The genetic exchange within the species provides the recovery of a species specific level of ecological stability that is lowered in particular individuals as a result of the accumulation of mutations in microevolutionary processes. It is supposed that the accumulation of the mutations that decrease organisms' ecological stability leads to the action of truncated selection. A chromosome through generations is not fixed, but rather it is "fluid," having many different combinations of alleles.
This allows nonfunctional less functional alleles to be cleared from a population. If recombination did not occur, then one deleterious mutant allele would cause an entire chromosome to be eliminated from the population. However, with recombination, the mutant allele can be separated from the other genes on that chromosome.
DSBs can trigger profound genomic rearrangements or, in contrast, generate genetic diversity in essential biological processes. In the latter case, programmed DSBs are physiologically produced through controlled cellular enzymes during meiotic differentiation, mating-type switching in Saccharomyces cerevisiae or in V D J and class switch recombination, which ensures the diversity of the immune response reviewed in Haber, ; Jung and Alt, ; Lieber et al.
Two primary strategies are used to repair DSBs: 1 HR, which requires a sequence-homologous partner and, in fact, corresponds to different processes involving both common and distinct mechanisms see below and Figure 1 ; and 2 NHEJ non-homologous end joining , which ligates the DNA ends of a DSB without requiring extended homologies Haber, Note that a highly mutagenic alternative end-joining pathway A-EJ has recently been identified for review Grabarz et al.
Figure 1. A The products of HR. Gene conversion left panel leading to non-reciprocal exchange of a DNA sequence in red. Crossing over right panel : reciprocal exchanges of adjacent sequences black and red.
Note that gene conversion can be associated with or without crossing over. B The double-strand break repair models through HR. Left panel: Gene conversion.
This process tolerates limited imperfect sequence homologies, thus creating heteroduplex intermediates bearing mismatches blue circle. The cruciform junctions Holliday junctions, HJ migrate.
Resolution or dissolution of the HJ occurs in two different orientations black or gray triangles , resulting in gene conversion either with or without crossing over.
Middle panel: Synthesis-dependent strand annealing. Initiation is similar to that of the previous model, but the invading strand de-hybridizes and re-anneals at the other end of the injured molecule; no HJ is formed. Right panel: Break-induced replication BIR. The initiation is similar to that of the previous models, but the synthesis continues over longer distances on the chromosome arms, even reaching the end of the chromosome.
Here, there is neither resolution of the HR nor crossover. C Single-strand annealing SSA. When a double-strand break is generated between two homologous sequences in tandem in the same orientation dotted arrows , an extended single-strand resection a reveals two complementary DNA strands that can hybridize b.
Careful examination of the data from the literature might challenge these assumptions, which requires revisiting the current view. Here, in a reciprocal view, we discuss the accuracy of HR and we present several situations of mutagenesis generated by HR. We conclude that HR is a double-edged sword, which on the one hand controls the equilibrium of genomic stability vs.
The importance of the versatility of HR and its impact on genomic integrity are discussed. Consistently with the implication of HR in genome stability maintenance, mutant cells that are defective in HR show elevated mutagenesis and genetic instability. However, in contrast, HR can appear as a mutagenic process per se , in many situations. Such concepts can be understood when considering the products and molecular mechanisms of HR.
The products of HR are gene conversion GC: non-reciprocal exchange of genetic material associated or not with crossing-over CO: reciprocal exchange of the adjacent sequences Figure 1A. Such products can account for genetic diversity or instability arising through HR. At this point, the HR processes differ in the processing of the intermediates, leading to either gene conversion, associated or not with crossing-over, or to SDSA synthesis-dependent strand annealing and BIR break-induced replication Figure 1B.
In addition, an alternative process SSA, single-strand annealing is also initiated by resection; however, the following step does not require Rad51 nor strand invasion of an intact duplex DNA, but the annealing of two complementary ssDNAs Figure 1C. SSA is a non-conservative process that systematically leads to the deletion of the intervening sequence between the two interacting DNA molecules reviewed in Haber, HR contributes to the robustness of DNA replication by multiple mechanisms Figure 2 and might be viewed as a pathway escorting fork progression reviewed in Costes and Lambert, Figure 2.
HR can act either at replication forks or at replicated chromatids to ensure the completion of chromosome duplication. Second, HR is involved in the recovery of arrested replication forks and has the potential to reassemble a functional replisome.
