Centric Misdivision Research Articles

Only the Rye Derived Part of the 1BL/1RS Hybrid Centromere Incorporates CENH3 of Wheat.

The precise assembly of the kinetochore complex at the centromere is epigenetically determined by substituting histone H3 with the centromere-specific histone H3 variant CENH3 in centromeric nucleosomes. A chromosome 1B reconstructed in wheat by centric misdivision from two wheat-rye centric translocations is known to carry a hybrid wheat-rye centromere. The resulting hybrid (dicentric)centromere is composed of both wheat and rye centromeric repeats. As CENH3 is a marker for centromere activity, we applied Immuno-FISH followed by ultrastructural super-resolution microscopy to address whether both or only parts of the hybrid centromere are active. Our study demonstrates that only the rye-derived centromere part incorporates CENH3 of wheat in the 1BL/1RS hybrid centromere. This finding supports the notion that one centromere part of a translocated chromosome undergoes inactivation, creating functional monocentric chromosomes to maintain chromosome stability.

Special features of transmission of markers for Aegilops biuncialis Vis. Chromosome 1U in common wheat hybrids

Aim. The wild wheat relative Aegilops biuncialis Vis. is a potential source of new storage protein alleles for enriching the common wheat gene pool. We have produced the wheat material from direct crossing of common wheat with Crimean samples of Ae. biuncialis followed by backcrossing with wheat. Using storage proteins as genetic markers, lines with introgressions of chromosome 1U have been selected. The objective of the investigation was to study the transmission of markers for chromosome 1U in the hybrid from crossing the common wheat cultivar Bezostaya 1 with a line carrying chromosome 1U. Methods. F2 seeds from the cross of the common wheat cultivar Besostaya 1 and the line NVG41 carrying Ae. biuncialis chromosome 1U were analyzed. Storage proteins encoded by Gli-U1 and Glu-U1 served as markers for chromosome 1U. Gliadins were analyzed by acid polyacrylamide gel electrophoresis by the procedure of Kozub et al. (2009). Electrophoresis of total seed proteins, including high-molecular-weight glutenin subunits, was carried out by the procedure of Laemmli (1970).Results. Among the F2 seeds analyzed, we identified four classes of genotypes with respect to the markers Gli-U1 and Glu-U1: 1) with complete chromosome 1U, 2) with the 1UL arm only, 3) with the 1US arm only, 4) without chromosome 1U. In the F1 hybrid NVG41 × Bezostaya 1, Ae. biuncialis chromosome 1U was transmitted with an increased frequency (with respect to the marker Glu-U1 and the simultaneous presence of the markers Gli-U1 and Glu-U1). The frequency of centric misdivision of chromosome 1U (univalent) in the F1 hybrid amounted about 9 %. This resulted in a significantly lower number of seeds with the 1US arm only (2 %) than that with the 1UL arm (6 %).Conclusions. In the progeny of F1 hybrids with Ae. biuncialis univalent 1U, a lower frequency of genotypes with the 1US arm only as compared with the 1UL arm only can be observed at the grain stage. This could be due to the loss of telocentrics with the 1US arm at gamete formation or reduced viability of genotypes with 1US only at the gamete stage or after zygote formation.Manuscript received 02.04.2020

Behavior of Centromeres in Univalents and Centric Misdivision in Wheat

Centromeres are responsible for the proper behavior of chromosomes in cell divisions. In meiosis the process is more complicated than in mitosis, as each chromosome in a bivalent has 2 sister centromeres and their behavior has to be strictly coordinated. Here, the behavior of sister centromeres in univalents in wheat is examined, showing that by metaphase I they often lose their coordination. This loss accelerates with the progression of anaphase I, leading to stable bipolar attachment and frequent separation of sister chromatids or to misdivision. Depending on the orientation of a univalent and its sister centromeres, misdivision may occur across the centromere region or across the pericentric chromatin. Chromosome fragments consisting of only the centromere region did not survive to the next generation. Midget chromosomes composed of the centromeres and parts of the pericentric chromatin did survive, but their transmission rates were low and appeared related to the amount of pericentric chromatin, probably because only the pericentric chromatin provides sister chromatid cohesion. As the cohesion of sister chromatids appears to be a function of the proximity to the kinetochore region, the definition of the centromere need not include pericentric regions.

