A recurrent translocation is mediated by homologous recombination between HERV-H elements

<p>Abstract</p> <p>Background</p> <p>Chromosome rearrangements are caused by many mutational mechanisms; of these, recurrent rearrangements can be particularly informative for teasing apart DNA sequence-specific factors. Some recurrent translocations are mediated by homologous recombination between large blocks of segmental duplications on different chromosomes. Here we describe a recurrent unbalanced translocation casued by recombination between shorter homologous regions on chromosomes 4 and 18 in two unrelated children with intellectual disability.</p> <p>Results</p> <p>Array CGH resolved the breakpoints of the 6.97-Megabase (Mb) loss of 18q and the 7.30-Mb gain of 4q. Sequencing across the translocation breakpoints revealed that both translocations occurred between 92%-identical human endogenous retrovirus (HERV) elements in the same orientation on chromosomes 4 and 18. In addition, we find sequence variation in the chromosome 4 HERV that makes one allele more like the chromosome 18 HERV.</p> <p>Conclusions</p> <p>Homologous recombination between HERVs on the same chromosome is known to cause chromosome deletions, but this is the first report of interchromosomal HERV-HERV recombination leading to a translocation. It is possible that normal sequence variation in substrates of non-allelic homologous recombination (NAHR) affects the alignment of recombining segments and influences the propensity to chromosome rearrangement.</p

Assembly and characterization of heterochromatin and euchromatin on human artificial chromosomes

BACKGROUND: Human centromere regions are characterized by the presence of alpha-satellite DNA, replication late in S phase and a heterochromatic appearance. Recent models propose that the centromere is organized into conserved chromatin domains in which chromatin containing CenH3 (centromere-specific H3 variant) at the functional centromere (kinetochore) forms within regions of heterochromatin. To address these models, we assayed formation of heterochromatin and euchromatin on de novo human artificial chromosomes containing alpha-satellite DNA. We also examined the relationship between chromatin composition and replication timing of artificial chromosomes. RESULTS: Heterochromatin factors (histone H3 lysine 9 methylation and HP1α) were enriched on artificial chromosomes estimated to be larger than 3 Mb in size but depleted on those smaller than 3 Mb. All artificial chromosomes assembled markers of euchromatin (histone H3 lysine 4 methylation), which may partly reflect marker-gene expression. Replication timing studies revealed that the replication timing of artificial chromosomes was heterogeneous. Heterochromatin-depleted artificial chromosomes replicated in early S phase whereas heterochromatin-enriched artificial chromosomes replicated in mid to late S phase. CONCLUSIONS: Centromere regions on human artificial chromosomes and host chromosomes have similar amounts of CenH3 but exhibit highly varying degrees of heterochromatin, suggesting that only a small amount of heterochromatin may be required for centromere function. The formation of euchromatin on all artificial chromosomes demonstrates that they can provide a chromosome context suitable for gene expression. The earlier replication of the heterochromatin-depleted artificial chromosomes suggests that replication late in S phase is not a requirement for centromere function

Elevated Rates of Sister Chromatid Exchange at Chromosome Ends

Chromosome ends are known hotspots of meiotic recombination and double-strand breaks. We monitored mitotic sister chromatid exchange (SCE) in telomeres and subtelomeres and found that 17% of all SCE occurs in the terminal 0.1% of the chromosome. Telomeres and subtelomeres are significantly enriched for SCEs, exhibiting rates of SCE per basepair that are at least 1,600 and 160 times greater, respectively, than elsewhere in the genome

