ISSN 0006-2979, Biochemistry (Moscow), 2025, Vol. 90, No. 11, pp. 1468-1483 © Pleiades Publishing, Ltd., 2025.
Russian Text © The Author(s), 2025, published in Biokhimiya, 2025, Vol. 90, No. 11, pp. 1561-1578.
1468
REVIEW
The Role of Non-Homologous End Joining
and Microhomology-Mediated End Joining
in Chromosomal Rearrangements
Nikolai A. Lomov
1,a
*, Nikolai A. Nikolaev
2,3
, Vladimir S. Viushkov
1
,
and Mikhail A. Rubtsov
1,4
1
Department of Molecular Biology, Faculty of Biology, Lomonosov Moscow State University,
119234 Moscow, Russia
2
Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University,
119234 Moscow, Russia
3
Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences,
117437 Moscow, Russia
4
Center for Industrial Technologies and Entrepreneurship,
I.M. Sechenov First Moscow State Medical University (Sechenov University), 119435 Moscow, Russia
a
e-mail: lomov13@gmail.com
Received July 29, 2025
Revised October 31, 2025
Accepted November 8, 2025
Abstract— Double-strand DNA break (DSB) repair mechanisms vary in their ability to prevent errors during
end joining. The joining of DSBs on different chromosomes can result in translocations, potentially leading
to tumorigenesis. This review examines the main mechanisms of DSB repair and factors influencing their
selection, as well as contribution of these mechanisms to the chromosomal rearrangements in human cells.
DOI: 10.1134/S0006297925602102
Keywords: double-strand DNA break repair, translocations, non-homologous end joining (NHEJ), microhomolo-
gy-mediated end joining (MMEJ)
* To whom correspondence should be addressed.
INTRODUCTION
Double-strand DNA breaks (DSBs) are the most
dangerous DNA lesions. They block replication and
transcription of damaged DNA, and many genes can
be lost during cell division due to chromosomal
breaks. Cells employ multiple DSB repair pathways
that often overlap for reliability. These pathways dif-
fer in the mechanisms involved, repair rate, and pro-
tection from errors. The most dangerous error is the
joining of DNA ends from different breaks, which can
lead to translocations and transformation of normal
cells into tumors [1-3]. The contribution of each DSB
repair mechanism to the formation of translocations
in human cells remains unclear. This review examines
the main DSB repair mechanisms, factors influencing
DNA repair pathway selection, and their association
with chromosomal rearrangements in mammalian
cells. Note that other chromosomal rearrangements–
large deletions and inversions – have the same mech-
anisms of formation as translocations, so the discussed
pathways apply to them as well.
GENERAL SCHEME
OF DSB REPAIR MECHANISMS
The general scheme of DSB repair pathways in
human cells is shown in Fig. 1. The first mechanism
is non-homologous end joining (NHEJ), which operates
throughout entire cell cycle [4-7]. Alternative end-join-
ing mechanisms that do not require a homology do-
nor, include microhomology-mediated end joining
(MMEJ) and single-strand annealing (SSA) [8]. Both
MMEJ and SSA use short homologous sequences (mi-
crohomologies) on both sides of the break, but differ
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Fig. 1. General scheme of DSB repair pathways in human cells: non-homologous end joining (NHEJ), microhomology-mediat-
ed end joining (MMEJ), single-strand annealing (SSA), and homology-directed repair (HDR). The final ligation stage for NHEJ,
MMEJ, and SSA is not shown; HDR variants resolving Holliday junctions are also not depicted. Orange, newly synthesized
DNA; red, homologous regions; dashed lines, nucleotide removal.
in the length of these homologous regions and pro-
teins involved [9]. However, these two pathways lack
a mechanism ensuring correct end selection for the
DNA ligation, making them error prone. Homology-
directed repair (HDR), also called homologous recom-
bination (HR) because it is involved in the crossing
over during meiotic recombination [10, 11], is more
reliable in preventing chromosomal rearrangements.
Because HDR uses a homologous template, typically,
sister chromatid, it is active mostly in the S and G2
phases of cell cycle [5]. The use of homology donor
prevents incorrect joining of chromosomal ends and
formation of translocations. Detailed description of
HDR can be found in reviews by Sanchez et al. [11],
Sun et al. [12], and Al-Zain and Symington [13].
SSA is a rare repair pathway because it requires
extensive homology regions (>20 nucleotides) at both
break sites [14, 15]. Therefore, translocations oc-
curring by SSA are uncommon. SSA is discussed in
Bhargava et al. [9], Blasiak [16], and Vu et al. [17].
We chose not to describe in detail the HDR and SSA
pathways in this review and focused on NHEJ and
MMEJ, their molecular mechanisms, and role in the
formation of chromosomal translocations.
NON-HOMOLOGOUS END JOINING (NHEJ)
NHEJ is a dominant DSB repair pathway in ver-
tebrates during the G1 phase, although it is active
throughout the entire cell cycle [5, 7, 18]. It is a uni-
versal mechanism for joining genomic fragments, in-
cluding during V(D)J recombination (immunoglobulin
and T-cell receptor gene formation). NHEJ had once
been considered a simple, linear process: break detec-
tion, end retention, processing, and ligation. However,
recent data have revealed a more complex organiza-
tion, with NHEJ protein complexes transiting between
multiple functional states [19, 20].
DSBs are recognized by the Ku70/Ku80 (XRCC6/
XRCC5, or X-ray repair cross-complementing protein
6/5) complex [21, 22], which is a heterodimer consist-
ing of two subunits with molecular masses of 70 and
80 kDa, respectively, that form a ring-like structure
with positively charged amino acids residues inside.
Ku70/Ku80 binds to the DNA ends [21, 23], both blunt
and overhanging, and recruits the catalytic sub-
unit (DNA-PKcs) of DNA-dependent protein kinase
(DNA-PK), forming the DNA-PK holoenzyme [24]. Two
break ends, each bound by Ku70/Ku80 and DNA-PKcs,
constitute the long-range synaptic complex (LR), which
may include additional factors, such as XRCC4 (X-ray
repair cross-complementing protein 4), XLF/Cernun-
nos, and LIG4 (DNA ligase IV). This complex holds the
break ends together.
Cryo-electron microscopy has revealed two struc-
tural variants of LR, differing in their architecture
and protein composition (Fig. 2). In the first of them,
dimerization occurs through the interaction of the
C-terminal domains of Ku80 with DNA-PKcs in trans
(referred to as Ku80-mediated LR or domain-swap LR).
In the second variant (referred to as XLF-mediated LR),
dimerization is mediated by a filament formed by aux-
iliary factors, such as XLF and two LIG4 complexes
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Fig. 2. NHEJ and MMEJ pathways of DSB repair in human cells. CTD, C-terminal domain; LR and SR, long- and short-range
synaptic complexes, respectively; HD and PD, helicase and DNA polymerase domains of POLQ, respectively. Small arrows
indicate the direction of the exonuclease and helicase activities of MRN and POLQ (see the text for explanation).
associated with XRCC4 [25-27]. So far, it remains un-
clear whether the Ku80- and XLF-mediated LR com-
plexes can interconvert or they form from different
monomeric structures of the DNA-PK holoenzyme
(described, for example, in Liu et al. [28]), or both
scenarios are possible.
The XLF-mediated LR can compact into a short-
range synaptic complex (SR) through mutual phos-
phorylation of two DNA-PKcs molecules, followed
by their dissociation from the complex [27, 29]. This
brings the break ends sufficiently close for the ligation
by LIG4. The organization of the complex appears to
be determined by the autophosphorylation of DNA-PK
and its interaction with DNA [19, 28].
During the NHEJ-mediated repair, the ends of the
DNA break can undergo processing. The nature and
extent of DNA processing in NHEJ, as well as addi-
tional proteins recruited for this purpose, vary de-
pending on the properties of DSB ends. Blunt ends
and compatible overhangs can be ligated directly [30].
However, if the ends contain incompatible overhangs,
chemical damage, or modifications, the NHEJ complex
employs specialized nucleases and polymerases to
prepare the ends for ligation. The major nuclease in
NHEJ is DNA cross-link repair 1C protein (ARTEMIS),
which possesses both endonuclease and 5′→3′ exonu-
clease activities [31, 32]. DNA polymerases involved
in NHEJ are polymerase µ (POLM) and polymerase λ
(POLL) (members of the Pol X family) that can add
nucleotides in a template-independent manner in ad-
dition to the standard polymerase activity [33].
Using in vitro systems, it was shown that incom-
patible 5′ ends are preferentially trimmed by ARTEMIS
[34], while incompatible 3′ ends undergo both deg-
radation by ARTEMIS and extension by POLM [34].
ARTEMIS also resolves the hairpin structures formed
at DNA ends during V(D)J recombination; mutations in
the ARTEMIS gene lead to severe combined immuno-
deficiency [35, 36]. Polynucleotide kinase/phosphatase
(PNKP) removes phosphate groups from 3′ ends and
adds phosphate groups to 5′ ends of DNA. Notably,
the presence at the overhangs of complementary nu-
cleotides (microhomology regions of 2-4 nucleotides)
facilitates end joining [34]. The processing of break
ends results in the appearance of small insertions and
deletions (indels) at the repair site [34, 37].
Different NHEJ complexes can include different
processing enzymes and activities. ARTEMIS is recruit-
ed by the DNA-PK holoenzyme either as a monomer
[28] or in the content of the Ku80-mediated LR [19].
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However, Stinson et al. [30] found that the removal
of 5′ overhangs in Xenopus laevis egg extracts was
catalyzed by an unidentified 5′→3′ exonuclease (not
ARTEMIS), whose activity depended on XLF and
XRCC4–LIG4. This exonuclease was present in XLF-
mediated LR or SR. There is an emerging consensus
that other types of processing occur either in the XLF-
mediated LR and SR or exclusively in the SR [19, 30,
38], which is supported by the fact that DNA-PKcs
and Ku70/80 protect the ends from most processing
enzymes until the SR is formed [30], allowing access
for ARTEMIS only [28].
Focusing the majority of processing activities at
the SR stage, i.e., when ligation takes place, appears
tobe an efficient way to minimize mutagenesis during
DNA repair, as the processed ends join at the earli-
est opportunity [39]. Therefore, new data and models
challenge the concept of NHEJ as a mechanism prone
to indels due to its simplistic nature.
