Biology Reference
In-Depth Information
CHAPTER
6
Targeting the Nucleotide Excision Repair
Pathway f
or Therapeutic A
pplications
John J. Turchi
*
, Steve M. Patrick
y
*
Indiana University School of Medicine, Indianapolis, IN
y
University of Toledo, Toledo, OH
OVER
VIEW OF THE NER PATH
WAY
and ERCC1-XPF, bind 3' and 5' of the damage, respec-
tively. Following the initial recognition, assembly of
a pre-incision complex precedes nuclease catalyzed inci-
sion of the damaged DNA strand 5' and 3' of the adduct.
Recent data suggest that the nuclease cleavage reactions
are coordinated by DNA polymerase extension with
ERCC1-XPF cleaving 5' of the lesion to generate a 3'
hydroxyl which can be extended by DNA polymerase
d
or
3
.
11
Extension proceeds in a traditional strand
displacement type activity until 3' cleavage by XPG
releases the damaged oligonucleotide. The final ligation
event necessary to create the intact duplex DNA origi-
nally demonstrated that DNA ligase I was responsible,
though more recently a role of DNA ligase III/ERCC3
has been described.
12,13
Biochemical pathway
NER is a highly conserved DNA repair pathway
employing an array of proteins and activities that enable
the removal and repair of a wide array of bulky DNA
adducts from genomic DNA. In lower eukaryotes, the
entire pathway can be accomplished with six proteins
while in higher eukaryotes, over 30 gene products are
required. The pathway can be divided into three essen-
tial steps, including recognition of the damage site, inci-
sion of a damaged oligonucleotide, and finally
resynthesis and ligation resulting in an intact, undam-
aged duplex DNA.
1,2
In addition to these essential
elements, numerous signaling and regulation events
occur often involving post-translational modifications
of NER and other proteins, including phosphorylation
and ubitiquination.
3
e
7
Although DNA damage-indepen-
dent incision has been reported, the NER pathway
achieves high specificity and low background DNA inci-
sion as a result of a stochastic process in recognition. To
allow preferential repair of coding DNA, recognition can
be coupled to transcription, termed transcription
coupled NER (TC-NER), while both transcribed and
non-transcribed DNA can be repaired via the global
genomic-NER (GG-NER) pathway.
8
e
10
Regardless of
which subpathway is activated, to accommodate the
differences in lesion structure and chemistry, eukaryotic
NER employs a complex recognition process involving
multiple gene products that assemble at the site of
damage prior to the introduction of any breaks in the
DNA phosphodiester backbone (
Figure 6.1
). The XPC/
Rad23B complex, DDB, XPA, RPA, and TFIIH engage
the DNA damage site with TFIIH, RPA, and XPA remain-
ing bound as the two structure-specific nucleases, XPG
Mutation in NER Proteins with Direct
Links to Three Human Diseases
Defects in the proteins involved in NER result in three
separate though inter-related autosomal recessive disor-
ders xeroderma pigmentosum (XP), Cocayne's
Syndrome (CS), and tricothoidystrophy (TTD).
14
The
association with NER with XP is probably the most
simple to reconcile as defects in any of the 7 XP comple-
mentation groups A-G result in the inability to repair
bulky adduct DNA damage such as the adducts caused
by exposure to UV-light. The increased UV sensitivity
can be directly attributed to the inability to repair UV
adducts which leads to a dramatic increased suscepti-
bility to UV induced skin cancers. In addition those
with XP also have an increased rate of other cancers
likely a result of the increased genetic instability result-
ing from the inability to catalyze NER. There is a fairly
large difference in UV sensitivity between the different
