Prometaphase I and anaphase I in an interchange heterozygote of Allium triquetrum (Liliaceae)

Four unstable malorientations in a chain quadrivalent, as well as maloriented bivalents, were studied in fixed prometaphase I pollen-mothercells from an interchange heterozygote of Allium triquetrum. The relative frequencies of these malorientations, in cells from pollen sacs of different developmental ages, suggest that maloriented bivalents, on the average, reorient before maloriented quadrivalents; and that, similarly, there are differences in the timing of reorientation amongst the four types of maloriented quadrivalents. — Further, the proportion of anaphase I cells derived from alternate orientation in the interchange quadrivalent is lower than expected in pollen sacs with a high percentage of cells in mid-anaphase; but higher than expected in pollen sacs in which relatively few cells have as yet proceeded into anaphase, and also in those in which most cells have already passed through anaphase. Arguably, these data are a direct outcome of the differential behavior of unstable maloriented quadrivalents in the preceeding prometaphase.     

Anther development and the orientation of the interchange quadrivalent in pearl millet

The effects of developmental stage of the anther, ageing of the plant, inbreeding and season on meiotic segregation of an interchange chain quadrivalent were studied in pearl millet, Pennisetum americanum (L.) Leeke. The frequency of adjacent orientation (and segregation) increased with developmental stage of the anther, independent of other factors. Plant age and degree of inbreeding had no demonstrable effects, but there was an indication that high temperature favoured adjacent orientation. Chromosome contraction as measured by change in chromosome length appeared to be negligible during metaphase-anaphase. Therefore, increased adjacent orientation cannot readily be explained by metaphase reorientation resulting from the straightening of chromosomes caused by an increase in their rigidity. It is probable that the unoriented or mal-oriented quadrivalents observed regularly at early metaphase I continue to straighten out prior to their delayed orientation. When they finally orientate late in metaphase, their orientation will more likely be adjacent than alternate.     

A quadrivalent studied in living and fixed grasshopper spermatocytes

A single specimen of the grasshopper Melanoplus differentialis was discovered which was heterozygous for an interchange involving two of the largest chromosomes of the complement. The resulting quadrivalent occurred as a ring-of-four, chain-of-four, and, rarely, as two heteromorphic bivalents. Metaphase orientation and anaphase disjunction of some of these quadrivalents were recorded in living cells. The chain-of-four was stable in the configuration in which two pairs of kinetochores were oriented to each pole. The configuration in which three pairs of kinetochores were oriented to one pole and one pair to the other pole was also stable, but was shifted off the equator toward the pole to which three pairs of kinetochores were oriented. Given the nature of this quadrivalent the unusual stability of the 31 configuration is expected from current hypotheses of chromosome orientation and reorientation.     

Chromosome mal-orientation and reorientation during mitosis.

We argue that mal-orientation of mitotic chromosomes is not as rare as once believed. However, unlike bivalents during meiosis I, the reorientation of a mal-oriented mitotic chromosome has yet to be observed. This appears to be due, in part, to the difficulty in differentiating mal-oriented chromosomes from mono-oriented ones which are common during spindle formation in living mitotic cells. We assume that mitotic cells possess mechanisms for correcting chromosome mal-orientations that are similar to those operating during meiosis. However, unlike meiosis, where reorientation appears to be triggered when tension on a K-fiber is relieved or reduced, other factors related to the close proximity of sister kinetochores may also induce reorientation in mal-oriented mitotic chromosomes. We favor a model in which the reorientation of a mitotic kinetochore depends on, and is initiated by, the kinetochore capturing MTs from the pole to which it is reorienting.     

