Inversion heterozygosity and sub-chromatid exchange in Agave stricta

A plant of Agave stricta Salm. (2n=60) has a bimodal complement of 10 L, 4 M and 46 S chromosomes. It is heterozygous for a paracentric inversion which involves the middle third of the long arm of one of the L chromosomes. It produces at anaphase I bridges and fragments and also loops and fragments, both single and double. Breakage and reunion at the sub-chromatid and at the chromatid level produce side-arm bridges and bridges and fragments respectively at anaphase I. A method is given, based on chiasma frequency, which will in certain cases of inversion heterozygosity provide a reasonable estimate of the position and the length in genetic map units of an inverted segment with respect to the whole chromatid arm.     

Incomplete sister chromatid separation of long chromosome arms

Chromosome segregation ensures the equal partitioning of chromosomes at mitosis. However, long chromosome arms may pose a problem for complete sister chromatid separation. In this paper we report on the analysis of cell division in primary cells from field vole Microtus agrestis, a species with 52 chromosomes including two giant sex chromosomes. Dual chromosome painting with probes specific for the X and the Y chromosomes showed that these long chromosomes are prone to mis-segregate, producing DNA bridges between daughter nuclei and micronuclei. Analysis of mitotic cells with incomplete chromatid separation showed that reassembly of the nuclear membrane, deposition of INner CENtromere Protein (INCENP)/Aurora B to the spindle midzone and furrow formation occur while the two groups of daughter chromosomes are still connected by sex chromosome arms. Late cytokinetic processes are not efficiently inhibited by the incomplete segregation as in a significant number of cell divisions cytoplasmic abscission proceeds while Aurora B is at the midbody. Live-cell imaging during late mitotic stages also revealed abnormal cell division with persistent sister chromatid connections. We conclude that late mitotic regulatory events do not monitor incomplete sister chromatid separation of the large X and Y chromosomes of Microtus agrestis, leading to defective segregation of these chromosomes. These findings suggest a limit in chromosome arm length for efficient chromosome transmission through mitosis.Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

The use of gene conversion to study synaptinemal complex structure and molecular details of chromatid pairing in meiosis

Summary Gene conversion can be used to study: the topography and pairing relationships of the four chromatids of a bivalent at the time of crossing over and hybrid DNA formation, the lengths of intimately paired segments and the frequency of intimate pairing at particular sites. Conversion ratios of different types, corresponding-site interference, co-conversion, and the range and distribution of conversion frequencies are discussed in relation to DNA and chromatid pairing, and synaptinemal complex organisation. Conversion data from Ascobolus immersus and other fungi are compared with electron microscope data from various organisms and with models of the synaptinemal complex.     

Relationship between sister chromatid exchanges and DNA replication in somatic cells of Drosophila melanogaster

In Drosophila melanogaster cell lines and larval neuroblast cells, two aspects of the phenomenon of sister chromatid exchanges were analyzed: (1) the frequency of SCEs in relation to the ploidy level (comparing diploid and tetraploid cells) and in relation to the cell type (comparing embryonic and larval cells) (2) the localization of the sites of exchange with reference to eu- and heterochromatin. A good correlation between SCE frequency and genome size in the same cell type (in distant species also), but a significant difference in the SCE rate between different cell types within the same species, were found. The results confirmed also the non-random distribution of SCEs in the different portions of the genome since a preferential localization in the euchromatin was clearly demonstrated. Moreover, a direct proportionality between SCE frequency and the length of the S phase was supposed, favouring the hypothesis of a relationship between the phenomenon of sister chromatid exchanges and DNA replication.     

