Ectopic Recombination Research Articles

Real-Time Evolution of a Subtelomeric Gene Family in Candida albicans.

Subtelomeric regions of the genome are notable for high rates of sequence evolution and rapid gene turnover. Evidence of subtelomeric evolution has relied heavily on comparisons of historical evolutionary patterns to infer trends and frequencies of these events. Here, we describe evolution of the subtelomeric TLO gene family in Candida albicans during laboratory passaging for over 4000 generations. C. albicans is a commensal and opportunistic pathogen of humans and the TLO gene family encodes a subunit of the Mediator complex that regulates transcription and affects a range of virulence factors. We identified 16 distinct subtelomeric recombination events that altered the TLO repertoire. Ectopic recombination between subtelomeres on different chromosome ends occurred approximately once per 5000 generations and was often followed by loss of heterozygosity, resulting in the complete loss of one TLO gene sequence with expansion of another. In one case, recombination within TLO genes produced a novel TLO gene sequence. TLO copy number changes were biased, with some TLOs preferentially being copied to novel chromosome arms and other TLO genes being frequently lost. The majority of these nonreciprocal recombination events occurred either within the 3′ end of the TLO coding sequence or within a conserved 50-bp sequence element centromere-proximal to TLO coding sequence. Thus, subtelomeric recombination is a rapid mechanism of generating genotypic diversity through alterations in the number and sequence of related gene family members.

Transposable Elements in Fungi: A Genomic Approach

Transposable elements (TEs) include a wide range of DNA sequences that can change positions in the genome. The accessibility of whole fungal genome sequences and analysis of TEs demonstrate the important role they play in genome evolution of fungi species. TE activity is a primary mechanism for high fitness, plasticity and adaptability in certain species of pathogenic fungi. Some of the deleterious effects of transposons may be due to ectopic recombination among transposons of the same family. To prevent possible damage caused by the transposons, some fungi possess TE-silencing mechanisms, for instance, RIP (Repeat Induced Pont mutation) and RNA silencing. In addition, TEs are efficient molecular markers due to their structure and transposition strategy. Copies or remnants of both class I (retrotransposons) and class II (DNA transposons) have been identified, and the number of distinct mobile elements continues to grow. Such repeated sequences vary in number as well as in size, and comprise 3 to 20% of the sequenced genome of most fungi. In contrast, the plant pathogenic fungus Blumeria graminis has a genome estimated at 174 Mb, and 85% of the genome includes TEs [3]. More than 215 genus-specific TEs were found in Laccaria bicolor in addition to many remaining degenerate copies. Additionally, 40 different TE families were detected in the genome of L. bicolor, and less than 5% nucleotide mutations had accumulated, which suggest that they are recently acquired elements, and possible activities can be inferred for such elements [4]. In general, the most abundant TEs in sequenced genomes of fungi are Gypsy, Copia and Tc1-Mariner (Figure 1).

The good, the bad, and the ugly: How to protect chromosome stability from potential threats

