Sex chromosome homologies between human and fish revealed by the chromosome painting method

Human sex chromosome-specific probes were hybridized to metaphase spreads of three fish species, Monopterus albus Zuiew, Danio rerio and Mastacembelus aculeatus Basilewsky, to reveal evolutionary conservation of sex chromosomal segments between distantly related species of vertebrates. The human X chromosomal paint disclosed 4, 8, and 6 conserved syntenic segments in the karyotypes of the three fish species respectively, which were scattered in several pairs of homologous chromosomes. But no conserved segment was identified in our experiments when the human Y chromosomal probes were used. The evolution of the X chromosome of vertebrates is discussed.     

Molecular differentiation of the homomorphic sex chromosomes inMegaselia scalaris (Diptera) detected by random DNA probes

Randomly cloned DNA fragments and a poly-(GATA) containing sequence were used as probes to identify sex chromosomal inheritance and to detect differences at the molecular level between the homomorphic X and Y in the phorid fly,Megaselia scalaris. Restriction fragment length differences between males and females and between two laboratory stocks of different geographic origin were used to differentiate between sex chromosomal and autosomal origin of the respective fragments. Five random probes detected X and Y chromosomal DNA loci and two others recognized autosomal DNA loci. One random probe and the poly(GATA) probe hybridized with both sex chromosomal and autosomal restriction fragments. Most of the Y chromosomal restriction fragments were conserved in length between the two stocks while most of the X chromosomal and autosomal fragments showed length polymorphism. It was concluded, therefore, that the Y chromosome contains a conserved segment in which crossover is suppressed and restriction site differences have accumulated relative to the X. These chromosomes, therefore, conform to a theoretically expected early stage of sex chromosome evolution.     

Multi-directional chromosome painting maps homologies between species belonging to three genera of New World monkeys and humans

We mapped chromosomal homologies in two species of Chiropotes (Pitheciini, Saki Monkeys) and one species of Aotus (Aotinae, Owl Monkey) by multi-directional chromosome painting. Human chromosome probes were hybridized to Chiropotes utahicki, C. israelita and Aotus nancymae metaphases. Wooly Monkey chromosome paints were also hybridized to Owl Monkey metaphases. We established Owl Monkey chromosome paint probes by flow sorting and reciprocally hybridized them to human chromosomes. The karyotypes of the Bearded Saki Monkeys studied here are close to the hypothesized ancestral platyrrhine karytoype, while that of the Owl Monkey appears to be highly derived. The A. nancymae karyotype is highly shuffled and only three human syntenic groups were found conserved coexisting with 17 derived human homologous associations. A minimum of 14 fissions and 13 fusions would be required to derive the A. nancymae karyotype from that of the ancestral New World primate karyotype. An inversion between homologs to segments of human 10 and 16 suggests a link between Callicebus and Chiropotes, while the syntenic association of 10/11 found in Aotus and Callicebus suggests a link between these two genera. Future molecular cytogenetic work will be needed to determine whether these rearrangements represent synapomorphic chromosomal traits.     

The origin and function of the mammalian Y chromosome and Y-borne genes – an evolving understanding

Mammals have an XX:XY system of chromosomal sex determination in which a small heterochromatic Y controls male development. The Y contains the testis determining factor SRY, as well as several genes important in spermatogenesis. Comparative studies show that the Y was once homologous with the X, but has been progressively degraded, and now consists largely of repeated sequences as well as degraded copies of X linked genes. The small original X and Y have been enlarged by cycles of autosomal addition to one partner, recombination onto the other and continuing attrition of the compound Y. This addition–attrition hypothesis predicts that the pseudoautosomal region of the human X is merely the last relic of the latest addition. Genes (including SRY) on the conserved or added region of the Y evolved functions in male sex determination and differentiation distinct from the general functions of their X-linked partners. Although the gonadogenesis pathway is highly conserved in vertebrates, its control has probably changed radically and rapidly in vertebrate – even mammalian – evolution.     

