Molecular Biology of the Cell by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morgan, Martin Raff, Keith Roberts, Peter Walter by by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morg

HOMOLOGOUS RECOMBINATION
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Figure 5–50 Repair of a broken replication fork by homologous
recombination. When a moving replication fork encounters a single-strand
break, it will collapse, but can be repaired by homologous recombination.
The process uses many of the same reactions shown in Figure 5–48 and
proceeds through the same basic steps. Green strands represent the new
DNA synthesis that takes place after the replication fork has broken. This
pathway allows the fork to move past the site that was nicked on the original
template by using the undamaged duplex as a template to synthesize DNA.
(Adapted from M.M. Cox, Proc. Natl Acad. Sci. USA 98:8173–8180, 2001.
With permission from National Academy of Sciences.)
replication
fork
DNA nick
MOVEMENT OF
REPLICATION FORK
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3′
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3′
some dangers to the cell as it sometimes “repairs” damage using the wrong bit
of the genome as the template. For example, sometimes a broken human chromosome
is “repaired” using the homolog from the other parent instead of the
sister chromatid as the template. Because maternal and paternal chromosomes
differ in DNA sequence at many positions along their lengths, this type of repair
can convert the sequence of the repaired DNA from the maternal to the paternal
sequence or vice versa. The result of this type of errant recombination is known as
loss of heterozygosity. It can have severe consequences if the homolog used for
repair contains a deleterious mutation, because the recombination event destroys
the “good” copy. Loss of heterozygosity, although rare, is a critical step in the formation
of many cancers (discussed in Chapter 20).
Cells go to great lengths to minimize the risk of mishaps of these types; indeed,
nearly every step of homologous recombination is carefully regulated. For example,
the first step, processing of the broken ends, is coordinated with the cell cycle:
the nuclease enzymes that carry out this process are activated (in part, by phosphorylation)
only in the S and G 2 phases of the cell cycle, when a daughter duplex
(either as a partially replicated chromosome or a fully replicated sister chromatid)
can serve as a template for repair (see Figure 5–50). The close proximity of the two
daughter chromosomes disfavors the use of other genome sequences in the repair
process.
The loading of RecA or Rad52 onto the processed DNA ends and the subsequent
strand-exchange reaction are also tightly controlled. Although these proteins
alone can carry out these steps in vitro, a series of accessory proteins, including
Rad52, is needed in eukaryotic cells to ensure that homologous recombination
is efficient and accurate (Figure 5–51). There are many such accessory proteins,
and exactly how they coordinate and control homologous recombination remains
a mystery. We do know that the enzymes that catalyze recombinational repair are
made at relatively high levels in eukaryotes and are dispersed throughout the
nucleus in an inactive form. In response to DNA damage, they rapidly converge
on the sites of DNA damage, become activated, and form “repair factories” where
many lesions are apparently brought together and repaired (Figure 5–52).
In Chapter 20, we shall see that both too much and too little homologous
recombination can lead to cancer in humans, the former through repair using the
“wrong” template (as described above) and the latter through an increased mutation
rate caused by inefficient DNA repair. Clearly, a delicate balance has evolved
that keeps this process in check on undamaged DNA, while still allowing it to act
efficiently and rapidly on DNA lesions as soon as they arise.
Not surprisingly, mutations in the components that carry out and regulate
homologous recombination are responsible for several inherited forms of cancer.
Two of these, the Brca1 and Brca2 proteins, were first discovered because
5′
REPLICATION FORK BREAKS
5′
3′
5′
3′
NUCLEASE DEGRADES
5′ END OF BROKEN STRAND
STRAND EXCHANGE
DNA SYNTHESIS
STRAND BREAKAGE
ADDITIONAL DNA
SYNTHESIS
REPLICATION FORK
RESTARTS
BLOCK TO REPLICATION OVERCOME
3′
5′
3′
5′
3′
5′
3′
5′
3′
Figure 5–51 Structure of a portion of the Rad52 protein. This doughnutshaped
structure is composed of 11 subunits. Single-strand DNA has been
modeled into the deep groove running along the protein surface. Rad52
helps load Rad51 onto single-strand DNA to form the nucleoprotein filament
that carries out strand exchange. Rad52 also acts later to re-form the double
helix and complete the homologous recombination reaction. (From
M.R. Singleton et al., Proc. Natl Acad. Sci. USA 99:13492–13497, 2002.
With permission from National Academy of Sciences.)
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