化合物上游的块在酶途径。表现为危及生命的酸中毒、呼吸窘迫、脑功能紊乱、高氨血症和酮症。幸存者发展出知之甚少的多器官损伤，特别是大脑和肾脏。通过CRISPR/Cas9技术工程化HEK 293细胞系以敲除MUT基因（MUT-KO）。鸟枪式无标记定量蛋白质组学和生物信息学分析揭示了MUT缺陷细胞中潜在的破坏性生物过程。MUT-KO诱导细胞结构和形态的改变以及ROS过度产生。我们发现参与细胞骨架和细胞粘附组织，细胞运输，线粒体和氧化过程的蛋白质的改变，通过Vim，EXT 2，SDC 2，FN 1，GLUL和CHD 1的调节进行验证。此外，开发了MUT拯救的细胞模型以控制MUT-KO效应的特异性。在全球范围内，MUT-KO的蛋白质组学景观表明，细胞模型通过线粒体功能受损和氧化还原过程中的不平衡，对丙酸盐和H2 O2诱导的应激具有增加的易感性。
Methylmalonic acidemia (MMA) is a rare inborn error of metabolism caused by deficiency of the methylmalonyl-CoA mutase (MUT) enzyme. Downstream MUT deficiency, methylmalonic acid accumulates together with toxic metabolites from propionyl-CoA and other compounds upstream of the block in the enzyme pathway. The presentation is with life-threatening acidosis, respiratory distress, brain disturbance, hyperammonemia, and ketosis. Survivors develop poorly understood multi-organ damage, notably to the brain and kidneys. The HEK 293 cell line was engineered by CRISPR/Cas9 technology to knock out theMUTgene (MUT-KO). Shotgun label-free quantitative proteomics and bioinformatics analyses revealed potential damaging biological processes in MUT-deficient cells. MUT-KO induced alteration of cellular architecture and morphology, and ROS overproduction. We found the alteration of proteins involved in cytoskeleton and cell adhesion organization, cell trafficking, mitochondrial, and oxidative processes, as validated by the regulation of VIM, EXT2, SDC2, FN1, GLUL, and CHD1. Additionally, a cell model of MUT-rescuing was developed in order to control the specificity of MUT-KO effects. Globally, the proteomic landscape of MUT-KO suggests the cell model to have an increased susceptibility to propionate- and H2O2-induced stress through an impairment of the mitochondrial functionality and unbalances in the oxidation-reduction processes.