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Sample records for centromeres

  1. Dynamic epigenetic states of maize centromeres

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    Yalin eLiu

    2015-10-01

    Full Text Available The centromere is a specialized chromosomal region identified as the major constriction, upon which the kinetochore complex is formed, ensuring accurate chromosome orientation and segregation during cell division. The rapid evolution of centromere DNA sequence and the conserved centromere function are two contradictory aspects of centromere biology. Indeed, the sole presence of genetic sequence is not sufficient for centromere formation. Various dicentric chromosomes with one inactive centromere have been recognized. It has also been found that de novo centromere formation is common on fragments in which centromeric DNA sequences are lost. Epigenetic factors play important roles in centromeric chromatin assembly and maintenance. Nondisjunction of the supernumerary B chromosome early prophase of meiosis I requires an active centromere. This review discusses recent studies in maize about genetic and epigenetic elements regulating formation and maintenance of centromere chromatin, as well as centromere behavior in meiosis.

  2. Engineered human dicentric chromosomes show centromere plasticity.

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    Higgins, Anne W; Gustashaw, Karen M; Willard, Huntington F

    2005-01-01

    The centromere is essential for the faithful distribution of a cell's genetic material to subsequent generations. Despite intense scrutiny, the precise genetic and epigenetic basis for centromere function is still unknown. Here, we have used engineered dicentric human chromosomes to investigate mammalian centromere structure and function. We describe three classes of dicentric chromosomes isolated in different cell lines: functionally monocentric chromosomes, in which one of the two genetically identical centromeres is consistently inactivated; functionally dicentric chromosomes, in which both centromeres are consistently active; and dicentric chromosomes heterogeneous with respect to centromere activity. A study of serial single cell clones from heterogeneous cell lines revealed that while centromere activity is usually clonal, the centromere state (i.e. functionally monocentric or dicentric) in some lines can switch within a growing population of cells. Because pulsed field gel analysis indicated that the DNA at the centromeres of these chromosomes did not change detectably, this switching of the centromere state is most likely due to epigenetic changes. Inactivation of one of the two active centromeres in a functionally dicentric chromosome was observed in a percentage of cells after treatment with Trichostatin A, an inhibitor of histone deacetylation. This study provides evidence that the activity of human centromeres, while largely stable, can be subject to dynamic change, most likely due to epigenetic modification.

  3. Centromere domain organization and histone modifications

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    P. Bjerling

    2002-05-01

    Full Text Available Centromere function requires the proper coordination of several subfunctions, such as kinetochore assembly, sister chromatid cohesion, binding of kinetochore microtubules, orientation of sister kinetochores to opposite spindle poles, and their movement towards the spindle poles. Centromere structure appears to be organized in different, separable domains in order to accomplish these functions. Despite the conserved nature of centromere functions, the molecular genetic definition of the DNA sequences that form a centromere in the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, in the fruit fly Drosophila melanogaster, and in humans has revealed little conservation at the level of centromere DNA sequences. Also at the protein level few centromere proteins are conserved in all of these four organisms and many are unique to the different organisms. The recent analysis of the centromere structure in the yeast S. pombe by electron microscopy and detailed immunofluorescence microscopy of Drosophila centromeres have brought to light striking similarities at the overall structural level between these centromeres and the human centromere. The structural organization of the centromere is generally multilayered with a heterochromatin domain and a central core/inner plate region, which harbors the outer plate structures of the kinetochore. It is becoming increasingly clear that the key factors for assembly and function of the centromere structure are the specialized histones and modified histones which are present in the centromeric heterochromatin and in the chromatin of the central core. Thus, despite the differences in the DNA sequences and the proteins that define a centromere, there is an overall structural similarity between centromeres in evolutionarily diverse eukaryotes.

  4. Structure, Function, and Evolution of Rice Centromeres

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    Jiang, Jiming

    2010-02-04

    The centromere is the most characteristic landmark of eukaryotic chromosomes. Centromeres function as the site for kinetochore assembly and spindle attachment, allowing for the faithful pairing and segregation of sister chromatids during cell division. Characterization of centromeric DNA is not only essential to understand the structure and organization of plant genomes, but it is also a critical step in the development of plant artificial chromosomes. The centromeres of most model eukaryotic species, consist predominantly of long arrays of satellite DNA. Determining the precise DNA boundary of a centromere has proven to be a difficult task in multicellular eukaryotes. We have successfully cloned and sequenced the centromere of rice chromosome 8 (Cen8), representing the first fully sequenced centromere from any multicellular eukaryotes. The functional core of Cen8 spans ~800 kb of DNA, which was determined by chromatin immunoprecipitation (ChIP) using an antibody against the rice centromere-specific H3 histone. We discovered 16 actively transcribed genes distributed throughout the Cen8 region. In addition to Cen8, we have characterized eight additional rice centromeres using the next generation sequencing technology. We discovered four subfamilies of the CRR retrotransposon that is highly enriched in rice centromeres. CRR elements are constitutively transcribed and different CRR subfamilies are differentially processed by RNAi. These results suggest that different CRR subfamilies may play different roles in the RNAi-mediated pathway for formation and maintenance of centromeric chromatin.

  5. Synaptonemal complex components persist at centromeres and are required for homologous centromere pairing in mouse spermatocytes.

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    C Gaston Bisig

    2012-06-01

    Full Text Available Recent studies in simple model organisms have shown that centromere pairing is important for ensuring high-fidelity meiotic chromosome segregation. However, this process and the mechanisms regulating it in higher eukaryotes are unknown. Here we present the first detailed study of meiotic centromere pairing in mouse spermatogenesis and link it with key events of the G2/metaphase I transition. In mouse we observed no evidence of the persistent coupling of centromeres that has been observed in several model organisms. We do however find that telomeres associate in non-homologous pairs or small groups in B type spermatogonia and pre-leptotene spermatocytes, and this association is disrupted by deletion of the synaptonemal complex component SYCP3. Intriguingly, we found that, in mid prophase, chromosome synapsis is not initiated at centromeres, and centromeric regions are the last to pair in the zygotene-pachytene transition. In late prophase, we first identified the proteins that reside at paired centromeres. We found that components of the central and lateral element and transverse filaments of the synaptonemal complex are retained at paired centromeres after disassembly of the synaptonemal complex along diplotene chromosome arms. The absence of SYCP1 prevents centromere pairing in knockout mouse spermatocytes. The localization dynamics of SYCP1 and SYCP3 suggest that they play different roles in promoting homologous centromere pairing. SYCP1 remains only at paired centromeres coincident with the time at which some kinetochore proteins begin loading at centromeres, consistent with a role in assembly of meiosis-specific kinetochores. After removal of SYCP1 from centromeres, SYCP3 then accumulates at paired centromeres where it may promote bi-orientation of homologous centromeres. We propose that, in addition to their roles as synaptonemal complex components, SYCP1 and SYCP3 act at the centromeres to promote the establishment and/or maintenance of

  6. A Positive Twist to the Centromeric Nucleosome

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    Josefina Ocampo

    2015-10-01

    Full Text Available Centromeric nucleosomes are critical for chromosome attachment to the mitotic spindle. In this issue of Cell Reports, Diaz-Ingelmo et al. (2015 propose that the yeast centromeric nucleosome is stabilized by a positively supercoiled loop formed by the sequence-specific CBF3 complex.

  7. Insights into centromeric transcription in mitosis.

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    Liu, Hong

    2016-01-01

    The major role of RNA polymerase II (RNAP II) is to generate mRNAs. I recently uncovered a novel function of RNAP II in chromosome segregation in mitosis, installing the cohesin protector, Shugoshin, at centromeres. Here I will discuss the current understanding of RNAP II-dependent centromeric transcription in mitosis.

  8. Chromatin ring formation at plant centromeres

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    Veit eSchubert

    2016-02-01

    Full Text Available We observed the formation of chromatin ring structures at centromeres of somatic rye and Arabidopsis chromosomes. To test whether this behavior is present also in other plant species and tissues we analyzed Arabidopsis, rye, wheat, Aegilops and barley centromeres during cell divisions and in interphase nuclei by immunostaining and FISH. Furthermore, structured illumination microscopy (super-resolution was applied to investigate the ultrastructure of centromere chromatin beyond the classical refraction limit of light. It became obvious, that a ring formation at centromeres may appear during mitosis, meiosis and in interphase nuclei in all species analyzed. However, varying centromere structures, as ring formations or globular organized chromatin fibers, were identified in different tissues of one and the same species. In addition, we found that a chromatin ring formation may also be caused by subtelomeric repeats in barley. Thus, we conclude that the formation of chromatin rings may appear in different plant species and tissues, but that it is not specific for centromere function. Based on our findings we established a model describing the ultrastructure of plant centromeres and discuss it in comparison to previous models proposed for animals and plants.

  9. Dicentric Chromosome Formation and Epigenetics of Centromere Formation in Plants

    Institute of Scientific and Technical Information of China (English)

    Shulan Fu; Zhi Gao; James Birchler; Fangpu Han

    2012-01-01

    Plant centromeres are generally composed of tandem arrays of simple repeats that form a complex chromosome locus where the kinetochore forms and microtubules attach during mitosis and meiosis.Each chromosome has one centromere region,which is essential for accurate division of the genetic material.Recently,chromosomes containing two centromere regions (called dicentric chromosomes)have been found in maize and wheat.Interestingly,some dicentric chromosomes are stable because only one centromere is active and the other one is inactivated.Because such arrays maintain their typical structure for both active and inactive centromeres,the specification of centromere activity has an epigenetic component independent of the DNA sequence.Under some circumstances,the inactive centromeres may recover centromere function,which is called centromere reactivation.Recent studies have highlighted the important changes,such as DNA methylation and histone modification,that occur during centromere inactivation and reactivation.

  10. Dicentric chromosome formation and epigenetics of centromere formation in plants.

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    Fu, Shulan; Gao, Zhi; Birchler, James; Han, Fangpu

    2012-03-20

    Plant centromeres are generally composed of tandem arrays of simple repeats that form a complex chromosome locus where the kinetochore forms and microtubules attach during mitosis and meiosis. Each chromosome has one centromere region, which is essential for accurate division of the genetic material. Recently, chromosomes containing two centromere regions (called dicentric chromosomes) have been found in maize and wheat. Interestingly, some dicentric chromosomes are stable because only one centromere is active and the other one is inactivated. Because such arrays maintain their typical structure for both active and inactive centromeres, the specification of centromere activity has an epigenetic component independent of the DNA sequence. Under some circumstances, the inactive centromeres may recover centromere function, which is called centromere reactivation. Recent studies have highlighted the important changes, such as DNA methylation and histone modification, that occur during centromere inactivation and reactivation.

  11. Atypical centromeres in plants – what they can tell us

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    Maria eCuacos

    2015-10-01

    Full Text Available The centromere, visible as the primary constriction of condensed metaphase chromosomes, is a defined chromosomal locus essential for genome stability. It mediates transient assembly of a multi-protein complex, the kinetochore, which enables interaction with spindle fibers and thus faithful segregation of the genetic information during nuclear divisions. Centromeric DNA varies in extent and sequence composition among organisms, but a common feature of almost all active eukaryotic centromeres is the presence of the centromeric histone H3 variant cenH3 (a.k.a. CENP-A.These typical centromere features apply to most studied species. However, a number of species display atypical centromeres, such as holocentromeres (centromere extension along almost the entire chromatid length or neocentromeres (ectopic centromere activity.In this review, we provide an overview of different atypical centromere types found in plants including holocentromeres, de novo formed centromeres and terminal neocentromeres as well as di-, tri- and metapolycentromeres (more than one centromere per chromosomes. We discuss their specific and common features and compare them to centromere types found in other eukaryotic species. We also highlight new insights into centromere biology gained in plants with atypical centromeres such as distinct mechanisms to define a holocentromere, specific adaptations in species with holocentromeres during meiosis or various scenarios leading to neocentromere formation.

  12. RNAi and heterochromatin repress centromeric meiotic recombination

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    Ellermeier, Chad; Higuchi, Emily C; Phadnis, Naina

    2010-01-01

    During meiosis, the formation of viable haploid gametes from diploid precursors requires that each homologous chromosome pair be properly segregated to produce an exact haploid set of chromosomes. Genetic recombination, which provides a physical connection between homologous chromosomes, is essen......During meiosis, the formation of viable haploid gametes from diploid precursors requires that each homologous chromosome pair be properly segregated to produce an exact haploid set of chromosomes. Genetic recombination, which provides a physical connection between homologous chromosomes....... Surprisingly, one mutant derepressed for recombination in the heterochromatic mating-type region during meiosis and several mutants derepressed for centromeric gene expression during mitotic growth are not derepressed for centromeric recombination during meiosis. These results reveal a complex relation between...... types of repression by heterochromatin. Our results also reveal a previously undemonstrated role for RNAi and heterochromatin in the repression of meiotic centromeric recombination and, potentially, in the prevention of birth defects by maintenance of proper chromosome segregation during meiosis....

  13. Role of transcription at centromeres in budding yeast.

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    Ohkuni, Kentaro; Kitagawa, Katsumi

    2012-01-01

    Centromeres are specialized chromosomal loci that are essential for proper chromosome segregation. Recent data show that a certain level of active transcription, regulated by transcription factors Cbf1 and Ste12, makes a direct contribution to centromere function in Saccharomyces cerevisiae. Here, we discuss the requirement and function of transcription at centromeres.

  14. Assembly of Drosophila centromeric nucleosomes requires CID dimerization.

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    Zhang, Weiguo; Colmenares, Serafin U; Karpen, Gary H

    2012-01-27

    Centromeres are essential chromosomal regions required for kinetochore assembly and chromosome segregation. The composition and organization of centromeric nucleosomes containing the essential histone H3 variant CENP-A (CID in Drosophila) is a fundamental, unresolved issue. Using immunoprecipitation of CID mononucleosomes and cysteine crosslinking, we demonstrate that centromeric nucleosomes contain CID dimers in vivo. Furthermore, CID dimerization and centromeric targeting require a residue implicated in formation of the four-helix bundle, which mediates intranucleosomal H3 dimerization and nucleosome integrity. Taken together, our findings suggest that CID nucleosomes are octameric in vivo and that CID dimerization is essential for correct centromere assembly.

  15. Stretching the rules: monocentric chromosomes with multiple centromere domains.

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    Neumann, Pavel; Navrátilová, Alice; Schroeder-Reiter, Elizabeth; Koblížková, Andrea; Steinbauerová, Veronika; Chocholová, Eva; Novák, Petr; Wanner, Gerhard; Macas, Jiří

    2012-01-01

    The centromere is a functional chromosome domain that is essential for faithful chromosome segregation during cell division and that can be reliably identified by the presence of the centromere-specific histone H3 variant CenH3. In monocentric chromosomes, the centromere is characterized by a single CenH3-containing region within a morphologically distinct primary constriction. This region usually spans up to a few Mbp composed mainly of centromere-specific satellite DNA common to all chromosomes of a given species. In holocentric chromosomes, there is no primary constriction; the centromere is composed of many CenH3 loci distributed along the entire length of a chromosome. Using correlative fluorescence light microscopy and high-resolution electron microscopy, we show that pea (Pisum sativum) chromosomes exhibit remarkably long primary constrictions that contain 3-5 explicit CenH3-containing regions, a novelty in centromere organization. In addition, we estimate that the size of the chromosome segment delimited by two outermost domains varies between 69 Mbp and 107 Mbp, several factors larger than any known centromere length. These domains are almost entirely composed of repetitive DNA sequences belonging to 13 distinct families of satellite DNA and one family of centromeric retrotransposons, all of which are unevenly distributed among pea chromosomes. We present the centromeres of Pisum as novel "meta-polycentric" functional domains. Our results demonstrate that the organization and DNA composition of functional centromere domains can be far more complex than previously thought, do not require single repetitive elements, and do not require single centromere domains in order to segregate properly. Based on these findings, we propose Pisum as a useful model for investigation of centromere architecture and the still poorly understood role of repetitive DNA in centromere evolution, determination, and function.

  16. Evolution of Centromeric Retrotransposons in Grasses

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    Sharma, Anupma; Presting, Gernot G.

    2014-01-01

    Centromeric retrotransposons (CRs) constitute a family of plant retroelements, some of which have the ability to target their insertion almost exclusively to the functional centromeres. Our exhaustive analysis of CR family members in four grass genomes revealed not only horizontal transfer (HT) of CR elements between the oryzoid and panicoid grass lineages but also their subsequent recombination with endogenous elements that in some cases created prolific recombinants in foxtail millet and sorghum. HT events are easily identifiable only in cases where host genome divergence significantly predates HT, thus documented HT events likely represent only a fraction of the total. If the more difficult to detect ancient HT events occurred at frequencies similar to those observable in present day grasses, the extant long terminal repeat retrotransposons represent the mosaic products of HT and recombination that are optimized for retrotransposition in their host genomes. This complicates not only phylogenetic analysis but also the establishment of a meaningful retrotransposon nomenclature, which we have nevertheless attempted to implement here. In contrast to the plant-centric naming convention used currently for CR elements, we classify elements primarily based on their phylogenetic relationships regardless of host plant, using the exhaustively studied maize elements assigned to six different subfamilies as a standard. The CR2 subfamily is the most widely distributed of the six CR subfamilies discovered in grass genomes to date and thus the most likely to play a functional role at grass centromeres. PMID:24814286

  17. Switching the centromeres on and off: epigenetic chromatin alterations provide plasticity in centromere activity stabilizing aberrant dicentric chromosomes.

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    Sato, Hiroshi; Saitoh, Shigeaki

    2013-12-01

    The kinetochore, which forms on a specific chromosomal locus called the centromere, mediates interactions between the chromosome and the spindle during mitosis and meiosis. Abnormal chromosome rearrangements and/or neocentromere formation can cause the presence of multiple centromeres on a single chromosome, which results in chromosome breakage or cell cycle arrest. Analyses of artificial dicentric chromosomes suggested that the activity of the centromere is regulated epigenetically; on some stably maintained dicentric chromosomes, one of the centromeres no longer functions as a platform for kinetochore formation, although the DNA sequence remains intact. Such epigenetic centromere inactivation occurs in cells of various eukaryotes harbouring 'regional centromeres', such as those of maize, fission yeast and humans, suggesting that the position of the active centromere is determined by epigenetic markers on a chromosome rather than the nucleotide sequence. Our recent findings in fission yeast revealed that epigenetic centromere inactivation consists of two steps: disassembly of the kinetochore initiates inactivation and subsequent heterochromatinization prevents revival of the inactivated centromere. Kinetochore disassembly followed by heterochromatinization is also observed in normal senescent human cells. Thus epigenetic centromere inactivation may not only stabilize abnormally generated dicentric chromosomes, but also be part of an intrinsic mechanism regulating cell proliferation.

  18. Strong nucleosomes of A. thaliana concentrate in centromere regions.

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    Salih, Bilal; Trifonov, Edward N

    2015-01-01

    Earlier identified strongest nucleosome DNA sequences of A. thaliana, those with visible 10-11 base sequence periodicity, are mapped along chromosomes. Resulting positional distributions reveal distinct maxima, one per chromosome, located in the centromere regions. Sequence-directed nucleosome mapping demonstrates that the strong nucleosomes (SNs) make tight arrays, several 'parallel' nucleosomes each, suggesting a columnar chromatin structure. The SNs represent a new class of centromeric nucleosomes, presumably, participating in synapsis of chromatids and securing the centromere architecture.