C-Myc—Dependent Formation of Robertsonian Translocation Chromosomes in Mouse Cells

Robertsonian (Rb) translocation chromosomes occur in human and murine cancers and involve the aberrant joining of two acrocentric chromosomes in humans and two telocentric chromosomes in mice. Mechanisms leading to their generation remain elusive, but models for their formation have been proposed. They include breakage of centromeric sequences and their subsequent fusions, centric misdivision, misparing between highly repetitive sequences of p-tel or p-arm repeats, and recombinational joining of centromeres and/or centromeric fusions. Here, we have investigated the role of the oncoprotein c-Myc in the formation of Rb chromosomes in mouse cells harboring exclusively telocentric chromosomes. In mouse plasmacytoma cells with constitutive c-Myc deregulation and in immortalized mouse lymphocytes with conditional c-Myc expression, we show that positional remodeling of centromeres in interphase nuclei coincides with the formation of Rb chromosomes. Furthermore, we demonstrate that c-Myc deregulation in a myc box II-dependent manner is sufficient to induce Rb translocation chromosomes. Because telomeric signals are present at all joined centromeres of Rb chromosomes, we conclude that c-Myc mediates Rb chromosome formation in mouse cells by telomere fusions at centromeric termini of telocentric chromosomes. Our findings are relevant to the understanding of nuclear chromosome remodeling during the initiation of genomic instability and tumorigenesis.

Unstable inheritance of maize B-type chromosomes that lack centric heterochromatin

The B chromosome of maize undergoes frequent non-disjunction at the second pollen mitosis. In B-A translocations, the B-A chromosome retains the capacity for non-disjunction. We have collected deletion-derivative TB-9Sb stocks. One derivative, the "type 1 telocentric", has a B-9 chromosome that lacks centric heterochromatin. It produces few recessive (non-disjunctional) phenotypes in pollen parent testcrosses of the translocation heterozygote, 9 9-B telo B-9. The finding helped demonstrate the role of centric heterochromatin in non-disjunction. An isochromo some derivative of the type 1 telocentric was also recovered. It was tested in the 9-B 9-B iso B-9 constitution. This is equivalent to 9 9-B telo B-9 in terms of chromosome 9 dosage. Surprisingly, crosses with the isochromosome gave significant levels of recessive phenotypes. In addition, high levels of variegated phenotypes were found. Recently, a circumstance was found that makes inheritance of the type 1 telocentric chromosome somewhat similar to that of the isochromosome. Crosses with hypoploid 9-B 9-B telo B-9 plants showed significant levels of recessive and variegated phenotypes. These crosses were investigated to help explain the source(s) of the phenotypes. Cytological and genetic studies were performed. Centric misdivision was found to account for the variegated phenotypes. A mixture of conventional B non-disjunction and centric misdivision produced the recessive phenotypes. The significance of conventional non-disjunction in the absence of centric heterochromatin is discussed.

Leaf rust and stripe rust resistance genes Lr54 and Yr37 transferred to wheat from Aegilops kotschyi

AbstractThe tendency of unpaired meiotic chromosomes to undergo centric misdivision was exploited to translocate leaf rust and stripe rust resistance genes from an Aegilops kotschyi addition chromosome to a group 2 chromosome of wheat. Monosomic and telosomic analyses showed that the translocation occurred to wheat chromosome arm 2DL. The introgressed region did not pair with the corresponding wheat 2DL telosome during meiosis suggesting that a whole arm may have been transferred. Female transmission of the resistance was about 55% whereas male transmission was strongly preferential (96%). The symbols Lr54 and Yr37 are proposed to designate the new resistance genes.