Chromosome micromanipulation

The relationship of kinetochore orientation and reorientation to orderly chromosome distribution in anaphase has been studied experimentally by micromanipulation of living grasshopper spermatocytes. Bivalents or the X chromosome at prometaphase or metaphase I can be detached from the spindle with a microneedle and moved to any desired location within the cell. Following a pause of variable duration the detached chromosome invariably moved, kinetochores foremost, back to the spindle, reassumed its characteristic metaphase position, and, with one exception, segregated normally at anaphase I. Detachment from the spindle is demonstrated unequivocally (1) by manipulation evidence for the absence of the firm spindle connections seen both before detachment and after reattachment and (2) by a functional criterion: a given kinetochore, oriented to one pole before detachment, often orients to the opposite pole after detachment. The segregation in anaphase was always as expected from the final, post-operation, orientation. Reorientation and prometaphase and anaphase movement after detachment cannot be distinguished from their counterparts in control cells. Kinetochore position after detachment is the primary determinant of the pole to which that kinetochore will orient. Therefore, since the experimenter determines kinetochore position, he can cause any given half-bivalent to segregate to a predetermined pole at anaphase I. Similarly, orientation of both half-bivalents to the same pole can be induced. These mal-oriented bivalents invariably reorient and normal anaphase segregation ensues. Non-disjunction can, however, be produced directly in late anaphase. These experiments are based upon current views of orderly chromosome distribution; their success confirms our understanding of the fundamental orientation process.     

The segregation pattern of a translocation quadrivalent

The segregation pattern of a translocation quadrivalent was studied in three hybrid families of the tetraploid (2n=28) oat, of the Avena strigosa polyploid complex. The rate of IV formation in F₁ was high and the fertility was normal. The adjacent-alternate orientation of this quadrivalent was very variable. Conspicuous variation in this frequency was found between anthers of the same floret and between florets. The alternate type was usually more common toward the end of MI. The adjacent-alternate ratio was found to be an unreliable measure for calculating the type of gametes produced by the F₁ and the F₂ plants derived from them. An attempt was made to determine the type of viable gametes produced on the F₁ and their frequency by examining the cytology of F₂ plants. The various combinations of the chromosomes of the translocation complex expected in the F₂ plants were derived and their expected frequencies calculated by taking into account chiasma formation at the chromosome ends and various restrictions of gamete viability. In none of the three hybrid families F₂ individuals were found to produce trivalents as the most complex chromosome configuration, indicating that one type of gamete derived from adjacent separation was not formed. The 11 ratio between F₂ plants having only bivalents (2II) and those with F ₁-like quadrivalent configuration R(c), expected when gametes resulting from alternate separation are fully functional, with the exclusion of other types, was not found. That ratio 2II/R(c), was 2-1/3 in the various families. The cytology of selected F₃ individuals basically followed the predictions based on chromosome association in the F₂.     

The distribution of male meiotic pairing sites on chromosome 2 of Drosophila melanogaster: meiotic pairing and segregation of 2-Y transpositions

The distribution of meiotic pairing sites on a Drosophila melanogaster autosome was studied by characterizing patterns of prophase pairing and anaphase segregation in males heterozygous for a number of 2-Y transpositions, collectively coveringall of chromosome arm 2R and one-fourth of chromosome arm 2L. It was found that all transpositions involving euchromatin from chromosome 2, even short stretches, increased the frequency of prophase I quadrivalents involving the sex and second chromosome bivalents above background levels. Quadrivalent frequencies were the same whether the males carried both elements of the transposition or just the Dp (2;Y) element along with two normal chromosome 2s, indicating that pairing is non-competitive. The frequency of quadrivalents was proportional to the size of the transposed region, suggesting that pairing sites are widely distributed on chromosome 2. Moreover, all but the smallest transpositions caused a detectable bias in the segregation ratio, in favor of alternate segregations, indicating that the prophase associations were effective in orienting centromeres to opposite poles. One transposition involving only heterochromatin of chromosome 2 had no effect on quadrivalent frequency, consistent with previous evidence that autosomal heterochromatin lacks meiotic pairing ability in males. One region at the base of chromosome arm 2L proved to be especially effective in stimulating quadrivalent formation and anaphase segregation, indicating the presence of a strong pairing site in this region. It is concluded that autosomal pairing in D. melanogaster males is based on general homology, despite the lack of homologous recombination.by A.C. Spradling     

