Characterization of chromosome structures of Falconinae (Falconidae, Falconiformes, Aves) by chromosome painting and delineation of chromosome rearrangements during their differentiation

Karyotypes of most bird species are characterized by around 2n = 80 chromosomes, comprising 7–10 pairs of large- and medium-sized macrochromosomes including sex chromosomes and numerous morphologically indistinguishable microchromosomes. The Falconinae of the Falconiformes has a different karyotype from the typical avian karyotype in low chromosome numbers, little size difference between macrochromosomes and a smaller number of microchromosomes. To characterize chromosome structures of Falconinae and to delineate the chromosome rearrangements that occurred in this subfamily, we conducted comparative chromosome painting with chicken chromosomes 1–9 and Z probes and microchromosome-specific probes, and chromosome mapping of the 18S–28S rRNA genes and telomeric (TTAGGG) _n sequences for common kestrel (Falco tinnunculus) (2n = 52), peregrine falcon (Falco peregrinus) (2n = 50) and merlin (Falco columbarius) (2n = 40). F. tinnunculus had the highest number of chromosomes and was considered to retain the ancestral karyotype of Falconinae; one and six centric fusions might have occurred in macrochromosomes of F. peregrinus and F. columbarius, respectively. Tandem fusions of microchromosomes to macrochromosomes and between microchromosomes were also frequently observed, and chromosomal locations of the rRNA genes ranged from two to seven pairs of chromosomes. These karyotypic features of Falconinae were relatively different from those of Accipitridae, indicating that the drastic chromosome rearrangements occurred independently in the lineages of Accipitridae and Falconinae.     

Direct detection of repetitive,whole chromosome paint and telomere DNA probes by immunogold electron microscopy

Biotinylated repetitive, whole chromosome paint and telomere DNA probes were investigated at the electron microscope level after non-isotopicin situ hybridization and direct immunogold detection. The protocol described allowed the visualization of a biotinylated chromosome 1 specific satellite DNA probe in the light microscope without silver intensification. This sensitive method was exploited to analyse factors contributing to signal strength in immunogold chromosome painting. Furthermore, it allowed us to investigate the distribution of (TTAGGG)n telomere repeats in human metaphase chromosomes and interphase nuclei. Telomeric and internal (TTAGGG)n repeats were detected at high spatial resolution in human metaphase chromosomes. In the periphery of lymphocyte interphase nuclei, two types of telomere hybridization signals were observed. They differed remarkably in compactness, indicating two types of chromatin conformation present at the interphase telomeresin situ.     

Karyotype evolution in monitor lizards: cross-species chromosome mapping of cDNA reveals highly conserved synteny and gene order in the Toxicofera clade

The water monitor lizard (Varanus salvator macromaculatus (VSA), Platynota) has a chromosome number of 2n?=?40: its karyotype consists of 16 macrochromosomes and 24 microchromosomes. To delineate the process of karyotype evolution in V. salvator macromaculatus, we constructed a cytogenetic map with 86 functional genes and compared it with those of the butterfly lizard (Leiolepis reevesii rubritaeniata (LRE); 2n?=?36) and Japanese four-striped rat snake (Elaphe quadrivirgata (EQU); 2n?=?36), members of the Toxicofera clade. The syntenies and gene orders of macrochromosomes were highly conserved between these species except for several chromosomal rearrangements: eight pairs of VSA macrochromosomes and/or chromosome arms exhibited homology with six pairs of LRE macrochromosomes and eight pairs of EQU macrochromosomes. Furthermore, the genes mapped to microchromosomes of three species were all located on chicken microchromosomes or chromosome 4p. No reciprocal translocations were found in the species, and their karyotypic differences were caused by: low frequencies of interchromosomal rearrangements, such as tandem fusions, or centric fissions/fusions between macrochromosomes and between macro- and microchromosomes; and intrachromosomal rearrangements, such as paracentric inversions or centromere repositioning. The chromosomal rearrangements that occurred in macrochromosomes of the Varanus lineage were also identified through comparative cytogenetic mapping of V. salvator macromaculatus and V. exanthematicus. Morphologic differences in chromosomes 6–8 between the two species could have resulted from pericentric inversion or centromere repositioning.     

