Higher-order repeats in the satellite DNA of the cave beetle Pholeuon proserpinae glaciale (Coleoptera: Cholevidae)

The present study characterizes the satellite DNA of the cave beetle Pholeuon proserpinae glaciale which represent about 3-5 % of its genome, and which is composed of monomers of 266 bp and 70.5 % A-T. Concerted evolution seems to act on a higher-order repeat, a dimer, composed of two types of 266-bp monomers that differ in three diagnostic sites. These dimers show a striking nucleotide identity (98.7 % similarity) suggesting strong homogenization processes. The presence of particular mutations shared by several dimers represents an early expansion of these types of repeats as proposed by the molecular drive model. Moreover, evidence of gene conversion tracts in P. proserpinae glaciale, which also could be the result of unequal sister chromatid exchange, would suggest that recombination is involved in the homogenization of stDNA sequences. The presence of a 17-bp-motif repeated six times, along another one of 31-bp repeated twice which also have embedded one 17-bp-motif, suggest that monomers have originated from those basic motifs.     

Molecular analysis of a polymorphic domain of alpha satellite from the human X chromosome.

Alpha satellite DNA, a diverse family of tandemly repeated DNA sequences located at the centromeric region of each human chromosome, is organized in a highly chromosome-specific manner and is characterized by a high frequency of restriction-fragment-length polymorphism. To examine events underlying the formation and spread of these polymorphisms within a tandem array, we have cloned and sequenced a representative copy of a polymorphic array from the X chromosome and compared this polymorphic copy with the predominant higher-order repeat form of X-linked alpha satellite. Sequence data indicate that the polymorphism arose by a single base mutation that created a new restriction site (for HindIII) in the sequence of the predominant repeat unit. This variant repeat unit, marked by the new HindIII site, was subsequently amplified in copy number to create a polymorphic domain consisting of approximately 500 copies of the variant repeat unit within the X-linked array of alpha satellite. We propose that a series of intrachromosomal recombination events between misaligned tandem arrays, involving multiple rounds of either unequal crossing-over or sequence conversion, facilitated the spread and fixation of this variant HindIII repeat unit.     

Chromosome-specific alpha satellite DNA from human chromosome 1: hierarchical structure and genomic organization of a polymorphic domain spanning several hundred kilobase pairs of centromeric DNA

The human alpha satellite repetitive DNA family is organized as distinct chromosome-specific subsets localized to the centromeric region of each chromosome. Here, we report he isolation and characterization of cloned repeat units which define a hierarchical subset of alpha satellite on human chromosome 1. This subset is characterized by a 1.9-kb higher-order repeat unit which consists of 11 tandem approximately 171-bp alpha satellite monomer repeat units. The higher-order repeat unit is itself tandemly repeated, present in at least 100 copies at the centromeric region of chromosome 1. Using pulsed-field gel electrophoresis we estimate the total array length of these tandem sequences at the centromere of chromosome 1 to be several hundred kilobase pairs. Under conditions of high stringency, the higher-order repeat probe hybridizes specifically to chromosome 1 and can be used to detect several associated restriction fragment length DNA polymorphisms. As such, this probe may be useful for molecular and genetic analyses of the centromeric region of human chromosome 1.     

Concerted evolution of primate alpha satellite DNA. Evidence for an ancestral sequence shared by gorilla and human X chromosome alpha satellite

