Sobo, a recently amplified satellite repeat of potato, and its implications for the origin of tandemly repeated sequences

Highly repetitive satellite DNA sequences are main components of heterochromatin in higher eukaryotic genomes. It is well known that satellite repeats can expand and contract dramatically, which may result in significant genome size variation among genetically related species. The origin of satellite repeats, however, is elusive. Here we report a satellite repeat, Sobo, from a diploid potato species, Solanum bulbocastanum. The Sobo repeat is mapped to a single location in the pericentromeric region of chromosome 7. This single Sobo locus spans approximately 360 kb of a 4.7-kb monomer. Sequence analysis revealed that the major part of the Sobo monomer shares significant sequence similarity with the long terminal repeats (LTRs) of a retrotransposon. The Sobo repeat was not detected in other Solanum species and is absent in some S. bulbocastanum accessions. Sobo monomers are highly homogenized and share >99% sequence identity. These results suggest that the Sobo repeat is a recently emerged satellite and possibly originated by a sudden amplification of a genomic region including the LTR of a retrotransposon and its flanking genomic sequences.     

Chromosomal location by in situ hybridization of the human Sau3A family of DNA repeats

Summary The Sau3A family is a human, clustered, highly repetitive, GC-rich DNA family. In situ hybridization studies with a plasmid carrying a Sau3A monomer as a probe have shown that Sau3A sequences are preferentially concentrated in the heterochromatic regions of human acrocentric chromosomes (D and G groups, both in pericentromeric regions and in cytological satellites) and in pericentromeric heterochromatin of chromosome 1. The same chromosomal locations were observed by using as probes two recombinant phages which carry Sau3A-positive genomic sectors. The two sectors differfor the relative proportions of monomer and multiples of Sau3A repeats, which show different extents of homology to the cloned monomer, and for the presence, in one of the two, of a samll amount of an unrelated repeat (alphoid DNA). The similarity of the results obtained with the three probes suggests that heterogeneous Sau3A repeats share the same chromosomal localizations and that the two analyzed genomic sectors may not contain significant amounts of repetitive DNAs other than the Sau3A family. A comparison between the chromosomal locations of Sau3A and EcoRI families of repeats has confirmed that each family is characterized by specific chromosomal locations and that single heterochromatic regions may contain both.     

Pericentromeric regions of soybean (Glycine max L. Merr.) chromosomes consist of retroelements and tandemly repeated DNA and are structurally and evolutionarily labile

Little is known about the physical makeup of heterochromatin in the soybean (Glycine max L. Merr.) genome. Using DNA sequencing and molecular cytogenetics, an initial analysis of the repetitive fraction of the soybean genome is presented. BAC 076J21, derived from linkage group L, has sequences conserved in the pericentromeric heterochromatin of all 20 chromosomes. FISH analysis of this BAC and three subclones on pachytene chromosomes revealed relatively strict partitioning of the heterochromatic and euchromatic regions. Sequence analysis showed that this BAC consists primarily of repetitive sequences such as a 102-bp tandem repeat with sequence identity to a previously characterized approximately 120-bp repeat (STR120). Fragments of Calypso-like retroelements, a recently inserted SIRE1 element, and a SIRE1 solo LTR were present within this BAC. Some of these sequences are methylated and are not conserved outside of G. max and G. soja, a close relative of soybean, except for STR102, which hybridized to a restriction fragment from G. latifolia. These data present a picture of the repetitive fraction of the soybean genome that is highly concentrated in the pericentromeric regions, consisting of rapidly evolving tandem repeats with interspersed retroelements.     

The centromeric regions of potato chromosomes contain megabase-sized tandem arrays of telomere-similar sequence

Telomere-similar sequences have been found in non-telomeric regions in various eukaryotic species. Centromeric regions often harbor such interstitial telomeric repeats (ITRs). We isolated a 2.8 kb ITR, pSbTC1, in a diploid potato species Solanum bulbocastanum. DNA sequences related to the pSbTC1 family are widely distributed in different Solanum species. The pSbTC1-related sequences are organized into tandem arrays and located mainly in the centromeric regions of potato chromosomes. Most notably, the pSbTC1-related sequences have undergone extensive amplification and a single array can span up to multiple megabases. These results suggest that the pSbTC1-related sequences are not simple relics of ancient events in karyotype evolution, such as chromosomal fusions. We also demonstrated that the pSbTC1-related sequences are heavily methylated and are associated with highly condensed centromeric heterochromatin.     

