Repeated sequences in the DNA of Drosophila and their localization in giant chromosomes

It is shown by isopycnic density gradient centrifugation that the DNAs of the sibling species Drosophila hydei, Drosophila neohydei and Drosophila pseudoneohydei differ regarding the numbers and proportions of satellite DNA bands. An overwhelming proportion of all repetitive nucleotide sequences of the DNA is contained in these satellite fractions. The majority of the satellites are species specific despite the close phylogenetic and cytological relationship between the three species studied. — By in situ hybridization experiments it is demonstrated that the various satellite sequences occupy different positions within the chromosomes. All types of localization patterns, from a wide spread occurrence in all chromosomes to an apparent restriction to kinetochore regions of single chromosomes, have been observed. Main band DNA, on the other hand, in its hybridization behavior reflects the DNA distribution according to the banding pattern in giant chromosomes. Generally satellite sequences seem to be included in -heterochromatic chromosome regions but no relation to the heterochromatin of the Y-chromosome was found. — Renaturation studies support various evidence that satellite sequences occur in tandemly repetitious units. At least some of this repetitious material seems to be linked to non-satellite DNA sequences or to DNA of other satellites.     

Comparative study of satellite sequences and phylogeny of five species from the genus Palorus (Insecta, Coleoptera).

Major satellite sequences are analysed in the three tenebrionid beetles Palorus cerylonoides, P. genalis, and P. ficicola, and compared with the ones from P. ratzeburgii and P. subdepressus reported elsewhere. All of them are A+T rich, pericentromerically located, and with lengths of about 150 bp, either in the form of monomers or formed by more complex repeating units. A preliminary phylogenetic analysis of Palorus species using the 3' end of the mitochondrial Cytochrome Oxidase I gene shows that the five Palorus species have been diverging for a considerable amount of evolutionary time, with the pair P. ratzeburgii and P. genalis being the most closely related. Only these two taxa showed some similarity between their respective high-copy-number satellite sequences, while other satellites are mutually unrelated and might have originated independently. However, all the satellites have in common tertiary structure induced by intrinsic DNA curvature, a characteristic which is conserved within the genus. Palorus major satellites were previously detected in the genomes of congeneric species as low-copy-number clusters (Mestrovi? et al., Mol. Biol. Evol. 15: 1062-1068. 1998). Given the divergences between the analysed species, the substitution rate deduced from high- and low-copy-number repeats is unexpectedly low. The presence of sequence-induced DNA curvature in all Palorus satellites and similar satellite DNAs in the species pair P. ratzeburgii and P. genalis suggest (i) that constraints are at the tertiary structure; and (ii) that the satellite DNA evolutionary turnover can be dependent on the history of the taxa under study, resulting in retention of similar satellites in related taxa.     

Evolution of satellite DNAs from the genus Palorus--experimental evidence for the "library" hypothesis

Satellite DNA profiles have been characterized in the congeneric species Palorus ratzeburgii, Palorus subdepressus, Palorus genalis, and Palorus ficicola (Coleoptera, Insecta), each of which contains a single, A + T-rich satellite DNA comprising a considerable portion of the genome (20%-40%). These satellites exhibit insignificant mutual sequence similarity. Using PCR assay, it has been shown that all four sequences are present in each of the tested Palorus species: one of them is amplified into a high copy number or a major satellite, while the three others are in the form of low-copy-number repeats estimated to make up approximately 0.05% of the genome. Each of the four satellites is interspecifically high conserved concerning the sequence, monomer length, and tandem repeat organization. Major, as well as low- copy-number, satellites are colocalized in the regions of pericentromeric heterochromatin on all chromosomes of the complement. The low-copy-number satellites are dispersed between the large arrays of the major satellite over the whole heterochromatic block. Our results explain satellite DNA evolution, confirming the hypothesis that related species share a "library" of conserved satellite sequences, some of which could be amplified into a major satellite. Due to the evolutionary dynamics of satellite DNAs, the content of the "library" is variable; the elimination of some sequences parallels the creation of the new ones. Quantitative changes in satellite DNAs, induced by occasional amplification of satellite repeat from the "library", could possibly occur in the course of the speciation process, thus forming a species-specific profile of satellite DNAs.      

