Meiotic Disjunction of Homologs in Saccharomyces Cerevisiae Is Directed by Pairing and Recombination of the Chromosome Arms but Not by Pairing of the Centromeres

We explored the behavior of meiotic chromosomes in Saccharomyces cerevisiae by examining the effects of chromosomal rearrangements on the pattern of disjunction and recombination of chromosome III during meiosis. The segregation of deletion chromosomes lacking part or all (telocentric) of one arm was analyzed in the presence of one or two copies of a normal chromosome III. In strains containing one normal and any one deletion chromosome, the two chromosomes disjoined in most meioses. In strains with one normal chromosome and both a left and right arm telocentric chromosome, the two telocentrics preferentially disjoined from the normal chromosome. Homology on one arm was sufficient to direct chromosome disjunction, and two chromosomes could be directed to disjoin from a third. In strains containing one deletion chromosome and two normal chromosomes, the two normal chromosomes preferentially disjoined, but in 4-7% of the tetrads the normal chromosomes cosegregated, disjoining from the deletion chromosome. Recombination between the two normal chromosomes or between the deletion chromosome and a normal chromosome increased the probability that these chromosomes would disjoin, although cosegregation of recombinants was observed. Finally, we observed that a derivative of chromosome III in which the centromeric region was deleted and CEN5 was integrated at another site on the chromosome disjoined from a normal chromosome III with fidelity. These studies demonstrate that it is not pairing of the centromeres, but pairing and recombination along the arms of the homologs, that directs meiotic chromosome segregation.     

马尾松染色体荧光带型的研究

对马尾松有丝分裂中期染色体荧光带纹的分析结果表明,其色霉素Ａ的染色体的荧光带赤;１对为着丝粒区和臂间我均有带纹的中间着丝染色体。６对为臂间区有带纺的中间着丝粒染色体;２对为着丝粒区有带纹的中间着丝粒染色体;３对无带纹的中间或近中着丝粒染色体;１对为着丝粒区有带纹的近中着丝粒染色体。     

Resolution of Dicentric Chromosomes by Ty-Mediated Recombination in Yeast

We have integrated a plasmid containing a yeast centromere, CEN5, into the HIS4 region of chromosome III by transformation. Of the three transformant colonies examined, none contained a dicentric chromosome, but all contained a rearranged chromosome III. In one transformant, rearrangement occurred by homologous recombination between two Ty elements; one on the left arm and the other on the right arm of chromosome III. This event produced a ring chromosome (ring chromosome III) of about 60 kb consisting of CEN3 and all other sequences between the two Ty elements. In addition, a linear chromosome (chromosome IIIA) consisting of sequences distal to the two Ty elements including CEN5, but lacking 60 kb of sequences from the centromeric region, was produced. Two other transformants also contain a similarly altered linear chromosome III as well as an apparently normal copy of chromosome III. These results suggest that dicentric chromosomes cannot be maintained in yeast and that dicentric structures must be resolved for the cell to survive.--The meiotic segregation properties of ring chromosome III and linear chromosome IIIA were examined in diploid cells which also contained a normal chromosome III. Chromosome IIIA and normal chromosome III disjoined normally, indicating that homology or parallel location of the centromeric regions of these chromosomes are not essential for proper meiotic segregation. In contrast, the 60-kb ring chromosome III, which is homologous to the centromeric region of the normal chromosome III, did not appear to pair with fidelity with chromosome III.     