Structural organization and functional analysis of centromeric DNA in the fission yeast Schizosaccharomyces pombe.

Centromeric DNA in the fission yeast Schizosaccharomyces pombe was isolated by chromosome walking and by field inversion gel electrophoretic fractionation of large genomic DNA restriction fragments. The centromere regions of the three chromosomes were contained on three SalI fragments (120 kilobases [kb], chromosome III; 90 kb, chromosome II; and 50 kb, chromosome I). Each fragment contained several repetitive DNA sequences, including repeat K (6.4 kb), repeat L (6.0 kb), and repeat B, that occurred only in the three centromere regions. On chromosome II, these repeats were organized into a 35-kb inverted repeat that included one copy of K and L in each arm of the repeat. Site-directed integration of a plasmid containing the yeast LEU2 gene into K repeats at each of the centromeres or integration of an intact K repeat into a chromosome arm had no effect on mitotic or meiotic centromere function. The centromeric repeat sequences were not transcribed and possessed many of the properties of constitutive heterochromatin. Thus, S. pombe is an excellent model system for studies on the role of repetitive sequence elements in centromere function.     

Generalized linear mixed model for segregation distortion analysis

Background

Concerted evolution refers to the pattern in which copies of multigene families show high intraspecific sequence homogeneity but high interspecific sequence diversity. Sequence homogeneity of these copies depends on relative rates of mutation and recombination, including gene conversion and unequal crossing over, between misaligned copies. The internally repetitive intergenic spacer (IGS) is located between the genes for the 28S and 18S ribosomal RNAs. To identify patterns of recombination and/or homogenization within IGS repeat arrays, and to identify regions of the IGS that are under functional constraint, we analyzed 13 complete IGS sequences from 10 individuals representing four species in the Daphnia pulex complex.

Results

Gene conversion and unequal crossing over between misaligned IGS repeats generates variation in copy number between arrays, as has been observed in previous studies. Moreover, terminal repeats are rarely involved in these events. Despite the occurrence of recombination, orthologous repeats in different species are more similar to one another than are paralogous repeats within species that diverged less than 4 million years ago. Patterns consistent with concerted evolution of these repeats were observed between species that diverged 8-10 million years ago. Sequence homogeneity varies along the IGS; the most homogeneous regions are downstream of the 28S rRNA gene and in the region containing the core promoter. The inadvertent inclusion of interspecific hybrids in our analysis uncovered evidence of both inter- and intrachromosomal recombination in the nonrepetitive regions of the IGS.

Conclusions

Our analysis of variation in ribosomal IGS from Daphnia shows that levels of homogeneity within and between species result from the interaction between rates of recombination and selective constraint. Consequently, different regions of the IGS are on substantially different evolutionary trajectories.     

High-resolution organization of mouse telomeric and pericentromeric DNA

We studied the organization of telomeric, major and minor satellite DNA sequences located in the pericentromeric regions of mouse telocentric and Robertsonian metacentric chromosomes by high-resolution fluorescence in situ hybridization. Molecular data have already proved that in telocentrics, from the physical chromosome end, telomeric sequences are followed by minor and then by major satellite DNA. We showed that the three families of repetitive DNA are organized as uninterrupted long-range cluster repeats and that there is no intermingling between telomeric and minor satellite DNA or between the major and the minor tandem repeats or with non-satellite DNA. The pericentromeric region of metacentric chromosomes consists of a small block of minor satellite DNA sandwiched between two blocks of major satellite DNA.     

