Genomic DNA insertions and deletions occur frequently between humans and nonhuman primates

Comparative DNA sequence studies between humans and nonhuman primates will be important for understanding the genetic basis of the phenotypic differences between these species. Here we compare approximately 27 Mb of human chromosome 21 with chimpanzee DNA sequences identifying 57 genomic rearrangements (deletions and insertions ranging in size from 0.2 to 8.0 kb) between the two species. These rearrangements are distributed along the entire length of chromosome 21, with approximately 35% found in genomic intervals encoding genes (genic intervals), and have occurred in the genomes of both humans and chimpanzees. Comparison of approximately 9 Mb of human chromosome 21 with orangutan, rhesus macaque, and woolly monkey DNA sequences identified a combined total of 114 genomic rearrangements between humans and nonhuman primates. Analysis of these rearrangements revealed that they are randomly distributed with respect to genic and nongenic intervals and identified one deletion that has likely resulted in the inactivation of a gene (beta1,3-galactosyltransferase) in the woolly monkey. Our data show that genomic rearrangements have occurred frequently during primate genome evolution and significantly contribute to the DNA differences between these species. These DNA rearrangements are commonly found in genic intervals, and thus provide natural starting points for focused investigations of qualitative and quantitative gene expression differences between humans and other primates.     

Insertional mutagenesis by transposable elements in the mammalian genome

Several mammalian repetitive transposable genetic elements were characterized in recent years, and their role in mutagenesis is delineated in this review. Two main groups have been described: elements with symmetrical termini such as the murine IAP sequences and the human THE 1 elements and elements characterized by a poly-A rich tail at the 3′ end such as the SINE and LINE sequences. The characteristic property of such mobile elements to spread and integrate in the host genome leads to insertional mutagenesis. Both germline and somatic mutations have been documented resulting from the insertion of the various types of mammalian repetitive transposable genetic elements. As foreseen by Barbara McClintock, such genetic events can cause either the activation or the inactivation of specific genes, resulting in their identification via an altered phenotype. Several disease states, such as hemophilia and cancer, are the result of this apparent aspect of genome instability. © 1993 Wiley-Liss, Inc.     

Diversity and evolution of transposable elements in Arabidopsis

Transposable elements are mobile genetic elements that have successfully populated eukaryotic genomes and show diversity in their structure and transposition mechanisms. Although first viewed solely as selfish, transposable elements are now known as important vectors to drive the adaptation and evolution of their host genome. Transposable elements can affect host gene structures, gene copy number, gene expression, and even as a source for novel genes. For example, a number of transposable element sequences have been co-opted to contribute to evolutionary innovation, such as the mammalian placenta and the vertebrate immune system. In plants, the need to adapt rapidly to changing environmental conditions is essential and is reflected, as will be discussed, by genome plasticity and an abundance of diverse, active transposon families. This review focuses on transposable elements in plants, particularly those that have beneficial effects on the host. We also emphasize the importance of having proper tools to annotate and classify transposons to better understand their biology.     

Control of transposable elements in Arabidopsis thaliana

Arabidopsis thaliana serves as a very good model organism to investigate the control of transposable elements (TEs) by genetic and genomic approaches. As TE movements are potentially deleterious to the hosts, hosts silence TEs by epigenetic mechanisms, such as DNA methylation. DNA methylation is controlled by DNA methyltransferases and other regulators, including histone modifiers and chromatin remodelers. RNAi machinery directs DNA methylation to euchromatic TEs, which is under developmental control. In addition to the epigenetic controls, some TEs are controlled by environmental factors. TEs often affect expression of nearby genes, providing evolutionary sources for epigenetic, developmental, and environmental gene controls, which could even be beneficial for the host.     

