Meiosis in Holocentric Chromosomes: Orientation and Segregation of an Autosome and Sex Chromosomes in Triatoma infestans (Heteroptera)

The meiotic behaviour of the X chromosome and one autosomal pair of the heteropteran Triatoma infestans was analysed by means of C-banding plus DAPI staining. At first metaphase, the X univalent is oriented with its long axis parallel to the equatorial plate, which suggests a holocentric interaction with the spindle fibres. After this initial orientation, kinetic activity is restricted to one of both chromatid ends. The election of the active chromatid end is random and it is independent of the end selected in the sister chromatid. At second metaphase, the X and Y chromatids associate side by side forming a pseudobivalent. After that, the kinetic activity is again restricted to either of both chromosomal ends in a random fashion. At first metaphase, the fourth autosomal bivalent shows two alternative random orientations depending on the chromosome end showing kinetic activity (DAPI positive or opposite). At second metaphase, half bivalents are oriented with their long axis parallel to the equatorial plate. Three different segregation patterns are observed. The kinetic activity can be localised: (i) in the end with the DAPI signal (46.9%), (ii) in the opposite end (44.6%) or (iii) in one DAPI-positive end in one chromatid and in the opposite end in the other one (8.5%). The existence of the last pattern indicates that the same end can show kinetic activity during both meiotic divisions. Our results provide new information on the comparative meiotic behaviour of autosomes and sex chromosomes in holocentric systems.     

Segregation of holocentric chromosomes at meiosis in the nematode,Caenorhabditis elegans

The meiotic segregation of the holocentric chromosomes ofCaenorhabditis elegans in both spermatogenesis and oogenesis is described. The extended kinetochore typical of the mitotic chromosome could not be differentiated on meiotic bivalents; instead microtubules appeared to project into the chromatin. The meiotic spindles formed during spermatogenesis contain centrioles and asters, while in oogenesis the spindles are acentriolar and barrel shaped. The formation of the acentriolar spindle was studied in fixed specimens by anti-tubulin immunofluorescence. Microtubule arrays were seen first to accumulate in the vicinity of the meiotic chromosomes prior to congression. At later stages, elongated spindle structures up to 13 in length were observed parallel to the surface of the embryo. Further development of the spindle appeared to involve its shortening into a barrel shape and rotation so that one spindle pole was opposed to the membrane. By anaphase the pole-to-pole spindle length reached a minimum of 3–4 . One end of each chromatid in the meiotic bivalent was labelled byin situ hybridization of a probe DNA to show that in oogenesis the chromatids were associated end-to-end in the bivalent. Furthermore, either the right or the left ends of the homologues could be held in association. At metaphase I the bivalents were oriented axially, such that kinetic activity was restricted to one end of each pair of sister chromatids. At metaphase II the chromosomes were also aligned axially.     

XY chromosome behaviour in the germ-line of the human male: A FISH analysis of spatial orientation,chromatin condensation and pairing

We have used multicolour fluorescencein situ hybridization to study the behaviour of the X and Y chromosomes in relation to a representative autosome, chromosome 1, on air-dried testicular preparations from normal fertile human males. In a proportion of Sertoli cells at interphase as well as spermatogonial metaphases there is an apparent selective undercondensation of the heterochromatic block of the long arm of the Y, which may be of functional significance with respect to Y-specific gene activity, initiating and maintaining spermatogenesis; we suggest that this may involve a mechanism similar to heterochromatin position-effect variegation inDrosophila. In the supporting Sertoli as well as pre-meiotic and leptotene cells the X and Y occupy relatively restricted domains at opposite poles of the nuclear membrane, while the chromosome 1 centromere regions are located interstitially and appear prealigned. The XY pairing and sex vesicle formation comprises a complex series of spatial movement and differential condensation patterns. On the basis of these observations we propose that: the XIST/Xist gene, known to be involved in somatic X inactivation, imposes a chromatin reorganization leading to bending at the X-inactivation centre both at first meiotic prophase in males and in the soma in females; and the differential X and Y segments are protected from potentially deleterious meiotic exchanges by their separate spatial orientation. In addition, there is an indication that the timing of pairing and first meiotic segregation of the sex chromosomes is different, and precocious in comparison to the pairing and segregation of the autosomes, which may explain the high incidence of sex chromosome aneuploidy in sperm.     

