位於人類粒線體第一蛋白質複合體的次單元 NDUFS7 (NADH dehydrogenase (ubiquinone) Fe-S protein 7)，是一個由細胞核基因所表現並且具有高度保留性的蛋白。NDUFS7 含有一個 [4Fe-4S] 的鐵硫中心 N2，其為第一複合體親水端上電子的最後接受者，同時也是 Quinone的還原者。在單純的模式生物以及病人檢體的研究中，NDUFS7 在第一蛋白質複合體的聚合和功能上扮演著重要的角色。NDUFS7 的突變目前已知與 Leigh 症候群 (Leigh syndrome)、躁鬱症有關。為了詳細探討 NDUFS7 在人類粒線體第一蛋白質複合體上的功能，本研究利用核糖核酸干擾技術 (RNAi) 降低 NDUFS7 在 T-REx293 細胞株的表現。結果顯示，NDUFS7 表現量的下降會導致細胞生長速度、三磷酸腺苷 (ATP) 產生量、耗氧量和第一蛋白質複合體活性的降低。這些證據指出，人類細胞中 NDUFS7 在第一蛋白質複合體的活性和能量的產生上扮演著重要的角色。

NADH dehydrogenase (ubiquinone) Fe-S protein 7 (NDUFS7) is one of the nuclear-encoded and highly conserved core subunits of mitochondrial complex I. This protein houses the tetranuclear iron-sulfur cluster N2, which is the terminal receptor of electrons and the reducer for quinone in complex I peripheral arm. NDUFS7 protein plays a critical role in complex I assembly and activity, according to the results of studies from simple organisms and human disease models. Mutations of NDUFS7 have been associated with Leigh Syndrome and bipolar disorder. To detailedly investigate the role of NDUFS7 in mitochondrial complex I of human cells, NDUFS7 was suppressed in human T-REx293 cells by RNA interference system in this study. In the NDUFS7 knockdown cells, the growth rate, ATP generation, oxygen consumption and complex I activity were reduced. These results provided the evidence of the essential role of NDUFS7 for complex I activity and energy production in human cells.

1. Anderson, S., Bankier, A. T., Barrell, B. G., de Bruijn, M. H. L., Coulson, A. R., Drouin, J., Eperon, I. C., Nierlich, D. P., Roe, B. A., Sanger, F., Schreier, P. H., Smith, A. J. H., Staden, R., and Young, I. G. (1981) Sequence and organization of the human mitochondrial genome, Nature 290, 457-465.
2. Yagi, T., and Matsuno-Yagi, A. (2003) The proton-translocating NADH−quinone oxidoreductase in the respiratory chain: the secret unlocked, Biochemistry 42, 2266-2274.
3. Carroll, J., Fearnley, I. M., Skehel, J. M., Shannon, R. J., Hirst, J., and Walker, J. E. (2006) Bovine complex I is a complex of forty-five different subunits, Journal of Biological Chemistry 281, 32724-32727.
4. Clason, T., Ruiz, T., Schägger, H., Peng, G., Zickermann, V., Brandt, U., Michel, H., and Radermacher, M. (2010) The structure of eukaryotic and prokaryotic complex I, Journal of Structural Biology 169, 81-88.
5. Ohnishi, T. (1998) Iron–sulfur clusters/semiquinones in complex I, Biochimica et Biophysica Acta (BBA) - Bioenergetics 1364, 186-206.
6. Dutton, P. L., Moser, C. C., Sled, V. D., Daldal, F., and Ohnishi, T. (1998) A reductant-induced oxidation mechanism for complex I, Biochimica et biophysica acta 1364, 245-257.
7. Sazanov, L. A., Peak-Chew, S. Y., Fearnley, I. M., and Walker, J. E. (2000) Resolution of the membrane domain of bovine complex I into subcomplexes: implications for the structural organization of the enzyme, Biochemistry 39, 7229-7235.
8. Friedrich, T. (2001) Complex I: a chimaera of a redox and conformation-driven proton pump?, J Bioenerg Biomembr 33, 169-177.
9. Holt, P. J., Morgan, D. J., and Sazanov, L. A. (2003) The location of NuoL and NuoM subunits in the membrane domain of the Escherichia coli complex I: implications for the mechanism of proton pumping, Journal of Biological Chemistry 278, 43114-43120.
10. Brandt, U. (2006) Energy converting NADH:quinone oxidoreductase (complex I), Annu. ReV. Biochem 75, 69-92.
11. Sazanov, L. A., and Hinchliffe, P. (2006) Structure of the hydrophilic domain of respiratory complex I from Thermus thermophilus, Science 311, 1430-1436.
12. Berrisford, J. M., and Sazanov, L. A. (2009) Structural basis for the mechanism of respiratory complex I, Journal of Biological Chemistry 284, 29773-29783.
