DNA Repair 8 (2009) 1038–1046
Contents lists available at ScienceDirect
journal homepage: www.elsevier.com/locate/dnarepair
The checkpoint response to replication stress
Dana Branzeia,∗, Marco Foiania,b
aFIRC Institute of Molecular Oncology Foundation (IFOM-IEO campus), Via Adamello 16, 20139 Milan, Italy
bDipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano, Via Celoria 26, 20133 Milan, Italy
a r t i c l ei n f o
Available online 23 May 2009
a b s t r a c t
Genome instability is a hallmark of cancer cells, and defective DNA replication, repair and recombination
have been linked to its etiology. Increasing evidence suggests that proteins influencing S-phase processes
roles in maintaining genome stability. DNA damage and replication stress activate a signal transduction
cascade, often referred to as the checkpoint response. A central goal of the replication checkpoint is to
maintain the integrity of the replication forks while facilitating replication completion and DNA repair
and coordinating these events with cell cycle transitions. Progression through the cell cycle in spite of
defective or incomplete DNA synthesis or unrepaired DNA lesions may result in broken chromosomes,
replication checkpoint during replication and in response to replication stress, as well as the enzymatic
activities that cooperate with the checkpoint pathway to promote fork resumption and repair of DNA
lesions thereby contributing to genome integrity.
© 2009 Elsevier B.V. All rights reserved.
1. Checkpoint function, genome integrity and cancer
Inadequate responses to replication stress or defects in DNA
repair underlie many forms of cancer [1,2]. Multiple significant
links between checkpoint proteins and genome stability have been
identified in human and other mammalian organisms, as well
as in various model systems [3,4]. The checkpoint cascade, also
often referred to as the DNA-damage response, is highly conserved
in eukaryotic organisms. Although initially most of the research
was conducted in budding and fission yeast , the past years of
research have shown that homologs exist for all the components
discovered in yeast, although the pathway is more elaborated in
Collapsed forks or fragile zones are prone to lead to chro-
mosome rearrangements or translocations and a large number
of studies document on the effect of checkpoint mutations on
genome-wide and site-specific stability [1,3,4,7]. It is also known
that chromosomal instability leading to different types of chro-
mosome rearrangements as well as chromosome loss plays an
checkpoint mutations are often found in cancer, and many human
genetic syndromes that lead to cancer predisposition are caused
E-mail address: firstname.lastname@example.org (D. Branzei).
by mutations in genes that protect the genome integrity during
This linkage between replication and cancer underscores the
importance of understanding how cells cope with aberrant repli-
cation forks. In the following sections we discuss on the molecular
mechanisms employed by replication checkpoints to stabilize the
replication forks and to assist and coordinate different damage-
tolerance mechanisms that contribute to repair and chromosome
integrity (Fig. 1).
2. The replication checkpoint cascade
The replication checkpoint is a sensor-response system acti-
vated by impeded replication forks or other types of lesions that
occur in S-phase, and is crucial for stabilizing replication forks and
fragile sites [8,9]. There are different types of independent molec-
ular complexes that sense and signal different types of damage, of
which the RPA-coated single stranded (ss) DNA is a central player,
although not always responsible or sufficient to activate the repli-
cation checkpoint [6,10]. Other checkpoint factors, such as TopB1,
the Mre11-Rad50-Xrs2 (MRX) complex in yeast (the Mre11-Rad50-
Nbs1 (MRN) complex in mammals), and the 9-1-1 complex have
also been implicated in the activation of the replication checkpoint
or the recruitment of sensor kinases to stalled forks [6,11] (Fig. 2).
and ATR, Tel1 and Mec1 in yeast, respectively. Tel1/ATM responds
mainly to double strand breaks (DSBs) whereas Mec1/ATR is
activated by ssDNA and stalled forks [11,12]. The Mec1/ATR path-
1568-7864/$ – see front matter © 2009 Elsevier B.V. All rights reserved.
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
Fig. 1. General scheme of the checkpoint responses to replication stress and the
primary impact of these functions on genomic stability.
way is the keystone of the replication checkpoint and a simplified
view of this transduction cascade is shown in Fig. 2.
or by the action of the MCM replicative helicases that may con-
tinue the unwinding ahead of the stalled replication fork [13,14].
Once formed, the ssDNA is bound by RPA, and the ssDNA–RPA
complex plays two critical roles in recruiting independently Mec1-
(Fig. 2). Physical interactions between RPA and checkpoint proteins
have been demonstrated and shown to be required for checkpoint
similarity to the large subunit of replication factor C (RFC), Rfc1,
interacts with the four smaller subunits of RFC, and this RFC-like
clamp loader complex is responsible for loading the PCNA-related
in mammals onto DNA . Although the order of the subsequent
events is not entirely clear, it is believed that co-localization of
Mec1/ATR and of the 9-1-1 complex allows interaction between
these proteins at damage sites and subsequent phosphorylation of
the 9-1-1 complex by ATR. Another important player in ATR acti-
vation appears to be Dpb11 in yeast and its functional ortholog
replication checkpoint in S-phase. Single stranded DNA coated with RPA triggers the
recruitment of Mec1-Ddc2 together with the one of Rad24 and of the checkpoint
clamp composed of Rad17-Mec3-Ddc1. Mrc1, Tof1, and Dpb11 are also associated
with replication forks. Interaction and phosphorylation events occurring between
Rad17-Mec3-Ddc1, Dpb11 and Mec1-Ddc2 lead to Mec1 activation. Subsequently,
Mrc1 and Rad9 function as adaptors to promote Rad53 hyperphosphorylation to
elicit a cell cycle response. Mec1 and Rad9 also promote Chk1 phosphorylation.
Mec1, together with Rad53 and to some extent also Chk1, mediate the response to
replication stress by promoting fork stabilization, fork processing and DNA repair
binds the 9-1-1 complex and is critical for Mec1/ATR activation
[17–19]. Following this step, additional Mec1/ATR substrates called
mediators are recruited and play an important role in amplify-
ing the checkpoint signal throughout the cell. In mammalian cells
this pathway is quite elaborated and several aspects are not well
understood. Here we will mainly focus on the checkpoint signal
amplification and the mediators in budding yeast (Fig. 2). The best-
characterized mediator in S. cerevisiae is Rad9, which functions as
an adaptor between Mec1 and the Rad53 checkpoint kinase .