MICROHOMOLOGY-MEDIATED
END JOINING (MMEJ)
The first evidence of an alternative end-joining
pathway came from the studies of Saccharomyces
cerevisiae Ku70 mutants. Even with additional Rad52
mutations, these cells retained their capacity for the
repair of DSBs caused by ionizing radiation, albe-
it this process was accompanied by the formation
of deletions [40]. Extracts from bovine thymus also
demonstrated the presence of two end-joining path-
ways: one joining blunt ends with no homology, and
another using short identical sequences (microhomol-
ogies) [41]. The studies of mammalian cells deficient
in NHEJ components confirmed the existence of an al-
ternative pathway, termed alt-NHEJ [42]. It was shown
that this mechanism acts as an alternative to NHEJ
and HDR, but can also function simultaneously with
these pathways. Later, it was renamed MMEJ to em-
phasize its reliance on microhomology [44]. NHEJ can
also use microhomologies of 2-4 nucleotides [34], but
this is not obligatory. In this review, the term MMEJ
refers specifically to the pathway described below,
while NHEJ refers to the canonical mechanism de-
pendent on Ku proteins, DNA-PKcs, and LIG4. Older
publications may use the terms alt-NHEJ, a-NHEJ, or
a-EJ to describe MMEJ. Due to the involvement of
DNA polymerase θ (POLQ), MMEJ is also called TMEJ
(theta-mediated end joining) [15, 45].
MMEJ typically begins with the DSB recognition
by the MRN complex composed of the MRE11 nucle-
ase, regulatory ATPase RAD50, and NBS1 (Nijmegen
breakage syndrome) scaffold protein [46]. MRN is
recruited by poly(ADP-ribose) polymerases (PARP1
and PARP2) recognizing the breaks in DNA [47].
Accumulation of MRN at the break sites can occur via
liquid-liquid phase separation, driven by the intrin-
sically disordered domains of MRNIP (MRN complex
interacting protein) [48, 49].
After recognition of the break, MRN resects its
ends. The activation of MRN requires its phosphor-
ylation by CtIP (CtBP-interacting protein/retinoblasto-
ma-binding protein 8), after which MRN binds to DNA
at a distance from the DSB [50, 51]. MRE11 creates
nick and then degrades the 5′ end back toward the
DSB end due to its 3′→5′ exonuclease activity (Fig. 2)
[14, 52]. The PARylation of histones, BRCA1 (BReast
CAncer gene 1), and other proteins is crucial for the
resection initiation [53-56].
The next step in MMEJ is the search for micro-
homologies and annealing. If 3′ overhangs contain
microhomologies (typically, <20 nucleotides), they can
anneal [14, 15]. DNA polymerase θ (POLQ in humans)
holds the 3′ ends while the search for the microho-
mologies takes place. POLQ is a central protein in the
MMEJ mechanism in mammals [57, 58]. Its distinctive
feature is the presence of the helicase domain [59,
60] that non-specifically binds single-stranded DNA
regions, competing with RPA (replication protein A).
Due to ATP hydrolysis, the helicase domain threads
the protruding 3′ ends of DSBs, displacing RPA pro-
teins [61]. During this process, single-stranded DNA
regions can anneal if the microhomology regions
are present. It is important to note that the helicase
domains of POLQ function as a dimer, thus holding
the break ends together [58, 62, 63]. After microho-
mology annealing, any protruding unpaired 3′ end
(3′-flap) is removed. According to different studies,
removal of 3′-flaps involves the XPF–ERCC1 com-
plex, APE2 (apurinic/apyrimidinic endonuclease 2)
protein, or 3′→5′ exonuclease activity of DNA poly-
merase δ [43, 64, 65].
POLQ then extends the complementary region via
non-processive, error-prone synthesis. This process is
accompanied by multiple dissociation/reassociation
of POLQ, formation and elongation of hairpins on
DNA single strands, etc., resulting in the generation
of templated insertions (TINSs) – a hallmark of MMEJ
(Fig. 3) [66, 67]. After addition of ~10 nucleotides,
POLQ is replaced by the less error-prone and more
processive POLδ–PCNA complex [65, 68]. After filling
the gaps, the breaks are ligated by LIG1 (replicative
ligase) or LIG3 [69, 70].
Since in MMEJ, the DNA ends are held together
due to the annealing in two microhomology regions,
one of these regions will be lost after DNA repair, as
well the DNA fragment between them. These micro-
homology-mediated deletions and TINSs are used to
identify MMEJ events [66, 67].
Recent studies have revealed MMEJ as a pre-
ferred DNA repair mechanism in mitosis [71-73].
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Fig. 3. Two examples of MMEJ mechanisms leading to the formation of templated insertions (TINSs). The choice of the
mechanism depends on the presence and orientation of homology regions (direct or inverted repeats shown in boxes).
Newly synthesized DNA is shown in (initially synthesized) and blue (synthesized after secondary annealing of DNA ends).
It was found that MMEJ is frequently involved in
rearrangements at common fragile sites, which are
prone to replication fork stalling and underreplica-
tion. As a result, these sites are prone to DNA breaks
that are repaired by MMEJ during mitosis [74, 75].
Therefore, MMEJ serves as a backup for DSBs
unrepaired by NHEJ or HDR, but its involvement re-
sults in the formation of deletions in the repaired
regions. For this reason, the activity of MMEJ is low
in normal cells. However, tumor cells often have im-
paired DNA repair pathways, making MMEJ the prin-
cipal mechanism of DNA repair in these cells, which
also makes POLQ a potential therapeutic target in the
antitumor therapy [76-78].
THE CHOICE OF DSB REPAIR MECHANISM
DSB repair pathways have been traditionally
viewed as independent molecular processes. Howev-
er, DNA repair can start via one pathway and end
via another. For example, the switch between the
mechanism can occur after recognition of the break
ends, as the binding of Ku70/80 does not prevent the
recruitment of MRN and initiation of end resection
via MMEJ or HDR [51, 79, 80], while recognition of
the break by the MRN complex can lead to the NHEJ-
mediated DNA repair [81, 82]. MMEJ, SSA, and HDR
share many similar steps; for example, they all start
with the MRN recruitment and end resection [83].
Despite the possibility of switching between the
DNA repair pathways, there are factors that predeter-
mine realization of particular mechanisms. The key
factor appears to be the end resection. Many proteins
affect the choice of the repair mechanism by modulat-
ing the extent of resection. Blunt ends or compatible
overhangs are optimal for NHEJ, while incompatible
ends require processing by nucleases/polymerases
[37, 84]. Extended single-stranded ends poorly bind
Ku70/80 [85]. MMEJ requires overhangs for the mi-
crohomology search. Therefore, once resection is ini-
tiated, the cells choose between HDR, SSA, or MMEJ.
Resection proceeds in two stages: short-range
resection (tens to hundreds of nucleotides) by MRN
followed by the long-range resection (hundreds to
thousands of nucleotides) by EXO1 and DNA2. Short-
range resection can occur even when the ends are
blocked by proteins or secondary structures; it starts
with a nick introduced at distance from the break,
after which DNA is degraded toward the break. Long-
range resection is faster, but more sensitive to ob-
stacles. Short-range resection is sufficient for MMEJ,
while HDR and SSA require long-rejection resection
[14, 28, 83, 86].
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Resection of DNA ends occurs during the S and
G2 phases of cell cycle and favors MMEJ, SSA, and
HDR. Cyclin-dependent kinases CDK1 and CDK2 phos-
phorylate CtIP, thus activating MRN and promoting
resection [87, 88]. Post-translational modifications of
repair proteins play an important role in the regu-
lation of resection. Thus, ataxia telangiectasia mu-
tated (ATM) kinase activated by DSBs, phosphory-
lates H2AX histone, resulting in the recruitment of
DNA repair proteins to the break. Ubiquitination of
H1 and H2A histones by ubiquitin ligases RNF8 and
RNF168 also signals for the repair protein recruit-
ment [81, 89].
53BP1 (p53-binding protein) and BRCA1 are key
regulators of resection. Both are recruited to the
chromatin regions containing histones ubiquitinated
by RNF8/RNF168 [81, 89]. 53BP1 inhibits resection
and directs DNA repair toward NHEJ [90, 91]. 53BP1
binds most DSBs by default, even in the G2 phase [92-
94]. Its antagonist BRCA1 modifies CtIP, BLM, WRN,
and EXO1, thus stimulating resection [95-97]. BRCA1
displaces 53BP1 during the S and G2 phases and re-
cruits the SMARCAD1 chromatin remodeling complex
[18, 85, 98-101]. The removal of ubiquitinated nucleo-
somes disrupts the binding between 53BP1 and DSBs
and facilitates resection of the break ends [18, 85,
98-101]. Recent data suggest that the state of chro-
matin influences the choice of DNA repair pathway
mechanism, e.g., euchromatin favors NHEJ over MMEJ
compared to heterochromatin [14, 102].
There is a mechanism that limits NHEJ in the
S and G2 phases. The micropeptide CYREN (a mem-
ber of a family of small regulatory peptides named
by analogy with microRNAs) binds Ku70/80, thus
restricting its participation in the DSB repair [103,
104]. Finaly, NHEJ can also occur “instantly,” without
involvement of ATM, RNF8, RNF168, or protein com-
plexes responsible for the choice of DNA repair mech-
anism [105].
The choice of MMEJ is strongly facilitated by
PARP1 [106]. Its activity varies during the cell cycle,
with the highest and lowest activities observed in the
S and G1 phases, respectively [107]. PARP1 recruits
MRN to DSBs [108], thus promoting resection. As a re-
sult, PARP1 inhibits NHEJ by competing with Ku70/80
for DNA binding and reducing the affinity of Ku70/80
for DNA (since the affinity of these proteins to the sin-
gle-stranded DNA formed by resection is lower) [109].
PARP1 also inhibits the long-range resection, which
shifts DNA repair toward MMEJ [106, 110, 111].
As mentioned above, MMEJ is a preferred DSB
repair pathway in mitosis, when NHEJ and early
stages of HDR are inhibited [90, 112, 113]. Polo-like
kinase 1 (PLK1), which controls cell entry to mitosis,
phosphorylates RHINO (Rad9, Hus1, Rad1 Interacting
Nuclear Orphan). RHINO is accumulated during mi-
tosis; its major function, together with the 9-1-1 com-
plex (Rad9, Hus1, Rad1), is triggering of ATR signaling
and cell cycle arrest in response to the replication
stress and DNA damage [114, 115]. However, it was
shown recently that phosphorylated RHINO recruits
POLQ to DSBs for MMEJ activation independently
of PARP1 [106, 116-119].
IMPACT OF DSB FORMATION
ON THE CHOICE OF DNA REPAIR MECHANISM
DSBs can appear in DNA for a variety of reasons.
Endogenous DSBs primarily occur during replication,
often in actively transcribed regions due to collisions
between the replication forks and RNA polymerase
or increased level of DNA damage [120-124]. Induc-
tion of DSBs can be programmed in certain cell types.
For instance, lymphocytes express RAG nuclease and
activation-induced cytidine deaminase (AID) for the
initiation of V(D)J recombination, class-switch re-
combination, and somatic hypermutation [125, 126].
In germ cells, SPO11 forms DSBs for meiotic recom-
bination [127-129]. Dysfunction of topoisomerase 2
(TOPO2) can also cause DSBs. TOPO2 decatenates rep-
licated chromosomes and relieves supercoiling [130]
by introducing transient DSBs in DNA molecule to
pass another DNA segment through [131, 132]. TOPO2
is a dimer; each monomer forms temporary covalent
bond with a DNA end in a DSB [133]. If TOPO2 is
nonfunctional, religation fails, and the break becomes
permanent [134, 135]. TOPO2 inhibitors are used in
chemotherapy to induce accumulation of DSBs in rap-
idly dividing cancer cells and cell death [136-138].