While the mechanism of origin-independent loading of a replisome by HR has been extensively characterized in bacteria, its counterpart in eukaryotic cells has only recently begun to emerge. Figure 2. Replication-maintenance by homologous recombination. Blue and red lines indicate parental and neo-synthesized strands, respectively.
A Replication-restart following collapse of the replication fork. B Repair of a broken replication fork. Star: DNA damage. Fork passage over a ssDNA nick or gaps in the parental DNA leads to a broken fork, with one of the sister chromatids being disconnected from the fork. Some components of the replisome are thus lost Roseaulin et al.
HR ensures the repair of such broken forks through a mechanism that is thought to be similar to BIR Bosco and Haber, ; Kraus et al. In Xenopus , HR-mediated fork repair leads to the reassembly of a replisome Hashimoto et al. But BIR that requires most of the components of canonic replisomes Lydeard et al. An inter-strand cross-link ICLs is a type of lesion that interferes with the progression of replication forks by preventing the unwinding of the parental DNA. ICLs are cleaved by specific nucleases, thus resulting in a broken fork that is then repaired by HR Long et al.
Many chromosomal elements can behave as fork obstacles, and it remains unclear whether fork breakages occur systematically. For example, DNA-bound proteins represent more than potential sites of fork arrest in budding yeast, and HR efficiently rescues replication forks blocked by protein complexes tightly bound to DNA in fission yeast Ivessa et al.
In this case, replication restart is initiated by the loading of HR factors at ssDNA exposed at blocked forks Mizuno et al. The mechanisms by which HR ensures replication restart remain to be determined. Nevertheless, the resumption of DNA synthesis at inactivated forks via the HR pathway is also mutagenic see below.
Finally, in addition to rescuing DNA synthesis at replication forks, HR is also involved in the stability and protection of forks that are impeded in their progression. HR defects lead to the accumulation of ssDNA gaps at replication forks, perhaps due to an uncoupling between lagging and leading strand synthesis Hashimoto et al.
Additionally, resection of neo-synthesized strands has been observed in mammalian and bacterial HR-deficient cells Courcelle and Hanawalt, ; Schlacher et al. While this fork-stabilizer function of HR during DNA replication appears to be evolutionarily conserved, its importance in ensuring the robustness of DNA replication remains to be established in eukaryotes. Therefore, because HR acts through multiple pathways at the replication fork or in its vicinity, it should play an essential role in protecting cells against spontaneous replication stress and thus against the resulting genetic instability, as discussed below.
In all organisms, HR-deficient cells exhibit a higher level of mutagenesis and genome rearrangements, both spontaneous and upon exposure to exogenous genotoxic agents Quah et al.
Replication stress covers many events that impact the accuracy of DNA replication and then jeopardize chromosome segregation during mitosis. Low levels of replication stress can generate mitotic defects, including anaphase bridges, supernumerary centrosomes and multipolar mitosis, which then lead to uneven chromosome segregation Wilhelm et al.
Because HR plays a pivotal role in the resumption of arrested replication forks, defects in HR should thus reveal endogenous replication stress. Consistently, HR-deficient cells are associated with spontaneous slowed replication fork progression Daboussi et al.
Similarly, fission yeast recombination factors are necessary to ensure successful chromosome segregation following the slowdown of fork progression Bailis et al. These data underline the essential role played by HR in protecting genome stability at the interface between replication and mitosis, as reviewed elsewhere Wilhelm et al. More surprisingly, several reports have noted a type of genome instability mediated by micro-homology in an HR-dependent manner. These types of genetic instability were initially assigned to the error-proneness of end joining.
Consequently, the actual view on the accuracy of HR has been challenged in many reports. The repair of these mismatched structures can transfer sequence polymorphisms and modify the genetic information of the recipient molecule, resulting in an apparent mutagenic event.
Additionally, the DNA synthesis initiated by the invading strand Figure 3A can duplicate a sequence that was absent in the donor molecule and thereby transfer this genetic information, resulting in modifications of the original recipient DNA sequence. Moreover, the resolution of the HR intermediate Holliday junctions can facilitate the exchange of adjacent sequences, leading to genetic rearrangements. Thus, both GC and CO intrinsically possess the capacity to modify genetic information.
This has been used to target gene replacement and gene correction using exogenous DNA. Note that when involving identical sequences for instance sister chromatids exchange: SCE , HR does not impact the genetic information.
Therefore, genome stability should not be strongly impacted by SCEs. In contrast, when involving repeated sequences which are not identical dispersed throughout the genome non-allelic recombination, NAHR , HR can affect the genetic information see below.
