Chi sites are examples of special sites enhancing homologous recombination in their region of the chromosome. Chi, 5′ G‐C‐T‐G‐G‐T‐G‐G3′, is a recognition site for the RecBC enzyme, which nicks DNA near Chi as it unwinds DNA. A molecular model of genetic recombination incorporating these features is reviewed.

Homologous recombination (HR) is required to protect and restart stressed replication forks. Paradoxically, the Mrc1 branch of the S phase checkpoints, which is activated by replicative stress, prevents HR repair at breaks and arrested forks. Indeed, the mechanisms underlying HR can threaten genome integrity if not properly regulated. Thus, understanding how cells avoid genetic instability associated with replicative stress, a hallmark of cancer, is still a challenge. Here I discuss recent results that support a model by which HR (...) responds to replication stress through replicative and repair activities that operate at different stages of the cell cycle (S and G2, respectively) and in distinct subnuclear structures. Remarkably, the replication checkpoint appears to control this scenario by inhibiting the assembly of HR repair centers at stressed forks during S phase, thereby avoiding genetic instability. (shrink)

Eukaryotic genomes harbor a large number of homologous repeat sequences that are capable of recombining. Their potential to disrupt genome stability highlights the need to understand how homologous recombination processes are coordinated. The Saccharomyces cerevisiae Rad1–Rad10 endonuclease performs an essential role in recombination between repeated sequences, by processing 3′ single‐stranded intermediates formed during single‐strand annealing and gene conversion events. Several recent studies have focused on factors involved in Rad1–Rad10‐dependent removal of 3′ nonhomologous tails during homologous (...) recombination, including Msh2–Msh3, Slx4, and the newly identified Saw1 protein. Together, this new work provides a model for how Rad1–Rad10‐dependent end processing is coordinated: Msh2–Msh3 stabilizes and prepares double‐strand/single‐strand junctions for Rad1–Rad10 cleavage, Saw1 recruits Rad1–Rad10 to 3′ tails, and Slx4 mediates crosstalk between the DNA damage checkpoint machinery and Rad1–Rad10. (shrink)

The nuclear pore complex (NPC) is emerging as a center for recruitment of a class of “difficult to repair” lesions such as double‐strand breaks without a repair template and eroded telomeres in telomerase‐deficient cells. In addition to such pathological situations, a recent study by Su and colleagues shows that also physiological threats to genome integrity such as DNA secondary structure‐forming triplet repeat sequences relocalize to the NPC during DNA replication. Mutants that fail to reposition the triplet repeat locus to the (...) NPC cause repeat instability. Here, we review the types of DNA lesions that relocalize to the NPC, the putative mechanisms of relocalization, and the types of recombinational repair that are stimulated by the NPC, and present a model for NPC‐facilitated repair. (shrink)

The function of meiotic recombination has remained controversial, despite recent inroads into mechanisms. Ideas concerning a possible role of recombination in the elimination or efficient incorporation of mutations have been backed by theoretical studies but have lacked empirical support. Recent investigations into the basis for local variations in recombination frequency in yeast have uncovered a strong association between recombination initiation sites and transcriptional regulatory sequences. Other recent studies indicate a strong correlation between transcription and mutation rates (...) in yeast genes. Taken together, these data imply that distributions of recombination and mutation frequencies may be strongly correlated. This suggests that recombination may be targeted to genomic sites of high mutation frequency; such a ‘mutation‐tracking’ function would clearly aid in the shuffling of mutations to break up unfavorable and create favorable allelic combinations. Moreover, recent insights into the mechanism of gene conversion in yeast reveal a very strong inherent bias in favor of alleles on the non‐initiating homolog. Combined with mutation tracking, these findings suggest a novel and general mechanism by which allelic gene conversion may act to eliminate mutations. (shrink)

Mitotic recombination is a repair process which is known to repair double strand breaks and to fill double‐strand gaps by copying a homologous sequence. Meiotic recombination is a process of heteroduplex formation which sometimes generates crossovers. Evidence is presented that the later stages of meiotic recombination have some characteristics of mitotic repair recombination, leading to the conclusion that mismatch repair may be a recombinogenic repair process. The evidence suggests that the recombinational repair process generates hetero‐duplex (...) bubbles which can move. Some bubbles become crossovers. Others cease to exist, perhaps because topoisomerase activity breaks them down. (shrink)

The repair of chromosomal double‐strand breaks (DSBs) by homologous recombination is essential to maintain genome integrity. The key step in DSB repair is the RecA/Rad51‐mediated process to match sequences at the broken end to homologous donor sequences that can be used as a template to repair the lesion. Here, in reviewing research about DSB repair, I consider the many factors that appear to play important roles in the successful search for homology by several homologous recombination (...) mechanisms. (shrink)