The mechanism of DSB induction affects the struc-
ture of break ends and, therefore, the choice of DNA
repair pathway [139, 140]. For example, this choice
depends on the type of ionization particles causing
the breaks in DNA: high-energy particles (e.g., in car-
bon-ion therapy) induce multiple DNA damage, with
DSBs surrounded by single-strand breaks and base
damage, which hinders Ku70/80 binding and favors
resection-dependent pathways. In contrast, lower-
energy photon-based radiotherapy causes DSBs that
can be directly ligated by NHEJ [15].
DNA breaks induced by the replication stress, the
so-called one-ended DSBs, are typically repaired by
HDR [141-143]. If unrepaired in the S or G2 phases,
they are repaired mostly by MMEJ (see above)
[74, 106].
The presence of covalently linked protein adducts
at the DSB ends can also affect the process of DNA
repair (as in the case of inhibition by TOPO2). The
repair of such DSBs requires removal of the DNA-TO-
PO2 complex by proteases or nucleases. Nucleolytic
removal involves MRN, which cleaves off the protein
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adduct together with the DNA fragment, thus initi-
ating resection of the break ends [144]. In the case
of proteolytic removal, tyrosyl-DNA phosphodies-
terase 2 (TDP2) cleaves the TOPO2-DNA bond. The
access of TDP2 to this bond requires denaturation
or partial proteolysis of TOPO2. The change in the
TOPO2 conformation is provided via sumoylation
by the SUMO ligase ZATT (Znf protein associated
with TDP2 and TOP2), while proteolytic cleavage is
catalyzed by proteasome or metallonuclease SPRTN
[144-147]. After the action of TDP2, the ends become
available for NHEJ or other pathways; however, such
complex mechanism of DNA end processing delays
the repair.
Different DNA repair mechanisms occur at differ-
ent rates, which affects the risk of errors during end
joining [148,149]. Slower repairs increase the chance
of ends drifting apart and encountering another
break, thus promoting translocations [150]. This may
explain why the TOPO2 inhibitor therapy is associat-
ed with the emergence of secondary leukemias fea-
turing chromosomal translocations. Etoposide-treated
cells often show DNA break ends outside the chro-
mosomal territories, which is a prerequisite for chro-
mosomal translocations [151, 152]. It was found that
the ends of etoposide-induced breaks are more mobile
than those of radiation-induced breaks [152, 153].
CONTRIBUTION OF NHEJ AND MMEJ
TO THE FORMATION OF TRANSLOCATIONS
The debates continue whether it is canonical
NHEJ or alternative mechanism (MMEJ) is the prima-
ry driver of chromosomal translocations in mammals
[154-158]. Numerous studies using various method-
ological approaches have yielded conflicting results.
Early studies of cells from leukemia patients
have shown that translocations are often formed
with the involvement of microhomologies at the
break junctions [154]. Inhibition or immune depletion
of NHEJ proteins in tumor cell extracts did not abol-
ish the joining of DNA fragments, and sequencing of
linked DNA fragments revealed the use of microho-
mologies, implicating MMEJ in the translocation for-
mation [154].
Another approach to evaluate the contribution of
NHEJ and MMEJ in the formation of translocations
is comparing DNA repair events in wild-type and
NHEJ-deficient cell lines. Simsek and Jasin [155] and
Weinstock et al. [159] used embryonic stem cells from
Xrcc4
–/–
and Ku70
–/–
transgenic mice containing two
transgenic cassettes with the I-SceI meganuclease site.
Transient expressed I-SceI induced DNA breaks, while
the fusion of two cassettes made cells antibiotic-re-
sistant, so colonies growing on antibiotic-containing
media were counted and sequenced. It was found that
microhomologies (≥4 nucleotides) were used in ~25%
of rearrangements. Experiments were performed in
both NHEJ gene-expressing and NHEJ gene-deficient
cells. In Ku70
–/–
or Xrcc4
–/–
cells, the frequency of
antibiotic-resistant (translocation-containing) colonies
was higher, suggesting that in the absence of NHEJ,
translocations form via MMEJ and that MMEJ is more
prone to erroneous joining [155].
In NIH3T3 mouse fibroblasts, the frequency of
translocation increased with the inhibition or knock-
down of DNA-PKcs [160]. The results were obtained
by invivo microscopy using the cells contained a sys-
tem for visualization of genomic loci, which allowed
to observe I-SceI-induced breaks and their conver-
gence with the following formation of translocations
[160]. DNA-PKcs inhibition increased the transloca-
tion frequency (as detected by PCR) in human lym-
phocytes with the integrated CRISPR/Cas9 system for
induction of rearrangements between the MYC and
IGH genes [161].
However, the study in human HCT116 and NALM6
cells yielded different results [156]. The breaks were
induced by programmable nucleases (ZFN, TALEN,
Cas9), and translocations were detected by PCR 48
hours after transfection. The PCR products were
cloned into plasmid vectors and sequenced. It was
found that most rearrangements occurred without the
use of microhomologies, as in the work by Simsek
and Jasin [155]. The difference was that in LIG4- or
XRCC4-deficient cells, the frequency of translocations
decreased, but the number of deletions and the use of
microhomologies at the translocation sites increased.
The authors concluded that under normal conditions,
translocations in human cells are formed mostly by
NHEJ, and the differences from the study by Simsek
and Jasin [155] may reflect species-specific (mouse vs.
human) or methodological variations [156]. For ex-
ample, during the 48-hour post-transfection period
(Ghezraoui et al. [156]), only a fraction of possible
translocations was formed, and their frequency was
even lower in NHEJ-deficient cells. In contrast, Sim-
sek and Jasin [155] allowed 7-10 days for all possible
translocations to occur, so the difference in the trans-
location frequency in the wild-type cell and cells with
mutations in the DNA repair system was the opposite.
Next-generation sequencing (NGS) of transloca-
tion junctions has provided deeper insights into the
contribution of particular DNA repair mechanisms.
Chiarle et al. [162] used mouse cells with integrated
I-SceI sites and lentiviral I-SceI expression, followed
by massively parallel sequencing of translocation
junctions to show that microhomology was used in
most translocations. Conversely, based on the results
of massively parallel sequencing of Cas9-induced
translocation junctions in human cell lines [157],
DOUBLE-STRAND DNA BREAK REPAIR AND TRANSLOCATIONS 1475
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
most translocations occurred without the use of mi-
crohomologies (>3 nucleotides), even in the case of
DNA-PKcs inhibition.
The studies conducted in 2011-2024, which em-
ployed different models, DSB induction methods, and
translocation detection techniques, yielded varying
results. Currently, it is commonly accepted that both
canonical NHEJ and MMEJ are involved in the for-
mation of translocations and normal DNA repair, and
the contribution of each pathway depends on the cell
type and pathway functionality. A frequent associa-
tion of translocations with MMEJ – more so than in
correct (cis) repair [154, 159, 162] – may reflect a
slower repair kinetics in MMEJ [163]. Normally, the
appearance of a break in DNA should immediately
initiate NHEJ [105]. If NHEJ is delayed, the risk of
ends drifting apart, getting “lost”, and forming trans-
locations increases. Such ends are also more likely
to undergo resection and MMEJ; even if Ku70/80 are
already bound, this does not preclude the MRN bind-
ing and resection [79]. In other words, DNA break
ends “getting lost” represent a common prerequisite
for both translocation formation and MMEJ-mediat-
ed joining. Further NGS studies comparing translo-
cations (trans repair) and normal repair (cis repair)
are needed for better understanding of DNA repair
mechanisms.
CONCLUSION
Cells employ multiple DSB repair mechanisms.
The choice of a particular pathway is influenced
by the DNA damage type and cell cycle phase and
is governed by the repair complex composition and
post-translational modifications of histones and re-
pair proteins. Chromosomal rearrangements most
commonly occur via NHEJ and MMEJ [3]. NHEJ does
not require extensive end processing, so rearrange-
ments typically occur at the break site with minimal
indel formation. MMEJ requires resection, while the
use of microhomologies inevitably causes deletions.
However, MMEJ activation does not necessarily result
in chromosomal rearrangements. The key factor in
joining the ends from the same break is their teth-
ering until ligation. Both translocations and the bias
toward the MMEJ pathway are consequences of end
separation. Further research is needed to clarify the
contribution of DNA repair pathways to the formation
of chromosomal rearrangements.
Abbreviations
DSB double-strand break
HDR homology-directed repair
LIG4 DNA ligase IV
LR long-range synaptic complex
MMEJ microhomology-mediated end joining
MRN MRE11–RAD50–NBS1 protein complex
NHEJ non-homologous end joining
POLQ DNA polymerase θ (Pol X family)
SR short-range synaptic complex
SSA single-strand annealing
Contributions
All authors contributed to writing this review.
Funding
This research was supported by the Russian Science
Foundation (project no.24-24-20019).
Ethics approval and consent to participate
This work does not contain any studies involving hu-
man and animal subjects.
Conflict of interest
The authors of this work declare that they have
noconflicts of interest.
REFERENCES
1. Greaves,M. (2015) When one mutation is all it takes,
Cancer Cell, 27, 433-434, https://doi.org/10.1016/
j.ccell.2015.03.016.
2. Andersson, A. K., Ma, J., Wang, J., Chen, X., Gedman,
A.L., Dang,J., Nakitandwe,J., Holmfeldt,L., Parker,M.,
Easton,J., Huether,R., Kriwacki,R., Rusch,M., Wu,G.,
Li, Y., Mulder, H., Raimondi, S., Pounds, S., Kang, G.,
Shi, L., Becksfort, J., Gupta, P., Payne- Turner, D.,
Vadodaria, B., Boggs, K., et al. (2015) The landscape
of somatic mutations in infant MLL-rearranged acute
lymphoblastic leukemias, Nat. Genet., 47, 330-337,
https://doi.org/10.1038/ng.3230.
3. Bunting, S.F., and Nussenzweig,A. (2013) End-joining,
translocations and cancer, Nat. Rev. Cancer, 13, 443-
454, https://doi.org/10.1038/nrc3537.
4. Beucher, A., Birraux, J., Tchouandong, L., Barton, O.,
Shibata, A., Conrad,S., Goodarzi, A. A., Krempler, A.,
Jeggo, P.A., and Löbrich,M. (2009) ATM and Artemis
promote homologous recombination of radiation-
induced DNA double-strand breaks in G2, EMBO J.,
28, 3413-3427, https://doi.org/10.1038/emboj.2009.276.
5. Karanam,K., Kafri,R., Loewer,A., and Lahav,G. (2012)
Quantitative live cell imaging reveals a gradual shift
between DNA repair mechanisms and a maximal use
of HR in mid-S phase, Mol. Cell, 47, 320-329, https://
doi.org/10.1016/j.molcel.2012.05.052.
