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.     

Metaphase I orientation of chain-forming interchange quadrivalents: a theoretical consideration

The orientation behavior of chain forming interchange quadrivalents at metaphase I was studied in three interchange heterozygotes of pearl millet [Pennisetum americanum (L.) Leeke] which involve chromosomes 1, 3, 6 and 7 in various combinations. Of these, two combinations predominantly produced rings and the third was a chain-forming type. The chain quadrivalents derived from the two ring-forming interchanges, as well as the chain quadrivalent generated by the third interchange, all showed one adjacent orientation at metaphase I (adjacent-1 or -2, depending upon the formation or failure of chiasmata and their positions in the different segments of the pachytene cross). Homologous centromere co-orientation leading to adjacent-1 and alternate-1 occurs following chiasma failure in the noncentric arms of the pachytene cross, and nonhomologous centromere co-orientation leading to adjacent-2 and alternate-2 occurs following chiasma failure in the centric arms of the pachytene cross. Thus, it has been proposed that, unlike in ring quadrivalents, a specific chain quadrivalent will have only homologous or nonhomologous centromere co-orientations at metaphase I.     

Chiasma interference and centromere co-orientation in a spontaneous translocation heterozygote of Euchorthippus pulvinatus gallicus (Acrididae; Orthoptera)

The meiosis of an individual of the species Euchorthippus pulvinatus gallicus heterozygous for a reciprocal translocation involving chromosomes 3 and 6 has been analysed using the Giemsa C-banding technique. It is concluded that: (i) Chiasma interference in the quadrivalent seems to act only at the arm level. There is no interference across the translocation break point. No interchromosomal chiasma interference could be demonstrated, (ii) The results concerning the co-orientation of the quadrivalent suggest that the length of the chromosomal segments between two adjacent centromeres at metaphase I is related with their orientation behaviour.     

Pairing competition between identical and homologous chromosomes in autotetraploid rye heterozygous for interstitial C-bands

Pairing competition between identical and homologous non-identical chromosomes is analysed in autotetraploid metaphase I cells of rye where one pair of identical partners bears an interstitial C-band in the long arm of chromosome 6R whereas the other pair of identical partners lacks such a C-band. This makes it possible to study pairing preferences in two distinct regions of the same chromosome arm. A significant excess of associations involving homologous partners is always observed in the proximal segment (from the centromere to the C-band), whereas a good fit with the expected random ratio, or else identical pairing preferences, is found in the distal segment (from the C-band to the telomere). Differences in the processes of pairing and chiasma formation in 6RL, and/or a readjustment in the pattern of chiasma distribution due to heterozygosity for the interstitial C-band as a result of homologous nonidentical pairing, may be responsible for the different behaviour detected in the two regions of the marked arm.     

Estimating meiotic chromosome pairing and recombination parameters in telocentric trisomics.

Telocentric trisomics (telotrisomics; one arm of a metacentric chromosome present in addition to two complete genomes) are used in theoretical studies of pairing affinities and chiasma formation in competitive situations and applied in genome analysis, gene localization, gene transfer, and breakage of close linkages. These applications require knowledge of the recombination characteristics of telotrisomics. Appropriate cytological and molecular markers and favorable chromosome morphology are not always available or applicable for quantitative analyses. We developed new mathematical models for extracting the maximum information from simple metaphase I observations. Two types of telotrisomics of the short arm of chromosome 1R of rye (Secale cereale), including several genotypes, were used as test material. In simple telotrisomics, pairing between morphologically identical complete chromosomes was more frequent than pairing between the telocentric and either of the normal chromosomes. In the telocentric substitution, morphologically identical telocentrics paired less frequently with each other than either one with the normal chromosome. Pairing partner switch was significant. Interaction between the two arms was variable. Variation within plants was considerable. Telotrisomics without markers are suitable for analyzing pairing preferences, for gene localization and gene transfer, and for breaking tight linkages, but less so for genome analysis.     

Pachytene pairing and metaphase I configurations in a tetraploid somatic Lycopersicon esculentum x L. peruvianum hybrid.

