Comparative chiasma analysis using a computerised optical digitiser

A new computerised technique has been devised for measuring the distribution of chiasmata along diplotene bivalents. The method involves the introduction into the field of view of the microscope, of a fine light spot which can be accurately manipulated along the chromosomes of each bivalent. The data recorded include (a) the positions of the chiasmata along the bivalent in terms of their relative distances from the centromere and (b) the individual bivalent and cellular chiasma frequencies. — The method has been applied to the analysis of chiasma distribution patterns in the two known species of the genus Caledia, C. species nova 1 and C. captiva and in two chromosomal races of the latter. Statistical tests indicate that within bivalents at least 40% of the comparative distribution patterns of chiasmata between races and species are significantly different. Similar comparisons between populations within races reveal only 18% significant differences. — The observed distribution patterns of chiasmata in this genus suggest that chiasma formation is sequential from centromere to telomere. — The variation in the frequency and distribution of chiasmata between races and species suggests that the interference distances between successive chiasmata are, at least partially, independent of chiasma frequency and position. — The interracial and interspecific differences in chromosome structure are correlated with changes in chiasma pattern.     

Chromosome association and pollen fertility of parental and interspecific hybrids of Lycopersicon esculentum X L. hirsutum and L. pennellii

Cytological studies were carried out on two wild species (L. hirsutum and L. pennellii) and the cultivated species (L. esculentum) of tomato and their F1 hybrids. Both parents and hybrids show a diploid chromosome number of 2n=24. The meiotic behaviour of the cultivated species showed a high degree of chromosome homology resulting in a high level of chiasmata frequency per bivalent. In contrast, the two wild species showed a slight increase in uniyalent frequency and a decrease in bivalent formation and chiasmata frequency. The meiotic behaviour of the hybrids showed a high level of univalents and low levels of bivalents as well as trivalents. Highly significant decreases in chiasmata frequency and increases in meiotic abnormalities, especially in the L. esculentum X L. pennellii hybrid, also were detected. The high meiotic irregularity and low chiasmata frequency recorded in the second hybrid indicated the disharmony and difference between its parental genomes and also served to predict its sterility. With regard to degree of pairing recorded in the hybrids, there is a possibility that sterility in such cases may refer to genetic factors in addition to the previously mentioned reasons. Pollen fertility showed no great difference between L. esculentum and L. hirsutum and their F1 hybrid, but a significant decrease was recorded in the L. esculentum X L. pennellii hybrid, which was clearly associated with high meiotic irregularity, low chiasmata frequency and chromosome association.     

Chiasmata distribution in the normal karyotype of mice

The number of chiasmata per cell and variance of chiasmata numbers were studied, as well as the recombinational interaction between different bivalents in CBA/Lac mice male line. No competition of bivalents for chiasmata was discovered in mice; at the same time, the chiasmata within one arm of the chromosome interfere with each other. The number of chiasmata per bivalent is estimated for each chromosome independently. The number of chiasmata per chromosome is limited both from below (minimum one chiasma independently of its size) and from above (positive interference of chiasmata).     

Chiasma distribution in mouse oocytes during diakinesis

The distribution of chiasmata in the mouse was examined by measurement of a single metacentric bivalent in 173 oocytes taken from 36 mice of the Rb3Bnr stock. Frequency distribution analysis revealed a well defined pattern of chiasma formation in both arms of the metacentric and, as in other organisms, interference and localization were thought to be major factors influencing this pattern. Despite the tendency for bivalents to form terminal associations at metaphase in the mouse and reported differences in chiasma frequency between early and late stages of meiosis, analysis of bivalents at diakinesis has produced no quantitative support for the concept of terminalization of chiasmata during meiosis.     

