Laboratory of Disease Models, School of Veterinary Medicine, Rakuno Gakuen University, Ebestsu 069-8501, Japan.
Laboratory of Immunology, Faculty of Pharmacy, Osaka Ohtani University, Osaka 584-8540, Japan.
Int J Mol Sci. 2024 Nov 6;25(22):11936. doi: 10.3390/ijms252211936.
Ubiquitin-specific protease 2 (USP2) maintains mitochondrial integrity in culture myoblasts. In this study, we investigated the molecular mechanisms underlying the protective role of USP2 in mitochondria. The knockout (KO) of the gene or the chemical inhibition of USP2 induced a robust accumulation of mitochondrial reactive oxygen species (ROS), accompanied by defects in mitochondrial membrane potential, in C2C12 myoblasts. ROS removal by N-acetyl-L-cysteine restored the mitochondrial dysfunction induced by USP2 deficiency. Comprehensive RT-qPCR screening and following protein analysis indicated that both the genetic and chemical inhibition of USP2 elicited a decrease in uncoupling protein 2 (UCP2) at mRNA and protein levels. Accordingly, the introduction of a -expressing construct effectively recovered the mitochondrial membrane potential, entailing an increment in the intracellular ATP level in KO C2C12 cells. In contrast, USP2 deficiency also decreased peroxisome proliferator-activated receptor γ coactivator 1α (PGC1α) protein in C2C12 cells, while it upregulated mRNA. Overexpression studies indicated that USP2 potentially stabilizes PGC1α in an isopeptidase-dependent manner. Given that PGC1α is an inducer of UCP2 in C2C12 cells, USP2 might ameliorate mitochondrial ROS by maintaining the PGC1α-UCP2 axis in myoblasts.
泛素特异性蛋白酶 2（USP2）在培养的成肌细胞中维持线粒体的完整性。在这项研究中，我们研究了 USP2 在保护线粒体中发挥作用的分子机制。基因敲除（KO）或 USP2 的化学抑制会导致 C2C12 成肌细胞中线粒体活性氧（ROS）的大量积累，同时伴随着线粒体膜电位的缺陷。用 N-乙酰-L-半胱氨酸去除 ROS 可以恢复 USP2 缺乏引起的线粒体功能障碍。综合 RT-qPCR 筛选和随后的蛋白分析表明，USP2 的基因和化学抑制都会导致解偶联蛋白 2（UCP2）在 mRNA 和蛋白水平上的减少。因此，-表达载体的引入有效地恢复了 KO C2C12 细胞中的线粒体膜电位，导致细胞内 ATP 水平升高。相比之下，USP2 的缺乏也会降低 C2C12 细胞中的过氧化物酶体增殖物激活受体 γ 共激活物 1α（PGC1α）蛋白，同时上调其 mRNA。过表达研究表明，USP2 可能以依赖于肽基水解酶的方式稳定 PGC1α。鉴于 PGC1α 是 C2C12 细胞中 UCP2 的诱导剂，USP2 可能通过维持成肌细胞中的 PGC1α-UCP2 轴来减轻线粒体 ROS。
