Identification and Characterization of Mitochondrial Targeting Sequence of Human Apurinic/Apyrimidinic Endonuclease 1

Cancer Center, Institute of Surgery, Daping Hospital, Third Military Medical University, Chongqing 400042, China.
Journal of Biological Chemistry (Impact Factor: 4.57). 03/2010; 285(20):14871-81. DOI: 10.1074/jbc.M109.069591
Source: PubMed


Dually targeted mitochondrial proteins usually possess an unconventional mitochondrial targeting sequence (MTS), which makes them difficult to predict by current bioinformatics approaches. Human apurinic/apyrimidinic endonuclease (APE1) plays a central role in the cellular response to oxidative stress. It is a dually targeted protein preferentially residing in the nucleus with conditional distribution in the mitochondria. However, the mitochondrial translocation mechanism of APE1 is not well characterized because it harbors an unconventional MTS that is difficult to predict by bioinformatics analysis. Two experimental approaches were combined in this study to identify the MTS of APE1. First, the interactions between the peptides from APE1 and the three purified translocase receptors of the outer mitochondrial membrane (Tom) were evaluated using a peptide array screen. Consequently, the intracellular distribution of green fluorescent protein-tagged, truncated, or mutated APE1 proteins was traced by tag detection. The results demonstrated that the only MTS of APE1 is harbored within residues 289-318 in the C terminus, which is normally masked by the intact N-terminal structure. As a dually targeted mitochondrial protein, APE1 possesses a special distribution pattern of different subcellular targeting signals, the identification of which sheds light on future prediction of MTSs.

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    • "Subsequently, the molecular chaperones allow proteins destined for the mitochondria to interact with translocase in the outer membrane of the TOM complex. Three subunits of this complex (Tom20, Tom22 and Tom70) function as import receptors for proteins containing an MTS [2] [5] [8] [11] [44]. To elucidate the interaction between rpS3 and TOM receptors in the cytosol, we performed a "
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    • "Recent studies have suggested the hypothesis of the involvement of a redox-mediating folding process of APE1 that would affect its subcellular distribution (Li et al. 2010; Vascotto et al. 2011). Increased cellular redox status would result in differential interaction of APE1 with mitochondrial transport proteins increasing the mitochondrial import into mitochondria after increased oxidative stress (Li et al. 2010; Vascotto et al. 2011). "
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    • "They showed that accumulation of APE1 within mitochondria, as a consequence of oxidative stress, may act as a protective mechanism facilitating mitochondrial genome maintenance and preventing apoptosis [17] [18]. APE1 is predominantly localized in the nucleus of most cell types, and mitochondrial APE1 is basically scarce and its mitochondrial translocation is largely conditional [19]. On the other hand, Singh et al. [8] found that mtDNA depletion promotes nuclear genomic instability and that the APE1 expression level is controlled by mtDNA copy number. "
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