期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 283, 期 14, 页码 8984-8994出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M709040200
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资金
- NCI NIH HHS [U54 CA091408, U54 CA 91408] Funding Source: Medline
- NCRR NIH HHS [U54 RR 019192-04, G12 RR003032, U54 RR019192, G12 RR 03032-19] Funding Source: Medline
- NHLBI NIH HHS [T32 HL 007737, HL 66109] Funding Source: Medline
- NIDDK NIH HHS [DK 07319] Funding Source: Medline
- NIGMS NIH HHS [S06 GM 08037, 5R25 GM 059994] Funding Source: Medline
- NINDS NIH HHS [U54 NS 041071-06, U54 NS041071] Funding Source: Medline
- PHS HHS [D34 HP 00003] Funding Source: Medline
Nuclear factor erythroid 2-related factor 2 ( Nrf2) mediates the transcriptional response of cells to oxidative stress and is translocated into the nucleus following, or concomitant with, its activation by electrophiles or reactive oxygen species. The mechanism of its translocation into the nucleus is not entirely elucidated. Here we have identified two novel nuclear localization signal ( NLS) motifs in murine Nrf2, one located near the N-terminal region ( amino acid residues 42-53) and the other ( residues 587-593) located near the C-terminal region. Imaging of green fluorescent protein ( GFP)-tagged Nrf2 revealed that mutation(s) in any of these sequences resulted in decreased nuclear fluorescence intensity compared with the wild-type Nrf2 when Nrf2 activation was induced with the electrophile tert-butylhydroquinone. The mutations also impaired Nrf2-induced transactivation of antioxidant response element-driven reporter gene expression to the same extent as the Nrf2 construct bearing mutation in a previously identified bipartite NLS that maps at residues 494-511. When linked to GFP or to GFPPEPCK-C each of the novel NLS motifs was sufficient to drive nuclear translocation of the fusion proteins. Co-immunoprecipitation assays demonstrated that importins alpha 5 and beta 1 associate with Nrf2, an interaction that was blocked by the nuclear import inhibitor SN50. SN50 also blocked tert-butylhydroquinone-induced nuclear fluorescence of GFP-Nrf2 in cells transfected with wild-type GFP-Nrf2. Overall these results reveal that multiple NLS motifs in Nrf2 function in its nuclear translocation in response to pro-oxidant stimuli and that the importin alpha-beta heterodimer nuclear import receptor system plays a critical role in the import process.
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