6. Rothkamm, K., Krüger, I., Thompson, L. H., and
Löbrich, M. (2003) Pathways of DNA double-strand
break repair during the mammalian cell cycle,
Mol. Cell Biol., 23, 5706-5715, https://doi.org/10.1128/
MCB.23.16.5706-5715.2003.
LOMOV et al.1476
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
7. Shahar, O. D., Raghu Ram, E. V. S., Shimshoni, E.,
Hareli,S., Meshorer, E., and Goldberg,M. (2012) Live
imaging of induced and controlled DNA double-strand
break formation reveals extremely low repair by ho-
mologous recombination in human cells, Oncogene,
31, 3495-3504, https://doi.org/10.1038/onc.2011.516.
8. Fishman-Lobell, J., Rudin, N., and Haber, J. E. (1992)
Two alternative pathways of double-strand break re-
pair that are kinetically separable and independent-
ly modulated, Mol. Cell Biol., 12, 1292-1303, https://
doi.org/10.1128/MCB.12.3.1292.
9. Bhargava, R., Onyango, D. O., and Stark, J. M. (2016)
Regulation of single-strand annealing and its role
in genome maintenance, Trends Genet., 32, 566-575,
https://doi.org/10.1016/j.tig.2016.06.007.
10. Li,W., and Ma,H. (2006) Double-stranded DNA breaks
and gene functions in recombination and meiosis, Cell
Res., 16, 402-412, https://doi.org/10.1038/sj.cr.7310052.
11. Sanchez, A., Reginato, G., and Cejka, P. (2021) Cross-
over or non-crossover outcomes: tailored processing
of homologous recombination intermediates, Curr.
Opin. Genet. Dev., 71, 39-47, https://doi.org/10.1016/
j.gde.2021.06.012.
12. Sun, Y., McCorvie, T. J., Yates, L. A., and Zhang, X.
(2020) Structural basis of homologous recombination,
Cell Mol. Life Sci., 77, 3-18, https://doi.org/10.1007/
s00018-019-03365-1.
13. Al-Zain, A.M., and Symington, L.S. (2021) Thedark side
of homology-directed repair, DNA Repair (Amst), 106,
103181, https://doi.org/10.1016/j.dnarep.2021.103181.
14. Ceccaldi,R., and Cejka,P. (2025) Mechanisms and reg-
ulation of DNA end resection in the maintenance of
genome stability, Nat. Rev. Mol. Cell Biol., 26, 586-599,
https://doi.org/10.1038/s41580-025-00841-4.
15. Van de Kamp, G., Heemskerk, T., Kanaar, R., and
Essers,J. (2021) DNA double strand break repair path-
ways in response to different types of ionizing radia-
tion, Front. Genet., 12, 738230, https://doi.org/10.3389/
fgene.2021.738230.
16. Blasiak, J. (2021) Single-strand annealing in can-
cer, Int.J. Mol. Sci., 22, 2167, https://doi.org/10.3390/
ijms22042167.
17. Vu, T. V., Das, S., Nguyen, C. C., Kim, J., and Kim, J.-
Y. (2022) Single-strand annealing: Molecular mecha-
nisms and potential applications in CRISPR-Cas-based
precision genome editing, Biotechnol.J., 17, e2100413,
https://doi.org/10.1002/biot.202100413.
18. Shibata, A. (2017) Regulation of repair pathway
choice at two-ended DNA double-strand breaks, Mu-
tat. Res., 803-805, 51-55, https://doi.org/10.1016/j.mrf-
mmm.2017.07.011.
19. Buehl, C. J., Goff, N. J., Hardwick, S. W., Gellert, M.,
Blundell, T.L., Yang,W., Chaplin, A.K., and Meek, K.
(2023) Two distinct long-range synaptic complexes
promote different aspects of end processing prior to
repair of DNA breaks by non-homologous end joining,
Mol. Cell, 83, 698-714.e4, https://doi.org/10.1016/j.mol-
cel.2023.01.012.
20. Zhao,F., Kim,W., Kloeber, J.A., and Lou,Z. (2020) DNA
end resection and its role in DNA replication and DSB
repair choice in mammalian cells, Exp. Mol. Med., 52,
1705-1714, https://doi.org/10.1038/s12276-020-00519-1.
21. Grundy, G. J., Moulding, H. A., Caldecott, K. W., and
Rulten, S. L. (2014) One ring to bring them all – the
role of Ku in mammalian non-homologous end joining,
DNA Repair (Amst), 17, 30-38, https://doi.org/10.1016/
j.dnarep.2014.02.019.
22. Thacker, J., and Zdzienicka, M. Z. (2003) The mam-
malian XRCC genes: their roles in DNA repair and
genetic stability, DNA Repair, 2, 655-672, https://
doi.org/10.1016/S1568-7864(03)00062-4.
23. Frit,P., Ropars,V., Modesti,M., Charbonnier, J.B., and
Calsou, P. (2019) Plugged into the Ku-DNA hub: The
NHEJ network, Progr. Biophys. Mol. Biol., 147, 62-76,
https://doi.org/10.1016/j.pbiomolbio.2019.03.001.
24. Davis, A. J., Chen, B. P. C., and Chen, D. J. (2014)
DNA-PK: a dynamic enzyme in a versatile DSB re-
pair pathway, DNA Repair (Amst), 17, 21-29, https://
doi.org/10.1016/j.dnarep.2014.02.020.
25. Chaplin, A.K., Hardwick, S.W., Stavridi, A.K., Buehl,
C.J., Goff, N.J., Ropars,V., Liang,S., De Oliveira, T.M.,
Chirgadze, D. Y., Meek, K., Charbonnier, J.-B., and
Blundell, T.L. (2021) Cryo-EM of NHEJ supercomplexes
provides insights into DNA repair, Mol. Cell, 81, 3400-
3409.e3, https://doi.org/10.1016/j.molcel.2021.07.005.
26. DeFazio, L. G., Stansel, R. M., Griffith, J. D., and
Chu, G. (2002) Synapsis of DNA ends by DNA-depen-
dent protein kinase, EMBO J., 21, 3192-3200, https://
doi.org/10.1093/emboj/cdf299.
27. Graham, T.G.W., Walter, J.C., and Loparo, J.J. (2016)
Two-stage synapsis of DNA ends during non-homol-
ogous end joining, Mol. Cell, 61, 850-858, https://
doi.org/10.1016/j.molcel.2016.02.010.
28. Liu,L., Chen,X., Li,J., Wang,H., Buehl, C.J., Goff, N.J.,
Meek, K., Yang, W., and Gellert, M. (2022) Autophos-
phorylation transforms DNA-PK from protecting to
processing DNA ends, Mol. Cell, 82, 177-189.e4, https://
doi.org/10.1016/j.molcel.2021.11.025.
29. Chen, S., Lee, L., Naila, T., Fishbain, S., Wang, A.,
Tomkinson, A. E., Lees-Miller, S. P., and He, Y. (2021)
Structural basis of long-range to short-range synap-
tic transition in NHEJ, Nature, 593, 294-298, https://
doi.org/10.1038/s41586-021-03458-7.
30. Stinson, B.M., Moreno, A.T., Walter, J.C., and Loparo,
J. J. (2020) A mechanism to minimize errors during
non-homologous end joining, Mol. Cell, 77, 1080-1091,
https://doi.org/10.1016/j.molcel.2019.11.018.
31. Jiang,W., Crowe, J.L., Liu,X., Nakajima,S., Wang,Y.,
Li,C., Lee, B.J., Dubois, R.L., Liu,C., Yu,X., Lan,L., and
Zha, S. (2015) Differential phosphorylation of DNA-
PKcs regulates the interplay between end-processing
and end-ligation during nonhomologous end-joining,
DOUBLE-STRAND DNA BREAK REPAIR AND TRANSLOCATIONS 1477
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
Mol. Cell, 58, 172-185, https://doi.org/10.1016/
j.molcel.2015.02.024.
32. Menon,V., and Povirk, L.F. (2016) End-processing nu-
cleases and phosphodiesterases: An elite supporting
cast for the non-homologous end joining pathway
of DNA double-strand break repair, DNA Repair, 43,
57-68, https://doi.org/10.1016/j.dnarep.2016.05.011.
33. Moon, A.F., Garcia-Diaz, M., Bebenek,K., Davis, B.J.,
Zhong,X., Ramsden, D.A., Kunkel, T.A., and Pedersen,
L.C. (2007) Structural insight into the substrate spec-
ificity of DNA Polymerase mu, Nat. Struct. Mol. Biol.,
14, 45-53, https://doi.org/10.1038/nsmb1180.
34. Chang, H. H. Y., Watanabe, G., Gerodimos, C. A.,
Ochi, T., Blundell, T. L., Jackson, S. P., and Lieber,
M.R. (2016) Different DNA end configurations dictate
which NHEJ components are most important for join-
ing efficiency, J.Biol. Chem., 291, 24377-24389, https://
doi.org/10.1074/jbc.M116.752329.
35. Ma, Y., Pannicke, U., Schwarz, K., and Lieber, M. R.
(2002) Hairpin opening and overhang processing by
an Artemis/DNA-dependent protein kinase complex
in nonhomologous end joining and V(D)J recom-
bination, Cell, 108, 781-794, https://doi.org/10.1016/
S0092-8674(02)00671-2.
36. Volk, T., Pannicke, U., Reisli, I., Bulashevska, A.,
Ritter, J., Björkman, A., Schäffer, A. A., Fliegauf, M.,
Sayar, E. H., Salzer, U., Fisch,P., Pfeifer, D., Di Virgil-
io, M., Cao, H., Yang, F., Zimmermann, K., Keles, S.,
Caliskaner, Z., Güner, S., Schindler, D., Hammar-
ström,L., Rizzi,M., Hummel,M., et al. (2015) DCLRE1C
(ARTEMIS) mutations causing phenotypes ranging
from atypical severe combined immunodeficiency
to mere antibody deficiency, Hum. Mol. Genet., 24,
7361-7372, https://doi.org/10.1093/hmg/ddv437.
37. Pannunzio, N. R., Watanabe, G., and Lieber, M. R.
(2018) Nonhomologous DNA end-joining for repair of
DNA double-strand breaks, J.Biol. Chem., 293, 10512-
10523, https://doi.org/10.1074/jbc.TM117.000374.
38. Goff, N.J., Mikhova, M., Schmidt, J. C., and Meek, K.
(2024) DNA-PK: A synopsis beyond synapsis, DNA
Repair (Amst), 141, 103716, https://doi.org/10.1016/
j.dnarep.2024.103716.
39. Stinson, B.M., and Loparo, J.J. (2021) Repair of DNA
Double-Strand Breaks by the Nonhomologous End
Joining Pathway, Annu. Rev. Biochem., 90, 137-164,
https://doi.org/10.1146/annurev-biochem-080320-110356.
40. Boulton, S. J., and Jackson, S. P. (1996) Saccharo-
myces cerevisiae Ku70 potentiates illegitimate DNA
double-strand break repair and serves as a barrier
to error-prone DNA repair pathways, EMBO J., 15,
5093-5103, https://doi.org/10.1002/j.1460-2075.1996.
tb00890.x.
