The absence of the somatic association of centromeres of homologous chromosomes in grass mitotic metaphases

Root-tip metaphases from Hordeum vulgare (19 cells), H. marinum (11 cells), Aegilops umbellulata (10 cells) and Zea mays (10 cells) were completely reconstructed from electron micrographs of serially sectioned nuclei. The identity of each chromosome was found by measuring the volumes of its two arms and the presence or absence of a secondary constriction at the nucleolar organising region. With the position of the centromere in three dimensions, these data were used to analyse the relative positions of homologous and heterologous centromeres. In 31 out of the 50 cells analysed, homologues were on average further apart than heterologues. Except for two nucleolar organising chromosomes, there was no evidence of any tendency for the distances between different homologue types to be differently distributed from distances between heterologues. Average distances between homologues of the single nucleolar organising chromosome (linkage group 6) of Zea (2n = 20) were lower than the average for heterologues and the interhomologue distances were distributed significantly differently from the separation distances of chromosome 6 to other chromosomes. Presumably this association occurred because of nucleolar fusion in the previous interphase. Homologues of one of the two nucleolar organising chromosomes of A. umbellulata were also distributed significantly differently from heterologues, with a tendency for homologues to lie farther apart than the average heterologous pair. These results do not support previous work using squashed and spread metaphase preparations (some including abnormal, marked chromosomes) for these species.     

Recovery of normal photosynthesis and respiration in common wheat with Agropyron cytoplasms by telocentric Agropyron chromosomes

Summary Alloplasmic common wheat (Triticum aestivum L. cultivais Penjamo 62 and Siete Cerros 66) with cytoplasms of wheatgrass (Agropyron trichophorum and Ag. glaucum) showed two aberrant phenotypes, i.e., gross reduction in plant vigor and male sterility. Plant vigor and male fertility were restored by cytoplasm-specific telocentric chromosomes (telosomes). Studies on carbon assimilation and consumption and on oxygen evolution and uptake showed that maximum rates of apparent photosynthesis were significantly lower in the alloplasmic lines than in their corresponding euplasmic lines and that the telosomes restored a normal level of photosynthesis. The decreased apparent photosynthetic rates in the alloplasmic lines were shown to be not due to decreased rates of true photosynthesis but to increased rates of dark respiration in the green leaves. In contrast, dark respiration in the roots was significantly low in the alloplasmic lines. The alloplasmic lines also showed decreased rates of respiratory consumption of new photosynthates. These results suggest that growth depression and male sterility in the alloplasmic lines are related to aberrant mitochondrial function, which is compensated for by cytoplasm-specific telosomes.     

Comparisons of recombination frequencies in hybrids involving telocentric and bibrachial wheat chromosomes

Telosomic stocks have been extensively used to map genes to chromosome arms and to determine gene-to-centromere genetic distances. It has been suggested that if a chromosome arm is present as a telosome, recombination frequencies will be drastically reduced in the centromeric region. However, previous studies have not considered the bias in recombination estimates due to selection against aneuploid gametes produced by failure of pairing at the first meiotic division. Formulas are derived here for adjusting recombination estimates for this bias. Adjusted recombination frequencies between markers located on both sides of the centromeres are analyzed in three different pairs of wheat (Triticum aestivum) isogenic segregating populations involving bibrachial and telocentric chromosomes. Recombination frequencies estimated from crosses involving telocentric chromosomes were not significantly different from recombination frequencies estimated from isogenic crosses involving bibrachial chromosomes. The implications of the present findings for karyotype evolution, and specifically for Robertsonian fissions and fusions, are discussed. Received: 10 March 1999 / Accepted: 17 June 1999     

Stacking pairs of disease resistance genes in wheat populations using telocentric chromosomes