Enrichment of short interspersed transposable elements to embryonic stem cell‐specific hypomethylated gene regions

Embryonic stem cells (ESCs) have a distinctive epigenome, which includes their genome‐wide DNA methylation modification status, as represented by the ESC‐specific hypomethylation of tissue‐dependent and differentially methylated regions (T‐DMRs) of Pou5f1 and Nanog. Here, we conducted a genome‐wide investigation of sequence characteristics associated with T‐DMRs that were differentially methylated between ESCs and somatic cells, by focusing on transposable elements including short interspersed elements (SINEs), long interspersed elements (LINEs) and long terminal repeats (LTRs). We found that hypomethylated T‐DMRs were predominantly present in SINE‐rich/LINE‐poor genomic loci. The enrichment for SINEs spread over 300 kb in cis and there existed SINE‐rich genomic domains spreading continuously over 1 Mb, which contained multiple hypomethylated T‐DMRs. The characterization of sequence information showed that the enriched SINEs were relatively CpG rich and belonged to specific subfamilies. A subset of the enriched SINEs were hypomethylated T‐DMRs in ESCs at Dppa3 gene locus, although SINEs are overall methylated in both ESCs and the liver. In conclusion, we propose that SINE enrichment is the genomic property of regions harboring hypomethylated T‐DMRs in ESCs, which is a novel aspect of the ESC‐specific epigenomic information.     

Polymorphism in Brucella strains detected by studying distribution of two short repetitive DNA elements.

Thirty-four Brucella reference or field strains representing all the species and biovars were studied by repetitive element sequence-based PCR, a PCR using primers complementary to two enterobacterial short repetitive sequences: repetitive extragenic palindromic and enterobacterial repetitive intergenic consensus sequences. All the stains showed a positive amplification, suggesting that the Brucella genome contains such sequences. Repetitive extragenic palindromic PCR was less discriminating than enterobacterial repetitive intergenic consensus PCR in terms of distinguishing strains, but a combination of the two methods was able to distinguish all the isolates, except for some strains belonging to biovars 3 and 9 of Brucella abortus. Repetitive element sequence-based PCR appears to be a simple and useful method for the study of brucellosis epidemiology.     

Widespread and frequent horizontal transfers of transposable elements in plants

Vertical, transgenerational transmission of genetic material occurs through reproduction of living organisms. In addition to vertical inheritance, horizontal gene transfer between reproductively isolated species has recently been shown to be an important, if not dominant, mechanism in the evolution of prokaryotic genomes. In contrast, only a few horizontal transfer (HT) events have been characterized so far in eukaryotes and mainly concern transposable elements (TEs). Whether these are frequent and have a significant impact on genome evolution remains largely unknown. We performed a computational search for highly conserved LTR retrotransposons among 40 sequenced eukaryotic genomes representing the major plant families. We found that 26 genomes (65%) harbor at least one case of horizontal TE transfer (HTT). These transfers concern species as distantly related as palm and grapevine, tomato and bean, or poplar and peach. In total, we identified 32 cases of HTTs, which could translate into more than 2 million among the 13,551 monocot and dicot genera. Moreover, we show that these TEs have remained functional after their transfer, occasionally causing a transpositional burst. This suggests that plants can frequently exchange genetic material through horizontal transfers and that this mechanism may be important in TE-driven genome evolution.Transposable elements (TEs) are mobile genomic DNA sequences that are found in almost all living organisms (Finnegan 1985). They so densely populate the genomes of many eukaryotic species that they are often the major components, as in human (>50%) (Prak and Kazazian 2000) or bread wheat (>95%) (Bennetzen 2000). In this regard, TEs have been shown to have a major impact on both structural and functional modifications of genomes (Bennetzen 2000; Feschotte 2008). They are usually classified into two distinct types. Class I elements (retrotransposons) transpose via an RNA intermediate through a copy and paste mechanism, whereas class II elements (transposons) transpose through a cut-and-paste mechanism (Wicker et al. 2007). Both classes comprise various types (orders and superfamilies in Wicker''s classification). Although most of these types can be found in all the plant genomes sequenced so far, LTR retrotransposons represent by far the major genomic constituents in the kingdom. In this regard, LTR retrotransposons have been shown to strongly impact genome structure (Piegu et al. 2006), whereas several reports have demonstrated their putative functional impact as epigenetic mediators (Kobayashi et al. 2004).TEs achieve their transposition cycle within their host and are thus considered to be lineage specific because, like genes, they are inherited vertically from one generation to another. However, unlike genes, they do not encode any information essential for their host, and their insertion into genes can in some cases have a negative effect on fitness. This “selfish” and potentially deleterious nature has raised the question of their persistence in eukaryotic lineages, especially after it was shown that TEs are strictly controlled by several silencing pathways (Slotkin and Martienssen 2007; Rigal and Mathieu 2011) and efficiently eliminated from their host genomes through deletions (Vitte and Panaud 2005). Horizontal transfers could allow TEs to escape this process by transposing into a new “naive” host genome, therefore ensuring their long-term survival. However, although horizontal gene transfers are very common in Bacteria (Rocha 2013), evidence of HTTs in eukaryotes remains scarce, although recent reports suggest their potential impact in genome evolution (Schaack et al. 2010). Three criteria have been defined for the detection of HTTs: patchy distributions of TEs in phylogenies; identification of TEs exhibiting high sequence similarity between distantly related taxa; and phylogenetic incongruence between the host and TEs (Gilbert et al. 2010; Kuraku et al. 2012; Wallau et al. 2012; Walsh et al. 2013). An exhaustive search for HTTs that meet these three criteria in a wide taxonomic range thus requires a comprehensive set of genomic resources. Next-generation sequencing (NGS) has made available full genome sequences for many organisms, enabling genome-wide comparative surveys for a large panel of evolutionary lineages. Using such resources, we surveyed HTTs across the plant kingdom, and we show that they are very frequent and widespread among monocots and dicots.     