Rad53 (Chk2 in mammals although its functional ortholog is Chk1)
plays pivotal roles in response to replication stress, and its phos-
phorylation is essential for checkpoint control [21,22]. Rad53 is
phosphorylated in a Mec1-dependent fashion in response to both
DNA damage and incomplete replication. Rad9 functions predom-
inantly in the G1/S and the G2/M transitions of the DNA damage
checkpoints, and only to some extent in response to replication
fork arrest  (Fig. 2). Rad9 is recruited to DSBs and hyperphos-
phorylated in a Mec1 and Tel1-dependent manner. It is proposed
that Rad9 catalyzes the activation of Rad53 by acting as a scaf-
fold to promote Rad53 autophosphorylation [24–26] (Fig. 2). In
response to replication stress, however, amplification of the repli-
cation checkpoint signal depends on Mrc1  (Fig. 2). Mec1
phosphorylates Mrc1 on multiple sites and mutation of these sites
in the mrc1AQmutant suppresses Rad53 hyperphosphorylation in
response to replication stress . However, experiments have so
far failed to demonstrate a physical interaction between Mrc1 and
Rad53 and thus the molecular mechanism through which Mrc1
activates Rad53 has yet to be characterized. The mammalian coun-
terpart of Mrc1, Claspin, interacts with the effector kinase Chk1
(the functional ortholog of Rad53) and is essential for its hyper-
phosphorylation [28–30]. The molecular mechanism of how the
checkpoint signal is amplified in mammalian cells is complex, and
it appears that different signals trigger the formation of alternative
Claspin–Chk1 complexes [6,10,30,31].
out the adaptor function between Mec1 and Rad53  (Fig. 2).
However, in the absence of Mrc1, the checkpoint mediator Rad9
can partially substitute for Mrc1 and promote Rad53 hyperphos-
phorylation, although likely leading to the formation of different
phosphoisoforms having distinct molecular functions [23,30]. Fol-
lowing autophosphorylation, Rad53 is released from the Rad9 (and
likely the Mrc1) complex, leading to an amplification of the check-
point signal [21,24]. Mec1 activates not only Rad53, but together
with Rad9, another checkpoint kinase, Chk1 [7,22]. Chk1 medi-
ates, in cooperation with Rad53, the G2/M checkpoint arrest ,
but recent reports have shown that Chk1 may also influence the
Before discussing the mechanism through which the replica-
tion checkpoint acts to stabilize replication forks, we like to note
that a number of different situations causing lesions or replica-
tion stress do not lead to formation of ssDNA and do not activate
and camptothecin (CPT) treatment as well as fork pausing at nat-
ural pause sites such as the rDNA replication fork barrier [10,14].
In such situations, the response to the stress factor or the repair of
the initial lesions occurs in a checkpoint-independent fashion or
requires additional proteins to activate the replication checkpoint.
3. Causes of fork stalling and general mechanisms through
which the replication checkpoint responds to replication
Replication fork progression is often impeded by exogenous
or endogenous DNA damage . In addition to lesion-induced
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
icantly when encountering tightly bound proteins, replication slow
zones, repetitive sequences, or aberrant DNA structures. Replica-
tion fork pausing can be experimentally induced by HU treatment,
which limits the dNTP pools, or by intra-S damage such as the
one caused by the methylating agent methyl methane sulphonate
(MMS). In most cases, the replication forks stall with a fully assem-
bled replisome but collapse occurs if the replisome dissociates. The
collapse is often induced by DNA breaks, at telomeres, or when
the pausing is prolonged, for instance by inefficient removal of
protein–DNA complexes [14,33,34]. Specialized proteins, such as
resumption by removing the impediments at specific replication
pausing sites [35,36].
The replication and damage checkpoints play crucial roles in
replication fork stabilization. Thus, while wild-type cells remain
competent to resume replication following a transient HU block
[37–39] and complete replication through alkylated DNA [38,40],
rad53 or mec1 mutants are unable to resume replication after HU
removal or to complete replication through damaged DNA, and
show lethality in the presence of replication stress [39,40]. The
replication checkpoint was proposed to mediate the response to
replication stress by promoting repair and facilitating replication
(Fig. 1). These functions are carried out by affecting the transcrip-
tion of repair genes and the pool of dNTPs [5,41], by regulating late
origin firing and fork rate [38,42] and by promoting fork stabiliza-
tion and DNA repair . Of these, fork stabilization appears as the
critical and most important role of Mec1 and Rad53 in response
to intra-S DNA damage, but all the functions mentioned above are
discussed and elaborated below.
4. The checkpoint effect on transcription
Following DNA damage or a block to DNA replication, Rad53,
together with Mec1, is required for the phosphorylation of Dun1,
a protein kinase that controls several DNA DNA damage-inducible
genes as well as genes encoding RNR (ribonucleotide reductase)
subunits involved in the modulation of dNTP pools . Although
the transcriptional response to DNA damage appears to share com-
ponents with the replication and damage checkpoint (Fig. 3), the
two pathways are different, and Dun1 is not a checkpoint protein
per se. The mechanism employed by Dun1 to induce transcription
and regulate dNTP pools is not yet fully understood but involves
inhibition of two key proteins, Crt1 and Sml1 [7,22,41,43] (Fig. 3).
Crt1 acts together with Ssn6 and Tup1 to repress gene transcription
in normally replicating cells . Crt1 phosphorylation by Mec1,
Rad53 and Dun1 causes it to dissociate from the “X box” sequence
to which it binds and which is present upstream of a number of
yeast repair genes . The dissociation of Crt1 from the promot-
ers leads to the transcription derepression of these genes (Fig. 3).
On the other hand, Sml1 binds and inhibits Rnr1, and therefore lim-
its the available dNTP pools . Dun1-mediated phosphorylation
of Sml1 induces Sml1 degradation , thus leading to activation
of Rnr1, which increases the concentration of dNTP pools neces-
sary for efficient DNA replication and repair [41,44]. The lethality
of mec1 and rad53 cells is rescued by SML1 deletion, although the
suggest that the lethality of null mec1 and rad53 mutants is due to
DNA replication without adequate accumulation of dNTP precur-
sors. Other phenotypes of mec1 and rad53 mutants have also been
formation, but these phenotypes are suppressed by the SML1 dele-
tion . However, the finding that blocking protein synthesis in
wild-type cells during S phase does not render them sensitive to
HU and does not prevent replication fork resumption following a
Fig. 3. Schematic representation of the checkpoint response mediating transcrip-
tion induction of repair genes. Following DNA damage or a block to DNA replication,
Mec1, Rad53, and Dun1 are required to phosphorylate Crt1, which leads to a tran-
sequence, to which Crt1 in its unphosphorylated state normally bind to repress their
transient HU block suggests that the effect of the checkpoint on
inducing transcription is not critical for fork stabilization or cell
5. The role of the checkpoint in regulating fork rate and
late origin firing
Mec1 and Rad53 regulate late origin firing during normal replica-
tion or in response to replication stress [40,46,47]. A recent work
has shown that rad53-1 and mec1-1 cells show only about 40% of
overlap for both early- and late-firing origins or for the early origins
of replication resembling fragile sites . Interestingly, although
mec1 null mutants are considerably more sensitive to HU and MMS
mutants [40,49], rad53-1 cells activate many more late-firing ori-
the mec1 and rad53 alleles studied, but nevertheless they suggest
that Mec1 and Rad53 control fork progression through both com-
mon and independent mechanisms. Considering that Mec1 targets
both Rad53 and Chk1 (Fig. 2), and that Rad53 regulates Cdc7/Dbf4
activity and chromatin structure independently of Mec1 [50–53],
the non-redundancy of Mec1 and Rad53 in regulating origin firing
becomes a concept easy to grasp. The role of late origin firing in
promoting the viability of cells experiencing HU-induced replica-
tion stress is also arguable as a mec1 mutant, mec1-100, that does
not block late origin firing in HU but can stabilize replication forks,
is much less sensitive to HU than mec1 null mutants . However,
we note that since this study  did not analyze origin activation
at a genome-wide scale, it is formally possible that many, although
not all late origins were still inhibited in the mec1-100 mutant.