To understand evolutionary events in the formation of higher-order repeat units in alpha satellite DNA, we have examined gorilla sequences homologous to human X chromosome alpha satellite. In humans, alpha satellite on the X chromosome is organized as a tandemly repeated, 2.0 x 10(3) base-pairs (bp) higher-order repeat unit, operationally defined by the restriction enzyme BamHI. Each higher-order repeat unit is composed of 12 tandem approximately 171 base-pair monomer units that have been classified into five distinct sequence homology groups. BamHI-digested gorilla genomic DNA hybridized with the cloned human 2 x 10(3) bp X alpha satellite repeat reveals three bands of sizes approximately 3.2 x 10(3), 2.7 x 10(3) and 2 x 10(3) bp. Multiple copies of all three repeat lengths have been isolated and mapped to the centromeric region of the gorilla X chromosome by fluorescence in situ hybridization. Long-range restriction mapping using pulsed-field gel electrophoresis shows that the 2.7 x 10(3) and 3.2 x 10(3) bp repeat arrays exist as separate but likely neighboring arrays on the gorilla X, each ranging in size from approximately 200 x 10(3) to 500 x 10(3) bp, considerably smaller than the approximately 2000 x 10(3) to 4000 x 10(3) bp array found on human X chromosomes. Nucleotide sequence analysis has revealed that monomers within all three gorilla repeat units can be classified into the same five sequence homology groups as monomers located within the higher-order repeat unit on the human X chromosome, suggesting that the formation of the five distinct monomer types predates the divergence of the lineages of contemporary humans and gorillas. The order of 12 monomers within the 2 x 10(3) and 2.7 x 10(3) bp repeat units from the gorilla X chromosome is identical with that of the 2 x 10(3) bp repeat unit from the human X chromosome, suggesting an ancestral linear arrangement and supporting hypotheses about events largely restricted to single chromosome types in the formation of alpha satellite higher-order repeat units.     

Interhomologue sequence variation of alpha satellite DNA from human chromosome 17: Evidence for concerted evolution along haplotypic lineages

Alpha satellite DNA is a family of tandemly repeated DNA found at the centromeres of all primate chromosomes. Different human chromosomes 17 in the population are characterized by distinct alpha satellite haplotypes, distinguished by the presence of variant repeat forms that have precise monomeric deletions. Pairwise comparisons of sequence diversity between variant repeat units from each haplotype show that they are closely related in sequence. Direct sequencing of PCR-amplified alpha satellite reveals heterogeneous positions between the repeat units on a chromosome as two bands at the same position on a sequencing ladder. No variation was detected in the sequence and location of these heterogeneous positions between chromosomes 17 from the same haplotype, but distinct patterns of variation were detected between chromosomes from different haplotypes. Subsequent sequence analysis of individual repeats from each haplotype confirmed the presence of extensive haplotype-specific sequence variation. Phylogenetic inference yielded a tree that suggests these chromosome 17 repeat units evolve principally along haplotypic lineages. These studies allow insight into the relative rates and/or timing of genetic turnover processes that lead to the homogenization of tandem DNA families. Correspondence to: H.F. Willard     

Differential homogenization and amplification of two satellite DNAs in the genus Cucurbita (Cucurbitaceae)

Two different satellite DNAs exist in the genus Cucurbita which are different with respect to repeat length (350 by and 170 bp), array size, and sequence homogenization. Whereas the 350-bp satellite DNA is prominent and very homogeneous in all species investigated except for C. maxima and C. lundelliana, the 170-bp satellite is rather evenly distributed in all species. In C. maxima and C. lundelliana the 350-bp satellite is present only in small amounts, but detectable by the sensitive PCR method. These repeats are also very homogeneous, reflecting a silent stage of satellite DNA. In contrast, the 170-bp satellite DNA is intra- and interspecifically heterogeneous. It is striking that the species with no detectable amount of 350-bp satellite contain 170-bp satellite DNA clusters with the highest degree of homogeneity. The evolution of satellite DNA repeats within cultivated and wild species in the genus Cucurbita is elucidated using the sequence data of both satellite DNAs from all species investigated. The value of satellite DNA for phylogenetic analysis between closely related species is discussed. Correspondence to: V. Hemleben     

Higher-Order Organization of Subrepeats and the Evolution of Cervid Satellite I DNA