Terminal 18q deletions are stabilized by neotelomeres

Background: All human chromosomes are capped by tandem repeat (TTAGGG)n sequences that protect them against end-to-end fusion and are essential to chromosomal replication and integrity. Therefore, after a chromosomal breakage, the deleted chromosomes must be stabilized by retaining the telomere or acquiring a new cap, by telomere healing or telomere capture. There are few reports with molecular approaches on the mechanisms involved in stabilization of 18q terminal deletions.Results: in this study we analyzed nine patients with 18q terminal deletion identified by G-banding and genomic array. FISH using PNA probe revealed telomeric signals in all deleted chromosomes tested. We fine-mapped breakpoints with customized arrays and sequenced six terminal deletion junctions. in all six deleted chromosomes sequenced, telomeric sequences were found directly attached to the breakpoints. Little or no microhomology was found at the breakpoints and none of the breaks sequenced were located in low copy repeat (LCR) regions, though repetitive elements were found around the breakpoints in five patients. One patient presented a more complex rearrangement with two deleted segments and an addition of 17 base pairs (bp).Conclusions: We found that all six deleted chromosomes sequenced were probably stabilized by the healing mechanism leading to a neotelomere formation.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Universidade Federal de São Paulo, Dept Morphol & Genet, BR-04023900 São Paulo, BrazilEmory Univ, Sch Med, Dept Human Genet, Atlanta, GA 30322 USAUniversidade Federal de São Paulo, Dept Biophys, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Pathol, Lab Citogenom, BR-05403000 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Morphol & Genet, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Biophys, BR-04023900 São Paulo, BrazilUniversidade Federal de São Paulo, Dept Pathol, Lab Citogenom, BR-05403000 São Paulo, BrazilFAPESP: 2012/51150-0FAPESP: 2012/15572-7Web of Scienc

Development of genomic resources for the prairie vole (Microtus ochrogaster): construction of a BAC library and vole-mouse comparative cytogenetic map

<p>Abstract</p> <p>Background</p> <p>The prairie vole (<it>Microtus ochrogaster</it>) is a premier animal model for understanding the genetic and neurological basis of social behaviors. Unlike other biomedical models, prairie voles display a rich repertoire of social behaviors including the formation of long-term pair bonds and biparental care. However, due to a lack of genomic resources for this species, studies have been limited to a handful of candidate genes. To provide a substrate for future development of genomic resources for this unique model organism, we report the construction and characterization of a bacterial artificial chromosome (BAC) library from a single male prairie vole and a prairie vole-mouse (<it>Mus musculus</it>) comparative cytogenetic map.</p> <p>Results</p> <p>We constructed a prairie vole BAC library (CHORI-232) consisting of 194,267 recombinant clones with an average insert size of 139 kb. Hybridization-based screening of the gridded library at 19 loci established that the library has an average depth of coverage of ~10×. To obtain a small-scale sampling of the prairie vole genome, we generated 3884 BAC end-sequences totaling ~2.8 Mb. One-third of these BAC-end sequences could be mapped to unique locations in the mouse genome, thereby anchoring 1003 prairie vole BAC clones to an orthologous position in the mouse genome. Fluorescence in situ hybridization (FISH) mapping of 62 prairie vole clones with BAC-end sequences mapping to orthologous positions in the mouse genome was used to develop a first-generation genome-wide prairie vole-mouse comparative cytogenetic map. While conserved synteny was observed between this pair of rodent genomes, rearrangements between the prairie vole and mouse genomes were detected, including a minimum of five inversions and 16 inter-chromosomal rearrangements.</p> <p>Conclusions</p> <p>The construction of the prairie vole BAC library and the vole-mouse comparative cytogenetic map represent the first genome-wide modern genomic resources developed for this species. The BAC library will support future genomic, genetic and molecular characterization of this genome and species, and the isolation of clones of high interest to the vole research community will allow for immediate characterization of the regulatory and coding sequences of genes known to play important roles in social behaviors. In addition, these resources provide an excellent platform for future higher resolution cytogenetic mapping and full genome sequencing.</p

The Evolutionary Origin of Man Can Be Traced in the Layers of Defunct Ancestral Alpha Satellites Flanking the Active Centromeres of Human Chromosomes

Alpha satellite domains that currently function as centromeres of human chromosomes are flanked by layers of older alpha satellite, thought to contain dead centromeres of primate progenitors, which lost their function and the ability to homogenize satellite repeats, upon appearance of a new centromere. Using cladistic analysis of alpha satellite monomers, we elucidated complete layer patterns on chromosomes 8, 17, and X and related them to each other and to primate alpha satellites. We show that discrete and chronologically ordered alpha satellite layers are partially symmetrical around an active centromere and their succession is partially shared in non-homologous chromosomes. The layer structure forms a visual representation of the human evolutionary lineage with layers corresponding to ancestors of living primates and to entirely fossil taxa. Surprisingly, phylogenetic comparisons suggest that alpha satellite arrays went through periods of unusual hypermutability after they became “dead” centromeres. The layer structure supports a model of centromere evolution where new variants of a satellite repeat expanded periodically in the genome by rounds of inter-chromosomal transfer/amplification. Each wave of expansion covered all or many chromosomes and corresponded to a new primate taxon. Complete elucidation of the alpha satellite phylogenetic record would give a unique opportunity to number and locate the positions of major extinct taxa in relation to human ancestors shared with extant primates. If applicable to other satellites in non-primate taxa, analysis of centromeric layers could become an invaluable tool for phylogenetic studies