In the tetraploid somatic hybrid between the diploid Lycopersicon species L. esculentum (tomato) and L. peruvianum, synaptonemal complexes formed quadrivalents in 73 of the 120 sets of four chromosomes (60.8%) in 10 cells studied in detail at pachytene. Of these, 43 had one pairing partner exchange, 22 had two, and 8 had three, very close to a Poisson distribution. The points of pairing partner exchange were concentrated at the middle of the two arms. The frequency per arm corresponded with physical arm length. There was a sharp drop around the centromere, and pericentric heterochromatin had a slightly lower probability of being involved in pairing partner exchange than euchromatin. The chromosomes align before pairing and there are several points of pairing initiation, with concentrations at or near the ends and the centromere. From zygotene to late pachytene the quadrivalent frequency decreased considerably. At late pachytene it was lower than expected with the observed high frequency of pairing partner exchange. Pairing affinity between species was only slightly lower than affinity within species, in spite of considerable genetic differentiation. The frequency of recombination nodules increased from early to late zygotene and then decreased strongly to full pachytene. There is a highly significant negative correlation between percent pairing and SC length. At metaphase I the frequency of quadrivalents was 0.444, and branched quadrivalents were rare, probably caused by interference and restriction of chiasma formation to distal euchromatin. Metaphase I quadrivalent frequency is a relatively good indication of pairing affinity in this material.     

Interaction between wheat chromosomes and rye telomeric heterochromatin on meiotic pairing of chromosome pair 1R of rye in wheat-rye derivatives

The effect of telomere heterochromatin on metaphase I association of chromosome pair 1R of rye was analyzed in normal diploid plants of rye (2n=14) and in wheat-rye derivatives with the chromosome constitution (0–7)A(0–7)BRR (2n=20, 21 and 22). The C-banding pattern of 1R was variable between plants. In diploid rye the presence or absence of telomeric heterochromatin in 1R does not influence its meiotic pairing. However, in wheat-rye derivatives the presence of telomeric heterochromatin decreases chiasma frequency in the 1R bivalent. This cannot be attributed to interference of heterochromatin with chiasma terminalization. This effect of heterochromatin is most pronounced in heterozygous condition. In plants heterozygous for telomeric C-bands the reduction of pairing is stronger in the short arm than in the long arm of the 1R bivalent.     

On the influence of decreased chiasma frequency on preferential MI pairing behaviour of rye chromosomes in wheat-rye derivatives

Meiotic behaviour of identical and homologous rye chromosomes was investigated in colchicine-induced duplicated meiocytes obtained from different wheat-rye derivatives. A great reduction in the amount of metaphase I (MI) associations accompanied by a strong tendency for identical over homologous nonidentical preferential MI pairing was found in all of the four rye chromosome arms analysed. Both of these features appear to be associated with a more distal chiasma localization where the presence of an interstitial C-band has allowed two distinct regions within the same chromosome arm to be studied separately. On the other hand, the MI pairing failure observed for the rye chromosomes under analysis does not seem to be an effect of telomeric or interstitial C-heterochromatin.by P.B. Moens     

Chiasma formation in the short arm of the nucleolar chromosome of the tomato

Summary A translocation heterozygote in tomato (Lycopersicon esculentum) is shown to have a cyclical type of interchange between the long arms of chromosomes 1, 2 (nucleolar) and 3. A study of chromosome association in this plant at metaphase I has indicated that in 21% of the cells a ring of six chromosomes is present. Since an open ring hexavalent can occur only if there is chiasma formation in all the translocated segments and in all the short arms of the three chromosomes, it is concluded that there is considerable frequency of chiasma formation in the short arm of the nucleolar chromosome. This conclusion contradicts the previous observations that chiasma formation is either absent or very rare in the entirely dark staining chromatic, sometimes referred to as heterochromatic, short arm of the nucleolar chromosome.Part of this investigation was carried out at the Department of Genetics, Agricultural University, Wageningen, when the author was serving a contract between the EURATOM-I.T.A.L. and the Agricultural University.     

Chromosome pairing in the allotetraploid Aegilops biuncialis and a triploid intergeneric hybrid.