41. Mason, R. M., Thacker, J., and Fairman, M. P. (1996)
The joining of non-complementary DNA double-strand
breaks by mammalian extracts, Nucleic Acids Res.,
24, 4946-4953, https://doi.org/10.1093/nar/24.24.4946.
42. Zhu,C., Mills, K.D., Ferguson, D.O., Lee,C., Manis,J.,
Fleming,J., Gao,Y., Morton, C.C., and Alt, F.W. (2002)
Unrepaired DNA breaks in p53-deficient cells lead to
oncogenic gene amplification subsequent to trans-
locations, Cell, 109, 811-821, https://doi.org/10.1016/
S0092-8674(02)00770-5.
43. Bennardo, N., Cheng, A., Huang, N., and Stark, J. M.
(2008) Alternative-NHEJ is a mechanistically distinct
pathway of mammalian chromosome break repair,
PLOS Genet., 4, e1000110, https://doi.org/10.1371/
journal.pgen.1000110.
44. Sfeir, A., Tijsterman, M., and McVey, M. (2024) Mi-
crohomology-mediated end-joining chronicles: trac-
ing the evolutionary footprints of genome protec-
tion, Ann. Rev. Cell Dev. Biol., 40, 195-218, https://
doi.org/10.1146/annurev-cellbio-111822-014426.
45. Schimmel,J., Van Schendel,R., Den Dunnen, J.T., and
Tijsterman, M. (2019) Templated insertions: Asmok-
ing gun for polymerase Theta-mediated end joining,
Trends Genet., 35, 632-644, https://doi.org/10.1016/
j.tig.2019.06.001.
46. Bian,L., Meng,Y., Zhang,M., and Li,D. (2019) MRE11-
RAD50-NBS1 complex alterations and DNA damage
response: implications for cancer treatment, Mol. Can-
cer, 18, 169, https://doi.org/10.1186/s12943-019-1100-5.
47. Conceição, C. J. F., Moe, E., Ribeiro, P. A., and
Raposo, M. (2025) PARP1: A comprehensive review
of its mechanisms, therapeutic implications and
emerging cancer treatments, Biochim. Biophys. Acta
Rev. Cancer, 1880, 189282, https://doi.org/10.1016/
j.bbcan.2025.189282.
48. Staples, C. J., Barone, G., Myers, K. N., Ganesh, A.,
Gibbs-Seymour, I., Patil, A. A., Beveridge, R. D.,
Daye, C., Beniston, R., Maslen, S., Ahel, I., Skehel,
J.M., and Collis, S. J. (2016) MRNIP/C5orf45 Interacts
with the MRN Complex and Contributes to the DNA
Damage Response, Cell Rep., 16, 2565-2575, https://
doi.org/10.1016/j.celrep.2016.07.087.
49. Wang, Y.-L., Zhao, W.-W., Bai, S.-M., Feng, L.-L., Bie,
S.-Y., Gong,L., Wang,F., Wei, M.-B., Feng, W.-X., Pang,
X.-L., Qin, C.-L., Yin, X.-K., Wang, Y.-N., Zhou, W.,
Wahl, D.R., Liu,Q., Chen, M., Hung, M.-C., and Wan,
X.-B. (2022) MRNIP condensates promote DNA dou-
ble-strand break sensing and end resection, Nat.
Commun., 13, 2638, https://doi.org/10.1038/s41467-022-
30303-w.
50. Deshpande, R. A., Myler, L. R., Soniat, M. M., Makha-
rashvili,N., Lee,L., Lees-Miller, S.P., Finkelstein, I.J.,
and Paull, T.T. (2020) DNA-dependent protein kinase
promotes DNA end processing by MRN and CtIP,
Sci. Adv., 6, eaay0922, https://doi.org/10.1126/sciadv.
aay0922.
51. Myler, L.R., Gallardo, I.F., Soniat, M.M., Deshpande,
R. A., Gonzalez, X. B., Kim, Y., Paull, T. T., and Fin-
kelstein, I. J. (2017) Single-molecule imaging reveals
how Mre11-Rad50-Nbs1 initiates DNA break repair,
LOMOV et al.1478
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
Mol. Cell, 67, 891-898.e4, https://doi.org/10.1016/
j.molcel.2017.08.002.
52. Symington, L. S. (2016) Mechanism and regulation
of DNA end resection in eukaryotes, Crit. Rev. Bio-
chem. Mol. Biol., 51, 195-212, https://doi.org/10.3109/
10409238.2016.1172552.
53. Hou, W.-H., Chen, S.-H., and Yu,X. (2019) Poly-ADP ri-
bosylation in DNA damage response and cancer ther-
apy, Mutat. Res., 780, 82-91, https://doi.org/10.1016/
j.mrrev.2017.09.004.
54. Khodyreva, S. N., and Lavrik, O. I. (2016) Poly(ADP-
Ribose) polymerase 1 as a key regulator of DNA re-
pair, Mol. Biol., 50, 580-595, https://doi.org/10.1134/
S0026893316040038.
55. Luedeman, M.E., Stroik,S., Feng,W., Luthman, A.J.,
Gupta, G. P., and Ramsden, D. A. (2022) Poly(ADP)
ribose polymerase promotes DNA polymerase theta-
mediated end joining by activation of end resec-
tion, Nat. Commun., 13, 4547, https://doi.org/10.1038/
s41467-022-32166-7.
56. Özdemir, C., Purkey, L. R., Sanchez, A., and Miller,
K. M. (2024) PARticular MARks: Histone ADP-ribo-
sylation and the DNA damage response, DNA Repair
(Amst), 140, 103711, https://doi.org/10.1016/j.dnarep.
2024.103711.
57. Shima, N., Munroe, R. J., and Schimenti, J. C. (2004)
The mouse genomic instability mutation chaos1 is
an allele of Polq that exhibits genetic interaction
with Atm, Mol. Cell Biol., 24, 10381-10389, https://
doi.org/10.1128/MCB.24.23.10381-10389.2004.
58. Fijen, C., Drogalis Beckham, L., Terino, D., Li, Y.,
Ramsden, D.A., Wood, R.D., Doublié,S., and Rothen-
berg, E. (2024) Sequential requirements for distinct
Polθ domains during theta-mediated end joining,
Mol. Cell, 84, 1460-1474.e6, https://doi.org/10.1016/
j.molcel.2024.03.010.
59. Newman, J. A., Cooper, C. D. O., Aitkenhead, H., and
Gileadi, O. (2015) Structure of the helicase domain
of DNA polymerase theta reveals a possible role in
the microhomology-mediated end-joining pathway,
Structure, 23, 2319-2330, https://doi.org/10.1016/
j.str.2015.10.014.
60. Vanson, S., Li, Y., Wood, R. D., and Doublié,S. (2022)
Probing the structure and function of polymerase θ
helicase-like domain, DNA Repair (Amst), 116, 103358,
https://doi.org/10.1016/j.dnarep.2022.103358.
61. Mateos-Gomez, P.A., Kent,T., Deng, S.K., McDevitt,S.,
Kashkina, E., Hoang, T. M., Pomerantz, R. T., and
Sfeir, A. (2017) The helicase domain of Polθ counter-
acts RPA to promote alt-NHEJ, Nat. Struct. Mol. Biol.,
24, 1116-1123, https://doi.org/10.1038/nsmb.3494.
62. Ito, F., Li, Z., Minakhin, L., Khant, H. A., Pomerantz,
R. T., and Chen, X. S. (2025) Structural basis for
Polθ-helicase DNA binding and microhomology-me-
diated end-joining, Nat. Commun., 16, 3725, https://
doi.org/10.1038/s41467-025-58441-x.
63. Schaub, J. M., Soniat, M. M., and Finkelstein, I. J.
(2022) Polymerase theta-helicase promotes end join-
ing by stripping single-stranded DNA-binding pro-
teins and bridging DNA ends, Nucleic Acids Res., 50,
3911-3921, https://doi.org/10.1093/nar/gkac119.
64. Fleury,H., MacEachern, M.K., Stiefel, C.M., Anand,R.,
Sempeck,C., Nebenfuehr,B., Maurer-Alcalá,K., Ball,K.,
Proctor, B., Belan, O., Taylor, E., Ortega, R., Dodd, B.,
Weatherly, L., Dansoko,D., Leung, J.W., Boulton, S. J.,
and Arnoult,N. (2023) TheAPE2 nuclease is essential
for DNA double-strand break repair by microhomol-
ogy-mediated end joining, Mol. Cell, 83, 1429-1445.e8,
https://doi.org/10.1016/j.molcel.2023.03.017.
65. Stroik, S., Carvajal-Garcia, J., Gupta, D., Edwards, A.,
Luthman, A., Wyatt, D. W., Dannenberg, R. L.,
Feng, W., Kunkel, T. A., Gupta, G. P., Hedglin, M.,
Wood, R., Doublié, S., Rothenberg, E., and Ramsden,
D. A. (2023) Stepwise requirements for polymerases
δ and θ in theta-mediated end joining, Nature, 623,
836-841, https://doi.org/10.1038/s41586-023-06729-7.
66. Carvajal-Garcia, J., Cho, J.-E., Carvajal-Garcia, P.,
Feng,W., Wood, R.D., Sekelsky,J., Gupta, G.P., Roberts,
S.A., and Ramsden, D.A. (2020) Mechanistic basis for
microhomology identification and genome scarring
by polymerase theta, Proc. Natl. Acad. Sci. USA, 117,
8476-8485, https://doi.org/10.1073/pnas.1921791117.
67. Stroik, S., Luthman, A. J., and Ramsden, D. A. (2024)
Templated insertions – DNA repair gets acrobat-
ic, Environ. Mol. Mutagen, 65, 82-89, https://doi.org/
10.1002/em.22564.
68. Kruchinin, A.A., and Makarova, A.V. (2023) Multifac-
eted nature of DNA polymeraseθ, Int.J. Mol. Sci., 24,
3619, https://doi.org/10.3390/ijms24043619.
69. Liang, L., Deng, L., Nguyen, S. C., Zhao, X., Maulion,
C.D., Shao,C., and Tischfield, J.A. (2008) Human DNA
ligases I and III, but not ligase IV, are required for
microhomology-mediated end joining of DNA dou-
ble-strand breaks, Nucleic Acids Res., 36, 3297-3310,
https://doi.org/10.1093/nar/gkn184.
70. Mengwasser, K. E., Adeyemi, R. O., Leng, Y., Choi,
M. Y., Clairmont, C., D’Andrea, A. D., and Elledge,
S. J. (2019) Genetic screens reveal FEN1 and APEX2
as BRCA2 synthetic lethal targets, Mol. Cell, 73, 885-
899.e6, https://doi.org/10.1016/j.molcel.2018.12.008.
