Biology Reference
In-Depth Information
oxidative stress in the livers of APE1 haploinsufficient
mice (Apex
þ
/-
).
113
2-NP is a hepatocarcinogen that
generates ROS promoting oxidative DNA damage.
114
In a previous study, Heydari and coworkers showed
that base excision repair activity is upregulated in
response to oxidative stress with a four-fold increase
in 8-hydroxydeoxyguanosine, 4
arrest and apoptosis were examined in response to
oxidative stress through monitoring levels of growth
arrest and DNA-damage inducible gamma, GADD45g,
p53, and caspase-3. Loss of APE1 was found to increase
apoptotic response due to significant increases in all
three of these proteins associated with cell cycle arrest
and apoptosis. Taken together, the data suggest an
important role for APE1's redox activity as well as its
DNA repair activity in animals subjected to oxidative
stress. This finding is consistent with previous reports
in which the redox activity of APE1 is subject to regu-
lation in response to various stimuli including oxida-
tive stress,
116,117
is essential for survival of neuronal
cells subjected to oxidative stress,
118
and is required
for normal embryonic hematopoiesis.
119
5-fold increases in
DNA single-strand breaks, two-fold increase in p53
levels, and a 42% increase in the level of DNA poly-
merase beta and BER capacity in C57BL/6 mice.
115
In
this study, they first examined induction of APE1 and
DNA-binding activity of NF-
k
B in response to treat-
ment with 2-NP. Both APE1 mRNA and protein levels
were increased in liver extracts of C57BL/6 mice as
was the DNA-binding activity of NF-
e
B, consistent
with stimulation by APE1's redox activity. They next
examined the effect of 2-NP on APE1 levels and NF-
k
k
Thus, an
optimal balance between NF-
B mediated and DNA
repair pathways is required for survival and is compro-
mised in Apex
þ
/-
haploinsufficient mice exposed to
oxidative stress. As the AP-endonuclease activity was
not found to be limiting in the overall base excision
repair activity in Apex
þ
/-
animals subjected to oxidative
stress, it is the reduced redox activity and 3'-phospho-
diesterase activity of APE1 that compromise cell
survival.
113
Thus, development of therapeutics target-
ing both APE1's redox and repair activities could well
potentiate the effectiveness of current cancer drugs for
which resistance is observed in response to increases
in APE1 levels.
9,120,121
k
B DNA-binding activity in Apex
þ
/-
and found as
expected that APE1 levels were significantly reduced
as was NF-
k
B DNA-binding activity. Comparisons
were then made between wt and Apex
þ
/-
animals under
normal and oxidative stress conditions. The levels of
DNA damage as measured by AP sites, single-strand
breaks, and aldehydic lesions were the same as wild-
type mice for the Apex
þ
/-
under normal conditions.
Under oxidative stress conditions, the number of
3'OH-containing single-strand breaks was significantly
increased whereas the number of aldehydic lesions
was significantly decreased for Apex
þ
/-
animals under
normal vs. oxidative stress conditions. However,
compared to wt animals under oxidative stress, the
number of aldehydic lesions was significantly increased
while the number of 3'OH-containing single-strand
breaks was significantly decreased. This finding argues
that APE1 is not rate-limiting under these conditions
and that upregulation of other base excision repair
enzymes such as DNA polymerase beta is in fact rate-
limiting. Thus, base excision repair activity was then
monitored through the use of a G:U mismatch repair
assay for which APE1 activity is essential in liver
nuclear extracts for animals treated with 2-NP.
Although overall BER activity decreased in Apex
þ
/-
animals in response to 2-NP, it was actually higher
than in the wild-type litter mates. The authors attribute
this unexpected result to alterations in APE1's redox
activity in the Apex
þ
/-
animals resulting in increased
ATF/CREB DNA-binding activity, which upregulates
DNA polymerease beta expression. Oxidative stress
was shown to increase monofunctional (UNG) but not
bifunctional (OGG1) DNA glycosylase activity in
Apex
þ
/-
animals. Upregulation of both UNG and DNA
polymerase beta is consistent with the increased overall
BER activity observed in the Apex
þ
/-
animals. However,
lack of upreguation of OGG1 is expected to result in
increased apoptosis. Accordingly, effects on cell cycle
ROLE OF REDOX REGULATION
IN DNA REPAIR
A number of transcription factors regulate expression
of DNA response genes and are themselves redox-regu-
lated by APE1. These include NF-
k
B, p53, AP-1, and
HIF-1
a
. Of these, p53 and AP-1 have been shown to
regulate the expression of a large number of DNA repair
genes (
Figure 11.8
). Maintenance of genomic integrity
is promoted by p53 through regulation of some of
the DNA repair pathways.
122,123
Both transactivation-
dependent and -independent mechanisms are involved
in p53's regulation of DNA repair genes (reviewed
in
122
) and p53 is itself regulated by both redox-depen-
dent and independent mechanisms involving Ref-1
(APE1), as discussed above.
DNA repair genes involved in direct repair, homolo-
gous recombination, global genome repair, mismatch
repair, and base excision repair (BER) (reviewed in
9
)
are regulated by p53. In response to oxidative or alkyla-
tive damage to DNA, BER is initiated by DNA glycosy-
lases that cleave the damaged DNA base creating an AP
site. Regulation of the BER pathway by p53 involves
both activation and negative regulation of BER enzymes
under different conditions and in different cell lines. In