SUB-CHROMATID REARRANGEMENTS IN TRILLIUM ERECTUM. I. ORIGIN AND NATURE OF CONFIGURATIONS INDUCED BY IONIZING RADIATION

Wilson , G. B. (Michigan State U., East Lansing.), A. H. Sparrow , and Virginia Pond . Subchromatid rearrangements in Trillium erectum. I. Origin and nature of configurations induced by ionizing radiation. Amer. Jour. Bot. 46(4): 309–316. Illus. 1959.—Microsporocytes of Trillium erectum were x-irradiated with 25 r at various stages of meiotic prophase and at first metaphase. Analysis of these cells at the following first and second anaphase revealed that post-pachytene irradiation produces 2-side-arm bridges which are indicative of half-chromatid exchanges. The occurrence of these bridges and knowledge of the structure and spatial relationship of chromatid strands in T. erectum have led to certain conclusions regarding the target and the number of strands broken by a single event: (1) the most likely target for primary effects is the 4 associated half-chromatids of a half-bivalent. The results of irradiation experiments suggest that the half-bivalent is effectively as well as structurally quadripartite at stages following pachytene. (2) Consideration of the configurations which would result from breakage and rejoining of 2, 3 or all 4 strands of the half-bivalent indicates that only 2 of the 4 half-chromatids are broken by a single event. Exchanges between 2 half-chromatids of sister chromatids will produce two recognizable types of 2-side-arm bridges: one with a true dicentric half-chromatid and one in which the bridge results merely from an interlocking of coils. Whether a 2-side-arm bridge appears at first or second meiotic anaphase is determined by the position and number of chiasmata between the point of breakage and the kinetochore. No 2-side-arm bridges have been detected at microspore anaphase following meiotic prophase irradiation. The types of configurations which might be expected at microspore metaphase as a result of broken 2-side-arm bridges are noted.     

Configurations resulting from iso-chromatid and iso-subchromatid unions after meiotic and mitotic prophase irradiation

Summary By making use of the chromosomes of Trillium erectum as a model, potential and actual configurations arising from presumed iso-chromatid and iso-subchromatid unions after irradiation of meiotic or mitotic prophase have been studied and analyzed. Diagrams and photographs of various recognizable types of chromatid or subchromatid rearrangements are presented. A minimum of two iso-chromatid unions within an arm of a single chromosome in meiotic prophase, if separated by a single chiasma, can give rise to a monocentric chromosome with a triplicated segment, the middle portion of which is an inversion. A minimum of two iso-subchromatid breaks within an arm at either meiotic or mitotic prophase also can result in the production of a monocentric chromosome containing a triplicated segment. The stage of appearance of dicentrics or bridges arising from chromatid or subchromatid unions in meiotic prophase is influenced by chiasma number and pattern and by the number of strands per chromosome or chromatid. Some of the rearrangements described may have genetic and evolutionary implication of considerable potential importance which has not been recognized previously.Research carried out at Brookhaven National Laboratory under the auspices of the U.S. Atomic Energy Commission.     

High density of REC8 constrains sister chromatid axes and prevents illegitimate synaptonemal complex formation

During meiosis, cohesin complexes mediate sister chromatid cohesion (SCC), synaptonemal complex (SC) assembly and synapsis. Here, using super‐resolution microscopy, we imaged sister chromatid axes in mouse meiocytes that have normal or reduced levels of cohesin complexes, assessing the relationship between localization of cohesin complexes, SCC and SC formation. We show that REC8 foci are separated from each other by a distance smaller than 15% of the total chromosome axis length in wild‐type meiocytes. Reduced levels of cohesin complexes result in a local separation of sister chromatid axial elements (LSAEs), as well as illegitimate SC formation at these sites. REC8 but not RAD21 or RAD21L cohesin complexes flank sites of LSAEs, whereas RAD21 and RAD21L appear predominantly along the separated sister‐chromatid axes. Based on these observations and a quantitative distribution analysis of REC8 along sister chromatid axes, we propose that the high density of randomly distributed REC8 cohesin complexes promotes SCC and prevents illegitimate SC formation.     

Salt bridges: geometrically specific, designable interactions

Salt bridges occur frequently in proteins, providing conformational specificity and contributing to molecular recognition and catalysis. We present a comprehensive analysis of these interactions in protein structures by surveying a large database of protein structures. Salt bridges between Asp or Glu and His, Arg, or Lys display extremely well-defined geometric preferences. Several previously observed preferences are confirmed, and others that were previously unrecognized are discovered. Salt bridges are explored for their preferences for different separations in sequence and in space, geometric preferences within proteins and at protein-protein interfaces, co-operativity in networked salt bridges, inclusion within metal-binding sites, preference for acidic electrons, apparent conformational side chain entropy reduction on formation, and degree of burial. Salt bridges occur far more frequently between residues at close than distant sequence separations, but, at close distances, there remain strong preferences for salt bridges at specific separations. Specific types of complex salt bridges, involving three or more members, are also discovered. As we observe a strong relationship between the propensity to form a salt bridge and the placement of salt-bridging residues in protein sequences, we discuss the role that salt bridges might play in kinetically influencing protein folding and thermodynamically stabilizing the native conformation. We also develop a quantitative method to select appropriate crystal structure resolution and B-factor cutoffs. Detailed knowledge of these geometric and sequence dependences should aid de novo design and prediction algorithms.     