71. Llorens-Agost,M., Ensminger,M., Le, H.P., Gawai,A.,
Liu, J., Cruz-García, A., Bhetawal, S., Wood, R. D.,
Heyer, W.-D., and Löbrich, M. (2021) POLθ-mediated
end joining is restricted by RAD52 and BRCA2 until
the onset of mitosis, Nat. Cell Biol., 23, 1095-1104,
https://doi.org/10.1038/s41556-021-00764-0.
72. Heijink, A. M., Stok, C., Porubsky, D., Manolika,
E. M., de Kanter, J. K., Kok, Y. P., Everts, M., de Boer,
H. R., Audrey, A., Bakker, F. J., Wierenga, E., Tijster-
man,M., Guryev,V., Spierings, D.C.J., Knipscheer,P.,
van Boxtel, R., Ray Chaudhuri, A., Lansdorp, P. M.,
and van Vugt, M. A. T. M. (2022) Sister chromatid
DOUBLE-STRAND DNA BREAK REPAIR AND TRANSLOCATIONS 1479
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
exchanges induced by perturbed replication can form
independently of BRCA1, BRCA2 and RAD51, Nat.
Commun., 13, 6722, https://doi.org/10.1038/s41467-
022-34519-8.
73. Ye, G., He, Y., Zhang, Y., Li, D., Liu, F., Li, Y., Ge, Q.,
Guo, Q., Han, S., Song, C., Chang, W., Zhang, H.,
Peng, Q., Sun, K., Ji, W., and Deng, L. (2025) Mitotic
DNA repair by TMEJ suppresses replication stress-
induced nuclear envelope reassembly defect, Nat.
Commun., 16, 8836, https://doi.org/10.1038/s41467-
025-63942-w.
74. Wilson, T.E., Ahmed,S., Winningham,A., and Glover,
T. W. (2024) Replication stress induces POLQ-mediat-
ed structural variant formation throughout common
fragile sites after entry into mitosis, Nat. Commun.,
15, 9582, https://doi.org/10.1038/s41467-024-53917-8.
75. Wu, T., Li, Y., Zhao, Y., Shah, S. B., Shi, L. Z., and
Wu, X. (2025) Break-induced replication is activat-
ed to repair R-loop-associated double-strand breaks
in SETX-deficient cells, Cell Rep., 44, 116386, https://
doi.org/10.1016/j.celrep.2025.116386.
76. Kunihisa, T., Inubushi, S., Tanino, H., and Hoffman,
R.M. (2024) Induction of the DNA-repair gene POLQ
only in BRCA1-mutant breast-cancer cells by methi-
onine restriction, Cancer Genom. Proteom., 21, 399-
404, https://doi.org/10.21873/cgp.20458.
77. Bazan Russo, T.D., Mujacic,C., Di Giovanni,E., Vitale,
M.C., Ferrante Bannera,C., Randazzo,U., Contino,S.,
Bono, M., Gristina, V., Galvano, A., Perez, A., Badala-
menti,G., Russo,A., Bazan,V., and Incorvaia,L. (2024)
Polθ: emerging synthetic lethal partner in homolo-
gous recombination-deficient tumors, Cancer Gene
Ther., 31, 1619-1631, https://doi.org/10.1038/s41417-
024-00815-2.
78. Schrempf, A., Slyskova, J., and Loizou, J. I. (2021)
Targeting the DNA repair enzyme polymerase θ in
cancer therapy, Trends Cancer, 7, 98-111, https://
doi.org/10.1016/j.trecan.2020.09.007.
79. Ackerson, S. M., Romney, C., Schuck, P. L., and Stew-
art, J.A. (2021) Tojoin or not to join: decision points
along the pathway to double-strand break repair vs.
chromosome end protection, Front. Cell Dev. Biol., 9,
708763, https://doi.org/10.3389/fcell.2021.708763.
80. Deshpande, R. A., Marin-Gonzalez, A., Barnes, H. K.,
Woolley, P. R., Ha, T., and Paull, T. T. (2023) Ge-
nome-wide analysis of DNA-PK-bound MRN cleavage
products supports a sequential model of DSB repair
pathway choice, Nat. Commun., 14, 5759, https://
doi.org/10.1038/s41467-023-41544-8.
81. Kumari,N., Kaur,E., Raghavan, S.C., and Sengupta,S.
(2025) Regulation of pathway choice in DNA re-
pair after double-strand breaks, Curr. Opin. Phar-
macol., 80, 102496, https://doi.org/10.1016/j.coph.
2024.102496.
82. Tan,J., Sun,X., Zhao,H., Guan,H., Gao,S., and Zhou,
P.-K. (2023) Double-strand DNA break repair: molec-
ular mechanisms and therapeutic targets, MedComm.
(2020), 4, e388, https://doi.org/10.1002/mco2.388.
83. Truong, L.N., Li,Y., Shi, L.Z., Hwang, P. Y.-H., He, J.,
Wang, H., Razavian, N., Berns, M. W., and Wu, X.
(2013) Microhomology-mediated End joining and
homologous recombination share the initial end re-
section step to repair DNA double-strand breaks in
mammalian cells, Proc. Natl. Acad. Sci. USA, 110,
7720-7725, https://doi.org/10.1073/pnas.1213431110.
84. Chang, H. H. Y., Pannunzio, N. R., Adachi, N., and
Lieber, M. R. (2017) Non-homologous DNA end join-
ing and alternative pathways to double-strand break
repair, Nat. Rev. Mol. Cell Biol., 18, 495-506, https://
doi.org/10.1038/nrm.2017.48.
85. Scully, R., Panday, A., Elango, R., and Willis, N. A.
(2019) DNA double-strand break repair-pathway
choice in somatic mammalian cells, Nat. Rev. Mol.
Cell Biol., 20, 698-714, https://doi.org/10.1038/s41580-
019-0152-0.
86. Sturzenegger, A., Burdova, K., Kanagaraj, R., Leviko-
va, M., Pinto, C., Cejka, P., and Janscak, P. (2014)
DNA2 cooperates with the WRN and BLM RecQ he-
licases to mediate long-range DNA end resection in
human cells, J. Biol. Chem., 289, 27314-27326, https://
doi.org/10.1074/jbc.M114.578823.
87. Ceppi, I., Howard, S. M., Kasaciunaite, K., Pinto, C.,
Anand,R., Seidel,R., and Cejka,P. (2020) CtIP promotes
the motor activity of DNA2 to accelerate long-range
DNA end resection, Proc. Natl. Acad. Sci. USA, 117,
8859-8869, https://doi.org/10.1073/pnas.2001165117.
88. Kciuk, M., Gielecińska, A., Mujwar, S., Mojzych, M.,
and Kontek, R. (2022) Cyclin-dependent kinases in
DNA damage response, Biochim. Biophys. Acta, 1877,
188716, https://doi.org/10.1016/j.bbcan.2022.188716.
89. Hu,Q., Zhao,D., Cui,G., Bhandari,J., Thompson, J.R.,
Botuyan, M. V., and Mer, G. (2024) Mechanisms of
RNF168 nucleosome recognition and ubiquitylation,
Mol. Cell, 84, 839-853.e12, https://doi.org/10.1016/
j.molcel.2023.12.036.
90. Hustedt, N., and Durocher, D. (2017) The control of
DNA repair by the cell cycle, Nat. Cell Biol., 19, 1-9,
https://doi.org/10.1038/ncb3452.
91. Setiaputra, D., Escribano-Diaz, C., Reinert, J. K.,
Sadana,P., Zong, D., Callen,E., Sifri, C., Seebacher,J.,
Nussenzweig, A., Thomä, N. H., and Durocher, D.
(2022) RIF1 acts in DNA repair through phosphopep-
tide recognition of 53BP1, Mol. Cell, 82, 1359-1371.e9,
https://doi.org/10.1016/j.molcel.2022.01.025.
92. Chapman, J. R., Taylor, M. R. G., and Boulton, S. J.
(2012) Playing the end game: DNA double-strand
break repair pathway choice, Mol. Cell, 47, 497-510,
https://doi.org/10.1016/j.molcel.2012.07.029.
93. Isono, M., Niimi, A., Oike, T., Hagiwara, Y., Sato, H.,
Sekine, R., Yoshida, Y., Isobe, S.-Y., Obuse, C.,
Nishi, R., Petricci, E., Nakada, S., Nakano, T., and
Shibata, A. (2017) BRCA1 directs the repair pathway
LOMOV et al.1480
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
to homologous recombination by promoting 53BP1
dephosphorylation, Cell Rep., 18, 520-532, https://
doi.org/10.1016/j.celrep.2016.12.042.
94. Ochs, F., Karemore, G., Miron, E., Brown, J., Sed-
lackova,H., Rask, M.-B., Lampe,M., Buckle,V., Scher-
melleh,L., Lukas,J., and Lukas,C. (2019) Stabilization
of chromatin topology safeguards genome integrity,
Nature, 574, 571-574, https://doi.org/10.1038/s41586-
019-1659-4.
95. Cruz-García, A., López-Saavedra, A., and Huertas, P.
(2014) BRCA1 accelerates CtIP-mediated DNA-end re-
section, Cell Rep., 9, 451-459, https://doi.org/10.1016/
j.celrep.2014.08.076.
96. Yu, X., Fu, S., Lai, M., Baer, R., and Chen, J. (2006)
BRCA1 ubiquitinates its phosphorylation-dependent
binding partner CtIP, Genes. Dev., 20, 1721-1726,
https://doi.org/10.1101/gad.1431006.
97. Ceppi, I., Dello Stritto, M. R., Mütze, M., Braunshi-
er,S., Mengoli,V., Reginato,G., Võ, H.M.P., Jimeno,S.,
Acharya,A., Roy,M., Sanchez,A., Halder,S., Howard,
S. M., Guérois, R., Huertas, P., Noordermeer, S. M.,
Seidel,R., and Cejka,P. (2024) Mechanism of BRCA1–
BARD1 function in DNA end resection and DNA
protection, Nature, 634, 492-500, https://doi.org/
10.1038/s41586-024-07909-9.
98. Densham, R.M., Garvin, A.J., Stone, H.R., Strachan,J.,
Baldock, R. A., Daza-Martin, M., Fletcher, A., Blair-
Reid, S., Beesley, J., Johal, B., Pearl, L. H., Neely, R.,
Keep, N.H., Watts, F. Z., and Morris, J. R. (2016) Hu-
man BRCA1-BARD1 ubiquitin ligase activity coun-
teracts chromatin barriers to DNA resection, Nat.
Struct. Mol. Biol., 23, 647-655, https://doi.org/10.1038/
nsmb.3236.
99. Bohgaki,M., Bohgaki,T., El Ghamrasni,S., Srikumar,T.,
Maire, G., Panier, S., Fradet-Turcotte, A., Stewart,
G. S., Raught, B., Hakem, A., and Hakem, R. (2013)
RNF168 ubiquitylates 53BP1 and controls its response
to DNA double-strand breaks, Proc. Natl. Acad. Sci.
USA, 110, 20982-20987, https://doi.org/10.1073/pnas.
1320302111.
