Mitochondrial oxidative phosphorylation system is recruited to detergent-resistant lipid rafts during myogenesis
Mitochondrial oxidative phosphorylation system is recruited to detergent-resistant lipid rafts during myogenesis
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DOI：
10.1002/pmic.200900826
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发表时间：
2010-07-01
期刊：
影响因子：
3.4
通讯作者：
Ko, Young-Gyu
中科院分区：
文献类型：
--
作者：
Kim, Bong-Woo;Lee, Joong-Won;Ko, Young-Gyu
由于耐去污剂脂筏在肌生成的信号转导中发挥重要作用，因此其全面的蛋白质组分析可以为了解其在肌管中的功能提供新的见解。在这里，从 C2C12 肌管中分离出耐去污剂的脂筏，并通过毛细管 RPLC/MS/MS 进行分析。在鉴定的 327 个蛋白质（或蛋白质组）中，28% 属于质膜或筏蛋白，29% 属于线粒体，20% 属于微粒体蛋白质，10% 属于其他蛋白质，13% 属于未知蛋白质。通过细胞分级、表面生物素标记、免疫荧光和脂筏分级，从 C2C12 肌管进一步证实了氧化磷酸化 (OXPHOS) 复合物在肌膜脂筏中的定位。添加外源细胞色素c后，从肌管分离的肌膜在NADH或琥珀酸盐存在的情况下具有消耗氧气的能力。通过抑制或下调肌管中的 OXPHOS I、III 和 IV，NADH 依赖性细胞外超氧化物的产生增加，表明 OXPHOS 蛋白是骨骼肌中细胞外 ROS 的主要来源。根据所有这些数据，我们可以得出结论，OXPHOS 蛋白在 C2C12 肌生成过程中与肌膜脂筏相关，以产生细胞外 ROS。
Since detergent-resistant lipid rafts play important roles in the signal transduction for myogenesis, their comprehensive proteomic analysis could provide new insights to understand their function in myotubes. Here, the detergent-resistant lipid rafts were isolated from C2C12 myotubes and analyzed by capillary RPLC/MS/MS Among the 327 proteins (or protein groups) identified, 28% were categorized to the plasma membrane or raft proteins, 29% to mitochondria, 20% to microsomal proteins, 10% to other proteins, and 13% to unknown proteins. The localization of oxidative phosphorylation (OXPHOS) complexes in the sarcolemma lipid rafts was further confirmed from C2C12 myotubes by cellular fractionation, surface-biotin labeling, immunofluorescence, and lipid raft fractionation. After adding exogenous cytochrome c, the sarcolemma isolated from myotubes had an ability to consume oxygen in the presence of NADH or succinate. The generation of NADH-dependent extracellular superoxide was increased by inhibiting or downregulating OXPHOS I, III, and IV in myotubes, indicating that OXPHOS proteins are major sources for extracellular ROS in skeletal muscle. With all these data, we can conclude that OXPHOS proteins are associated with the sarcolemma lipid rafts during C2C12 myogenesis to generate extracellular ROS.