Based on sequence analyses of 17 complete centromeric DNA monomers from ten different deer species, a model is proposed for the genesis, evolution, and genomic organization of cervid satellite I DNA. All cervid satellite I DNA arose from the initial amplification of a 31-bp DNA sequence. These 31-bp subrepeats were organized in a hierarchical fashion as 0.8-kb monomers in plesiometacarpalia deer and 1-kb monomers in telemetacarpalia deer. The higher-order repeat nature of cervid centromeric satellite DNA monomers accounts for their high intragenomic and intraspecific sequence conservation. Such high intraspecific sequence conservation validates the use of a single cervid satellite I DNA monomer from each deer species for interspecific sequence comparisons to elucidate phylogenetic relationships. Also, a specific 0.18-kb tandem duplication was observed in all 1-kb monomers, implying that 1-kb cervid satellite I DNA monomers arose from an unequal crossover event between two similar 0.8-kb ancestral DNA sequences. Received: 28 May 1996 / Accepted: 24 October 1996     

Structure, organization, and sequence of alpha satellite DNA from human chromosome 17: evidence for evolution by unequal crossing-over and an ancestral pentamer repeat shared with the human X chromosome.

The centromeric regions of all human chromosomes are characterized by distinct subsets of a diverse tandemly repeated DNA family, alpha satellite. On human chromosome 17, the predominant form of alpha satellite is a 2.7-kilobase-pair higher-order repeat unit consisting of 16 alphoid monomers. We present the complete nucleotide sequence of the 16-monomer repeat, which is present in 500 to 1,000 copies per chromosome 17, as well as that of a less abundant 15-monomer repeat, also from chromosome 17. These repeat units were approximately 98% identical in sequence, differing by the exclusion of precisely 1 monomer from the 15-monomer repeat. Homologous unequal crossing-over is suggested as a probable mechanism by which the different repeat lengths on chromosome 17 were generated, and the putative site of such a recombination event is identified. The monomer organization of the chromosome 17 higher-order repeat unit is based, in part, on tandemly repeated pentamers. A similar pentameric suborganization has been previously demonstrated for alpha satellite of the human X chromosome. Despite the organizational similarities, substantial sequence divergence distinguishes these subsets. Hybridization experiments indicate that the chromosome 17 and X subsets are more similar to each other than to the subsets found on several other human chromosomes. We suggest that the chromosome 17 and X alpha satellite subsets may be related components of a larger alphoid subfamily which have evolved from a common ancestral repeat into the contemporary chromosome-specific subsets.     

Genomic organization of alpha satellite DNA on human chromosome 7: evidence for two distinct alphoid domains on a single chromosome.

A complete understanding of chromosomal disjunction during mitosis and meiosis in complex genomes such as the human genome awaits detailed characterization of both the molecular structure and genetic behavior of the centromeric regions of chromosomes. Such analyses in turn require knowledge of the organization and nature of DNA sequences associated with centromeres. The most prominent class of centromeric DNA sequences in the human genome is the alpha satellite family of tandemly repeated DNA, which is organized as distinct chromosomal subsets. Each subset is characterized by a particular multimeric higher-order repeat unit consisting of tandemly reiterated, diverged alpha satellite monomers of approximately 171 base pairs. The higher-order repeat units are themselves tandemly reiterated and represent the most recently amplified or fixed alphoid sequences. We present evidence that there are at least two independent domains of alpha satellite DNA on chromosome 7, each characterized by their own distinct higher-order repeat structure. We determined the complete nucleotide sequences of a 6-monomer higher-order repeat unit, which is present in approximately 500 copies per chromosome 7, as well as those of a less-abundant (approximately 10 copies) 16-monomer higher-order repeat unit. Sequence analysis indicated that these repeats are evolutionarily distinct. Genomic hybridization experiments established that each is maintained in relatively homogeneous tandem arrays with no detectable interspersion. We propose mechanisms by which multiple unrelated higher-order repeat domains may be formed and maintained within a single chromosomal subset.     