43
13. Efremov, R. G., Baradaran, R., and Sazanov, L. A. (2010) The architecture of respiratory complex I, Nature 465, 441-445.
14. Hunte, C., Zickermann, V., and Brandt, U. (2010) Functional modules and structural basis of conformational coupling in mitochondrial complex I, Science 329, 448-451.
15. Efremov, R. G., and Sazanov, L. A. (2011) Structure of the membrane domain of respiratory complex I, Nature 476, 414-420.
16. Hyslop, S. J., Duncan, A. M., Pitkänen, S., and Robinson, B. H. (1996) Assignment of the PSST subunit gene of human mitochondrial complex I to chromosome 19p13, Genomics 37, 375-380.
17. Ingledew, W. J., and Ohnishi, T. (1980) An analysis of some thermodynamic properties of iron-sulphur centres in site I of mitochondria, Biochemical Journal 186, 111-117.
18. Wang, D.-C., Meinhardt, S. W., Sackmann, U., Weiss, H., and Ohnishi, T. (1991) The iron-sulfur clusters in the two related forms of mitochondrial NADH: ubiquinone oxidoreductase made by Neurospora crassa, European Journal of Biochemistry 197, 257-264.
19. Hellwig, P., Scheide, D., Bungert, S., Mäntele, W., and Friedrich, T. (2000) FT-IR spectroscopic characterization of NADH:ubiquinone oxidoreductase (complex I) from Escherichia coli: oxidation of FeS cluster N2 is coupled with the protonation of an aspartate or glutamate side chain, Biochemistry 39, 10884-10891.
20. Brandt, U. (1997) Proton-translocation by membrane-bound NADH:ubiquinone-oxidoreductase (complex I) through redox-gated ligand conduction, Biochimica et Biophysica Acta (BBA) - Bioenergetics 1318, 79-91.
21. Schuler, F., Yano, T., Di Bernardo, S., Yagi, T., Yankovskaya, V., Singer, T. P., and Casida, J. E. (1999) NADH-quinone oxidoreductase: PSST subunit couples electron transfer from iron–sulfur cluster N2 to quinone, Proceedings of the National Academy of Sciences 96, 4149-4153.
22. Ohnishi, S. T., Ohnishi, T., Muranaka, S., Fujita, H., Kimura, H., Uemura, K., Yoshida, K.-i., and Utsumi, K. (2005) A possible site of superoxide generation in the complex I segment of rat heart mitochondria, J Bioenerg Biomembr 37, 1-15.
23. Garofano, A., Zwicker, K., Kerscher, S., Okun, P., and Brandt, U. (2003) Two aspartic acid residues in the PSST-homologous NUKM subunit of complex I from Yarrowia lipolytica are essential for catalytic activity, Journal of Biological Chemistry 278, 42435-42440.
24. Duarte, M., Pópulo, H., Videira, A., Friedrich, T., and Schulte, U. (2002)
44
Disruption of iron-sulphur cluster N2 from NADH: ubiquinone oxidoreductase by site-directed mutagenesis, Biochemical Journal 364, 833-839.
25. Flemming, D., Schlitt, A., Spehr, V., Bischof, T., and Friedrich, T. (2003) Iron-sulfur cluster N2 of the Escherichia coli NADH:ubiquinone oxidoreductase (Complex I) is located on subunit NuoB, Journal of Biological Chemistry 278, 47602-47609.
26. Yano, T., and Yagi, T. (1999) H+-translocating NADH-quinone oxidoreductase (NDH-1) of Paracoccus denitrificans : studies on topology and stoichiometry of the peripheral subunits, Journal of Biological Chemistry 274, 28606-28611.
27. Di Bernardo, S., and Yagi, T. (2001) Direct interaction between a membrane domain subunit and a connector subunit in the H+-translocating NADH-quinone oxidoreductase, FEBS Letters 508, 385-388.
28. Ahlers, P. M., Garofano, A., Kerscher, S. J., and Brandt, U. (2000) Application of the obligate aerobic yeast Yarrowia lipolytica as a eucaryotic model to analyse Leigh syndrome mutations in the complex I core subunits PSST and TYKY, Biochimica et biophysica acta 1459, 258-265.
29. Lebon, S., Rodriguez, D., Bridoux, D., Zerrad, A., Rötig, A., Munnich, A., Legrand, A., and Slama, A. (2007) A novel mutation in the human complex I NDUFS7 subunit associated with Leigh syndrome, Molecular Genetics and Metabolism 90, 379-382.