The effect of colchicine on meiosis in Lilium speciosum cv. “Rosemede”

Chromosome pairing and chiasma frequency were studied in meiocytes at diakinesis of Lilium speciosum cv. Rosemede fixed up to 21 days after the start of either continuous or 3 day pulse colchicine treatment. The two treatments gave similar results. In pulse treated pollen mother cells (PMCs) the mean chiasma frequency per cell fell from 26.4 in controls to 8.5 after fourteen days while the mean number of univalents per cell increased from 0.05 to 17.58. There was a negative correlation between mean chiasma frequency per bivalent and per PMC in colchicine treated buds; univalents were preferentially induced in bivalents with one chiasma, and preferentially excluded in bivalents with 4 chiasmata. Some chiasmata were redistributed to surviving bivalents despite the concurrent reduction in chiasma frequency per meiocyte. — Colchicine sensitivity began in premeiotic interphase and extended to mid or late zygotene in PMCs; ongoing synapsis was unaffected. However, susceptibility to univalency was asynchronous between bivalents occurring at zygotene in short chromosomes but at late premeiotic interphase in the longest chromosomes. The number of chiasmata per bivalent could be altered by colchicine without inducing univalents, but the ultimate effect was to reduce the number of chiasmata per bivalent (or per chromosome arm) directly to zero. The major factors determining the order and extent of reduced pairing and chiasma number were total chromosome length and arm length. Pairing and chiasma formation in embryo sac mother cells were less sensitive to colchicine than in PMCs, but their behavior was otherwise similar.     

Double insertion of homogeneously staining regions in chromosome 1 of wild Mus musculus musculus: effects on chromosome pairing and recombination

An examination of the meiotic pattern of chromosome 1 isolated from a feral mouse population and containing a double insertion (Is) of homogeneously staining regions (HSRs) was carried out. The region delineated by the proximal breakpoint of Is(HSR;1C5) 1Icg and the distal breakpoint of Is(HSR;1E3)2Icg is desynapsed during the early pachytene stage and heterosynapsed at the midpachytene, as shown by electron microscopic analysis of synaptonemal complexes. The HSRs have no effect on the segregation of chromosome 1 in heterozygous mice. The lack of homosynapsis in the region under study causes chiasmata redistribution in heteromorphic bivalents. In normal males, single chiasmata are located in the medial part of the chromosome. In heterozygotes, this segment is heterosynapsed and unavailable for recombination. This leads to a significant decrease in the frequency of bivalents bearing single chiasmata. The total number of chiasmata per bivalent is much higher in heterozygous males than in normal ones. The recombination frequency between proximal markers fz and In also is higher in heterozygous animals. The increase in the total chiasma number in the heteromorphic bivalent is due to the addition of double chiasmata located mostly at precentromeric and pretelomeric regions of the chromosome.     

Induced Bivalent Interlocking and the Course of Meiotic Chromosome Synapsis

WHEN chromosomes pair at meiosis the bivalents so formed do not normally interlock. Heat-treatments can, however, induce bivalent interlocking in the locust Locusta migratoria. Only the longest bivalents interlock and usually only two are found per cell; two “rod” bivalents, with single chiasmata, two “ring” bivalents, each with two or three chiasmata, or one “rod” and one “ring” bivalent (Fig. 1a, b and c). The nature of this interlocking and the metaphase orientational and congressional properties of interlocked bivalents are analysed in detail elsewhere¹.     

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.     

Chiasma frequency,distribution and interference maps of mouse autosomes

Chiasma frequencies were analysed and chiasma positions measured in diakinesis/metaphase I autosomal bivalents from oocytes and spermatocytes of F₁ hybrid C3H/HeH×101/H mice. Twenty chromosome size ranks, including the presumptive X bivalent, could be distinguished in oocytes, and nineteen autosomal ranks plus the XY pair spermatocytes. Overall, mean cell chiasma frequencies of the two sexes did not differ significantly once the contribution of the presumptive X bivalent and the XY pair were taken into account. Sex related differences in chiasma distribution patterns were evident, however. In monochiasmate bivalents, the chiasma was most commonly located interstitially in oocytes while in spermatocytes it could be either interstitial or distal. In dichiasmate bivalents, the chiasmata tended to be more centrally located in oocytes than in spermatocytes. Minimum inter-chiasma distances did not appear to show any great variation in chromosome pairs of different sizes, however, mean inter-chiasma distances did increase with the bivalent length. The minimum-inter chiasma distance data suggest that chiasma interference is complete over a chromosomal segment equating to approximately 60 Mb. Measurement of the positions of chiasmata along chromosome arms open up the possibility of producing chiasma-based genetic maps for all the autosomes of the mouse.     