  19. Identification of novel Drosophila centromere-associated proteins.

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    Barth, Teresa K; Schade, Georg O M; Schmidt, Andreas; Vetter, Irene; Wirth, Marc; Heun, Patrick; Thomae, Andreas W; Imhof, Axel

    2014-10-01

    Centromeres are chromosomal regions crucial for correct chromosome segregation during mitosis and meiosis. They are epigenetically defined by centromeric proteins such as the centromere-specific histone H3-variant centromere protein A (CENP-A). In humans, 16 additional proteins have been described to be constitutively associated with centromeres throughout the cell cycle, known as the constitutive centromere-associated network (CCAN). In contrast, only one additional constitutive centromeric protein is known in Drosophila melanogaster (D.mel), the conserved CCAN member CENP-C. To gain further insights into D.mel centromere composition and biology, we analyzed affinity-purified chromatin prepared from D.mel cell lines expressing green fluorescent protein tagged histone three variants by MS. In addition to already-known centromeric proteins, we identified novel factors that were repeatedly enriched in affinity purification-MS experiments. We analyzed the cellular localization of selected candidates by immunocytochemistry and confirmed localization to the centromere and other genomic regions for ten factors. Furthermore, RNA interference mediated depletion of CG2051, CG14480, and hyperplastic discs, three of our strongest candidates, leads to elevated mitotic defects. Knockdowns of these candidates neither impair the localization of several known kinetochore proteins nor CENP-A(CID) loading, suggesting their involvement in alternative pathways that contribute to proper centromere function. In summary, we provide a comprehensive analysis of the proteomic composition of Drosophila centromeres. All MS data have been deposited in the ProteomeXchange with identifier PXD000758 (http://proteomecentral.proteomexchange.org/dataset/PXD000758).

  20. Centromere activity in dicentric small supernumerary marker chromosomes.

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    Ewers, Elisabeth; Yoda, Kinya; Hamid, Ahmed B; Weise, Anja; Manvelyan, Marina; Liehr, Thomas

    2010-07-01

    Twenty-five dicentric small supernumerary marker chromosomes (sSMC) derived from #13/21, #14, #15, #18, and #22 were studied by immunohistochemistry for their centromeric activity. Centromere protein (CENP)-B was applied as marker for all centromeres and CENP-C to label the active ones. Three different 'predominant' activation patterns could be observed, i.e., centric fusion or either only one or all two centromeres were active. In one inherited case, the same activation pattern was found in mother and son. In acrocentric-derived sSMC, all three activation patterns could be present. In contrary, in chromosome 18-derived sSMC, only the fusion type was observed. In concordance with previous studies a certain centromeric plasticity was observed in up to 13% of the cells of an individual case. Surprisingly, the obtained data suggests a possible influence of the sSMC carrier's gender on the implementation of the predominant activation pattern; especially, only one active centromere was found more frequently in female than in male carriers. Also, it might be suggested that dicentric sSMC with one active centromere could be less stable than such with two active ones-centromeric plasticity might have an influence here, as well. Also, centromere activity in acrocentric-derived dicentrics could be influenced by heteromorphisms of the corresponding short arms. Finally, evidence is provided that the closer the centromeres of a dicentric are and if they are not fused, the more likely it was that both of them became active. In concordance and refinement with previous studies, a distance of 1.4 Mb up to about 13 Mb the two active centromere state was favored, while centromeric distance of over approximately 15 Mb lead to inactivation of one centromere. Overall, here, the first and largest ever undertaken study in dicentric sSMC is presented, providing evidence that the centromeric activation pattern is, and parental origin may be of interest for their biology. Influence of

  1. Centromere positions in chicken and Japanese quail chromosomes: de novo centromere formation versus pericentric inversions

    NARCIS (Netherlands)

    Zlotina, A.; Galkina, S.; Krasikova, A.; Crooijmans, R.P.M.A.; Groenen, M.; Gaginskaya, E.; Deryusheva, S.

    2012-01-01

    Chicken (Gallus gallus domesticus, GGA) and Japanese quail (Coturnix coturnix japonica, CCO) karyotypes are very similar. They have identical chromosome number (2n = 78) and show a high degree of synteny. Centromere positions on the majority of orthologous chromosomes are different in these two spec

  2. Neocentrics and holokinetics (holocentrics): chromosomes out of the centromeric rules.

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    Guerra, M; Cabral, G; Cuacos, M; González-García, M; González-Sánchez, M; Vega, J; Puertas, M J

    2010-07-01

    The centromere appears as a single constriction at mitotic metaphase in most eukaryotic chromosomes. Holokinetic chromosomes are the exception to this rule because they do not show any centromeric constrictions. Holokinetic chromosomes are usually forgotten in most reviews about centromeres, despite their presence in a number of animal and plant species. They are generally linked to very intriguing and unusual mechanisms of mitosis and meiosis. Holokinetic chromosomes differ from monocentric chromosomes not only in the extension of the kinetochore plate, but also in many other peculiar karyological features, which could be understood as the 'holokinetic syndrome' that is reviewed in detail. Together with holokinetic chromosomes we review neocentromeric activity, a similarly intriguing case of regions able to pull chromosomes towards the poles without showing the main components reported to be essential to centromeric function. A neocentromere is a chromosomal region different from the true centromere in structure, DNA sequence and location, but is able to lead chromosomes to the cell poles in special circumstances. Neocentromeres have been reported in plants and animals showing different features. Both in humans and Drosophila, neocentric activity appears in somatic cells with defective chromosomes lacking a functional centromere. In most cases in plants, neocentromeres appear in chromosomes which have normal centromeres, but are active only during meiosis. Because of examples such as spontaneous or induced neocentromeres and holokinetic chromosomes, it is becoming less surprising that different structures and DNA sequences of centromeres appear in evolution.

  3. The Past, Present, and Future of Human Centromere Genomics

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    Megan E. Aldrup-MacDonald

    2014-01-01

    Full Text Available The centromere is the chromosomal locus essential for chromosome inheritance and genome stability. Human centromeres are located at repetitive alpha satellite DNA arrays that compose approximately 5% of the genome. Contiguous alpha satellite DNA sequence is absent from the assembled reference genome, limiting current understanding of centromere organization and function. Here, we review the progress in centromere genomics spanning the discovery of the sequence to its molecular characterization and the work done during the Human Genome Project era to elucidate alpha satellite structure and sequence variation. We discuss exciting recent advances in alpha satellite sequence assembly that have provided important insight into the abundance and complex organization of this sequence on human chromosomes. In light of these new findings, we offer perspectives for future studies of human centromere assembly and function.

  4. HACking the centromere chromatin code: insights from human artificial chromosomes.

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    Bergmann, Jan H; Martins, Nuno M C; Larionov, Vladimir; Masumoto, Hiroshi; Earnshaw, William C

    2012-07-01

    The centromere is a specialized chromosomal region that serves as the assembly site of the kinetochore. At the centromere, CENP-A nucleosomes form part of a chromatin landscape termed centrochromatin. This chromatin environment conveys epigenetic marks regulating kinetochore formation. Recent work sheds light on the intricate relationship between centrochromatin state, the CENP-A assembly pathway and the maintenance of centromere function. Here, we review the emerging picture of how chromatin affects mammalian kinetochore formation. We place particular emphasis on data obtained from Human Artificial Chromosome (HAC) biology and the targeted engineering of centrochromatin using synthetic HACs. We discuss implications of these findings, which indicate that a delicate balance of histone modifications and chromatin state dictates both de novo centromere formation and the maintenance of centromere identity in dividing cell populations.

  5. Validation of the micronucleus-centromere assay for biological dosimetry

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    Wojcik A.

    2000-01-01

    Full Text Available The micronucleus assay is frequently used for purposes of biological dosimetry. Due to high interindividual variability in the spontaneous frequency of micronuclei, its sensitivity in the low dose region is poor. It has been suggested that this problem can be mitigated by selectively analyzing the frequency of those micronuclei which contain only acentric fragments. Using a pan-centromeric FISH probe we have studied the dose dependence of micronuclei with centromeres in peripheral lymphocytes of human donors. In contrast to previous publications, our approach is based on determining the relative frequency of micronuclei with and without centromeric signals. Our results confirm previous observations that in the low dose range of ionizing radiation, the micronucleus-centromere assay is more sensitive than the conventional micronucleus test.

  6. Dicentric chromosomes: unique models to study centromere function and inactivation

    OpenAIRE

    Kaitlin M Stimpson; Matheny, Justyne E.; Sullivan, Beth A.

    2012-01-01

    Dicentric chromosomes are products of genome rearrangement that place two centromeres on the same chromosome. Depending on the organism, dicentric stability varies after formation. In humans, dicentrics occur naturally in a substantial portion of the population and usually segregate successfully in mitosis and meiosis. Their stability has been attributed to inactivation of one of the two centromeres, creating a functionally monocentric chromosome that can segregate normally during cell divisi...

  7. Tetrameric structure of centromeric nucleosomes in interphase Drosophila cells.

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    Yamini Dalal

    2007-08-01

    Full Text Available Centromeres, the specialized chromatin structures that are responsible for equal segregation of chromosomes at mitosis, are epigenetically maintained by a centromere-specific histone H3 variant (CenH3. However, the mechanistic basis for centromere maintenance is unknown. We investigated biochemical properties of CenH3 nucleosomes from Drosophila melanogaster cells. Cross-linking of CenH3 nucleosomes identifies heterotypic tetramers containing one copy of CenH3, H2A, H2B, and H4 each. Interphase CenH3 particles display a stable association of approximately 120 DNA base pairs. Purified centromeric nucleosomal arrays have typical "beads-on-a-string" appearance by electron microscopy but appear to resist condensation under physiological conditions. Atomic force microscopy reveals that native CenH3-containing nucleosomes are only half as high as canonical octameric nucleosomes are, confirming that the tetrameric structure detected by cross-linking comprises the entire interphase nucleosome particle. This demonstration of stable half-nucleosomes in vivo provides a possible basis for the instability of centromeric nucleosomes that are deposited in euchromatic regions, which might help maintain centromere identity.

  8. Multilocus analysis for gene-centromere mapping using first polar bodies and secondary oocytes

    Energy Technology Data Exchange (ETDEWEB)

    Da, Y.; Jarrell, V.L.; Wang, T.; Fernando, R.L.; Wheeler, M.B.; Lewin, H.A. [Univ. of Illinois, Urbana, IL (United States)

    1995-02-01

    Polar body and oocyte typing is a new technique for gene-centromere mapping and for generating female linkage maps. A maximum likelihood approach is presented for ordering multiple markers relative to the centromere and for estimating recombination frequencies between markers and between the centromere and marker loci. Three marker-centromere orders are possible for each pair of markers: two orders when the centromere flanks the two markers and one order when the centromere is flanked by the two markers. For each possible order, the likelihood was expressed as a function of recombination frequencies for two adjacent intervals. LOD score for recombination frequency between markers or between the centromere and a marker locus was derived based on the likelihood for each gene-centromere order. The methods developed herein provide a general solution to the problem of multilocus gene-centromere mapping that involves all theoretical crossover possibilities, including four-strand double crossovers. 24 refs., 2 figs., 2 tabs.

  9. Adaptive evolution of centromere proteins in plants and animals

    Directory of Open Access Journals (Sweden)

    Henikoff Steven

    2004-08-01

    Full Text Available Abstract Background Centromeres represent the last frontiers of plant and animal genomics. Although they perform a conserved function in chromosome segregation, centromeres are typically composed of repetitive satellite sequences that are rapidly evolving. The nucleosomes of centromeres are characterized by a special H3-like histone (CenH3, which evolves rapidly and adaptively in Drosophila and Arabidopsis. Most plant, animal and fungal centromeres also bind a large protein, centromere protein C (CENP-C, that is characterized by a single 24 amino-acid motif (CENPC motif. Results Whereas we find no evidence that mammalian CenH3 (CENP-A has been evolving adaptively, mammalian CENP-C proteins contain adaptively evolving regions that overlap with regions of DNA-binding activity. In plants we find that CENP-C proteins have complex duplicated regions, with conserved amino and carboxyl termini that are dissimilar in sequence to their counterparts in animals and fungi. Comparisons of Cenpc genes from Arabidopsis species and from grasses revealed multiple regions that are under positive selection, including duplicated exons in some grasses. In contrast to plants and animals, yeast CENP-C (Mif2p is under negative selection. Conclusions CENP-Cs in all plant and animal lineages examined have regions that are rapidly and adaptively evolving. To explain these remarkable evolutionary features for a single-copy gene that is needed at every mitosis, we propose that CENP-Cs, like some CenH3s, suppress meiotic drive of centromeres during female meiosis. This process can account for the rapid evolution and the complexity of centromeric DNA in plants and animals as compared to fungi.

  10. No longer a nuisance: long non-coding RNAs join CENP-A in epigenetic centromere regulation.

    Science.gov (United States)

    Rošić, Silvana; Erhardt, Sylvia

    2016-04-01

    Centromeres represent the basis for kinetochore formation, and are essential for proper chromosome segregation during mitosis. Despite these essential roles, centromeres are not defined by specific DNA sequences, but by epigenetic means. The histone variant CENP-A controls centromere identity epigenetically and is essential for recruiting kinetochore components that attach the chromosomes to the mitotic spindle during mitosis. Recently, a new player in centromere regulation has emerged: long non-coding RNAs transcribed from repetitive regions of centromeric DNA function in regulating centromeres epigenetically. This review summarizes recent findings on the essential roles that transcription, pericentromeric transcripts, and centromere-derived RNAs play in centromere biology.

  11. Chromosomes. CENP-C reshapes and stabilizes CENP-A nucleosomes at the centromere.

    Science.gov (United States)

    Falk, Samantha J; Guo, Lucie Y; Sekulic, Nikolina; Smoak, Evan M; Mani, Tomoyasu; Logsdon, Glennis A; Gupta, Kushol; Jansen, Lars E T; Van Duyne, Gregory D; Vinogradov, Sergei A; Lampson, Michael A; Black, Ben E

    2015-05-01

    Inheritance of each chromosome depends upon its centromere. A histone H3 variant, centromere protein A (CENP-A), is essential for epigenetically marking centromere location. We find that CENP-A is quantitatively retained at the centromere upon which it is initially assembled. CENP-C binds to CENP-A nucleosomes and is a prime candidate to stabilize centromeric chromatin. Using purified components, we find that CENP-C reshapes the octameric histone core of CENP-A nucleosomes, rigidifies both surface and internal nucleosome structure, and modulates terminal DNA to match the loose wrap that is found on native CENP-A nucleosomes at functional human centromeres. Thus, CENP-C affects nucleosome shape and dynamics in a manner analogous to allosteric regulation of enzymes. CENP-C depletion leads to rapid removal of CENP-A from centromeres, indicating their collaboration in maintaining centromere identity.

  12. Three wise centromere functions: see no error, hear no break, speak no delay.

    OpenAIRE

    Tanaka, Tomoyuki U; Clayton, Lesley; Natsume, Toyoaki

    2013-01-01

    The main function of the centromere is to promote kinetochore assembly for spindle microtubule attachment. Two additional functions of the centromere, however, are becoming increasingly clear: facilitation of robust sister-chromatid cohesion at pericentromeres and advancement of replication of centromeric regions. The combination of these three centromere functions ensures correct chromosome segregation during mitosis. Here, we review the mechanisms of the kinetochore–microtubule interaction,...

  13. The cotton centromere contains a Ty3-gypsy-like LTR retroelement.

    Directory of Open Access Journals (Sweden)

    Song Luo

    Full Text Available The centromere is a repeat-rich structure essential for chromosome segregation; with the long-term aim of understanding centromere structure and function, we set out to identify cotton centromere sequences. To isolate centromere-associated sequences from cotton, (Gossypium hirsutum we surveyed tandem and dispersed repetitive DNA in the genus. Centromere-associated elements in other plants include tandem repeats and, in some cases, centromere-specific retroelements. Examination of cotton genomic survey sequences for tandem repeats yielded sequences that did not localize to the centromere. However, among the repetitive sequences we also identified a gypsy-like LTR retrotransposon (Centromere Retroelement Gossypium, CRG that localizes to the centromere region of all chromosomes in domestic upland cotton, Gossypium hirsutum, the major commercially grown cotton. The location of the functional centromere was confirmed by immunostaining with antiserum to the centromere-specific histone CENH3, which co-localizes with CRG hybridization on metaphase mitotic chromosomes. G. hirsutum is an allotetraploid composed of A and D genomes and CRG is also present in the centromere regions of other AD cotton species. Furthermore, FISH and genomic dot blot hybridization revealed that CRG is found in D-genome diploid cotton species, but not in A-genome diploid species, indicating that this retroelement may have invaded the A-genome centromeres during allopolyploid formation and amplified during evolutionary history. CRG is also found in other diploid Gossypium species, including B and E2 genome species, but not in the C, E1, F, and G genome species tested. Isolation of this centromere-specific retrotransposon from Gossypium provides a probe for further understanding of centromere structure, and a tool for future engineering of centromere mini-chromosomes in this important crop species.

  14. SHUGOSHINs and PATRONUS protect meiotic centromere cohesion in Arabidopsis thaliana.

    Science.gov (United States)

    Zamariola, Linda; De Storme, Nico; Vannerum, Katrijn; Vandepoele, Klaas; Armstrong, Susan J; Franklin, F Christopher H; Geelen, Danny

    2014-03-01

    In meiosis, chromosome cohesion is maintained by the cohesin complex, which is released in a two-step manner. At meiosis I, the meiosis-specific cohesin subunit Rec8 is cleaved by the protease Separase along chromosome arms, allowing homologous chromosome segregation. Next, in meiosis II, cleavage of the remaining centromere cohesin results in separation of the sister chromatids. In eukaryotes, protection of centromeric cohesion in meiosis I is mediated by SHUGOSHINs (SGOs). The Arabidopsis genome contains two SGO homologs. Here we demonstrate that Atsgo1 mutants show a premature loss of cohesion of sister chromatid centromeres at anaphase I and that AtSGO2 partially rescues this loss of cohesion. In addition to SGOs, we characterize PATRONUS which is specifically required for the maintenance of cohesion of sister chromatid centromeres in meiosis II. In contrast to the Atsgo1 Atsgo2 double mutant, patronus T-DNA insertion mutants only display loss of sister chromatid cohesion after meiosis I, and additionally show disorganized spindles, resulting in defects in chromosome segregation in meiosis. This leads to reduced fertility and aneuploid offspring. Furthermore, we detect aneuploidy in sporophytic tissue, indicating a role for PATRONUS in chromosome segregation in somatic cells. Thus, ploidy stability is preserved in Arabidopsis by PATRONUS during both meiosis and mitosis.

  15. Recombination patterns reveal information about centromere location on linkage maps

    DEFF Research Database (Denmark)

    Limborg, Morten T.; McKinney, Garrett J.; Seeb, Lisa W.

    2016-01-01

    . mykiss) characterized by low and unevenly distributed recombination – a general feature of male meiosis in many species. Further, a high frequency of double crossovers along chromosome arms in barley reduced resolution for locating centromeric regions on most linkage groups. Despite these limitations...

  16. Dicentric chromosomes: unique models to study centromere function and inactivation.

    Science.gov (United States)

    Stimpson, Kaitlin M; Matheny, Justyne E; Sullivan, Beth A

    2012-07-01

    Dicentric chromosomes are products of genome rearrangement that place two centromeres on the same chromosome. Depending on the organism, dicentric stability varies after formation. In humans, dicentrics occur naturally in a substantial portion of the population and usually segregate successfully in mitosis and meiosis. Their stability has been attributed to inactivation of one of the two centromeres, creating a functionally monocentric chromosome that can segregate normally during cell division. The molecular basis for centromere inactivation is not well understood, although studies in model organisms and in humans suggest that genomic and epigenetic mechanisms can be involved. Furthermore, constitutional dicentric chromosomes ascertained in patients presumably represent the most stable chromosomes, so the spectrum of dicentric fates, if it exists, is not entirely clear. Studies of engineered or induced dicentrics in budding yeast and plants have provided significant insight into the fate of dicentric chromosomes. And, more recently, studies have shown that dicentrics in humans can also undergo multiple fates after formation. Here, we discuss current experimental evidence from various organisms that has deepened our understanding of dicentric behavior and the intriguingly complex process of centromere inactivation.