In contrast to the results suggesting an active role for the check-
point in inhibiting firing of late origins during exposure to HU or
to MMS [40,46,47], a recent study found that in yeast cells the ori-
gins were activated in the same order in the presence or absence of
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
HU, except that the time scale was different in the two cases, con-
sistent with a general reduction of the fork rate by HU . Thus,
this study implies that the late-firing origins are not preferentially
inhibited in the presence of HU. The picture emerging suggests that
checkpoint mutants affect the temporal program of origin firing
such that the late origins fire early, at the same time as early ori-
gins. The mechanism for this is not understood but one possibility
suggested by a recent study is that in checkpoint mutants treated
with HU, late/dormant origin firing occurs as a consequence of the
collapse of replication forks arising from early replicons .
Regulation of origin firing could in principle affect also fork rate,
and it has been proposed that one function of the intra-S check-
point is to slow down replication fork progression in the presence
of DNA damage . However, in such experiments it is important
to distinguish between the passive effect of DNA lesions that act as
checkpoints slowing replication fork progression. In budding yeast,
DNA combing experiments showed that fork rates are normal in
rad53-11 mutants, suggesting that the Mec1/Rad53 pathway may
not generally regulate fork progression . Interestingly, Mrc1
is required for normal fork progression and promotes replication
fork stability at regions containing inverted repeats that fold into
hairpin-like DNA structures . Considering also that Mrc1 and
Rad53 are required for the maintenance of artificial trinucleotide
repeats inserted in the yeast genome , one possibility is that
Mrc1 and Rad53 are especially required for normal fork rates at
regions that are intrinsically difficult to replicate. In agreement
with this view, chicken DT40 cells mutated for Chk1 (the func-
, and Chk1 mutants also show low levels of the Mrc1 ortholog,
Claspin , which is also required for normal fork rates ; fur-
thermore the human genome is much more complex than the one
of yeast being enriched in repetitive sequences or regions that are
difficult to replicate.
6. Role of the checkpoint in promoting replication fork
An important function of the replication checkpoint is to main-
tain the integrity of existing replication forks. The molecular
mechanism through which checkpoints regulate replication forks
is not yet clear, but chromatin immunoprecipitation (ChIP) exper-
iments have indicated that S. cerevisiae mec1 and rad53 mutants
are defective in replisome stabilization [63,64]. That is, when repli-
cation checkpoint mutants are exposed to replication stress, for
instance by treatment with HU, the replisome dissociates from the
stalled replication fork [63,64] (Fig. 4). This breakdown of the repli-
some is believed to trigger the nascent DNA chains to engage in
pathological transitions [39,65]. Consistent with this view, long
patches of ssDNA accumulate in checkpoint mutants treated with
HU, presumably because of DNA degradation [65,66] (Fig. 4). Fur-
(2D) gel electrophoresis of replicating DNA, it has been shown
that one unusual type DNA structure represented by four-branched
cells but accumulates to about 8% of the replication intermediates
in rad53 cells, is [39,65] (Fig. 4). These structures are recombino-
genic and can be further subjected to nuclease-mediated cleavage,
giving rise to double strand breaks (DSBs) or long stretches of
ssDNA regions representing gapped or hemireplicated molecules
[39,65,67] (Fig. 4).
In regard to the enzymatic activities that mediate processing
of the replication forks from which the replisome has dissociated,
studies have shown that Exo1 plays an important role . Exo1
was demonstrated to counteract reversed fork formation, as in the
absence of Exo1 that is, in rad53 exo1 mutants, a much larger accu-
mulation of reversed forks is observed as compared to the one of
rad53 single mutants [65,67] (Fig. 4). However, an alternate but not
exclusive explanation is that in replication checkpoint mutants the
lagging strand apparatus may be mis-regulated [65,68], leading to
defects in lagging strand synthesis, and thus to a larger accumula-
. A recent study showed that EXO1 deletion can suppress the
a primary target of Rad53 and that Exo1 phosphorylation might act
to restrain Exo1-dependent replication fork breakdown or resec-
tion events at the forks . In line with this view, a recent study
indicated that Rad53-dependent phosphorylation of Exo1 may act
to limit ssDNA accumulation and simultaneously act as a feedback
loop to restrain DNA damage checkpoint activation . The effect
Fig. 4. Transitions at stalled replication forks. In the absence of a functional replication checkpoint, the replisome dissociates from the stalled forks, and the forks degenerate
by experiencing fork reversal and processing, at least in part mediated by Exo1. Reversed forks can be also subjects to other nucleolytic events leading to double strand break
formation. Alternatively, specialized enzymes may promote the regression of the reversed forks and replication resumption.
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
of exo1 mutation in suppressing rad53 replication defects appears
to be specific for intra-S damage (MMS) and is not observed when
The answer of how the reversed forks accumulating in replica-
tion checkpoint mutants treated with HU are formed in the first
place is not yet clear but several explanations are possible. One
hypothesis is that the sister chromatid junctions resembling hemi-
deprived of replisome, as it has been shown to be the case of rad53
mutants . Another mechanism envisages a topology-mediated
transition, as the superhelical strain building up at the replication
fork may cause the formation of four-way junctions at replication
of replisome are annealed to form four-way junctions by the activ-
ity of a specialized enzyme. Recently, the repair protein Rad5 was
shown in vitro to possess helicase activity and to act by reannealing
the nascent chains of a stalled fork, thereby giving rise to four-way
known. Once reversed forks are formed, they need to be regressed
after repair synthesis, in order to prevent abnormal and aberrant
recombination events that may endanger genome stability (Fig. 4).
The factors promoting this transition remain to be uncovered.