Distributive Disjunction of Authentic Chromosomes in Saccharomyces Cerevisiae

Distributive disjunction is defined as the first division meiotic segregation of either nonhomologous chromosomes that lack homologs or homologous chromosomes that have not recombined. To determine if chromosomes from the yeast Saccharomyces cerevisiae were capable of distributive disjunction, we constructed a strain that was monosomic for both chromosome I and chromosome III and analyzed the meiotic segregation of the two monosomic chromosomes. In addition, we bisected chromosome I into two functional chromosome fragments, constructed strains that were monosomic for both chromosome fragments and examined meiotic segregation of the chromosome fragments in the monosomic strains. The two nonhomologous chromosomes or chromosome fragments appeared to segregate from each other in approximately 90% of the asci analyzed, indicating that yeast chromosomes were capable of distributive disjunction. We also examined the ability of a small nonhomologous centromere containing plasmid to participate in distributive disjunction with the two nonhomologous monosomic chromosomes. The plasmid appeared to efficiently participate with the two full length chromosomes suggesting that distributive disjunction in yeast is not dependent on chromosome size. Thus, distributive disjunction in S. cerevisiae appears to be different from Drosophila melanogaster where a different sized chromosome is excluded from distributive disjunction when two similar size nonhomologous chromosomes are present.     

M chromosome of the wild silkworm, Bombyx mandarina (n = 27), corresponds to two chromosomes in the domesticated silkworm, Bombyx mori (n = 28).

Chromosomes of Bombyx mori (n = 28) and of Bombyx mandarina (n = 27) were studied cytogenetically to resolve the origin of the large M chromosome in the Japaneses type of B. mandarina. In the F1 progeny from the reciprocal cross between B. mandarina and B. mori, the mitotic chromosome number was 2n = 55, and a chromosome configuration of 26 bivalents plus 1 trivalent was observed at metaphase I of germ cells. The trivalent chromosome consisted of the M chromosome from B. mandarina and two chromosomes from B. mori. When males of B. mori were mated to the F1 females, nuclei with two types of chromosome number (2n = 55 and 2n = 56) and two sets of chromosome pairs (26 bivalents plus 1 trivalent versus 28 bivalents) were observed in the metaphase I stage. Linkage analysis showed that the 14th chromosome of B. mori was involved in these two types of chromosome segregation. This result indicates that the M chromosome in B. mandarina arose from a fusion between a chromosome corresponding to the 14th linkage group and another, yet unidentified linkage group.     

Fishing for a diagnosis

A family with primary infertility and two members with mental retardation and subtle facial dysmorphism is described. In the two retarded persons chromosomal rearrangements (partial monosomy of chromosome 5 and partial trisomy of chromosome 7) were detected. One member of the family had died with major congenital malformations. Her fibroblasts had been stored and her chromosomes showed the inverse pattern (partial trisomy of chromosome 5 and partial monosomy of chromosome 7). It appeared that in familial mental retardation with or without congenital malformations FISH-techniques should be used to detect submicroscopic chromosomal aberrations, which are not detectable by routine chromosome studies.     

The late-replicating X chromosome in digynous mouse triploid embryos

Using BrdU-labeling and acridine orange staining, the behavior of X-chromosome replication was studied in 28 XXX and 19 XXY digynous mouse triploids. In some of these the paternal and maternal X chromosome could by cytologically distinguished. Such embryos were obtained by mating chromosomally normal females with males carrying Cattanach's X chromosome which contains an autosomal insertion that substantially increases the length of this chromosome. In the XXX triploids there were two distinct cell lines, one with two late-replicating X chromosomes, and the other with only one late-replicating X. The XXY triploids were also composed of two cell populations, one with a single late-replicating X and the other with no late replicating X chromosome. Assuming that the late-replicating X is genetically inactive, in both XXX and XXY triploids, cells from the embryonic region tended to have only one active X chromosome, whereas those from the extra-embryonic membranes tended to have two active X chromosomes. The single active X chromosome was either paternal or maternal in origin, but two active X chromosomes were overwhelmingly maternal in origin, suggesting paternal X-inactivation in extra-embryonic tissues.     

Chromosomal origin of small ring marker chromosomes in man: characterization by molecular genetics.