The maintenance of chromosome stability depends on the precise duplication of the genome, efficient DNA repair and recombination, and proper interactions between the centromeres/kinetochores of the duplicated chromosomes and the spindle apparatus (Fig. ​(Fig.1).1). Errors in these processes lead to genomic instability and aneuploidy—a hallmark of many diseases, including cancers. Knowledge of these processes at the molecular level is also essential for understanding principles of genome evolution, organization, and variability. We organized a conference on chromosome stability at the Jawaharlal Nehru Centre for Advanced Scientific Research (JNCASR) in Bangalore, India to facilitate interactions between the fast growing Indian community interested in areas related to chromosome stability with investigators from other countries. The meeting featured talks from 40 principal investigators and postdoctoral fellows representing India, the United States of America (USA), Europe, and Japan. Approximately 80 graduate and undergraduate students from India participated in the meeting. We present here highlights of the meeting. Figure 1 Aspects of chromosome stability during mitosis and meiosis discussed during the meeting. A diploid cell containing a homologous pair of chromosomes (red and green) is shown at the lower most step of the DNA ladder. As a cell progresses through the cell ... DNA repair and recombination pathways Accurate DNA repair and recombination is critical for chromosome stability. Jim Haber (Brandeis University, USA) discussed genome stability during repair of a broken chromosome in budding yeast. Double‐strand breaks (DSBs) can be repaired by nonhomologous end‐joining (NHEJ) or by homologous recombination in which a intact, identical, or nearly identical sequence is used as a template. However, even conservative repair of DSBs by homologous recombination is associated with a highly elevated risk of mutation. Such mutations often have a “signature” associated with slippage or template switching of the repair DNA polymerases. His group showed that pairs of such inter‐chromosomal template switching events can occur between ectopic (non‐allelic) homologous sequences as often as once every 100 repair events. These types of instability associated with ectopic recombination may underlie some of the complex rearrangements seen in human developmental diseases or in cancer cell chromosomes exhibiting chromothripsis in which individual chromosomes are shattered and then reassembled. Eric Alani (Cornell University, USA) continued with the theme of homologous recombination between diverged DNA sequence in yeast. He discussed how cells make the decision to either unwind and reject recombination intermediates containing mismatches (heteroduplex rejection), or to maintain the intermediates and correct the mismatches. He showed that overexpression of the mismatch repair protein Msh6 can decrease or even stimulate heteroduplex rejection, depending on whether the recombination occurs in the context of a replication fork or independent of it. Thus, even though higher levels of Msh6 may improve recombination fidelity in general, this protein appears to reduce recombination fidelity during replication. The identification of genomic regions prone to instability due to mitotic homologous recombination is important. Tom Petes (Duke University, USA) described evidence that spontaneous mitotic crossovers in yeast are often initiated by a break in unreplicated DNA and presented genome‐wide mapping of mitotic crossovers in yeast. Analysis of mitotic crossover hotspots showed that they are often associated with closely‐linked inverted repeats. Jennifer Surtees (University at Buffalo, USA) summarized her analysis of the mismatch repair complex (Msh2/Msh3) on expansions of trinucleotide repeat (TNR) tracts in yeast. She showed that msh3 mutations result in a reduction in tract expansions, and proposed a model by which the Msh2/Msh3 complex stabilized small DNA loops to promote tract expansion. These results demonstrate unforeseen consequences of active MMR in the context of TNR tracts and have important implications for understanding diseases caused by TNR expansions. The above talks discussed the potential of homologous recombination to generate genome rearrangements and instability. Wolf Heyer (University of California‐Davis, USA) discussed how recombination has evolved as a pathway with metastable, reversible intermediates to avoid inappropriate DNA strand invasions in non‐allelic targets and thus minimize the potential for genomic instability. Using in vitro assays, he demonstrated how the anti‐recombination helicase Srs2 dissociates the strand invasion catalysed by Rad51‐ssDNA filaments, whereas Topoisomerase 3 reverses D‐loops exerting anti‐recombination activity. These results suggest that recombination and repair pathways should not be seen as a fixed sequence of events, rather as displaying reversibility at multiple steps. Pathway reversibility creates novel circumstances where synthetic lethality can potentially be exploited in targeting recombination pathways during anti‐cancer therapy. Sathees Raghavan (IISc, India) discussed DNA damage and chromosomal instability in mammalian cells with a focus on the less‐studied mitochondrial DSB repair pathway. He showed that Microhomology Mediated End Joining (MMEJ) or alternative‐NHEJ are the preferred pathways for rejoining DNA with DSBs in mitochondria.

Low levels of LTR retrotransposon deletion by ectopic recombination in the gigantic genomes of salamanders.