Intragenic duplication and divergence in the spectrin superfamily of proteins

Many structural, signaling, and adhesion molecules contain tandemly repeated amino acid motifs. The alpha-actinin/spectrin/dystrophin superfamily of F-actin-crosslinking proteins contains an array of triple alpha-helical motifs (spectrin repeats). We present here the complete sequence of the novel beta-spectrin isoform beta(Heavy)- spectrin (beta H). The sequence of beta H supports the origin of alpha- and beta-spectrins from a common ancestor, and we present a novel model for the origin of the spectrins from a homodimeric actin-crosslinking precursor. The pattern of similarity between the spectrin repeat units indicates that they have evolved by a series of nested, nonuniform duplications. Furthermore, the spectrins and dystrophins clearly have common ancestry, yet the repeat unit is of a different length in each family. Together, these observations suggest a dynamic period of increase in repeat number accompanied by homogenization within each array by concerted evolution. However, today, there is greater similarity of homologous repeats between species than there is across repeats within species, suggesting that concerted evolution ceased some time before the arthropod/vertebrate split. We propose a two-phase model for the evolution of the spectrin repeat arrays in which an initial phase of concerted evolution is subsequently retarded as each new protein becomes constrained to a specific length and the repeats diverge at the DNA level. This evolutionary model has general applicability to the origins of the many other proteins that have tandemly repeated motifs.      

Chromosome-specific alpha satellites: two distinct families on human chromosome 18.

Two types of human chromosome 18-specific alpha satellite fragments have been cloned and sequenced. They represent closely related but distinct alphoid families formed by two different types of the higher-order repeated units (1360-bp EcoRI and 1700-bp HindIII fragments) that do not alternate in the genome. The individual repeats within each family are 99% identical and interfamily homology is about 78%. Sequence analysis shows that both repeats belong to alphoid suprachromosomal family 2, but their homology is not higher than that of family members located on different chromosomes. Therefore, the two repeats shared a common origin in the recent past, although they are not the direct offspring of one ancestral sequence. Our data indicate that these two 18-specific domains have appeared as a result of two separate amplification events. Despite the high degree of homology, they are not undergoing intrachromosomal homogenization, although some variation of this process might take place within each domain.     

Tandem-repetitive noncoding DNA: forms and forces

A model of sequence-dependent, unequal crossing-over and gene amplification (slippage replication) has been stimulated in order to account for various structural features of tandemly repeated DNA sequences. It is shown that DNA whose sequence is not maintained by natural selection will exhibit repetitive patterns over a wide range of recombination rates as a result of the interaction of unequal crossing-over and slippage replication, processes that depend on sequence similarity. At high crossing-over frequencies, the nucleotide patterns generated in the simulations are simple and highly regular, with short, nearly identical sequences repeated in tandem. Decreasing recombination rates increase the tendency to longer and more-complex repeat units. Periodicities have been observed down to very low recombination rates (one or more orders of magnitude lower than mutation rate). At such low rates, most of the sequences contain repeats which have an extensive substructure and a high degree of heterogeneity among each other; often higher-order structures are superimposed on a tandem array. These results are compared with various structural properties of tandemly repeated DNAs known from eukaryotes, the spectrum ranging from simple-sequence DNAs, particularly the hypervariable mini-satellites, to the classical satellite DNAs, located in chromosomal regions of low recombination, e.g., heterochromatin.     

Variation in satellite DNA profiles--causes and effects

Heterochromatic regions of the eukaryotic genome harbour DNA sequences that are repeated many times in tandem, collectively known as satellite DNAs. Different satellite sequences co-exist in the genome, thus forming a set called a satellite DNA library. Within a library, satellite DNAs represent independent evolutionary units. Their evolution can be explained as a result of change in two parameters: copy number and nucleotide sequence, both of them ruled by the same mechanisms of concerted evolution. Individual change in either of these two parameters as well as their simultaneous evolution can lead to the genesis of species-specific satellite profiles. In some cases, changes in satellite DNA profiles can be correlated with chromosomal evolution and could possibly influence the evolution of species.     