Bioinformatics and genomic analysis of transposable elements in eukaryotic genomes

A major portion of most eukaryotic genomes are transposable elements (TEs). During evolution, TEs have introduced profound changes to genome size, structure, and function. As integral parts of genomes, the dynamic presence of TEs will continue to be a major force in reshaping genomes. Early computational analyses of TEs in genome sequences focused on filtering out “junk” sequences to facilitate gene annotation. When the high abundance and diversity of TEs in eukaryotic genomes were recognized, these early efforts transformed into the systematic genome-wide categorization and classification of TEs. The availability of genomic sequence data reversed the classical genetic approaches to discovering new TE families and superfamilies. Curated TE databases and their accurate annotation of genome sequences in turn facilitated the studies on TEs in a number of frontiers including: (1) TE-mediated changes of genome size and structure, (2) the influence of TEs on genome and gene functions, (3) TE regulation by host, (4) the evolution of TEs and their population dynamics, and (5) genomic scale studies of TE activity. Bioinformatics and genomic approaches have become an integral part of large-scale studies on TEs to extract information with pure in silico analyses or to assist wet lab experimental studies. The current revolution in genome sequencing technology facilitates further progress in the existing frontiers of research and emergence of new initiatives. The rapid generation of large-sequence datasets at record low costs on a routine basis is challenging the computing industry on storage capacity and manipulation speed and the bioinformatics community for improvement in algorithms and their implementations.     

Very short DNA segments can be detected and handled by the repair machinery during germline chromothriptic chromosome reassembly

Analyses at nucleotide resolution reveal unexpected complexity of seemingly simple and balanced chromosomal rearrangements. Chromothripsis is a rare complex aberration involving local shattering of one or more chromosomes and reassembly of the resulting DNA segments. This can influence gene expression and cause abnormal phenotypes. We studied the structure and mechanism of a seemingly balanced de novo complex rearrangement of four chromosomes in a boy with developmental and growth delay. Microarray analysis revealed two paternal de novo deletions of 0.7 and 2.5 Mb at two of the breakpoints in 1q24.3 and 6q24.1‐q24.2, respectively, which could explain most symptoms of the patient. Subsequent whole‐genome mate‐pair sequencing confirmed the chromothriptic nature of the rearrangement. The four participating chromosomes were broken into 29 segments longer than 1 kb. Sanger sequencing of all breakpoint junctions revealed additional complexity compatible with the involvement of different repair pathways. We observed translocation of a 33 bp long DNA fragment, which may have implications for the definition of the lower size limit of structural variants. Our observations and literature review indicate that even very small fragments from shattered chromosomes can be detected and handled by the repair machinery during germline chromothriptic chromosome reassembly.     