Ten cases of small ring chromosomes which did not stain with distamycinA/DAPI and did not possess satellite regions associated with nucleolus-organizing regions are described. In situ hybridization with a battery of biotinylated pericentric repeat probes specific either for individual chromosomes or for groups of chromosomes allowed the identification of the chromosomal origin of these marker chromosomes. There was one example of a marker derived from each of chromosomes 1, 3, 6, 14, 16, 18, 20, 13 or 21, and the X, and there were two examples of markers derived from chromosome 12. One case possessed two markers, one derived from chromosome 6, and one derived from the X. The mechanism of generation of ring marker chromosomes is discussed. Five of seven cases who could be phenotypically assessed were abnormal. Three of these--the first with a ring chromosome derived from chromosome 1; the second with two markers, one derived from chromosome 6 and the other from the X chromosome; and the third with a ring chromosome derived from chromosome 20--each possessed distinctive facies. Additional cases with identified rings may allow the delineation of new chromosomal syndromes.     

Stabilization of dicentric chromosomes in Saccharomyces cerevisiae by telomere addition to broken ends or by centromere deletion.

We introduced CEN6 DNA via integrative transformation into the right arm of chromosome II in a haploid Saccharomyces cerevisiae strain thus creating a dicentric chromosome. The majority of the transformed cells did not grow into colonies as concluded from control transformations with mutated CEN6 DNA. Five percent of the initial transformants with the wild-type centromere gave rise to well growing cells. We analysed the probable fate of the dicentric chromosome in two transformants by electrophoretic separation of chromosome sized DNA and by hybridizations with chromosome II DNA probes. We found two different mechanisms which generated cells lacking dicentric chromosomes. The first mechanism is breakage of the chromatid between the two-centromeres and healing of the new ends to functional telomeres thus creating progeny cells with the chromosome II information split into two genetically stable new chromosomes one carrying CEN2 and the other CEN6. The second mechanism is loss of the resident CEN2 by a 30-50 kb deletion event which resulted in a genetically stable but shortened chromosome II. Both mechanisms operated in the two transformants studied.     

Chromosome rearrangements between chicken and guinea fowl defined by comparative chromosome painting and FISH mapping of DNA clones

Chromosome homology between chicken (Gallus gallus) and guinea fowl (Numida meleagris) was investigated by comparative chromosome painting with chicken whole chromosome paints for chromosomes 1-9 and Z and by comparative mapping of 38 macrochromosome-specific (chromosomes 1-8 and Z) and 30 microchromosome-specific chicken cosmid DNA clones. The comparative chromosome analysis revealed that the homology of macrochromosomes is highly conserved between the two species except for two inter-chromosomal rearrangements. Guinea fowl chromosome 4 represented the centric fusion of chicken chromosome 9 with the q arm of chicken chromosome 4. Guinea fowl chromosome 5 resulted from the fusion of chicken chromosomes 6 and 7. A pericentric inversion was found in guinea fowl chromosome 7, which corresponded to chicken chromosome 8. All the chicken microchromosome-specific DNA clones were also localized to microchromosomes of guinea fowl except for several clones localized to the short arm of chromosome 4. These results suggest that the cytogenetic genome organization is highly conserved between chicken and guinea fowl.     

Chromosome evolution in fish: BrdU replication patterns demonstrate chromosome homeologies in two species of the genus Astyanax

A comparison of R-banding patterns obtained by 5-bromodeoxyuridine incorporation was made between the chromosomes of two fish species of the genus Astyanax (Characiformes: Tetragonopterinae), A. altiparanae with 2n = 50 chromosomes, and A. schubarti with 2n = 36 chromosomes. The two species present the highest and the lowest chromosome numbers found in this fish genus, respectively, for which the modal chromosome number is 50. R-band homeology was detected, involving eleven chromosomes of A. schubarti and seventeen chromosomes of A. altiparanae, indicating a close chromosomal relationship between the two species, in spite of their great difference in chromosome number. A chromosome fusion in the past history of the group was hypothesized as a possible cause of the discrepant chromosome numbers of the two species.     