If not appropriately resolved, the structures arising at stalled
replication forks or in attempts to restart replication following
encounter with DNA lesions, may trigger inappropriate recombina-
checkpoint, and reversed fork formation prevented [39,65]. If we
hypothesize that to some extent fork reversal may be a physiolog-
ical process mediated for instance by the Rad5 helicase , then
regression of these structures should subsequently occur in order
to prevent their cleavage by nucleases and to complete replication
(Fig. 4). RecQ helicases have also been proposed to promote regres-
sion of reversed forks based on their biochemical properties ,
but there is yet no evidence for accumulation of reversed forks in
In addition to Exo1 [32,67], several other nucleases may be impli-
cated in processing reversed forks, but their nature remain to be
elucidated; favorite candidates for such functions remain the Sae2
and the Mre11 complex implicated in early steps of recombination
repair and replication [55,67,75,76] as well as Mus81/Mms4 and
Slx1/Slx4 nucleases that play important functions in recombina-
tion repair and genome stability, and whose activity appears to be
regulated by the replication checkpoint [77,78].
7. Checkpoints facilitate replication in the presence of DNA
Although the S-phase checkpoint is activated both by stalled
forks and by DNA lesions, its role in regulating the processing of
replication forks appears to be different in these two situations.
Thus, while HU-treated rad53 cells accumulate by 2D gel analysis
reversed forks [39,65], no pathological structures are detected dur-
ing replication in the presence of DNA lesions induced by MMS
or UV treatment. Nevertheless, EM analysis of replicating DNA
from UV-irradiated cells revealed that rad53 mutants contained a
high fraction of large gapped forks . Since checkpoint mutants
are unable to resume replication following treatment with either
HU or DNA-damaging agents [38–40], it follows that checkpoints
also play an important role in promoting completion of chromo-
some replication in the presence of, or following, genotoxic stress.
Recent evidence has shown that Mec1 and Rad53 have separable
roles in promoting replication through alkylated DNA, and that
Rad53 regulation of Exo1 is crucial to facilitate replication in the
presence of damaged DNA . Exo1 is phosphorylated in vivo in
response to MMS treatment , but whether Exo1 phosphory-
lation directly influences the stability of replication forks remains
unknown. Recently, it has been shown that in addition to Mec1 and
Rad53, Chk1 can also promote replication through damaged DNA
. Consistent with the data obtained in yeast, a role for ATM/ATR
in restart of damaged replication forks was also concluded from
studies of replicating chromosomes in Xenopus laevis egg extracts
8. The checkpoint pathway and intra-S repair
generated by repriming downstream of the lesion on both leading
and lagging strands . These gaps can then be filled-in by means
of two different mechanisms. One pathway uses a combination of
replicative and translesion synthesis (TLS) polymerases to replicate
across the lesion, and the outcome can sometimes be error-prone.
The other gap-filling mechanism, referred to as template switch
(TS), is essentially error-free and uses the undamaged information
of the sister duplex to bypass the damage  (Fig. 5). TS utilizes a
mechanism that shares similarities with homologous recombina-
tion (HR) [83,84] (Fig. 5). Genetically, error-free mediated damage
Rad5-Mms2 and the other one mediated by members belonging
to the Rad51-Rad52 HR pathway . The TS process is thought to
give rise to transient, hemicatenane-like or pseudo-double Holli-
[74,85,86], together with Ubc9- and Mms21-dependent sumoyla-
tion  (Fig. 5). Recently, it has been shown that both Rad18-Rad5
and the Rad51 pathway promote the formation of sister chromatid
junctions (SCJs) that have the biochemical properties of pseudo-
double HJs at damaged replication forks . Rad18 and Rad51
appear to cooperate to promote TS, and the Rad18 functionality in
TS requires previous sumoylation of PCNA  (Fig. 5).
Most factors of the RAD18 PRR pathway have ubiquitin con-
jugating or ligating activities [89–91] and an important target of
the Rad18 pathway in DNA repair is PCNA . In response to
DNA-damage, PCNA is mono- or polyubiquitinated at the highly
conserved lysine residue K164 . Monoubiquitination of PCNA
depends on Rad6 and Rad18  and promotes translesion synthe-
sis [93,94], whereas PCNA modification through Lys (K)-63-linked
polyubiquitin chains requires Ubc13-Mms2 and Rad5 functional-
ities  and is required for error-free PRR [92,95,96] and for
TS-mediated by SCJ formation .
The damage and replication checkpoints have multiple connec-
tions with both the TS and the TLS pathways of gap-filling, and
phosphorylation events mediated by the checkpoint kinases are
likely to control the choice of the repair pathway. Genetic analysis
has suggested that the DNA damage checkpoint functions in PRR
and likely promotes both error-free and error-prone branches of
PRR  (Fig. 5). The DNA damage checkpoint, 9-1-1, was shown to
facilitate the recruitment and damage bypass mediated by Pol zeta
(Pol?) in budding yeast and DinB in fission yeast [98,99] (Fig. 5). In
budding yeast, deletion of DNA damage checkpoint genes reduces
damage-induced mutagenesis [97,100], and the mms2 spontaneous
and damage-induced mutagenesis is partially dependent on the
RAD9 gene .
The checkpoint also influences the error-free PRR. In fission
yeast, phosphorylation of Rad9 of the 9-1-1 complex by the ATM
and ATR-related Rad3 checkpoint kinase at Thr225 located within
the PCNA like domain, is enhanced by DNA damage and promotes
the physical interaction between Rad9 and the post-replication
repair protein Mms2, contributing to gap-filling in an error-free
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
Fig. 5. Schematic representation of processes that mediate gap-filling repair during replication. Gaps generated during replication by repriming downstream of the lesion
can be filled-in by using specialized translesion synthesis (TLS) polymerases or the newly synthesized strand as a template in a process referred to as template switch.
The checkpoint clamp 9-1-1 or Rad17-Mec3-Ddc1 promotes TLS-induced mutagenesis in the presence of DNA damage. Two mechanisms can contribute to template switch
events, one requiring the joint action of the Rad18-Rad5-Mms2 post-replication repair pathway together with the one of homologous recombination, while the other one is
independent of Rad18 but requires homologous recombination. The choice of these two template switch pathways is regulated by Siz1-dependent and PCNA sumoylation. In
fission yeast, Rad9 phosphorylation at Thr225 by Rad3 also promotes Mms2-dependent repair while inhibiting mutagenesis and inappropriate recombination. The pseudo-
double Holliday Junction intermediates generated by template switch events are resolved by the action of Sgs1 and Top3, whose functionality is modulated by Ubc9 and
Mms21 dependent sumoylation.
as well as hyperrecombination events are induced  (Fig. 5).