Note that, if the final product of an equal SCE is error-free, this is not due to the accuracy of the HR process, but to the fact that the DNA are identical indeed HR can efficiently processes with imperfectly homologous sequences and because associated mechanisms orientate such kinds of events: 1-HR is restricted to the S and G2 phases which correspond to the cell cycle phases presenting sister chromatids and 2-the tight cohesion of the sister chromatids, through the cohesins complex, orientates the event to an equal SCE.
Thus, the structure of the DNA and accessory associated mechanisms, rather than HR itself, favor such an error-free event. In addition, HR can initiate mutagenic DNA synthesis even when the interacting DNA molecules are fully identical such as sister chromatids see discussion below. Finally, we can point out that, in yeast as well as in mammalian cells, spontaneous SCE have been described to be largely independent of the main actors of HR Rad51, Rad52, Rad54 , in contrast with induced SCE Dronkert et al.
Noteworthy, at meiosis, which aims at creating genetic diversity, equal SCEs are repressed and HR between homologous chromosomes which are not identical is favored.
Therefore, in this situation, HR is used to generate genetic diversity. Figure 3. A Copy of one sequence of the donor absent on the recipient molecule. One of two homologous molecules red and black can contain one heterologous sequence blue. Upon gene conversion or SDSA see Figure 1 the heterologous blue sequence can be copied and transferred from the donor sequence red to the homologous recipient sequence black , resulting in a genetic modification of the recipient sequence.
B Sister chromatid exchanges. Between repeat sequences blue boxes without misalignment upper panel or with misalignment resulting in unequal sister chromatid exchanges lower panel and amplification and loss of genetic material. C Impact of gene conversion. Non-reciprocal exchange of genetic information between two heteroalleles, leading to a loss of heterozygosity upper panel and between a pseudogene hatched , which often contains nonsense mutations and a gene in red , leading to the inactivation of the latter lower panel.
D Chromosomal rearrangements resulting from crossing-over CO between repeat sequences. Thus, in the cases discussed above, associated processes, rather than the HR machinery itself, in fact control the accuracy of the final outcome of HR.
Gene conversion is able to transfer genetic information in a non-reciprocal manner between two hetero-alleles, resulting in loss of heterozygosity; gene conversion can also transfer one stop codon from a pseudogene to a related coding sequence, leading to its extinction Figure 3C Amor et al.
Moreover, crossing over between repeated sequences that are dispersed throughout the genome non-allelic HR could lead to genomic rearrangements, such as translocations, deletions, amplifications and inversions Figure 3D. These models account for genome rearrangements responsible for different human pathologies, attesting to the existence of these processes in vivo Purandare and Patel, ; Chen et al. In Saccharomyces cerevisiae , using an intron-based chromosomal translocation assay, it has been reported that DSB-induced translocation occurs via triparental recombination events.
A short homologous sequence in the third chromosome serves as a bridge template for recombination events occurring between two non-homologous chromosomes. Rad59 and Srs2 are also required, although to a lesser extent, whereas KU70 plays no role. These data suggest that BIR-mediated triparental recombination could be a major mechanism for chromosomal translocations in eukaryotic cells Schmidt et al.
Using a newly designed substrate for the analysis of DSB-induced chromosomal translocation, the group of Aguilera shows that Mus81 and Yen1 endonucleases promote BIR, thus causing non-reciprocal translocations.
These endonucleases, as well as Slx4, promote replication template switching during BIR, thus participate in the generation of complex rearrangements when repeated sequences dispersed throughout the genome are involved Pardo and Aguilera, BIR can also induce genome instability in mammalian cells. It was recently reported that replicative stress induced by the overexpression of cyclin E in human cells led to copy number alteration CNA.
The depletion of Pol D3, which encodes a subunit of pol delta, decreases the frequency of these events. The authors propose that BIR repair of damaged replication forks might explain the presence of segmental genomic duplication in human cancers. Replication fork arrest has also been reported to promote non-allelic HR between repeated sequences. In budding yeast, a reduced level of replicative polymerases, which can potentially alter the progression of replication forks, leads to recombination between an inverted Ty element and translocation Lemoine et al.
A more direct connection between fork arrest and HR-mediated genome rearrangements has been established in fission yeast, in which the block of a single replication fork leads to translocation and genomic deletion that results from HR between repeated sequences Lambert et al. Such chromosomal rearrangements are a direct consequence of replication restart at unbroken forks by HR and not a consequence of failure in restarting forks and subsequent aberrant processing Mizuno et al. Given the potential role of HR in mediating chromosomal rearrangement, factors that prevent non-allelic HR might thus be considered as factors protecting against homology-mediated genomic instability.