Across the tree of life, species vary dramatically in nuclear genome size. Mutations that add or remove sequences from genomes-insertions or deletions, or indels-are the ultimate source of this variation. Differences in the tempo and mode of insertion and deletion across taxa have been proposed to contribute to evolutionary diversity in genome size. Among vertebrates, most of the largest genomes are found within the salamanders, an amphibian clade with genome sizes ranging from ~14 to ~120 Gb. Salamander genomes have been shown to experience slower rates of DNA loss through small (i.e., <30 bp) deletions than do other vertebrate genomes. However, no studies have addressed DNA loss from salamander genomes resulting from larger deletions. Here, we focus on one type of large deletion-ectopic-recombination-mediated removal of LTR retrotransposon sequences. In ectopic recombination, double-strand breaks are repaired using a "wrong" (i.e., ectopic, or non-allelic) template sequence-typically another locus of similar sequence. When breaks occur within the LTR portions of LTR retrotransposons, ectopic-recombination-mediated repair can produce deletions that remove the internal transposon sequence and the equivalent of one of the two LTR sequences. These deletions leave a signature in the genome-a solo LTR sequence. We compared levels of solo LTRs in the genomes of four salamander species with levels present in five vertebrates with smaller genomes. Our results demonstrate that salamanders have low levels of solo LTRs, suggesting that ectopic-recombination-mediated deletion of LTR retrotransposons occurs more slowly than in other vertebrates with smaller genomes.

Structural and sequence diversity of the transposon Galileo in the Drosophila willistoni genome.

BackgroundGalileo is one of three members of the P superfamily of DNA transposons. It was originally discovered in Drosophila buzzatii, in which three segregating chromosomal inversions were shown to have been generated by ectopic recombination between Galileo copies. Subsequently, Galileo was identified in six of 12 sequenced Drosophila genomes, indicating its widespread distribution within this genus. Galileo is strikingly abundant in Drosophila willistoni, a neotropical species that is highly polymorphic for chromosomal inversions, suggesting a role for this transposon in the evolution of its genome.ResultsWe carried out a detailed characterization of all Galileo copies present in the D. willistoni genome. A total of 191 copies, including 133 with two terminal inverted repeats (TIRs), were classified according to structure in six groups. The TIRs exhibited remarkable variation in their length and structure compared to the most complete copy. Three copies showed extended TIRs due to internal tandem repeats, the insertion of other transposable elements (TEs), or the incorporation of non-TIR sequences into the TIRs. Phylogenetic analyses of the transposase (TPase)-encoding and TIR segments yielded two divergent clades, which we termed Galileo subfamilies V and W. Target-site duplications (TSDs) in D. willistoni Galileo copies were 7- or 8-bp in length, with the consensus sequence GTATTAC. Analysis of the region around the TSDs revealed a target site motif (TSM) with a 15-bp palindrome that may give rise to a stem-loop secondary structure.ConclusionsThere is a remarkable abundance and diversity of Galileo copies in the D. willistoni genome, although no functional copies were found. The TIRs in particular have a dynamic structure and extend in different ways, but their ends (required for transposition) are more conserved than the rest of the element. The D. willistoni genome harbors two Galileo subfamilies (V and W) that diverged ~9 million years ago and may have descended from an ancestral element in the genome. Galileo shows a significant insertion preference for a 15-bp palindromic TSM.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-792) contains supplementary material, which is available to authorized users.

Evidence for the persistence of an active endogenous retrovirus (ERVE) in humans.

Transposable elements (TEs) account for nearly half (44 %) of the human genome. However, their overall activity has been steadily declining over the past 35-50 million years, so that <0.05 % of TEs are presumably still "alive" (potentially transposable) in human populations. All the active elements are retrotransposons, either autonomous (LINE-1 and possibly the endogenous retrovirus ERVK), or non-autonomous (Alu and SVA, whose transposition is dependent on the LINE-1 enzymatic machinery). Here we show that a lineage of the endogenous retrovirus ERVE was recently engaged in ectopic recombination events and may have at least one potentially fully functional representative, initially reported as a novel retrovirus isolated from blood cells of a Chinese patient with chronic myeloid leukemia, which bears signals of positive selection on its envelope region. Altogether, there is strong evidence that ERVE should be included in the short list of potentially active TEs, and we give clues on how to identify human specific insertions of this element that are likely to be segregating in some of our populations.