Chromosome pairing behaviour of the natural allotetraploid Aegilops biuncialis (genome UUMM) and a triploid hybrid Ae. biuncialis x Secale cereale (genome UMR) was analyzed by electron microscopy in surface-spread prophase I nuclei. Synaptonemal-complex analysis at zygotene and pachytene revealed that synapsis in the allotetraploid was mostly between homologous chromosomes, although a few quadrivalents were also formed. Only homologous bivalents were observed at metaphase I. In contrast, homoeologous and heterologous chromosome associations were common at prophase I and metaphase I of the triploid hybrid. It is concluded that the mechanism controlling bivalent formation in Ae. biuncialis acts mainly at zygotene by restricting pairing to homologous chromosomes, but also acts at pachytene by preventing chiasma formation in the homoeologous associations. In the hybrid the mechanism fails at both stages. Key words : Aegilops biuncialis, allotetraploid, intergeneric hybrid, pairing control, synaptonemal complex.     

Meiosis in allopolyploid Crepis capillaris. II. Autotetraploids.

Meiotic chromosome pairing of autotetraploid Crepis capillaris was analysed by electron microscopy of surface-spread prophase I nuclei and compared with light microscopic observations of metaphase I chromosome configurations. Prophase I quadrivalent frequencies are high in all three tetrasomes. (A, D, and C) and partially dependent on chromosome size. At metaphase I quadrivalent frequencies are much lower and strongly dependent on chromosome size. There is no evidence for multivalent elimination during prophase I in this system, and the reduction in multivalent frequency at metaphase I can be explained by an insufficiency of appropriately placed chiasmata. The high frequencies of prophase I quadrivalents far exceed the two-thirds expected on a simple model with two terminal independent pairing initiation sites per tetrasome, suggesting that multiple pairing initiation occurs. Direct observations reveal relatively high frequencies of pairing partner switches (PPSs) at prophase I, which confirms this suggestion. The numbers of PPSs per tetrasome show a good fit to the Poisson distribution, and their positional distribution along chromosomes is random and nonlocalized. These observations favour a model of pairing initiation based on a large number of evenly distributed autonomous pairing sites each with a uniform and low probability of generating a PPS.     

Non-homologous chromosome pairing and crossover formation in haploid rice meiosis

While many studies have provided significant insight into homolog pairing during meiosis, information on non-homologous pairing is much less abundant. In the present study, fluorescence in situ hybridization (FISH) was used to investigate non-homologous pairing in haploid rice during meiosis. At pachytene, non-homologous chromosomes paired and formed synaptonemal complexes. FISH analysis data indicated that chromosome pairing could be grouped into three major types: (1) single chromosome paired fold-back as the univalent structure, (2) two non-homologous chromosomes paired as the bivalent structure, and (3) three or more non-homologous chromosomes paired as the multivalent structure. In the survey of 70 cells, 65 contained univalents, 45 contained bivalents, and 49 contained multivalent. Moreover, chromosomes 9 and 10 as well as chromosomes 11 and 12 formed non-homologous bivalents at a higher frequency than the other chromosomes. However, chiasma was always detected in the bivalent only between chromosomes 11 and 12 at diakinesis or metaphase I, indicating the pairing between these two chromosomes leads non-homologous recombination during meiosis. The synaptonemal complex formation between non-homologs was further proved by immunodetection of RCE8, PAIR2, and ZEP1. Especially, ZEP1 only loaded onto the paired chromosomes other than the un-paired chromosomes at pachytene in haploid.     

Chiasma formation in duplicated segments of the haploid rye genome

In meiosis of haploid rye associations of two or more chromosomes are observed. In order to investigate whether these associations are chiasmate, metaphase I and anaphase I associations were analysed after Giemsa banding. — At anaphase I chromatid exchanges between differently marked chromosome arms were observed, which proved the presence of real chiasmata. The association between banded and unbanded arms shows that the heterochromatic telomeres do not act as secondary pairing sources. Different statistical approaches were used to test randomness of chiasma formation. It appeared to be non-random, which showed that the segments involved were non-randomly located and probably limited in number. The nature of these segments is discussed.     