A new chromosome instability disorder

Chromosome analysis in a 31-year-old woman referred for primary amenorrhea, revealed a very high incidence of chromosome aberrations. She had microcephaly and immunodeficiency. Her healthy parents were consanguineous (1/32) and a younger sister, also with primary amenorrhea, died when 20 years old with a malignant lymphoma. Chromosome studies were performed on lymphocytes and fibroblasts and in both tissues a high proportion of metaphases with multiple chromosome aberrations was found. Clonal and sporadic rearrangements, consisting of balanced and unbalanced translocations and dicentric chromosomes were more numerous than chromatid and chromosome breaks. In the lymphocytes the same unbalanced translocation t(8q;21q) was present in about 59% of the metaphases. Rearrangements involving chromosomes 7 and 14, similar to those described in patients with ataxia-telangiectasia were found, but with a lower frequency. Sister Chromatid Exchanges were not increased. Chromosome and chromatid abnormalities were enhanced after exposure of cells to mitomycin C but not after exposure to the radiomimetic drug bleomycin. Clinical and cytogenetic characteristics of the patient are compared with those of syndromes (Ataxia-Telangiectasia and Werner's syndrome) or isolated cases (Weemaes et al. 1981, Sperling 1983, Spinner et al. 1985) whose features are similar to those of our patient. This case might represent a new chromosome instability syndrome due to a recessive mutation.     

伴复杂核型异常的髓系恶性血液病中17号染色体异常分析

目的 研究伴复杂核型异常(complex chromosomal abnormalities,CCAs)的髓系恶性血液病中17号染色体的异常特征.方法 经R显带常规细胞遗传学分析显示CCAs的73例髓系恶性血液病,包括21例急性髓系白血病(acute myeloid leukemia,AML)、36例慢性髓系白血病(chronic myeloid leukemia,CML)、16例骨髓增生异常综合征(myelodysplastic syndrome,MDS),并进一步多重荧光原位杂交分析.结果 73例伴CCAs的髓系恶性血液病中,17号染色体异常最常见,占46.5%(34/73),其中AML12例,CML13例,MDS9例,9例CML慢性期患者均未见17号染色体异常.结构异常较多见,总发生率为43.8%(32/73);AML、CML、MDS3组发生率分别为52.4%(11/21)、33.3%(12/36)、56.3%(9/16);所有病例中发生数目异常共15.1%(11/73),三组发生率分别为25.0%(3/12)、38.5%(5/13)、33.3%(3/9),11例数目异常均为-17.有9例同时出现数目异常和结构异常.伴有17号染色体的结构异常中,以非平衡易位多见,3组分别为16、15、8个;平衡易位2个,分别为发生于AML中的t(15;17)及发生于CML中的t(15;17;22).17号染色体结构易位的对手染色体多变,包括了除5号、6号和22号外的所有染色体.结构易位频率最高的对手染色体是15号,占8.2%(6/73);其次为2号,占5.4%(4/73).6例存在17号与15号易位的病例中5例为急性早幼粒白血病,1例为CML急变期.结论 伴CCAs的髓系恶性血液病中17号染色体异常发生率高,以结构异常为主.所有的数目异常均为-17;结构异常以非平衡易位多见.     