on Thr225 functions to direct repair through a Pli1 (Siz1)-mediated
sumoylation pathway into the error-free branch of PRR . This
model is in agreement with other genetic data obtained in budding
yeast where lack of PCNA sumoylation was shown to suppress the
damage sensitivity of error-free PRR mutants [87,102,103] and with
the finding that, in budding yeast, the ability of the Rad18-Mms2
pathway to promote gap-filling by means of SCJs requires Siz1-
dependent sumoylation . In the absence of Siz1-dependent and
PCNA sumoylation, TS occurs solely by means of HR and inde-
pendently of the Rad18 pathway  (Fig. 5). In fission yeast,
Thr225-dependent Rad9 phosphorylation prevents inappropriate
Rad51-dependent recombination, and these recombination events
might be related to the ones promoting SCJ formation in bud-
ding yeast but occurring independently of Rad18-Mms2 in the
absence of sumoylation . Whether similar to the fission yeast
situation, the Rad18 error-free pathway in other eukaryotes is
controlled by phosphorylation is not known, but it is a likely pos-
sibility considering Rad18 interaction with RPA , the finding
that Rad18 is an ATR-ATM target , and that MMS-induced
accumulation of human Rad18 at stalled forks involves protein
phosphorylation that is inhibited by wortmannin treatment and
which, therefore, may be performed by S-phase checkpoint kinases
. Genetic evidence in yeast suggests that Srs2 together with
sumoylated PCNA might provide the activity that promotes the
DNA repair function of Rad18-Rad5 [102,103,107,108] and likely
the TS events by means of SCJs at damaged replication forks
. Since Srs2 is a checkpoint target , it will be of interest
to address whether the checkpoint-mediated phosphorylation of
Srs2 plays any role in coordinating these repair events at repli-
cation forks. In addition to the possibility of checkpoint kinases
regulating the functionality of PRR, it is also plausible that Rad18-
repair. In regard to this, Rad18 was recently reported to promote
ubiquitination of Rad17 , but whether this event influences
the gap-filling ability of cells remains to be addressed by future
Recombination is also used to restart replication forks and
to promote repair of replication-associated lesions, but the role
of checkpoint proteins in controlling recombination is complex
. Recombination plays a major role in promoting replication of
telomeric and subtelomeric regions and in the repair of double-
strand breaks (DSBs) generated during replication . DSB repair
by gene conversion or break induced replication is thought to be
a late S or G2 event, and is under the control of cyclin depen-
dent kinases, Cdks [11,112]. CDK activity influences HR through
phosphorylation of several repair and checkpoint proteins .
Damage checkpoints are required for damage-induced recombi-
nation , but checkpoint-dependent phosphorylation of repair
proteins acts differentially to either inhibit or activate recombina-
tion repair events (reviewed in ref. ). In a recent study it has
been shown that Tel1 (the ATM ortholog), together with Sae2 and
tures when the fork encounters the DSB site . Thus, Tel1/ATM
and the MRX/MRN complex control the integrity of forks encoun-
tering a DSB site and the cruciform intermediates formed in the
absence of these pathways may also influence the formation of
sister telomere fusions in ATM- or MRN-deficient cells [55,114].
Finally, all the repair events mentioned above are likely to be
DNA lesion and the cellular context of where the repair events take
place . Although the mechanisms are very intricate and many
aspects are still to be detailed, the checkpoint-mediated control of
the cell cycle.
9. Concluding remarks
The past years have brought significant contributions in under-
standing the mechanisms through which checkpoints contribute
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
to replication fork stabilization or DNA repair following replication
stress. Different factors and regulatory pathways are intertwined
in various processes that occur at replication forks. Identification
of genetic links and substrates has proved useful for our under-
standing of the mechanisms through which checkpoint signaling
translates into increased replication fork stability and regulation of
the increasing evidence that suggests a link between cancer and
the chromosomal abnormalities caused by faulty DNA replication,
understanding the mechanisms that process the stalled replication
forks and promote DNA repair will continue to be an increasing
focus of research.
Conflict of interest
The work in authors’ laboratories is supported by grants from
the Associazione Italiana per la Ricerca sul Cancro, Association for
International Cancer Research to D.B. and M.F., and by European
Community DNA Repair and GENICA grants, Telethon, MIUR, and
Ministry of Health to M.F.
 M.B. Kastan, J. Bartek, Cell-cycle checkpoints and cancer, Nature 432 (2004)
 B. Vogelstein, K.W. Kinzler, Cancer genes and the pathways they control, Nat.
Med. 10 (2004) 789–799.
 R.D. Kolodner, C.D. Putnam, K. Myung, Maintenance of genome stability in
Saccharomyces cerevisiae, Science 297 (2002) 552–557.
 B.B. Zhou, S.J. Elledge, The DNA damage response: putting checkpoints in per-
spective, Nature 408 (2000) 433–439.
 S.J. Elledge, Cell cycle checkpoints: preventing an identity crisis, Science 274
 J.W. Harper, S.J. Elledge, The DNA damage response: ten years after, Mol. Cell
28 (2007) 739–745.
 K.A. Nyberg, R.J. Michelson, C.W. Putnam, T.A. Weinert, Toward maintaining
the genome: DNA damage and replication checkpoints, Annu. Rev. Genet. 36
 D. Branzei, M. Foiani, Interplay of replication checkpoints and repair proteins
at stalled replication forks, DNA Repair (Amst.) 6 (2007) 994–1003.
Cell 111 (2002) 779–789.
 H. Tourriere, P. Pasero, Maintenance of fork integrity at damaged DNA and
natural pause sites, DNA Repair (Amst.) 6 (2007) 900–913.
 D. Branzei, M. Foiani, Regulation of DNA repair throughout the cell cycle, Nat.
Rev. Mol. Cell Biol. 9 (2008) 297–308.
 J. Bartek, J. Lukas, DNA damage checkpoints: from initiation to recovery or
adaptation, Curr. Opin. Cell Biol. 19 (2007) 238–245.
 T.S. Byun, M. Pacek, M.C. Yee, J.C. Walter, K.A. Cimprich, Functional uncoupling
of MCM helicase and DNA polymerase activities activates the ATR-dependent
checkpoint, Genes Dev. 19 (2005) 1040–1052.
 D. Branzei, M. Foiani, The DNA damage response during DNA replication, Curr.
Opin. Cell Biol. 17 (2005) 568–575.
 X. Xu, S. Vaithiyalingam, G.G. Glick, D.A. Mordes, W.J. Chazin, D. Cortez, The
basic cleft of RPA70N binds multiple checkpoint proteins, including RAD9, to
regulate ATR signaling, Mol. Cell Biol. 28 (2008) 7345–7353.
 J. Majka, P.M. Burgers, Yeast Rad17/Mec3/Ddc1: a sliding clamp for the DNA
damage checkpoint, Proc. Natl. Acad. Sci. U. S. A. 100 (2003) 2249–2254.
 A. Kumagai, J. Lee, H.Y. Yoo, W.G. Dunphy, TopBP1 activates the ATR-ATRIP
complex, Cell 124 (2006) 943–955.
 D.A. Mordes, E.A. Nam, D. Cortez, Dpb11 activates the Mec1-Ddc2 complex,
Proc. Natl. Acad. Sci. U. S. A. 105 (2008) 18730–18734.