30. Lebon, S., Minai, L., Chretien, D., Corcos, J., Serre, V., Kadhom, N., Steffann, J., Pauchard, J.-Y., Munnich, A., Bonnefont, J.-P., and Rötig, A. (2007) A novel mutation of the NDUFS7 gene leads to activation of a cryptic exon and impaired assembly of mitochondrial complex I in a patient with Leigh syndrome, Molecular Genetics and Metabolism 92, 104-108.
31. Efremov, R., and Sazanov, L. (2012) Structure of complex I, In A Structural Perspective on Respiratory Complex I (Sazanov, L., Ed.), pp 3-21, Springer Netherlands.
32. Danis, L. (1951) Subacute necrotizing encephalomyelopathy in an infant, J Neurol Neurosurg Psychiatry 14, 216.
33. Dahl, H. H. (1998) Getting to the nucleus of mitochondrial disorders identification of respiratory chain–enzyme genes causing Leigh syndrome, Am J Hum Genet 63, 1594.
34. Triepels, R. H., van den Heuvel, L. P., Loeffen, J. L., Buskens, C. A., Smeets, R. J., Rubio Gozalbo, M. E., Budde, S. M., Mariman, E. C., Wijburg, F. A., Barth, P. G., Trijbels, J. M., and Smeitink, J. A. (1999) Leigh syndrome associated with a mutation in the NDUFS7 (PSST) nuclear encoded subunit of complex I, Neurol 45, 787.
45
35. Visch, H.-J., Rutter, G. A., Koopman, W. J. H., Koenderink, J. B., Verkaart, S., de Groot, T., Varadi, A., Mitchell, K. J., van den Heuvel, L. P., Smeitink, J. A. M., and Willems, P. H. G. M. (2004) Inhibition of mitochondrial Na+-Ca2+ exchange restores agonist-induced ATP production and Ca2+ handling in human complex I deficiency, Journal of Biological Chemistry 279, 40328-40336.
36. Sun, X., Wang, J., Tseng, M., and Young, L. T. (2006) Downregulation in components of mitochondrial electron transport chain in postmortem frontal cortex from subjects with bipolar disorder, J Psychiatry Neurosci 31(3), 189.
37. Andreazza Ac, S. L., W. J., Y. L. (2010) Mitochondrial complex I activity and oxidative damage to mitochondrial proteins in the prefrontal cortex of patients with bipolar disorder, Archives of General Psychiatry 67, 360-368.
38. Rera, M., Azizi, M. J., and Walker, D. W. (2012) Organ-specific mediation of lifespan extension: more than a gut feeling?, Ageing Research Reviews, (Epub ahead of print).
39. Copeland, J. M., Cho, J., Lo Jr, T., Hur, J. H., Bahadorani, S., Arabyan, T., Rabie, J., Soh, J., and Walker, D. W. (2009) Extension of Drosophila life span by RNAi of the mitochondrial respiratory chain, Current Biology 19, 1591-1598.
40. Chou, T. F. (2009) Human mitochondrial complex I NDUFS7 subunit has a dual distribution both in mitochondria and nuclei. Master Thesis, National Tsing Hua University.
41. Chen, R. H., Sarnecki, C., and Blenis, J. (1992) Nuclear localization and regulation of erk- and rsk-encoded protein kinases, Molecular and Cellular Biology 12, 915-927.
42. Madison, D. L., Yaciuk, P., Kwok, R. P. S., and Lundblad, J. R. (2002) Acetylation of the adenovirus-transforming rrotein E1A determines nuclear localization by disrupting association with importin-α, Journal of Biological Chemistry 277, 38755-38763.
43. Ulrich, H. D. (2002) Degradation or maintenance: actions of the ubiquitin system on eukaryotic chromatin, Eukaryotic Cell 1, 1-10.
44. Cho, G., Lim, Y., and Golden, J. A. (2009) SUMO interaction motifs in Sizn1 are required for promyelocytic leukemia protein nuclear body localization and for transcriptional activation, Journal of Biological Chemistry 284, 19592-19600.
45. Rossignol, R., Faustin, B., Rocher, C., Malgat, M., Mazat, J. P., and Letellier, T. (2003) Mitochondrial threshold effects, Journal of Biochemistry 370, 751.
46 James, A., Wei, Y., Pang, C. and Murphy, M. (1996) Altered mitochondrial
46
function in fibroblasts containing MELAS or MERRF mitochondrial DNA mutations. Biochem. J. 318, 401-407
47 Lodi, R., Carelli, V., Lotti, S., Valentino, L., Barboni, P., Palloti, F., Montagna, P. and Barbiroli, B. (2001) The biochemical expression of the 3460/ND1 leber's hereditary optic neuropathy mtDNA mutation is tissue specific. An in vivo phosphorus MR spectroscopy study of one pedigree. In The Fifth European Meeting on Mitochondrial Pathology, Venice, Italy, 19-23 September 2001, abstr. book, p. 194