Human (Homo sapiens) and chimpanzee (Pan troglodytes) share similar ancestral centromeric alpha satellite DNA sequences but other fractions of heterochromatin differ considerably

The euchromatic regions of chimpanzee (Pan troglodytes) genome share approximately 98% sequence similarity with the human (Homo sapiens), while the heterochromatic regions display considerable divergence. Positive heterochromatic regions revealed by the CBG-technique are confined to pericentromeric areas in humans, while in chimpanzees, these regions are pericentromeric, telomeric, and intercalary. When human chromosomes are digested with restriction endonuclease AluI and stained by Giemsa (AluI/Giemsa), positive heterochromatin is detected only in the pericentromeric regions, while in chimpanzee, telomeric, pericentromeric, and in some chromosomes both telomeric and centromeric, regions are positive. The DA/DAPI technique further revealed extensive cytochemical heterogeneity of heterochromatin in both species. Nevertheless, the fluorescence in situ hybridization technique (FISH) using a centromeric alpha satellite cocktail probe revealed that both primates share similar pericentromeric alpha satellite DNA sequences. Furthermore, cross-hybridization experiments using chromosomes of gorilla (Gorilla gorilla) and orangutan (Pongo pygmaeus) suggest that the alphoid repeats of human and great apes are highly conserved, implying that these repeat families were present in their common ancestor. Nevertheless, the orangutan's chromosome 9 did not cross-hybridize with human probe. The euchromatic regions of chimpanzee (Pan troglodytes) genome share approximately 98% sequence similarity with the human (Homo sapiens), while the heterochromatic regions display considerable divergence. Positive heterochromatic regions revealed by the CBG-technique are confined to pericentromeric areas in humans, while in chimpanzees, these regions are pericentromeric, telomeric, and intercalary. When human chromosomes are digested with restriction endonuclease AluI and stained by Giemsa (AluI/Giemsa), positive heterochromatin is detected only in the pericentromeric regions, while in chimpanzee, telomeric, pericentromeric, and in some chromosomes both telomeric and centromeric, regions are positive. The DA/DAPI technique further revealed extensive cytochemical heterogeneity of heterochromatin in both species. Nevertheless, the fluorescence in situ hybridization technique (FISH) using a centromeric alpha satellite cocktail probe revealed that both primates share similar pericentromeric alpha satellite DNA sequences. Furthermore, cross-hybridization experiments using chromosomes of gorilla (Gorilla gorilla) and orangutan (Pongo pygmaeus) suggest that the alphoid repeats of human and great apes are highly conserved, implying that these repeat families were present in their common ancestor. Nevertheless, the orangutan's chromosome 9 did not cross-hybridize with human probe. © 1995 Wiley-Liss, Inc.     

Repeats and subrepeats in the intergenic spacer of rDNA from the nematode Meloidogyne arenaria

Summary Ribosomal DNA (rDNA) repeats of the plant-parasitic nematode Meloidogyne arenaria are heterogeneous in size and appear to contain 5S rRNA gene sequences. Moreover, in a recA ⁺ bacterial host, plasmid clones of a 9 kb rDNA repeat show deletion events within a 2 kb intergenic spacer (IGS), between 28S and 5S DNA sequences. These deletions appear to result from a reduction in the number of tandem 129 by repeats in the IGS. The loss of such repeats might explain how rDNA length heterogeneity, observed in the Meloidogyne genome, could have arisen. Each 129 by repeat also contains three copies of an 8 by subrepeat, which has sequence similarity to an element found in the IGS repeats of some plant rDNAs.     

Constitutive heterochromatin polymorphisms in human chromosomes identified by whole comparative genomic hybridization

Whole comparative genomic hybridization (W-CGH) is a new technique that reveals cryptic differences in highly repetitive DNA sequences, when different genomes are compared using metaphase or interphase chromosomes. W-CGH provides a quick approach to identify differential expansion of these DNA sequences at the single-chromosome level in the whole genome. In this study, we have determined the frequency of constitutive chromatin polymorphisms in the centromeric regions of human chromosomes using a whole-genome in situ cross-hybridization method to compare the whole genome of five different unrelated individuals. Results showed that the pericentromeric constitutive heterochromatin of chromosome 6 exhibited a high incidence of polymorphisms in repetitive DNA families located in pericentromeric regions. The constitutive heterochromatin of chromosomes 5 and 9 was also identified as highly polymorphic. Although further studies are necessary to corroborate and assess the overall incidence of these polymorphisms in human populations, the use of W-CGH could be pertinent and of clinical relevance to assess rapidly, from a chromosomal viewpoint, genome similarities and differences in closely related genomes such as those of relatives, or in more specific situations such as bone marrow transplantation where chimerism is produced in the recipient.     