 J. Majka, P.M. Burgers, Clamping the Mec1/ATR checkpoint kinase into action,
Cell Cycle 6 (2007) 1157–1160.
 L. Zou, D. Cortez, S.J. Elledge, Regulation of ATR substrate selection by Rad17-
dependent loading of Rad9 complexes onto chromatin, Genes Dev. 16 (2002)
 A. Pellicioli, M. Foiani, Signal transduction: how Rad53 kinase is activated,
Curr. Biol. 15 (2005) R769–771.
 D. Branzei, M. Foiani, The Rad53 signal transduction pathway: replication fork
 A.A. Alcasabas, A.J. Osborn, J. Bachant, F. Hu, P.J. Werler, K. Bousset, K. Furuya,
J.F. Diffley, A.M. Carr, S.J. Elledge, Mrc1 transduces signals of DNA replication
stress to activate Rad53, Nat. Cell Biol. 3 (2001) 958–965.
Rad53 activating machine, Mol. Cell 8 (2001) 129–136.
 F.D. Sweeney, F. Yang, A. Chi, J. Shabanowitz, D.F. Hunt, D. Durocher, Saccha-
Biol. 15 (2005) 1364–1375.
 G.W. Toh, N.F. Lowndes, Role of the Saccharomyces cerevisiae Rad9 protein
in sensing and responding to DNA damage, Biochem. Soc. Trans. 31 (2003)
 A.J. Osborn, S.J. Elledge, Mrc1 is a replication fork component whose phospho-
rylation in response to DNA replication stress activates Rad53, Genes Dev. 17
 A. Kumagai, W.G. Dunphy, Claspin, a novel protein required for the activation
Mol. Cell 6 (2000) 839–849.
the regulated binding of Chk1, Nat. Cell Biol. 5 (2003) 161–165.
20 (2006) 772–783.
 C.C. Chini, J. Chen, Repeated phosphopeptide motifs in human Claspin are
phosphorylated by Chk1 and mediate Claspin function, J. Biol. Chem. 281
kinases in stabilizing DNA replication forks, Genes Dev. 22 (2008) 1816–1827.
 S. Lambert, A. Watson, D.M. Sheedy, B. Martin, A.M. Carr, Gross chromoso-
mal rearrangements and elevated recombination at an inducible site-specific
replication fork barrier, Cell 121 (2005) 689–702.
site promotes recombination in fission yeast, EMBO J. 24 (2005) 2011–2023.
helicase promotes genome integrity by preventing replication fork stalling:
viability of rrm3 cells requires the intra-S-phase checkpoint and fork restart
activities, Mol. Cell Biol. 24 (2004) 3198–3212.
 K.H. Schmidt, R.D. Kolodner, Requirement of Rrm3 helicase for repair of spon-
taneous DNA lesions in cells lacking Srs2 or Sgs1 helicase, Mol. Cell. Biol. 24
 B.A. Desany, A.A. Alcasabas, J.B. Bachant, S.J. Elledge, Recovery from DNA repli-
cational stress is the essential function of the S-phase checkpoint pathway,
Genes Dev. 12 (1998) 2956–2970.
 J.A. Tercero, M.P. Longhese, J.F. Diffley, A central role for DNA replication forks
in checkpoint activation and response, Mol. Cell 11 (2003) 1323–1336.
C.S. Newlon, M. Foiani, The DNA replication checkpoint response stabilizes
stalled replication forks, Nature 412 (2001) 557–561.
damaged DNA by the Mec1/Rad53 checkpoint, Nature 412 (2001) 553–557.
 X. Zhao, R. Rothstein, The Dun1 checkpoint kinase phosphorylates and regu-
lates the ribonucleotide reductase inhibitor Sml1, Proc. Natl. Acad. Sci. U. S. A.
99 (2002) 3746–3751.
promote replication fork progression and recovery independently of Rad53,
Mol. Cell 19 (2005) 699–706.
 M. Huang, Z. Zhou, S.J. Elledge, The DNA replication and damage checkpoint
pathways induce transcription by inhibition of the Crt1 repressor, Cell 94
 X. Zhao, E.G. Muller, R. Rothstein, A suppressor of two essential checkpoint
genes identifies a novel protein that negatively affects dNTP pools, Mol. Cell 2
 R.S. Cha, N. Kleckner, ATR homolog Mec1 promotes fork progression,
thus averting breaks in replication slow zones, Science 297 (2002) 602–
 C. Santocanale, J.F.X. Diffley, A Mec1- and Rad53-dependent checkpoint con-
trols late-firing origins of DNA replication, Nature 395 (1998) 615–618.
 K. Shirahige, Y. Hori, K. Shiraishi, M. Yamashita, K. Takahashi, C. Obuse, T. Tsu-
rimoto, H. Yoshikawa, Regulation of DNA-replication origins during cell-cycle
progression, Nature 395 (1998) 618–621.
 M. Raveendranathan, S. Chattopadhyay, Y.T. Bolon, J. Haworth, D.J. Clarke, A.K.
Bielinsky, Genome-wide replication profiles of S-phase checkpoint mutants
reveal fragile sites in yeast, EMBO J. 25 (2006) 3627–3639.
 R. Gardner, C.W. Putnam, T. Weinert, RAD53, DUN1 and PDS1 define two
parallel G2/M checkpoint pathways in budding yeast, EMBO J. 18 (1999)
 P.R. Dohrmann, R.A. Sclafani, Novel role for checkpoint Rad53 protein kinase
in the initiation of chromosomal DNA replication in Saccharomyces cerevisiae,
Genetics 174 (2006) 87–99.
 A. Gunjan, A. Verreault, A Rad53 kinase-dependent surveillance mechanism
that regulates histone protein levels in S. cerevisiae, Cell 115 (2003) 537–549.
 A. Emili, D.M. Schieltz, J.R. Yates, L.H. Hartwell 3rd, Dynamic interaction of
DNA damage checkpoint protein Rad53 with chromatin assembly factor Asf1,
Mol. Cell 7 (2001) 13–20.
 A.A. Franco, W.M. Lam, P.M. Burgers, P.D. Kaufman, Histone deposition protein
Asf1 maintains DNA replisome integrity and interacts with replication factor
C, Genes Dev. 19 (2005) 1365–1375.
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
 G.M. Alvino, D. Collingwood, J.M. Murphy, J. Delrow, B.J. Brewer, M.K. Raghura-
 Y. Doksani, R. Bermejo, S. Fiorani, J.E. Haber, M. Foiani, Replicon dynamics,
dormant origin firing, and terminal fork integrity after double-strand break
formation, Cell 137 (2009) 247–258.
