| Summary: | 碩士 === 國立清華大學 === 分子醫學研究所 === 104 === NADH-dehydrogenase (ubiquinone) Fe-S protein 7 (NDUFS7), a highly conserved protein encoded by the nuclear DNA, is one of the core subunits in the mitochondrial NADH-coenzyme Q oxidoreductase (Complex I). It contains a tetranuclear iron-sulfur cluster N2 that assists electron transfer and serves as the final electron donor for ubiquinone and thus contributes to energy generation in the oxidative phosphorylation (OXPHOS) system. Mutations in NDUFS7 have been reported in patients suffering from neurodegenerative diseases such as Leigh syndrome and might also been assoicated with bipolar disorder. In addition to being present as an individual complex, the assembly of component OXPHOS complexes (Complex I~V) into several larger supercomplexes is considered to have functional significance not only for maintaining the stability of complexes but also for increasing the efficiency of electron transfer. In this study, we applied the RNA interference technique to suppress NDUFS7 expression in T-REx293 cells and then analyzed the obtained knockdown cell lines O10 and U21 with a series of functional assays. Our results showed that knockdown of NDUFS7 caused a significant decrease in the cellular oxygen consumption rate, mitochondrial membrane potential, ATP generation and Complex I activity. In addition, the amount of harmful reactive oxygen species was apparently increased in mitochondria. On the other hand, by conducting the high resolution clear native electrophoresis (HrCNE) analyses, we found that the deficiency of NDUFS7 or NDUFV2 influenced individual complex assembly and reduced the stability of Complex I subunits. By applying a milder condition for extraction of protein complexes from the mitochondrial samples, we also found that the deficiency of NDUFS7 could hamper supercomplex formation. Based on these findings, we conclude that NDUFS7 plays an indispensable role in maintaining the functions of electron transport chain and the assembly of individual Complex I, III, and IV and the supercomplex.
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