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.     

Restriction endonuclease and molecular analyses of three rat genomes with special reference to chromosome rearrangement and speciation problems

When differences are found between related species of organisms, it is often assumed that the differences themselves are causal factors either in speciation itself or in processes related to speciation. Two recent proposals on the functions of satellite DNA (Hatch et al., 1976 and Fry and Salser, 1977) are that (a) large amounts of satellite DNA are important in facilitating chromosome rearrangements and hence cytogenetic evolution, and (b) satellite DNA differences between homologous chromosomes lead to pairing difficulties and are important in generating infertility barriers and hence speciation. If these proposals were to have some generality, one could expect organisms with very low amounts of highly repeated DNA to exhibit few chromosome rearrangements and to be evolutionarily conservative in a cyto-genetic sense. — We have chosen two very closely related species of rat which are phenotypically almost indistinguishable and which have undergone massive genome reorganization. They differ by 11 major centric rearrangements (2n=32, 2n=50). We have characterised their genomes by restriction endonuclease digestions, thermal denaturations, analytical ultracentrifugations and reassociation techniques, and have found that they have virtually no highly repeated DNA. Thus the 11 major chromosomal rearrangements have been fixed in present day genomes with hardly any highly repeated DNA, centric or otherwise. — It appears therefore that a large amount of highly repeated DNA is not obligatory for the formation and fixation of chromosome rearrangements. In addition, the existing literature reveals that one can find almost any situation at all, from species groups with high amounts of satellite DNA and no gross chromosomal rearrangements, to ones such as those described here, with tiny amounts of highly repeated DNA and massive chromosomal reorganisation. Since direct experimental data indicates that satellite DNA differences per se between homologous chromosomes do not cause infertility, speculations concerning modes of speciation based on satellite DNA differences between otherwise homologous chromosomes would appear to be ill founded.     

Sex chromosome associated satellite DNA: Evolution and conservation

Satellites visible in female but not in male DNA were isolated from the snakesElaphe radiata (satellite IV, p = 1.708 g · cm^–3) andBungarus fasciatus (BK¹ minor, p=1.709 g · cm^–3). The satellites cross hybridize. Hybridization of³H labelled nick translated BK minor satellite DNA with the total male and female DNA and/or chromosomes in situ of different species of snakes revealed that its sequences are conserved throughout the snake group and are mainly concentrated on the W chromosome. Snakes lacking sex chromosomes do possess related sequences but there is no sex difference and visible related satellites are absent. The following conclusions have been reached on the basis of these results. 1. The W chromosome associated satellite DNA is related to similar sequences scattered in the genome. 2. The origin and increment in the number of the W satellite DNA sequence on the W chromosome is associated with the heterochromatinization of the W. 3. Satellite sequences have become distributed along the length of the W and resulted in morphological differentiation of sex chromosomes. 4. Evolutionary conservation of W satellite DNA strongly suggests that functional constraints may have limited sequence divergence.     

Analysis of alphoid DNA variation and kinetochore size in human chromosome 21: evidence against pathological significance of alphoid satellite DNA diminutions.

Centromeric alpha satellite DNA sequences are linked to the kinetochore CENP-B proteins and therefore may be involved in the centromeric function. The high heterogeneity of size of the alphoid blocks raises the question of whether small amount of alphoid DNA or "deletion" of this block may have a pathological significance in the human centromere. In the present study, we analysed the correlation between size variations of alphoid DNA and kinetochore sizes in human chromosome 21 by molecular cytogenetic and immunochemical techniques. FISH analyses of alpha satellite DNA sizes in chromosome 21 homologues correlated well with the variation of their physical size as determined by pulsed field gel electrophoresis (PFGE). By contrast, the immunostaining study of the same homologous chromosomes with antikinetochore antibodies suggested that there is no positive correlation between the alpha satellite DNA block and kinetochore sizes. FISH analysis of chromosome 21-specific alphoid DNA and immunostaining of kinetochore extended interphase chromatin fibers indicate that centromeric kinetochore-specific proteins bind to restricted areas of centromeric DNA arrays. Thus, probably, restricted regions of centromeric DNA play an important role in kinetochore formation, centromeric function and abnormal chromosome segregation leading to non-disjunction.     

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.     

Characterization and chromosomal distribution of satellite DNA sequences of the water buffalo (Bubalus bubalis).