Detection and location of three simple sequence DNAs in polytene chromosomes from virilis group species of Drosophila

In vitro synthesized RNAs complementary to the three satellite DNAs of Drosophila virilis have been used in a series of in situ hybridization experiments with polytene chromosomes from virilis group species. Gall and Atherton (1974) demonstrated that each of the satellites of D. virilis is comprised of many repeats of a distinct, seven base pair long, simple sequence. With few exceptions, copies of each of these simple sequences are detected in the chromocenters of all virilis group species. This is true even in species which do not possess satellite DNAs at buoyant densities corresponding to those of the satellite DNAs of D. virilis. Small quantities of the three simple sequences are also detected in euchromatic arms of several different species. The same euchromatic location may contain detectable copies of one, two, or all three simple sequence DNAs. The amounts of simple sequences at each location in the euchromatin may vary between species, between different stocks of the same species, and even between individuals of the same stock. The simple sequences located in the euchromatin appear to undergo DNA replication during formation of polytene chromosomes unlike those in heterochromatin. The locations of the euchromatic sequences are not the results of single chromosomal inversion events involving heterochromatic and euchromatic breakpoints.     

Compositional patterns in the nuclear genome of cold-blooded vertebrates

Summary DNA preparations obtained from 122 species of fishes, 5 species of amphibians, and 13 species of reptiles were investigated in their compositional properties by analytical equilibrium centrifugation in CsCl density gradients. These species represented 21 orders of Osteichthyes, 3 orders of Chondrichthyes, 2 orders of amphibians, and 3 orders of reptiles. Modal buoyant densities of fish DNAs ranged from 1.696 to 1.707 g/cm³, the vast majority of values falling, however, between 1.699 and 1.704 g/cm³, which is the range covered by the DNAs of amphibians and reptiles. In all cases, DNA bands in CsCl were only weakly asymmetrical and only very rarely were accompanied by separate satellite bands (mostly on the GC-rich side). Intermolecular compositional heterogeneities were low in the vast majority of cases, and, like CsCl band asymmetries, at least partially due to cryptic or poorly resolved satellites. The present findings indicate, therefore, that DNAs from cold-blooded vertebrates are characterized by a number of common properties, namely a very wide spectrum of modal buoyant densities, low intermolecular compositional heterogeneities, low CsCl band asymmetries, and, in most cases, small amounts of satellite DNAs. In the case of fish DNAs a negative correlation was found between the GC level and the haploid size (c value) of the genome. If polyploidization is neglected, this phenomenon appears to be mainly due to the fact that increases and decreases in GC are associated with contraction and expansion phenomena, respectively, of intergenic noncoding sequences, which are GC poor relative to coding sequences.     

Behaviour of sex chromosome associated satellite DNAs in somatic and germ cells in snakes

Sex chromosome associated satellite DNAs isolated from the snakes Elaphe radiata (sat III) (Singh et al., 1976) and Bungarus fasciatus (Elapidae) (minor satellite) are evolutionarily conserved throughout the suborder Ophidia. An autosome limited satellite DNA (B. fasciatus major satellite) is not similarly conserved. Both types of satellites have been studied by in situ hybridisation in various somatic tissues and germ cells where it has been observed that the W sex chromosome remains condensed in interphase nuclei. In growing oocytes however, the W chromosome satellite rich heterochromatin decondenses completely whilst the autosomal satellite rich regions remain condensed. Later, the cycle is reversed and the W chromosome condenses whilst the autosomal satellite regions decondense. In a primitive snake (Eryx johni johni) where the sex chromosomes are not differentiated and where there is no satellite DNA specific to them, these phenomena are absent. — The differential behaviour of autosomal and sex chromosome associated satellite DNAs is discussed in the light of gene regulation.     

Interspecies comparison of the highly-repeated DNA of Australasian Luzula (Juncaceae)

The woodrush genus Luzula is characterised by having holocentric chromosomes. DNA of nine related Australasian species shows similar satellite DNAs which re very similar in nucleotide sequence content and unit length. Differences between the repetitive DNAs are evident as either the presence or absence of particular restriction enzyme sites. Sequence variants have probably been introduced into the repeated DNA components of ancestral species and particular variants reamplified during the evolution of the genus. Sequence amplification appears to be restricted to sequences already present in the genome rather thant the de novo generation of repeats. The evolution of highly-repeated DNA sequences dispersed throughout the holocentric chromosomes of Luzula thus appears to be very similar to that known in eukaryotes with the more normal monocentric chromosome organisation.     

Evolution of Satellite DNAs in a Radiation of Endemic Hawaiian Spiders: Does Concerted Evolution of Highly Repetitive Sequences Reflect Evolutionary History?