100. Markert,J., Zhou,K., and Luger,K. (2021) SMARCAD1
is an ATP-dependent histone octamer exchange factor
with de novo nucleosome assembly activity, Sci. Adv.,
7, eabk2380, https://doi.org/10.1126/sciadv.abk2380.
101. Lo, C.S.Y., van Toorn,M., Gaggioli, V., Paes Dias,M.,
Zhu, Y., Manolika, E. M., Zhao, W., van der Does, M.,
Mukherjee,C., Souto Gonçalves, J.G.S.C., van Royen,
M. E., French, P. J., Demmers, J., Smal, I., Lans, H.,
Wheeler, D., Jonkers, J., Chaudhuri, A. R., Marteijn,
J. A., and Taneja, N. (2021) SMARCAD1-mediated ac-
tive replication fork stability maintains genome in-
tegrity, Sci. Adv., 7, eabe7804, https://doi.org/10.1126/
sciadv.abe7804.
102. Vergara, X., Manjón, A. G., de Haas, M., Morris, B.,
Schep,R., Leemans,C., Friskes,A., Beijersbergen, R.L.,
Sanders, M. A., Medema, R. H., and van Steensel, B.
(2024) Widespread chromatin context-dependencies
of DNA double-strand break repair proteins, Nat.
Commun., 15, 5334, https://doi.org/10.1038/s41467-
024-49232-x.
103. Arnoult, N., Correia, A., Ma, J., Merlo, A., Garcia-
Gomez, S., Maric, M., Tognetti, M., Benner, C. W.,
Boulton, S.J., Saghatelian,A., and Karlseder,J. (2017)
Regulation of DNA repair pathway choice in S and
G2 phases by the NHEJ inhibitor CYREN, Nature, 549,
548-552, https://doi.org/10.1038/nature24023.
104. Xie,L., Bowman, M.E., Louie, G.V., Zhang,C., Ardejani,
M. S., Huang, X., Chu, Q., Donaldson, C. J., Vaughan,
J.M., Shan,H., Powers, E.T., Kelly, J.W., Lyumkis,D.,
Noel, J. P., and Saghatelian, A. (2023) Biochemistry
and protein interactions of the CYREN microprotein,
Biochemistry, 62, 3050-3060, https://doi.org/10.1021/
acs.biochem.3c00397.
105. Kieffer, S. R., and Lowndes, N. F. (2022) Immedi-
ate-early, early, and late responses to DNA double
stranded breaks, Front. Genet., 13, 793884, https://
doi.org/10.3389/fgene.2022.793884.
106. Ortega, R., Bitler, B. G., and Arnoult, N. (2025) Mul-
tiple functions of PARP1 in the repair of DNA dou-
ble strand breaks, DNA Repair (Amst), 152, 103873,
https://doi.org/10.1016/j.dnarep.2025.103873.
107. Yamashita, S., Tanaka, M., Ida, C., Kouyama, K.,
Nakae,S., Matsuki,T., Tsuda,M., Shirai,T., Kamemu-
ra,K., Nishi,Y., Moss,J., and Miwa,M. (2022) Physio-
logical levels of poly(ADP-ribose) during the cell cycle
regulate HeLa cell proliferation, Exp. Cell Res., 417,
113163, https://doi.org/10.1016/j.yexcr.2022.113163.
108. Liu,C., Vyas,A., Kassab, M.A., Singh, A.K., and Yu,X.
(2017) The role of poly ADP-ribosylation in the first
wave of DNA damage response, Nucleic Acids Res.,
45, 8129-8141, https://doi.org/10.1093/nar/gkx565.
109. Wang, M., Wu, W., Wu, W., Rosidi, B., Zhang, L.,
Wang, H., and Iliakis, G. (2006) PARP-1 and Ku com-
pete for repair of DNA double strand breaks by dis-
tinct NHEJ pathways, Nucleic Acids Res., 34, 6170-
6182, https://doi.org/10.1093/nar/gkl840.
110. Caron, M.-C., Sharma, A.K., O’Sullivan,J., Myler, L.R.,
Ferreira, M. T., Rodrigue, A., Coulombe, Y., Ethier, C.,
Gagné, J.-P., Langelier, M.-F., Pascal, J. M., Finkelstein,
I. J., Hendzel, M. J., Poirier, G. G., and Masson, J.-Y.
(2019) Poly(ADP-ribose) polymerase-1 antagonizes
DNA resection at double-strand breaks, Nat. Com-
mun., 10, 2954, https://doi.org/10.1038/s41467-019-
10741-9.
111. Lodovichi, S., Quadri, R., Sertic, S., and Pellicioli, A.
(2023) PARylation of BRCA1 limits DNA break resec-
tion through BRCA2 and EXO1, Cell Rep., 42, 112060,
https://doi.org/10.1016/j.celrep.2023.112060.
112. Blackford, A. N., and Stucki, M. (2020) How cells
respond to DNA breaks in mitosis, Trends Bio-
chem. Sci., 45, 321-331, https://doi.org/10.1016/j.tibs.
2019.12.010.
DOUBLE-STRAND DNA BREAK REPAIR AND TRANSLOCATIONS 1481
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
113. Rieder, C.L., and Cole, R.W. (1998) Entry into mitosis in
vertebrate somatic cells is guarded by a chromosome
damage checkpoint that reverses the cell cycle
when triggered during early but not late prophase,
J. Cell Biol., 142, 1013-1022, https://doi.org/10.1083/
jcb.142.4.1013.
114. Lindsey-Boltz, L. A., Kemp, M. G., Capp, C., and
Sancar,A. (2015) RHINO forms a stoichiometric com-
plex with the 9-1-1 checkpoint clamp and mediates
ATR-Chk1 signaling, Cell Cycle, 14, 99-108, https://
doi.org/10.4161/15384101.2014.967076.
115. Hara,K., Tatsukawa,K., Nagata,K., Iida,N., Hishiki,A.,
Ohashi,E., and Hashimoto,H. (2024) Structural basis
for intra- and intermolecular interactions on RAD9
subunit of 9-1-1 checkpoint clamp implies functional
9-1-1 regulation by RHINO, J.Biol. Chem., 300, 105751,
https://doi.org/10.1016/j.jbc.2024.105751.
116. Brambati,A., Sacco,O., Porcella,S., Heyza,J., Kareh,M.,
Schmidt, J.C., and Sfeir,A. (2023) RHINO directs MMEJ
to repair DNA breaks in mitosis, Science, 381, 653-660,
https://doi.org/10.1126/science.adh3694.
117. Gelot,C., Kovacs, M.T., Miron,S., Mylne,E., Haan,A.,
Boeffard-Dosierre,L., Ghouil,R., Popova,T., Dingli,F.,
Loew, D., Guirouilh-Barbat, J., Del Nery, E., Zinn-
Justin, S., and Ceccaldi, R. (2023) Polθ is phosphor-
ylated by PLK1 to repair double-strand breaks in
mitosis, Nature, 621, 415-422, https://doi.org/10.1038/
s41586-023-06506-6.
118. Iliaki, S., Beyaert, R., and Afonina, I. S. (2021) Polo-
like kinase 1 (PLK1) signaling in cancer and be-
yond, Biochem. Pharmacol., 193, 114747, https://
doi.org/10.1016/j.bcp.2021.114747.
119. van Vugt, M.A.T.M., and Tijsterman,M. (2023) POLQ
to the rescue for double-strand break repair during
mitosis, Nat. Struct. Mol. Biol., 30, 1828-1830, https://
doi.org/10.1038/s41594-023-01168-4.
120. Aguilera, A., and Gaillard, H. (2014) Transcription
and recombination: when RNA meets DNA, Cold
Spring Harb. Perspect. Biol., 6, a016543, https://
doi.org/10.1101/cshperspect.a016543.
121. Corazzi, L., Ionasz, V. S., Andrejev, S., Wang, L.-C.,
Vouzas, A., Giaisi, M., Di Muzio, G., Ding, B., Marx,
A.J.M., Henkenjohann,J., Allers, M.M., Gilbert, D.M.,
and Wei, P.-C. (2024) Linear interaction between
replication and transcription shapes DNA break dy-
namics at recurrent DNA break clusters, Nat. Com-
mun., 15, 3594, https://doi.org/10.1038/s41467-024-
47934-w.
122. Goehring,L., Huang, T.T., and Smith, D.J. (2023) Tran-
scription-replication conflicts as a source of genome
instability, Annu. Rev. Genet., 57, 157-179, https://
doi.org/10.1146/annurev-genet-080320-031523.
123. Lin, Y., Dent, S. Y. R., Wilson, J. H., Wells, R. D., and
Napierala, M. (2010) R loops stimulate genetic insta-
bility of CTG·CAG repeats, Proc. Natl. Acad. Sci. USA,
107, 692-697, https://doi.org/10.1073/pnas.0909740107.
124. Mehta, A., and Haber, J. E. (2014) Sources of DNA
double-strand breaks and models of recombination-
al DNA repair, Cold Spring Harb. Perspect. Biol., 6,
a016428, https://doi.org/10.1101/cshperspect.a016428.
125. Chaudhuri, J., Basu,U., Zarrin,A., Yan,C., Franco,S.,
Perlot, T., Vuong, B., Wang, J., Phan, R. T., Datta, A.,
Manis, J., and Alt, F. W. (2007) Evolution of the im-
munoglobulin heavy chain class switch recombina-
tion mechanism, Adv. Immunol., 94, 157-214, https://
doi.org/10.1016/S0065-2776(06)94006-1.
126. Schatz, D.G., and Swanson, P.C. (2011) V(D)J Recom-
bination: mechanisms of initiation, Annu. Rev. Gen-
et., 45, 167-202, https://doi.org/10.1146/annurev-genet-
110410-132552.
127. Baudat, F., Manova, K., Yuen, J. P., Jasin, M., and
Keeney, S. (2000) Chromosome synapsis defects and
sexually dimorphic meiotic progression in mice
lacking Spo11, Mol. Cell, 6, 989-998, https://doi.org/
10.1016/S1097-2765(00)00098-8.
128. Romanienko, P. J., and Camerini-Otero, R. D. (2000)
The mouse Spo11 gene is required for meiotic
chromosome synapsis, Mol. Cell, 6, 975-987, https://
doi.org/10.1016/S1097-2765(00)00097-6.
129. Tang, X., Hu, Z., Ding, J., Wu, M., Guan, P., Song, Y.,
Yin, Y., Wu, W., Ma, J., Huang, Y., and Tong, M.-H.
(2025) Invitro reconstitution of meiotic DNA double-
strand-break formation, Nature, 639, 800-807, https://
doi.org/10.1038/s41586-024-08551-1.
130. Champoux, J.J. (2001) DNA topoisomerases: structure,
function, and mechanism, Annu. Rev. Biochem., 70, 369-
413, https://doi.org/10.1146/annurev.biochem.70.1.369.
131. Liu, L.F., Rowe, T.C., Yang,L., Tewey, K.M., and Chen,
G. L. (1983) Cleavage of DNA by mammalian DNA
topoisomerase II, J.Biol. Chem., 258, 15365-15370,
https://doi.org/10.1016/S0021-9258(17)43815-4.