Meiotic behaviour of holocentric chromosomes: orientation and segregation of autosomes in Triatoma infestans (Heteroptera)

The meiotic behaviour of the holocentric chromosomes of the heteropteran species Triatoma infestans has been analysed by means of orcein staining and C-banding on squashed spermatocytes. We have focused our analysis on chromosome 3, which shows a large distal heterochromatic band at one of the ends of both homologues. At metaphase I,and independently of the chiasma position, two alternative orientations have been observed: either the hetero-chromatic or the euchromatic ends of both homologues are directed to opposite poles. At anaphase I, the kinetic activity is restricted to the same chromosome end (euchromatic or heterochromatic) of each homologue. The frequencies of these two alternatives are not random and differ significantly among the five individuals analysed. However, the euchromatic ends present kinetic activity at a higher frequency than the heterochromatic ends. At metaphase II, half-bivalents also show the kinetic activity restricted to either of the chromosome ends (euchromatic orheterochromatic). The frequencies of each alternative are inverted in anaphase II compared with those scored in anaphase I. Accordingly, those ends that present kinetic activity at anaphase I segregate reductionally during the first meiotic division and equationally during the second meiotic division. These results provide sound evidence on the meiotic behaviour of holocentric chromosomes, as regards the absence of chiasma terminalization and the modes of orientation and segregation. This revised version was published online in August 2006 with corrections to the Cover Date.     

Behaviour of ring bivalents in holokinetic systems: Alternative sites of spindle attachment in Pachylis argentinus and Nezara viridula (Heteroptera)

Heteropteran chromosomes are holokinetic; during mitosis, sister chromatids segregate parallel to each other but, during meiosis, kinetic activity is restricted to one pair of telomeric regions. This meiotic behaviour has been corroborated for all rod bivalents. For ring bivalents, we have previously proposed that one of the two chiasmata releases first, and a telokinetic activity is also achieved. In the present work we analyse the meiotic behaviour of ring bivalents in Pachylis argentinus (Coreidae) and Nezara viridula (Pentatomidae) and we describe for the first time the chromosome complement and male meiosis of the former (2n = 12 + 2m + X0, pre-reduction of the X). Both species possess a large chromosome pair with a secondary constriction which is a nucleolus organizer region as revealed by in-situ hybridization. Here we propose a new mode of segregation for ring bivalents: when the chromosome pair bears a secondary constriction, it is not essential that one of the chiasmata releases first since these regions or repetitive DNA sequences adjacent to them become functional as alternative sites for microtubule attachment and they undertake chromosome segregation to the poles during anaphase I.     

Nonrandom sister chromatid segregation of sex chromosomes in Drosophila male germline stem cells

Sister chromatids are the product of DNA replication, which is assumed to be a very precise process. Therefore, sister chromatids should be exact copies of each other. However, reports have indicated that sister chromatids are segregated nonrandomly during cell division, suggesting that sister chromatids are not the same, although their DNA sequences are the same. Researchers have speculated that stem cells may retain template strands to avoid replication-induced mutations. An alternative proposal is that cells may segregate distinct epigenetic information carried on sister chromatids. Recently, we found that Drosophila male germline stem cells segregate sister chromatids of X and Y chromosomes with a strong bias. We discuss this finding in relation to existing models for nonrandom sister chromatid segregation.     