Centromere Orientation of Quadrivalents of Heterozygous Translocations and an Autoploid of GOSSYPIUM HIRSUTUM L

Cytological observations of quadrivalents of heterozygous translocations in Gossypium hirsutum L. demonstrate that, in addition to alternate-1 and alternate-2 orientations, a third alternate orientation (alternate-3), which occurs as a three-dimensional, V-type configuration, can be identified.—Two additional types of disjunctions, the centromere orientations of which are rotational modifications of either adjacent or alternate configurations, were also observed in quadrivalents of a translocation heterozygote. These two types are rare, and both appear in the form of the Roman numeral X . The X and the alternate-3 types also occur in quadrivalents of an autoploid of G. hirsutum.—The two X types, along with adjacent-1, adjacent-2, alternate-1 and alternate-2 orientations, represent the six possible types of planar 2 x 2 random orientation of the four centromeres of a quadrivalent. Including the three-dimensional alternate-3 type, there are seven types of orientation.     

Meiotic behaviour of chromosomes 1R, 2R and 5R in autotetraploid rye

The meiotic behaviour of chromosomes 1R, 2R and 5R was studied in C-banded preparations of autotetraploid rye. Analysis of pairing and chiasma formation was based on metaphase I configurations, using the model designed by Sybenga, with slight modifications. Frequencies of two modes of pairing (one quadrivalent or two bivalents) differed from those expected for random pairing. Although preferential pairing for some arm pairs of chromosome 2R was detected, this did not seem to be the cause of the increased bivalent pairing. This increase was attributed to either the spatial separation of the four homologous chromosomes in some premeiotic cells into two groups of two, or a correction of the synaptonemal complex, or both. The number of chiasmate associations showed variation between chromosomes and between arms within the same chromosome. It was closely related to arm length, but different after quadrivalent and bivalent pairing. This is suggested to be a consequence of partner exchange interfering with pairing and, consequently, with chiasma formation, and a different chiasma distribution after quadrivalent pairing. Variation between chromosomes in the frequencies of alternate and adjacent co-orientation in metaphase I quadrivalents without interstitial chiasmata suggests that the relative positions of the centromeres in the quadrivalent influence their co-orientation.     

Modification of the properties of a translocation in the German cockroach by selection

Summary A stock of Blattella germanica bearing the interchange T(3; 12)/3;12 was subjected to close inbreeding with selection for random disjunction at metaphase I. After 3–4 generations of selection, interchange quadrivalent chiasma frequency decreased, variability in free bivalent chiasma frequency increased sharply, and individuals with either random or directed disjunction were present in the stock. Random disjunction was modified from a ratio of 2112 (adj.-1; alt.-1; adj.-2; alt.-2) to a ratio of 1111. After 7–8 generations of selection, chiasma frequency appeared to stabilize at lower than normal levels and variability decreased for both quadrivalents and free bivalents. Directed disjunction was modified from a ratio of 2114 to 1112, and no individuals with the original high level of directed disjunction were detected. Chains-of-four tended to orient randomly, especially in individuals where the ring quadrivalents showed directed disjunction. Relaxation of inbreeding, but not selection, produced an increase in chiasma frequency and variability in both free bivalents and quadrivalents, but the modified ratios for both random and directed disjunction were retained. These results are discussed with respect to inbreeding effects and genetic control of chiasma frequency and metaphase I disjunction in interchange quadrivalents.     

Effect of inbreeding and selection for vigour and fertility on meiotic behaviour in autotetraploid job's tears,Coix lacryma-jobi L.