Diversity of a major repetitive DNA sequence in diploid and polyploid Triticeae

About 90 members of a major tandemly repeated DNA sequence family originally described in rye as pSc119.2 have been isolated from 11 diploid and polyploid Triticeae species using primers from along the length of the sequence for PCR amplification. Alignment and similarity analysis showed that the 120-bp repeat unit family is diverse with single nucleotide changes and few insertions and deletions occurring throughout the sequence, with no characteristic genome or species-specific variants having developed during evolution of the extant genomes. Fluorescent in situ hybridization showed that each of the large blocks of the repeat at chromosomal sites harboured many variants of the 120-bp repeat. There were substantial copy number differences between genomes, with abundant sub-terminal sites in rye, interstitial sites in the B genome of wheat, and relatively few sites in the A and D genome. We conclude that sequence homogenization events have not been operative in this repeat and that the common ancestor of the Triticeae tribe had multiple sequences of the 120-bp repeat with a range of variation not unlike that seen within and between species today. This diversity has been maintained when sites are moved within the genome and in all species since their divergence within the Triticeae.     

Molecular analysis of a deletion polymorphism in alpha satellite of human chromosome 17: evidence for homologous unequal crossing-over and subsequent fixation.

The human alpha satellite DNA family is organized into chromosome-specific subsets characterized by distinct higher-order repeats based on a approximately 171 basepair monomer unit. On human chromosome 17, the predominant form of alpha satellite is a 16-monomer (16-mer) higher-order repeat present in 500-1000 copies per chromosome 17. In addition, less abundant 15-monomer and 14-monomer repeats are also found constitutively on chromosome 17. Polymorphisms in the form of different higher-order repeat lengths have been described for this subset, the most prominent polymorphism being a 13-monomer (13-mer) higher-order repeat present on approximately 35% of all chromosomes 17. To investigate the nature of this polymorphism, we have cloned, sequenced and compared the relevant regions of the 13-mer to the previously characterized 16-mer repeat. The results show that the repeats are virtually identical, with the principal difference being the exclusion of three monomers from the 13-mer repeat. We propose that the 13-mer is the product of an isolated homologous recombination event between two monomers of the 16-mer repeat. Sequence comparisons reveal the approximate site of recombination and flanking regions of homology. This recombination site corresponds to a position within the alphoid monomer which has been previously implicated in an independent homologous recombination event, suggesting that there may exist a preferred register for recombination in alphoid DNA. We suggest that these events are representative of an ongoing process capable of reorganizing the satellite subset of a given chromosome, thereby contributing to the establishment of chromosome-specific alpha satellite subsets.     

Chromosome-specific alpha satellite DNA from the centromere of human chromosome 16

To examine the molecular organization of DNA sequences located in the centromeric region of human chromosome 16 we have isolated and characterized a chromosome 16-specific member of the alpha satellite DNA family. The probe obtained is specific for the centromere of chromosome 16 by somatic cell hybrid analysis and by fluorescence in situ hybridization and allows detection of specific hybridizing domains in interphase nuclei. Nucleotide sequence analysis indicates that this class of chromosome 16 alpha satellite (D16Z2) is organized as a series of diverged 340-bp dimers arranged in a tandem array of 1.7-kb higher-order repeat units. As measured by pulsed-field gel electrophoresis, the total D16Z2 array spans approximately 1,400-2,000 kb of centromeric DNA. These sequences are highly polymorphic, both by conventional agarose-gel electrophoresis and by pulsed-field gel electrophoresis. Investigation of this family of alpha satellite should facilitate the further genomic, cytogenetic, and genetic analysis of chromosome 16.     

Comparative structure and evolution of goat and sheep satellite I DNAs.