THE EVOLUTION OF POLLEN TETRADS IN ONAGRACEAE

In Onagraceae, pollen is shed in mature tetrads in most Epilobieae, many species of Ludwigia (Jussiaeeae), and two closely related species of the large genus Camissonia (Onagreae). Mature tetrads of Camissonia cardiophylla and representative species of Epilobium and Ludwigia were examined with light, scanning, and transmission electron microscopes. Morphological diagnoses of monad units indicated that individual taxa could be readily distinguished. Statistical analyses of tetrads which remained after acetolysis treatment revealed significant differences in the strength of the binding mechanisms. Mechanisms of tetrad cohesion were found to consist of two principal types. Common to all taxa is cohesion of pollen wall surfaces at the aperture margins; this mechanism is well known in many angiosperm groups. With the exception of Camissonia, the remaining taxa also display binding by means of short exine fragments between adjacent pollen units. These fragments, termed bridges and reported here for the first time, are located in the area extending from the aperture margins to near the center of the proximal exine faces. Thin sections reveal that layers of the bridges are identical with those of the exine. Comparisons were made between bridges and viscin threads, both of which occur on the proximal faces of the grains. Viscin threads are present on all pollen grains in Onagraceae and exhibit distinctive morphologies, and bridges were viewed morphogenetically as related to viscin threads but including an endexine layer and occupying a position near the apertures where cohesion of wall surfaces also occurs. In an evolutionary sense, the formation of mature tetrads almost certainly occurred independently in Camissonia and may have done so in Ludwigia and the Epilobieae.     

Meiotic chromosomal aberrations in wild populations of Podophyllum peltatum

Meiotic chromosomal aberrations observed in wild populations of the plant Podophyllum peltatum include incomplete homologous pairing, non-homologous pairing, and inversion heterozygosity in pachytene; univalents, asymmetrical bivalents, and translocation heterozygosity in metaphase-I; bridge and fragments in anaphase-I; and non-disjunction as detected in anaphase-II. Incomplete homologous pachytene pairing is believed to result in non-homologous pairing and in the formation of metaphase-I univalents. The unequal distribution and precocious division of univalents in anaphase-I leads to non-disjunction. Non-disjunction chromosomes (varying in frequency from 0.0 to 24.6%) appear to be distributed among the genome on the basis of chromosome length. Asymmetrical bivalents and anaphase-I side-arm bridges are believed to be caused by chromatid breakage and fusion rather than inversion heterozygosity. Of the 135 clones examined, 20 were found to be heterozygous for translocations. The possibility of widespread distribution of some translocations is suggested.     

Studies on graft unions

Summary The occurrence of plasmodesmata in the graft interfaces of two heteroplastic grafts (Impatiens walleriana onImpatiens olivieri andHelianthus annum onVicia faba) has been studied. For both systems two types of intercellular strand are described: 1. Continuous plasmodesmata interconnecting the cells of stock and scion and 2. half plasmodesmata traversing the wall part of one partner cell without connection to the abutting cell. Single strands or branched forms occur in both types of plasmodesma. In the case of half plasmodesmata, branchings with extended median nodules predominate. The distribution of half and continuous plasmodesmata varies with the different areas of a graft interface: in the region of bridging vascular tissues most cell connections are continuous. In areas where cortex or pith-derived callus cells and those of misaligned tissues (cortex/vascular tissue; cortex/pith; pith/vascular tissue) match, discontinuous strands predominate.Branched half plasmodesmata also occur in presumably fused walls between related callus cells; they are typical structures secondarily formed in non-division walls.The results are discussed with regard to compatibility/incompatibility phenomena in heterografts and the development and function of interspecific cell bridges.     

Homolog pairing and sister chromatid cohesion in heterochromatin in <Emphasis Type="Italic">Drosophila</Emphasis> male meiosis I

Drosophila males undergo meiosis without recombination or chiasmata but homologous chromosomes pair and disjoin regularly. The X–Y pair utilizes a specific repeated sequence within the heterochromatic ribosomal DNA blocks as a pairing site. No pairing sites have yet been identified for the autosomes. To search for such sites, we utilized probes targeting specific heterochromatic regions to assay heterochromatin pairing sequences and behavior in meiosis by fluorescence in situ hybridization (FISH). We found that the small fourth chromosome pairs at heterochromatic region 61 and associates with the X chromosome throughout prophase I. Homolog pairing of the fourth chromosome is disrupted when the homolog conjunction complex is perturbed by mutations in SNM or MNM. On the other hand, six tested heterochromatic regions of the major autosomes proved to be largely unpaired after early prophase I, suggesting that stable homolog pairing sites do not exist in heterochromatin of the major autosomes. Furthermore, FISH analysis revealed two distinct patterns of sister chromatid cohesion in heterochromatin: regions with stable cohesion and regions lacking cohesion. This suggests that meiotic sister chromatid cohesion is incomplete within heterochromatin and may occur at specific preferential sites.     