Identification and chromosomal distribution of DNA sequence segments conserved since divergence of Escherichia coli and Bacillus subtilis

DNA sequence segments conserved since divergence of Escherichia coli and Bacillus subtilis were identified, using the GenBank sequence database. Chromosomal locations of the conserved segments were compared between the two bacteria, and the following three features were observed. (1) Although the two genomes are nearly identical in size, chromosomal arrangements of the conserved segments are considerably different from each other. (2) In many cases, chromosomal locations of a conserved segment in the two species have deviated from each other by a multiple of 60°. (3) There are many instances in which a contiguous segment in one genome is split into two or more segments located at distinct positions in the other genome, and these split segments were found to tend to lie on the E. coli or B. subtilis genome separated by distances of multiples of 60°. On the basis of these observations, genome organizations of the two bacteria were discussed in terms of genome doublings as well as random chromosomal rearrangements.     

Stable karyotypes: a general rule for the fish of the family Prochilodontidae?

Cytogenetic studies involving the family Prochilodontidae have shown that these fish can be characterized by a constant diploid number and a conserved karyotypic macrostructure. This study focused on comparative physical chromosomal mapping using 18S and 5S rDNA to compare the species Semaprochilodus insignis and S. taeniurus. Our results indicated the conservation of large number of conventional chromosomal markers. The molecular cytogenetic analyses of the location of the 18S rDNA indicated the maintenance of a chromosome pair bearing these sites in both species analyzed, and it appears to be a conserved character among the majority of the species of this family. The stability of the number of 5S ribosomal DNA sites and their chromosomal localization as has been reported for the Prochilodontidae was not, however, confirmed for S. insignis and S. taeniurus, as these species showed multiple specific rDNA 5S sites. As such, and in spite of the fact that a number of studies indicate that the family Prochilodontidae has a conserved karyotypic structure, the utilization of molecular tools that use chromosomal segments as markers revealed that this presumed stability cannot be extended to the genome level for the species S. insignis and S. taeniurus.     

Cytogenetic mapping of immunoglobulin heavy chain genes in Antarctic fish

The chromosomal location of the IgH locus has been analyzed in several bony fish of the Antarctic perciform group Notothenioidei. Two IgH probes were prepared from the species Trematomus bernacchii (family Nototheniidae, tribe Trematominae) and mapped onto the chromosomes of ten species belonging to the same genus (Trematomus) and in two outgroups, through one-color and two-color FISH. A single location of the IgH locus was found in the majority of the species examined, including the outgroups, whereas in four of them the IgH genes splited to two chromosomal loci. RT-PCR experiments revealed the presence of three allelic sequences in T. newnesi, a species in which the IgH genes were organized in two chromosomal loci. Possible pathways leading to IgH genes duplication during the diversification of trematomine fishes were inferred from the analysis of the FISH patterns in a phylogenetic context. The present work provides the first comprehensive picture of IgH genes organization at chromosomal level in a bony fish group.     

Whole chromosome painting reveals independent origin of sex chromosomes in closely related forms of a fish species

The wolf fish Hoplias malabaricus includes well differentiated sex systems (XY and X₁X₂Y in karyomorphs B and D, respectively), a nascent XY pair (karyomorph C) and not recognized sex chromosomes (karyomorph A). We performed the evolutionary analysis of these sex chromosomes, using two X chromosome-specific probes derived by microdissection from the XY and X₁X₂Y sex systems. A putative-sex pair in karyomorph A was identified, from which the differentiated XY system was evolved, as well as the clearly evolutionary relationship between the nascent XY system and the origin of the multiple X₁X₂Y chromosomes. The lack of recognizable signals on the sex chromosomes after the reciprocal cross-FISH experiments highlighted that they evolved independently from non-homologous autosomal pairs. It is noteworthy that these distinct pathways occur inside the same nominal species, thus exposing the high plasticity of sex chromosome evolution in lower vertebrates. Possible mechanisms underlying this sex determination liability are also discussed.     