Myoglobinuria caused by multiple deletions of mitochondrial DNA

We report two brothers with inherited recurrent myoglobinuria associated with distinct morphological abnormalities of muscle mitochondria and multiple deletions of muscle mitochondrial DNA. Patient 1 (26 years old) and Patient 2 (21 years old) had recurrent episodes of myoglobinuria provoked by strenuous exercise or alcohol intake. Histochemistry of their biopsied limb muscles showed ragged-red fibers and cytochrome c oxidase-negative fibers as well as degenerating and regenerating fibers. Electron microscopy showed a pronounced accumulation of abnormal mitochondria containing paracrystalline inclusions and moderate increases of glycogen particles. Southern blot analysis revealed multiple deletions of mitochondrial DNA, some of which were common to both patients. By the primer shift polymerase chain reaction method, we detected multiple abnormal fragments indicating mitochondrial DNA deletions. Nucleotide sequencing of the deleted regions disclosed directly repeated sequences of 1 to 12 bp on each side of the deletions. Since the end points of mitochondrial DNA deletions were within 20 bp of the major non-coding region, probable mutations in this region contribute to the pathogenesis of multiple mitochondrial DNA deletions found in these patients. We propose that a defect of the mitochondrial energy-transducing system due to multiple mitochondrial DNA deletions is a novel genetic cause of inherited recurrent myoglobinuria.     

Somatically heritable switches in the DNA modification of Mu transposable elements monitored with a suppressible mutant in maize

Many transposable elements in maize alternate between active and inactive phases associated with the modification of their DNA. Elements in an inactive phase lose their ability to transpose, their ability to excise from reporter alleles and, in some cases, their ability to enhance or suppress mutant phenotypes caused by their insertion. The maize mutant hcf106 is a recessive pale green seedling lethal caused by the insertion of the transposable element Mu1. We show that the hcf106 mutant phenotype is suppressed in lines that have lost Mu activity. That is, homozygous hcf106 seedlings are dark green and viable when transposable elements belonging to the Robertson's Mutator family are modified in their terminal inverted repeats, a diagnostic feature of inactive lines. This property of the mutant phenotype has been used to follow clonal leaf sectors containing modified Mu elements that arise from single somatic cells during plant development. The distribution of these sectors indicates that epigenetic switches involving Mu DNA modification occur progressively as the meristem ages.     

The Trickster in the genome: contribution and control of transposable elements

In mythology, the Trickster is an archetype who typically behaves selfishly and delights in playing tricks and breaking ordinary rules. In many myths and folktales, however, the Trickster also brings new knowledge and, ultimately, has positive effects on the community. Transposable elements (TEs) might have played such a role in the story of genome evolution. TEs can cause nonroutine genetic events like insertional mutations and ectopic recombination that provide a fundamental source of genetic variation, but they can also be a potential threat to genome integrity. Thus, the activity of TEs is usually controlled by an array of sophisticated mechanisms for genome defense. Recent findings indicate that TEs are important components of eukaryotic genomes, often to a much larger extent than ever anticipated. In this review, I focus on the contributions of TEs to various aspects of genome evolution. In addition, why TEs are specific targets for the genome defense mechanisms is discussed.     

Evolutionary dynamics of transposable elements in the short-tailed opossum Monodelphis domestica