  17. High-resolution mapping and transcriptional activity analysis of chicken centromere sequences on giant lampbrush chromosomes.

    Science.gov (United States)

    Krasikova, Alla; Fukagawa, Tatsuo; Zlotina, Anna

    2012-12-01

    Exploration into morphofunctional organisation of centromere DNA sequences is important for understanding the mechanisms of kinetochore specification and assembly. In-depth epigenetic analysis of DNA fragments associated with centromeric nucleosome proteins has demonstrated unique features of centromere organisation in chicken karyotype: there are both mature centromeres, which comprise chromosome-specific homogeneous arrays of tandem repeats, and recently evolved primitive centromeres, which consist of non-tandemly organised DNA sequences. In this work, we describe the arrangement and transcriptional activity of chicken centromere repeats for Cen1, Cen2, Cen3, Cen4, Cen7, Cen8, and Cen11 and non-repetitive centromere sequences of chromosomes 5, 27, and Z using highly elongated lampbrush chromosomes, which are characteristic of the diplotene stage of oogenesis. The degree of chromatin packaging and fine spatial organisations of tandemly repetitive and non-tandemly repetitive centromeric sequences significantly differ at the lampbrush stage. Using DNA/RNA FISH, we have demonstrated that during the lampbrush stage, DNA sequences are transcribed within the centromere regions of chromosomes that lack centromere-specific tandem repeats. In contrast, chromosome-specific centromeric repeats Cen1, Cen2, Cen3, Cen4, Cen7, Cen8, and Cen11 do not demonstrate any transcriptional activity during the lampbrush stage. In addition, we found that CNM repeat cluster localises adjacent to non-repetitive centromeric sequences in chicken microchromosome 27 indicating that centromere region in this chromosome is repeat-rich. Cross-species FISH allowed localisation of the sequences homologous to centromeric DNA of chicken chromosomes 5 and 27 in centromere regions of quail orthologous chromosomes.

  18. Bacterial scaffold directs pole-specific centromere segregation.

    Science.gov (United States)

    Ptacin, Jerod L; Gahlmann, Andreas; Bowman, Grant R; Perez, Adam M; von Diezmann, Alexander R S; Eckart, Michael R; Moerner, W E; Shapiro, Lucy

    2014-05-13

    Bacteria use partitioning systems based on the ParA ATPase to actively mobilize and spatially organize molecular cargoes throughout the cytoplasm. The bacterium Caulobacter crescentus uses a ParA-based partitioning system to segregate newly replicated chromosomal centromeres to opposite cell poles. Here we demonstrate that the Caulobacter PopZ scaffold creates an organizing center at the cell pole that actively regulates polar centromere transport by the ParA partition system. As segregation proceeds, the ParB-bound centromere complex is moved by progressively disassembling ParA from a nucleoid-bound structure. Using superresolution microscopy, we show that released ParA is recruited directly to binding sites within a 3D ultrastructure composed of PopZ at the cell pole, whereas the ParB-centromere complex remains at the periphery of the PopZ structure. PopZ recruitment of ParA stimulates ParA to assemble on the nucleoid near the PopZ-proximal cell pole. We identify mutations in PopZ that allow scaffold assembly but specifically abrogate interactions with ParA and demonstrate that PopZ/ParA interactions are required for proper chromosome segregation in vivo. We propose that during segregation PopZ sequesters free ParA and induces target-proximal regeneration of ParA DNA binding activity to enforce processive and pole-directed centromere segregation, preventing segregation reversals. PopZ therefore functions as a polar hub complex at the cell pole to directly regulate the directionality and destination of transfer of the mitotic segregation machine.

  19. Three wise centromere functions: see no error, hear no break, speak no delay.

    Science.gov (United States)

    Tanaka, Tomoyuki U; Clayton, Lesley; Natsume, Toyoaki

    2013-12-01

    The main function of the centromere is to promote kinetochore assembly for spindle microtubule attachment. Two additional functions of the centromere, however, are becoming increasingly clear: facilitation of robust sister-chromatid cohesion at pericentromeres and advancement of replication of centromeric regions. The combination of these three centromere functions ensures correct chromosome segregation during mitosis. Here, we review the mechanisms of the kinetochore-microtubule interaction, focusing on sister-kinetochore bi-orientation (or chromosome bi-orientation). We also discuss the biological importance of robust pericentromeric cohesion and early centromere replication, as well as the mechanisms orchestrating these two functions at the microtubule attachment site.

  20. A regulatory effect of INMAP on centromere proteins: antisense INMAP induces CENP-B variation and centromeric halo.

    Directory of Open Access Journals (Sweden)

    Tan Tan

    Full Text Available CENP-B is a highly conserved protein that facilitates the assembly of specific centromere structures both in interphase nuclei and on mitotic chromosomes. INMAP is a conserved protein that localizes at nucleus in interphase cells and at mitotic apparatus in mitotic cells. Our previous results showed that INMAP over-expression leads to spindle defects, mitotic arrest and formation of polycentrosomal and multinuclear cells, indicating that INMAP may modulate the function of (a key protein(s in mitotic apparatus. In this study, we demonstrate that INMAP interacts with CENP-B and promotes cleavage of the N-terminal DNA binding domain from CENP-B. The cleaved CENP-B cannot associate with centromeres and thus lose its centromere-related functions. Consistent with these results, CENP-B in INMAP knockdown cells becomes more diffused around kinetochores. Although INMAP knockdown cells do not exhibit gross defects in mitotic spindle formation, these cells go through mitosis, especially prophase and metaphase, with different relative timing, indicating subtle abnormality. These results identify INMAP as a model regulator of CENP-B and support the notion that INMAP regulates mitosis through modulating CENP-B-mediated centromere organization.

  1. A regulatory effect of INMAP on centromere proteins: antisense INMAP induces CENP-B variation and centromeric halo.

    Science.gov (United States)

    Tan, Tan; Chen, Zhe; Lei, Yan; Zhu, Yan; Liang, Qianjin

    2014-01-01

    CENP-B is a highly conserved protein that facilitates the assembly of specific centromere structures both in interphase nuclei and on mitotic chromosomes. INMAP is a conserved protein that localizes at nucleus in interphase cells and at mitotic apparatus in mitotic cells. Our previous results showed that INMAP over-expression leads to spindle defects, mitotic arrest and formation of polycentrosomal and multinuclear cells, indicating that INMAP may modulate the function of (a) key protein(s) in mitotic apparatus. In this study, we demonstrate that INMAP interacts with CENP-B and promotes cleavage of the N-terminal DNA binding domain from CENP-B. The cleaved CENP-B cannot associate with centromeres and thus lose its centromere-related functions. Consistent with these results, CENP-B in INMAP knockdown cells becomes more diffused around kinetochores. Although INMAP knockdown cells do not exhibit gross defects in mitotic spindle formation, these cells go through mitosis, especially prophase and metaphase, with different relative timing, indicating subtle abnormality. These results identify INMAP as a model regulator of CENP-B and support the notion that INMAP regulates mitosis through modulating CENP-B-mediated centromere organization.

  2. Euchromatic subdomains in rice centromeres are associated with genes and transcription.

    Science.gov (United States)

    Wu, Yufeng; Kikuchi, Shinji; Yan, Huihuang; Zhang, Wenli; Rosenbaum, Heidi; Iniguez, A Leonardo; Jiang, Jiming

    2011-11-01

    The presence of the centromere-specific histone H3 variant, CENH3, defines centromeric (CEN) chromatin, but poorly understood epigenetic mechanisms determine its establishment and maintenance. CEN chromatin is embedded within pericentromeric heterochromatin in most higher eukaryotes, but, interestingly, it can show euchromatic characteristics; for example, the euchromatic histone modification mark dimethylated H3 Lys 4 (H3K4me2) is uniquely associated with animal centromeres. To examine the histone marks and chromatin properties of plant centromeres, we developed a genomic tiling array for four fully sequenced rice (Oryza sativa) centromeres and used chromatin immunoprecipitation-chip to study the patterns of four euchromatic histone modification marks: H3K4me2, trimethylated H3 Lys 4, trimethylated H3 Lys 36, and acetylated H3 Lys 4, 9. The vast majority of the four histone marks were associated with genes located in the H3 subdomains within the centromere cores. We demonstrate that H3K4me2 is not a ubiquitous component of rice CEN chromatin, and the euchromatic characteristics of rice CEN chromatin are hallmarks of the transcribed sequences embedded in the centromeric H3 subdomains. We propose that the transcribed sequences located in rice centromeres may provide a barrier preventing loading of CENH3 into the H3 subdomains. The separation of CENH3 and H3 subdomains in the centromere core may be favorable for the formation of three-dimensional centromere structure and for rice centromere function.

  3. Total centromere size and genome size are strongly correlated in ten grass species.

    Science.gov (United States)

    Zhang, Han; Dawe, R Kelly

    2012-05-01

    It has been known for decades that centromere size varies across species, but the factors involved in setting centromere boundaries are unknown. As a means to address this question, we estimated centromere sizes in ten species of the grass family including rice, maize, and wheat, which diverged 60~80 million years ago and vary by 40-fold in genome size. Measurements were made using a broadly reactive antibody to rice centromeric histone H3 (CENH3). In species-wide comparisons, we found a clear linear relationship between total centromere size and genome size. Species with large genomes and few chromosomes tend to have the largest centromeres (e.g., rye) while species with small genomes and many chromosomes have the smallest centromeres (e.g., rice). However, within a species, centromere size is surprisingly uniform. We present evidence from three oat-maize addition lines that support this claim, indicating that each of three maize centromeres propagated in oat are not measurably different from each other. In the context of previously published data, our results suggest that the apparent correlation between chromosome and centromere size is incidental to a larger trend that reflects genome size. Centromere size may be determined by a limiting component mechanism similar to that described for Caenorhabditis elegans centrosomes.

  4. SWI/SNF-like chromatin remodeling factor Fun30 supports point centromere function in S. cerevisiae.

    Directory of Open Access Journals (Sweden)

    Mickaël Durand-Dubief

    2012-09-01

    Full Text Available Budding yeast centromeres are sequence-defined point centromeres and are, unlike in many other organisms, not embedded in heterochromatin. Here we show that Fun30, a poorly understood SWI/SNF-like chromatin remodeling factor conserved in humans, promotes point centromere function through the formation of correct chromatin architecture at centromeres. Our determination of the genome-wide binding and nucleosome positioning properties of Fun30 shows that this enzyme is consistently enriched over centromeres and that a majority of CENs show Fun30-dependent changes in flanking nucleosome position and/or CEN core micrococcal nuclease accessibility. Fun30 deletion leads to defects in histone variant Htz1 occupancy genome-wide, including at and around most centromeres. FUN30 genetically interacts with CSE4, coding for the centromere-specific variant of histone H3, and counteracts the detrimental effect of transcription through centromeres on chromosome segregation and suppresses transcriptional noise over centromere CEN3. Previous work has shown a requirement for fission yeast and mammalian homologs of Fun30 in heterochromatin assembly. As centromeres in budding yeast are not embedded in heterochromatin, our findings indicate a direct role of Fun30 in centromere chromatin by promoting correct chromatin architecture.

  5. Uncoupling of satellite DNA and centromeric function in the genus Equus.

    Directory of Open Access Journals (Sweden)

    Francesca M Piras

    2010-02-01

    Full Text Available In a previous study, we showed that centromere repositioning, that is the shift along the chromosome of the centromeric function without DNA sequence rearrangement, has occurred frequently during the evolution of the genus Equus. In this work, the analysis of the chromosomal distribution of satellite tandem repeats in Equus caballus, E. asinus, E. grevyi, and E. burchelli highlighted two atypical features: 1 several centromeres, including the previously described evolutionary new centromeres (ENCs, seem to be devoid of satellite DNA, and 2 satellite repeats are often present at non-centromeric termini, probably corresponding to relics of ancestral now inactive centromeres. Immuno-FISH experiments using satellite DNA and antibodies against the kinetochore protein CENP-A demonstrated that satellite-less primary constrictions are actually endowed with centromeric function. The phylogenetic reconstruction of centromere repositioning events demonstrates that the acquisition of satellite DNA occurs after the formation of the centromere during evolution and that centromeres can function over millions of years and many generations without detectable satellite DNA. The rapidly evolving Equus species gave us the opportunity to identify different intermediate steps along the full maturation of ENCs.

  6. Epigenetically-inherited centromere and neocentromere DNA replicates earliest in S-phase.

    Directory of Open Access Journals (Sweden)

    Amnon Koren

    2010-08-01

    Full Text Available Eukaryotic centromeres are maintained at specific chromosomal sites over many generations. In the budding yeast Saccharomyces cerevisiae, centromeres are genetic elements defined by a DNA sequence that is both necessary and sufficient for function; whereas, in most other eukaryotes, centromeres are maintained by poorly characterized epigenetic mechanisms in which DNA has a less definitive role. Here we use the pathogenic yeast Candida albicans as a model organism to study the DNA replication properties of centromeric DNA. By determining the genome-wide replication timing program of the C. albicans genome, we discovered that each centromere is associated with a replication origin that is the first to fire on its respective chromosome. Importantly, epigenetic formation of new ectopic centromeres (neocentromeres was accompanied by shifts in replication timing, such that a neocentromere became the first to replicate and became associated with origin recognition complex (ORC components. Furthermore, changing the level of the centromere-specific histone H3 isoform led to a concomitant change in levels of ORC association with centromere regions, further supporting the idea that centromere proteins determine origin activity. Finally, analysis of centromere-associated DNA revealed a replication-dependent sequence pattern characteristic of constitutively active replication origins. This strand-biased pattern is conserved, together with centromere position, among related strains and species, in a manner independent of primary DNA sequence. Thus, inheritance of centromere position is correlated with a constitutively active origin of replication that fires at a distinct early time. We suggest a model in which the distinct timing of DNA replication serves as an epigenetic mechanism for the inheritance of centromere position.

  7. The chromosomal passenger complex activates Polo kinase at centromeres.

    Directory of Open Access Journals (Sweden)

    Mar Carmena

    2012-01-01

    Full Text Available The coordinated activities at centromeres of two key cell cycle kinases, Polo and Aurora B, are critical for ensuring that the two sister kinetochores of each chromosome are attached to microtubules from opposite spindle poles prior to chromosome segregation at anaphase. Initial attachments of chromosomes to the spindle involve random interactions between kinetochores and dynamic microtubules, and errors occur frequently during early stages of the process. The balance between microtubule binding and error correction (e.g., release of bound microtubules requires the activities of Polo and Aurora B kinases, with Polo promoting stable attachments and Aurora B promoting detachment. Our study concerns the coordination of the activities of these two kinases in vivo. We show that INCENP, a key scaffolding subunit of the chromosomal passenger complex (CPC, which consists of Aurora B kinase, INCENP, Survivin, and Borealin/Dasra B, also interacts with Polo kinase in Drosophila cells. It was known that Aurora A/Bora activates Polo at centrosomes during late G2. However, the kinase that activates Polo on chromosomes for its critical functions at kinetochores was not known. We show here that Aurora B kinase phosphorylates Polo on its activation loop at the centromere in early mitosis. This phosphorylation requires both INCENP and Aurora B activity (but not Aurora A activity and is critical for Polo function at kinetochores. Our results demonstrate clearly that Polo kinase is regulated differently at centrosomes and centromeres and suggest that INCENP acts as a platform for kinase crosstalk at the centromere. This crosstalk may enable Polo and Aurora B to achieve a balance wherein microtubule mis-attachments are corrected, but proper attachments are stabilized allowing proper chromosome segregation.

  8. The chromosomal passenger complex activates Polo kinase at centromeres.

    Science.gov (United States)

    Carmena, Mar; Pinson, Xavier; Platani, Melpi; Salloum, Zeina; Xu, Zhenjie; Clark, Anthony; Macisaac, Fiona; Ogawa, Hiromi; Eggert, Ulrike; Glover, David M; Archambault, Vincent; Earnshaw, William C

    2012-01-01

    The coordinated activities at centromeres of two key cell cycle kinases, Polo and Aurora B, are critical for ensuring that the two sister kinetochores of each chromosome are attached to microtubules from opposite spindle poles prior to chromosome segregation at anaphase. Initial attachments of chromosomes to the spindle involve random interactions between kinetochores and dynamic microtubules, and errors occur frequently during early stages of the process. The balance between microtubule binding and error correction (e.g., release of bound microtubules) requires the activities of Polo and Aurora B kinases, with Polo promoting stable attachments and Aurora B promoting detachment. Our study concerns the coordination of the activities of these two kinases in vivo. We show that INCENP, a key scaffolding subunit of the chromosomal passenger complex (CPC), which consists of Aurora B kinase, INCENP, Survivin, and Borealin/Dasra B, also interacts with Polo kinase in Drosophila cells. It was known that Aurora A/Bora activates Polo at centrosomes during late G2. However, the kinase that activates Polo on chromosomes for its critical functions at kinetochores was not known. We show here that Aurora B kinase phosphorylates Polo on its activation loop at the centromere in early mitosis. This phosphorylation requires both INCENP and Aurora B activity (but not Aurora A activity) and is critical for Polo function at kinetochores. Our results demonstrate clearly that Polo kinase is regulated differently at centrosomes and centromeres and suggest that INCENP acts as a platform for kinase crosstalk at the centromere. This crosstalk may enable Polo and Aurora B to achieve a balance wherein microtubule mis-attachments are corrected, but proper attachments are stabilized allowing proper chromosome segregation.

  9. Molecular characterization of flow-sorted mammalian centromeres

    Energy Technology Data Exchange (ETDEWEB)

    Hamkalo, B.A.; Henschen, A.; Parseghian, M.H. [Univ. of Calfornia, Irvine, CA (United States). Dept. of Molecular Biology and Biochemistry] [and others

    1998-12-31

    This is the final report of a three-year, Laboratory Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). The project involved experiments directed towards developing a molecular characterization of the centromere region of mammalian chromosomes. Attempts to purify this essential chromosomal locus by conventional methods have thus far been unsuccessful. However, preliminary data obtained in collaboration with the National Flow Cytometry Resource (NFCR) showed that it is possible to purify a chromosome fragment that is present in certain cultured mouse cell lines and has all the properties expected of an intact centromere region. To begin sorting this minichromosome for the identification of proteins preferentially associated with centromere regions, standard buffers utilized in chromosome sorting were evaluated for potential effects on maintenance of chromosomal proteins during sorting. The data indicate that the presence of several buffer constituents results in the extraction of all but a few chromosomal proteins. The subsequent use of a magnesium sulfate buffer resulted in the sorting of mouse chromosomes that do not suffer a significant loss of proteins. Several DNA stains were also evaluated for causing protein dissociation, but no significant losses were observed. Although flow-sorted chromosomes have been used extensively for DNA analysis and cloning, this is a pioneering effort by the NFCR, and its collaborators, to exploit chromosome sorting capabilities for the analysis of chromosomal proteins.