Non-homologous sex chromosomes in two species of the genus Eigenmannia (Teleostei: Gymnotiformes)

The Neotropical genus Eigenmannia is a fish group with unknown species diversity where representatives possess a broad range of chromosomal sex determining systems namely XY/XX, X(1)X(2)Y/X(1)X(1)X(2)X(2), ZZ/ZW as well as homomorphic sex chromosomes. To test the homology of two heteromorphic XY sex chromosome systems present in two sympatric populations, reciprocal cross-species FISH experiments were performed using probes derived by microdissection of X and Y chromosomes present in analyzed specimens of Eigenmannia virescens and Eigenmannia sp.2, respectively. While X and Y paint probes hybridized to species-specific sex chromosomes, in reciprocal cross-FISH both probes hybridized exclusively to autosomes. The result suggests multiple independent origins of the XY systems in the analyzed populations.     

X-linked genetic homologies between mouse and man

X-linked genes are conserved among all mammalian species, but the organization of genes on the X chromosome varies from one species to another. This review summarizes the evidence for established gene homologies between mice and human beings. It also describes genes that are possible homologies because of their locations in the human and murine X chromosomes and similarities in the phenotypes they produce. Based on current knowledge of homologous gene location, the human and murine X chromosomes appear to contain four highly conserved segments and differ in organization by only three to four simple chromosomal rearrangements.     

A Comparative Study of Pygmy Hippopotamus (<Emphasis Type="Italic">Choeropsis liberiensis</Emphasis>) Karyotype by Cross-Species Chromosome Painting

The family Hippopotamidae is comprised of two genera with two living species, the common hippo (Hippopotamus amphibius) and the pygmy hippo (Choeropsis liberiensis). Unlike the common hippo, the karyotype of C. liberiensis has not yet been investigated via cross-species chromosome painting methods. We established chromosomal homologies between the pygmy hippo, pig, and cattle by fluorescence in situ hybridization using whole chromosome, arm-specific, region specific, and bacterial artificial chromosome (BAC) probes. Probes from the 18 pig autosomes painted 45 conserved chromosomal segments in the pygmy hippo genome. The pygmy hippo and cattle homology map was deduced from our hybridization results of painting probes to pygmy hippo chromosomes with a combination of previously published dromedary hybridization data. On the pygmy hippo and cattle homology map, 29 cattle autosomes revealed 39 conservative segments on pygmy hippo chromosomes. For a more detailed structural analysis of genome rearrangements and X chromosome structure, we used cattle region specific and BAC probes. Our report demonstrates that cattle probes are useful not only in comparative studies within Ruminantia, but also in more phylogenetically distant Artiodactyla species.     

人类性染色体特异ＤＮＡ对三种鱼类染色体的描绘

染色体描绘是研究基因组进化的强有力手段之一,用人Ｘ和Ｙ染色体文库特异ＤＮＡ为探针,对３种硬骨鱼类－刺鳅、黄鳝和斑马鱼的有丝分裂中期裂染色体进行了描绘研究,结果表明,在这３种鱼类的染色体组中都发现有人Ｘ染色体特DN的同源片段,它们散布在几对同源染色体中,但用人Ｙ染色体ＤＮ描绘这３种鱼类染色体时,则没有检测出可见的同源片段。同时对从低等脊椎动物到人类的Ｘ染色体进化过程进行了进一步探讨。     

Complete homology maps of the rabbit (Oryctolagus cuniculus) and human by reciprocal chromosome painting