Cells mitigate the detrimental consequences of DNA damage on genome stability by attempting high fidelity repair. Homologous recombination templates DNA double‐strand break (DSB) repair on an identical or near identical donor sequence in a process that can in principle access the entire genome. Other physiological processes, such as homolog recognition and pairing during meiosis, also harness the HR machinery using programmed DSBs to physically link homologs and generate crossovers. A consequence of the homology search process by a long (...) nucleoprotein filament is the formation of multi‐invasions (MI), a joint molecule in which the damaged ssDNA has invaded more than one donor molecule. Processing of MI joint molecules can compromise the integrity of both donor sites and lead to their rearrangement. Here, two mechanisms for the generation of rearrangements as a pathological consequence of MI processing are detailed and the potential relevance for non‐allelic homologous recombination discussed. Finally, it is proposed that MI‐induced crossover formation may be a feature of physiological recombination. (shrink)

Mammalian cells frequently depend on homologous recombination (HR) to repair DNA damage accurately and to help rescue stalled or collapsed replication forks. The essence of HR is an exchange of nucleotides between identical or nearly identical sequences. Although HR fulfills important biological roles, recombination between inappropriate sequence partners can lead to translocations or other deleterious rearrangements and such events must be avoided. For example, the recombination machinery must follow stringent rules to preclude recombination between the (...) many repetitive elements in a mammalian genome that share significant but imperfect homology. This paper takes a conceptual approach in addressing the homology requirements for recombination in mammalian genomes as well as the general strategy used by cells to reject recombination between similar but imperfectly matched sequences. A mechanism of heteroduplex rejection that involves the unwinding of recombination intermediates that may form between mismatched sequences is discussed. BioEssays 30:1163–1171, 2008. © 2008 Wiley Periodicals, Inc. (shrink)

The nature of the relationship between the two types of meiotic recombination outcomes, exchange (crossing‐over) and simple gene conversion, has been debated for years. I here propose that these two types of events are not necessarily causally related and hypothesize that the primary role of events detected as simple gene conversion is in the recognition of homology during synapsis.

The recombinational repair of chromosomal double‐strand breaks (DSBs) is of critical importance to all organisms, who devote considerable genetic resources to ensuring such repair is accomplished. In Saccharomyces cerevisiae, DSB‐mediated recombination can be initiated synchronously by the conditional expression of two site‐specific endonucleases, HO or I‐Scel. DNA undergoing recombination can then be extracted at intervals and analyzed. Recombination initiated by meiotic‐specific DSBs can be followed in a similar fashion. This type of ‘in vivo biochemistry’ has been used (...) to describe several discrete steps in two different homologous recombination pathways: gene conversion and single‐strand annealing. The roles of specific proteins during recombination can be established by examining DNA in strains deleted for the corresponding gene. These same approaches are now becoming available for the study of recombination in both higher plants and animals. Physical monitoring can also be used to analyze nonhomologous recombination events, whose mechanisms appear to be conserved from yeast to mammals. (shrink)

Homologous chromosome pairing is required for proper chromosome segregation and recombination during meiosis. The mechanism by which a pair of homologous chromosomes contact each other to establish pairing is not fully understood. When pairing occurs during meiotic prophase in the fission yeast, Schizosaccharomyces pombe, the nucleus oscillates between the cell poles and telomeres remain clustered at the leading edge of the moving nucleus. These meiosis‐specific activities produce movements of telomere‐bundled chromosomes. Several lines of evidence suggest that these (...) movements facilitate homologous chromosome pairing by aligning homologous chromosomes and promoting contact between homologous regions. Since telomere clustering and nuclear or chromosome movements in meiotic prophase have been observed in a wide range of eukaryotic organisms, it is suggested that telomere‐mediated chromosome movements are general activities that facilitate homologous chromosome pairing. BioEssays 23:526–533, 2001. © 2001 John Wiley & Sons, Inc. (shrink)

Replication protein A (RPA) is the main eukaryotic single‐stranded DNA (ssDNA) binding protein, having essential roles in all DNA metabolic reactions involving ssDNA. RPA binds ssDNA with high affinity, thereby preventing the formation of secondary structures and protecting ssDNA from the action of nucleases, and directly interacts with other DNA processing proteins. Here, we discuss recent results supporting the idea that one function of RPA is to prevent annealing between short repeats that can lead to chromosome rearrangements by microhomology‐mediated end (...) joining or the formation of hairpin structures that are substrates for structure‐selective nucleases. We suggest that replication fork catastrophe caused by depletion of RPA could result from cleavage of secondary structures by nucleases, and that failure to cleave hairpin structures formed at DNA ends could lead to gene amplification. These studies highlight the important role RPA plays in maintaining genome integrity. (shrink)

Unlike the most well‐characterized prokaryotic polymerase, E. Coli DNA pol I, none of the eukaryotic polymerases have their own 5′ to 3′ exonuclease domain for nick translation and Okazaki fragment processing. In eukaryotes, FEN‐1 is an endo‐and exonuclease that carries out this function independently of the polymerase molecules. Only seven nucleases have been cloned from multicellular eukaryotic cells. Among these, FEN‐1 is intriguing because it has complex structural preferences; specifically, it cleaves at branched DNA structures. The cloning of FEN‐1 permitted (...) establishment of the first eukaryotic nuclease family, predicting that S. cerevisiae RAD2 (S. pombe Rad13) and its mammalian homolog, XPG, would have similar structural specficity. The FEN‐1 nuclease family includes several similar enzymes encoded by bacteriophages. The crystal structures of two enzymes in the FEN‐1 nuclease family have been solved and they provide a structural basis for the interesting steric requirements of FEN‐1 substrates. Because of their unique structural specificities, FEN‐1 and its family members have important roles in DNA replication, repair and, potentially, recombination. Recently, FEN‐1 was found to specifically associate with PCNA, explaining some aspects of FEN‐1 function during DNA replication and potentially in DNA repair. (shrink)