Lampbrush Chromosomes in the Japanese Quail Coturnix coturnix japonica: Cytological Map of Macrochromosomes and Meiotic Crossing-over Frequency in Females

Cytological map of lampbrush macrobivalents of the Japanese quail (Coturnix coturnix japonica) were constructed. Investigation of chiasmata allowed to estimate the frequency of reciprocal genetic recombination (crossing over) in Japanese quail female meiosis. The total chiasma number in bivalents of Japanese quail oocyte nuclei was determined to be 53–58. Macrobivalents 1–5 and Z of the Japanese quail had on average 3.3 chiasmata per bivalent, and microbivalents, 1.0–1.1 chiasmata per bivalent. The chiasmata (crossover) frequency in Japanese quail females was lower than in chicken. In macrochromosomes of Japanese quail females, one crossover occurred per 43.9 Mb, and in chicken, per 30.0 Mb. Judging from chiasma frequency, the genetic length of the Japanese quail genome is likely to be 2650–2900 cM. Crossover frequency in the species was 0.023 per Mb in macrobivalents and 0.07–0.08 Mb in microbivalents and for the total genome, 0.041 crossing over per Mb. The genetic length of one Mb (recombination rate ) in female Japanese quails was 1.14 cM in macrochromosomes, 3.60–4.12 cM in microchromosomes, and about 1.96–2.15 cM averaged over the genome.     

Chromosome-specific desynapsis in the n = 2 race of Haplopappus gracilis (Compositae)

During cytological screening for pollen sterility in a wild population of Haplopappus gracilis (n = 2), several partially sterile plants were found that had good pachytene pairing but varying numbers of univalents. Some plants had chromosome A bivalents or A univalents, while in the same cells chromosome B had only bivalents. In other plants the reverse condition occurred; the B chromosome had B bivalents or B univalents and only A bivalents. This demonstrates a chromosome-specific effect for the desynapsis genes. Hybridization between the two homozygous mutant genotypes produced only normal bivalents; this indicates the two mutants are not alleles and each is recessive. An F2 generation showed independent assortment of the desynaptic mutations. The chromosome A bivalent is the larger of the two and normally has one or two chiasmata; the B bivalent normally has a single chiasma. Chiasmata distribution was tested in the desynaptic mutant A bivalents and showed an acceptable fit to a binomial distribution. This occurs also in heterozygous, asynaptic pairing control gene mutations. Analysis of the NOR bivalent in two hologenomic desynaptic mutations in tomato also showed a good fit to a binomial distribution of chiasmata. This indicates the same methods are applicable to diverse species.     

Relationship between pachytene synapsis, metaphase I associations, and transmission of 2B and 4B chromosomes in rye.

2B rye plants selected for high (H) or low (L) B transmission rate were studied at pachytene and metaphase I of meiosis to determine the relationship between synapsis, bivalents at metaphase I, and B transmission rate. The results show that the 2 B chromosomes (Bs) form bivalents at pachytene in both the H and L lines, whereas the frequency of bivalents at metaphase I is much higher in the H than in the L line. This demonstrates that B transmission is mainly related to the proper association of Bs at metaphase I, as well as that synapsis of the 2 Bs in the L line is normal, but the bivalent is not consolidated by a chiasma in most cases. Crosses were made between 2B plants of the H and L lines in all combinations (H x H, H x L, L x H, and L x L) to obtain 4B plants. Similarly, bivalent formation at pachytene and metaphase I was studied. The results show that 4B plants of the H x H and L x L classes differ significantly at pachytene and metaphase I since the former forms more bivalents. The heterozygous 4 Bs of the H x L and L x H classes show intermediate values. The relation H x H > H x L > L x H > L x L was consistently found for the variables transmission rate, bivalents at pachytene, bivalents at metaphase I, and B mean chiasma frequency. A maternal effect was also found. Our data suggest that there are two separate mechanisms acting upon synapsis and chiasma formation in H and L B chromosomes: (i) there is variable efficiency of the control of synapsis at early stages of meiosis; and (ii) there is variable efficiency of the control of the number of chiasmata.     