Fluorescence in situ hybridization (FISH) was used to construct a homology map to analyse the extent of evolutionary conservation of chromosome segments between human and rabbit (Oryctolagus cuniculus, 2n = 44). Chromosome-specific probes were established by bivariate fluorescence activated flow sorting followed by degenerate oligonucleotide-primed PCR (DOP-PCR). Painting of rabbit probes to human chromosomes and vice versa allowed a detailed analysis of the homology between these species. All rabbit chromosome paints, except for the Y paint, hybridized to human chromosomes. All human chromosome paints, except for the Y paint, hybridized to rabbit chromosomes. The results obtained revealed extensive genome conservation between the two species. Rabbit chromosomes 12, 19 and X were found to be completely homologous to human chromosomes 6, 17 and X, respectively. All other human chromosomes were homologous to two or sometimes three rabbit chromosomes. Many conserved chromosome segments found previously in other mammals (e.g. cat, pig, cattle, Indian muntjac) were also found to be conserved in rabbit chromosomes.     

The karyotype of Alligator mississippiensis,and chromosomal mapping of the ZFY/X homologue,Zfc

Comparative mapping studies of X-linked genes in mammals have provided insights into the evolution of the X chromosome. Many reptiles including the American alligator, Alligator mississippiensis, do not appear to possess heteromorphic sex chromosomes, and sex is determined by the incubation temperature of the egg during embryonic development. Mapping of homologues of mammalian X-linked genes in reptiles could lead to a greater understanding of the evolution of vertebrate sex chromosomes. One of the genes used in the mammalian mapping studies was ZFX, an X-linked copy of the human ZFY gene which was originally isolated as a candidate for the mammalian testis-determining factor (TDF). ZFX is X-linked in eutherians, but maps to two autosomal locations in marsupials and monotremes, close to other genes associated with the eutherian X. The alligator homologue of the ZFY/ZFX genes, Zfc, has been isolated and described previously. A detailed karyotype of A. mississippiensis is presented, together with chromosomal in situ hybridisation data localising the Zfc gene to chromosome 3. Further chromosomal mapping studies using eutherian X-linked genes may reveal conserved chromosomal regions in the alligator that have become part of the eutherian X chromosome during evolution.     

Comparative Chromosome Painting in Six Species of Oligoryzomys (Rodentia,Sigmodontinae) and the Karyotype Evolution of the Genus

Oligoryzomys belongs to the tribe Oryzomyini, and contains about 22 species. Diploid numbers range from 2n = 44 in Oligoryzomys sp. 2 to 2n = 72 in O. utiaritensis and phylogenetic relationships are not well defined. The high morphological convergence leads to misidentification of taxonomic entities and the species are often identified by chromosomal characters. Until now, the genus has been studied only by classical cytogenetic approaches. To understand the chromosomal evolution of Oligoryzomys, we developed chromosome probes from a female of Oligoryzomys moojeni (OMO) with 2n = 70 and hybridized to other five Oligoryzomys species. The probes painted 31 segments on O. fornesi (OFO) with 2n = 62; 32 segments on O. microtis (OMI), 2n = 64; 33 segments on O. nigripes (ONI), 2n = 62 and on O. rupestris (ORU), 2n = 46; and 34 on Oligoryzomys sp. 2 (OSP), 2n = 44. OMO probes 4 and 5 showed a syntenic association in O. fornesi, O. microtis and O. nigripes and were also presented in the same pair, although disrupted, in O. rupestris and Oligoryzomys sp. 2. Concerning O. rupestris and Oligoryzomys sp. 2, species with the lowest diploid numbers of the genus, a total of 8 probes hybridized to 11 segments on the largest pair of ORU 1 and 9 probes hybridized to 12 segments on OSP 1. Also, OMO 6 painted three segments in ORU, corresponding to the proximal segment of ORU 2q, and the whole of ORU 19 and 20. In OSP, the segment corresponding to ORU 20 was homologous to OSP 1p. OMO X showed signals of hybridization in both X and Y chromosomes. Extensive chromosomal rearrangements, that could not be detected by classical cytogenetic techniques, such as pericentric inversions or repositioning of centromeres, Robertsonian rearrangements and tandem fusions/fissions, as well as gain/activation or loss/inactivation of centromeres and telomeric sequences have driven the huge genome reshuffling in these closely related species.     