Satellite DNAs are known for an unusual and nonuniform evolution characterized by rapid evolutionary change between species and concerted evolution leading to molecular homogeneity within species. In this paper we use satellite DNAs for phylogenetic analysis of a rapidly evolving lineage of spiders and compare the phylogeny with a hypothesis previously generated based on mitochondrial DNA and allozymes. The spiders examined include almost all species within a monophyletic clade of endemic Hawaiian Tetragnatha species, the spiny-leg clade. The phylogeny based on satellite sequences is largely congruent to those produced by mtDNA and allozymes, except that the satellite DNA yields much longer branches, with higher levels of support for any given node. Closely related species that have differentiated ecologically within an island are well resolved with satellite DNA but much less so with mtDNA. These results suggest that Tetragnatha stDNA repeats seem to be evolving gradually and cohesively during the diversification of these endemic Hawaiian spiders. The study also reveals gain–loss of satellite DNA copies during species diversification. We conclude that satellite DNA sequences may potentially be very useful for resolving relationships between rapidly evolving taxa within an adaptive radiation. In addition, satellite DNA as a nuclear marker suggests that hybridization or peripatry could play a possible role in species formation that cannot be revealed by mitochondrial markers due to its maternal inheritance.This article contains online supplementary data.Reviewing Editor: Dr. Rafael Zardoya     

Satellite DNA in the sheep and goat

Sheep and goats possess closely similar satellite DNAs which seem to be concentrated in the heterochromatin and the nucleolus. In both species, the major and minor satellites have buoyant densities of 1.711 g·cm ^–3 and 1.718 g·cm ^–3 and represent 12% and 2.5%, respectively, of the total DNA. Major satellite concentration in the combined heterochromatin and nucleolar fraction was 19%, and in the pure nucleolar fraction it was 51%. Most hitherto reported satellite DNAs have varied markedly in density and concentration from species to species, even within the same genus. The close similarity of the sheep and goat satellite is discussed in the light of what is known about the evolutionary relationship between the sheep and the goat and in the light of various hypotheses concerning the possible functions of satellite DNA.     

Analysis of DNA from related and diverse species of Barley

DNAs of three closely related diploid species Hordeum vulgare, H. agriocrithon and H. spontaneum were compared among themselves and with a group of three species related to each other H. californicum (2x), H. jubatum (4x), and H. arizonicum (6x) having one genome in common.Buoyant densities of the DNAs from these species fall within a narrow range (1.700–1.701 g cm⁻³). Melting temperatures (T_m) of the DNAs are also similar (85.2–86.2°C). None of the DNAs showed any satellite band in neutral Cesium chloride gradients. In silver-cesium sulphate gradients, however, DNAs of all the species formed satellites on both the light and heavy sides of the main. band The diploid series have similar but not identical satellite patterns. The members of the polyploid series showed similar satellite patterns among themselves but distinct from those of the diploid series. In the polyploid group the amount of satellite showed a progressive decrease with the increase in the ploidy.The amount of repeated DNA (reassociating up to Cot 100) in H. vulgare was 69% and in H. arizonicum 65% of the total. The two species differed in their Cot curves. The low Cot fractions of H. arizonicum contained components of low buoyant densities which were absent in the corresponding Cot fractions of H. vulgare DNA.     

The Alu I-induced bands in metaphase chromosomes of orangutan (Pongo pygmaeus)

Summary Restriction endonucleases have been recently proved to be active on fixed chromosomes, thus they are useful in chromatin structure studies. Within this class of enzymes, Alu I is able to detect the presence and localization of highly repetitive DNA sequences in human and in other mammalian and dipteran species. In this paper the pattern obtained on fixed metaphase chromosomes of orangutan (Pongo pygmaeus) by Alu I digestion and Giemsa staining is shown. The results are discussed in the light of the distribution, in this species, of the I–IV human satellite DNAs. It is also suggested that in Pongo some highly repetitive sequences, different from the major human satellites, are present.     