In a recent publication, d'Alençon et al.(1) presented evidence that a form of non‐homologous DNA recombination involving direct repeats is dependent upon the replication of the DNA. In addition, density‐labeling experiments showed that after recombination was stimulated, progenies were present only in molecules that had undergone complete replication. These observations are consistent with a replicative and not a breakage‐and‐rejoining model for the DNA recombination events. These two models had of course been contrasted many years ago in (...) mechanistic studies of homologous DNA recombination. It is therefore interesting to place the latest findings of d'Alençon et al. in this historical context. (shrink)

DNA junctions are by‐products of recombinational repair, during which a damaged DNA sequence, assisted by RecA filament, invades an intact homologous DNA to form a joint molecule. The junctions are three‐strand or four‐strand depending on how many single DNA strands participate in joint molecules. In E. coli, at least two independent pathways to remove the junctions are proposed to operate. One is via RuvAB‐promoted migration of four‐strand junctions with their subsequent resolution by RuvC. In vivo, RuvAB and RuvC enzymes (...) might work in a single complex, a resolvasome, to clean DNA from used RecA filaments and to resolve four‐strand junctions. An alternative pathway for junction removal could be via RecG‐promoted branch migration of three‐strand junctions, provided that an as yet uncharacterized endonuclease activity incises one of the strands in the joint molecules. (shrink)

The multiple activities of the RecA protein in DNA metabolism have inspired over a decade of research in dozens of laboratories around the world. This effort has nevertheless failed to yield an understanding of the mechanism of several RecA protein‐mediated processes, the DNA strand exchange reactions prominent among them. The major factors impeding progress are the invalid constraints placed upon the problem by attempting to understand RecA protein‐mediated DNA strand exchange within the context of an inappropriate biological paradigm – namely, (...) homologous genetic recombination as a mechanism for generating genetic diversity. In this essay I summarize genetic and biochemical data demonstrating that RecA protein evolved as the central component of a recombinational DNA repair system, with the generation of genetic diversity being a sometimes useful byproduct, and review the major in vitro activities of RecA protein from a repair perspective. While models proposed for both recombination and recombinational repair often make use of DNA strand cleavage and transfer steps that appear to be quite similar, the molecular and thermodynamic requirements of the two processes are very different. The recombinational repair function provides a much more logical and informative framework for thinking about the biochemical properties of RecA and the strand exchange reactions it facilitates. (shrink)

RNA polymerase II is recruited to DNA double‐strand breaks (DSBs), transcribes the sequences that flank the break and produces a novel RNA type that has been termed damage‐induced long non‐coding RNA (dilncRNA). DilncRNAs can be processed into short, miRNA‐like molecules or degraded by different ribonucleases. They can also form double‐stranded RNAs or DNA:RNA hybrids. The DNA:RNA hybrids formed at DSBs contribute to the recruitment of repair factors during the early steps of homologous recombination (HR) and, in this way, (...) contribute to the accuracy of the DNA repair. However, if not resolved, the DNA:RNA hybrids are highly mutagenic and prevent the recruitment of later HR factors. Here recent discoveries about the synthesis, processing, and degradation of dilncRNAs are revised. The focus is on RNA clearance, a necessary step for the successful repair of DSBs and the aim is to reconcile contradictory findings on the effects of dilncRNAs and DNA:RNA hybrids in HR. (shrink)

Homologous recombination (HR) is essential for the accurate repair of DNA double‐strand breaks and damaged replication forks. However, inappropriate or aberrant HR can also result in genome rearrangements. The maintenance of cell viability is, therefore, a careful balancing act between the benefits of HR (the error‐free repair of DNA strand breaks) and the potential detrimental outcomes of HR (chromosomal rearrangements). Two papers have recently provided a mechanistic insight into how HR may be tempered by RecQ helicases to prevent (...) genome instability and diseases that are a consequence of this, such as cancer.1,2 BioEssays 30:291–295, 2008. © 2008 Wiley Periodicals, Inc. (shrink)

Diploid germ cells produce haploid gametes through meiosis, a unique type of cell division. Independent reassortment of parental chromosomes and their recombination leads to ample genetic variability among the gametes. Importantly, new mutations also occur during meiosis, at frequencies much higher than during the mitotic cell cycles. These meiotic mutations are associated with genetic recombination and depend on double‐strand breaks (DSBs) that initiate crossing over. Indeed, sequence variation among related strains is greater around recombination hotspots than elsewhere (...) in the genome, presumably resulting from recombination‐associated mutations. Significantly, enhanced mutagenicity in meiosis may lead to faster divergence during evolution, as germ‐line mutations are the ones that are transmitted to the progeny and thus have an evolutionary impact. The molecular basis for mutagenicity in meiosis may be related to the repair of meiotic DSBs by polymerases, or to the exposure of single‐strand DNA to mutagenic agents during its repair. (shrink)