Comparative FISH analysis of distribution of B chromosome repetitive DNA in A an d B chromosomes in two subspecies of Podisma sapporensis (Orthoptera, Acrididae)

FISH analysis of B chromosome repetitive DNA distribution in A and B chromosomes of two subspecies of Podisma sapporensis (P. s. sapporensis and P. s. krylonensis) was performed. In the B chromosomes, C-positive regions contained homologous DNA repeats present also in some C-positive A chromosome regions. Most C-negative regions contained DNA repeats characteristic of A chromosome euchromatic regions. The two subspecies analyzed differed in the location of A chromosome regions enriched with repeats homologous to repeats of B chromosomes. The only common region enriched with these B chromosome repeats in both subspecies was the X chromosome pericentromeric region. The origin of B chromosomes in P. sapporensis is discussed.     

Centromere inactivation and epigenetic modifications of a plant chromosome with three functional centromeres

A chromosome with two functional centromeres is cytologically unstable and can only be stabilized when one of the two centromeres becomes inactivated via poorly understood mechanisms. Here, we report a transmissible chromosome with multiple centromeres in wheat. This chromosome encompassed one large and two small domains containing the centromeric histone CENH3. The two small centromeres are in a close vicinity and often fused as a single centromere on metaphase chromosomes. This fused centromere contained approximately 30% of the CENH3 compared to the large centromere. An intact tricentric chromosome was transmitted to about 70% of the progenies, which was likely a consequence of the dominating pulling capacity of the large centromere during anaphases of meiosis. The tricentric chromosome showed characteristics typical to dicentric chromosomes, including chromosome breaks and centromere inactivation. Remarkably, inactivation was always associated with the small centromeres, indicating that small centromeres are less likely to survive than large ones in dicentric chromosomes. The inactivation of the small centromeres also coincided with changes of specific histone modifications, including H3K27me2 and H3K27me3, of the pericentromeric chromatin.     

Analysis of reciprocal translocations by chromosome painting: applications and limitations of the technique.

Fluorescent in situ hybridization with chromosome-specific DNA libraries (chromosome painting) is an important new method for assessing chromosome rearrangements. In the research presented in this paper, two familial reciprocal translocations have been studied in the balanced and unbalanced forms, using both traditional G-banding techniques and chromosome painting. Although for each case two chromosomes were involved in the rearrangement, we found that only one chromosome library was suitable for detecting the translocation. These findings illustrate both the potential and the limitations of chromosome painting as a diagnostic tool in cytogenetics.     

Locating a site on the maize B chromosome that controls preferential fertilization.

In maize, the B chromosome can undergo nondisjunction at the second pollen mitosis, producing sperm with two B chromosomes and sperm with zero B chromosomes. Preferential fertilization is the ability of the sperm carrying two B chromosomes to transmit more frequently to the embryo of a kernel than the sperm lacking the B chromosome. A translocation involving the B chromosome and chromosome 9, TB-9Sb, has been used to study preferential fertilization. The B-9 chromosome has the same properties of nondisjunction and preferential fertilization as the standard B chromosome. Deletion derivatives of B-9, which lack the centric heterochromatin and possibly some adjacent euchromatin, were tested for their ability to induce preferential fertilization. They were found to lack the capacity for preferential fertilization.     

Isolabeling of the long arm of the human Y chromosome demonstrated by the FPG technique

Isolabeling segments were found in the distal region of the long arm of Y chromosomes derived from human leukocytes grown through two replication cycles in medium containing BrdU and stained by the FPG technique. Three main types of Y chromosome staining patterns were demonstrated: I-Y chromosome with typical SCD, II-Y chromosome with weakly stained distal regions of long arms (isolabeling segments), III-Y chromosome with both terminal regions displaying SCD interrupted by one isolabeled segment. The existence of different types of Y chromosome staining patterns was explained on the basis of the previously described hypothesis of unequal distribution of thymine residues between two DNA polynucleotide chains in the distal part of the long arms of human Y chromosomes.     