  10. Organization and evolution of Gorilla centromeric DNA from old strategies to new approaches

    OpenAIRE

    Catacchio, C. R.; Ragone, R.; Chiatante, G.; M. Ventura

    2015-01-01

    The centromere/kinetochore interaction is responsible for the pairing and segregation of replicated chromosomes in eukaryotes. Centromere DNA is portrayed as scarcely conserved, repetitive in nature, quickly evolving and protein-binding competent. Among primates, the major class of centromeric DNA is the pancentromeric α-satellite, made of arrays of 171 bp monomers, repeated in a head-to-tail pattern. α-satellite sequences can either form tandem heterogeneous monomeric arrays or assemble in h...

  11. Hematopoietic stem cell transplantation corrects the immunologic abnormalities associated with immunodeficiency-centromeric instability-facial dysmorphism syndrome.

    NARCIS (Netherlands)

    Gennery, A.R.; Slatter, M.A.; Bredius, R.G.; Hagleitner, M.M.; Weemaes, C.M.R.; Cant, A.J.; Lankester, A.C.

    2007-01-01

    Immunodeficiency-centromeric instability-facial dysmorphism syndrome, characterized by variable immunodeficiency, centromeric instability, and facial anomalies caused by epigenetic dysregulation resulting in hypomethylation, is caused in many patients by mutations in DNMT3B, a DNA methyltransferase

  12. Chromosome segregation regulation in human zygotes : Altered mitotic histone phosphorylation dynamics underlying centromeric targeting of the chromosomal passenger complex

    NARCIS (Netherlands)

    Van De Werken, C.; Avo Santos, M.; Laven, J. S E; Eleveld, C.; Fauser, B. C J M; Lens, S. M A; Baart, E. B.

    2015-01-01

    STUDY QUESTION Are the kinase feedback loops that regulate activation and centromeric targeting of the chromosomal passenger complex (CPC), functional during mitosis in human embryos? SUMMARY ANSWER Investigation of the regulatory kinase pathways involved in centromeric CPC targeting revealed normal

  13. Organization and evolution of Gorilla centromeric DNA from old strategies to new approaches.

    Science.gov (United States)

    Catacchio, C R; Ragone, R; Chiatante, G; Ventura, M

    2015-09-21

    The centromere/kinetochore interaction is responsible for the pairing and segregation of replicated chromosomes in eukaryotes. Centromere DNA is portrayed as scarcely conserved, repetitive in nature, quickly evolving and protein-binding competent. Among primates, the major class of centromeric DNA is the pancentromeric α-satellite, made of arrays of 171 bp monomers, repeated in a head-to-tail pattern. α-satellite sequences can either form tandem heterogeneous monomeric arrays or assemble in higher-order repeats (HORs). Gorilla centromere DNA has barely been characterized, and data are mainly based on hybridizations of human alphoid sequences. We isolated and finely characterized gorilla α-satellite sequences and revealed relevant structure and chromosomal distribution similarities with other great apes as well as gorilla-specific features, such as the uniquely octameric structure of the suprachromosomal family-2 (SF2). We demonstrated for the first time the orthologous localization of alphoid suprachromosomal families-1 and -2 (SF1 and SF2) between human and gorilla in contrast to chimpanzee centromeres. Finally, the discovery of a new 189 bp monomer type in gorilla centromeres unravels clues to the role of the centromere protein B, paving the way to solve the significance of the centromere DNA's essential repetitive nature in association with its function and the peculiar evolution of the α-satellite sequence.

  14. High frequency of centromere inactivation resulting in stable dicentric chromosomes of maize.

    Science.gov (United States)

    Han, Fangpu; Lamb, Jonathan C; Birchler, James A

    2006-02-28

    Somatic chromosome spreads from maize (Zea mays L.) plants containing B-A translocation chromosomes undergoing the chromosome type breakage-fusion-bridge cycle were examined by FISH. The size and type of extra chromosomes varied among cells of the same individual. A collection of minichromosomes derived from the chromosome type breakage-fusion-bridge cycle was examined for the presence of stable dicentric chromosomes. Six of 23 chromosomes in the collection contained two regions with DNA sequences typical of centromeres. Functional analysis and immunolabeling of CENH3, the centromere-specific histone H3 variant, revealed only one functional centromere per chromosome, despite the duplicate centromere sequences. One plant was found with an inactive B centromere that had been translocated to the short arm of chromosome 9. The translocated centromere region appeared identical to that of a normal B chromosome. The inactivation of the centromeres was stable for at least four generations. By using dicentrics from dispensable chromosomes, centromere inactivation was found to be quite common under these circumstances.

  15. Centromere architecture breakdown induced by the viral E3 ubiquitin ligase ICP0 protein of herpes simplex virus type 1.

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    Sylvain Gross

    Full Text Available The viral E3 ubiquitin ligase ICP0 protein has the unique property to temporarily localize at interphase and mitotic centromeres early after infection of cells by the herpes simplex virus type 1 (HSV-1. As a consequence ICP0 induces the proteasomal degradation of several centromeric proteins (CENPs, namely CENP-A, the centromeric histone H3 variant, CENP-B and CENP-C. Following ICP0-induced centromere modification cells trigger a specific response to centromeres called interphase Centromere Damage Response (iCDR. The biological significance of the iCDR is unknown; so is the degree of centromere structural damage induced by ICP0. Interphase centromeres are complex structures made of proximal and distal protein layers closely associated to CENP-A-containing centromeric chromatin. Using several cell lines constitutively expressing GFP-tagged CENPs, we investigated the extent of the centromere destabilization induced by ICP0. We show that ICP0 provokes the disappearance from centromeres, and the proteasomal degradation of several CENPs from the NAC (CENP-A nucleosome associated and CAD (CENP-A Distal complexes. We then investigated the nucleosomal occupancy of the centromeric chromatin in ICP0-expressing cells by micrococcal nuclease (MNase digestion analysis. ICP0 expression either following infection or in cell lines constitutively expressing ICP0 provokes significant modifications of the centromeric chromatin structure resulting in higher MNase accessibility. Finally, using human artificial chromosomes (HACs, we established that ICP0-induced iCDR could also target exogenous centromeres. These results demonstrate that, in addition to the protein complexes, ICP0 also destabilizes the centromeric chromatin resulting in the complete breakdown of the centromere architecture, which consequently induces iCDR.

  16. Structure and function of centromeric and pericentromeric heterochromatin in Arabidopsis thaliana

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    Lauriane eSimon

    2015-11-01

    Full Text Available The centromere is a specific chromosomal region where the kinetochore assembles to ensure the faithful segregation of sister chromatids during mitosis and meiosis. Centromeres are defined by a local enrichment of the specific histone variant CenH3 mostly at repetitive satellite sequences. A larger pericentromeric region containing repetitive sequences and transposable elements surrounds the centromere that adopts a particular chromatin state characterized by specific histone variants and post-translational modifications and forms a transcriptionally repressive chromosomal environment. In the model organism Arabidopsis thaliana centromeric and pericentromeric domains form conspicuous heterochromatin clusters called chromocenters in interphase. Here we discuss, using Arabidopsis as example, recent insight into mechanisms involved in maintenance and establishment of centromeric and pericentromeric chromatin signatures as well as in chromocenter formation.

  17. The centromere geometry essential for keeping mitosis error free is controlled by spindle forces.

    Science.gov (United States)

    Loncarek, Jadranka; Kisurina-Evgenieva, Olga; Vinogradova, Tatiana; Hergert, Polla; La Terra, Sabrina; Kapoor, Tarun M; Khodjakov, Alexey

    2007-11-29

    Accurate segregation of chromosomes, essential for the stability of the genome, depends on 'bi-orientation'-simultaneous attachment of each individual chromosome to both poles of the mitotic spindle. On bi-oriented chromosomes, kinetochores (macromolecular complexes that attach the chromosome to the spindle) reside on the opposite sides of the chromosome's centromere. In contrast, sister kinetochores shift towards one side of the centromere on 'syntelic' chromosomes that erroneously attach to one spindle pole with both sister kinetochores. Syntelic attachments often arise during spindle assembly and must be corrected to prevent chromosome loss. It is assumed that restoration of proper centromere architecture occurs automatically owing to elastic properties of the centromere. Here we test this assumption by combining laser microsurgery and chemical biology assays in cultured mammalian cells. We find that kinetochores of syntelic chromosomes remain juxtaposed on detachment from spindle microtubules. These findings reveal that correction of syntelic attachments involves an extra step that has previously been overlooked: external forces must be applied to move sister kinetochores to the opposite sides of the centromere. Furthermore, we demonstrate that the shape of the centromere is important for spindle assembly, because bipolar spindles do not form in cells lacking centrosomes when multiple chromosomes with juxtaposed kinetochores are present. Thus, proper architecture of the centromere makes an important contribution to achieving high fidelity of chromosome segregation.

  18. Widespread Positive Selection Drives Differentiation of Centromeric Proteins in the Drosophila melanogaster subgroup.

    Science.gov (United States)

    Beck, Emily A; Llopart, Ana

    2015-11-25

    Rapid evolution of centromeric satellite repeats is thought to cause compensatory amino acid evolution in interacting centromere-associated kinetochore proteins. Cid, a protein that mediates kinetochore/centromere interactions, displays particularly high amino acid turnover. Rapid evolution of both Cid and centromeric satellite repeats led us to hypothesize that the apparent compensatory evolution may extend to interacting partners in the Condensin I complex (i.e., SMC2, SMC4, Cap-H, Cap-D2, and Cap-G) and HP1s. Missense mutations in these proteins often result in improper centromere formation and aberrant chromosome segregation, thus selection for maintained function and coevolution among proteins of the complex is likely strong. Here, we report evidence of rapid evolution and recurrent positive selection in seven centromere-associated proteins in species of the Drosophila melanogaster subgroup, and further postulate that positive selection on these proteins could be a result of centromere drive and compensatory changes, with kinetochore proteins competing for optimal spindle attachment.

  19. Centromere inactivation on a neo-Y fusion chromosome in threespine stickleback fish.

    Science.gov (United States)

    Cech, Jennifer N; Peichel, Catherine L

    2016-12-01

    Having one and only one centromere per chromosome is essential for proper chromosome segregation during both mitosis and meiosis. Chromosomes containing two centromeres are known as dicentric and often mis-segregate during cell division, resulting in aneuploidy or chromosome breakage. Dicentric chromosome can be stabilized by centromere inactivation, a process which reestablishes monocentric chromosomes. However, little is known about this process in naturally occurring dicentric chromosomes. Using a combination of fluorescence in situ hybridization (FISH) and immunofluorescence combined with FISH (IF-FISH) on metaphase chromosome spreads, we demonstrate that centromere inactivation has evolved on a neo-Y chromosome fusion in the Japan Sea threespine stickleback fish (Gasterosteus nipponicus). We found that the centromere derived from the ancestral Y chromosome has been inactivated. Our data further suggest that there have been genetic changes to this centromere in the two million years since the formation of the neo-Y chromosome, but it remains unclear whether these genetic changes are a cause or consequence of centromere inactivation.

  20. High quality maize centromere 10 sequence reveals evidence of frequent recombination events

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    Thomas Kai Wolfgruber

    2016-03-01

    Full Text Available The ancestral centromeres of maize contain long stretches of the tandemly arranged CentC repeat. The abundance of tandem DNA repeats and centromeric retrotransposons (CR have presented a significant challenge to completely assembling centromeres using traditional sequencing methods. Here we report a nearly complete assembly of the 1.85 Mb maize centromere 10 from inbred B73 using PacBio technology and BACs from the reference genome project. The error rates estimated from overlapping BAC sequences are 7 x 10-6 and 5 x 10-5 for mismatches and indels, respectively. The number of gaps in the region covered by the reassembly was reduced from 140 in the reference genome to three. Three expressed genes are located between 92 and 477 kb of the inferred ancestral CentC cluster, which lies within the region of highest centromeric repeat density. The improved assembly increased the count of full-length centromeric retrotransposons from 5 to 55 and revealed a 22.7 kb segmental duplication that occurred approximately 121,000 years ago. Our analysis provides evidence of frequent recombination events in the form of partial retrotransposons, deletions within retrotransposons, chimeric retrotransposons, segmental duplications including higher order CentC repeats, a deleted CentC monomer, centromere-proximal inversions, and insertion of mitochondrial sequences. Double-strand DNA break (DSB repair is the most plausible mechanism for these events and may be the major driver of centromere repeat evolution and diversity. This repair appears to be mediated by microhomology, suggesting that tandem repeats may have evolved to facilitate the repair of frequent DSBs in centromeres.

  1. Rad51-Rad52 mediated maintenance of centromeric chromatin in Candida albicans.

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    Sreyoshi Mitra

    2014-04-01

    Full Text Available Specification of the centromere location in most eukaryotes is not solely dependent on the DNA sequence. However, the non-genetic determinants of centromere identity are not clearly defined. While multiple mechanisms, individually or in concert, may specify centromeres epigenetically, most studies in this area are focused on a universal factor, a centromere-specific histone H3 variant CENP-A, often considered as the epigenetic determinant of centromere identity. In spite of variable timing of its loading at centromeres across species, a replication coupled early S phase deposition of CENP-A is found in most yeast centromeres. Centromeres are the earliest replicating chromosomal regions in a pathogenic budding yeast Candida albicans. Using a 2-dimensional agarose gel electrophoresis assay, we identify replication origins (ORI7-LI and ORI7-RI proximal to an early replicating centromere (CEN7 in C. albicans. We show that the replication forks stall at CEN7 in a kinetochore dependent manner and fork stalling is reduced in the absence of the homologous recombination (HR proteins Rad51 and Rad52. Deletion of ORI7-RI causes a significant reduction in the stalled fork signal and an increased loss rate of the altered chromosome 7. The HR proteins, Rad51 and Rad52, have been shown to play a role in fork restart. Confocal microscopy shows declustered kinetochores in rad51 and rad52 mutants, which are evidence of kinetochore disintegrity. CENP-ACaCse4 levels at centromeres, as determined by chromatin immunoprecipitation (ChIP experiments, are reduced in absence of Rad51/Rad52 resulting in disruption of the kinetochore structure. Moreover, western blot analysis reveals that delocalized CENP-A molecules in HR mutants degrade in a similar fashion as in other kinetochore mutants described before. Finally, co-immunoprecipitation assays indicate that Rad51 and Rad52 physically interact with CENP-ACaCse4 in vivo. Thus, the HR proteins Rad51 and Rad52

  2. Clinical spectrum of immunodeficiency, centromeric instability and facial dysmorphism (ICF syndrome).

    NARCIS (Netherlands)

    Hagleitner, M.M.; Lankester, A.; Maraschio, P.; Hulten, M.; Fryns, J.P.; Schuetz, C.; Gimelli, G.; Davies, E.G.; Gennery, A.; Belohradsky, B.H.; Groot, R. de; Gerritsen, E.J.; Mattina, T.; Howard, P.J.; Fasth, A.; Reisli, I.; Furthner, D.; Slatter, M.A.; Cant, A.J.; Cazzola, G.; Dijken, P.J. van; Deuren, M. van; Greef, J.C. de; Maarel, S.M. van der; Weemaes, C.M.R.

    2008-01-01

    BACKGROUND: Immunodeficiency, centromeric instability and facial dysmorphism (ICF syndrome) is a rare autosomal recessive disease characterised by facial dysmorphism, immunoglobulin deficiency and branching of chromosomes 1, 9 and 16 after PHA stimulation of lymphocytes. Hypomethylation of DNA of a

  3. The nucleoplasmin homolog NLP mediates centromere clustering and anchoring to the nucleolus.

    Science.gov (United States)

    Padeken, Jan; Mendiburo, María José; Chlamydas, Sarantis; Schwarz, Hans-Jürgen; Kremmer, Elisabeth; Heun, Patrick

    2013-04-25

    Centromere clustering during interphase is a phenomenon known to occur in many different organisms and cell types, yet neither the factors involved nor their physiological relevance is well understood. Using Drosophila tissue culture cells and flies, we identified a network of proteins, including the nucleoplasmin-like protein (NLP), the insulator protein CTCF, and the nucleolus protein Modulo, to be essential for the positioning of centromeres. Artificial targeting further demonstrated that NLP and CTCF are sufficient for clustering, while Modulo serves as the anchor to the nucleolus. Centromere clustering was found to depend on centric chromatin rather than specific DNA sequences. Moreover, unclustering of centromeres results in the spatial destabilization of pericentric heterochromatin organization, leading to partial defects in the silencing of repetitive elements, defects during chromosome segregation, and genome instability.

  4. Mislocalization of the Drosophila centromere-specific histone CIDpromotes formation of functional ectopic kinetochores

    Energy Technology Data Exchange (ETDEWEB)

    Heun, Patrick; Erhardt, Sylvia; Blower, Michael D.; Weiss,Samara; Skora, Andrew D.; Karpen, Gary H.

    2006-01-30

    The centromere-specific histone variant CENP-A (CID in Drosophila) is a structural and functional foundation for kinetochore formation and chromosome segregation. Here, we show that overexpressed CID is mislocalized into normally non-centromeric regions in Drosophila tissue culture cells and animals. Analysis of mitoses in living and fixed cells reveals that mitotic delays, anaphase bridges, chromosome fragmentation, and cell and organismal lethality are all direct consequences of CID mislocalization. In addition, proteins that are normally restricted to endogenous kinetochores assemble at a subset of ectopic CID incorporation regions. The presence of microtubule motors and binding proteins, spindle attachments, and aberrant chromosome morphologies demonstrate that these ectopic kinetochores are functional. We conclude that CID mislocalization promotes formation of ectopic centromeres and multicentric chromosomes, which causes chromosome missegregation, aneuploidy, and growth defects. Thus, CENP-A mislocalization is one possible mechanism for genome instability during cancer progression, as well as centromere plasticity during evolution.

  5. DNMT3B interacts with constitutive centromere protein CENP-C to modulate DNA methylation and the histone code at centromeric regions.

    Science.gov (United States)

    Gopalakrishnan, Suhasni; Sullivan, Beth A; Trazzi, Stefania; Della Valle, Giuliano; Robertson, Keith D

    2009-09-01

    DNA methylation is an epigenetically imposed mark of transcriptional repression that is essential for maintenance of chromatin structure and genomic stability. Genome-wide methylation patterns are mediated by the combined action of three DNA methyltransferases: DNMT1, DNMT3A and DNMT3B. Compelling links exist between DNMT3B and chromosome stability as emphasized by the mitotic defects that are a hallmark of ICF syndrome, a disease arising from germline mutations in DNMT3B. Centromeric and pericentromeric regions are essential for chromosome condensation and the fidelity of segregation. Centromere regions contain distinct epigenetic marks, including dense DNA hypermethylation, yet the mechanisms by which DNA methylation is targeted to these regions remains largely unknown. In the present study, we used a yeast two-hybrid screen and identified a novel interaction between DNMT3B and constitutive centromere protein CENP-C. CENP-C is itself essential for mitosis. We confirm this interaction in mammalian cells and map the domains responsible. Using siRNA knock downs, bisulfite genomic sequencing and ChIP, we demonstrate for the first time that CENP-C recruits DNA methylation and DNMT3B to both centromeric and pericentromeric satellite repeats and that CENP-C and DNMT3B regulate the histone code in these regions, including marks characteristic of centromeric chromatin. Finally, we demonstrate that loss of CENP-C or DNMT3B leads to elevated chromosome misalignment and segregation defects during mitosis and increased transcription of centromeric repeats. Taken together, our data reveal a novel mechanism by which DNA methylation is targeted to discrete regions of the genome and contributes to chromosomal stability.