Interspecific hybridization as a genomic stressor inducing mobilization of transposable elements in Drosophila.

Transposable elements (TEs) are DNA sequences able to be mobilized in host genomes. They are currently recognized as the major mutation inducers because of their insertion in the target, their effect on neighboring regions, or their ectopic recombination. A large number of factors including chemical and physical factors as well as intraspecific crosses have traditionally been identified as inducers of transposition. Besides environmental factors, interspecific crosses have also been proposed as promoters of transposition of particular TEs in plants and different animals. Our previous published work includes a genome-wide survey with the set of genomic TEs and shows that interspecific hybridization between the species Drosophila buzzatii and Drosophila koepferae induces genomic instability by transposition bursts. A high percentage of this instability corresponds to TEs belonging to classes I and II. The detailed study of three TEs (Osvaldo, Helena, and Galileo), representative of the different TE families, shows an increase of transposition in hybrids compared with parental species, that varies depending on the element. This study suggests ample variation in TE regulation mechanisms and the question is why this variation occurs. Interspecific hybridization is a genomic stressor that disrupts the stability of TEs probably contributing to a relaxation of the mechanisms controlling TEs in the Drosophila genome. In this commentary paper we will discuss these results and the molecular mechanisms that could explain these increases of transposition rates observed in interspecific Drosophila hybrids.

Characterization and potential evolutionary impact of transposable elements in the genome of Cochliobolus heterostrophus.

BackgroundCochliobolus heterostrophus is a dothideomycete that causes Southern Corn Leaf Blight disease. There are two races, race O and race T that differ by the absence (race O) and presence (race T) of ~ 1.2-Mb of DNA encoding genes responsible for the production of T-toxin, which makes race T much more virulent than race O. The presence of repetitive elements in fungal genomes is considered to be an important source of genetic variability between different species.ResultsA detailed analysis of class I and II TEs identified in the near complete genome sequence of race O was performed. In total in race O, 12 new families of transposons were identified. In silico evidence of recent activity was found for many of the transposons and analyses of expressed sequence tags (ESTs) demonstrated that these elements were actively transcribed. Various potentially active TEs were found near coding regions and may modify the expression and structure of these genes by acting as ectopic recombination sites. Transposons were found on scaffolds carrying polyketide synthase encoding genes, responsible for production of T-toxin in race T. Strong evidence of ectopic recombination was found, demonstrating that TEs can play an important role in the modulation of genome architecture of this species. The Repeat Induced Point mutation (RIP) silencing mechanism was shown to have high specificity in C. heterostrophus, acting only on transposons near coding regions.ConclusionsNew families of transposons were identified. In C. heterostrophus, the RIP silencing mechanism is efficient and selective. The co-localization of effector genes and TEs, therefore, exposes those genes to high rates of point mutations. This may accelerate the rate of evolution of these genes, providing a potential advantage for the host. Additionally, it was shown that ectopic recombination promoted by TEs appears to be the major event in the genome reorganization of this species and that a large number of elements are still potentially active. So, this study provides information about the potential impact of TEs on the evolution of C. heterostrophus.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-536) contains supplementary material, which is available to authorized users.

Characterization of the breakpoints of a polymorphic inversion complex detects strict and broad breakpoint reuse at the molecular level.

Inversions are an integral part of structural variation within species, and they play a leading role in genome reorganization across species. Work at both the cytological and genome sequence levels has revealed heterogeneity in the distribution of inversion breakpoints, with some regions being recurrently used. Breakpoint reuse at the molecular level has mostly been assessed for fixed inversions through genome sequence comparison, and therefore rather broadly. Here, we have identified and sequenced the breakpoints of two polymorphic inversions-E1 and E2 that share a breakpoint-in the extant Est and E1 + 2 chromosomal arrangements of Drosophila subobscura. The breakpoints are two medium-sized repeated motifs that mediated the inversions by two different mechanisms: E1 via staggered breaks and subsequent repair and E2 via repeat-mediated ectopic recombination. The fine delimitation of the shared breakpoint revealed its strict reuse at the molecular level regardless of which was the intermediate arrangement. The occurrence of other rearrangements in the most proximal and distal extended breakpoint regions reveals the broad reuse of these regions. This differential degree of fragility might be related to their sharing the presence outside the inverted region of snoRNA-encoding genes.