Association of inter- and intrachromosomal exchanges with the distribution of low- and high-LET radiation-induced breaks in chromosomes

To study the effects of low- and high-linear energy transfer (LET) radiation on break locations within a chromosome, we exposed human epithelial cells in vitro to (137)Cs γ rays at both low and high dose rates, secondary neutrons at a low dose rate, and 600 MeV/u iron ions at a high dose rate. Breakpoints were identified using multicolor banding in situ hybridization (mBAND), which paints chromosome 3 in 23 different colored bands. For all four radiation scenarios, breakpoint distributions were found to be different from the predicted distribution based on band width. Detailed analysis of chromosome fragment ends involved in inter- or intrachromosomal exchanges revealed that the distributions of fragment ends participating in interchromosomal exchanges were similar between the two low-LET radiation dose rates and between the two high-LET radiation types, but the distributions were less similar between low- and high-LET radiations. For fragment ends participating in intrachromosomal exchanges, the distributions for all four radiation scenarios were similar, with clusters of breaks found in three regions. Analysis of the locations of the two fragment ends in chromosome 3 that joined to form an intrachromosomal exchange demonstrated that two breaks with a greater genomic separation can be more likely to rejoin than two closer breaks, indicating that chromatin folding can play an important role in the rejoining of chromosome breaks. Comparison of the breakpoint distributions to the distributions of genes indicated that the gene-rich regions do not necessarily contain more breaks. In general, breakpoint distributions depend on whether a chromosome fragment joins with another fragment in the same chromosome or with a fragment from a different chromosome.     

Nonhistone nuclear proteins specific to certain mouse embryonal carcinoma clones having an inactive X chromosome

By means of one-dimensional polyacrylamide slab gel electrophoresis nonhistone nuclear proteins were compared in murine embryonal carcinoma cell clones with two X chromosomes; both are active in some clones and one of them is inactive in others and in a population of cells having only one X chromosome. Under our experimental conditions, we succeeded in finding two extra bands at approximately 46,000 Da in cells having an inactive X chromosome. Furthermore, a band at approximately 71,000 Da was significantly heavier in cells having an inactive X chromosome than in those having two active X or those having only one X chromosome.     

Human chromosome 21-encoded cDNA clones

We have employed two strategies to isolate random cDNA clones encoded by chromosome 21. In the first approach, a cDNA library representing expressed genes of WA17, a mouse-human somatic cell hybrid carrying chromosome 21 as its sole human chromosome, was screened with total human DNA to identify human chromosome 21-specific cDNAs. The second approach utilized previously characterized single-copy genomic fragments from chromosome 21 as probes to retrieve homologous coding sequences from a human fetal brain cDNA library. Six cDNA clones on chromosome 21 were obtained in this manner. Two were localized to the proximal long arm of chromosome 21, two to the distal portion of the long arm, and one to the region of 21q22 implicated in the pathology of Down syndrome.     

Antibody labelling and flow cytometric analysis of metaphase chromosomes reveals two discrete structural forms

Metaphase chromosomes from cultured Chinese Hamster Ovary cells were labelled in suspension with a monoclonal antibody to histone 2B, counterstained with propidium iodide (PI) and analysed by flow cytometry. Contour plots of antibody binding (FITC fluorescence) against DNA content (PI fluorescence) revealed two discrete forms of each individual chromosome, showing high and low levels of antibody binding respectively. The two types of chromosome were easily distinguishable by immunofluorescence microscopy. The distribution of individual chromosomes between the two populations was related to chromosome size, with larger chromosomes predominating in the low-labelling population and vice versa. Variations in ionic strength, pH, divalent cation concentration or preparation procedure influenced the absolute level of antibody binding but did not eliminate the two populations. In contrast, preincubation with intercalating dyes, such as ethidium and propidium, inhibited antibody binding and generated a single, low-labelling population. Preliminary evidence suggests that in vivo changes in chromosome structure can affect the distribution of chromosomes between the two populations. Prolonged exposure of cells to Colcemid prior to chromosome isolation, a procedure known to increase chromosome condensation, resulted in a progressive shift into the low-labelling population. Our results suggest that chromosomes undergo a relatively rapid transition from the high-labelling to the low-labelling form during the prometaphase-metaphase stage of mitosis. The timing of this transition appears to be size dependent, with the larger chromosomes preceding the smaller. The transition may represent a change in chromosome condensation.