It has been recently demonstrated that yeast cells are able to partially regress chromosome segregation in telophase as a response to DNA double‐strand breaks (DSBs), likely to find a donor sequence for homology‐directed repair (HDR). This regression challenges the traditional concept that establishes anaphase events as irreversible, hence opening a new field of research in cell biology. Here, the nature of this new behavior in yeast is summarized and the underlying mechanisms are speculated about. It is also discussed whether it (...) can be reproduced in other eukaryotes. Overall, this work brings forwards the need of understanding how cells attempt to repair DSBs when transiting the latest stages of mitosis, i.e., anaphase and telophase. (shrink)

The notion that eukaryotes are ancestrally sexual has been gaining attention. This idea comes in part from the discovery of sets of “meiosis‐specific genes” in the genomes of protists. The existence of these genes has persuaded many that these organisms may be engaging in sex, even though this has gone undetected. The involvement of sex in protists is supported by the view that asexual reproduction results in the accumulation of mutations that would inevitably result in the decline and extinction of (...) such lineages. It is argued that this phenomenon can be obviated by polyploidy and that the “meiosis‐specific genes” are used in other processes, including polyploidy control and homologous recombination, independent of meiosis. These phenomena account for the finding that these genes are expressed in cultures devoid of apparent cell fusion events. Hence, it is also proposed that asexual, and not sexual, reproduction is the ancestral condition. (shrink)

Several DNA-damage detection and repair mechanisms have evolved to repair double-strand breaks induced by mutagens. Later in evolutionary history, DNA single- and double-strand cuts made possible immune diversity by V(D)J recombination and recombination at meiosis. Such cuts are induced endogenously and are highly regulated and controlled. In meiosis, DNA cuts are essential for the initiation of homologous recombination, and for the formation of joint molecule and crossovers. Many proteins that function during somatic DNA-damage detection and repair (...) are also active during homologous recombination. However, their meiotic functions may be altered from their somatic roles through localization, posttranslational modifications and/or interactions with meiosis-specific proteins. Presumably, somatic repair functions and meiotic recombination diverged during evolution, resulting in adaptations specific to sexual reproduction. BioEssays 27:795–808, 2005. © 2005 Wiley Periodicals, Inc. (shrink)

The genomes of virtually all sexually reproducing species contain transposable elements. Although active elements generally transpose more rapidly than they are inactivated by mutation or excision, their number can be kept in check by purifying selection if its effectiveness becomes disproportionately greater as their copy number increases. In sexually reproducing species, such synergistic selection can result from ectopic crossing-over or from homologous recombination under negative epistasis. In addition, there may be controls on transposon activity that are associated with (...) meiosis. Because a sexual lineage that abandons sex must lack such mechanisms, it may be driven to extinction by the unchecked proliferation of deleterious transposons inherited from its sexual progenitor. An important component of the evolutionary advantage of sex over asex may therefore lie in the ability of sex, despite facilitating the spread of deleterious elements within interbreeding populations, also to restrain their intragenomic proliferation. BioEssays 27:76–85, 2005. © 2004 Wiley Periodicals, Inc. (shrink)

Replication forks inevitably stall at damaged DNA in every cell cycle. The ability to overcome DNA lesions is an essential feature of the replication machinery. A variety of specialized polymerases have recently been discovered, which enable cells to replicate past various forms of damage by a process termed translesion synthesis. Alternatively, homologous recombination can be used to restart DNA replication across the lesion. Genetic and biochemical studies have shed light on the impact of these two post‐replication repair pathways (...) in bacteria and yeast. In vertebrates, however, a genetic approach to study post‐replication repair has been compromised because many of the genes involved appear to be essential for embryonic development. We have taken advantage of the chicken cell line DT40 to perform a genetic analysis of translesion synthesis and homologous recombination and to characterize genetic interactions between these two pathways in vertebrates. In this article, we aim to summarize our current understanding of post‐replication repair in DT40 in the perspective of bacterial, yeast and mammalian genetics. BioEssays 26:151–158, 2004. © 2004 Wiley Periodicals, Inc. (shrink)

The RecA family proteins mediate homologous recombination, a ubiquitous mechanism for repairing DNA double‐strand breaks (DSBs) and stalled replication forks. Members of this family include bacterial RecA, archaeal RadA and Rad51, and eukaryotic Rad51 and Dmc1. These proteins bind to single‐stranded DNA at a DSB site to form a presynaptic nucleoprotein filament, align this presynaptic filament with homologous sequences in another double‐stranded DNA segment, promote DNA strand exchange and then dissociate. It was generally accepted that RecA family (...) proteins function throughout their catalytic cycles as right‐handed helical filaments with six protomers per helical turn. However, we recently reported that archaeal RadA proteins can also form an extended right‐handed filament with three monomers per helical turn and a left‐handed protein filament with four monomers per helical turn. Subsequent structural and functional analyses suggest that RecA family protein filaments, similar to the F1‐ATPase rotary motor, perform ATP‐dependent clockwise axial rotation during their catalytic cycles. This new hypothesis has opened a new avenue for understanding the molecular mechanism of RecA family proteins in homologous recombination. BioEssays 30:48–56, 2008. © 2007 Wiley Periodicals, Inc. (shrink)