Genetic mapping of Y-chromosomal DNA markers in Pacific salmon

Sex chromosomes in fish provide an intriguing view of how sex-determination mechanisms evolve in vertebrates. Many fish species with single-factor sex-determination systems do not have cytogenetically-distinguishable sex chromosomes, suggesting that few sex-specific sequences or chromosomal rearrangements are present and that sex-chromosome evolution is thus at an early stage. We describe experiments examining the linkage arrangement of a Y-chromosomal GH pseudogene (GH-Y) sequence in four species of salmon (chum, Oncorhynchus keta; pink, O. gorbuscha; coho, O. kisutch; chinook, O. tshawytscha). Phylogenetic analysis indicates that GH-Y arose early in Oncorhynchus evolution, after this genus had diverged from Salmo and Salvelinus. However, GH-Y has not been detected in some Oncorhynchus species (O. nerka, O. mykiss and O. clarki), consistent with this locus being deleted in some lineages. GH-Y is tightly linked genetically to the sex-determination locus on the Y chromosome and, in chinook salmon, to another Y-linked DNA marker OtY1. GH-Y is derived from an ancestral GH2 gene, but this latter functional GH locus is autosomal or pseudoautosomal. YY chinook salmon are viable and fertile, indicating the Y chromosome is not deficient of vital genetic functions present on the X chromosome, consistent with sex chromosomes that are in an early stage of divergence.     

点带石斑鱼染色体显带技术及其描绘研究

点带石斑鱼(Epinephelus malabaricus)属于鲈形目, 科、石斑鱼亚科、石斑鱼属, 是中国东南沿海暖水性礁栖的名贵海产经济鱼类. 采用PHA活体注射结合秋水仙素培养, 取点带石斑鱼全肾, 低渗处理, 空气干燥制片法制作染色体标本, 利用Alu I 限制性内切酶介导的原位切口平移显带技术, 在点带石斑鱼有丝分裂中期染色体上诱导出带纹清晰、分散良好的多重带. 结果显示, 多数染色体显现出8-10条带纹, 最少的一对染色体也有4条带纹, 同源染色体带纹基本一致, 在每对染色体上的数目及其分布具明显特征性且相对稳定, 同时发现不同分裂相的同一号染色体上, 特征带纹鲜明一致, 带纹数目基本吻合, 具有可重复性和可操作性; 然后用人X和Y染色体文库特异DNA为探针, 对点带石斑鱼的有丝分裂中期分裂相染色体进行了描绘研究. 结果表明, 点带石斑鱼染色体组中测出了人X染色体特异DNA同源片段的3个保守同线群, 分别在点带石斑鱼的第7、第13和第22号同源染色体上, 它们的杂交信号最近边距着丝粒的百分比距离分别大约为62.3%、43.4%及44.4%; 人X染色质同源片段的大小约占点带石斑鱼基因组的4.63％. 但用人Y染色体DNA描绘点带石斑鱼染色体时, 没有检测出可见的同源片段. 研究结果可以为从低等脊椎动物到人类性染色体的进化过程提供一种新的研究思路.       

In situ hybridization (FISH) maps chromosomal homologies between Alouatta belzebul (Platyrrhini,Cebidae) and other primates and reveals extensive interchromosomal rearrangements between howler monkey genomes