Frequent expansions of the bitter taste receptor gene repertoire during evolution of mammals in the Euarchontoglires clade.

Genome studies of mammals in the superorder Euarchontoglires (a clade that comprises the orders Primates, Dermoptera, Scandentia, Rodentia, and Lagomorpha) are important for understanding the biological features of humans, particularly studies of medical model animals such as macaques and mice. Furthermore, the dynamic ecoevolutionary signatures of Euarchontoglires genomes may be discovered because many species in this clade are characterized by their successful adaptive radiation to various ecological niches. In this study, we investigated the evolutionary trajectory of bitter taste receptor genes (TAS2Rs) in 28 Euarchontoglires species based on homology searches of 39 whole-genome assemblies. The Euarchontoglires species possessed variable numbers of intact TAS2Rs, which ranged from 16 to 40, and their last common ancestor had at least 26 intact TAS2Rs. The gene tree showed that there have been at least seven lineage-specific events involving massive gene duplications. Gene duplications were particularly evident in the ancestral branches of anthropoids (the anthropoid cluster), which may have promoted the adaptive evolution of anthropoid characteristics, such as a trade-off between olfaction and other senses and the development of herbivorous characteristics. Subsequent whole-gene deletions of anthropoid cluster TAS2Rs in hominoid species suggest ongoing ectopic homologous recombination in the anthropoid cluster. These findings provide insights into the roles of adaptive sensory evolution in various ecological niches and important clues related to the molecular mechanisms that underlie taste diversity in Euarchontoglires mammalian species, including humans.

Generation and characterization of Ins1-cre-driver C57BL/6N for exclusive pancreatic beta cell-specific Cre-loxP recombination.

Cre/loxP system-mediated site-specific recombination is utilized to study gene function in vivo. Successful conditional knockout of genes of interest is dependent on the availability of Cre-driver mice. We produced and characterized pancreatic β cell-specific Cre-driver mice for use in diabetes mellitus research. The gene encoding Cre was inserted into the second exon of mouse Ins1 in a bacterial artificial chromosome (BAC). Five founder mice were produced by microinjection of linearized BAC Ins1-cre. The transgene was integrated between Mafa and the telomere on chromosome 15 in one of the founders, BAC Ins1-cre25. To investigate Cre-loxP recombination, BAC Ins1-cre25 males were crossed with two different Cre-reporters, R26R and R26GRR females. On gross observation, reporter signal after Cre-loxP recombination was detected exclusively in the adult pancreatic islets in both F1 mice. Immunohistological analysis indicated that Cre-loxP recombination-mediated reporter signal was colocalized with insulin in pancreatic islet cells of both F1 mice, but not with glucagon. Moreover, Cre-loxP recombination signal was already observed in the pancreatic islets at E13.5 in both F1 fetuses. Finally, we investigated ectopic Cre-loxP recombination for Ins1, because the ortholog Ins2 is also expressed in the brain, in addition to the pancreas. However, there was no Cre-loxP recombination-mediated reporter signal in the brain of both F1 mice. Our data suggest that BAC Ins1-cre25 mice are a useful Cre-driver C57BL/6N for pancreatic β cell-specific Cre-loxP recombination, except for crossing with knock-in mice carrying floxed gene on chromosome 15.