  6. Premitotic assembly of human CENPs -T and -W switches centromeric chromatin to a mitotic state.

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    Lisa Prendergast

    2011-06-01

    Full Text Available Centromeres are differentiated chromatin domains, present once per chromosome, that direct segregation of the genome in mitosis and meiosis by specifying assembly of the kinetochore. They are distinct genetic loci in that their identity in most organisms is determined not by the DNA sequences they are associated with, but through specific chromatin composition and context. The core nucleosomal protein CENP-A/cenH3 plays a primary role in centromere determination in all species and directs assembly of a large complex of associated proteins in vertebrates. While CENP-A itself is stably transmitted from one generation to the next, the nature of the template for centromere replication and its relationship to kinetochore function are as yet poorly understood. Here, we investigate the assembly and inheritance of a histone fold complex of the centromere, the CENP-T/W complex, which is integrated with centromeric chromatin in association with canonical histone H3 nucleosomes. We have investigated the cell cycle regulation, timing of assembly, generational persistence, and requirement for function of CENPs -T and -W in the cell cycle in human cells. The CENP-T/W complex assembles through a dynamic exchange mechanism in late S-phase and G2, is required for mitosis in each cell cycle and does not persist across cell generations, properties reciprocal to those measured for CENP-A. We propose that the CENP-A and H3-CENP-T/W nucleosome components of the centromere are specialized for centromeric and kinetochore activities, respectively. Segregation of the assembly mechanisms for the two allows the cell to switch between chromatin configurations that reciprocally support the replication of the centromere and its conversion to a mitotic state on postreplicative chromatin.

  7. The Role of Dicentric Chromosome Formation and Secondary Centromere Deletion in the Evolution of Myeloid Malignancy

    OpenAIRE

    MacKinnon, Ruth N.; Campbell, Lynda J.

    2011-01-01

    Dicentric chromosomes have been identified as instigators of the genome instability associated with cancer, but this instability is often resolved by one of a number of different secondary events. These include centromere inactivation, inversion, and intercentromeric deletion. Deletion or excision of one of the centromeres may be a significant occurrence in myeloid malignancy and other malignancies but has not previously been widely recognized, and our reports are the first describing centrom...

  8. CenH3 evolution reflects meiotic symmetry as predicted by the centromere drive model.

    Science.gov (United States)

    Zedek, František; Bureš, Petr

    2016-09-15

    The centromere drive model explaining rapid evolution of eukaryotic centromeres predicts higher frequency of positive selection acting on centromeric histone H3 (CenH3) in clades with asymmetric meiosis compared to the clades with only symmetric meiosis. However, despite the impression one might get from the literature, this key prediction of the centromere drive model has not only never been confirmed, but it has never been tested, because all the previous studies dealt only with the presence or absence instead of the frequency of positive selection. To provide evidence for or against different frequencies of positively selected CenH3 in asymmetrics and symmetrics, we have inferred the selective pressures acting on CenH3 in seventeen eukaryotic clades, including plants, animals, fungi, ciliates and apicomplexa, using codon-substitution models, and compared the inferred frequencies between asymmetrics and symmetrics in a quantitative manner. We have found that CenH3 has been evolving adaptively much more frequently in clades with asymmetric meiosis compared with clades displaying only symmetric meiosis which confirms the prediction of centromere drive model. Our findings indicate that the evolution of asymmetric meiosis required CenH3 to evolve adaptively more often to counterbalance the negative consequences of centromere drive.

  9. Identification of Drosophila centromere associated proteins by quantitative affinity purification-mass spectrometry

    Science.gov (United States)

    Barth, Teresa K.; Schade, Georg O.M.; Schmidt, Andreas; Vetter, Irene; Wirth, Marc; Heun, Patrick; Imhof, Axel; Thomae, Andreas W.

    2015-01-01

    Centromeres of higher eukaryotes are epigenetically defined by the centromere specific histone H3 variant CENP-ACID. CENP-ACID builds the foundation for the assembly of a large network of proteins. In contrast to mammalian systems, the protein composition of Drosophila centromeres has not been comprehensively investigated. Here we describe the proteome of Drosophila melanogaster centromeres as analyzed by quantitative affinity purification-mass spectrometry (AP-MS). The AP-MS input chromatin material was prepared from D. melanogaster cell lines expressing CENP-ACID or H3.3 fused to EGFP as baits. Centromere chromatin enriched proteins were identified based on their relative abundance in CENP-ACID–GFP compared to H3.3-GFP or mock affinity-purifications. The analysis yielded 86 proteins specifically enriched in centromere chromatin preparations. The data accompanying the manuscript on this approach (Barth et al., 2015, Proteomics 14:2167-78, DOI: 10.1002/pmic.201400052) has been deposited to the ProteomeXchange Consortium (http://www.proteomexchange.org) via the PRIDE partner repository with the dataset identifier PXD000758. PMID:26306323

  10. Nucleolar activity and CENP-C regulate CENP-A and CAL1 availability for centromere assembly in meiosis.

    Science.gov (United States)

    Kwenda, Lucretia; Collins, Caitriona M; Dattoli, Anna A; Dunleavy, Elaine M

    2016-04-15

    The centromere-specific histone CENP-A is the key epigenetic determinant of centromere identity. Whereas most histones are removed from mature sperm, CENP-A is retained to mark paternal centromeres. In Drosophila males we show that the centromere assembly factors CAL1 and CENP-C are required for meiotic chromosome segregation, CENP-A assembly and maintenance on sperm, as well as fertility. In meiosis, CENP-A accumulates with CAL1 in nucleoli. Furthermore, we show that CENP-C normally limits the release of CAL1 and CENP-A from nucleoli for proper centromere assembly in meiotic prophase I. Finally, we show that RNA polymerase I transcription is required for efficient CENP-A assembly in meiosis, as well as centromere tethering to nucleoli.

  11. The Role of Dicentric Chromosome Formation and Secondary Centromere Deletion in the Evolution of Myeloid Malignancy

    Directory of Open Access Journals (Sweden)

    Ruth N. MacKinnon

    2011-01-01

    Full Text Available Dicentric chromosomes have been identified as instigators of the genome instability associated with cancer, but this instability is often resolved by one of a number of different secondary events. These include centromere inactivation, inversion, and intercentromeric deletion. Deletion or excision of one of the centromeres may be a significant occurrence in myeloid malignancy and other malignancies but has not previously been widely recognized, and our reports are the first describing centromere deletion in cancer cells. We review what is known about dicentric chromosomes and the mechanisms by which they can undergo stabilization in both constitutional and cancer genomes. The failure to identify centromere deletion in cancer cells until recently can be partly explained by the standard approaches to routine diagnostic cancer genome analysis, which do not identify centromeres in the context of chromosome organization. This hitherto hidden group of primary dicentric, secondary monocentric chromosomes, together with other unrecognized dicentric chromosomes, points to a greater role for dicentric chromosomes in cancer initiation and progression than is generally acknowledged. We present a model that predicts and explains a significant role for dicentric chromosomes in the formation of unbalanced translocations in malignancy.

  12. A cytogenetic study of nuclear power plant workers using the micronucleus-centromere assay.

    Science.gov (United States)

    Thierens, H; Vral, A; Barbé, M; Aousalah, B; De Ridder, L

    1999-09-15

    A cytogenetic study was performed in 215 nuclear power plant workers occupationally exposed to radiation using the micronucleus-centromere assay for peripheral blood lymphocytes. As control population served administrative staff with yearly doses below 1 mSv. The increase of the micronucleus frequency with age, observed in the non-smoking control population, is mainly due to an enhanced number of centromere-positive micronuclei, pointing to an increased chromosome loss. No differences in the number of micronuclei, centromere-positive and centromere-negative micronuclei between smokers and non-smokers are observed. An analysis of the micronucleus data vs. the dose accumulated over the 10 years preceding the venepuncture shows no significant clastogenic or aneuploidogenic effects of the exposure in the studied population which is representative for workers in the nuclear industry at present. According to the linear fits to our data an increase of the micronucleus frequency pro rata 0.5 per 1000 binucleated cells per year, related to the centromere-negative micronuclei, may be expected for workers with the maximal tolerable dose of 20 mSv/year.

  13. Centromere-independent accumulation of cohesin at ectopic heterochromatin sites induces chromosome stretching during anaphase.

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    Raquel A Oliveira

    2014-10-01

    Full Text Available Pericentric heterochromatin, while often considered as "junk" DNA, plays important functions in chromosome biology. It contributes to sister chromatid cohesion, a process mediated by the cohesin complex that ensures proper genome segregation during nuclear division. Long stretches of heterochromatin are almost exclusively placed at centromere-proximal regions but it remains unclear if there is functional (or mechanistic importance in linking the sites of sister chromatid cohesion to the chromosomal regions that mediate spindle attachment (the centromere. Using engineered chromosomes in Drosophila melanogaster, we demonstrate that cohesin enrichment is dictated by the presence of heterochromatin rather than centromere proximity. This preferential accumulation is caused by an enrichment of the cohesin-loading factor (Nipped-B/NIPBL/Scc2 at dense heterochromatic regions. As a result, chromosome translocations containing ectopic pericentric heterochromatin embedded in euchromatin display additional cohesin-dependent constrictions. These ectopic cohesion sites, placed away from the centromere, disjoin abnormally during anaphase and chromosomes exhibit a significant increase in length during anaphase (termed chromatin stretching. These results provide evidence that long stretches of heterochromatin distant from the centromere, as often found in many cancers, are sufficient to induce abnormal accumulation of cohesin at these sites and thereby compromise the fidelity of chromosome segregation.

  14. Sisters unbound is required for meiotic centromeric cohesion in Drosophila melanogaster.

    Science.gov (United States)

    Krishnan, Badri; Thomas, Sharon E; Yan, Rihui; Yamada, Hirotsugu; Zhulin, Igor B; McKee, Bruce D

    2014-11-01

    Regular meiotic chromosome segregation requires sister centromeres to mono-orient (orient to the same pole) during the first meiotic division (meiosis I) when homologous chromosomes segregate, and to bi-orient (orient to opposite poles) during the second meiotic division (meiosis II) when sister chromatids segregate. Both orientation patterns require cohesion between sister centromeres, which is established during meiotic DNA replication and persists until anaphase of meiosis II. Meiotic cohesion is mediated by a conserved four-protein complex called cohesin that includes two structural maintenance of chromosomes (SMC) subunits (SMC1 and SMC3) and two non-SMC subunits. In Drosophila melanogaster, however, the meiotic cohesion apparatus has not been fully characterized and the non-SMC subunits have not been identified. We have identified a novel Drosophila gene called sisters unbound (sunn), which is required for stable sister chromatid cohesion throughout meiosis. sunn mutations disrupt centromere cohesion during prophase I and cause high frequencies of non-disjunction (NDJ) at both meiotic divisions in both sexes. SUNN co-localizes at centromeres with the cohesion proteins SMC1 and SOLO in both sexes and is necessary for the recruitment of both proteins to centromeres. Although SUNN lacks sequence homology to cohesins, bioinformatic analysis indicates that SUNN may be a structural homolog of the non-SMC cohesin subunit stromalin (SA), suggesting that SUNN may serve as a meiosis-specific cohesin subunit. In conclusion, our data show that SUNN is an essential meiosis-specific Drosophila cohesion protein.

  15. The role of dicentric chromosome formation and secondary centromere deletion in the evolution of myeloid malignancy.

    Science.gov (United States)

    Mackinnon, Ruth N; Campbell, Lynda J

    2011-01-01

    Dicentric chromosomes have been identified as instigators of the genome instability associated with cancer, but this instability is often resolved by one of a number of different secondary events. These include centromere inactivation, inversion, and intercentromeric deletion. Deletion or excision of one of the centromeres may be a significant occurrence in myeloid malignancy and other malignancies but has not previously been widely recognized, and our reports are the first describing centromere deletion in cancer cells. We review what is known about dicentric chromosomes and the mechanisms by which they can undergo stabilization in both constitutional and cancer genomes. The failure to identify centromere deletion in cancer cells until recently can be partly explained by the standard approaches to routine diagnostic cancer genome analysis, which do not identify centromeres in the context of chromosome organization. This hitherto hidden group of primary dicentric, secondary monocentric chromosomes, together with other unrecognized dicentric chromosomes, points to a greater role for dicentric chromosomes in cancer initiation and progression than is generally acknowledged. We present a model that predicts and explains a significant role for dicentric chromosomes in the formation of unbalanced translocations in malignancy.

  16. Cultivation and differentiation change nuclear localization of chromosome centromeres in human mesenchymal stem cells.

    Science.gov (United States)

    Voldgorn, Yana I; Adilgereeva, Elmira P; Nekrasov, Evgeny D; Lavrov, Alexander V

    2015-01-01

    Chromosome arrangement in the interphase nucleus is not accidental. Strong evidences support that nuclear localization is an important mechanism of epigenetic regulation of gene expression. The purpose of this research was to identify differences in the localization of centromeres of chromosomes 6, 12, 18 and X in human mesenchymal stem cells depending on differentiation and cultivating time. We analyzed centromere positions in more than 4000 nuclei in 19 mesenchymal stem cell cultures before and after prolonged cultivation and after differentiation into osteogenic and adipogenic directions. We found a centromere reposition of HSAX at late passages and after differentiation in osteogenic direction as well as of HSA12 and HSA18 after adipogenic differentiation. The observed changes of the nuclear structure are new nuclear characteristics of the studied cells which may reflect regulatory changes of gene expression during the studied processes.

  17. Cultivation and differentiation change nuclear localization of chromosome centromeres in human mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Yana I Voldgorn

    Full Text Available Chromosome arrangement in the interphase nucleus is not accidental. Strong evidences support that nuclear localization is an important mechanism of epigenetic regulation of gene expression. The purpose of this research was to identify differences in the localization of centromeres of chromosomes 6, 12, 18 and X in human mesenchymal stem cells depending on differentiation and cultivating time. We analyzed centromere positions in more than 4000 nuclei in 19 mesenchymal stem cell cultures before and after prolonged cultivation and after differentiation into osteogenic and adipogenic directions. We found a centromere reposition of HSAX at late passages and after differentiation in osteogenic direction as well as of HSA12 and HSA18 after adipogenic differentiation. The observed changes of the nuclear structure are new nuclear characteristics of the studied cells which may reflect regulatory changes of gene expression during the studied processes.

  18. Formation of functional CENP-B boxes at diverse locations in repeat units of centromeric DNA in New World monkeys.

    Science.gov (United States)

    Kugou, Kazuto; Hirai, Hirohisa; Masumoto, Hiroshi; Koga, Akihiko

    2016-06-13

    Centromere protein B, which is involved in centromere formation, binds to centromeric repetitive DNA by recognizing a nucleotide motif called the CENP-B box. Humans have large numbers of CENP-B boxes in the centromeric repetitive DNA of their autosomes and X chromosome. The current understanding is that these CENP-B boxes are located at identical positions in the repeat units of centromeric DNA. Great apes also have CENP-B boxes in locations that are identical to humans. The purpose of the present study was to examine the location of CENP-B box in New World monkeys. We recently identified CENP-B box in one species of New World monkeys (marmosets). In this study, we found functional CENP-B boxes in CENP-A-assembled repeat units of centromeric DNA in 2 additional New World monkeys (squirrel monkeys and tamarins) by immunostaining and ChIP-qPCR analyses. The locations of the 3 CENP-B boxes in the repeat units differed from one another. The repeat unit size of centromeric DNA of New World monkeys (340-350 bp) is approximately twice that of humans and great apes (171 bp). This might be, associated with higher-order repeat structures of centromeric DNA, a factor for the observed variation in the CENP-B box location in New World monkeys.

  19. Replication initiator DnaA binds at the Caulobacter centromere and enables chromosome segregation.

    Science.gov (United States)

    Mera, Paola E; Kalogeraki, Virginia S; Shapiro, Lucy

    2014-11-11

    During cell division, multiple processes are highly coordinated to faithfully generate genetically equivalent daughter cells. In bacteria, the mechanisms that underlie the coordination of chromosome replication and segregation are poorly understood. Here, we report that the conserved replication initiator, DnaA, can mediate chromosome segregation independent of replication initiation. It does so by binding directly to the parS centromere region of the chromosome, and mutations that alter this interaction result in cells that display aberrant centromere translocation and cell division. We propose that DnaA serves to coordinate bacterial DNA replication with the onset of chromosome segregation.

  20. Physical Characterization of human centromeric regions using transformation-associated recombination cloning technology

    Energy Technology Data Exchange (ETDEWEB)

    Vladimir Larionov, Ph D

    2007-06-05

    A special interest in the organization of human centromeric DNA was stimulated a few years ago when two independent groups succeeded in reconstituting a functional human centromere, using constructs carrying centromere-specific alphoid DNA arrays. This work demonstrated the importance of DNA components in mammalian centromeres and opened a way for studying the structural requirements for de novo kinetochore formation and for construction of human artificial chromosomes (HACs) with therapeutic potential. To elucidate the structural requirements for formation of HACs with a functional kinetochore, we developed a new method for cloning of large DNA fragments for human centromeric regions that can be used as a substrate for HAC formation. This method exploits in vivo recombination in yeast (TAR cloning). In addition, a new strategy for the construction of alphoid DNA arrays was developed in our lab. The strategy involves the construction of uniform or hybrid synthetic alphoid DNA arrays by the RCA-TAR technique. This technique comprises two steps: rolling circle amplification of an alphoid DNA dimer and subsequent assembling of the amplified fragments by in vivo homologous recombination in yeast (Figure 1). Using this system, we constructed a set of different synthetic alphoid DNA arrays with a predetermined sequence varying in size from 30 to 140 kb and demonstrated that some of the arrays are competent in HAC formation. Because any nucleotide can be changed in a dimer before its amplification, this new technique is optimal for identifying the structural requirements for de novo kinetochore formation in HACs. Moreover, the technique makes possible to introduce into alphoid DNA arrays recognition sites for DNA-binding proteins. We have made the following progress on the studying of human centromeric regions using transformation-associated recombination cloning technology: i) minimal size of alphoid DNA array required for de novo kinetochore formation was estimated; ii

  1. Recognition of the centromere-specific histone Cse4 by the chaperone Scm3

    OpenAIRE

    Cho, Uhn-Soo; Harrison, Stephen C.

    2011-01-01

    A specialized nucleosome is a component of all eukaryotic kinetochores. The core of this nucleosome contains a centromere-specific histone, CENP-A (the Cse4 gene product in budding yeast), instead of the usual H3. Assembly of a centromeric nucleosome depends on a specific chaperone, called Scm3 in yeast and HJURP in higher eukaryotes. We describe here the structure of a complex formed by an N-terminal fragment of Scm3 with the histone-fold domains of Cse4, and H4, all prepared as recombinant ...

  2. Inter-domain Cooperation in INCENP Promotes Aurora B Relocation from Centromeres to Microtubules

    Directory of Open Access Journals (Sweden)

    Armando van der Horst

    2015-07-01

    Full Text Available The chromosomal passenger complex is essential for error-free chromosome segregation and proper execution of cytokinesis. To coordinate nuclear division with cytoplasmic division, its enzymatic subunit, Aurora B, relocalizes from centromeres in metaphase to the spindle midzone in anaphase. In budding yeast, this requires dephosphorylation of the microtubule-binding (MTB domain of the INCENP analog Sli15. The mechanistic basis for this relocalization in metazoans is incompletely understood. We demonstrate that the putative coiled-coil domain within INCENP drives midzone localization of Aurora B via a direct, electrostatic interaction with microtubules. Furthermore, we provide evidence that the CPC multimerizes via INCENP’s centromere-targeting domain (CEN box, which increases the MTB affinity of INCENP. In (prometaphase, the MTB affinity of INCENP is outcompeted by the affinity of its CEN box for centromeres, while at anaphase onset—when the histone mark H2AT120 is dephosphorylated—INCENP and Aurora B switch from centromere to microtubule localization.