 A.G. Paulovich, L.H. Hartwell, A checkpoint regulates the rate of progression
through S phase in S. cerevisiae in response to DNA damage, Cell 82 (1995)
 G. Versini, I. Comet, M. Wu, L. Hoopes, E. Schwob, P. Pasero, The yeast Sgs1
helicase is differentially required for genomic and ribosomal DNA replication,
EMBO J. 22 (2003) 1939–1949.
 I. Voineagu, V. Narayanan, K.S. Lobachev, S.M. Mirkin, Replication stalling at
proteins, Proc. Natl. Acad. Sci. U. S. A. 105 (2008) 9936–9941.
 C.H. Freudenreich, M. Lahiri, Structure-forming CAG/CTG repeat sequences
are sensitive to breakage in the absence of Mrc1 checkpoint function and S-
phase checkpoint signaling: implications for trinucleotide repeat expansion
diseases, Cell Cycle 3 (2004) 1370–1374.
 E. Petermann, A. Maya-Mendoza, G. Zachos, D.A. Gillespie, D.A. Jackson, K.W.
Caldecott, Chk1 requirement for high global rates of replication fork pro-
gression during normal vertebrate S phase, Mol. Cell Biol. 26 (2006) 3319–
 E. Petermann, K.W. Caldecott, Evidence that the ATR/Chk1 pathway maintains
normal replication fork progression during unperturbed S phase, Cell Cycle 5
fork rates in human cells, Mol. Biol. Cell 19 (2008) 2373–2378.
 C. Lucca, F. Vanoli, C. Cotta-Ramusino, A. Pellicioli, G. Liberi, J. Haber, M. Foiani,
Checkpoint-mediated control of replisome-fork association and signalling in
response to replication pausing, Oncogene 23 (2004) 1206–1213.
 J.A. Cobb, L. Bjergbaek, K. Shimada, C. Frei, S.M. Gasser, DNA polymerase stabi-
lization at stalled replication forks requires Mec1 and the RecQ helicase Sgs1,
EMBO J. 22 (2003) 4325–4336.
replication forks owing to checkpoint defects, Science 297 (2002) 599–602.
 W. Feng, D. Collingwood, M.E. Boeck, L.A. Fox, G.M. Alvino, W.L. Fangman,
M.K. Raghuraman, B.J. Brewer, Genomic mapping of single-stranded DNA in
hydroxyurea-challenged yeasts identifies origins of replication, Nat. Cell Biol.
8 (2006) 148–155.
 C. Cotta-Ramusino, D. Fachinetti, C. Lucca, Y. Doksani, M. Lopes, J. Sogo, M.
Foiani, Exo1 processes stalled replication forks and counteracts fork reversal
in checkpoint-defective cells, Mol. Cell 17 (2005) 153–159.
 A. Pellicioli, C. Lucca, G. Liberi, F. Marini, M. Lopes, P. Plevani, A. Romano, P.P. Di
Fiore, M. Foiani, Activation of Rad53 kinase in response to DNA damage and its
effect in modulating phosphorylation of the lagging strand DNA polymerase,
EMBO J. 18 (1999) 6561–6572.
 I. Morin, H.P. Ngo, A. Greenall, M.K. Zubko, N. Morrice, D. Lydall, Checkpoint-
dependent phosphorylation of Exo1 modulates the DNA damage response,
EMBO J. 27 (2008) 2400–2410.
 M. Lopes, C. Cotta-Ramusino, G. Liberi, M. Foiani, Branch migrating sister
chromatid junctions form at replication origins through Rad51/Rad52-
independent mechanisms, Mol. Cell 12 (2003) 1499–1510.
 L. Postow, N.J. Crisona, B.J. Peter, C.D. Hardy, N.R. Cozzarelli, Topological chal-
lenges to DNA replication: conformations at the fork, Proc. Natl. Acad. Sci. U.
S. A. 98 (2001) 8219–8226.
 A. Blastyak, L. Pinter, I. Unk, L. Prakash, S. Prakash, L. Haracska, Yeast Rad5
protein required for postreplication repair has a DNA helicase activity specific
for replication fork regression, Mol. Cell 28 (2007) 167–175.
 I.D. Hickson, RecQ helicases: caretakers of the genome, Nat. Rev. Cancer 3
 G. Liberi, G. Maffioletti, C. Lucca, I. Chiolo, A. Baryshnikova, C. Cotta-
Ramusino, M. Lopes, A. Pellicioli, J.E. Haber, M. Foiani, Rad51-dependent
DNA structures accumulate at damaged replication forks in sgs1 mutants
defective in the yeast ortholog of BLM RecQ helicase, Genes Dev. 19 (2005)
 D. D’Amours, S.P. Jackson, The yeast Xrs2 complex functions in S phase check-
point regulation, Genes Dev. 15 (2001) 2238–2249.
 H.L. Klein, Molecular biology: DNA endgames, Nature 455 (2008) 740–741.
 M. Kai, M.N. Boddy, P. Russell, T.S. Wang, Replication checkpoint kinase Cds1
Dev. 19 (2005) 919–932.
 S. Flott, J. Rouse, Slx4 becomes phosphorylated after DNA damage in a
Mec1/Tel1-dependent manner and is required for repair of DNA alkylation
damage, Biochem. J. 391 (2005) 325–333.
 M. Lopes, M. Foiani, J.M. Sogo, Multiple mechanisms control chromosome
Mol. Cell 21 (2006) 15–27.
 M.B. Smolka, C.P. Albuquerque, S.H. Chen, H. Zhou, Proteome-wide identifica-
tion of in vivo targets of DNA damage checkpoint kinases, Proc. Natl. Acad. Sci.
U. S. A. 104 (2007) 10364–10369.
 K. Trenz, E. Smith, S. Smith, V. Costanzo, ATM and ATR promote Mre11 depen-
dent restart of collapsed replication forks and prevent accumulation of DNA
breaks, EMBO J. 25 (2006) 1764–1774.
nascent leading strand, Nature 439 (2006) 557–562.
 N.P. Higgins, K. Kato, B. Strauss, A model for replication repair in mammalian
cells, J Mol Biol 101 (1976) 417–425.
 S.J. Goldfless, A.S. Morag, K.A. Belisle, V.A. Sutera Jr., S.T. Lovett, DNA repeat
21 (2006) 595–604.
 L. Wu, I.D. Hickson, The Bloom’s syndrome helicase suppresses crossing over
during homologous recombination, Nature 426 (2003) 870–874.
 H.W. Mankouri, H.P. Ngo, I.D. Hickson, Shu proteins promote the formation of
homologous recombination intermediates that are processed by Sgs1-Rmi1-
Top3, Mol Biol Cell 18 (2007) 4062–4073.
 D. Branzei, J. Sollier, G. Liberi, X. Zhao, D. Maeda, M. Seki, T. Enomoto, K. Ohta,
M. Foiani, Ubc9- and Mms21-Mediated Sumoylation Counteracts Recombino-
genic Events at Damaged Replication Forks, Cell 127 (2006) 509–522.