Satellite DNA sequences were isolated from the water buffalo (Bubalus bubalis) after digestion with two restriction endonucleases, BamHI and StuI. These satellite DNAs of the water buffalo were classified into two types by sequence analysis: one had an approximately 1,400 bp tandem repeat unit with 79% similarity to the bovine satellite I DNA; the other had an approximately 700 bp tandem repeat unit with 81% similarity to the bovine satellite II DNA. The chromosomal distribution of the satellite DNAs were examined in the river-type and the swamp-type buffaloes with direct R-banding fluorescence in situ hybridization. Both the buffalo satellite DNAs were localized to the centromeric regions of all chromosomes in the two types of buffaloes. The hybridization signals with the buffalo satellite I DNA on the acrocentric autosomes and X chromosome were much stronger than that on the biarmed autosomes and Y chromosome, which corresponded to the distribution of C-band-positive centromeric heterochromatin. This centromere-specific satellite DNA also existed in the interstitial region of the long arm of chromosome 1 of the swamp-type buffalo, which was the junction of the telomere-centromere tandem fusion that divided the karyotype in the two types of buffaloes. The intensity of the hybridization signals with buffalo satellite II DNA was almost the same over all the chromosomes, including the Y chromosome, and no additional hybridization signal was found in noncentromeric sites.     

Cervid satellite DNA and karyotypic evolution of Indian muntjac

Five satellite DNA families (designated as satellite I?CV) have been identified in the Cervidae so far. Among those, satellite I, II and IV are centromere specific. Satellite I and II are shared by large number of deer species, where satellite IV is highly conserved among several deer species examined. Satellite III was initially thought to be roe deer specific but later identified in Chinese water deer as well. SatelliteV is Y-chromosome specific for several Asian deer species examined but also found in the pericentric region of Indian muntjac chromosome 3 and in X chromosome of Chinese water deer. The observation of interstitial hybridization sites on Indian muntjac chromosomes with satellite DNA I probe generated from Chinese muntjac provides the first molecular evidence supporting the tandem fusion theory that 2n=6??/7??of Indian muntjac karyotype could derive from an ancestral Chinese muntjac-like species with 2n=46. Interspecies chromosome painting study and the maximum number of interstitial hybridization detected with satellite I and satellite II DNA probes lend support to the hypothesis that the Indian muntjac karyotype could evolve directly from an ancestral Chinese water deer-like species with 2n=70. Such hypothesis is further substantiated by the finding of satellite V signals presented in specific chromosome regions between the Chinese water deer and the Indian muntjac chromosomes.     

Molecular organization and haplotype analysis of centromeric DNA from human chromosome 17: Implications for linkage in neurofibromatosis

The alpha satellite DNA subset located at the centromere of human chromosome 17 has been shown to be tightly linked genetically to the gene for von Recklinghausen neurofibromatosis (NF1). The centromeric DNA polymorphisms used for linkage analyses in NF1 are complex and involve a "locus" (D17Z1) that spans over one million base pairs of satellite DNA. To understand more completely the basis for these polymorphisms and how they might be best scored and used in the analysis of NF1, we have examined the molecular composition of the alpha satellite array on individual copies of chromosome 17 by two complementary approaches. First, we have analyzed segregation of chromosome 17 alpha satellite haplotypes in large, three-generation families that provide information on the different types of alpha satellite segregating in a block fashion. Second, we have analyzed directly the extent of variation in different D17Z1 arrays by genomic blotting analysis of haploid copies of chromosome 17 isolated in rodent/human somatic cell hybrids. The data indicate the existence of a wide range of different alpha satellite variants on individual copies of chromosome 17, each haplotype differing in the size, restriction map, and relative proportion of particular polymorphic repeat forms. Despite this complexity, the D17Z1 markers provide a potentially useful and genetically close starting point for the molecular and clinical analysis of NF1.     

赤链华游蛇的染色体组型与银带带型研究

该文研究赤链华游蛇的染色体组型与NOR     

Fine Mapping of Satellite DNA Sequences along the Y Chromosome of Drosophila Melanogaster: Relationships between Satellite Sequences and Fertility Factors