Resistance of wheat to diseases such as fusarium head blight (FHB) and leaf rust is more effective and durable when resistance genes are stacked. This study investigated whether pairs of disease resistance genes will become fixed at higher frequencies in subsequent generations when placed in the hemizygous condition using telocentric chromosomes. Three pairs of telocentric chromosomes were tested for their male and female transmission to predict the fixation rate of hemizygous chromosome arms using reciprocal testcrosses. Hemizygous arm transmission was about 50% through ovules and about 75% through pollen because of pollen certation. To test if a corresponding increase in disease resistance could be observed in populations utilizing telocentric chromosomes, three resistance gene pairs were analyzed separately in three populations. These pairs were Lr16/Lr34 and Lr22a/Lr52 for resistance to leaf rust and Fhb1/Qfhs.ifa-5A for FHB resistance. Each of these gene combinations was involved in a crossing and selection scheme that identified F₁ plants that were either dihybrid or double monotelodisomic (DMTD). For each resistance gene combination F₃ families were produced for phenotypic testing. The Lr16/Lr34 and Lr22a/Lr52 F₃ populations both showed a sharp increase in leaf rust resistance among families derived from DMTD F₁ plants compared to those from dihybrid F₁ plants. A smaller increased resistance was found in the FHB population. The increased frequency of resistance was attributed to pollen certation and zygotic selection against the ditelosomic and double ditelosomic conditions. We conclude that telocentric chromosomes are a viable breeding tool to fix gene stacks.     

Somatic association of chromosomes in diploid and hexaploid Avena

The frequency distributions of certain homologous and non-homologous chromosomes in somatic cells of diploid and liexaploid species of Avena were studied and compared with the theoretical random distribution.All homologous chromosomes studied irrespective of shape and size showed non-random distribution in both diploid and hexaploid species. In all cases the homologous chromosomes were closer than would be expected with random distribution. Chromosomal characters such as size, shape, and presence or absence of nucleolar organizing regions did not exert appreciable influence on the somatic association of homologues.While the non-homologues followed the theoretical random distribution in diploid species, a significant deviation from the random curve was noted for non-homologues in hexaploid species. However, diploid and hexaploid non-homologous chromosomes had characteristic S-shaped cumulative frequency distributions which were distinct from the half-moon-shaped ones obtained for homologous chromosomes.The different regions (short arm, long arm, centromere and mid-points) of two pairs of homologous chromosomes (one of them being nucleolar) studied showed non-random distribution with the exception of the long arm of the non-nucleolar chromosome. From these results the role of the centromere or mutual attraction of homologous segments could not be assessed with certainty.Contribution No. 232 from the Research Station, Central Experimental Farm, Ottawa, Ontario, Canada.     

On the origin of telocentric chromosomes in mammals

The origin of mammalian telocentric chromosomes is considered under the classical (fusion) and fission hypotheses using both theoretical analyses of the mechanisms proposed under the two hypotheses, and the published chromosomal data for 723 mammal species. Telocentrics are defined on the basis of short arm size (S_w) as chromosomes with S_w < 0·1(Imai, 1976). The fusion hypothesis lacks adequate models for producing these telocentrics, but their origin is readily understood under the fission hypothesis. Based on these analyses, I propose a cyclical model of chromosome change, symbolized:

in which T, A, and $\text{M}$ are, respectively, telocentric, acrocentric, and meta-, submeta- and subtelocentric chromosomes. The chief elements of this model are centric fission $\text{(}\text{M}\text{→ T + T)}$, tandem growth of constitutive heterochromatin (T → A), and pericentric inversion $\text{(A →}\text{M}\text{)}$. Under this model, therefore, mammalian karyotypes have an overall tendency, with occasional reversals, to evolve higher numbers of both chromosomes and chromosome arms.     

Cytogenetic relationship ofAegilops longissima chromosomes with common wheat chromosomes

The relationships of three wheat-Aegilops longissima chromosome addition lines A, C, and D with homoeologous wheat chromosomes were studied in PMC meiosis. Substitutions of alien chromosome A for wheat chromosome 6 B, chromosome C for 1 B and chromosome D for 4 B were obtained. The production of 4 BS/C and 7 BS/D chromosome translocations indicated cytogenetic relationships of C partially to homoeologous wheat chromosomes of group 1 and 4, and D partially to homoeologous wheat chromosomes of group 4 and 7.     

Somatic association of chromosomes in asynaptic genotypes of Avena sativa L.