DNA secondary structures are associated with recombination in majorPlasmodium falciparumvariable surface antigen gene families

Many bacterial, viral and parasitic pathogens undergo antigenic variation to counter host immune defense mechanisms. In Plasmodium falciparum, the most lethal of human malaria parasites, switching of var gene expression results in alternating expression of the adhesion proteins of the Plasmodium falciparum-erythrocyte membrane protein 1 class on the infected erythrocyte surface. Recombination clearly generates var diversity, but the nature and control of the genetic exchanges involved remain unclear. By experimental and bioinformatic identification of recombination events and genome-wide recombination hotspots in var genes, we show that during the parasite’s sexual stages, ectopic recombination between isogenous var paralogs occurs near low folding free energy DNA 50-mers and that these sequences are heavily concentrated at the boundaries of regions encoding individual Plasmodium falciparum-erythrocyte membrane protein 1 structural domains. The recombinogenic potential of these 50-mers is not parasite-specific because these sequences also induce recombination when transferred to the yeast Saccharomyces cerevisiae. Genetic cross data suggest that DNA secondary structures (DSS) act as inducers of recombination during DNA replication in P. falciparum sexual stages, and that these DSS-regulated genetic exchanges generate functional and diverse P. falciparum adhesion antigens. DSS-induced recombination may represent a common mechanism for optimizing the evolvability of virulence gene families in pathogens.

Sex-specific aspects of endogenous retroviral insertion and deletion

BackgroundWe wish to understand how sex and recombination affect endogenous retroviral insertion and deletion. While theory suggests that the risk of ectopic recombination will limit the accumulation of repetitive DNA in areas of high meiotic recombination, the experimental evidence so far has been inconsistent. Under the assumption of neutrality, we examine the genomes of eighteen species of animal in order to compute the ratio of solo-LTRs that derive from insertions occurring down the male germ line as opposed to the female one (male bias). We also extend the simple idea of comparing autosome to allosome in order to predict the ratio of full-length proviruses we would expect to see under conditions of recombination linked deletion or otherwise.ResultsUsing our model, we predict the ratio of allosomal to autosomal full-length proviruses to lie between and under increasing male bias in mammals and between 1 and 2 under increasing male bias in birds. In contrast to our expectations, we find that a pattern of male bias is not universal across species and that there is a frequent overabundance of full-length proviruses on the allosome beyond the ratios predicted by our model.ConclusionsWe use our data as a whole to argue that full-length proviruses should be treated as deleterious mutations or as effectively neutral mutations whose persistence in a full-length state is linked to the rate of meiotic recombination and whose origin is not universally male biased. These conclusions suggest that retroviral insertions on the allosome may be more prolific and that it might be possible to identify mechanisms of replication that are enhanced in the female sex.

R Loops Are Linked to Histone H3 S10 Phosphorylation and Chromatin Condensation

R loops are transcription byproducts that constitute athreat to genome integrity. Here we show that R loops are tightly linked to histone H3 S10 phosphorylation (H3S10P), a mark of chromatin condensation.Chromatin immunoprecipitation (ChIP)-on-chip (ChIP-chip) analyses reveal H3S10P accumulation at centromeres, pericentromeric chromatin, and a large number of active open reading frames (ORFs) in R-loop-accumulating yeast cells, better observed in G1. Histone H3S10 plays a key role in maintaining genome stability, as scored by ectopic recombination and plasmid loss, Rad52 foci, and Rad53 checkpoint activation. H3S10P coincides with the presence of DNA-RNA hybrids, is suppressed by ribonucleaseH overexpression, and causes reduced accessibility of restriction endonucleases, implying a tight connection between R loops, H3S10P, and chromatin compaction. Such histone modifications were also observed in R-loop-accumulating Caenorhabditis elegans and HeLa cells. We therefore provide a role of RNA in chromatin structure essential to understand how R loops modulate genome dynamics.