The entirely heterochromatic Y chromosome of Drosophila melanogaster contains a series of simple sequence satellite DNAs which together account for about 80% of its length. Molecular cloning of the three simple sequence satellite DNAs of D. melanogaster (1.672, 1.686 and 1.705 g/ml) revealed that each satellite comprises several distinct repeat sequences. Together 11 related sequences were identified and 9 of them were shown to be located on the Y chromosome. In the present study we have finely mapped 8 of these sequences along the Y by in situ hybridization on mitotic chromosome preparations. The hybridization experiments were performed on a series of cytologically determined rearrangements involving the Y chromosome. The breakpoints of these rearrangements provided an array of landmarks along the Y which have been used to localize each sequence on the various heterochromatic blocks defined by Hoechst and N-banding techniques. The results of this analysis indicate a good correlation between the N-banded regions and 1.705 repeats and between the Hoechst-bright regions and the 1.672 repeats. However, the molecular basis for banding does not appear to depend exclusively on DNA content, since heterochromatic blocks showing identical banding patterns often contain different combinations of satellite repeats. The distribution of satellite repeats has also been analyzed with respect to the male fertility factors of the Y chromosome. Both loop-forming (kl-5, kl-3 and ks-1) and non-loop-forming (kl-2 and ks-2) fertility genes contain substantial amounts of satellite DNAs. Moreover, each fertility region is characterized by a specific combination of satellite sequences rather than by an homogeneous array of a single type of repeat.(ABSTRACT TRUNCATED AT 250 WORDS)     

The distribution of repetitive DNAs between regular and supernumerary chromosomes in Species of Glossina (Tsetse): a two-step process in the origin of supernumeraries

Several species of tsetse fly within the Morsitans and Fusca subgenera of Glossina contain supernumerary (B) chromosomes. Previous studies on the meiotic behaviour of chromosomes (Southern and Pell, 1973) and the C-band patterns (Jordan et al., 1977) have indicated a close similarity between the Y chromosome and the supernumeraries. The distributions of the highly abundant families of DNA (satellite DNAs) between the autosomes, sex chromosomes and B chromosomes of G.m. morsitans, G. austeni and G. pallidipes have been examined by in situ hybridisation. In addition, the organisation and sequence homologies of satellite DNAs have been examined by restriction enzymes and heterologous hybridisations in in situ and Southern transfer conditions. The majority of satellite sequences that are homologous between species are distributed in several different arrangements between A and B chromosome telomeres with minority sequences at some centromeric and intercalary locations. There is no extensive satellite DNA similarity between the Y and B chromosomes. We suggest that the Y and B chromosome associations and synchronous allocycly during meiosis are the result of extensive heterochromatinisation of these two chromosome types, that is probably a reflection of two separate stages involved in the generation of the B chromosomes in the genus. The independent evolution of satellites and supernumeraries is discussed.     

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.     

A human chromosome 9-specific alphoid DNA repeat spatially resolvable from satellite 3 DNA by fluorescent in situ hybridization.

We have isolated a DNA clone (pMR9A) that identifies an alphoid DNA subset specific for chromosome 9. This alphoid subset is characterized by a dimeric organization as revealed by Southern blot analysis after digestion with HaeIII, HinfI, or StuI. Nonradioactive in situ hybridization demonstrated that pMR9A hybridizes only to the centromeric region of chromosome 9 and reveals chromosome 9 aneuploidies in interphase nuclei. In addition, the probe detects quantitative differences in alpha satellite DNA on chromosome 9, but these quantitative differences are not correlated with the size of the heterochromatic region. Double-labeling experiments, using a chromosome 9-specific satellite 3 clone and pMR9A, enabled us spatially to distinguish the alphoid and satellite 3 domains on metaphase chromosomes after treatment of the cultures with 5-azacytidine.     

Satellite association frequency and number of nucleoli depend on cell cycle duration and NOR-activity

Summary In human lymphocyte cultures the frequencies of satellite associations in first, second, and third mitoses were investigated using the BUDR-method. A marked decrease of the association frequency with increasing numbers of cell cycles was found. The number of nucleoli seen in interphase is correlated with the satellite association frequency in the respective metaphase. Satellite association is positively correlated to Ag-staining intensity of the NORs. Individual differences in satellite associations are due to differences in NOR activity and in lymphocyte activation. BUDR diminishes somewhat the Ag-staining intensity of the NORs but has no effect on satellite association frequencies. The main reason for the decrease of satellite association frequency in second and third lymphocyte mitoses is presumably a certain dislocation of the original chromosome position during mitosis and a decreased possibility of association during the short interphases. The high association frequency in first mitosis resembles the chromosome position in the long interphase of G₀-lymphocytes.     