Summary Meiotic behaviour of the colchicine-induced raw autotetraploid (4n = 40) Job's tears (Coix lacryma-jobi L.) was compared with that of the tetraploid envolved from it through selection for vigour and fertility over a 4 year period and selfing of the selected plants for 3 generations. A significant decrease in quadrivalent frequency and an increase in bivalent frequency per cell, greater frequency of ring quadrivalents, more cells with regular separation of chromosomes and fewer cells with laggards at anaphase I, fewer irregularities in meiosis II, fewer pollen quartets with micronuclei and fewer aneuploids in the progenies were found in evolved tetraploid. The average chiasma frequency per cell, per quadrivalent and per bivalent were more or less the same in both tetraploids. All these facts indicate that inbreeding and selection for vigour and fertility have brought about a shift towards regular meiosis in the evolved tetraploid. The increase in fertility during the period of selection was not, however, significant, suggesting that its response to selection is slow, that a number of factors are probably involved and that, besides multivalents, genic factors also govern sterility in the tetraploid.     

Cytogenetics of double cross hybrids between Pennisetum americanum — P. purpureum amphiploids and P. americanum x Pennisetum squamulatum interspecific hybrids

Summary Pearl millet, Pennisetum americanum L. Leeke-napiergrass, Pennisetum purpureum Schum. amphiploids (2n=42) were crossed with pearl millet X Pennisetum squamulatum Fresen. interspecific hybrids (2n=41) to study the potential of germplasm transfer from wild Pennisetum species to pearl millet. These two interspecific hybrids were highly cross-compatible and more than two thousand trispecific progenies were produced from 17 double crosses. All doublecross hybrids were perennial and showed a wide range of morphological variations intermediate to both parents in vegetative and inflorescence characteristics. Some crosses resulted in sublethal progenies. Chromosomes paired mainly as bivalents (¯x15.88) or remained as univalents. At metaphase I, trivalents, quadrivalents, an occasional hexavalent and a high frequency of bivalents indicated some homeology among the genomes of the three species. Delayed separation of bivalents, unequal segregation of multivalents, lagging chromosomes, and chromatin bridges were observed at anaphase I. Although approximately 93% of the double-cross hybrids were male-sterile, pollen stainability in male-fertile plants ranged up to 94%. Seed set ranged from 0 to 37 seed per inflorescence in 71 plants under open-pollinated conditions. Apomictic embryo sac development was observed in double-cross progenies when crosses involved a pearl millet x P. squamulatum apomictic hybrid as pollen parent. These new double-cross hybrids may serve as bridging hybrids to transfer genes controlling apomixis and other plant characteristics from the wild Pennisetum species to pearl millet.     

Meiotic pairing and segregation of translocation quadrivalents in yeast

Meiotic pairing and segregation were studied in three different heterozygous reciprocal translocation strains of the baker’s yeast, Saccharomyces cerevisiae. Pachytene translocation quadrivalents were identified by a combination of immunofluorescence and fluorescence in situ hybridization and the karyotypes of meiotic products were determined by pulsed-field gel electrophoresis. The translocations differed with respect to the relative sizes of the chromosomes involved and the positions of translocation breakpoints, and produced translocation quadrivalents of widely different shapes. This allowed us to study the influence of the morphology of quadrivalents on their segregation behaviour. In all cases alternate predominated over adjacent segregation. 3:1 disjunction of chromosomes was more frequent when translocation breakpoints were close to the centromeres. If a translocation breakpoint was distant from the centromere, the occurrence of an intervening chiasma influenced the pattern of segregation. In general, quadrivalent formation and segregation resembled the behaviour of translocation heterozygotes in most higher eukaryotes. We therefore conclude that, although chromosome condensation does not occur in yeast metaphase, centromere orientation and chromosome disjunction are governed in a way similar to that of higher eukaryotes. Received: 6 February 1998; in revised form: 19 May 1998 / Accepted: 23 May 1998     

Cytological studies on meiosis and male gametophyte development in autotetraploid cucumber