Alleles of the homologous recombination gene, RAD59, identify multiple responses to disrupted DNA replication in Saccharomyces cerevisiae

BackgroundIn Saccharomyces cerevisiae, Rad59 is required for multiple homologous recombination mechanisms and viability in DNA replication-defective rad27 mutant cells. Recently, four rad59 missense alleles were found to have distinct effects on homologous recombination that are consistent with separation-of-function mutations. The rad59-K166A allele alters an amino acid in a conserved α-helical domain, and, like the rad59 null allele diminishes association of Rad52 with double-strand breaks. The rad59-K174A and rad59-F180A alleles alter amino acids in the same domain and have genetically similar effects on homologous recombination. The rad59-Y92A allele alters a conserved amino acid in a separate domain, has genetically distinct effects on homologous recombination, and does not diminish association of Rad52 with double-strand breaks.ResultsIn this study, rad59 mutant strains were crossed with a rad27 null mutant to examine the effects of the rad59 alleles on the link between viability, growth and the stimulation of homologous recombination in replication-defective cells. Like the rad59 null allele, rad59-K166A was synthetically lethal in combination with rad27. The rad59-K174A and rad59-F180A alleles were not synthetically lethal in combination with rad27, had effects on growth that coincided with decreased ectopic gene conversion, but did not affect mutation, unequal sister-chromatid recombination, or loss of heterozygosity. The rad59-Y92A allele was not synthetically lethal when combined with rad27, stimulated ectopic gene conversion and heteroallelic recombination independently from rad27, and was mutually epistatic with srs2. Unlike rad27, the stimulatory effect of rad59-Y92A on homologous recombination was not accompanied by effects on growth rate, cell cycle distribution, mutation, unequal sister-chromatid recombination, or loss of heterozygosity.ConclusionsThe synthetic lethality conferred by rad59 null and rad59-K166A alleles correlates with their inhibitory effect on association of Rad52 with double-strand breaks, suggesting that this may be essential for rescuing replication lesions in rad27 mutant cells. The rad59-K174A and rad59-F180A alleles may fractionally reduce this same function, which proportionally reduced repair of replication lesions by homologous recombination and growth rate. In contrast, rad59-Y92A stimulates homologous recombination, perhaps by affecting association of replication lesions with the Rad51 recombinase. This suggests that Rad59 influences the rescue of replication lesions by multiple recombination factors.

Transposable Elements: Powerful Contributors to Angiosperm Evolution and Diversity

Transposable elements (TEs) are a dominant feature of most flowering plant genomes. Together with other accepted facilitators of evolution, accumulating data indicate that TEs can explain much about their rapid evolution and diversification. Genome size in angiosperms is highly correlated with TE content and the overwhelming bulk (>80%) of large genomes can be composed of TEs. Among retro-TEs, long terminal repeats (LTRs) are abundant, whereas DNA-TEs, which are often less abundant than retro-TEs, are more active. Much adaptive or evolutionary potential in angiosperms is due to the activity of TEs (active TE-Thrust), resulting in an extraordinary array of genetic changes, including gene modifications, duplications, altered expression patterns, and exaptation to create novel genes, with occasional gene disruption. TEs implicated in the earliest origins of the angiosperms include the exapted Mustang, Sleeper, and Fhy3/Far1 gene families. Passive TE-Thrust can create a high degree of adaptive or evolutionary potential by engendering ectopic recombination events resulting in deletions, duplications, and karyotypic changes. TE activity can also alter epigenetic patterning, including that governing endosperm development, thus promoting reproductive isolation. Continuing evolution of long-lived resprouter angiosperms, together with genetic variation in their multiple meristems, indicates that TEs can facilitate somatic evolution in addition to germ line evolution. Critical to their success, angiosperms have a high frequency of polyploidy and hybridization, with resultant increased TE activity and introgression, and beneficial gene duplication. Together with traditional explanations, the enhanced genomic plasticity facilitated by TE-Thrust, suggests a more complete and satisfactory explanation for Darwin’s “abominable mystery”: the spectacular success of the angiosperms.

Gene duplication in the major insecticide target site, Rdl, in Drosophila melanogaster.