The telochore: A telomeric differentiation of the chromosome axis

We have analysed by means of silver staining the structure of the chromosome axis at the telomeres of meiotic chromosomes in three different grasshopper species. At metaphase I the chromatid axes run the length of the chromatids although they do not reach the chromosome ends. The axes of sister chromatids are associated and show a round differentiation at their distal ends that we have named the telochore. Telochores never contact the chromosome ends: there is always some chromatin beyond them. In late metaphase I bivalents with a distal chiasma, anaphase I and metaphase II half-bivalents and anaphase II chromatids, the axes clearly possess one telochore in each chromosome end. These results seem to indicate that telochores are differentiations of the distal ends of chromatids. We discuss the possible structural significance of telochores according to the current scaffold/radial loop model of chromatin organization of eukaryotic metaphase chromosomes. Additionally, we suggest the possible functional role of the telochore as a nucleoprotein domain forming a protective cap for telomeric DNA.     

Holocentric chromosomes in meiosis. II. The modes of orientation and segregation of a trivalent

The modes of orientation and segregation of the sex chromosome trivalent X₁X₂Y in male meiosis of Cacopsylla mali (Psylloidea, Homoptera) were analysed. Males with an X₁X₂Y sex chromosome system coexist with males displaying a neo-XY system in populations of this species. The fusion chromosome resulting in the formation of a trivalent in meiosis originates from the fusion of an autosome with the neo-Y chromosome. In the majority of metaphase I cells (92.4%) the X₁X₂Y trivalent showed co-orientation; X₁ and X₂ chromosomes oriented towards one pole whereas the Y oriented towards the opposite pole. In the rest of the cells (7.6%) the trivalent with subterminal chiasmata was oriented parallel to the equatorial plane. From this orientation the trivalent produced triple chromatids joined together by undivided telomeric parts of chromosomes and hence by sister chromatid cohesion at anaphase I. In the majority of metaphase II cells the orientation of triple chromatids suggested the production of unbalanced gametes. However, in a small number of cells (1.7%) the trivalent showed co-orientation of X₁X₂ with Y. Both the first division and second division co-orientations, or 94.1% of divisions as a whole, were estimated to yield balanced gametes, containing either X₁ and X₂ chromosomes or Y chromosome. It was concluded that, since the triple chromatid contained undivided telomere regions at metaphase II, which divided at anaphase II, the orientation of the trivalent with its longitudinal axis parallel to the equatorial plane in metaphase I also represents co-orientation and results in pre-reduction. The existence of post-reductional behaviour of holocentric bivalents and multivalents is discussed.     

The distribution of α-kleisin during meiosis in the holocentromeric plant <Emphasis Type="Italic">Luzula elegans</Emphasis>

Holocentric chromosomes occur in a number of independent eukaryotic lineages, and they form holokinetic kinetochores along the entire poleward chromatid surfaces. Due to this alternative chromosome structure, Luzula elegans sister chromatids segregate already in anaphase I followed by the segregation of the homologues in anaphase II. However, not yet known is the localization and dynamics of cohesin and the structure of the synaptonemal complex (SC) during meiosis. We show here that the α-kleisin subunit of cohesin localizes at the centromeres of both mitotic and meiotic metaphase chromosomes and that it, thus, may contribute to assemble the centromere in L. elegans. This localization and the formation of a tripartite SC structure indicate that the prophase I behaviour of L. elegans is similar as in monocentric species.     

Meiotic analysis of XX/XY and neo-XX/XY sex chromosomes in Phyllostomidae by cross-species chromosome painting revealing a common chromosome 15-XY rearrangement in Stenodermatinae

We analyzed the meiotic behavior of the sex chromosomes of the bats Glossophaga soricina (XX/XY), Artibeus cinereus and Uroderma magnirostrum (neo-XX/XY) using multicolor FISH. The X chromosome and pair 15 autosome probes are from Phyllostomus hastatus and the Y from Carollia brevicauda. On both species with the neo-XX/XY system, the autosome translocated to the sex chromosomes is the pair 15 in P. hastatus, a synapomorphy. The analysis of meiosis confirms that the X and Y have a pseudo-autosomal region, with a typical end-to-end pairing. The autosomal regions of the neo-XX/XY shows different levels of condensation when compared to the original XX/XY. The compound system presented a characteristic shape, as if it was a closed ring with a tail. The ring represents the non-paired segments of the X and Y and the small pairing region of the original sex chromosomes. The tail corresponds to the pairing of the 15 P. hastatus autosomal bivalent, which are translocated to the sex chromosomes. Probably the non-pairing is responsible for the meiotic silencing of these segments.     