With improved staining and chromosome preparation techniques, meiosis of pollen mother cells (PMCs) and male gametophyte development in autotetraploid cucumber (Cucumis sativus L.) was studied to understand the correlation between chromosomes behaviour and fertility. Various chromosome configurations, e.g. multivalent, quadrivalents, trivalents, bivalents and univalents were observed in most PMCs at metaphase I. Lagging chromosomes were frequently observed at anaphase in both meiotic divisions. In addition, chromosomes segregations were not synchronous and equal in some PMCs during anaphase II and telophase II. Dyads, triads, tetrads with micronuclei and polyads were observed at tetrad stage, and the frequencies of normal tetrad with four microcytes were only 55.4 %. The frequency of abnormal behaviour in each stage of meiosis was counted, and the average value was 37.2 %. The normal meiotic process could be accomplished to form the microspore tetrads via simultaneous cytokinesis. Most microspores could develop into fertile gametophytes with 2 cells and 3 germ pores through the following stages: single-nucleus early stage, single-nucleus late stage and 2-celled stage. The frequency of abnormalities was low during the process of male gametophyte development. The germination rate of pollen grains was 46.9 %. These results suggested that abnormal meiosis in PMCs was the reason for low pollen fertility in the autotetraploid cucumber.     

Meiotic studies of a human male carrier of the common translocation, t(11;22), suggests postzygotic selection rather than preferential 3:1 MI segregation as the cause of liveborn offspring with an unbalanced translocation

The t(11;22)(q23;q11) translocation is the only non-Robertsonian rearrangement for which there are a large number of unrelated families, apparently with the same breakpoints. These families most often have been ascertained through an abnormal child with the karyotype 47,XX or XY, +der(22) t(11;22)(q23;q11). To explain the high incidence of 3:1 segregants, rarely seen in offspring of carriers of other reciprocal translocations, a number of theoretical models have been suggested. We have used both electron microscope analysis of the synaptonemal complex (SC) and dual-color FISH to investigate the meiotic chromosome behavior in a male carrier of the translocation who has the karyotype 46,XY, t(11;22)(q23;q11). Chromosome synapsis, first-meiotic chiasma configuration, and segregation behavior of this translocation have been analyzed directly. Examination of SCs by electron microscopy showed pachytene-cross formation in 49/50 nuclei. Approximately 50% (26/50) revealed a classical fully synapsed quadrivalent. A proportion of these (10/26), however, showed some central asymmetry, suggesting heterologous synapsis. The remaining cells appeared to have incomplete synapsis. FISH analysis showed only quadrivalents in all 100 metaphase I nuclei. The chiasma frequency was increased within the interstitial segments, in comparison with the same region in normal bivalents. All types of segregation category were found in metaphase II nuclei. There was no indication of preferential 3:1 anaphase I segregation. We conclude that the +der(22) constitution in offspring of carriers of t(11;22)(q23;q11) is not likely to be due to meiotic 3:1 segregation being especially common. Rather, the +der(22) constitution is more likely to be the result of postzygotic selection against other unbalanced karyotypes.     

Meiotic segregation of five different reciprocal translocations in the onion fly,Hylemya antiqua (Meigen)

For one translocation (T14) with short interstitial segments in Hylemya antiqua significant differences in segregation behaviour between males and females were observed. In males the ratio of alternate:adjacent 1:adjacent 2 was approximately 730 and in females about 813. This difference is attributed to the difference in type of chromosome association. Female meiosis is chiasmate and male meiosis is achiasmate. It is suggested that meiotic pairing in males results in relative short Coorientation Determing Distances (CDDs) between homologous centromeres which favours alternate and adjacent 1 segregation. In females because of non-localized chiasmata on the average no differences in CDD between homologous and nonhomologous centromeres are expected. This might explain the occurrence of coorientation between non-homologous centromeres resulting in adjacent 2 segregations. Four other translocations with longer interstitial segments than T14 showed in males as well as females predominantly an alternate and adjacent 1 segregation, adjacent 2 was hardly found (0–3.6%). The longer distance between non-homologous centromeres is probably the reason.     