Identification of chromosome rearrangements between the laboratory mouse (Mus musculus) and the Indian spiny mouse (Mus platythrix) by comparative FISH analysis

Comparative chromosome painting was applied to the Indian spiny mouse (Mus platythrix) with mouse (M. musculus) chromosome-specific probes for understanding the process of chromosome rearrangements between the two species. The chromosome locations of the 5S and 18S-28S ribosomal RNA genes and the order of the 119 and Tcp-1 genes in the In(17)2 region of the t-complex were also compared. All the painting probes were successfully hybridized to the Indian spiny mouse chromosomes, and a total of 27 segments homologous to mouse chromosomes were identified. The comparative FISH analysis revealed that tandem fusions were major events in the chromosome evolution of the Indian spiny mouse. In addition, other types of chromosome rearrangements, i.e. reciprocal translocations and insertions, were also included.     

Molecular cytogenetic detection of paternal chromosome fragments in allogynogenetic gibel carp, Carassius auratus gibelio Bloch

In gynogenesis, sperm from related species activates egg and embryonic development, but normally does not contribute genetically to the offspring. In gibel carp, Carassius auratus gibelio Bloch, however, gynogenetic offspring often show some phenotypes apparently derived from the heterologous sperm donor. This paternal effect of allogynogenesis is outstanding in an artificial clone F produced by cold treatment of clone E eggs after insemination with blunt-nose black bream (Megaloabrama amblycephala Yin) sperm. Karyotype analysis revealed 5–15 supernumerary microchromosomes in different individuals of clone F in addition to 156 normal chromosomes inherited from the maternal clone E. A painting probe was prepared from the microdissected microchromosomes, and used to investigate the origin of these microchromosomes. Strong positive signals were detected on each microchromosomes of clone F and on 4 pairs of chromosomes in blunt-nose black bream, whereas no signals were detected on the chromosomes of clone E. This result indicates that some paternal chromosome fragments of blunt-nose black bream have been incorporated into the artificial clone F. Therefore, the manipulation of allogynogenesis may provide a unique method to transfer DNA between diverse species for fish breeding. This revised version was published online in July 2006 with corrections to the Cover Date.     

Karyology of the Antarctic chiton Nuttallochiton mirandus (Thiele, 1906) (Mollusca: Polyplacophora) with some considerations on chromosome evolution in chitons

We describe the karyotype, location of nucleolus-organizing regions (NORs) and heterochromatin distribution and composition in the Antarctic chiton Nuttallochiton mirandus. Specimens had a karyotype of 2n = 32 chromosomes, of which two were microchromosomes. Among macrochromosomes, the elements of the first and fourth pair were bi-armed, the others were telocentric. At least six NOR sites were detected with NOR-FISH, but only four were Ag-NOR-banding-positive. The two microchromosomes were essentially euchromatic, while all macrochromosomes exhibited clear pericentromeric C bands that were found to be AT-rich (being quinacrine- and DAPI-positive) and resistant to digestion with AluI and HaeIII. N. mirandus has the largest number of chromosomes (2n = 32) and telocentric elements (26) of all the chiton species studied to date. The karyological results of our study agree with previous molecular data indicating N. mirandus as a sister taxon of Acanthochitona crinita. The karyotypes of the two species could be related as a result of Robertsonian rearrangements. According to the more parsimonious hypothesis, the former would be the primitive karyotype, although other evolutionary events cannot be ruled out.     

The molecular basis of chromosome orthologies and sex chromosomal differentiation in palaeognathous birds

Palaeognathous birds (Struthioniformes and Tinamiformes) have morphologically conserved karyotypes and less differentiated ZW sex chromosomes. To delineate interspecific chromosome orthologies in palaeognathous birds we conducted comparative chromosome painting with chicken (Gallus gallus, GGA) chromosome 1–9 and Z chromosome paints (GGA1–9 and GGAZ) for emu, double-wattled cassowary, ostrich, greater rhea, lesser rhea and elegant crested tinamou. All six species showed the same painting patterns: each probe was hybridized to a single pair of chromosomes with the exception that the GGA4 was hybridized to the fourth largest chromosome and a single pair of microchromosomes. The GGAZ was also hybridized to the entire region of the W chromosome, indicating that extensive homology remains between the Z and W chromosomes on the molecular level. Comparative FISH mapping of four Z- and/or W-linked markers, the ACO1/IREBP, ZOV3 and CHD1 genes and the EE0.6 sequence, revealed the presence of a small deletion in the proximal region of the long arm of the W chromosome in greater rhea and lesser rhea. These results suggest that the karyotypes and sex chromosomes of palaeognathous birds are highly conserved not only morphologically, but also at the molecular level; moreover, palaeognathous birds appear to retain the ancestral lineage of avian karyotypes.     