Satellite DNA and chromosomes in Neotropical fishes: methods,applications and perspectives

Constitutive heterochromatin represents a substantial portion of the eukaryote genome, and it is mainly composed of tandemly repeated DNA sequences, such as satellite DNAs, which are also enriched by other dispersed repeated elements, including transposons. Studies on the organization, structure, composition and in situ localization of satellite DNAs have led to consistent advances in the understanding of the genome evolution of species, with a particular focus on heterochromatic domains, the diversification of heteromorphic sex chromosomes and the origin and maintenance of B chromosomes. Satellite DNAs can be chromosome specific or species specific, or they can characterize different species from a genus, family or even representatives of a given order. In some cases, the presence of these repeated elements in members of a single clade has enabled inferences of a phylogenetic nature. Genomic DNA restriction, using specific enzymes, is the most frequently used method for isolating satellite DNAs. Recent methods such as C₀t–1 DNA and chromosome microdissection, however, have proven to be efficient alternatives for the study of this class of DNA. Neotropical ichthyofauna is extremely rich and diverse enabling multiple approaches with regard to the differentiation and evolution of the genome. Genome components of some species and genera have been isolated, mapped and correlated with possible functions and structures of the chromosomes. The 5SHindIII‐DNA satellite DNA, which is specific to Hoplias malabaricus of the Erythrinidae family, has an exclusively centromeric location. The As51 satellite DNA, which is closely correlated with the genome diversification of some species from the genus Astyanax, has also been used to infer relationships between species. In the Prochilodontidae family, two repetitive DNA sequences were mapped on the chromosomes, and the SATH 1 satellite DNA is associated with the origin of heterochromatic B chromosomes in Prochilodus lineatus. Among species of the genus Characidium and the Parodontidae family, amplifications of satellite DNAs have demonstrated that these sequences are related to the differentiation of heteromorphic sex chromosomes. The possible elimination of satellite DNA units could explain the genome compaction that occurs among some species of Neotropical Tetraodontiformes. These topics are discussed in the present review, showing the importance of satellite DNA analysis in the differentiation and karyotype evolution of Actinopterygii.     

A tandemly repetitive centromeric DNA sequence of the fish Hoplias malabaricus (Characiformes: Erythrinidae) is derived from 5S rDNA

A substantial fraction of the eukaryotic genome consists of repetitive DNA sequences that include satellites, minisatellites, microsatellites, and transposable elements. Although extensively studied for the past three decades, the molecular forces that generate, propagate and maintain repetitive DNAs in the genomes are still discussed. To further understand the dynamics and the mechanisms of evolution of repetitive DNAs in vertebrate genome, we searched for repetitive sequences in the genome of the fish species Hoplias malabaricus. A satellite sequence, named 5SHindIII-DNA, which has a conspicuous similarity with 5S rRNA genes and spacers was identified. FISH experiments showed that the 5S rRNA bona fide gene repeats were clustered in the interstitial position of two chromosome pairs of H. malabaricus, while the satellite 5SHindIII-DNA sequences were clustered in the centromeric position in nine chromosome pairs of the species. The presence of the 5SHindIII-DNA sequences in the centromeres of several chromosomes indicates that this satellite family probably escaped from the selective pressure that maintains the structure and organization of the 5S rDNA repeats and become disperse into the genome. Although it is not feasible to explain how this sequence has been maintained in the centromeric regions, it is possible to hypothesize that it may be involved in some structural or functional role of the centromere organization.     

Satellite DNA relationships in man and the primates

We have investigated the genomes of a series of primates to identify the presence of sequences related to human satellite DNAs I, II and III by restriction enzyme digestion and hybridisation with probes of these satellite DNAs. Where we have found such related sequences we have examined the extent to which they have diverged by measuring the stability of the hybrids. DNA satellite III is the oldest sequence being common to species which have diverged some 24 million years ago. In contrast DNA satellites I and II are of much more recent origin. Our results permit us to draw conclusions about the way these sequences have evolved, and how the evolution of repeated DNA sequences may be related to the evolution of the primate lineage.     

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     

The biological significance of variation in satellite DNA and heterochromatin in newts of the genus Triturus: an evolutionary perspective

The functional and evolutionary significance of highly repetitive, simple sequence (satellite) DNA is analysed by examining available information on the patterns of variation of heterochromatin and cloned satellites among newts (family Salamandridae), and particularly species of the European genus Triturus. This information is used to develop a model linking evolutionary changes in satellite DNAs and chromosome structure. In this model, satellites accumulate initially in large tandem blocks around centromeres of some or all of the chromosomes, mainly by repeated chromosomal exchanges in these regions. Centromeric blocks later become broken up and dispersed by small, random chromosome rearrangements in these regions. They are dispersed first to pericentric locations and then gradually more distally into the chromosome arms and telomeres. Dispersal of a particular satellite is accompanied by changes in sequence structure (for example, base substitutions, deletions, etc.) and a corresponding decrease in its detectability at either the molecular or cytological level. On the basis of this model, observed satellites in newt species may be classified as 'old', 'young', or of 'intermediate' phylogenetic age. The functions and effects of satellite DNA and heterochromatin at the cellular and organismal levels are also discussed. It is suggested that satellite DNA may have an impact on cell proliferation through the effect of late-replicating satellite-rich heterochromatin on the duration of S-phase of the cell cycle. It is argued that even small alterations in cell cycle time due to changes in heterochromatin amount may have magnified effects on organismal growth that may be of adaptive significance.     

