Abstract
Human NADH dehydrogenase (ubiquinone) Fe-S protein 7 (NDUFS7) is one of 44 subunits in mitochondrial complex I. The N-terminal 1-60 amino acids of NDUFS7 have been defined as a mitochondrial targeting sequence (MTS) and the C-terminus contains a nuclear localization signal (NLS) and a nuclear export signal (NES). The sequence of NDUFS7 is highly conserved in the motif: CCXXE(X)60C(X)30CP. It can bind to an iron-sulfur cluster (a [4Fe-4S] cluster) called N2, a redox center in the terminus of complex I electron transfer pathway. In previous study, we identified NDUFS7 as a protein substrate of sumoylation. Posttranslational modifications of proteins by the small ubiquitin-like modifier (SUMO) have been found to be associated with various cellular processes. The SUMO protein can be conjugated to a target protein by sequential actions of enzyme E1 (SAE1/2), enzyme E2 (UBC9) and E3 ligases. The proteins conjugated by SUMO could be involved in subcellular localization, function or responding to stresses. In this study, NDUFS7, SUMO-1 and UBC9 were co-expressed in HEK293 cells to clarify its subcellular localization by cell fractionation. The result showed that the sumoylated NDUFS7 were present in both the nuclear and cytosol fractions. Then, we established a new construct which can express the NDUFS7-SUMO-1 fusion protein, and found that the fusion protein can block the mitochondrial import. Furthermore, the effect of different stress response on sumoylation of NDUFS7 was explored by treating cells with various stress inducers, such as CoCl2, H2O2, starvation and apoptosis reagents. The results suggested that CoCl2-induced hypoxia increases the sumoylation of NDUFS7. The effects of NDUFS7 sumoylation on the subcellular localization of the protein and its physiological consequence would be further explored.