Comparative chromosome mapping of sex-linked genes and identification of sex chromosomal rearrangements in the Japanese wrinkled frog (Rana rugosa, Ranidae) with ZW and XY sex chromosome systems

There are regional variations of sex chromosome morphologies in the Japanese wrinkled frog, Rana rugosa (2n = 26): heterogametic ZZ/ZW-type and XX/XY-type sex chromosomes, and two different types of homomorphic sex chromosomes. To search for homology between the ZW and XY sex chromosomes and the chromosome rearrangements that have occurred during sex chromosomal differentiation in R. rugosa, we performed chromosome mapping of sexual differentiation genes for R. rugosa by FISH. Three genes, AR, SF-1/Ad4BP and Sox3, were localized to both the ZW and XY chromosomes, and their locations were all different between the Z and W and between the X and Y. AR and SF-1/Ad4BP were located on the short arms of the W and X and the long arms of Z and Y, and Sox3 was mapped to the different locations on the long arms between the Z and W and between the X and Y, probably as a result of multiple rearrangements that occurred during the process of sex chromosome differentiation. However, the chromosomal locations of three genes were almost consistent between the Z and Y and between the W and X, indicating that the Z and Y chromosomes and the W and X chromosomes were respectively derived from the same origins. Dmrt1, which is located on avian sex chromosomes, was localized to autosomes in R. rugosa with both the ZW and XY sex chromosomes, suggesting that Dmrt1 might not be related to sex determination in this species.     

An XX/XY heteromorphic sex chromosome system in the Australian chelid turtle <Emphasis Type="Italic">Emydura macquarii</Emphasis>: A new piece in the puzzle of sex chromosome evolution in turtles

Chromosomal sex determination is the prevalent system found in animals but is rare among turtles. In fact, heteromorphic sex chromosomes are known in only seven of the turtles possessing genotypic sex determination (GSD), two of which correspond to cryptic sex microchromosomes detectable only with high-resolution cytogenetic techniques. Sex chromosomes were undetected in previous studies of Emydura macquarii, a GSD side-necked turtle. Using comparative genomic hybridization (CGH) and GTG-banding, a heteromorphic XX/XY sex chromosome system was detected in E. macquarii. The Y chromosome appears submetacentric and somewhat larger than the metacentric X, the first such report for turtles. CGH revealed a male-specific chromosomal region, which appeared heteromorphic using GTG-banding, and was restricted to the telomeric region of the p arm. Based on our observations and the current phylogeny of chelid turtles, we hypothesize that the sex chromosomes of E. macquarii might be the result of a translocation of an ancestral Y microchromosome as found in a turtle belonging to a sister clade, Chelodina longicollis, onto the tip of an autosome. However, in the absence of data from an outgroup, the opposite (fission of a large XY into an autosome and a micro-XY) is theoretically equally likely. Alternatively, the sex chromosome systems of E. macquarii and C. longicollis may have evolved independently. We discuss the potential causes and consequences of such putative chromosome rearrangements in the evolution of sex chromosomes and sex-determining systems of turtles in general.     