Cohesin establishes sister‐chromatid cohesion during S phase to ensure proper chromosome segregation in mitosis. It also facilitates postreplicative homologous recombination repair of DNA double‐strand breaks by promoting local pairing of damaged and intact sister chromatids. In G2 phase, cohesin that is not bound to chromatin is inactivated, but its reactivation can occur in response to DNA damage. Recent papers by Koshland's and Sjögren's groups describe the critical role of the known cohesin cofactor Eco1 (Ctf7) and ATR checkpoint kinase (...) in damage‐induced reactivation of cohesin, revealing an intricate mechanism that regulates sister‐chromatid pairing to maintain genome integrity.1,2 BioEssays 30:5–9, 2008. © 2007 Wiley Periodicals, Inc. (shrink)

Mouse embryonic stem cell lines offer an attractive route for introducing rare genetic alternations into the gene pool since the cells can be pre‐screened in culture and the mutations then transmitted into the germline through chimera production. Two applications of this technique seem ideally suited for a genetic analysis of development are enhancer and gene trap screens for loci expressed during gastrulation and production of targeted mutations using homologous recombination. These approaches should greatly increase the number of mouse (...) developmental mutants available and help to elucidate the genetic hierarchy controlling embryogenesis. (shrink)

The eukaryotic genome is packaged into a periodic nucleoprotein structure termed chromatin. The repeating unit of chromatin, the nucleosome, consists of DNA that is wound nearly two times around an octamer of histone proteins. To facilitate DNA‐directed processes in chromatin, it is often necessary to rearrange or to mobilize the nucleosomes. This remodeling of the nucleosomes is achieved by the action of chromatin‐remodeling complexes, which are a family of ATP‐dependent molecular machines. Chromatin‐remodeling factors share a related ATPase subunit and participate (...) in transcriptional regulation, DNA repair, homologous recombination and chromatin assembly. In this review, we provide an overview of chromatin‐remodeling enzymes and discuss two possible mechanisms by which these factors might act to reorganize nucleosome structure. BioEssays 25:1192–1200, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

The main focus of this article is to present the practical aspect of the code rules of variation and the search for a second set of genomic rules, including comparison of sequences to understand how to preserve compatible organisms in danger of extinction and how to generate biodiversity. Three new rules of variation are introduced: 1) homologous recombination, 2) a healthy fertile offspring, and 3) comparison of compatible genomes. The novel search in the natural world for fully compatible (...) genomes capable of homologous recombination is explored by using examples of human polymorphisms and by the production of fertile offspring by crossbreeding. By a rational control of: natural crossbreeding of organisms with compatible genomes (something already happening in nature), the current work focuses on the generation of new varieties after a careful plan. This study is presented within the context of biosemiotics, which studies the processing of information, signalling and signs by living systems. I define a group of organisms having compatible genomes as a single theme: the genomic species or population, able to speak the same molecular language through different accents, with each variety within a theme being a different version of the same book. These studies have a molecular, compatible genetics context. Population and ecosystem biosemiotics will be exemplified by a possible genetic damage capable of causing mutations by breaking the rules of variation through the coordinated patterns of atoms present in the 9/11 World Trade Center contaminated dust (U, Ba, La, Ce, Sr, Rb, K, Mn, Mg, etc.), combination that may be able to overload the molecular quality control mechanisms of the human body. I introduce here the balance of codons in the circular genetic code: 2[1(1)+1(3)+1(4)+4(2)]=2[2(2)+3(4)]. (shrink)

The generation of panels of mutant mice by homologous recombination has greatly increased the ability to assess the function of particular gene products in vivo. The ability to control the developmental stage, the tissue and the nature of the mutation would be an important innovation. A recent report(1) demonstrates that the conservative site‐specific recombination of bacteriophage P1, namely Cre‐lox, can be used successfuly in combination with homologous recombination to generate temporal‐and cell‐restricted mutations in vivo. The (...) technical advance allows a greater flexibility in gene targeting and will have a significant impact on how complex gene functions are studied in vivo. (shrink)

Human DNA can be cloned as yeast artificial chromosomes (YACs), each of which contains several hundred kilobases of human DNA. This DNA can be manipulated in the yeast host using homologous recombination and yeast selectable markers. In relatively few steps it is possible to make virtually any change in the cloned human DNA from single base pair changes to deletions and insertions. In order to study the function of the cloned DNA and the effects of the changes made (...) in the yeast, the human DNA must be transferred back into mammalian cells. Recent experiments indicate that large genes can be transferred from the yeast host to mammalian cells in tissue culture and that the genes are transferred intact and are expressed. Using the same methods it may soon be possible to transfer YAC DNA into the mouse germ line so that the expression and function of genes cloned in YACs can be studied in developing and adult mammalian animals. (shrink)