For example, increasing the distance between repeated sequences reduced the frequency of non-allelic HR Lichten and Haber, ; Godwin et al. Mutagenesis associated with HR was first reported in E. This mechanism limits genetic instability to the stress response and to regions near a DSB, and therefore produces localized mutations rather than dispersed mutations.
This could be an important evolutionary strategy, both for the minimization of deleterious mutations in cells that acquire a rare adaptive mutation and also for concerted evolution within genes and gene clusters reviewed in Rosenberg et al. In addition, it has recently been shown that the DNA synthesis step during elongation of the invading strand is highly mutagenic in Saccharomyces cerevisiae , with the mutation rate increasing by up to fold, and exhibits a mutation signature primarily microhomology-mediated inter-strand template switching.
Indeed, BIR, one of the HR-type processes that are thought to restart replication forks, duplicates DNA over a long distance, even to the end of the chromosome arm, by establishing a replication fork-like structure Figure 1B. Strikingly, in Saccharomyces cerevisiae , DNA synthesis that is induced through BIR is highly inaccurate over the entire path of the replication fork.
The high level of mutation results from the combinatorial effects of an increase of the nucleotide pool induced by the DDR, the uncoupling of DNA synthesis with mismatch repair, and the exposure of ssDNA Deem et al. The migration of the D-loop results in the extrusion of the synthesized DNA and the exposure of a long stretch of ssDNA, which can become a hot spot for lesions leading to mutations Saini et al. One essential role of HR is to reactivate arrested replication forks.
In Schizosaccharomyces pombe , this process is error-prone. As mentioned above, replication restart by HR mediates non-allelic HR. More surprisingly, it also leads to small deletions and duplications flanked by micro-homology. Indeed, replication forks restarted by HR are associated with error-prone DNA synthesis, liable to template switch events at micro-homologies Iraqui et al.
When progressing across small inverted repeats or palindromes, forks recovered by HR are prone to generate large chromosomal inversions Mizuno et al. Indeed, sister chromatids are identical, thus GC cannot transfer mutation and CO will not have any genetic impact. This is done by associating two processes as discussed above : 1 restriction of HR in S and G2 phase and 2 the cohesion of the sister chromatids.
Excess HR can also lead to the accumulation of HR intermediates, which generates genomic instability and cell death Gangloff et al. Thus, HR is a double-edged sword; on the one hand, it protects against genetic instability, but on the other hand, it can trigger cell lethality as well as profound genomic rearrangements and point mutations. Therefore, the HR process should be tightly controlled to avoid unnecessary HR events. Helicases, by destabilizing abortive HR intermediates, protect against the genomic instability generated by HR reviewed in Barber et al.
Additionally, it has been proposed that restricting the initiation of unscheduled HR can also prevent against the accumulation of such toxic HR intermediates. Of note, the fact that protective systems have evolved to counteract excess HR underlines the potential risks of this pathway.
The classical theory of cancer development proposed that cells gradually and randomly accumulate mutations and rearrangements that increase their survival reviewed in Stratton et al. However, recent studies have revealed that critical aspects of cancer development can occur on a much shorter timescale. In a process called chromothripsis from the Greek chromos for chromosome and thripsis, shattering into pieces , tens to thousands of genomic rearrangements occur in one cellular crisis Berger et al.
In kataegis, mutations accumulate in hotspots of hundreds of bases to megabases in a single cell cycle Nik-Zainal et al. Interestingly, both processes are linked to DSB repair events. In chromothripsis, cells undergo tens to thousands of genomic rearrangements clustered into discrete subchromosomal territories, as first described in a small set of tumors Berger et al.
What causes such a dramatic remodeling of the genome is still unknown. However, the implicated regions are sharply circumscribed and this suggests that the original DNA damage occurs during mitosis when DNA is highly condensed. Although several mechanisms have been suggested to explain the clustered rearrangements, the most plausible cause is replicative stress on regions difficult to replicate e.
In particular, replication intermediates that do not expose long stretches of ssDNA and therefore do not activate the checkpoints allow cells to enter mitosis in their presence Chan et al. Revoke Cancel. Keywords Keywords for this Article. Save Cancel. Flag Inappropriate The Content is: Objectionable. Flag Content Cancel.
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