Robertsonian translocations in wheat arise by centric misdivision of univalents at anaphase I and rejoining of broken centromeres during interkinesis of meiosis II

The mechanism of origin of Robertsonian translocations was investigated in plants monosomic for chromosome 1A of wheat and 1H^t of Elymus trachycaulus by GISH. Chromosomes 1A and 1H^t stayed univalent in all metaphase I cells analyzed, suggesting that Robertsonian translocations do not originate from meiotic recombination in centromeric regions with shared DNA sequence homology. At ana-/telophase I, the 1H^t and 1A univalents underwent either chromosome or chromatid segregation and misdivided in 6–7% of the pollen mother cells. None of the ana-/telophases I analyzed had Robertsonian translocations, which were only observed in 2% of the “half tetrads” at ana-/telophase II. The frequency of Robertsonian translocations observed at ana-/telophase II corresponds well with the number of Robertsonian translocations (1–4%) detected in progenies derived from plants monosomic for group-1 chromosomes of wheat (1A, 1B, and 1D) and 1H^t of E. trachycaulus. Our data suggest that Robertsonian translocations arise from centric misdivision of univalents at ana-/telophase I, followed by segregation of the derived telocentric chromosomes to the same nucleus, and fusion of the broken ends during the ensuing interkinesis.

Molecular distinction between true centric fission and pericentric duplication-fission

Centromere (centric) fission, also known as transverse or lateral centric misdivision, has been defined as the splitting of one functional centromere of a metacentric or submetacentric chromosome to produce two derivative centric chromosomes. It has been observed in a range of organisms and has been ascribed an important role in karyotype evolution; however, the underlying mechanisms remain unknown. We have investigated four cases of apparent centric fission in humans. Two cases show a missing chromosome 22 or 18 that is replaced by two centric ring products, a third case shows two chromosome-10-derived telocentric chromosomes, whereas a fourth case involves the formation of two chromosome-18-derived isochromosomes. In all four cases, results of gross cytogenetic and fluorescence in situ hybridisation analyses were consistent with a simple centric fission event. However, detailed molecular analyses provided evidence in support of centromere duplication as a predisposing mechanism for the observed chromosomal breakage in two of the cases. Results for the third case are consistent with direct centric fission not involving centromere pre-duplication as the likely mechanism. Insufficient material has precluded the further study of the fourth case. The data provide the first molecular evidence for centromere pre-duplication as a possible mechanism to explain the classically assumed simple "centric fission" events in clinical cytogenetics, karyotype evolution and speciation.

An asymptotic determination of minimum centromere size for the maize B chromosome

The maize B chromosome is a dispensable chromosome and therefore serves as a model system to study centromere function. The B centromere region is estimated to be approximately 9,000 kb in size and contains a 1.4 kb repeat that is specific to this centromere. When maintained as a univalent, the B chromosome occasionally undergoes centric misdivision. Consecutive misdivision analysis of the maize B chromosome centromere has generated a collection of functional centromeres that are greatly reduced in complexity. These small centromeres are often correlated with strongly reduced meiotic transmission. Molecular analyses of the misdivision collection have revealed that the smallest functional maize B centromere is a minimum of 110 kb in size. Considering the collection as a whole, meiotic transmission becomes severely compromised when the estimated centromere size is reduced to a few hundred kilobases.

The centromere structure in Robertsonian wheat-rye translocation chromosomes indicates that centric breakage-fusion can occur at different positions within the primary constriction.