The behavior of nucleolus organizers in structurally changed karyotypes of barley

Two standard karyotype barley lines and 18 lines with karyotypes reconstructed by means of induced reciprocal translocations have been studied with respect to nucleolus formation. The standard karyotype contains two pairs of satellite chromosomes (pairs 6 and 7). Five of the structurally changed karyotypes contain, as a result of reciprocal translocations between the standard satellite chromosomes, only one satellite chromosome pair, each chromosome with two satellites and two nucleolus organizing regions. Under these circumstances, only two of the four NORs are active in nucleolus formation while the other two — probably the transposed ones — remain inactive; hence the maximum number of primary nucleoli per nucleus is two. — When NORs are translocated to chromosomes with no NOR in the standard karyotyp, the normal pattern of nucleolus formation remains unchanged. The same is true after transposition of segments from other chromosomes to the satellites of the standard SAT-chromosome pairs 6 and 7. The results obtained are discussed with respect to effects of translocations on the activity and behaviour of nucleolus organizing regions.     

DNA composition in South American camelids I. Characterization and in situ hybridization of satellite DNA fractions

The DNA composition and the in situ hybridization of satellite fractions were analysed in the New World camelids llama, alpaca, guanaco and vicuña. In the four camelid forms, it was possible to identify a similar main band DNA and five satellite fractions (I–V) with G+C base contents ranging from 32% to 66%. Satellites II–V from llama were in situ reannealed on chromosomes from the four camelid forms. The results obtained were: (a) the four satellites hybridized with regions of C-banding (centromeric regions of all chromosomes and short arms of some autosomes); (b) in general, homologous hybridizations (llama DNA versus llama chromosomes) were more efficient than heterologous reassociations; there were however three exceptions to this rule (vicuña and alpaca satellite fraction II, chromosome group B; vicuña fraction V, chromosome groups A and B); (c) X chromosomes from the four camelids had satellites III–V but lacked satellite II, (d) no satellite fraction was detected on chromosome Y. The analysis of the in situ hybridization patterns allowed to conclude that most or all C-banded chromosome regions comprise several satellite DNA fractions. It is, moreover, proposed that there is an ample interspecies variation in the number of chromosomes that cross-react with a given satellite fraction. Our data give further support to the close genomic kinship of New World camelids.     

Alpha satellite DNAs on chromosomes 10 and 12 are both members of the dimeric suprachromosomal subfamily, but display little identity at the nucleotide sequence level

We have investigated the organization and complexity of alpha satellite DNA on chromosomes 10 and 12 by restriction endonuclease mapping, in situ hybridization (ISH), and DNA-sequencing methods. Alpha satellite DNA on both chromosomes displays a basic dimeric organization, revealed as a 6- and an 8-mer higher-order repeat (HOR) unit on chromosome 10 and as an 8-mer HOR on chromosome 12. While these HORs show complete chromosome specificity under high-stringency ISH conditions, they recognize an identical set of chromosomes under lower stringencies. At the nucleotide sequence level, both chromosome 10 HORs are 50% identical to the HOR on chromosome 12 and to all other alpha satellite DNA sequences from the in situ cross-hybridizing chromosomes, with the exception of chromosome 6. An 80% identity between chromosome 6- and chromosome 10-derived alphoid sequences was observed. These data suggest that the alphoid DNA on chromosomes 6 and 10 may represent a distinct subclass of the dimeric subfamily. These sequences are proposed to be present, along with the more typical dimeric alpha satellite sequences, on a number of different human chromosomes.     

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.     

Organization of the 378 bp satellite repeat in terminal heterochromatin of Allium fistulosum

Telomeres, DNA-protein structures, are important elements of the eukaryotic chromosome. Telomeric regions of the majority of higher plants contain heptanucleotides TTTAGGG arranged into a tandem repeat. However, some taxa have no such repeats. These are some species of lilies (Lilium) and onions (Allium). For example, terminal regions of chromosomes of Spanish onion (Allium fistulosum) contain satellite DNA whose unit repeats are 380 bp in length, and the short arm of its chromosome 8 contains rDNA repeats. This study deals with the terminal heterochromatin and organization of the satellite repeat in A. fistulosum. Fluorescent in situ hybridization (FISH) was used to locate the satellite DNA on chromosomes and on extended DNA of A. fistulosum. Nonsatellite DNA was found in the structure of telomeric repeat. Polymerase chain reaction (PCR) and Southern hybridization were used for analysis of terminal heterochromatin. Various rearrangements were found in the satellite repeat. The roles of retrotransposones and microsatellites in the formation of terminal heterochromatin are discussed.