We hybridized whole human chromosome specific probes to metaphases of the black-and-red howler monkey Alouatta belzebul in order to establish chromosomal homology between humans and black-and-red howlers. The results show that the black-and-red howler monkey has a highly rearranged genome and that the human chromosome homologs are often fragmented and translocated. The number of hybridization signals we obtained per haploid set was 40. Nine human chromosome probes gave multiple signals on different howler chromosomes, showing that their synteny is disturbed in A. Belzebul. Fourteen black-and-red howler autosomes were completely hybridized by one human autosomal paint, six had two signals, three had three signals, and one chromosome had four signals. Howler chromosomes with multiple signals have produced 12 chromosomal syntenies or hybridization associations which differ from those found in humans: 1/2, 2/20, 3/21, 4/15, 4/16, 5/7, 5/11, 8/18, 9/12, 10/16, 14/15, and 15/22. The hybridization pattern was then compared with those found in two red howler taxa and other mammals. The comparison shows that even within the genus Alouatta numerous interchromosomal rearrangements differentiate each taxa: A. belzebul has six unique apomorphic associations, A. seniculus sara and A. seniculus arctoidea share seven derived associations, and additionally A. seniculus sara has four apomorphic associations and A. seniculus arctoideaseven apomorphic associations. A. belzebul appears to have a more conserved karyotype than the red howlers. Both red and black-and-red howlers are characterized by Y-autosome translocations; the peculiar chromosomal sex system found in the red howler taxa could be considered a further transformation of the A. belzebul sex system. The finding that apparently morphologically similar or even identical taxa have such extreme genomic differences has important implications for speciation theory and neotropical primate conservation. Am. J. Primatol. 46:119–133, 1998. © 1998 Wiley-Liss, Inc.     

Chromosome Painting Shows That Pygathrix nemaeus Has the Most Basal Karyotype Among Asian Colobinae

We mapped the chromosomal homology of Pygathrix namaeus (douc) with human and other primates by in situ hybridization of human chromosome paints. The synteny of 3 human chromosomes (1, 2, 19) is fragmented in the douc karyotype and the 23 human probes (autosomes plus X) provided 26 signals. There are associations between human chromosomes 14/15, 21/22, and 1/19. Human chromosomes 1 and 19 are divided in two segments and associated on douc chromosomes 8 and 10. The fragmentation and association of human chromosomes 1 and 19 is best explained as the result of a reciprocal translocation, which occurs in all documented Asian colobines studied, but not in the African species Colobus guereza. However, the homologs to douc chromosome 10 in all other Asian documented colobines show an additional pericentric inversion. Our results indicate that Pygathrix nemeus is karyologically the most conservative colobine species yet studied and that this species probably diverged early after the separation of Asian and African Colobinae. The data reinforce the monophyly of the Colobinae and their division into an African and an Asian clade.     

The origin of an unusual sex chromosome constitution in <Emphasis Type="Italic">Acomys</Emphasis> sp. (Rodentia,Muridae) from Tanzania

This paper describes a case which presents an evident variation from the “standard” XX/XY sex chromosomal constitution in a rodent, Acomys sp. This species known to be found in three localities of East Africa has only recently been separated from A. spinosissimus, its closest relative. In our study, five specimens of Acomys sp. and eight specimens of A. spinosissimus were live-trapped in five localities. Comparisons between the two taxa assed by G- banding show a complete homology in the chromosomal shape and banding pattern for 29 pairs of chromosomes corresponding to the complete autosomal set of A. spinosissimus. However, while all the A. spinosissimus analysed have 2n = 60 and a XY-XX system, in Acomys sp. males and females constitute mosaics for sex chromosomes in the bone marrow cells. Females (2n = 59, 60) have an excess (97%) of aneuploid cells with one single giant X chromosome, and males (2n = 60, 61) show X0/XY cells occurring in somatic tissues and XY cells in the germinal lineage. In addition, an odd heterochromatic submetacentric chromosome was identified in all the cells examined in two males and a female of Acomys sp. Since this chromosome was not related to sex determination and it is not present in all the analysed specimens, it can be considered as a B chromosome. Finally, the in situ fluorescence hybridisation (FISH) with telomeric probes showed a very intense interstitial telomeric signal (ITS) at the medial part on the long heterochromatic arm of the X chromosome. This could be due to recent chromosomal rearrangement.