Current theories of breast cancer progression have been greatly influenced by the development and refinement of mouse transgenic and gene targeting technologies. Early transgenic mouse models confirmed the involvement of oncogenes, previously implicated in human breast cancer, by establishing a causal relationship between overexpression or activation of these genes and mammary tumorigenesis. More recently, the importance of genes located at sites of loss of heterozygosity in human breast cancer have been examined in mice by their targeted disruption via homologous (...) recombination. The union of these two approaches allows the generation of complex animal models that more accurately reflect the multistep nature of human breast cancer. This review will examine how the study of transgenic mice has increased our understanding of the molecular events responsible for oncogenic transformation of the mammary gland. BioEssays 22:554–563, 2000. © 2000 John Wiley & Sons, Inc. (shrink)

It is somewhat ironic that animals that are the prime choice for detailed genetic analysis, such as the fruit fly and the nematode, have thus far been largely refractory to reverse genetic analysis. Their detailed genetic map, and small genome size have made them subjects of ambitious genome analysis projects, but there is still no strategy to introduce desired changes into their genomes by homologous recombination. Some alternative approaches have recently become available; this review describes possibilities and unsolved (...) problems for reverse genetics in the nematode Caenorhabditis elegans. The transposon Tc1 could prove to be very useful for the isolation of knock out mutants, and possibly also for introduction of more subtle alterations. (shrink)

The RecA family proteins mediate homologous recombination, a ubiquitous mechanism for repairing DNA double‐strand breaks (DSBs) and stalled replication forks. Members of this family include bacterial RecA, archaeal RadA and Rad51, and eukaryotic Rad51 and Dmc1. These proteins bind to single‐stranded DNA at a DSB site to form a presynaptic nucleoprotein filament, align this presynaptic filament with homologous sequences in another double‐stranded DNA segment, promote DNA strand exchange and then dissociate. It was generally accepted that RecA family (...) proteins function throughout their catalytic cycles as right‐handed helical filaments with six protomers per helical turn. However, we recently reported that archaeal RadA proteins can also form an extended right‐handed filament with three monomers per helical turn and a left‐handed protein filament with four monomers per helical turn. Subsequent structural and functional analyses suggest that RecA family protein filaments, similar to the F1‐ATPase rotary motor, perform ATP‐dependent clockwise axial rotation during their catalytic cycles. This new hypothesis has opened a new avenue for understanding the molecular mechanism of RecA family proteins in homologous recombination. BioEssays 30:48–56, 2008. © 2007 Wiley Periodicals, Inc. (shrink)

DNA double‐strand breaks (DSBs) are extremely hazardous lesions for all DNA‐bearing organisms and the mechanisms of DSB repair are highly conserved. In the eukaryotic mitotic cell cycle, DSBs are often present following DNA replication while, in meiosis, hundreds of DSBs are generated as a prelude to the reshuffling of the maternally and paternally derived genomes. In both cases, the DSBs are repaired by a process called homologous recombinational repair (HRR), which utilises an intact DNA molecule as the repair template. (...) Mitotic and meiotic HRR are managed by ‘checkpoints’ that inhibit cell division until DSB repair is complete. Here we attempt to summarise the substantial recent progress in understanding the checkpoint management of HRR in mitosis (focussing mainly on mammals) and then go on to use this information as a framework for understanding the presumed checkpoint management of HRR in mammalian meiosis. BioEssays 29:974–986, 2007. © 2007 Wiley Periodicals, Inc. (shrink)

The special properties of the Y chromosome stem form the fact that it is a non‐recombining degenerate derivative of the X chromosome. The absence of homologous recombination between the X and the Y chromosome leads to gradual degeneration of various Y chromosome genes on an evolutionary timescale. The absence of recombination, however, also favors the accumulation of transposable elements on the Y chromosome during its evolution, as seen with both Drosophila and mammalian Y chromosomes. Alongside these processes, (...) the acquisition and amplification of autosomal male benefit genes occur. This review will focus on recent studies that reveal the autosome‐acquired genes on the Y chromosome of both Drosophila and humans. The evolution of the acquired and amplified genes on the Y chromosome is also discussed. Molecular and comparative analyses of Y‐linked repeats in the Drosophila melanogaster genome demonstrate that there was a period of their degeneration followed by a period of their integration into RNAi silencing, which was beneficial for male fertility. Finally, the function of non‐coding RNA produced by amplified Y chromosome genetic elements will be discussed. BioEssays 27:1256–1262, 2005. © 2005 Wiley Periodicals, Inc. (shrink)

Meiotic recombination in eukaryotic cells requires two homologs of E. coli RecA protein, Rad51 and Dmc1. Until recently, the role of Dmc1 in meiotic recombination was mostly attributed to genetic studies as purified Dmc1 was found to be a much weaker recombinase than Rad51 in the test tube. Now, Sehorn and colleagues1 have reported that, like Rad51, human Dmc1 is an efficient recombinase in vitro. Dmc1 forms helical nucleoprotein filaments—the signature of classical recombinases such as Rad51. These observations (...) reveal a high level of similitude between the Dmc1 and the Rad51 family of recombination enzymes in higher eukaryotes. BioEssays 26:1151–1155, 2004. © 2004 Wiley Periodicals, Inc. (shrink)

