Transcription factor GABP/NRF-2 controlling biogenesis of mitochondria
regulates basal expression of peroxiredoxin V but the mitochondrial function
of peroxiredoxin V is dispensable in the dog
Nadezhda Usmanovaa, Nikolai Tomilina, Boris Zhivotovskyb, Andrey Kropotova,*
aInstitute of Cytology, Russian Academy of Sciences, Tikhorestkii Avenue 4, 194064 St. Petersburg, Russ
bInstitute of Environmental Medicine, Division of Toxicology, Karolinska Institutet, Stockholm, Sweden
a r t i c l e i n f o
Received 7 June 2010
Accepted 1 October 2010
Available online xxx
Gene expression regulation
a b s t r a c t
Peroxiredoxins (PRDXs) represent a conserved family of six antioxidant proteins which are widely
expressed in different organisms. Human PRDX5 is detected in the cytosol and nucleus and can also
target peroxisomes and mitochondria. However, it remains unknown if mitochondrial localization of
PRDX5 is essential for its functions. Here we studied whether the known regulator of mitochondrial
biogenesis, transcription factor GABP/NRF-2, is required for the basal expression of the human PRDX5
gene and what the significance is of the mitochondrial targeting of the PRDX5 protein. It was found that
mutation-mediated inactivation of all potential binding sites for GAPB in the PRDX5 promoter lead to
w80% inhibition of its basal activity in a reporter gene assay. Co-transfection of plasmids expressing
GABP-alpha and GABP-beta stimulated activity of the non-mutated PRDX5 promoter but had no effect on
the mutated promoter, suggesting that basal expression of the human PRDX5 gene is regulated by GABP.
We found that the dog c-Myc-tagged PRDX5 did not target the mitochondria of human cells. Endoge-
nously expressed PRDX5 also showed no association with mitochondria in the dog cells. It appears,
therefore, that during evolution the dog PRDX5 gene lost its upstream ATG codon and mitochondrial
targeting signal without major functional consequences.
? 2010 Published by Elsevier Masson SAS.
reduce hydrogen peroxide and organic hydroperoxides by electron
transfer from thioredoxins, glutathione, or cyclophilins . Perox-
iredoxin proteins are present in most living species requiring
oxygen. Mammalian cells express six isoforms (PRDX1 to PRDX6)
that are encoded by different nuclear genes. They are important in
antioxidant defense and in hydrogen peroxide-mediated signalling
[2e4]. Furthermore, the importance of mammalian PRDXs was
shown in other cellular processes, including apoptosis, cell prolif-
eration and differentiation [5,6]. PRDX1 is mainly located in the
cytoplasm and knockout of its gene in mice leads to haemolytic
anaemia and a shortened life span . PRDX2 was found to bind to
integral membrane proteins or cell membranes via its C-terminal
region and knockout of its gene in mice also leads to haemolytic
anaemia . PRDX3 is located in mitochondria and targeting of its
gene in mice results in the accumulation of reactive oxygen species
(ROS) in macrophages and increased lung sensitivity to inflam-
mation-inducing agents . PRDX4 is present as a secretory protein
in most tissues. In sexually mature testis it is anchored to the
endoplasmic reticulum membrane of spermatogenic cells via an
uncleaved N-terminal hydrophobic peptide. PRDX4 knockout
results in elevated spermatogenic cell death via oxidative stress
. PRDX6 was identified as a secretory antioxidant protein of the
olfactory epithelium  and PRDX6 gene knockout mice show
lung pathology and increased mortality with hyperoxia .
PRDX5 is a thioredoxin peroxidase, which is highly expressed in
many tissues. Human PRDX5 (hPRDX5) contains N-terminal mito-
chondrial and C-terminal peroxisome targeting (PTS1) signals,
allowing its localization to mitochondria [13,16] and peroxisomes
[13,14,16]. A significant amount of PRDX5 is also present in the
cytosol and in the nucleus . Similar to other 2-Cys peroxir-
edoxins, PRDX5 requires a thioredoxin  or cyclophilin A  as
a reducing partner. The peroxidase function of PRDX5 in vivo was
Abbreviations: ETS, a family of transcription factors; EBS, ETS binding site; PRDX,
peroxiredoxin; NRF-1, nuclear respiratory factor 1; GABP, GA-binding protein; aTIS,
alternative translation initiation sites; GFP, green fluorescent protein; dPRDX5, dog
PRDX5; hPRDX5, human PRDX5; PBS, phosphate-buffered saline.
* Corresponding author. Tel.: þ7 812 2970326; fax: þ7 812 2970341.
E-mail address: firstname.lastname@example.org (A. Kropotov).
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0300-9084/$ e see front matter ? 2010 Published by Elsevier Masson SAS.
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