  3. Inter-domain Cooperation in INCENP Promotes Aurora B Relocation from Centromeres to Microtubules

    NARCIS (Netherlands)

    van der Horst, Armando; Vromans, Martijn J M; Bouwman, Kim; van der Waal, Maike S; Hadders, Michael A; Lens, Susanne M A; Lens, SMA

    2015-01-01

    The chromosomal passenger complex is essential for error-free chromosome segregation and proper execution of cytokinesis. To coordinate nuclear division with cytoplasmic division, its enzymatic subunit, Aurora B, relocalizes from centromeres in metaphase to the spindle midzone in anaphase. In buddin

  4. Chromatin determinants of the inner-centromere rely on replication factors with functions that impart cohesion.

    Science.gov (United States)

    Abe, Takuya; Kawasumi, Ryotaro; Arakawa, Hiroshi; Hori, Tetsuya; Shirahige, Katsuhiko; Losada, Ana; Fukagawa, Tatsuo; Branzei, Dana

    2016-10-18

    Replication fork-associated factors promote genome integrity and protect against cancer. Mutations in the DDX11 helicase and the ESCO2 acetyltransferase also cause related developmental disorders classified as cohesinopathies. Here we generated vertebrate model cell lines of these disorders and cohesinopathies-related genes. We found that vertebrate DDX11 and Tim-Tipin are individually needed to compensate for ESCO2 loss in chromosome segregation, with DDX11 also playing complementary roles with ESCO2 in centromeric cohesion. Our study reveals that overt centromeric cohesion loss does not necessarily precede chromosome missegregation, while both these problems correlate with, and possibly originate from, inner-centromere defects involving reduced phosphorylation of histone H3T3 (pH3T3) in the region. Interestingly, the mitotic pH3T3 mark was defective in all analyzed replication-related mutants with functions in cohesion. The results pinpoint mitotic pH3T3 as a postreplicative chromatin mark that is sensitive to replication stress and conducts with different kinetics to robust centromeric cohesion and correct chromosome segregation.

  5. Centromeres cluster de novo at the beginning of meiosis in Brachypodium distachyon.

    Directory of Open Access Journals (Sweden)

    Ruoyu Wen

    Full Text Available In most eukaryotes that have been studied, the telomeres cluster into a bouquet early in meiosis, and in wheat and its relatives and in Arabidopsis the centromeres pair at the same time. In Arabidopsis, the telomeres do not cluster as a typical telomere bouquet on the nuclear membrane but are associated with the nucleolus both somatically and at the onset of meiosis. We therefore assessed whether Brachypodium distachyon, a monocot species related to cereals and whose genome is approximately twice the size of Arabidopsis thaliana, also exhibited an atypical telomere bouquet and centromere pairing. In order to investigate the occurrence of a bouquet and centromere pairing in B distachyon, we first had to establish protocols for studying meiosis in this species. This enabled us to visualize chromosome behaviour in meiocytes derived from young B distachyon spikelets in three-dimensions by fluorescent in situ hybridization (FISH, and accurately to stage meiosis based on chromatin morphology in relation to spikelet size and the timing of sample collection. Surprisingly, this study revealed that the centromeres clustered as a single site at the same time as the telomeres also formed a bouquet or single cluster.

  6. Mutations in ZBTB24 are associated with immunodeficiency, centromeric instability, and facial anomalies syndrome type 2

    NARCIS (Netherlands)

    Greef, J.C. de; Wang, J.; Balog, J.; Dunnen, J.T. den; Frants, R.R.; Straasheijm, K.R.; Aytekin, C.; Burg, M. van der; Duprez, L.; Ferster, A.; Gennery, A.R.; Gimelli, G.; Reisli, I.; Schuetz, C.; Schulz, A.; Smeets, D.F.C.M.; Sznajer, Y.; Wijmenga, C.; Eggermond, M.C. van; Ostaijen-ten Dam, M.M. van; Lankester, A.C.; Tol, M.J. van; Elsen, P.J. van den; Weemaes, C.M.R.; Maarel, S.M. van der

    2011-01-01

    Autosomal-recessive immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome is mainly characterized by recurrent, often fatal, respiratory and gastrointestinal infections. About 50% of patients carry mutations in the DNA methyltransferase 3B gene (DNMT3B) (ICF1). The remaining

  7. Tension sensing by Aurora B kinase is independent of survivin-based centromere localization.

    Science.gov (United States)

    Campbell, Christopher S; Desai, Arshad

    2013-05-01

    Accurate segregation of the replicated genome requires chromosome biorientation on the spindle. Biorientation is ensured by Aurora B kinase (Ipl1), a member of the four-subunit chromosomal passenger complex (CPC). Localization of the CPC to the inner centromere is central to the current model for how tension ensures chromosome biorientation: kinetochore-spindle attachments that are not under tension remain close to the inner centromere and are destabilized by Aurora B phosphorylation, whereas kinetochores under tension are pulled away from the influence of Aurora B, stabilizing their microtubule attachments. Here we show that an engineered truncation of the Sli15 (known as INCENP in humans) subunit of budding yeast CPC that eliminates association with the inner centromere nevertheless supports proper chromosome segregation during both mitosis and meiosis. Truncated Sli15 suppresses the deletion phenotypes of the inner-centromere-targeting proteins survivin (Bir1), borealin (Nbl1), Bub1 and Sgo1 (ref. 6). Unlike wild-type Sli15, truncated Sli15 localizes to pre-anaphase spindle microtubules. Premature targeting of full-length Sli15 to microtubules by preventing Cdk1 (also known as Cdc28) phosphorylation also suppresses the inviability of Bir1 deletion. These results suggest that activation of Aurora B kinase by clustering either on chromatin or on microtubules is sufficient for chromosome biorientation.

  8. [Chromosomal organization of centromeric Ty3/gypsy retrotransposons in Allium cepa L. and Allium fistulosum L].

    Science.gov (United States)

    Kiseleva, A V; Kirov, I V; Khrustaleva, L I

    2014-06-01

    This is the first report on the presence of Ty3/gypsy-like retrotransposons in the centromeric region of Allium cepa and Allium fistulosum. The paper identifies the putative Ty3/gypsy centromeric retrotransposons (CR) among the DNA sequences of A. cepa present in the NCBI database and evaluates their copy number in the genomes of Allium cepa and Allium fistulosum. The putative copy number of Ty3/gypsy CR constituted about 26000 for A. cepa and about 7000 for A. fistulosum. The chromosomal organization of Ty3/gypsy CR was analyzed with the help of fluorescent in situ hybridization (FISH). The 300-bp PCR products synthesized with genomic DNA of Allium cepa and Allium fistulosum and primers designed for the sequence ET645811 of A. cepa (Genome Survey Sequence database), displaying similarity to the reverse transcriptase of the CR Ty3/gypsy family, served as FISH hybridization probes. On the chromosomes of A. cepa, hybridization signals were mainly localized in the centromeric region. On the chromosomes of A. fistulosum the signals were less expressed in the centromeric regions, though they were abundant in other chromosomal regions. The pathways of evolution in these closely related species are discussed.

  9. Novel Centromeric Loci of the Wine and Beer Yeast Dekkera bruxellensis CEN1 and CEN2

    DEFF Research Database (Denmark)

    Ishchuk, Olena P.; Vojvoda Zeljko, Tanja; Schifferdecker, Anna J.

    2016-01-01

    The wine and beer yeast Dekkera bruxellensis thrives in environments that are harsh and limiting, especially in concentrations with low oxygen and high ethanol. Its different strains' chromosomes greatly vary in number (karyotype). This study isolates two novel centromeric loci (CEN1 and CEN2...

  10. Survivin mediates targeting of the chromosomal passenger complex to the centromere and midbody

    NARCIS (Netherlands)

    Vader, G; Kauw, JJW; Medema, RH; Lens, SMA

    2006-01-01

    The chromosomal passenger complex (CPC) coordinates chromosomal and cytoskeletal events of mitosis. The enzymatic core of this complex (Aurora-B) is guided through the mitotic cell by its companion chromosomal passenger proteins, inner centromere protein (INCENP), Survivin and Borealin/Dasra-B, ther

  11. Correlation between centromere protein-F autoantibodies and cancer analyzed by enzyme-linked immunosorbent assay

    DEFF Research Database (Denmark)

    Welner, Simon; Trier, Nicole Hartwig; Morten Frisch, Morten

    2013-01-01

    Centromere protein-F (CENP-F) is a large nuclear protein of 367 kDa, which is involved in multiple mitosis-related events such as proper assembly of the kinetochores, stabilization of heterochromatin, chromosome alignment and mitotic checkpoint signaling. Several studies have shown a correlation...

  12. Loss of pRB causes centromere dysfunction and chromosomal instability.

    Science.gov (United States)

    Manning, Amity L; Longworth, Michelle S; Dyson, Nicholas J

    2010-07-01

    Chromosome instability (CIN) is a common feature of tumor cells. By monitoring chromosome segregation, we show that depletion of the retinoblastoma protein (pRB) causes rates of missegregation comparable with those seen in CIN tumor cells. The retinoblastoma tumor suppressor is frequently inactivated in human cancers and is best known for its regulation of the G1/S-phase transition. Recent studies have shown that pRB inactivation also slows mitotic progression and promotes aneuploidy, but reasons for these phenotypes are not well understood. Here we describe the underlying mitotic defects of pRB-deficient cells that cause chromosome missegregation. Analysis of mitotic cells reveals that pRB depletion compromises centromeric localization of CAP-D3/condensin II and chromosome cohesion, leading to an increase in intercentromeric distance and deformation of centromeric structure. These defects promote merotelic attachment, resulting in failure of chromosome congression and an increased propensity for lagging chromosomes following mitotic delay. While complete loss of centromere function or chromosome cohesion would have catastrophic consequences, these more moderate defects allow pRB-deficient cells to proliferate but undermine the fidelity of mitosis, leading to whole-chromosome gains and losses. These observations explain an important consequence of RB1 inactivation, and suggest that subtle defects in centromere function are a frequent source of merotely and CIN in cancer.

  13. Aurora B kinase controls the separation of centromeric and telomeric heterochromatin

    OpenAIRE

    Gachet, Yannick; Reyes, Celine; Tournier, Sylvie

    2015-01-01

    The segregation of chromosomes is coordinated at multiple levels to prevent chromosome loss, a phenotype frequently observed in cancers. We recently described an essential role for telomeres in the physical separation of chromosomes and identified Aurora B kinase as a double agent involved in the separation of centromeric and telomeric heterochromatin.

  14. Aurora B kinase controls the separation of centromeric and telomeric heterochromatin.

    Science.gov (United States)

    Gachet, Yannick; Reyes, Celine; Tournier, Sylvie

    2016-03-01

    The segregation of chromosomes is coordinated at multiple levels to prevent chromosome loss, a phenotype frequently observed in cancers. We recently described an essential role for telomeres in the physical separation of chromosomes and identified Aurora B kinase as a double agent involved in the separation of centromeric and telomeric heterochromatin.

  15. Characterization of centromeric histone H3 (CENH3 variants in cultivated and wild carrots (Daucus sp..

    Directory of Open Access Journals (Sweden)

    Frank Dunemann

    Full Text Available In eukaryotes, centromeres are the assembly sites for the kinetochore, a multi-protein complex to which spindle microtubules are attached at mitosis and meiosis, thereby ensuring segregation of chromosomes during cell division. They are specified by incorporation of CENH3, a centromere specific histone H3 variant which replaces canonical histone H3 in the nucleosomes of functional centromeres. To lay a first foundation of a putative alternative haploidization strategy based on centromere-mediated genome elimination in cultivated carrots, in the presented research we aimed at the identification and cloning of functional CENH3 genes in Daucus carota and three distantly related wild species of genus Daucus varying in basic chromosome numbers. Based on mining the carrot transcriptome followed by a subsequent PCR-based cloning, homologous coding sequences for CENH3s of the four Daucus species were identified. The ORFs of the CENH3 variants were very similar, and an amino acid sequence length of 146 aa was found in three out of the four species. Comparison of Daucus CENH3 amino acid sequences with those of other plant CENH3s as well as their phylogenetic arrangement among other dicot CENH3s suggest that the identified genes are authentic CENH3 homologs. To verify the location of the CENH3 protein in the kinetochore regions of the Daucus chromosomes, a polyclonal antibody based on a peptide corresponding to the N-terminus of DcCENH3 was developed and used for anti-CENH3 immunostaining of mitotic root cells. The chromosomal location of CENH3 proteins in the centromere regions of the chromosomes could be confirmed. For genetic localization of the CENH3 gene in the carrot genome, a previously constructed linkage map for carrot was used for mapping a CENH3-specific simple sequence repeat (SSR marker, and the CENH3 locus was mapped on the carrot chromosome 9.

  16. Transgenerational propagation and quantitative maintenance of paternal centromeres depends on Cid/Cenp-A presence in Drosophila sperm.

    Directory of Open Access Journals (Sweden)

    Nitika Raychaudhuri

    Full Text Available In Drosophila melanogaster, as in many animal and plant species, centromere identity is specified epigenetically. In proliferating cells, a centromere-specific histone H3 variant (CenH3, named Cid in Drosophila and Cenp-A in humans, is a crucial component of the epigenetic centromere mark. Hence, maintenance of the amount and chromosomal location of CenH3 during mitotic proliferation is important. Interestingly, CenH3 may have different roles during meiosis and the onset of embryogenesis. In gametes of Caenorhabditis elegans, and possibly in plants, centromere marking is independent of CenH3. Moreover, male gamete differentiation in animals often includes global nucleosome for protamine exchange that potentially could remove CenH3 nucleosomes. Here we demonstrate that the control of Cid loading during male meiosis is distinct from the regulation observed during the mitotic cycles of early embryogenesis. But Cid is present in mature sperm. After strong Cid depletion in sperm, paternal centromeres fail to integrate into the gonomeric spindle of the first mitosis, resulting in gynogenetic haploid embryos. Furthermore, after moderate depletion, paternal centromeres are unable to re-acquire normal Cid levels in the next generation. We conclude that Cid in sperm is an essential component of the epigenetic centromere mark on paternal chromosomes and it exerts quantitative control over centromeric Cid levels throughout development. Hence, the amount of Cid that is loaded during each cell cycle appears to be determined primarily by the preexisting centromeric Cid, with little flexibility for compensation of accidental losses.

  17. Esperanto for histones: CENP-A, not CenH3, is the centromeric histone H3 variant.

    Science.gov (United States)

    Earnshaw, W C; Allshire, R C; Black, B E; Bloom, K; Brinkley, B R; Brown, W; Cheeseman, I M; Choo, K H A; Copenhaver, G P; Deluca, J G; Desai, A; Diekmann, S; Erhardt, S; Fitzgerald-Hayes, M; Foltz, D; Fukagawa, T; Gassmann, R; Gerlich, D W; Glover, D M; Gorbsky, G J; Harrison, S C; Heun, P; Hirota, T; Jansen, L E T; Karpen, G; Kops, G J P L; Lampson, M A; Lens, S M; Losada, A; Luger, K; Maiato, H; Maddox, P S; Margolis, R L; Masumoto, H; McAinsh, A D; Mellone, B G; Meraldi, P; Musacchio, A; Oegema, K; O'Neill, R J; Salmon, E D; Scott, K C; Straight, A F; Stukenberg, P T; Sullivan, B A; Sullivan, K F; Sunkel, C E; Swedlow, J R; Walczak, C E; Warburton, P E; Westermann, S; Willard, H F; Wordeman, L; Yanagida, M; Yen, T J; Yoda, K; Cleveland, D W

    2013-04-01

    The first centromeric protein identified in any species was CENP-A, a divergent member of the histone H3 family that was recognised by autoantibodies from patients with scleroderma-spectrum disease. It has recently been suggested to rename this protein CenH3. Here, we argue that the original name should be maintained both because it is the basis of a long established nomenclature for centromere proteins and because it avoids confusion due to the presence of canonical histone H3 at centromeres.

  18. Genetic and physical mapping of two centromere-proximal regions of chromosome IV in Aspergillus nidulans

    DEFF Research Database (Denmark)

    Aleksenko, Alexei Y.; Nielsen, Michael Lynge; Clutterbuck, A.J.

    2001-01-01

    The centromere-proximal portion of the chromosome was mapped physically using overlapping clones of a cosmid genomic library. Two contiguous segments of a physical map, based on restriction mapping of cosmid clones, were generated, together covering more than 0.4 Mb DNA. A reverse genetic mapping...... approach was used to establish a correlation between physical and genetic maps; i,e., marker genes were integrated into physically mapped segments and subsequently mapped by mitotic and meiotic recombination. The resulting data, together with additional classical genetic mapping, lead to a substantial...... revision of the genetic map of the chromosome, including the position of the centromere, Comparison of physical and genetic maps indicates that meiotic recombination is low in subcentromeric DNA, its frequency being reduced from 1 crossover per 0.8 Mb to approximately 1 crossover per 5 Mb per meiosis...

  19. Posttranslational modification of CENP-A influences the conformation of centromeric chromatin.

    Science.gov (United States)

    Bailey, Aaron O; Panchenko, Tanya; Sathyan, Kizhakke M; Petkowski, Janusz J; Pai, Pei-Jing; Bai, Dina L; Russell, David H; Macara, Ian G; Shabanowitz, Jeffrey; Hunt, Donald F; Black, Ben E; Foltz, Daniel R

    2013-07-16

    Centromeres are chromosomal loci required for accurate segregation of sister chromatids during mitosis. The location of the centromere on the chromosome is not dependent on DNA sequence, but rather it is epigenetically specified by the histone H3 variant centromere protein A (CENP-A). The N-terminal tail of CENP-A is highly divergent from other H3 variants. Canonical histone N termini are hotspots of conserved posttranslational modification; however, no broadly conserved modifications of the vertebrate CENP-A tail have been previously observed. Here, we report three posttranslational modifications on human CENP-A N termini using high-resolution MS: trimethylation of Gly1 and phosphorylation of Ser16 and Ser18. Our results demonstrate that CENP-A is subjected to constitutive initiating methionine removal, similar to other H3 variants. The nascent N-terminal residue Gly1 becomes trimethylated on the α-amino group. We demonstrate that the N-terminal RCC1 methyltransferase is capable of modifying the CENP-A N terminus. Methylation occurs in the prenucleosomal form and marks the majority of CENP-A nucleosomes. Serine 16 and 18 become phosphorylated in prenucleosomal CENP-A and are phosphorylated on asynchronous and mitotic nucleosomal CENP-A and are important for chromosome segregation during mitosis. The double phosphorylation motif forms a salt-bridged secondary structure and causes CENP-A N-terminal tails to form intramolecular associations. Analytical ultracentrifugation of phospho-mimetic CENP-A nucleosome arrays demonstrates that phosphorylation results in greater intranucleosome associations and counteracts the hyperoligomerized state exhibited by unmodified CENP-A nucleosome arrays. Our studies have revealed that the major modifications on the N-terminal tail of CENP-A alter the physical properties of the chromatin fiber at the centromere.