Satellite DNA-correlated nucleosomal proteins in Drosophila virilis

Three major satellite DNAs comprise 40–45% of the genome of Drosophila virilis. Since these satellites are not substrates for most restriction enzymes, we were able to digest D. virilis nuclei with HaeIII and micrococcal nuclease and isolate chromatin fractions containing variable levels of satellite DNA. Electrophoretic analysis of these chromatin fractions revealed that the level of the acid-soluble chromosomal protein, cp17.3, was directly related to the percentage of satellite DNA in chromatin. The correlation between cp17.3 and satellite DNA abundance suggests that cp17.3 is involved in the heterochromatic condensation of satellite DNAs. cp17.3 occurs at a frequency of one molecule per 10–20 nucleosomes. It is detected in an electrophoretically distinguishable class of mononucleosomes, provisionally identified as MN1uH2A, which contains ubiquitinated histone H2A (uH2a) but lacks histone H1. It is not detected in MN1, a second class of mononucleosomes, which lacks uH2A and H1. Since cp17.3 is correlated with satellite DNAs and present in nucleosome cores, it might be a histone variant specifically associated with satellite DNAs.This work was supported by Grant GM22138 from the National Institutes of Health. G.A.V. was a predoctoral trainee supported by Grant GM07094 from the National Institutes of Health.     

Isolation and comparison of tribe-specific centromeric repeats within Bovidae

A taxonomic division of the family Bovidae (Artiodactyla) is difficult and the evolutionary relationships among most bovid subfamilies remain uncertain. In this study, we isolated the cattle satellite I clone BTREP15 (1.715 satellite DNA family) and autosomal centromeric DNAs of members of ten bovid tribes. We wished to determine whether the analysis of fluorescence in situ hybridization patterns of the cattle satellite I clone (BTREP15) and tribe-specific centromeric repeats isolated by laser microdissection would help to reveal some of the ambiguities occurring in the systematic classification of the family Bovidae. The FISH study of the presence and distribution of the cattle satellite I clone BTREP15 (1.715 satellite DNA family) within members of ten bovid tribes was not informative. FISH analysis of autosomal centromeric DNA probes in several species within one tribe revealed similar hybridization patterns in autosomes confirming tribal homogeneity of these probes. Sex chromosomes showed considerable variation in sequence composition and arrangement not only between tribes but also between species of one tribe. According to our findings it seems that Oreotragus oreotragus developed its own specific satellite DNA which does not hybridize to any other bovid species analysed. Our results suggest O. oreotragus as well as Aepyceros melampus may be unique species not particularly closely related to any of the recognized bovid tribes. This study indicates the isolation of tribe-specific centromeric DNAs by laser microdissection and cloning the sequence representing the main motif of these repetitive DNAs could offer the perspectives for comparative phylogenetic studies.     

Wide distribution of related satellite DNA families within the genus <Emphasis Type="Italic">Pimelia</Emphasis> (Tenebrionidae)

Major satellites of species in the genus Pimelia comprise large portions of their genomes and belong to seven major satellite families which all originate from a common ancestral sequence. Here we present the results of comprehensive screening of 26 Pimelia species belonging to three distinct geographic groups (Ibero-Balearic, African and Canary Islands) for the presence of different Pimelia satellite families in their genomes. Dot-blot hybridization experiments suggest that together with one dominant, highly abundant satellite family, other families are also present in genomes of the majority of examined Pimelia species, but as low-copy number repeats. The estimated abundance of these underrepresented repeats is about 4,000 copies per haploid genome. Signals of highly abundant satellite family from P. scabrosa (PSCA) in examined congeneric species, obtained after PCR amplification and Southern hybridization under high stringency conditions, corroborate sequence preservation of low-copy representatives of satellite families. PRINS localized low-copy repeats within the pericentromeric regions of all chromosomes. These results point to the existence of an extensive library of repetitive DNAs that was already present in the genome of the common ancestor of extant Pimelia taxa, and shifts the period of diversification of Pimelia satellites far in the history of this genus.