Some cytologic aspects of meiosis in female guinea pig

Cytogenetic preparations from young, adult, random bred female guinea pig oocytes were made on days 5–8 of vaginal cycles with an in vitro technique. First meiotic metaphase was harvested at 8 hours and second meiotic metaphase at 14 hours. The mean chiasmata count per bivalent was 1.035. A microscopic appearance suggesting lampbrushing was seen in some metaphase I bivalents.     

Male and female meiosis in grasshoppers II. Chorthippus brunneus

The frequencies and distribution patterns of chiasmata in the autosomal bivalents of Chorthippus brunneus are very similar in male and female meiosis. The mean cell chiasma frequency of oocytes is however significantly higher than that of spermatocytes, but this difference is almost certainly due to the incremental effect of the X bivalent on the chiasma frequencies of oocytes. The implications of these findings in terms of chiasma control are briefly discussed.     

Lampbrush chromosomes and chiasmata of sex-reversed crested newts

Triturus cristatus carnifex provides a particularly clear example of sexual dimorphism for chiasma frequency and localisation. Oocytes from normal XX females routinely carry one proximal chiasma on each arm of their lampbrush bivalents. Spermatocytes from normal XY males have more numerous and relatively distal chiasmata. Lampbrush chromosomes from the oocytes of sex-reversed XY neofemales are found to resemble those from normal oocytes in having one proximal chiasma on each bivalent arm. A comparison of particular markers on the heteromorphic long arm of chromosome 1 provides evidence to equate the lampbrush 1A to somatic 1A, and confirms previous reports that lampbrush chromosome 1A is slightly longer than 1B. The XY sex bivalent of neofemales does not show any obvious heteromorphy of recognised marker loops. Received: 9 September 1997 / Accepted: 16 October 1997     

Absence of chiasmata from the heteromorphic region of chromosome I during spermatogenesis in Triturus cristatus carnifex

Analysis of squash preparations of spermatocytes from crested newts, Triturus cristatus carnifex, has shown that in most cells at least one large bivalent regularly fails to form chiasmata in one arm-pair. Feulgen microphotometry of diplotene and metaphase bivalents has shown that it is the largest bivalent in each cell which shows chiasma failure in one arm-pair. A C-banding technique which identifies chromosome I by virtue of a long, darkly stained region in its long arm, was used to confirm the absence of chiasmata from one arm-pair of the longest bivalent, and specifically from the darkly stained region. The achiasmate region which chromosome I exhibits during spermatogenesis, corresponds to the heteromorphic region of oocyte lampbrush bivalent I in which chiasmata never form. A possible correlation between the complete absence of crossing-over from the heteromorphic region and unusual cytological and molecular features which it exhibits, are discussed.     

Lampbrush chromosomes in the japanese quail Coturnix coturnix japonica: cytological maps of macro chromosomes and meiotic crossover frequency in females

Cytological maps of lampbrush macrobivalents of the Japanese quail (Coturnix coturnix japonica) were constructed. Investigation of chiasmata allowed determination of the meiotic frequency of reciprocal genetic recombination (crossing over) in Japanese quail females. The total chiasma number in bivalents of Japanese quail oocyte nuclei was determined to be 53-58. Macrobivalents 1-5 and Z of the Japanese quail had on average 3.3 chiasmata per bivalent, and microbivalents, 1.0-1.1 chiasmata per bivalent. The chiasmata (crossover) frequency in Japanese quail females was lower than in chicks. In macrochromosomes of Japanese quail females, one crossover occurred per 43.9 Mb, and in chicken, per 30.0 Mb. Judging from chiasma frequency, the genetic length of the Japanese quail genome is likely to be 2650-2900 cM. Crossover frequency in the species was 0.023 per Mb in macrobivalents and 0.07-0.08 Mb in microbivalents and for the total genome, 0.041 crossovers per Mb. The genetic length of one Mb (theta) in female Japanese quails was 1.14 cM in macrochromosomes, 3.60-4.12 cM in microchromosomes, and about 1.96-2.15 cM averaged over the genome.     