  20. Promiscuous stimulation of ParF protein polymerization by heterogeneous centromere binding factors.

    Science.gov (United States)

    Machón, Cristina; Fothergill, Timothy J G; Barillà, Daniela; Hayes, Finbarr

    2007-11-16

    The segrosome is the nucleoprotein complex that mediates accurate segregation of bacterial plasmids. The segrosome of plasmid TP228 comprises ParF and ParG proteins that assemble on the parH centromere. ParF, which exemplifies one clade of the ubiquitous ParA superfamily of segregation proteins, polymerizes extensively in response to ATP binding. Polymerization is modulated by the ParG centromere binding factor (CBF). The segrosomes of plasmids pTAR, pVT745 and pB171 include ParA homologues of the ParF subgroup, as well as diverse homodimeric CBFs with no primary sequence similarity to ParG, or each other. Centromere binding by these analogues is largely specific. Here, we establish that the ParF homologues of pTAR and pB171 filament modestly with ATP, and that nucleotide hydrolysis is not required for this polymerization, which is more prodigious when the cognate CBF is also present. By contrast, the ParF homologue of plasmid pVT745 did not respond appreciably to ATP alone, but polymerized extensively in the presence of both its cognate CBF and ATP. The co-factors also stimulated nucleotide-independent polymerization of cognate ParF proteins. Moreover, apart from the CBF of pTAR, the disparate ParG analogues promoted polymerization of non-cognate ParF proteins suggesting that filamentation of the ParF proteins is enhanced by a common mechanism. Like ParG, the co-factors may be modular, possessing a centromere-specific interaction domain linked to a flexible region containing determinants that promiscuously stimulate ParF polymerization. The CBFs appear to function as bacterial analogues of formins, microtubule-associated proteins or related ancillary factors that regulate eucaryotic cytoskeletal dynamics.

  1. Identification of a centromeric exchange of acrocentric chromosomes by fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Yu, C.W.; Immken, L.; Curry, C.J.R. [UCSF, Fresno, CA (United States)] [and others

    1994-09-01

    Exchanges of the peri-centromeric area of acrocentric chromosomes are difficult to identify using the conventional cytogenetic techniques. Fluorescence in situ hybridization (FISH) provides a new way for precisely identifying such rearrangements. Here we report a case of centromeric rearrangement in an amniotic fluid specimen with an extra marker chromosome. M.G., a 41-year-old G1, was referred for advanced maternal age. Chromosome studies revealed a 47,XX +mar karyotype. The marker appeared to be bi-satallited with a single C band. Chromosome studies from the parents were normal. The parents elected to terminate the pregnancy. Anatomical examination of the abortus revealed a very short neck, posteriorly rotated ears, high set cecum, absent hepatic lobation and low abdominal kidneys with short ureters. FISH studies with alpha-satellite probes of 13/21, 14/22, and 15, and the DiGeorge probe, indicated that there is a translocation of 21 alpha-satellite to the 22, and that the marker chromosome probably consists of 14/22 alpha-satellite material. FISH analysis of the parents chromosome revealed that father had the translocation of 21 alpha-satellite to the 22 as well. Exchanges of centromeric material among the acrocentric chromosomes may not be an uncommon event in humans. Although it probably has no clinical significance, it may result in non-disjunction or marker chromosome formation from an uncommon satellite association. With the use of FISH techniques, exchanges involving the centromeric regions of acrocentric chromosomes can be identified.

  2. A stable acentric marker chromosome: Possible existence of an intercalary ancient centromere at distal 8p

    Energy Technology Data Exchange (ETDEWEB)

    Ohashi, Hirofumi; Fukushima, Yoshimitsu; Wakui, Keiko; Ogawa, Kioyshi [Saitama Children`s Medical Center, Iwatsuki (Japan); Okano, Tetsuroh [Kitazato Univ., Tokyo (Japan); Niikawa, Norio [Nagasaki Univ. School of Medicine (Japan)

    1994-12-01

    A centromere is considered to be an essential chromosomal component where microtubule-kinetochore interaction occurs to segregate sister chromatids faithfully and acentric chromosomes are unstable and lost through cell divisions. We report a novel marker chromosome that was acentric but stable through cell divisions. The patient was a 2-year-old girl with mental retardation, patent ductus arteriosus, and mild dysmorphic features. G-banded chromosome analysis revealed that an additional small marker chromosome was observed in all 100 cells examined. By the reverse-chromosome-painting method, the marker was found to originate from the distal region of 8p, and a subsequent two-color FISH analysis with cosmid probes around the region revealed that the marker was an inverted duplication interpreted as 8pter {yields} p23.1::p23.1 {yields} 8pter. No centromeric region was involved in the marker. By FISH, no {alpha}-satellite sequence was detected on the marker, while a telomere sequence was detected at each end. Anti-kinetochore immunostaining, using a serum from a patient with CREST (calcinosis, Raynaud syndrome, esophageal dismotility, sclerodactyly, and telangiectasia) syndrome, showed a pair of signals on the marker, which indicated that a functional kinetochore was present on the marker. The analysis of this patient might suggest the possibility that an ancient centromere sequence exists at distal 8p (8p23.1-pter) and was activated through the chromosome rearrangement in the patient.

  3. Centromere binding and a conserved role in chromosome stability for SUMO-dependent ubiquitin ligases.

    Directory of Open Access Journals (Sweden)

    Loes A L van de Pasch

    Full Text Available The Saccharomyces cerevisiae Slx5/8 complex is the founding member of a recently defined class of SUMO-targeted ubiquitin ligases (STUbLs. Slx5/8 has been implicated in genome stability and transcription, but the precise contribution is unclear. To characterise Slx5/8 function, we determined genome-wide changes in gene expression upon loss of either subunit. The majority of mRNA changes are part of a general stress response, also exhibited by mutants of other genome integrity pathways and therefore indicative of an indirect effect on transcription. Genome-wide binding analysis reveals a uniquely centromeric location for Slx5. Detailed phenotype analyses of slx5Δ and slx8Δ mutants show severe mitotic defects that include aneuploidy, spindle mispositioning, fish hooks and aberrant spindle kinetics. This is associated with accumulation of the PP2A regulatory subunit Rts1 at centromeres prior to entry into anaphase. Knockdown of the human STUbL orthologue RNF4 also results in chromosome segregation errors due to chromosome bridges. The study shows that STUbLs have a conserved role in maintenance of chromosome stability and links SUMO-dependent ubiquitination to a centromere-specific function during mitosis.

  4. Shearing of the CENP-A dimerization interface mediates plasticity in the octameric centromeric nucleosome.

    Science.gov (United States)

    Winogradoff, David; Zhao, Haiqing; Dalal, Yamini; Papoian, Garegin A

    2015-11-25

    The centromeric nucleosome is a key epigenetic determinant of centromere identity and function. Consequently, deciphering how CENP-A containing nucleosomes contribute structurally to centromere function is a fundamental question in chromosome biology. Here, we performed microsecond timescale all-atom molecular dynamics (MD) simulations of CENP-A and H3 nucleosomes, and report that the octameric CENP-A core particles and nucleosomes display different dynamics from their canonical H3-containing counterparts. The most significant motion observed is within key interactions at the heart of the CENP-A octameric core, wherein shearing of contacts within the CENP-A:CENP-A' dimerization interface results in a weaker four helix bundle, and an extrusion of 10-30 bp of DNA near the pseudo-dyad. Coupled to other local and global fluctuations, the CENP-A nucleosome occupies a more rugged free energy landscape than the canonical H3 nucleosome. Taken together, our data suggest that CENP-A encodes enhanced distortability to the octameric nucleosome, which may allow for enhanced flexing of the histone core in vivo.

  5. ParB Partition Proteins: Complex Formation and Spreading at Bacterial and Plasmid Centromeres

    Directory of Open Access Journals (Sweden)

    Barbara Funnell

    2016-08-01

    Full Text Available In bacteria, active partition systems contribute to the faithful segregation of both chromosomes and low-copy-number plasmids. Each system depends on a site-specific DNA binding protein to recognize and assemble a partition complex at a centromere-like site, commonly called parS. Many plasmid and all chromosomal centromere-binding proteins are dimeric helix-turn-helix DNA binding proteins, which are commonly named ParB. Although the overall sequence conservation among ParBs is not high, the proteins share similar domain and functional organization, and they assemble into similar higher-order complexes. In vivo, ParBs spread; that is, DNA binding extends away from the parS site into the surrounding nonspecific DNA, a feature that reflects higher-order complex assembly. ParBs bridge and pair DNA at parS and nonspecific DNA sites. ParB dimers interact with each other via flexible conformations of an N-terminal region. This review will focus on the properties of the HTH centromere-binding protein, in light of recent experimental evidence and models that are adding to our understanding of how these proteins assemble into large and dynamic partition complexes at and around their specific DNA sites.

  6. Separase Is Required for Homolog and Sister Disjunction during Drosophila melanogaster Male Meiosis, but Not for Biorientation of Sister Centromeres.

    Directory of Open Access Journals (Sweden)

    Ariane C Blattner

    2016-04-01

    Full Text Available Spatially controlled release of sister chromatid cohesion during progression through the meiotic divisions is of paramount importance for error-free chromosome segregation during meiosis. Cohesion is mediated by the cohesin protein complex and cleavage of one of its subunits by the endoprotease separase removes cohesin first from chromosome arms during exit from meiosis I and later from the pericentromeric region during exit from meiosis II. At the onset of the meiotic divisions, cohesin has also been proposed to be present within the centromeric region for the unification of sister centromeres into a single functional entity, allowing bipolar orientation of paired homologs within the meiosis I spindle. Separase-mediated removal of centromeric cohesin during exit from meiosis I might explain sister centromere individualization which is essential for subsequent biorientation of sister centromeres during meiosis II. To characterize a potential involvement of separase in sister centromere individualization before meiosis II, we have studied meiosis in Drosophila melanogaster males where homologs are not paired in the canonical manner. Meiosis does not include meiotic recombination and synaptonemal complex formation in these males. Instead, an alternative homolog conjunction system keeps homologous chromosomes in pairs. Using independent strategies for spermatocyte-specific depletion of separase complex subunits in combination with time-lapse imaging, we demonstrate that separase is required for the inactivation of this alternative conjunction at anaphase I onset. Mutations that abolish alternative homolog conjunction therefore result in random segregation of univalents during meiosis I also after separase depletion. Interestingly, these univalents become bioriented during meiosis II, suggesting that sister centromere individualization before meiosis II does not require separase.

  7. Identification and Preliminary Analysis of Several Centromere-associated Bacterial Artificial Chromosome Clones from a Diploid Wheat Library

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Although the centromeres of some plants have been investigated previously, our knowledge of the wheat centromere is still very limited. To understand the structure and function of the wheat centromere, we used two centromeric repeats (RCS1 and CCS1-5ab) to obtain some centromere-associated bacterial artificial chromosome (BAC) clones in 32 RCS1-related BAC clones that had been screened out from a diploid wheat (Triticum boeoticum Boiss.; 2n=2x=14) BAC library. Southern hybridization results indicated that, of the 32 candidates,there were 28 RCS1-positive clones. Based on gel blot patterns, the frequency of RCS1 was approximately one copy every 69.4 kb in these 28 RCS1-positive BAC clones. More bands were detected when the same filter was probed with CCS1-5ab. Furthermore, the CCS1 bands covered all the bands detected by RCS1, which suggests that some CCS1 repeats were distributed together with RCS1. The frequency of CCS1 families was once every 35.8 kb, nearly twice that of RCS1. Fluorescence in situ hybridization (FISH) analysis indicated that the five BAC clones containing RCS1 and CCS1 sequences all detected signals at the centromeric regions in hexaploid wheat, but the signal intensities on the A-genome chromosomes were stronger than those on the B- and/or D-genome chromosomes. The FISH analysis among nine Triticeae cereals indicated that there were A-genomespecific (or rich) sequences dispersing on chromosome arms in the BAC clone TbBAC5. In addition, at the interphase cells, the centromeres of diploid species usually clustered at one pole and formed a ring-like allocation in the period before metaphase.

  8. Overexpression of SETβ, a protein localizing to centromeres, causes precocious separation of chromatids during the first meiosis of mouse oocytes.

    Science.gov (United States)

    Qi, Shu-Tao; Wang, Zhen-Bo; Ouyang, Ying-Chun; Zhang, Qing-Hua; Hu, Meng-Wen; Huang, Xin; Ge, Zhaojia; Guo, Lei; Wang, Ya-Peng; Hou, Yi; Schatten, Heide; Sun, Qing-Yuan

    2013-04-01

    Chromosome segregation in mammalian oocyte meiosis is an error-prone process, and any mistake in this process may result in aneuploidy, which is the main cause of infertility, abortion and many genetic diseases. It is now well known that shugoshin and protein phosphatase 2A (PP2A) play important roles in the protection of centromeric cohesion during the first meiosis. PP2A can antagonize the phosphorylation of rec8, a member of the cohesin complex, at the centromeres and thus prevent cleavage of rec8 and so maintain the cohesion of chromatids. SETβ is a protein that physically interacts with shugoshin and inhibits PP2A activity. We thus hypothesized that SETβ might regulate cohesion protection and chromosome segregation during oocyte meiotic maturation. Here we report for the first time the expression, subcellular localization and functions of SETβ during mouse oocyte meiosis. Immunoblotting analysis showed that the expression level of SETβ was stable from the germinal vesicle stage to the MII stage of oocyte meiosis. Immunofluorescence analysis showed SETβ accumulation in the nucleus at the germinal vesicle stage, whereas it was targeted mainly to the inner centromere area and faintly localized to the interchromatid axes from germinal vesicle breakdown to MI stages. At the MII stage, SETβ still localized to the inner centromere area, but could relocalize to kinetochores in a process perhaps dependent on the tension on the centromeres. SETβ partly colocalized with PP2A at the inner centromere area. Overexpression of SETβ in mouse oocytes caused precocious separation of sister chromatids, but depletion of SETβ by RNAi showed little effects on the meiotic maturation process. Taken together, our results suggest that SETβ, even though it localizes to centromeres, might not be essential for chromosome separation during mouse oocyte meiotic maturation, although its forced overexpression causes premature chromatid separation.

  9. Separase Is Required for Homolog and Sister Disjunction during Drosophila melanogaster Male Meiosis, but Not for Biorientation of Sister Centromeres.

    Science.gov (United States)

    Blattner, Ariane C; Chaurasia, Soumya; McKee, Bruce D; Lehner, Christian F

    2016-04-01

    Spatially controlled release of sister chromatid cohesion during progression through the meiotic divisions is of paramount importance for error-free chromosome segregation during meiosis. Cohesion is mediated by the cohesin protein complex and cleavage of one of its subunits by the endoprotease separase removes cohesin first from chromosome arms during exit from meiosis I and later from the pericentromeric region during exit from meiosis II. At the onset of the meiotic divisions, cohesin has also been proposed to be present within the centromeric region for the unification of sister centromeres into a single functional entity, allowing bipolar orientation of paired homologs within the meiosis I spindle. Separase-mediated removal of centromeric cohesin during exit from meiosis I might explain sister centromere individualization which is essential for subsequent biorientation of sister centromeres during meiosis II. To characterize a potential involvement of separase in sister centromere individualization before meiosis II, we have studied meiosis in Drosophila melanogaster males where homologs are not paired in the canonical manner. Meiosis does not include meiotic recombination and synaptonemal complex formation in these males. Instead, an alternative homolog conjunction system keeps homologous chromosomes in pairs. Using independent strategies for spermatocyte-specific depletion of separase complex subunits in combination with time-lapse imaging, we demonstrate that separase is required for the inactivation of this alternative conjunction at anaphase I onset. Mutations that abolish alternative homolog conjunction therefore result in random segregation of univalents during meiosis I also after separase depletion. Interestingly, these univalents become bioriented during meiosis II, suggesting that sister centromere individualization before meiosis II does not require separase.

  10. [Sop proteins can cause transcriptional silencing of genes located close to the centromere sites of linear plasmid N15].

    Science.gov (United States)

    Mardanov, A V; Lane, D; Ravin, N V

    2010-01-01

    Stable inheritance of bacterial chromosomes and low copy number plasmids is ensured by accurate partitioning of replicated molecules between the daughter cells at division. Partitioning of the prophage of the temperate bacteriophage N15, which exists as a linear plasmid molecule with covalently closed ends, depends on the sop locus, comprising genes sopA and sopB, as well as four centromere sites located in different regions of the N15 genome essential for replication and the control of lysogeny. We found that binding of SopB to the centromere can silence centromere-proximal promoters, presumably due to subsequent polymerizing of SopB along the DNA. Close to the IR4 centromere site we identified a promoter, P59, able to drive expression of phage late genes encoding the structural proteins of virion. We found that following binding to IR4 the N15 Sop proteins can cause repression of this promoter. The repression depends on SopB and became stronger in the presence of SopA. Sop-dependent silencing of centromere-proximal promoters control gene expression in phage N15, particularly preventing undesired expression of late genes in the N15 prophage. Thus, the phage N15 sop system not only ensures plasmid partitioning but is also involved in the genetic network controlling prophage replication and the maintenance of lysogeny.

  11. Characterization of the genomic organization of the region bordering the centromere of chromosome V of Podospora anserina by direct sequencing.

    Science.gov (United States)

    Silar, Philippe; Barreau, Christian; Debuchy, Robert; Kicka, Sébastien; Turcq, Béatrice; Sainsard-Chanet, Annie; Sellem, Carole H; Billault, Alain; Cattolico, Laurence; Duprat, Simone; Weissenbach, Jean

    2003-08-01

    A Podospora anserina BAC library of 4800 clones has been constructed in the vector pBHYG allowing direct selection in fungi. Screening of the BAC collection for centromeric sequences of chromosome V allowed the recovery of clones localized on either sides of the centromere, but no BAC clone was found to contain the centromere. Seven BAC clones containing 322,195 and 156,244bp from either sides of the centromeric region were sequenced and annotated. One 5S rRNA gene, 5 tRNA genes, and 163 putative coding sequences (CDS) were identified. Among these, only six CDS seem specific to P. anserina. The gene density in the centromeric region is approximately one gene every 2.8kb. Extrapolation of this gene density to the whole genome of P. anserina suggests that the genome contains about 11,000 genes. Synteny analyses between P. anserina and Neurospora crassa show that co-linearity extends at the most to a few genes, suggesting rapid genome rearrangements between these two species.

  12. Regulation of centromere localization of the Drosophila Shugoshin MEI-S332 and sister-chromatid cohesion in meiosis.

    Science.gov (United States)

    Nogueira, Cristina; Kashevsky, Helena; Pinto, Belinda; Clarke, Astrid; Orr-Weaver, Terry L

    2014-07-31

    The Shugoshin (Sgo) protein family helps to ensure proper chromosome segregation by protecting cohesion at the centromere by preventing cleavage of the cohesin complex. Some Sgo proteins also influence other aspects of kinetochore-microtubule attachments. Although many Sgo members require Aurora B kinase to localize to the centromere, factors controlling delocalization are poorly understood and diverse. Moreover, it is not clear how Sgo function is inactivated and whether this is distinct from delocalization. We investigated these questions in Drosophila melanogaster, an organism with superb chromosome cytology to monitor Sgo localization and quantitative assays to test its function in sister-chromatid segregation in meiosis. Previous research showed that in mitosis in cell culture, phosphorylation of the Drosophila Sgo, MEI-S332, by Aurora B promotes centromere localization, whereas Polo phosphorylation promotes delocalization. These studies also suggested that MEI-S332 can be inactivated independently of delocalization, a conclusion supported here by localization and function studies in meiosis. Phosphoresistant and phosphomimetic mutants for the Aurora B and Polo phosphorylation sites were examined for effects on MEI-S332 localization and chromosome segregation in meiosis. Strikingly, MEI-S332 with a phosphomimetic mutation in the Aurora B phosphorylation site prematurely dissociates from the centromeres in meiosis I. Despite the absence of MEI-S332 on meiosis II centromeres in male meiosis, sister chromatids segregate normally, demonstrating that detectable levels of this Sgo are not essential for chromosome congression, kinetochore biorientation, or spindle assembly.