The gametocidal chromosome as a tool for chromosome manipulation in wheat

Many alien chromosomes have been introduced into common wheat (the genus Triticum) from related wild species (the genus Aegilops). Some alien chromosomes have unique genes that secure their existence in the host by causing chromosome breakage in the gametes lacking them. Such chromosomes or genes, called gametocidal (Gc) chromosomes or Gc genes, are derived from different genomes (C, S, S^l and M^g) and belong to three different homoeologous groups 2, 3 and 4. The Gc genes of the C and M^g genomes induce mild, or semi-lethal, chromosome mutations in euploid and alien addition lines of common wheat. Thus, induced chromosomal rearrangements have been identified and established in wheat stocks carrying deletions of wheat and alien (rye and barley) chromosomes or wheat–alien translocations. The gametocidal chromosomes isolated in wheat to date are reviewed here, focusing on their feature as a tool for chromosome manipulation.     

Cohesion proteins are present in centromere protein bodies associated with avian lampbrush chromosomes

Proteins of sister chromatid cohesion are important for maintenance of meiotic chromosome structure and retention of homologous chromosomes in bivalents during diplotene. Localization of the cohesion proteins within nuclei of growing oocytes merits special attention, particularly in avian oocytes, in which diplotene chromosomes assume the form of lampbrush chromosomes (LBCs). We performed indirect immunostaining using antibodies against cohesins SMC1α, SMC1β, SMC3, Rad21, and the SA/STAG family on chaffinch, pigeon and duck LBCs spreads, and frozen ovary sections. On LBCs spreads, antibodies to the majority of cohesins showed punctate staining on chromosome axes. LBC lateral loops, where sister chromatids are separated, did not show cohesin components. The spherical entities attached to the LBCs centromeres in avian germinal vesicles, the so-called protein bodies (PBs), were enriched in SMC1α, SMC3, Rad21, STAG1 and STAG2. The synaptonemal complex component SYCP3, which also participates in cohesion, was detected in the axes of avian lampbrush bivalents and, to a greater degree, in the PBs. In vitellogenic oocytes, cohesion proteins persist in the PBs associated with condensing bivalents when they concentrate into the karyosphere. These results indicate that cohesion proteins accumulate in centromere PBs in avian oocytes and are involved into structural maintenance of lampbrush chromosome axes.     

Multidirectional chromosome painting between the Hirola antelope (Damaliscus hunteri, Alcelaphini, Bovidae), sheep and human

Chromosome specific painting probes of human, sheep and the Hirola antelope ( Damaliscus hunteri ) derived by flow sorting of chromosomes were used in multi directional chromosome painting experiments to better define the karyological relationship within Bovidae species (specifically, Caprini and Alcelaphini tribes) and humans. Although not all chromosomes of Damaliscus hunteri could be resolved into single peaks by flow-sorting we managed to present a complete homology map for chromosomes between the three species. When comparing the karyotype of Damaliscus hunteri with human all of the main known motives in mammalian chromosome evolution are present (i.e. associations of human homologous chromosomes 3-21, 4-8, 7-16, 14-15, 16-19 and two forms of 12-22) which were also confirmed with the sheep paint probes. Further, we observed those patterns that have been described as common derived traits for artiodactyls (i.e. associations of human homologous chromosomes 5/19 and a complex alternating pattern of hybridizations with human chromosome 14 and 15 probes). As known from classical karyotyping some of the Damaliscus chromosomes are biarmed and were supposedly involved in Robertsonian translocations frequently found in karyotype evolution of bovids. We refined these rearrangements with the molecular probes and also delineated a chromosome painting pattern that should be the result of a paracentric inversion in the Damaliscus hunteri karyotype. This study demonstrates that multidirectional chromosome painting will be a valuable tool for the investigation of the dynamics of chromosome evolution in exotic bovid species.     