The amino acid sequence of the major parvalbumin of the whiting (Gadus merlangus).

1. The amino acid sequence of the major parvalbumin of the Whiting has been determined; the polypeptide chain is made of 108 residues, the terminal amino acid group is acetylated, there is no disulfide bridges, the structure of the two calcium binding sites is preserved and the distribution along the polypeptide chain of the hydrophobic residues implicated in the compact hydrophobic core of the protein is also maintained. 2. The comparison of this amino acid sequence with other parvalbumins indicates that it belongs to the beta type and that within the Gadidae family two types of parvalbumins also occur.     

Embryogenic Response and Mitotic Instability in Callus Cultures Derived from Maize Inbred Lines Differing in Heterochromatic Knob Content of Chromosomes

Four families of sister inbred lines derived from a tropicalmaize variety have been evaluated for their ability to formcallus cultures with a morphogenetic response. Lines were homozygousfor heterochromatic knobs at 6L₂, 6L₃, 7L and 8L₁but differedfor the presence or absence of K2L, K3L, K7S and K9S. Cleardifferences in embryogenic response were observed between thefamilies of inbreds. Only one family formed friable, highlyembryogenic type II calli; the other families formed slow growingnon-embryogenic or poorly embryogenic cultures. All the genotypesscreened showed a similar response to the two culture mediatested, suggesting that genetic factors are responsible forthe major differences among the families. Mitotic abnormalitieswere investigated in Feulgen preparations of most cultures.Anaphase bridges resulting from delayed chromatid separation,typical bridges and fragments were observed. In a previous study,delayed chromatids were shown to be held together at heterochromaticknob sites, while typical bridges would be formed by dicentricchromatids arising from breakage-fusion-bridge cycles initiatedby chromosome arms broken during the primary event. In the presentstudy, the frequency of both types of bridges was not strictlycorrelated with the knob content of the genotypes analysed.This suggests that knobs may undergo alterations in cultureleading to mitotic disturbance, and that this response may begenotype dependent.Copyright 1998 Annals of Botany Company Zea maysL., maize, plant tissue culture, somatic embryogenesis, somaclonal variation, heterochromatin, C-banding, breakage-fusion-bridge cycle.     

Fine structure of anaphase bridges in meiotic chromosomes of the crane fly Pales

Chromatin bridges of autosomal bivalents in anaphase I were observed in spermatocytes of Pales ferruginea. The bridges are formed without simultaneous production of akinetochoric (akinetic) fragments. A bridge consists of a single fiber up to approximately. 500 Å in diameter. Filamentous substructures of approximately 100 Å diameter can be visualized. It is suggested that these bridges represent a low order coiling of the chromatid, and may be caused by non-separation of the terminal segments of the chromatids (telomeres).     

Synapsis and crossing over within a paracentric inversion in the grasshopper,Camnula pellucida

A male grasshopper, Camnula pellucida (Scudder), was found to be heterozygous for a paracentric inversion occupying approximately 10% of the length of one of the two longest chromosomes. Analysis of 297 cells in pachytene revealed inversion loops, suspected inversion loops, asynapsis, and straight pairing in 1.0, 2.7, 8.4, and 87.9% of the analyzable cells, respectively. The frequency of straight pairing (87.9%) indicated a high degree of non-homologous pairing. Analysis of 603 cells in anaphase I and II, and in telophase I and II for the presence of acentric fragments and dicentric chromatid bridges indicated that crossing over within the inversion region occured in about 8% of the cells. The difference between the frequency of the observed plus suspected inversion loops in pachytene and that of the dicentric chromatid bridges and acentric fragments in anaphase I or subsequent stages was not statistically significant. The correspondence between the presence of inversion loops and crossovers within the region of the inversion is thus similar to that observed by Maguire (1966) for a short paracentric inversion in maize. The reasons for this correspondence are considered.Supported by grants GB 1585 and GB 6745 from the National Science Foundation, Washington, D.C.     