The distribution of distances between homologous chromosomes in root tip cells of Avena sativa was studied in synaptic and asynaptic genotypes. The distances between homologous chromosomes were smaller than that calculated for two randomly distributed chromosomes, while non-homologous chromosomes did not deviate from the random theoretical distribution. The distances between homologous chromosomes in the asynaptic genotype were significantly greater than in synaptic plants. The loose association of homologous chromosomes in somatic tissue is correlated with the failure of chromosome pairing at meiosis in asynaptic plants.     

The theory of the pairing of telocentric chromosomes in triploids and trisomics

The possible pairing patterns of telocentric chromosomes in triploids and trisomics are considered and expressions are derived allowing the prediction of expected meiotic, chromosomal, and cellular pairing patterns. The calculation of the relative affinity of the homoeologous chromosomes involved in the pairing patterns with the telocentrics is discussed.     

Cytogenetic identification of Triticum peregrinum chromosomes added to common wheat.

C-banded karyotypes of a complete set of 14 Triticum peregrinum whole chromosome addition lines and 25 telosomic addition lines are reported. The added T. peregrinum chromosomes were not structurally rearranged compared with the corresponding chromosomes of the donor accession. Comprehensive karyotypic analysis confirmed Triticum umbellulatum as the donor species of the Uv genome and identified Triticum longissimum as the donor species of the Sv genome of T. peregrinum. Neither the Uv nor Sv genome chromosomes of the T. peregrinum accession showed large modifications when compared with the ancestral U and S1 genomes. Key words : Triticum aestivum, Triticum peregrinum, Triticum umbellulatum, Triticum longissimum, chromosome addition lines, C-banding.     

Stable telocentric chromosomes produced by centric fission in chinese hamster cells in vitro

Metaphase examination of pseudodiploid Chinese hamster cells revealed that spontaneous breaks or fission occurred rather frequently (2.9%) at the centromeric regions of subtelo- or metacentric chromosomes, resulting in the production of telocentric chromosomes. The centromeric fission appeared to occur in every member of the chromosome complement. An attempt was made to isolate cells possessing thus derived telocentrics from the cell population and gave two clonal lines which were retaining one and two telocentric chromosomes, respectively. Both banding and labeling patterns of these chromosomes indicated unequivocally their X chromosome origin. They were transmitted successively to the daughter cells during a 3-month culture period, showing no tendency to fuse to produce a metacentric chromosome.Contribution No. 897 from the National Institute of Genetics, Japan.     

The loss of centromeres from chromosomes of aged women

Both aneuploid cells in mitosis and nondisjunction in meiosis increase with advancing age. The cause(s) of these phenomena remains unknown. Here, it was confirmed that a positive Cd-band reflects the presence of a functioning centromere while a negative reaction is indicative of its inactivation or loss. We applied the Cd-banding technique to mitotic spreads obtained from 14 aged and 13 control females (peripheral blood culture). Of 6,474 scored chromosomes from the aged women, 62(0.96%) were Cd-negative; this was the case in 12 of 3,861 (0.31%) chromosomes from the younger controls. The difference was highly significant (P less than .001). About 60% of the 62 Cd-negative chromosomes from aged women belong to the C group. Chromosomes from six of the 14 aged females were additionally examined by either the C-band or the distamycin-DAPI technique; of 2,080 chromosomes, 13 (0.63%) showed premature separation of their centromeres (control: 7/5, 080, 0.14%). Our findings suggest that in aged women, the chromosomes tend to lose their Cd-positive material and the function of the centromere.     

Construction and behavior of circularly permuted and telocentric chromosomes in Saccharomyces cerevisiae.

We developed techniques that allow us to construct novel variants of Saccharomyces cerevisiae chromosomes. These modified chromosomes have precisely determined structures. A metacentric derivative of chromosome III which lacks the telomere-associated X and Y' elements, which are found at the telomeres of most yeast chromosomes, behaves normally in both mitosis and meiosis. We made a circularly permuted telocentric version of yeast chromosome III whose closest telomere was 33 kilobases from the centromere. This telocentric chromosome was lost at a frequency of 1.6 X 10(-5) per cell compared with a frequency of 4.0 X 10(-6) for the natural metacentric version of chromosome III. An extremely telocentric chromosome whose closet telomere was only 3.5 kilobases from the centromere was lost at a frequency of 6.0 X 10(-5). The mitotic stability of telocentric chromosomes shows that the very high frequency of nondisjunction observed for short linear artificial chromosomes is not due to inadequate centromere-telomere separation.     