Reassessment of a chromosome 12q + marker by fluorescent in situ hybridization (FISH)

We present a case previously described by Jenkins et al. (1983) as atypical Down syndrome (DS). The initial diagnosis was first made on the basis of phenotypic and cytogenetic data. This analysis was supported by studies of superoxide dismutase (SOD1) activity that maps to band 21q22.1. Results from phenotypic, chromosome banding and SODI studies suggested a karyotype of 46,XX,—12, + t(12pter to 12qter::21q21 to 21q22.?2). Using fluorescent in situ hybridization (FISH) for chromosome painting with DNA libraries derived from sorted human chromosomes to stain selectively the chromosomes No. 21 and No. 12, we demonstrate that the marker chromosome 12q+ has no chromosome 21 content but it is derived from chromosome 12.     

Chromosomal polymorphism and comparative painting analysis in the zebra finch

The zebra finch (Taeniopygia guttata) is often studied because of its interesting behaviour and neurobiology. Genetic information on this species has been lacking, making analysis of informative mutants difficult. Here we report on an improved cytological method for preparation of metaphase chromosomes suitable for fluorescent in situ hybridization of adult birds. We found that individual chicken chromosome paints usually hybridized to single zebra finch chromosomes, indicating only minor chromosomal rearrangements since the evolutionary divergence of these two species, and suggesting that the genomic location of chicken genes will predict the location of zebra finch orthologues. Chicken chromosome 1 appears to have split into two macrochromosomes in zebra finches, and chicken chromosome 4 paint hybridizes to a zebra finch macrochromosome and a microchromosome. This pattern was confirmed by mapping the androgen receptor (AR), which is located on chicken chromosome 4 but on a zebra finch microchromosome. We detected a telocentric/submetacentric polymorphism of chromosome 6 in our colony of zebra finches, and found that the polymorphism was inherited in a Mendelian pattern     

Reciprocal chromosome painting between a New World primate, the woolly monkey, and humans

We employed fluorescence-activated chromosome sorting (FACS) to construct chromosome paint sets for the woolly monkey (Lagothrix lagotricha) and then FISH to reciprocally paint human and woolly monkey metaphases. Reciprocal chromosome painting between humans and the woolly monkey allowed us to assign subchromosomal homologies between these species. The reciprocal painting data between humans and the woolly monkey also allow a better interpretation of the chromosomal difference between humans and platyrrhines, and refine hypotheses about the genomic rearrangements that gave origin to the genome of New World monkeys. Paints of woolly monkey chromosomes were used to paint human metaphases and forty-five clear signals were detected. Paints specific to each human chromosome were used to paint woolly monkey metaphases. The 23 human paints gave 39 clear signals on the woolly monkey karyotype. The woolly monkey chromosomes painted by human paints produced 7 associations of segments homologous to human chromosomes or human chromosome segments: 2/16, 3/21, 4sol;15, 5/7, 8/18, 10/16 and 14/15. A derived translocation between segments homologous to human chromosomes 4 and 15 is a synapomorphic marker linking all Atelines. These species may also be linked by fragmentation of homologs to human 1, 4, and 15.     

Interstitial telomeric sites and NORs in Hartmann's zebra (Equus zebra hartmannae) chromosomes

Interstitial telomeric sites (ITSs) are considered as signatures of chromosomal rearrangements that take place during karyotype evolution. Understanding that equids have undergone rapid karyotype evolution compared with the average in other mammals, a search of these signatures was carried out in the Hartmann's mountain zebra (Equus zebra hartmannae; EZH) chromosomes. Six consistent ITSs were identified on five of the zebra chromosomes (EZH1p, 1q, 2q, 5q, 6q and 11q). The location of these ITSs coincided with fusion points of some of the evolutionarily conserved human-Hartmann's zebra chromosomal segments suggesting that the sequences are remnants of fusion events between ancestral chromosomes. Incidentally, three of the ITSs also matched with the presence of constitutive heterochromatin. Further, ribosomal gene clusters were localized on five zebra chromosomes and the data were compared with those in other equid species. The findings offer preliminary evidence on the likely evolution of some of the Hartmann's zebra chromosomes and add to the current search for clues that lead to the ancestral chromosomal configuration in equids. This revised version was published online in July 2006 with corrections to the Cover Date.