Homologous centromeres compete for segregation to the secondary oocyte nucleus at female meiosis I. Centromeric repeats also compete with each other to populate centromeres in mitotic cells of the germline and have become adapted to use the recombinational machinery present at centromeres to promote their own propagation. Repeats are not needed at centromeres, rather centromeres appear to be hospitable habitats for the colonization and proliferation of repeats. This is probably an indirect consequence of two distinctive features of centromeric DNA. (...) Centromeres are subject to breakage by the mechanical forces exerted by microtubules and meiotic crossing‐over is suppressed. Centromeric proteins acting in trans are under selection to mitigate the costs of centromeric repeats acting in cis. Collateral costs of mitotic competition at centromeres may help to explain the high rates of aneuploidy observed in early human embryos. (shrink)

Premeiotic association of homologous chromosomes in the yeast, Saccharomyces cerevisiae has been shown, by means of fluorescent in situ hybridization (FISH)(1,2). Time course and mutant studies show that the premeiotic associations are disrupted upon entry into meiosis, to be reestablished shortly before synapsis. The data are consistent with a model in which multiple, unstable interactions bring homologues together, prior to stable joining by recombination(3).

During meiosis, homologous chromosomes must pair in order to permit recombination and correct chromosome segregation to occur. Two recent papers1,2 show that meiotic pairing is also important for correct gene expression during meiosis. They describe data for the filamentous fungus Neurospora crassa that show that a lack of pairing generated by ectopic integration of genes can result in silencing of genes expressed during meiosis. This can result in aberrant meioses whose defects are specific to the function of the (...) unpaired gene. Furthermore, mutations affecting the silencing mechanism have been selected in a gene encoding a putative RNA‐dependent RNA polymerase. This finding indicates the involvement of a meiotic specific post‐transcriptional gene silencing mechanism (PTGS) similar to that observed in vegetative cells in N. crassa and other organisms. Finally, this gene product is essential for normal meiosis, suggesting that RNA‐dependent processes are fundamental to the sexual cycle. BioEssays 25:99–103, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

Mammalian telomeres have evolved safeguards to prevent their recognition as DNA double‐stranded breaks by suppressing the activation of various DNA sensing and repair proteins. We have shown that the telomere‐binding proteins TRF2 and RAP1 cooperate to prevent telomeres from undergoing aberrant homology‐directed recombination by mediating t‐loop protection. Our recent findings also suggest that mammalian telomere‐binding proteins interact with the nuclear envelope to maintain chromosome stability. RAP1 interacts with nuclear lamins through KU70/KU80, and disruption of RAP1 and TRF2 function result (...) in nuclear envelope rupture, promoting telomere‐telomere recombination to form structures termed ultrabright telomeres. In this review, we discuss the importance of the interactions between shelterin components and the nuclear envelope to maintain telomere homeostasis and genome stability. (shrink)

Mammals have an XX:XY system of chromosomal sex determination in which a small heterochromatic Y controls male development. The Y contains the testis determining factor SRY, as well as several genes important in spermatogenesis. Comparative studies show that the Y was once homologous with the X, but has been progressively degraded, and now consists largely of repeated sequences as well as degraded copies of X linked genes. The small original X and Y have been enlarged by cycles of autosomal (...) addition to one partner, recombination onto the other and continuing attrition of the compound Y. This addition–attrition hypothesis predicts that the pseudoautosomal region of the human X is merely the last relic of the latest addition. Genes (including SRY) on the conserved or added region of the Y evolved functions in male sex determination and differentiation distinct from the general functions of their X‐linked partners. Although the gonadogenesis pathway is highly conserved in vertebrates, its control has probably changed radically and rapidly in vertebrate – even mammalian – evolution. (shrink)

Somatic diversification of antigen receptor genes depends on the activity of enzymes whose homologs participate in a mutagenic DNA repair in unicellular species. Indeed, by engaging error-prone polymerases, gap filling molecules and altered mismatch repair pathways, lymphocytes utilize conserved components of genomic stress response systems, which can already be found in bacteria and archaea. These ancient systems of mutagenesis and repair act to increase phenotypic diversity of microbial cell populations and operate to enhance their ability to produce fit variants during (...) stress. Coopted by lymphocytes, the ancient mutagenic processing systems retained their diversification functions instilling the adaptive immune cells with enhanced evolvability and defensive capacity to resist infection and damage. As reviewed here, the ubiquity and conserved character of specialized variation-generating mechanisms from bacteria to lymphocytes highlight the importance of these mechanisms for evolution of life in general. (shrink)