The Resistance to Dieldrin gene, Rdl, encodes a GABA-gated chloride channel subunit that is targeted by cyclodiene and phenylpyrazole insecticides. The gene was first characterized in Drosophila melanogaster by genetic mapping of resistance to the cyclodiene dieldrin. The 4,000-fold resistance observed was due to a single amino acid replacement, Ala(301) to Ser. The equivalent change was subsequently identified in Rdl orthologs of a large range of resistant insect species. Here, we report identification of a duplication at the Rdl locus in D. melanogaster. The 113-kb duplication contains one WT copy of Rdl and a second copy with two point mutations: an Ala(301) to Ser resistance mutation and Met(360) to Ile replacement. Individuals with this duplication exhibit intermediate dieldrin resistance compared with single copy Ser(301) homozygotes, reduced temperature sensitivity, and altered RNA editing associated with the resistant allele. Ectopic recombination between Roo transposable elements is involved in generating this genomic rearrangement. The duplication phenotypes were confirmed by construction of a transgenic, artificial duplication integrating the 55.7-kb Rdl locus with a Ser(301) change into an Ala(301) background. Gene duplications can contribute significantly to the evolution of insecticide resistance, most commonly by increasing the amount of gene product produced. Here however, duplication of the Rdl target site creates permanent heterozygosity, providing unique potential for adaptive mutations to accrue in one copy, without abolishing the endogenous role of an essential gene.

DNA in motion during double-strand break repair

DNA organization and dynamics profoundly affect many biological processes such as gene regulation and DNA repair. In this review, we present the latest studies on DNA mobility in the context of DNA damage. Recent studies demonstrate that DNA mobility is dramatically increased in the presence of double-strand breaks (DSBs) in the yeast Saccharomyces cerevisiae. As a consequence, chromosomes explore a larger nuclear volume, facilitating homologous pairing but also increasing the rate of ectopic recombination. Increased DNA dynamics is dependent on several homologous recombination (HR) proteins and we are just beginning to understand how chromosome dynamics is regulated after DNA damage.

The Role of Recombination in the Origin and Evolution of Alu Subfamilies

Alus are the most abundant and successful short interspersed nuclear elements found in primate genomes. In humans, they represent about 10% of the genome, although few are retrotransposition-competent and are clustered into subfamilies according to the source gene from which they evolved. Recombination between them can lead to genomic rearrangements of clinical and evolutionary significance. In this study, we have addressed the role of recombination in the origin of chimeric Alu source genes by the analysis of all known consensus sequences of human Alus. From the allelic diversity of Alu consensus sequences, validated in extant elements resulting from whole genome searches, distinct events of recombination were detected in the origin of particular subfamilies of AluS and AluY source genes. These results demonstrate that at least two subfamilies are likely to have emerged from ectopic Alu-Alu recombination, which stimulates further research regarding the potential of chimeric active Alus to punctuate the genome.

Analysis of plastid and mitochondrial DNA insertions in the nucleus (NUPTs and NUMTs) of six plant species: size, relative age and chromosomal localization

We analysed the size, relative age and chromosomal localization of nuclear sequences of plastid and mitochondrial origin (NUPTs-nuclear plastid DNA and NUMTs-nuclear mitochondrial DNA) in six completely sequenced plant species. We found that the largest insertions showed lower divergence from organelle DNA than shorter insertions in all species, indicating their recent origin. The largest NUPT and NUMT insertions were localized in the vicinity of the centromeres in the small genomes of Arabidopsis and rice. They were also present in other chromosomal regions in the large genomes of soybean and maize. Localization of NUPTs and NUMTs correlated positively with distribution of transposable elements (TEs) in Arabidopsis and sorghum, negatively in grapevine and soybean, and did not correlate in rice or maize. We propose a model where new plastid and mitochondrial DNA sequences are inserted close to centromeres and are later fragmented by TE insertions and reshuffled away from the centromere or removed by ectopic recombination. The mode and tempo of TE dynamism determines the turnover of NUPTs and NUMTs resulting in their species-specific chromosomal distributions.