Segregation of chromosomes into the spermatozoa of a man heterozygous for a 14;21 Robertsonian translocation

Twenty-four spermatozoa from a man heterozygous for a Robertsonian translocation (45,XY,-14,-21,+t(14q;21q) were studied cytogenetically in order to determine the meiotic segregation of the translocation. When compared to the expected 1:1 ratio we observed a greater number of chromosomally normal sperm than sperm with the balanced translocation. Three sperm carried the translocation in an unbalanced form.     

Protein immunolocalization supports the presence of identical mechanisms of XY body formation in eutherians and marsupials

The meiotic sex chromosomes of the American marsupials Monodelphis dimidiata and Didelphis albiventris were studied with electron microscopy (EM) and with immunofluorescence localization of meiotic proteins SYCP1 and SYCP3, and proteins essential for meiotic sex chromosome inactivation (MSCI), γ-H2AX and BRCA1. The chromatin of the non-synaptic X and Y chromosomes contains γ-H2AX, first as foci and then as homogeneous staining at late stages. The thick and split X and Y axes are labelled with BRCA1 except at one terminus. The bulgings of the axes contain SYCP1 as well as the inner side of the dense plate. The evenly spaced and highly packed chromatin fibres of the conjoined XY body in these species have the same behaviour and the same components (γ-H2AX in the chromatin, BRCA1 in the axes) as in the XY body of eutherian species. These observations and recent data from the literature suggest that XY body formation is ancestral to the metatherian–eutherian divergence.     

Characterization of rec15, an early meiotic recombination gene in Schizosaccharomyces pombe

In S. pombe strains mutant for rec15 aberrant ascus morphology, reduced spore viability and severe reduction of meiotic recombination was detected. Genetic and cytological analysis identified frequent interruption of meiosis after the first division, and nondisjunction I, as the main segregation errors in the mutant. Chromosome segregation at meiosis I was not random in rec15, suggesting the presence of a backup system for correct segregation of achiasmate chromosomes. The analysis of meiotic progression in time-course experiments revealed that the major meiotic events, such as the onset of premeiotic DNA synthesis, of horse-tail nuclear movement, and of the first meiotic division occurred earlier in rec15 than in wild-type. The early onset of meiotic events is a novel observation for an early recombination mutant and implies a function of rec15 protein already at or before DNA synthesis.     

Behavior of chloroplast genes during the early zygotic divisions of Chlamydomonas reinhardtii

Summary Chloroplast mutations in the green alga Chlamydomonas reinhardtii exhibit a predominantly maternal pattern of inheritance and this pattern can be perturbed by UV irradiation of the maternal gametes prior to mating. In a series of crosses over a range of UV doses, the transmission, segregation, and recombination of mutations at three closely linked chloroplast loci have been examined by pedigree analysis of products arising from the first three post-zygotic divisions. Stocks used in these crosses were constructed to permit identification of the nuclear products of each of the two meiotic divisions and the first post-meiotic mitotic division.A bias toward maternal alleles at all three chloroplast loci was observed in all pedigrees and in zygote clones analyzed from the same crosses many generations after meiosis. This bias decreased with increasing UV dose and with each subsequent division. Segregation of chloroplast genes was rapid during the first three post-zygotic divisions. The type of segregation event from which a given heteroplasmic cell arose had a significant effect on its most likely segregation. pattern in the subsequent division. The results presented here have been discussed in terms of published models of chloroplast gene segregation.     