Unexpected behavior of an inverted rye chromosome arm in wheat

Distal location of chiasmata in chromosome arms is thought to be a consequence of the distal initiation of synapsis. Observations of meiotic behavior of a rye chromosome with an inverted arm show that patterns of chiasma distribution and frequency are also inverted; therefore, the patterns of synapsis and chiasma distribution are independent, and recombination frequency along a chromosome is position-independent and segment-specific. Since cases of random distribution of chiasmata and recombination are known in rye, a genetic mechanism must be present that licenses specific chromosome regions for recombination. Large differences in the metaphase I pairing of the inversion in various combinations of two armed and telocentric chromosomes confirm the major role of the telomere bouquet in early homologue recognition. However, occasional synapsis and chiasmate pairing of the distal regions of normal arms with the proximal regions of the inversion suggest that an alternative mechanism for juxtaposing of homologues must also be present. Synapsis in inversion heterozygotes was mostly complete but in the antiparallel orientation, hence defying homology, but non-homologues never synapsed. Instances of synapsis strictly limited to the chiasma-capable segments of the arm suggest that, in rye, both recombination-dependent and recombination-independent mechanisms for homologue recognition must be present.     

Chromosomal diversity and morphological dimorphism in Moroccan wild oat, Avena agadiriana

Chromosomal diversity and morphological dimorphism were examined by breeding the intraspecific hybrids among six accessions of Avena agadiriana Baum et Fedak. The accession M55 occasionally showed a quadrivalent, which indicated that it was heterogeneous for a reciprocal translocation. The chromosome pairings in the intra-early-flowering ecotype hybrids were almost normal with high chiasma frequency at metaphase I. However, the intra-late-flowering ecotype hybrids showed a quadrivalent or a trivalent with a univalent, similar to the inter-ecotype hybrids. The marginal population of the northeastern end, M74, always showed a quadrivalent or a trivalent in all the hybrids due to a unique large reciprocal translocation. This result was consistent with the late-flowering ecotype. There was a genocline of the chiasma frequency resulting from chromosomal rearrangements between adjacent populations. There was also a significant negative correlation between the mean chiasma frequency and the annual rainfall in each collection site (r = ?0.880). The two distinct ecotypes were characterized by the different magnitude and number of chromosomal rearrangements, reciprocal translocation and loss of satellite chromosomes, and they were adapted to the wide range of environments along the Atlantic coast of Morocco.     

The relation between pairing preference and chiasma frequency in tetrasomics of rye.

The association pattern of marked tetrasomes of Secale chromosome 1R at meiotic first metaphase was analyzed. Two of the four chromosomes were identical with terminal C-bands at both arms; the other two were also identical but lacked C-bands and were homologous or homeologous with the first two. Four different types of heterozygotes for 1R were studied: (i). autotetraploid hybrids between genetic variants within Secale cereale subsp. cereale, (ii). tetraploid hybrids between subspecies of Secale cereale, (iii). tetraploid hybrids between species of Secale, and (iv). autotetrasomes of S. cereale in a wheat background. Earlier observations that heterozygous associations (banded with unbanded) had consistently higher chiasma frequencies than homozygous associations were extended and confirmed. To analyze this phenomenon more closely, the possible relations between this correlation and several other meiotic phenomena were studied. For this analysis, three genetically different autotetraploid hybrids within S. cereale were selected that differed with respect to the relation between pairing type and chiasma frequency. Special attention was given to different patterns of interference and other meiotic phenomena in the two chromosome arms of chromosome 1R. No relations between such phenomena and the relation between pairing type and chiasma frequency could be established. A hypothesis is formulated assuming that long-distance homologue attraction is concentrated in a limited number of sites and that in different genotypes, different patterns of active sites are present. Moderately weak attraction sites can pair with strong homologous sites under favorable genetic conditions, but two weak sites cannot. Then, heterozygotes have more effective pairing initiation and consequently chiasma formation than homozygotes. Under less favorable conditions, only strong sites are effective, and then, homozygotes pair better, but the chiasma frequency is lower. A model of the forces involved in homologue attraction is presented.     