Cytogenetic studies on <Emphasis Type="Italic">Metasequoia glyptostroboides</Emphasis>, a living fossil species

The chromosome morphology and meiotic pairing behavior in the pollen mother cells (PMCs) of Metasequoia glyptostroboides were investigated. The results showed that: (1) The chromosome number of the PMCs was 2n=22. (2) The PMCs developed in the successive manner, and the nucleoids in the dynamic development were similar to those of the other gymnosperms. (3) At prophase, most of the chromosomes were unable to be identified distinctively because the chromosomes were long and tangled together. The chromosome segments were paired non-synchronously. At pachytene, the interstitial or terminal regions of some bivalents did not form synapsis and the paired chromosomes showed difference in sizes, indicating that there were structure differences between the homologous chromosomes. (4) At diakinesis, the ring bivalents showed complicated configurations due to the differences in location and number of chiasmata. In addition, there were cross-linked bivalents. (5) At metaphase I, the chromosome configuration of each cell was 8.2II ⁰ + 1.1II + 1.3II ⁺ + 0.8I. Most of the chromosomes were ring bivalents, but some were cross-linked bivalents, rod bivalents, or univalents. (6) 15\% PMCs at anaphase I and 22\% PMCs at anaphase II presented chromosome bridges, chromosome fragments, micronuclei, and lagging chromosomes. Twenty seven percent microspores finally moved into one to three micronuclei. Twenty five percent pollens were abortive. The results indicated that the observed individual of M. glyptostroboideswas probably a parpcentric inversion heterozygote, and there were structural and behavioral differences between the homologous chromosomes. The chromosomal aberration of M. glyptostroboidesmay play an important role in the evolution of this relict species, which is known as a living fossil. Further evidence is needed to test whether the differences between homologous chromosomes were due to hybridization.     

The quantitative analysis of chromosome pairing and chiasma formation based on the relative frequencies of M I configurations

In autotetraploids, chromosome pairing may be in the form of quadrivalents or bivalent pairs. Whether or not the quadrivalents are maintained until first meiotic metaphase depends on the formation of chiasmata. The relative frequencies of M I configurations thus contain information both on pairing and on chiasma formation. With distal chiasma localisation six configurations can be recognised and their relative frequencies determined: ring quadrivalents, chain quadrivalents, trivalents (with univalent), ring bivalents, open (rod) bivalents, univalent pairs. These represent five degrees of freedom permitting five parameters to be estimated: the frequency (f) of quadrivalent pairing; the frequencies of chiasmate association of the two ends (arms in metacentrics), a′, b′, after quadrivalent pairing, and a, b after bivalent pairing. — The appropriate formulae have been derived and applied to observations on Tradescantia virginiana (4n=24) which has pronounced distal chiasma localisation. Slight modifications make the model applicable to autotetraploids with interstitial in addition to distal chiasmata. In T. virginiana, chromosome pairing appeared to be random between homologues (65.8% quadrivalent pairing; 55.4% observed at M I). After quadrivalent pairing chiasmate association is frequent in the “average long” arm (95.0%) and much less so in the other arm (60.5%). This is attributed to partner exchange. After bivalent pairing chiasma frequencies are still different for the two arms (93.8% and 83.5% association respectively) but much less pronounced. Various complications are discussed.     

Checkpoint control in crane-fly spermatocytes: unattached chromosomes induced by cytochalasin D or latrunculin treatment do not prevent or delay the start of anaphase

Summary Variable numbers of bivalents and sex chromosomes do not attach to the spindle when prophase or early prometaphase cranefly spermatocytes (2n=8) are treated with cytochalasin D or latrunculin. The unattached bivalents lie in the cytoplasm or at the spindle pole, and they do not delay onset of autosomal anaphase; sometimes they disjoin at the same time as the attached bivalents, so they respond to the global signals that initiate anaphase. Unattached sex chromosomes do not delay autosomal anaphase, either. Of various interpretations of these data, we think the best explanation is that the checkpoint system responds to physical rather than chemical cues; we think that the spindle is a tensegral structure, that chromosomes need to interact with the spindle in order to be recognised by the anaphase-onset checkpoint control, and that the physical interaction of chromosomes with spindle acts as a signalling network. Cytochalasin D and latrunculin treatments delay onset of sex chromosome anaphase (which normally occurs about 15 min after autosomal anaphase) and cause altered patterns of sex-chromosome segregation.