A model for chromosome structure during the mitotic and meiotic cell cycles

The chromosome scaffold model in which loops of chromatin are attached to a central, coiled chromosome core (scaffold) is the current paradigm for chromosome structure. Here we present a modified version of the chromosome scaffold model to describe chromosome structure and behavior through the mitotic and meiotic cell cycles. We suggest that a salient feature of chromosome structure is established during DNA replication when sister loops of DNA extend in opposite directions from replication sites on nuclear matrix strands. This orientation is maintained into prophase when the nuclear matrix strand is converted into two closely associated sister chromatid cores with sister DNA loops extending in opposite directions. We propose that chromatid cores are contractile and show, using a physical model, that contraction of cores during late prophase can result in coiled chromatids. Coiling accounts for the majority of chromosome shortening that is needed to separate sister chromatids within the confines of a cell. In early prophase I of meiosis, the orientation of sister DNA loops in opposite directions from axial elements assures that DNA loops interact preferentially with homologous DNA loops rather than with sister DNA loops. In this context, we propose a bar code model for homologous presynaptic chromosome alignment that involves weak paranemic interactions of homologous DNA loops. Opposite orientation of sister loops also suppresses crossing over between sister chromatids in favor of crossing over between homologous non-sister chromatids. After crossing over is completed in pachytene and the synaptonemal complex breaks down in early diplotene (= diffuse stage), new contractile cores are laid down along each chromatid. These chromatid cores are comparable to the chromatid cores in mitotic prophase chromosomes. As an aside, we propose that leptotene through early diplotene represent the missing G2 period of the premeiotic interphase. The new chromosome cores, along with sister chromatid cohesion, stabilize chiasmata. Contraction of cores in late diplotene causes chromosomes to coil in a configuration that encourages subsequent syntelic orientation of sister kinetochores and amphitelic orientation of homologous kinetochore pairs on the spindle at metaphase I.     

Nucleolus size variation during meiosis and NOR activity of a B chromosome in the grasshopper <Emphasis Type="Italic">Eyprepocnemis plorans</Emphasis>

The number of nucleoli and nucleolar area were measured in meiotic cells from males of the grasshopper Eyprepocnemis plorans collected in three natural populations. Number of nucleoli per cell showed no significant correlation among cells in different meiotic stages, but there was strong positive correlation for nucleolar area between leptotene and interkinesis cells in individuals from distant populations (Salobreña in Spain, and Smir in Morocco). No correlation was, however, observed for both parameters between the meiotic stages analysed in individuals from the population of Torrox (Spain). The number of nucleoli at leptotene was about double the number at interkinesis, as expected from the double ploidy level at leptotene and the corresponding double number of rDNA clusters. Leptotene nucleolar area, however, was about fourfold that in interkinesis, presumably due to higher requirements for ribosome biogenesis in meiosis I than meiosis II. In Torrox, diplotene cells showed a lower number of nucleoli but larger nucleolar area than in leptotene cells, suggesting an increase in nucleolus size during prophase I. Significant differences were found among populations for nucleolar area but not for number of nucleoli, the smallest nucleolar area being observed in Torrox, which is the population harbouring the most parasitic B chromosome variant. No clear effects on nucleolar area or number of nucleoli were associated with the B-chromosome number. However, B-chromosome effects on the nucleolar area were apparent in the Torrox population when data were analysed with respect to a B-chromosome odd–even pattern in leptotene and interkinesis cells. However, in diplotene cells no odd–even pattern was observed for both nucleolar parameters, suggesting that the increase in nucleolar size from leptotene to diplotene dilutes the leptotene odd–even pattern. The rDNA distally located in the B chromosome was associated with a nucleolus in 6.5% out of the 247 diplotene cells analysed. The implications of these findings are discussed in the context of B chromosomes as stress-causing genome parasites and the nucleolus as a sensor of stress.     

Diagnosing and preventing inherited disease: The use of first polar bodies for preimplantation diagnosis of aneuploidy