Univalent chromosomes at meiotic metaphase I have a tendency to misdivide at the centromeres. Fusion of the misdivision products may produce Robertsonian translocations. The fine structure of the centromeres in Robertsonian wheat-rye translocation chromosomes was analyzed by fluorescence in situ hybridization (FISH) using two centromere-specific DNA clones: pRCS1, derived from rice, and pAWRC1, derived from rye. Clone pRCS1 hybridizes to the centromeres of all grasses including wheat and rye, whereas clone pAWRC1 is rye specific and hybridizes only to the centromeres of rye. Four of the six wheat-rye translocations derived from a single centric misdivision event (1st generation translocations) had hybrid centromeres, with approximately half of the centromere derived from rye and half from wheat. In the two other 1st generation translocations, the entire centromere was derived from rye. Among eight reconstructed wheat and rye chromosomes that originated from two consecutive centric misdivision-fusion events (2nd generation translocations), T1BS.1BL (derived from T1BS.1RL and T1RS.1BL) and one of three T2BS.2BL (derived from T2RS.2BL and T2BS.2RL) had hybrid centromeres. T1RS.1RL (derived from T1BS.1RL and T1RS.1BL), two of three T2BS.2BL, and all three T2RS.2RL (derived from T2RS.2BL and T2BS.2RL) had rye centromeres. All three 3rd generation translocations had hybrid centromeres with approximately half of the centromere derived from rye. There were no indications that the composite structure of the centromere in these chromosomes affected their behavior in mitosis or meiosis. These observations support the notion of a compound structure of the centromere in higher organisms, and indicate that during the centric breakage-fusion event, centromere breakage may occur in different positions along the segment of the chromosome that interacts with the spindle fibers. Normal behavior of the 1st, 2nd, and 3rd generation centric translocations in mitosis and meiosis indicates that, at least in wheat and rye, centromeres are not chromosome specific.

Development of Arm Specific RAPD Markers for Rye Chromosome 2R in Wheat

ABSTRACTRye (Secale cereale L.) chromosome 2R carries several genes for resistance to biotic and abiotic stresses, which could be used in wheat (Triticum aestivum L.) improvement. To make these genes accessible in breeding, translocations of 2R to wheat chromosome 2B have been generated. To compensate for the lack of clearly identifiable cytological markers on chromosome 2R, random amplified polymorphic DNAs (RAPDs) were employed as markers. Wheat lines homozygous for centric translocations 2RS.2BL and 2BS.2RL were screened with 489 primers, resulting in 65 clear and reproducible RAPD polymorphisms. To assign the polymorphisms to individual arms of chromosomes 2R and 2B, the arms of the translocations were separated by centric misdivision and the resulting progeny with desired chromosome constructs were rescreened. As a result, 17 markers were assigned to chromosome arm 2RS, 15 to 2RL, and the remaining 33 most likely located on 2B. Preliminary mapping studies (not shown here) have indicated that some of these markers are evenly distributed along the chromosome arms. These markers should be useful for the localization of translocation breakpoints in recombinant wheat‐rye chromosomes.

Construction of midget chromosomes in wheat

To test the usefulness of breakage-fusion-bridge (BFB) cycles in generating new chromosome aberrations in bread wheat (Triticum aestivum L.) and to extend the range of aberrations available, a series of midget chromosomes was produced from the long arm of chromosome 1B. Using a reverse tandem duplication initiated chromatid type BFB cycle, the 1BL arm was broken and fused with centromeres of either chromosome 5BL or 1RS to form dicentric chromosomes. The 1R and 5B centromeres were broken by centric misdivision. Among the progenies of plants with dicentric chromosomes, two classes of monocentric chromosomes were selected: deficient chromosomes 1B and chromosomes that had 1RS or 5BL for one arm and various fragments of 1BL for the other arm. Following centric misdivision of these monocentrics, midget chromosomes 1BL were isolated: deficient and deletion telocentrics and telocentrics derived from interstitial regions of 1BL. By chance, one deficient chromosome 1BS and one deletion chromosome 1BS were identified in unrelated lines of the same wheat. Following centric misdivision of these chromosomes, two midget chromosomes covering the whole of 1BS were added to the set.