The satellite I DNAs of domestic goat (Capra hircus) and domestic sheep (Ovis aries) have been studied using molecular hybridisation and restriction enzyme analysis. Both satellite DNAs are composed of repeat units of 820 base pairs in length, but their restriction maps, although similar, differ in certain respects. Thus the majority of sheep satellite I repeat units have two EcoRI sites and one AluI site, whereas the majority of goat satellite I repeat units have one EcoRI site and two AluI sites. The sheep satellite I repeat units with the two EcoRI sites are much more homogeneous than the repeats forming the remainder of the satellite, as judged by the difference in the melting temperatures of native and reassociated molecules. DNAs from species of wild sheep and goats were screened for the presence of these repeat units, and they appear to have been amplified during the radiation of the Ovis genus. Goat satellite I is composed of a single sequence type which has changed through base substitution until the sequence now shows considerable heterogeneity. It is proposed that the major sequence types of these two satellite DNAs were amplified by different saltatory replication events at different times in the evolution of the group.     

Concerted evolution,a slow process for ant satellite DNA: study of the satellite DNA in the <Emphasis Type="Italic">Aphaenogaster</Emphasis> genus (Hymenoptera,Formicidae)

A family of satellite DNA is analyzed in seven ant species from the genus Aphaenogaster. This satellite DNA is organized as tandemly repeated sequences with a consensus sequence of 160 bp in length. The sampled sequences show a high similarity and belong to the same family of satellite DNA. However in Aphaenogaster spinosa, two types of repeat clearly differentiated have been found. Phylogenetic analyses using satellite DNA show that sequences do not cluster in a species-specific way, with one exception. Concretely, the second type of repeats of A. spinosa (APSP-II) which constitutes a new satellite DNA subfamily. The obtained results with satellite DNA are compared with those obtained using mitochondrial and nuclear DNA to determine the correlation between evolution of satellite DNA and phylogenetic relationships among the analyzed ants. The high interspecific similarity for the satellite DNA seems not to be in concordance with the concerted evolution pattern, commonly accepted to explain the evolution of satellite DNA. However, the accumulated data suggest that evolution of satellite DNA in ants follows the concerted evolution pattern but that this process is slow in relation with other organisms, probably due to the eusociality and haplodiploidy of these insects.     

Evolution and recombination of bovine DNA repeats

The history of the abundant repeat elements in the bovine genome has been studied by comparative hybridization and PCR. The Bov-A and Bov-B SINE elements both emerged just after the divergence of the Camelidae and the true ruminants. A 31-bp subrepeat motif in satellites of the Bovidae species cattle, sheep, and goat is also present in Cervidae (deer) and apparently predates the Bovidae. However, the other components of the bovine satellites were amplified after the divergence of the cattle and the Caprinae (sheep and goat). A 23-bp motif, which as subrepeat of two major satellites occupies 5% of the cattle genome, emerged only after the split of the water buffalo and other cattle species. During the evolution of the Bovidae the satellite repeat units were shaped by recombination events involving subrepeats, other satellite components, and SINE elements. Differences in restriction sites of homologous satellites indicate a continuing rapid horizontal spread of new sequence variants. Correspondence to: J.A. Lenstra     

L1 repeat elements in the human epsilon-G gamma-globin gene intergenic region: sequence analysis and concerted evolution within this family

We have deduced the sequence of a composite long interspersed repeated DNA in primates and herein describe its relationship to a complex repeat element (L1Heg) located in the interval linking the human epsilon- and G gamma-globin genes. The main element of L1Heg is 3' truncated and interrupted by the insertion of the 3' end of a second L1 element. Transposition of L1Heg into this intergenic locus generated a 62-bp duplication of flanking sequences. In contrast, insertion of the second repeat may have been mediated by homology between donor and target sequences. The main repeat represents a novel class of abundant elements whose sequences have diverged from other rodent and primate LINES approximately 1.3 kb downstream from the 5' terminus of L1Heg. Comparison of L1Heg with the sequences of two other related L1 members revealed a complex set of rearrangements confined within a region that resembles the long terminal repeats of other types of retroposons. The boundaries of conversion-like events were defined on the basis of the clustering of nucleotide sequence variants common to two or more nonallelic 3' L1H elements. Several of these events are apparently initiated or resolved within a common 150-bp region that coincides with the 3' terminus of a pan-mammalian open reading frame. This analysis showed that concerted genetic interactions and random drift both contribute appreciably to sequence variation within this set of L1H members.