Characterization of a highly repeated DNA sequence family in five species of the genus Eulemur

The karyotypes of Eulemur species exhibit a high degree of variation, as a consequence of the Robertsonian fusion and/or centromere fission. Centromeric and pericentromeric heterochromatin of eulemurs is constituted by highly repeated DNA sequences (including some telomeric TTAGGG repeats) which have so far been investigated and used for the study of the systematic relationships of the different species of the genus Eulemur. In our study, we have cloned a set of repetitive pericentromeric sequences of five Eulemur species: E. fulvus fulvus (EFU), E. mongoz (EMO), E. macaco (EMA), E. rubriventer (ERU), and E. coronatus (ECO). We have characterized these clones by sequence comparison and by comparative fluorescence in situ hybridization analysis in EMA and EFU. Our results showed a high degree of sequence similarity among Eulemur species, indicating a strong conservation, within the five species, of these pericentromeric highly repeated DNA sequences.     

Junctions between repetitive DNAs on the PSR chromosome of Nasonia vitripennis: Association of palindromes with recombination

The Paternal-Sex-Ratio (PSR) chromosome of Nasonia vitripennis contains several families of repetitive DNAs that show significant sequence divergence but share two palindromic regions. This study reports on the analysis of junctions between two of these repetitive DNA families (psr2 and psr18). Three lambda clones that hybridized to both repeat families were isolated from PSR-genomic DNA libraries through multiple screenings and analyzed by Southern blots. Analysis of clones showed a region in which the two repeat types are interspersed, flanked by uniform blocks of each repeat type. PCR amplification of genomic DNA confirmed the contiguous arrangement of psr2 and psr18 on PSR and identified an additional junction region between these repeats that was not present in the lambda inserts. We isolated and sequenced 41 clones from the lambda inserts and genomic PCR products containing junction sequences. Sequence analysis showed that all transitions between psr2 and psr18 repeats occurred near one of the two palindromes. Based on the inheritance pattern of PSR, recombination between repeats on this chromosome must be mitotic (rather than meiotic) in origin. The occurrence of exchanges near the palindromes suggests that these sequences enhance recombination between repeat units. Rapid amplification of repetitive DNA may have been an important factor in the evolution of the PSR chromosome. Correspondence to: John H. Werren     

Localization of repetitive DNA sequences in the pericentromeric heterochromatin of malarial mosquitoes of the "Anopheles maculipennis" complex

Distribution of eight fragments of conserved repetitive DNA from pericentromeric heterochromatin of chromosome 2 of Anopheles atroparvus has been investigated by in situ hybridization on polytene chromosomes of An. atroparvus and An. messeae. We have shown that heterochromatic regions of all chromosomes both in An. atroparvus and An. messeae vary in combinations of, at least, conserved repeats. Some repeats have been found only in pericentromeric heterochromatic regions of chromosomes 2 (clones Atr2R-46a, Atr2R-73, Atr2R-85a in An. atroparvus and Atr2R-25 in An. messeae). Others have been found in two (clones Atr2R-25a and Atr2R-90 in An. atroparvus, Atr2R-25a in An. messeae) and more (clones Atr2R-118, Atr2R-136 in An. atroparvus, Atr2R-73 in An. messeae) pericentromeric heterochromatic regions of chromosomes. DNA comparison of pericentromeric heterochromatic regions of chromosomes in species of the "Anopheles maculipennis" complex is species- and chromosome-specific, due, in particular, to different maintenance of conserved repeates.