Further manipulation by centric misdivision of the 1RS.1BL translocation in wheat

Complete chromosomes 1R and 1B were reconstructed in wheat from the centric wheat-rye translocation 1RS.1BL. Three substitutions: 1R(1A), 1R(1B), 1R(1D), and three new centric translocations: 1RS.1AL, 1RS.1BL, 1RS.1DL were produced from the reconstructed chromosome 1R. Each one of these has the same rye chromosome arm 1RS which was present in the original translocation 1RS.1BL of ‘Kavkaz’ wheat. Reconstructed chromosome 1B and a normal chromosome 1R were used to produce a new 1RS.1BL translocation. This translocation has the long arm from the original 1RS.1BL translocation of ‘Kavkaz’, but a different 1RS arm. The third generation centric translocations were mitotically stable and were normally transmitted to progeny. Misdivision frequency of the reconstructed chromosomes 1R did not change relative to normal 1R, whereas the misdivision frequency of the two reconstructed chromosomes 1B tested was significantly higher relative to normal 1B. These experiments demonstrate that repeated cycles of centric breakage and fusion do not impair the function of centromeres in wheat and rye but may change chromosome's susceptibility to misdivision.

Reconstruction in wheat of complete chromosomes 1B and 1R from the 1RS.1BL translocation of 'Kavkaz' origin

Translocation 1RS.1BL originally from the 'Aurora' and 'Kavkaz' wheats has spread to bread wheats all over the world. It offers several resistance genes and appears to significantly increase yields. For future study of the arm location of the yield-increasing factors, complete chromosomes 1B and 1R were reconstructed from the 1RS.1BL translocation. This was accomplished by centric misdivision and fusion in double monosomies 20 II + 1RS.1BL I + 1BS.1RL I. The frequency of misdivision and misdivision-fusion of the two unpaired chromosomes was 23.1%, which was much higher than expected but comparable with that observed in other stocks grown under similar conditions. Transmission of the reconstructed chromosomes to progeny was normal. In addition to all possible misdivision and misdivision-fusion products, two chromosomes of ambiguous origin were recovered: chromosome 1B with a proximal insert of about 40% of 1RS and chromosome 1R with a proximal insert of about 30% of 1BS. The study demonstrates that centric translocations can misdivide and fuse again to produce stable chromosomes with new arm combinations.

Cytogenetic Studies of Plasmodial Sporocytes, Chromatin Disintegration, Desynapsis and Centromere Breakage in Pearl Millet.

In pearl millet (Pennisetum americanum (L.) Leeke, n=7), in the selfed progenies of a line L. S. 326-3, a mutant with plasmodial sporocyte (syncyte) formation, chromatin disintegration, desynapsis and centromere breakage was identified and its cytogenetic features were studied. Two to 24-nucleate syncytes were formed due to suppression of cytokinesis during premeiotic mitoses. In syncytes, chromatin degeneration occurred from prepachytene to telophase I stages, thereafter only masses of empty cytoplasm were found. In uninucleate PMCs, chromatin extruded in prepachytene stages in some, degenerated upto diakinesis in several, and either bivalents or univalents (ranging from 1 to 14) were formed in the rest. Centric misdivision leading to telocentric chromosomes (ranging from 2 to 14 per cell) occurred in a proportion of PMCs at anaphase I. There was high sterility of male and female gametes.F2 and test cross progenies of mutant X normal segregated respectively to 9:3:3:1 and 1:1:1:1 ratios of (i) completely normal meiotic behaviour, (ii) syncyte formation and chromatin disintegration but without desynapsis and centromere breakage, (iii) desynapsis and centromere breakage but without syncyte formation and chromatin disintegration and (iv) meiotic behaviour like the original mutant. Crosses between the recombinants (ii) and (iii) above yielded progeny with completely normal cytological behaviour. The original mutant thus was a double mutant. The genes were designated as snd (syncyte formation and chromatin disintegration) and ds cb (desynapsis and centromere breakage).