Chiasma frequency effects of structural chromosome change

Three structural chromosome changes in the plant Hypochoeris radicata 2n = 8 have been tested for their effects on chiasma formation: (1) centric fission of chromosome 1, (2) a whole arm exchange between chromosomes 1 and 3, and (3) an interchange between the long arm of chromosome 1 and the short arm of 2 which gives an effectively three-armed pachytene multiple. Mean chiasma frequencies were compared between full-sibs in families segregating for the rearrangements. In each family the chiasma frequency was higher in heterozygotes than basic homozygotes. The size of the chiasma increase is dependant on the number of additional potentially-paired segments in the complement at pachytene. Fission heterozygotes and 1/2 interchange heterozygotes, with one extra pairing region, both form about 0.45 more chiasmata per PMC than full-sib basic homozygotes. The 1/3 exchange, with two additional pairing regions, increases chiasma frequency by twice this, about 0.85 per PMC. Individuals homozygous for the centric fission maintain the raised chiasma level. The chiasma increase appears limited to the chromosome(s) affected by structural change with no detectable interchromosomal effect.     

Chiasma patterns in a translocation derived duplication heterozygote of rye

Relative multivalent and bivalent configuration frequencies at first meiotic metaphase of a translocation derived duplication heterozygote of rye have been used to study recombination (chiasma) patterns. After multivalent pairing chiasma frequency is greatly reduced in the segments proximal to the duplication even when no chiasma is formed. There is positive interference after multivalent pairing between the duplication and the two adjacent segments, possibly especially the interstitial segment of the donor chromosome. A variegated type of across-centromere interference is inferred for both chromosomes. The duplication can in principle be applied in a hybrid variety using chromosomal male sterility genes. The restriction of recombination is not as effective as claimed for some other systems working with excess chromosomal material.     

Investigation of Homologous Crossing over and Sister Chromatid Exchange in the Wheat Nor-B2 Locus Coding for Rrna and Gli-B2 Locus Coding for Gliadins

Recombination was investigated within the Nor-B2 locus of wheat chromosome 6B that contains several thousand of the 18S-5.8S-26S rRNA (rDNA) repeated units. Additionally, recombination was assessed for several chromosome regions, in arm 6Bq between the centromere and the B2 locus (awn suppressor) and in arm 6Bp between the centromere and Nor-B2, between Nor-B2 and a distal C-band and between Nor-B2 and Gli-B2 coding for gliadins. The experimental design permitted the distinction between crossing over between homologous chromosomes and exchange between sister chromatids. No homologous crossing over within the Nor-B2 locus was found in a sample of 446 chromosomes, but one exchange with the attributes of unequal sister chromatid exchange was identified. The molecular characteristics of this presumed sister chromatid exchange indicate that the spacer variants present in the Nor-B2 locus are clustered. No homologous recombination was detected within the distal Gli-B2 locus containing repeated genes coding for gliadin seed-storage proteins. Both arms of chromosome 6B showed low crossing-over frequency in the proximal regions. The distance from the centromere to Nor-B2 was only from 0.3 to 2.2 cM although it accounts for about two-thirds of the metaphase chromosome arm, which shows a great distortion of the metaphase map of the arm. The level of homologous recombination within the Nor-B2 locus is lower than in the chromosome region immediately distal to it. Whether it is comparable to that in the chromosome region proximal to it could not be determined. Recombination frequencies of different pairs of chromosome 6B in all but one interval paralleled the frequencies of their metaphase I pairing: Lower pairing at metaphase I was paralleled by lower crossing-over frequency. This relationship indicated that reduced metaphase I pairing between 6B chromosomes from different populations is due to impaired crossing-over and not due to precocious chiasma terminalization.     

Chromosome structure and pairing preferences in autotetraploid rye (Secale cereale).

The influence of chromosome structure upon pairing behaviour during meiosis was investigated by comparing four autotetraploid genotypes of rye (Secale cereale) containing homologous chromosome sets with different degrees of structural similarity. The series provided a range of genotypes that, at one extreme, contained structurally identical chromosome sets and, at the other extreme, sets that are certainly more heterozygous in the genic sense and probably also more diverse from a purely structural viewpoint. Relative frequencies of pairing configurations at meiotic prophase and metaphase I were compared by electron microscopy of whole-mount surface-spread synaptonemal complex complements and light microscopy of squash preparations. Despite unexpectedly low quadrivalent frequencies over all four genotypes, higher mean bivalent frequencies appeared to be associated with greater homologue diversity. In other words, greater structural divergence between chromosome sets appears to facilitate more efficient discrimination between homologous and identical chromosomes that drives the formation of bivalents. Statistical comparisons were not able to confirm in some cases the significance of the observed pattern of pairing behaviour.