Localised chiasmata due to partial pairing: A 3D reconstruction of synaptonemal complexes in male Stethophyma grossum

The possible role of localised pairing as a mechanism producing localised chiasmata in Stethophyma grossum spermatocytes has been examined ultrastructurally. Nuclei at four successive stages of meiosis from leptotene to pachytene were reconstructed from a series of ultrathin sections and the extent of synapsis as demonstrated by synaptonemal complex (SC) formation was calculated. On the basis of the relative lengths of SCs and condensed chromosomes it was reasoned that only the centromeric ends of the long and medium length bivalents paired, and only one end of these SCs was found attached to the nuclear envelope. Only the three shortest bivalents paired completely, and both their SC ends were attached to the nuclear envelope. Thus pairing was directly related to the distribution of chiasmata. The extent of pairing at different stages suggests that the shortest bivalent paired very quickly, the longer ones progressively slower, and that pairing proceeded zip-like from a point at or very close to the end attached to the nuclear envelope, since all stretches of SC were attached to the envelope, and there were never more than 11 pieces, one for each bivalent. Chromosome decondensation and axial core formation did not occur far in advance of SC formation, and synapsis appeared to be much slower in S. grossum than in other species with non-localised chiasmata, as a larger proportion of the meiotic cysts were in zygotene, compared to Stauroderus scalaris and Locusta migratoria, although this was not quantified.     

Meiosis VII: “Detorsive bending” as a basis for geometric shapes of late prophase bivalents

A new model depending on mechanical properties of chromosomes is adduced as a basis for diplotene opening-out and for curvature occurring in grasshopper bivalents, during and subsequent to diplotene. Conditions underlying the model are: (1) rigid physical binding exists between sister chromatids, (2) each chromatid remains free of torsional strain if its pairing face is straight, i.e. the chromatid is bilaterally symmetrical, (3) reciprocal exchange together with stiff binding between sisters produces twist in each chromated before diplotene begins, (4) stiffening of the bivalent during late meiotic prophase removes the twist resulting from reciprocal exchange, (5) since sister binding prevents untwisting of chromatids about their long axes, untwisting would be achieved only in conjunction withbending of each chromated. It is shown that this bending, called detorsive bending, automatically produces opening out, not only in bivalents with one chiasma but also in those with more than one, especially if the chiasmata are interstitial.In bivalonts with two chiasmata, classes of curvature resulting when both chiasmata are interstitial (II), when one is interstitial and one terminal (IT) and when both are terminal (TT) are attributed to differences in strength of opening out at interstitial and at terminal chiasmata respectively. It is postulated that mechanisms responsible for opening out at terminal chiasmata are basically different from those at interstitial chiasmata.A theoretical basis of a method for cytological detection of chromatid interference is outlined and arguments are presented against the electrostatic hypothesis.     

Myths and mechanisms of meiosis

A comparative analysis of the meiotic secquence in a wide variety of organisms indicates there is no convincing evidence that: (1) Premeiotic pairing plays any role in the synapsis of homologues. (2) Heterochromatic association facilitates homologous pairing. (3) Chiasmata ever form within segments which are positively heteropycnotic at zygotenepachytene. (4) Localisation of chiasmata depends on prior localisation of pairing or on the occurrence of euchromatin-heterochromatin boundaries. (5) Prior association of centromeres plays any role in determining co-orientation. (6) Any form of supra-chromosomal organisation exists involving permanent association between the members of a haploid complement, and (7) Unequal progeny ratios recovered from structurally modified Drosophila complements arise as a consequence of distributive pairing. — On the other hand there is good evidence that: (1) Interlocking of bivalents can occur regularly in species with a chiasma frequency sufficiently high to regularly produce ring bivalents and in which the chiasmata are localised to the ends of the bivalent. (2) Some forms of terminal association cannot represent terminalised chiasmata. (3) U-type exchanges present at diplotene result from errors in crossing over. (4) Pairing and chiasma formation are not necessary for coorientation, and (5) at least some types of elastic constrictions present at first metaphase represent extended nucleolar organisers.In memory of the late Stanley G. Smith and his signal contribution to the science of cytogenetics.