132. Schmidt, B. H., Osheroff, N., and Berger, J. M. (2012)
Structure of a topoisomeraseII–DNA–nucleotide com-
plex reveals a new control mechanism for ATPase
activity, Nat. Struct. Mol. Biol., 19, 1147-1154, https://
doi.org/10.1038/nsmb.2388.
133. Nitiss, J.L. (2009) Targeting DNA topoisomeraseII in
cancer chemotherapy, Nat. Rev. Cancer, 9, 338-350,
https://doi.org/10.1038/nrc2607.
134. Canela,A., Maman,Y., Huang, S.N., Wutz,G., Tang,W.,
Zagnoli-Vieira,G., Callen,E., Wong,N., Day,A., Peters,
J.-M., Caldecott, K. W., Pommier, Y., and Nussenz-
weig, A. (2019) Topoisomerase II-induced chromo-
some breakage and translocation is determined by
chromosome architecture and transcriptional activ-
ity, Mol. Cell, 75, 252-266.e8, https://doi.org/10.1016/
j.molcel.2019.04.030.
135. Robinson, M. J., and Osheroff, N. (1991) Effects of
antineoplastic drugs on the post-strand-passage DNA
cleavage/religation equilibrium of topoisomerase II,
Biochemistry, 30, 1807-1813, https://doi.org/10.1021/
bi00221a012.
LOMOV et al.1482
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
136. Cowell, I. G., Sondka, Z., Smith, K., Lee, K. C., Man-
ville, C. M., Sidorczuk-Lesthuruge, M., Rance, H. A.,
Padget,K., Jackson, G.H., Adachi,N., and Austin, C.A.
(2012) Model for MLL translocations in therapy-relat-
ed leukemia involving topoisomerase IIβ-mediated
DNA strand breaks and gene proximity, Proc. Natl.
Acad. Sci. USA, 109, 8989-8994, https://doi.org/10.1073/
pnas.1204406109.
137. Smith, K. A., Cowell, I. G., Zhang, Y., Sondka, Z., and
Austin, C.A. (2014) Therole of topoisomerase II beta
on breakage and proximity of RUNX1 to partner al-
leles RUNX1T1 and EVI1, Genes Chromosomes Cancer,
53, 117-128, https://doi.org/10.1002/gcc.22124.
138. Zhang,Y., Strissel, P., Strick,R., Chen,J., Nucifora,G.,
Le Beau, M. M., Larson, R. A., and Rowley, J. D.
(2002) Genomic DNA breakpoints in AML1/RUNX1
and ETO cluster with topoisomerase II DNA cleavage
and DNaseI hypersensitive sites in t(8;21) leukemia,
Proc. Natl. Acad. Sci. USA, 99, 3070-3075, https://
doi.org/10.1073/pnas.042702899.
139. Reginato, G., and Cejka, P. (2020) The MRE11 com-
plex: Aversatile toolkit for the repair of broken DNA,
DNA Repair, 91-92, 102869, https://doi.org/10.1016/
j.dnarep.2020.102869.
140. Sasanuma, H., Yamada, S., Tsuda, M., and Takeda, S.
(2020) Restoration of ligatable “clean” double-strand
break ends is the rate-limiting step in the rejoin-
ing of ionizing-radiation-induced DNA breakage,
DNA Repair, 93, 102913, https://doi.org/10.1016/
j.dnarep.2020.102913.
141. Britton,S., Chanut,P., Delteil,C., Barboule,N., Frit,P.,
and Calsou,P. (2020) ATM antagonizes NHEJ proteins
assembly and DNA-ends synapsis at single-ended
DNA double strand breaks, Nucleic Acids Res., 48,
9710-9723, https://doi.org/10.1093/nar/gkaa723.
142. Kockler, Z. W., Osia, B., Lee, R., Musmaker, K., and
Malkova,A. (2021) Repair of DNA breaks by break-in-
duced replication, Annu. Rev. Biochem., 90, 165-191,
https://doi.org/10.1146/annurev-biochem-081420-
095551.
143. Han,J., and Huang,J. (2020) DNA double-strand break
repair pathway choice: the fork in the road, Genome
Instab. Dis., 1, 10-19, https://doi.org/10.1007/s42764-
019-00002-w.
144. Wojtaszek, J. L., and Williams, R. S. (2024) From the
TOP: Formation, recognition and resolution of to-
poisomerase DNA protein crosslinks, DNA Repair
(Amst), 142, 103751, https://doi.org/10.1016/j.dnarep.
2024.103751.
145. Lomov, N. A., Viushkov, V. S., and Rubtsov, M. A.
(2023) Mechanisms of secondary leukemia develop-
ment caused by treatment with DNA topoisomer-
ase inhibitors, Biochemistry (Moscow), 88, 892-911,
https://doi.org/10.1134/S0006297923070040.
146. Riccio, A. A., Schellenberg, M. J., and Williams, R. S.
(2020) Molecular mechanisms of topoisomerase 2
DNA–protein crosslink resolution, Cell. Mol. Life Sci.,
77, 81-91, https://doi.org/10.1007/s00018-019-03367-z.
147. Swan, R. L., Cowell, I. G., and Austin, C. A. (2022)
Mechanisms to repair stalled topoisomerase II-DNA
covalent complexes, Mol. Pharmacol., 101, 24-32,
https://doi.org/10.1124/molpharm.121.000374.
148. Löbrich, M., and Jeggo, P. (2017) A process of resec-
tion-dependent nonhomologous end joining involv-
ing the goddess artemis, Trends Biochem. Sci., 42,
690-701, https://doi.org/10.1016/j.tibs.2017.06.011.
149. Qi,Y., Warmenhoven, J. W., Henthorn, N. T., Ingram,
S. P., Xu, X. G., Kirkby, K. J., and Merchant, M. J.
(2021) Mechanistic modelling of slow and fast NHEJ
dna repair pathways following radiation for G0/G1
normal tissue cells, Cancers, 13, 2202, https://doi.org/
10.3390/cancers13092202.
150. Lieber, M.R. (2010) NHEJ and its backup pathways: re-
lation to chromosomal translocations, Nat. Struct. Mol.
Biol., 17, 393-395, https://doi.org/10.1038/nsmb0410-393.
151. Glukhov, S. I., Rubtsov, M. A., Alexeyevsky, D. A.,
Alexeevski, A.V., Razin, S.V., and Iarovaia, O.V. (2013)
The broken MLL gene is frequently located outside
the inherent chromosome territory in human lym-
phoid cells treated with DNA topoisomeraseII poison
etoposide, PLoS One, 8, e75871, https://doi.org/10.1371/
journal.pone.0075871.
152. Lomov, N.A., Viushkov, V.S., Ulianov, S.V., Gavrilov,
A. A., Alexeyevsky, D. A., Artemov, A. V., Razin, S. V.,
and Rubtsov, M.A. (2022) Recurrent translocations in
topoisomerase inhibitor-related leukemia are deter-
mined by the features of DNA breaks rather than by
the proximity of the translocating genes, Int. J. Mol.
Sci., 23, 9824, https://doi.org/10.3390/ijms23179824.
153. Krawczyk, P.M., Borovski,T., Stap,J., Cijsouw,A., Ten
Cate, R., Medema, J. P., Kanaar, R., Franken, N. A. P.,
and Aten, J.A. (2012) Chromatin mobility is increased
at sites of DNA double-strand breaks, J.Cell Sci., 125,
2127-2133, https://doi.org/10.1242/jcs.089847.
154. Bentley, J., Diggle, C. P., Harnden, P., Knowles, M. A.,
and Kiltie, A. E. (2004) DNA double strand break re-
pair in human bladder cancer is error prone and
involves microhomology-associated end-joining, Nu-
cleic Acids Res., 32, 5249-5259, https://doi.org/10.1093/
nar/gkh842.
155. Simsek,D., and Jasin,M. (2010) Alternative end-join-
ing is suppressed by the canonical NHEJ component
Xrcc4–ligase IV during chromosomal translocation
formation, Nat. Struct. Mol. Biol., 17, 410-416, https://
doi.org/10.1038/nsmb.1773.
156. Ghezraoui,H., Piganeau,M., Renouf,B., Renaud, J.-B.,
Sallmyr,A., Ruis,B., Oh,S., Tomkinson, A.E., Hendrick-
son, E.A., Giovannangeli,C., Jasin,M., and Brunet,E.
(2014) Chromosomal translocations in human cells
are generated by canonical nonhomologous end-join-
ing, Mol. Cell, 55, 829-842, https://doi.org/10.1016/
j.molcel.2014.08.002.
DOUBLE-STRAND DNA BREAK REPAIR AND TRANSLOCATIONS 1483
BIOCHEMISTRY (Moscow) Vol. 90 No. 11 2025
157. Wang, J., Sadeghi, C.A., and Frock, R.L. (2024) DNA-
PKcs suppresses illegitimate chromosome rearrange-
ments, Nucleic Acids Res., 52, 5048-5066, https://
doi.org/10.1093/nar/gkae140.
158. Zhang, Y., and Jasin, M. (2011) An essential role
for CtIP in chromosomal translocation formation
through an alternative end-joining pathway, Nat.
Struct. Mol. Biol., 18, 80-84, https://doi.org/10.1038/
nsmb.1940.
159. Weinstock, D. M., Brunet, E., and Jasin, M. (2007)
Formation of NHEJ-derived reciprocal chromosomal
translocations does not require Ku70, Nat. Cell Biol.,
9, 978-981, https://doi.org/10.1038/ncb1624.
160. Roukos, V., Voss, T. C., Schmidt, C. K., Lee, S., Wang-
sa,D., and Misteli,T. (2013) Spatial dynamics of chro-
mosome translocations in living cells, Science, 341,
660-664, https://doi.org/10.1126/science.1237150.
161. Shmakova, A., Lomov, N., Viushkov, V., Tsfasman, T.,
Kozhevnikova, Y., Sokolova, D., Pokrovsky, V., Syrki-
na, M., Germini, D., Rubtsov, M., and Vassetzky, Y.
(2023) Cell models with inducible oncogenic trans-
locations allow to evaluate the potential of drugs to
favor secondary translocations, Cancer Commun., 43,
154-158, https://doi.org/10.1002/cac2.12370.
162. Chiarle, R., Zhang, Y., Frock, R. L., Lewis, S. M., Mo-
linie, B., Ho, Y.-J., Myers, D. R., Choi, V. W., Compag-
no, M., Malkin, D. J., Neuberg, D., Monti, S., Gial-
lourakis, C.C., Gostissa, M., and Alt, F. W. (2011) Ge-
nome-wide translocation sequencing reveals mech-
anisms of chromosome breaks and rearrangements
in B cells, Cell, 147, 107-119, https://doi.org/10.1016/
j.cell.2011.07.049.
163. Dueva,R., and Iliakis,G. (2013) Alternative pathways
of non-homologous end joining (NHEJ) in genomic in-
stability and cancer, Translat. Cancer Res., 2, 163-177.
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