Differential response of constitutive and facultative heterochromatin in the manifestation of mitomycin induced chromosome aberrations in Chinese hamster cells in vitro

The distribution of chromatid aberrations induced by mitomycin C among the individual chromosomes of female and male Chinese hamster cells in vitro was studied. The aberrations were found to be non-randomly distributed. Among the autosomes, the chromosomes possessing constitutive heterochromatin were more often involved in aberrations as well as in homologous exchanges. The inactivated X chromosomes in the female cells offer a situation where the short arm is facultatively heterochromatic and the long arm constitutively heterochromatic, thus enabling an analysis of their response for aberration formation. The short arm was seldom found to be involved in the aberration. The long arm of the inactivated X was more often affected (5 to 10 times) than the long arm of the functional X though both are constitutively heterochromatic. The possible role of (a) structure of heterochromatin, (b) the chromocenter formation and their association, (c) allocycly, and (d) the qualitative differences in the DNA of different types of heterochromatin are discussed in relation to the formation of chromatid aberrations.     

Classification of chromosome segregation errors in cancer

Abnormal chromosome segregation at mitosis is one way by which neoplastic cells accumulate the many genetic abnormalities required for tumour development. In this paper, a straightforward morphology-based classification of chromosome segregation errors in cancer is suggested. This classification distinguishes between abnormalities in spindle symmetry (spindle multipolarity, size-asymmetry of ana-telophase poles) and abnormalities in sister chromatid segregation (chromosome bridges, chromatid bridges, chromosome lagging, acentric fragment lagging). Often, these categories of errors must be combined to accurately describe the events in a single abnormal mitotic cell. The suggested categories can to some extent be distinguished by standard chromatin staining. However, labelling of abnormal mitotic figures by fluorescence in situ hybridization and immunofluorescence enhances the accuracy of classification and also allows visualisation of the segregation of individual chromosomes, making it possible to detect non-disjunction also in the absence of gross alterations in mitotic morphology. Further characterisation of the molecular alterations leading to abnormal chromosome segregation together with the current developments in nano-level and real-time imaging will undoubtedly lead to an improved understanding of chromosome dynamics in cancer cells. Any morphology-based classification of chromosome segregation errors in cancer must therefore be taken as provisional, anticipating a satisfactory integration of morphology and molecular biology.     

GFP tagging of budding yeast chromosomes reveals that protein–protein interactions can mediate sister chromatid cohesion

Background Precise control of sister chromatid separation is essential for the accurate transmission of genetic information. Sister chromatids must remain linked to each other from the time of DNA replication until the onset of chromosome segregation, when the linkage must be promptly dissolved. Recent studies suggest that the machinery that is responsible for the destruction of mitotic cyclins also degrades proteins that play a role in maintaining sister chromatid linkage, and that this machinery is regulated by the spindle-assembly checkpoint. Studies on these problems in budding yeast are hampered by the inability to resolve its chromosomes by light or electron microscopy.Results We have developed a novel method for visualizing specific DNA sequences in fixed and living budding yeast cells. A tandem array of 256 copies of the Lac operator is integrated at the desired site in the genome and detected by the binding of a green fluorescent protein (GFP)–Lac repressor fusion expressed from the HIS3 promoter. Using this method, we show that sister chromatid segregation precedes the destruction of cyclin B. In mad or bub cells, which lack the spindle-assembly checkpoint, sister chromatid separation can occur in the absence of microtubules. The expression of a tetramerizing form of the GFP–Lac repressor, which can bind Lac operators on two different DNA molecules, can hold sister chromatids together under conditions in which they would normally separate.Conclusions We conclude that sister chromatid separation in budding yeast can occur in the absence of microtubule-dependent forces, and that protein complexes that can bind two different DNA molecules are capable of holding sister chromatids together.     

Sister chromatid exchanges in Drosophila melanogaster cell lines in vitro

The frequency of sister chromatid exchanges (SCEs) in two cell lines of Drosophila melanogaster with different karyotypes (XX and XY) was determined, considering (1) the distribution of SCEs within each chromosome, with reference to eu- and heterochromatin and (2) the distribution of SCEs in different chromosomes. A comparison was made between chromosome pairs within each karyotype and between the two different karyotypes. The following results were obtained. The SCEs are not randomly distributed along chromosomes, since exchanges were never observed in heterochromatin. SCEs are more frequent in XY than in XX cells; moreover, in both cell types there exists a significantly higher frequency of SCEs in the X chromosome than in the autosomes. These findings are discussed in relation to chromosome aberrations and mitotic recombination.