The B-chromosome system of Allium schoenoprasum

Nine morphologically distinct euchromatic B-chromosomes have been identified in Allium schoenoprasum from the River Wye, South Wales. The most common type (89%) is telocentric (Bt−1) and it is likely that the non-standard Bs are derivatives of Bt−1 by deletion, centric shift and/or centric misdivision. New B-types have also been produced from standard Bs in controlled crosses. In general, the Bs are mitotically extremely stable, although occasional plants, particularly those carrying non-standard Bs, are conspicuously variable in their B-constitution between root-tip cells. In addition, B-chromosome number is enhanced in some anthers of about one third of plants. Behaviour of B-chromosomes during meiosis is described. Although there is little bivalent formation, less than 5% of the Bs are lost during meiosis in anthers. There is, however, no evidence of B-chromosome accumulation in the offspring of controlled crosses, usually a slight loss, and Bs have deleterious effects on aspects of vigour and fertility. Thus, no satisfactory explanation for populations with up to 65% B-containing individuals has yet been found.

Unstable telocentric chromosome produced after centric misdivision of a 21q/21q translocated element

An unstable telocentric chromosome was found in an individual with Down's syndrome and an unusual chromosomal mosaic, 46,XX, t(21q21q)+,21-/46,XX,21q-/45,XX,21-. As the 21q/21q chromosome was of paternal origin, based on the characteristics of its centromeric heterochromatin and on the characteristics of both 21 chromosomes of the father, it was concluded that its formation involved centric breakage and loss of centromeric material. The cell line with the 21q- chromosome may have originated from the translocation by an asymmetric misdivision of the reduced centromeric material. Of the two telocentrics produced by this fracture, one, possessing the smaller amount of centromeric apparatus, would be immediately lost; the other would be retained, but complete activity of its centromere would not be restored. It would therefore be unstable and might be lost.

A human family suggesting evidence for centric fission and stability of a telocentric chromosome.

One of the female members of a family with hereditary D/D-translocation chromosome was found to be mosaic for two major cell lines. One of her cell lines was consistent with balanced D/D-translocation, i.e., 45, XX, D^–D^–, t (DqDq). Her other cell line had each cell with 46chromosomes. In these cells, however, a normal D-chromosome was replaced by a telocentric ‘marker’ chromosome with comparable size of the long arm of a D-chromosome. The mosaic carrier has transmitted this ‘marker’ chromosome to her son. It is interpreted that during the embryonic life of the female carrier, one of the cells with the translocation D/D-chromosome underwent centric misdivision that was followed by a pericentric inversion in one of the telocentrics. These two processes apparently gave to a stable telocentric D-chromosome and a regular D-group chromosome.

Karyotypic fissioning and Canid phylogeny

A theory is advanced which holds that mammalian evolution has been largely dependent on extensive chromosomal changes. These take the form of occasional simultaneous massive increase in diploid number through centric mis-division, here called karyotypic fissioning. This event occurs in a single individual and subsequently spreads through a population depending on chance, meiotic compatibility, dynamics of the population and natural selection. The possibility that fusions also occur sporadically is not precluded, although such events are not seen as abundant. This theory accounts for the wide variation in diploid number among the Mammalia and relates the presumed episodes of karyotypic fissioning with known periods of explosive speciation and adaptive radiation. Finally, the traditional evolutionary concept of mutant allele substitution through gene frequency shift under the influence of natural selection is placed in a new perspective. While it is still seen as a primary mechanism of evolution, it is seen as more significant as a “fine-tuning” mechanism, perhaps often responding to exigencies precipitated by chromosomal changes. The theory is tested by integrating its assumptions with the phylogeny of the recent Canidae as previously reconstructed from the fossil record and classical studies of anatomy. The correlation between increase in diploid number and adaptive radiation within this group is obvious. Within the family it is possible to specify approximate times and places of karyotypic fissioning which have been found to be consistent with known paleontogical and zoogeographic facts. It has also been possible to suggest relationships among living species which had not been previously recognized, but which are testable by independent techniques.