A large proportion of patients undergoing in-vitro fertilization(IVF) are aged 35 years. It has been estimated that in thisage group, 50% of embryos are chromosomally abnormal, with aneuploidybeing the major contributing factor. Since the origin of mostaneuploidies is maternal meiosis I non-disjunction, unfertilizedoocytes could be safely screened for aneuploidy by analysingtheir first polar bodies. To determine the feasibility of firstpolar body aneuploidy analysis, polar bodies were analysed byfluorescence in-situ hybridization (FISH) using probes simultaneouslyfor chromosomes X, Y, 18, 13/21 or X, Y, 18 and 16. Within 6h of retrieval, 88% showed a normal segregation involving asingle chromosome of each kind, with double-dotted hybridizationsignals, corresponding to dyads (chromosomes in metaphase Icomposed of two chromatids). The rest showed non-disjunctionof full dyads (6%), or an unbalanced pre-division of dyads (6%),which gives a segregation of one chromatid or one dyad and achromatid with the first polar body. But only 34% of polar bodiesanalysed 24 h after retrieval or later showed a normal segregation,with most of the other polar bodies showing balanced pre-division,with two separated hybridization signals for all the chromosomesanalysed. The rates of non-disjunction and unbalanced pre-divisionafter 24 h in culture were similar to the rates in fresh oocytes.When both types of aneuploidy were considered together, an increaseof aneuploidy with maternal age was detected, which althoughslight, was significant (P = 0.025). Because dyads seem to undergorapid pre-division shortly after polar body retrieval, performanceof FISH aneuploidy analysis of polar bodies is therefore onlyrecommended when conducted within 6 h of their retrieval.     

Microsatellite analysis in Turner syndrome: Parental origin of X chromosomes and possible mechanism of formation of abnormal chromosomes

Turner syndrome is a chromosomal disorder in which all or part of one X chromosome is missing. The meiotic or mitotic origin of most cases remains unknown due to the difficulty in detecting hidden mosaicism and to the lack of meiotic segregation studies. We analyzed 15 Turner patients, 10 with a 45,X whereas the rest had a second cell line with abnormal X‐chromosomes: a pseudodicentric, an isochromosome, one large and one small ring, and the last with a long arm deletion. Our aims were: to detect X cryptic mosaicism in patients with a 45,X constitution; to determine the parental origin of the abnormality; to infer the zygotic origin of the karyotype and to suggest the timing and mechanism of the error(s) leading to the formation of abnormal X chromosomes from maternal origin. Molecular investigation did not revealed heterozygosity for any microsatellite, excluding X mosaicism in the 45,X cases. Parental origin of the single X chromosome was maternal in 90% of these patients. Three of the structurally abnormal Xs were maternally derived whereas the other two were paternal. These results allowed us to corroborate breakpoints in these abnormal X chromosomes and suggest that the pseudodicentric chromosome originated from post‐zygotic sister chromatid exchange, whereas the Xq deleted chromosome probably arose after a recombination event during maternal meiosis. © 2001 Wiley‐Liss, Inc.     

Unbiased segregation of yeast chromatids in Saccharomyces cerevisiae

The budding yeast Saccharomyces cerevisiae is characterized by asymmetric cell division and the asymmetric inheritance of spindle components during normal vegetative growth and during certain specialized cell divisions. There has been a longstanding interest in the possibility that yeast chromosomes segregate non-randomly during mitosis and that some of the differences between mother and daughter cells could be explained by selective chromatid segregation. This review traces the history of the experiments to determine if there is biased chromatid segregation in yeast. The special aspects of spindle morphogenesis and behavior in yeast that could accommodate a mechanism for biased segregation are discussed. Finally, a recent experiment demonstrated that yeast chromatids segregate randomly without mother–daughter bias in a common laboratory strain grown under routine laboratory conditions.     

Hybridization of Pichia stipitis with its presumptive imperfect partner Candida shehatae

Summary A triauxotrophic strain of the D-xylose fermenting yeast Pichia stipitis, was hybridized with diauxotrophic strains of its presumptive imperfect partner Candida shehatae through polyethylene glycol (PEG) induced protoplast fusion. A small fraction of prototrophic clones, selected on specific media, appeared to be partial hybrids as determined by cellular diphenylamine DNA quantitations after three passages on a complex medium. The hybrid nature of the Pichia-resembling fusants was confirmed by cell volume estimation, analysis of nuclear condition and the isolation of a variety of mutant recombinant phenotypic segregants by meiotic segregation, as well as induced and spontaneous mitotic segregation.