 D. Branzei, F. Vanoli, M. Foiani, SUMOylation regulates Rad18-mediated tem-
plate switch, Nature 456 (2008) 915–920.
 S. Jentsch, J.P. McGrath, A. Varshavsky, The yeast DNA repair gene RAD6
encodes a ubiquitin-conjugating enzyme, Nature 329 (1987) 131–134.
 R.M. Hofmann, C.M. Pickart, Noncanonical MMS2-encoded ubiquitin-
conjugating enzyme functions in assembly of novel polyubiquitin chains for
DNA repair, Cell 96 (1999) 645–653.
ubiquitin-conjugating enzymes in DNA repair, EMBO J. 19 (2000) 3388–
 C. Hoege, B. Pfander, G.L. Moldovan, G. Pyrowolakis, S. Jentsch, RAD6-
dependent DNA repair is linked to modification of PCNA by ubiquitin and
SUMO, Nature 419 (2002) 135–141.
 P. Stelter, H.D. Ulrich, Control of spontaneous and damage-induced mutagen-
esis by SUMO and ubiquitin conjugation, Nature 425 (2003) 188–191.
 P.L. Kannouche, J. Wing, A.R. Lehmann, Interaction of human DNA polymerase
eta with monoubiquitinated PCNA: a possible mechanism for the polymerase
switch in response to DNA damage, Mol Cell 14 (2004) 491–500.
 C.A. Torres-Ramos, S. Prakash, L. Prakash, Requirement of RAD5 and MMS2 for
postreplication repair of UV-damaged DNA in Saccharomyces cerevisiae, Mol.
Cell Biol. 22 (2002) 2419–2426.
 L. Haracska, C.A. Torres-Ramos, R.E. Johnson, S. Prakash, L. Prakash, Opposing
effects of ubiquitin conjugation and SUMO modification of PCNA on replica-
tional bypass of DNA lesions in Saccharomyces cerevisiae, Mol. Cell Biol. 24
 L. Barbour, L.G. Ball, K. Zhang, W. Xiao, DNA damage checkpoints are involved
in postreplication repair, Genetics 174 (2006) 1789–1800.
 M. Kai, T.S. Wang, Checkpoint activation regulates mutagenic translesion syn-
thesis, Genes Dev. 17 (2003) 64–76.
 S. Sabbioneda, B.K. Minesinger, M. Giannattasio, P. Plevani, M. Muzi-Falconi, S.
and is partially required for spontaneous polzeta-dependent mutagenesis in
Saccharomyces cerevisiae, J. Biol. Chem. 280 (2005) 38657–38665.
 A.G. Paulovich, R.U. Margulies, B.M. Garvik, L.H. Hartwell, RAD9, RAD17, and
RAD24 are required for S phase regulation in Saccharomyces cerevisiae in
response to DNA damage, Genetics 145 (1997) 45–62.
 M. Kai, K. Furuya, F. Paderi, A.M. Carr, T.S. Wang, Rad3-dependent phosphory-
lation of the checkpoint clamp regulates repair-pathway choice, Nat. Cell Biol.
9 (2007) 691–697.
 B. Pfander, G.L. Moldovan, M. Sacher, C. Hoege, S. Jentsch, SUMO-modified
PCNA recruits Srs2 to prevent recombination during S phase, Nature 436
 E. Papouli, S. Chen, A.A. Davies, D. Huttner, L. Krejci, P. Sung, H.D. Ulrich,
Crosstalk between SUMO and Ubiquitin on PCNA Is Mediated by Recruitment
of the Helicase Srs2p, Mol. Cell 19 (2005) 123–133.
dependent DNA damage bypass is mediated by replication protein a, Mol. Cell
29 (2008) 625–636.
 S. Matsuoka, B.A. Ballif, A. Smogorzewska, E.R. McDonald 3rd, K.E. Hurov, J.
Luo, C.E. Bakalarski, Z. Zhao, N. Solimini, Y. Lerenthal, Y. Shiloh, S.P. Gygi, S.J.
Elledge, ATM and ATR substrate analysis reveals extensive protein networks
responsive to DNA damage, Science 316 (2007) 1160–1166.
N. Tomilin, DNA damage-induced accumulation of Rad18 protein at stalled
replication forks in mammalian cells involves upstream protein phosphoryla-
tion, Biochem. Biophys. Res. Commun. 323 (2004) 831–837.
 L. Rong, F. Palladino, A. Aguilera, H.L. Klein, The hyper-gene conversion hpr5-
1 mutation of Saccharomyces cerevisiae is an allele of the SRS2/RADH gene,
Genetics 127 (1991) 75–85.
 A. Aboussekhra, R. Chanet, A. Adjiri, F. Fabre, Semidominant suppressors of
Srs2 helicase mutations of Saccharomyces cerevisiae map in the RAD51 gene,
whose sequence predicts a protein with similarities to procaryotic RecA pro-
teins, Mol. Cell Biol. 12 (1992) 3224–3234.
 G. Liberi, I. Chiolo, A. Pellicioli, M. Lopes, P. Plevani, M. Muzi-Falconi, M.
requires a functional Mec1-dependent pathway and Cdk1 activity, EMBO J. 19
 Y. Fu, Y. Zhu, K. Zhang, M. Yeung, D. Durocher, W. Xiao, Rad6-Rad18 mediates
a eukaryotic SOS response by ubiquitinating the 9-1-1 checkpoint clamp, Cell
133 (2008) 601–611.
 F. Paques, J.E. Haber, Multiple pathways of recombination induced by double-
strand breaks in Saccharomyces cerevisiae, Microbiol Mol Biol Rev. 63 (1999)
1046 Download full-text
D. Branzei, M. Foiani / DNA Repair 8 (2009) 1038–1046
 G. Ira, A. Pellicioli, A. Balijja, X. Wang, S. Fiorani, W. Carotenuto, G. Liberi, D.
Bressan, L. Wan, N.M. Hollingsworth, J.E. Haber, M. Foiani, DNA end resection,
homologous recombination and DNA damage checkpoint activation require
CDK1, Nature 431 (2004) 1011–1017.
 H. Ogiwara, A. Ui, F. Onoda, S. Tada, T. Enomoto, M. Seki, Dpb11,
the budding yeast homolog of TopBP1, functions with the checkpoint
clamp in recombination repair, Nucleic Acids Res. 34 (2006) 3389–
 L. Ciapponi, G. Cenci, J. Ducau, C. Flores, D. Johnson-Schlitz, M.M. Gorski, W.R.
Engels, M. Gatti, The Drosophila Mre11/Rad50 complex is required to pre-
vent both telomeric fusion and chromosome breakage, Curr. Biol. 14 (2004)