Despite their seemingly endless diversity, proteins adopt a limited number of structural forms. It has been estimated that 80% of proteins will be found to adopt one of only about 400 folds, most of which are already known. These folds are largely formed by a limited ‘vocabulary’ of recurring supersecondary structure elements, often by repetition of the same element and, increasingly, elements similar in both structure and sequence are discovered. This suggests that modern proteins evolved by fusion and recombination (...) from a more ancient peptide world and that many of the core folds observed today may contain homologous building blocks. The peptides forming these building blocks would not in themselves have had the ability to fold, but would have emerged as cofactors supporting RNA‐based replication and catalysis (the ‘RNA world’). Their association into larger structures and eventual fusion into polypeptide chains would have allowed them to become independent of their RNA scaffold, leading to the evolution of a novel type of macromolecule: the folded protein. BioEssays 25:837–846, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

Enzymatic misrepair of ionizing‐radiation‐induced DNA damage can produce large‐scale rearrangements of the genome, such as translocations and dicentrics. These and other chromosome exchange aberrations can cause major phenotypic alterations, including cell death, mutation and neoplasia. Exchange formation requires that two (or more) genomic loci come together spatially. Consequently, the surprisingly rich aberration spectra uncovered by recently developed techniques, when combined with biophysically based computer modeling, help characterize large‐scale chromatin architecture in the interphase nucleus. Most results are consistent with a picture (...) whereby chromosomes are mainly confined to territories, chromatin motion is limited, and interchromosomal interactions involve mainly territory surfaces. Aberration spectra and modeling also help characterize DNA repair/misrepair mechanisms. Quantitative results for mammalian cells are best described by a breakage‐and‐reunion model, suggesting that the dominant recombinational mechanism during the G0/G1 phase of the cell cycle is non‐homologous end‐joining of radiogenic DNA double strand breaks. In turn, better mechanistic and quantitative understanding of aberration formation gives new insights into health‐related applications. BioEssays 24:714–723, 2002. © 2002 Wiley Periodicals, Inc. (shrink)

Since intelligent design (ID) advocates claimed the ubiquitous mouse trap as an example of systems that cannot have evolved, mouse trap history is doubly relevant to studying material culture. On the one hand, debunking ID claims about mouse traps and, by implication, also about other irreducibly complex systems has a high educational value. On the other hand, a case study of mouse trap history may contribute insights to the academic discussion about material culture evolution. Michael Behe argued that mouse traps (...) cannot trap mice with any part missing; therefore, they cannot have a precursor with one part less, therefore, cannot have a continuous history, and therefore, cannot have evolved. The patented and seminal precursor of current flat snap traps, however, had one part less, because spring and striker were formed of one wire. Secondly, historical records that reach back into the Bronze Age suggest that its history continued for a very long time. Thirdly, all prerequisites for evolution (variation, transmission, and selection) abound in mouse trap populations. Hence, Behe’s triple-jump conclusion about mouse traps is false each step. There is no, in principle, impossibility for mouse traps to evolve. An evolutionary account of mouse trap history also has academic merits beyond its educational value. Three important conclusions can be drawn: (1) reticulate phylogenies of artifact systems may be resolvable as overlapping, but branching, phylogenies of parts; (2) homologous ideas may be realized by analogous material, that is, phylogenies of information do not necessarily coincide with those of material parts; (3) recombination of parts between different artifact systems increases the cumulative nature of cultural evolution. (shrink)

Chromosome transmission in S. cerevisiae requires the activities of many structural and regulatory proteins required for the replication, repair, recombination and segregation of chromosomal DNA, and co‐ordination of the chromosome cycle with progression through the cell cycle. An important structural domain on each chromosome is the kinetochore (centromere DNA and associated proteins), which provides the site of attachment of chromosomes to the spindle microtubules. Stoler et al.(1) have recently reported the cloning of an essential gene CSE4, mutations in which (...) cause chromosome nondisjunction of a marked chromosome bearing a centromere DNA mutation. The cse4–1 mutation causes cells to arrest in the G2/M phase of the cell cycle with a 2N DNA content in a RAD9 checkpoint‐independent manner. The carboxyl terminus of Cse4p and the human centromere‐localized protein CENP‐A have a high degree of homology to the C‐terminal domain of histone H3. Since both CENP‐A and Cse4p also have biochemical properties similar to histones H3 and H4, it is tempting to speculate that these histone H3‐like proteins are components of specialized nucleosomes, a class of which may be unique to the centromeres. (shrink)

Type II DNA topoisomerases (Topo II) are essential enzymes implicated in key nuclear processes. The recent discovery of a novel kind of Topo II (DNA topoisomerase VI) in Archaea led to a division of these enzymes into two non‐homologous families, (Topo IIA and Topo IIB) and to the identification of the eukaryotic protein that initiates meiotic recombination, Spo11. In the present report, we have updated the distribution of all Topo II in the three domains of life by a (...) phylogenomic approach. Both families exhibit an atypical distribution by comparison with other informational proteins, with predominance of Topo IIA in Bacteria, Eukarya and viruses, and Topo IIB in Archaea. However, plants and some Archaea contain Topo II from both families. We confront this atypical distribution with current hypotheses on the evolution of the three domains of life and origin of DNA genomes. BioEssays 25:232–242, 2003. © 2003 Wiley Periodicals, Inc. (shrink)