The effect of additions ofAegilops longissima chromosomes on grain protein in common wheat

Summary The effect of various chromosomes ofAegilops longissima when added to the common wheat cultivar Chinese Spring was evaluated at two levels of nitrogen fertilization for absolute and relative amount of protein in the grain. All the added chromosomes ofAe. longissima increased protein percentage: protein increase by chromosomes D, C and A averaged 3.8% while that by chromosomes F, E, G and B averaged 1.7%. Addition lines F, D and C had a significantly higher protein weight per grain. On the other hand, lines A, E and G had reduced grain protein weight per grain as compared with that of Chinese Spring. Line C carries the HMW glutenin and some of the gliadin subunits ofAe. longissima. The effect of this line, however, and obviously that of the other lines on protein content was through genes controlling the level of storage protein rather than through genes that code directly for these proteins. Nitrogen fertilization affected protein content and the relative amount of the various protein fractions in a similar manner in every addition line. When high levels of nitrogen fertilization were compared to low ones, the relative amount of the HMW glutenins remained constant while that of HMW gliadins increased and that of the LMW subunits decreased. In contrast to the nitrogen effect, increase in protein content by the addition oflongissima chromosomes did not change the relative amounts of the various protein fractions.The paper is based on a portion of a dissertation to be submitted by A.A.L. in partial fulfillment of the PhD requirements in the Feinberg Graduate School, The Weizmann Institute of Science, RehovotThe Marshall and Edith Korshak Professor of Plant Cytogenetics     

Flow sorting of mitotic chromosomes in common wheat (Triticum aestivum L.)

The aim of this study was to develop an improved procedure for preparation of chromosome suspensions, and to evaluate the potential of flow cytometry for chromosome sorting in wheat. Suspensions of intact chromosomes were prepared by mechanical homogenization of synchronized root tips after mild fixation with formaldehyde. Histograms of relative fluorescence intensity (flow karyotypes) obtained after the analysis of DAPI-stained chromosomes were characterized and the chromosome content of all peaks on wheat flow karyotype was determined for the first time. Only chromosome 3B could be discriminated on flow karyotypes of wheat lines with standard karyotype. Remaining chromosomes formed three composite peaks and could be sorted only as groups. Chromosome 3B could be sorted at purity >95% as determined by microscopic evaluation of sorted fractions that were labeled using C-PRINS with primers for GAA microsatellites and for Afa repeats, respectively. Chromosome 5BL/7BL could be sorted in two wheat cultivars at similar purity, indicating a potential of various wheat stocks for sorting of other chromosome types. PCR with chromosome-specific primers confirmed the identity of sorted fractions and suitability of flow-sorted chromosomes for physical mapping and for construction of small-insert DNA libraries. Sorted chromosomes were also found suitable for the preparation of high-molecular-weight DNA. On the basis of these results, it seems realistic to propose construction of large-insert chromosome-specific DNA libraries in wheat. The availability of such libraries would greatly simplify the analysis of the complex wheat genome.     

Retrotransposon-related DNA sequences in the centromeres of grass chromosomes.

Several distinct DNA fragments were subcloned from a sorghum (Sorghum bicolor) bacterial artificial chromosome clone 13I16 that was derived from a centromere. Three fragments showed significant sequence identity to either Ty3/gypsy- or Ty1/copia-like retrotransposons. Fluorescence in situ hybridization (FISH) analysis revealed that the Ty1/copia-related DNA sequences are not specific to the centromeric regions. However, the Ty3/gypsy-related sequences were present exclusively in the centromeres of all sorghum chromosomes. FISH and gel-blot hybridization showed that these sequences are also conserved in the centromeric regions of all species within Gramineae. Thus, we report a new retrotransposon that is conserved in specific chromosomal regions of distantly related eukaryotic species. We propose that the Ty3/gypsy-like retrotransposons in the grass centromeres may be ancient insertions and are likely to have been amplified during centromere evolution. The possible role of centromeric retrotransposons in plant centromere function is discussed.