Biology Reference
In-Depth Information
CHAPTER
7
Targeting Homologous Recombination
Repair in Cancer
Henning Willers, Heike N. Pf
ยจ
ffle, Lee Zou
Massachusetts General Hospital, Boston, MA
MECHANISMS AND REGULATION
OF HOMOLOGOUS
RECOMBINATION REPAIR
pathways may be used in a context-specific manner.
During the cell cycle, the HRR pathway predominantly
functions in S and G2 phases, and it is particularly
important for genomic stability during DNA replication.
The process of HRR can be roughly divided into four
phases: (1) resection of DNA ends; (2) formation of
RAD51 filament; (3) strand invasion and Holliday junc-
tion (HJ) formation; (4) dissolution of HJs (
Figure 7.2
).
Depending on whether the second DSB end can be
captured in phase 3 and how the HJs are resolved by
the specialized endonucleases in phase 4, HR can
generate either crossover or non-crossover repair prod-
ucts. During the process of HRR, the DNA structures
generated at DSBs activate the ATM and ATR check-
point kinases. These kinases not only trigger the
signaling pathways that lead to cell cycle arrest, but
also locally phosphorylate numerous DNA repair
proteins at or around sites of DSBs. In the following
sections, we will describe the principal biochemical
events during HR, and the potential regulatory func-
tions of the ATM and ATR checkpoint kinases.
An Overview of Homologous Recombination
Repair (HRR)
A double-stranded DNA break (DSB) is one of the
most deleterious forms of DNA damage. If not repaired
properly, DSBs could lead to mutations, deletions, trans-
locations, and amplifications in the genome. In human
cells, DSBs are repaired by multiple repair pathways,
including HRR, non-homologous end joining (NHEJ),
alternative NHEJ (alt-NHEJ; also known as the microho-
mology-mediated end joining or MMEJ), and single-
strand annealing (SSA).
1
e
3
The NHEJ pathway, which
directly rejoins broken DNA ends in the absence of
DNA sequence homology, is an error-prone mechanism.
The HRR, SSA, and alt-NHEJ pathways are dependent
on homologous DNA sequences, and they require nucle-
olytic resection of DNA ends to generate single-stranded
DNA (ssDNA) (
Figure 7.1
). The alt-NHEJ pathway
requires only limited amount of ssDNA and sequence
homology, and is error-prone. The SSA pathway
involves more extensively resected DNA ends and typi-
cally operates at DSBs flanked by direct repeats. Repair
of DSBs by SSA leads to deletions of the sequences
between the direct repeats.
4
In contrast to NHEJ, alt-
NHEJ, and SSA, HRR is a pathway with high fidelity.
In this pathway, the ssDNA generated by nucleolytic
resection of DNA ends interacts with the RAD51 recom-
binase to form RAD51 filament (
Figure 7.2
). As
described below, RAD51 filament enables DNA ends
to search for homologous sequences in the genome
and use these sequences to repair DSBs. The different
DSB repair pathways may function in concert to maxi-
mize the efficiency of DSB repair. Alternatively, these
The Principal Events during HRR
Resection of DNA Ends
The budding yeast
Saccharomyces cerevisiae
has been
used as a model system to study the resection of DNA
ends.
5
Most, if not all, of the factors involved in resection
are conserved from yeast to humans (see
Figure 7.1
).
6
When DSBs are generated in yeast, they are directly
recognized by the Mre11-Rad50-Xrs2 (MRN) complex.
The MRX complex promotes resection through at least
two distinct mechanisms. In the first mechanism, MRX
helps to recruit the Top3 (topoisomerase 3)-Rmi1
complex to DNA ends.
7,8
The Top3-Rmi1 complex,
together with RPA, stimulates unwinding of DNA ends