Integrating 5-Hydroxymethylcytosine into the Epigenomic Landscape of Human Embryonic Stem Cells

Covalent modification of DNA distinguishes cellular identities and is crucial for regulating the pluripotency and differentiation of embryonic stem (ES) cells. The recent demonstration that 5-methylcytosine (5-mC) may be further modified to 5-hydroxymethylcytosine (5-hmC) in ES cells has revealed a novel regulatory paradigm to modulate the epigenetic landscape of pluripotency. To understand the role of 5-hmC in the epigenomic landscape of pluripotent cells, here we profile the genome-wide 5-hmC distribution and correlate it with the genomic profiles of 11 diverse histone modifications and six transcription factors in human ES cells. By integrating genomic 5-hmC signals with maps of histone enrichment, we link particular pluripotency-associated chromatin contexts with 5-hmC. Intriguingly, through additional correlations with defined chromatin signatures at promoter and enhancer subtypes, we show distinct enrichment of 5-hmC at enhancers marked with H3K4me1 and H3K27ac. These results suggest potential role(s) for 5-hmC in the regulation of specific promoters and enhancers. In addition, our results provide a detailed epigenomic map of 5-hmC from which to pursue future functional studies on the diverse regulatory roles associated with 5-hmC

The evolutionary dynamics of α-satellite

α-Satellite is a family of tandemly repeated sequences found at all normal human centromeres. In addition to its significance for understanding centromere function, α-satellite is also a model for concerted evolution, as α-satellite repeats are more similar within a species than between species. There are two types of α-satellite in the human genome; while both are made up of ∼171-bp monomers, they can be distinguished by whether monomers are arranged in extremely homogeneous higher-order, multimeric repeat units or exist as more divergent monomeric α-satellite that lacks any multimeric periodicity. In this study, as a model to examine the genomic and evolutionary relationships between these two types, we have focused on the chromosome 17 centromeric region that has reached both higher-order and monomeric α-satellite in the human genome assembly. Monomeric and higher-order α-satellites on chromosome 17 are phylogenetically distinct, consistent with a model in which higher-order evolved independently of monomeric α-satellite. Comparative analysis between human chromosome 17 and the orthologous chimpanzee chromosome indicates that monomeric α-satellite is evolving at approximately the same rate as the adjacent non-α-satellite DNA. However, higher-order α-satellite is less conserved, suggesting different evolutionary rates for the two types of α-satellite

Human Artificial Chromosomes with Alpha Satellite-Based De Novo Centromeres Show Increased Frequency of Nondisjunction and Anaphase Lag

Human artificial chromosomes have been used to model requirements for human chromosome segregation and to explore the nature of sequences competent for centromere function. Normal human centromeres require specialized chromatin that consists of alpha satellite DNA complexed with epigenetically modified histones and centromere-specific proteins. While several types of alpha satellite DNA have been used to assemble de novo centromeres in artificial chromosome assays, the extent to which they fully recapitulate normal centromere function has not been explored. Here, we have used two kinds of alpha satellite DNA, DXZ1 (from the X chromosome) and D17Z1 (from chromosome 17), to generate human artificial chromosomes. Although artificial chromosomes are mitotically stable over many months in culture, when we examined their segregation in individual cell divisions using an anaphase assay, artificial chromosomes exhibited more segregation errors than natural human chromosomes (P < 0.001). Naturally occurring, but abnormal small ring chromosomes derived from chromosome 17 and the X chromosome also missegregate more than normal chromosomes, implicating overall chromosome size and/or structure in the fidelity of chromosome segregation. As different artificial chromosomes missegregate over a fivefold range, the data suggest that variable centromeric DNA content and/or epigenetic assembly can influence the mitotic behavior of artificial chromosomes