The genome of the gray short-tailed opossum Monodelphis domestica is notable for its large size ( approximately 3.6 Gb). We characterized nearly 500 families of interspersed repeats from the Monodelphis. They cover approximately 52% of the genome, higher than in any other amniotic lineage studied to date, and may account for the unusually large genome size. In comparison to other mammals, Monodelphis is significantly rich in non-LTR retrotransposons from the LINE-1, CR1, and RTE families, with >29% of the genome sequence comprised of copies of these elements. Monodelphis has at least four families of RTE, and we report support for horizontal transfer of this non-LTR retrotransposon. In addition to short interspersed elements (SINEs) mobilized by L1, we found several families of SINEs that appear to use RTE elements for mobilization. In contrast to L1-mobilized SINEs, the RTE-mobilized SINEs in Monodelphis appear to shift from G+C-rich to G+C-low regions with time. Endogenous retroviruses have colonized approximately 10% of the opossum genome. We found that their density is enhanced in centromeric and/or telomeric regions of most Monodelphis chromosomes. We identified 83 new families of ancient repeats that are highly conserved across amniotic lineages, including 14 LINE-derived repeats; and a novel SINE element, MER131, that may have been exapted as a highly conserved functional noncoding RNA, and whose emergence dates back to approximately 300 million years ago. Many of these conserved repeats are also present in human, and are highly over-represented in predicted cis-regulatory modules. Seventy-six of the 83 families are present in chicken in addition to mammals.     

Intramolecular recombination of a mitochondrial minicircular plasmid-like DNA of date-palm mediated by a set of short direct-repeat sequences

A molecular clone containing the complete sequence of a mitochondrial circular plasmid-like DNA (the R plasmid) isolated from the date-palm variety V3DP was used as a probe in Southern analyses of mitochondrial DNA prepared from other varieties. Another circular structure (the S plasmid) was detected in some of these varieties, and sequenced from variety V2DP. It appears that the R plasmid could have arisen from the S plasmid by an intermolecular recombination event at a set of 26-bp imperfect short direct repeats.     

Genomic structure of chromosome 17 deletions in BRCA1-associated ovarian cancers

The aim of this study was to determine the genomic structure of the deletions on chromosome 17 in ovarian carcinomas from women with inherited BRCA1 mutations. Normal and tumor DNA from 14 ovarian tumors associated with inherited BRCA1 mutations were extracted and tested for loss of heterozygosity (LOH) at microsatellite markers along chromosome 17. Finer mapping using more microsatellite markers and single nucleotide polymorphisms helped further define the LOH margins. The genomic repeated elements within the LOH breakpoint regions were identified using the University of California Santa Cruz Genome Database, and the frequencies were compared to regions of equal GC percentages across the genome. Of the 14 ovarian tumors, 12 showed LOH of the entire chromosome 17. The other two tumors lost the distal end of the 17q arm. The breakpoints of these two tumors occurred in regions with significantly high frequencies of short interspersed nuclear elements (SINE), specifically Alu elements. Ovarian tumors of high grade and stage have large regions of LOH along chromosome 17, with most tumors showing loss of the entire chromosome. In those tumors with retention of part of chromosome 17, LOH margins suggest that a high Alu content may have a role in the deletions.     

Genomic deletions in MSH2 or MLH1 are a frequent cause of hereditary non-polyposis colorectal cancer: identification of novel and recurrent deletions by MLPA

Gene dosage abnormalities account for a significant proportion of the mutations in genes tested in DNA diagnostic laboratories. Detection of these changes has proved a challenge as the methods available to date are time consuming or unreliable. The multiplex ligation-dependent probe assay (MLPA) is a new technique allowing relative quantification of up to 40 different nucleic acid sequences in a single reaction tube. We have evaluated MLPA for potential use in the diagnostic setting against the following criteria: accuracy, reagent cost, hands-on time, reliability, and retests required. A total of 215 UK patients referred for genetic testing on the basis of a family history consistent with autosomal dominant hereditary non-polyposis colorectal cancer (HNPCC or Lynch syndrome) were tested by MLPA. Of these, 12 cases with deletions of one or more exons were identified, six with MLH1 deletions and six with MSH2 deletions. Test failure rates were less than 5% and overall mutation detection sensitivity in this series was increased by approximately 50% by the inclusion of MLPA for an additional testing cost of about 10%. Two novel mutations in MSH2 and 10 novel point mutations in MLH1 were also identified during the course of this study. We conclude that MLPA is a cost effective and robust gene dosage method that can be readily adopted by diagnostic services. Comprehensive mutation scanning for MSH2 and MLH1 is incomplete without gene dosage analysis.