  13. Differing requirements for RAD51 and DMC1 in meiotic pairing of centromeres and chromosome arms in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Olivier Da Ines

    Full Text Available During meiosis homologous chromosomes pair, recombine, and synapse, thus ensuring accurate chromosome segregation and the halving of ploidy necessary for gametogenesis. The processes permitting a chromosome to pair only with its homologue are not fully understood, but successful pairing of homologous chromosomes is tightly linked to recombination. In Arabidopsis thaliana, meiotic prophase of rad51, xrcc3, and rad51C mutants appears normal up to the zygotene/pachytene stage, after which the genome fragments, leading to sterility. To better understand the relationship between recombination and chromosome pairing, we have analysed meiotic chromosome pairing in these and in dmc1 mutant lines. Our data show a differing requirement for these proteins in pairing of centromeric regions and chromosome arms. No homologous pairing of mid-arm or distal regions was observed in rad51, xrcc3, and rad51C mutants. However, homologous centromeres do pair in these mutants and we show that this does depend upon recombination, principally on DMC1. This centromere pairing extends well beyond the heterochromatic centromere region and, surprisingly, does not require XRCC3 and RAD51C. In addition to clarifying and bringing the roles of centromeres in meiotic synapsis to the fore, this analysis thus separates the roles in meiotic synapsis of DMC1 and RAD51 and the meiotic RAD51 paralogs, XRCC3 and RAD51C, with respect to different chromosome domains.

  14. Stem-loop structures of the repetitive DNA sequences located at human centromeres

    Energy Technology Data Exchange (ETDEWEB)

    Gupta, G.; Garcia, A.E.; Ratliff, R.; Moyzis, R.K. [Los Alamos National Lab., NM (United States); Catasti, P.; Hong, Lin; Yau, P. [California Univ., Davis, CA (United States). Dept. of Biological Chemistry; Bradbury, E.M. [Los Alamos National Lab., NM (United States)]|[California Univ., Davis, CA (United States). Dept. of Biological Chemistry

    1993-09-01

    The presence of the highly conserved repetitive DNA sequences in the human centromeres argues for a special role of these sequences in their biological functions - most likely achieved by the formation of unusual structures. This prompted us to carry out quantitative one- and two-dimensional nuclear magnetic resonance (lD/2D NMR) spectroscopy to determine the structural properties of the human centromeric repeats, d(AATGG){sub n.d}(CCATT){sub n}. The studies on centromeric DNAs reveal that the complementary sequence, d(AATGG){sub n.d}(CCATT){sub n}, adopts the usual Watson-Crick B-DNA duplex and the pyrimidine-rich d(CCATT){sub n} strand is essentially a random coil. However, the purine-rich d(AATGG){sub n} strand is shown to adopt unusual stem-loop structures for repeat lengths, n=2,3,4, and 6. In addition to normal Watson-Crick A{center_dot}T pairs, the stem-loop structures are stabilized by mismatch A{center_dot}G and G{center_dot}G pairs in the stem and G-G-A stacking in the loop. Stem-loop structures of d(AATGG)n are independently verified by gel electrophoresis and nuclease digestion studies. Thermal melting studies show that the DNA repeats, d(AATGG){sub n}, are as stable as the corresponding Watson-Crick duplex d(AATGG){sub n.d}(CCATT){sub n}. Therefore, the sequence d(AATGG){sub n} can, indeed, nucleate a stem-loop structure at little free-energy cost and if, during mitosis, they are located on the chromosome surface they can provide specific recognition sites for kinetochore function.

  15. Cytological identification of an isotetrasomic in rice and its application to centromere mapping

    Institute of Scientific and Technical Information of China (English)

    CHENGZHUKUAN; HENGXIUYU; 等

    1997-01-01

    The aneuploid with isochromosome or telochromosome is ideal material for exploring the position of centromere in lingkage map.For obtaining these aneuploids in rice,the primary trisomics from triplo-1 to triplo-12 and the aneuploids derived from a triploid of indica rice variety Zhongxiao 3037 were carefully investigated.From the offsprings of triplo-10,a primary trisomic of chromosome 10 of the variety,an isotetrasomic “triplo-10-1” was obtained.Cytological investigation revealed that a pair of extra isochromosomes of triplo-10-1 were come from the short arm of chromosome 10.In the offsprings of the isotetrasomic,a secondary trisomic “triplo-10-2”,in which the extra-chromosome was an isochromosome derived from the short arm of chromosome 10,was identified.With the isotetrasomic,secondary trisomic,primary trisomic and diploid of variety Zhongxiao 3037,different molecular markers were used for exploring the position of the centromere of chromosome 10.Based on the DNA dosage effect,it was verified that the molecular markers G1125,G333 and L169 were Located on the short arm,G1084 and other 16 available molecular markers were on the long arm of chromosome 10.So the centromere of chromosome 10 was located somewhere between G1125 and G1084 according to the RFLP linkage map given by Kurata et al[1].The distance from G1125 to G1084 was about 3.2cM.

  16. RNA Pol II promotes transcription of centromeric satellite DNA in beetles.

    Directory of Open Access Journals (Sweden)

    Zeljka Pezer

    Full Text Available Transcripts of centromeric satellite DNAs are known to play a role in heterochromatin formation as well as in establishment of the kinetochore. However, little is known about basic mechanisms of satellite DNA expression within constitutive heterochromatin and its regulation. Here we present comprehensive analysis of transcription of abundant centromeric satellite DNA, PRAT from beetle Palorus ratzeburgii (Coleoptera. This satellite is characterized by preservation and extreme sequence conservation among evolutionarily distant insect species. PRAT is expressed in all three developmental stages: larvae, pupae and adults at similar level. Transcripts are abundant comprising 0.033% of total RNA and are heterogeneous in size ranging from 0.5 kb up to more than 5 kb. Transcription proceeds from both strands but with 10 fold different expression intensity and transcripts are not processed into siRNAs. Most of the transcripts (80% are not polyadenylated and remain in the nucleus while a small portion is exported to the cytoplasm. Multiple, irregularly distributed transcription initiation sites as well as termination sites have been mapped within the PRAT sequence using primer extension and RLM-RACE. The presence of cap structure as well as poly(A tails in a portion of the transcripts indicate RNA polymerase II-dependent transcription and a putative polymerase II promoter site overlaps the most conserved part of the PRAT sequence. The treatment of larvae with alpha-amanitin decreases the level of PRAT transcripts at concentrations that selectively inhibit pol II activity. In conclusion, stable, RNA polymerase II dependant transcripts of abundant centromeric satellite DNA, not regulated by RNAi, have been identified and characterized. This study offers a basic understanding of expression of highly abundant heterochromatic DNA which in beetle species constitutes up to 50% of the genome.

  17. Novel Centromeric Loci of the Wine and Beer Yeast Dekkera bruxellensis CEN1 and CEN2

    Science.gov (United States)

    Ishchuk, Olena P.; Vojvoda Zeljko, Tanja; Schifferdecker, Anna J.; Mebrahtu Wisén, Sofia; Hagström, Åsa K.; Rozpędowska, Elżbieta; Rørdam Andersen, Mikael; Hellborg, Linda; Ling, Zhihao; Sibirny, Andrei A.

    2016-01-01

    The wine and beer yeast Dekkera bruxellensis thrives in environments that are harsh and limiting, especially in concentrations with low oxygen and high ethanol. Its different strains’ chromosomes greatly vary in number (karyotype). This study isolates two novel centromeric loci (CEN1 and CEN2), which support both the yeast’s autonomous replication and the stable maintenance of plasmids. In the sequenced genome of the D. bruxellensis strain CBS 2499, CEN1 and CEN2 are each present in one copy. They differ from the known “point” CEN elements, and their biological activity is retained within ~900–1300 bp DNA segments. CEN1 and CEN2 have features of both “point” and “regional” centromeres: They contain conserved DNA elements, ARSs, short repeats, one tRNA gene, and transposon-like elements within less than 1 kb. Our discovery of a miniature inverted-repeat transposable element (MITE) next to CEN2 is the first report of such transposons in yeast. The transformants carrying circular plasmids with cloned CEN1 and CEN2 undergo a phenotypic switch: They form fluffy colonies and produce three times more biofilm. The introduction of extra copies of CEN1 and CEN2 promotes both genome rearrangements and ploidy shifts, with these effects mediated by homologous recombination (between circular plasmid and genome centromere copy) or by chromosome breakage when integrated. Also, the proximity of the MITE-like transposon to CEN2 could translocate CEN2 within the genome or cause chromosomal breaks, so promoting genome dynamics. With extra copies of CEN1 and CEN2, the yeast’s enhanced capacities to rearrange its genome and to change its gene expression could increase its abilities for exploiting new and demanding niches. PMID:27560164

  18. Centromere binding specificity in assembly of the F plasmid partition complex

    OpenAIRE

    2011-01-01

    The segregation of plasmid F of Escherichia coli is highly reliable. The Sop partition locus, responsible for this stable maintenance, is composed of two genes, sopA and sopB and a centromere, sopC, consisting of 12 direct repeats of 43 bp. Each repeat carries a 16-bp inverted repeat motif to which SopB binds to form a nucleoprotein assembly called the partition complex. A database search for sequences closely related to sopC revealed unexpected features that appeared highly conserved. We hav...

  19. Domain architectures of the Scm3p protein provide insights into centromere function and evolution

    OpenAIRE

    Aravind, L.; Lakshminarayan, M. Iyer; Wu, Carl

    2007-01-01

    Recently, Scm3p has been shown to be a nonhistone component of centromeric chromatin that binds stoichiometrically to CenH3–H4 histones, and to be required for the assembly of kinetochores in S. cerevisiae. Scm3p is conserved across fungi, and displays a remarkable variation in protein size, ranging from ~200 amino acids in Saccharomyces cerevisiae to ~1300 amino acids in Neurospora crassa. This is primarily due a variable C-terminal segment that is linked to a conserved N-terminal, CenH3-int...

  20. A new model of sperm nuclear architecture following assessment of the organization of centromeres and telomeres in three-dimensions

    Science.gov (United States)

    Ioannou, Dimitrios; Millan, Nicole M.; Jordan, Elizabeth; Tempest, Helen G.

    2017-01-01

    The organization of chromosomes in sperm nuclei has been proposed to possess a unique “hairpin-loop” arrangement, which is hypothesized to aid in the ordered exodus of the paternal genome following fertilization. This study simultaneously assessed the 3D and 2D radial and longitudinal organization of telomeres, centromeres, and investigated whether chromosomes formed the same centromere clusters in sperm cells. Reproducible radial and longitudinal non-random organization was observed for all investigated loci using both 3D and 2D approaches in multiple subjects. We report novel findings, with telomeres and centromeres being localized throughout the nucleus but demonstrating roughly a 1:1 distribution in the nuclear periphery and the intermediate regions with exodus of paternal chromosomes following fertilization. PMID:28139771

  1. Organization and evolution of primate centromeric DNA from whole-genome shotgun sequence data.

    Directory of Open Access Journals (Sweden)

    Can Alkan

    2007-09-01

    Full Text Available The major DNA constituent of primate centromeres is alpha satellite DNA. As much as 2%-5% of sequence generated as part of primate genome sequencing projects consists of this material, which is fragmented or not assembled as part of published genome sequences due to its highly repetitive nature. Here, we develop computational methods to rapidly recover and categorize alpha-satellite sequences from previously uncharacterized whole-genome shotgun sequence data. We present an algorithm to computationally predict potential higher-order array structure based on paired-end sequence data and then experimentally validate its organization and distribution by experimental analyses. Using whole-genome shotgun data from the human, chimpanzee, and macaque genomes, we examine the phylogenetic relationship of these sequences and provide further support for a model for their evolution and mutation over the last 25 million years. Our results confirm fundamental differences in the dispersal and evolution of centromeric satellites in the Old World monkey and ape lineages of evolution.

  2. Point Mutations in Centromeric Histone Induce Post-zygotic Incompatibility and Uniparental Inheritance.

    Directory of Open Access Journals (Sweden)

    Sundaram Kuppu

    2015-09-01

    Full Text Available The centromeric histone 3 variant (CENH3, aka CENP-A is essential for the segregation of sister chromatids during mitosis and meiosis. To better define CENH3 functional constraints, we complemented a null allele in Arabidopsis with a variety of mutant alleles, each inducing a single amino acid change in conserved residues of the histone fold domain. Many of these transgenic missense lines displayed wild-type growth and fertility on self-pollination, but exhibited frequent post-zygotic death and uniparental inheritance when crossed with wild-type plants. The failure of centromeres marked by these missense mutation in the histone fold domain of CENH3 reproduces the genome elimination syndromes described with chimeric CENH3 and CENH3 from diverged species. Additionally, evidence that a single point mutation is sufficient to generate a haploid inducer provide a simple one-step method for the identification of non-transgenic haploid inducers in existing mutagenized collections of crop species. As proof of the extreme simplicity of this approach to create haploid-inducing lines, we performed an in silico search for previously identified point mutations in CENH3 and identified an Arabidopsis line carrying the A86V substitution within the histone fold domain. This A87V non-transgenic line, while fully fertile on self-pollination, produced postzygotic death and uniparental haploids when crossed to wild type.

  3. Characterization of the oncogenic function of centromere protein F in hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Yongdong; Liu, Lulu; Zeng, Tingting; Zhu, Ying-Hui [State Key Laboratory of Oncology in Southern China, Sun Yat-Sen University Cancer Center, Guangzhou (China); Li, Jiangchao [Vascular Biology Research Institute, Guangdong Pharmaceutical University, Guangzhou (China); Chen, Leilei [Department of Clinical Oncology, The University of Hong Kong, Pokfulam, Hong Kong (China); Li, Yan; Yuan, Yun-Fei [State Key Laboratory of Oncology in Southern China, Sun Yat-Sen University Cancer Center, Guangzhou (China); Ma, Stephanie, E-mail: stefma@hku.hk [Department of Clinical Oncology, The University of Hong Kong, Pokfulam, Hong Kong (China); State Key Laboratory for Liver Research, The University of Hong Kong, Pokfulam, Hong Kong (China); Guan, Xin-Yuan, E-mail: xyguan@hkucc.hku.hk [State Key Laboratory of Oncology in Southern China, Sun Yat-Sen University Cancer Center, Guangzhou (China); Department of Clinical Oncology, The University of Hong Kong, Pokfulam, Hong Kong (China); State Key Laboratory for Liver Research, The University of Hong Kong, Pokfulam, Hong Kong (China)

    2013-07-12

    Highlights: •Overexpression of CENPF is frequently detected in HCC. •Upregulation of CENPF serves as an independent prognosis factor in HCC patients. •CENPF functions as an oncogene in HCC by promoting cell G2/M transition. -- Abstract: Centromere protein F (CENPF) is an essential nuclear protein associated with the centromere-kinetochore complex and plays a critical role in chromosome segregation during mitosis. Up-regulation of CENPF expression has previously been detected in several solid tumors. In this study, we aim to study the expression and functional role of CENPF in hepatocellular carcinoma (HCC). We found CENPF was frequently overexpressed in HCC as compared with non-tumor tissue. Up-regulated CENPF expression in HCC was positively correlated with serum AFP, venous invasion, advanced differentiation stage and a shorter overall survival. Cox regression analysis found that overexpression of CENPF was an independent prognosis factor in HCC. Functional studies found that silencing CENPF could decrease the ability of the cells to proliferate, form colonies and induce tumor formation in nude mice. Silencing CENPF also resulted in the cell cycle arrest at G2/M checkpoint by down-regulating cell cycle proteins cdc2 and cyclin B1. Our data suggest that CENPF is frequently overexpressed in HCC and plays a critical role in driving HCC tumorigenesis.

  4. Identification of a high frequency of chromosomal rearrangements in the centromeric regions of prostate cancer patients

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The aim of the present investigation was to study the major chromosomal aberrations (CA) like deletion, translocation,inversion and mosaic in prostate cancer patients of Tamilnadu, Southern India. Totally 45 blood samples were collected from various hospitals in Tamilnadu, Southern India. Equal numbers of normal healthy subjects were chosen after signing a consent form. Volunteers provided blood samples (5 ml) to establish leukocyte cultures. Cytogenetic studies were performed by using Giemsa-banding technique and finally the results were ensured by spectral karyotyping (SKY) technique. In the present investigation, major CA like deletion, translocation, inversion and mosaic were identified in experimental subjects. Results showed frequent CA in chromosomes 1, 3, 5, 6, 7, 9, 13, 16, 18 and X. In comparison with experimental subjects, the control subjects exhibited very low levels of major CA (P<0.05). In the present study, the high frequency of centromeric rearrangements indicates a potential role for mitotic irregularities associated with the centromere in prostate cancer tumorigenesis. Identification of chromosome alterations may be helpful in understanding the molecular basis of the disease in better manner.

  5. High-affinity binding of Chp1 chromodomain to K9 methylated histone H3 is required to establish centromeric heterochromatin.

    Science.gov (United States)

    Schalch, Thomas; Job, Godwin; Noffsinger, Victoria J; Shanker, Sreenath; Kuscu, Canan; Joshua-Tor, Leemor; Partridge, Janet F

    2009-04-10

    In fission yeast, assembly of centromeric heterochromatin requires the RITS complex, which consists of Ago1, Tas3, Chp1, and siRNAs derived from centromeric repeats. Recruitment of RITS to centromeres has been proposed to depend on siRNA-dependent targeting of Ago1 to centromeric sequences. Previously, we demonstrated that methylated lysine 9 of histone H3 (H3K9me) acts upstream of siRNAs during heterochromatin establishment. Our crystal structure of Chp1's chromodomain in complex with a trimethylated lysine 9 H3 peptide reveals extensive sites of contact that contribute to Chp1's high-affinity binding. We found that this high-affinity binding is critical for the efficient establishment of centromeric heterochromatin, but preassembled heterochromatin can be maintained when Chp1's affinity for H3K9me is greatly reduced.

  6. Genome-wide mapping of cytosine methylation revealed dynamic DNA methylation patterns associated with genes and centromeres in rice.

    Science.gov (United States)

    Yan, Huihuang; Kikuchi, Shinji; Neumann, Pavel; Zhang, Wenli; Wu, Yufeng; Chen, Feng; Jiang, Jiming

    2010-08-01

    We conducted genome-wide mapping of cytosine methylation using methylcytosine immunoprecipitation combined with Illumina sequencing. The chromosomal distribution pattern of methylated DNA is similar to the heterochromatin distribution pattern on rice chromosomes. The DNA methylation patterns of rice genes are similar to those in Arabidopsis thaliana, including distinct methylation patterns asssociated with gene bodies and promoters. The DNA sequences in the core domains of rice Cen4, Cen5 and Cen8 showed elevated methylation levels compared with sequences in the pericentromeric regions. In addition, elevated methylation levels were associated with the DNA sequences in the CENH3-binding subdomains, compared with the sequences in the flanking H3 subdomains. In contrast, the centromeric domain of Cen11, which is composed exclusively of centromeric satellite DNA, is hypomethylated compared with the pericentromeric domains. Thus, the DNA sequences associated with functional centromeres can be either hypomethylated or hypermethylated. The methylation patterns of centromeric DNA appear to be correlated with the composition of the associated DNA sequences. We propose that both hypomethylation and hypermethylation of CENH3-associated DNA sequences can serve as epigenetic marks to distinguish where CENH3 deposition will occur within the surrounding H3 chromatin.

  7. Mutations in Z8T824 Are Associated with Immunodeficiency, Centromeric Instability, and Facial Anomalies Syndrome Type 2

    NARCIS (Netherlands)

    de Greef, Jessica C.; Wang, Jun; Balog, Judit; den Dunnen, Johan T.; Frants, Rune R.; Straasheijm, Kirsten R.; Aytekin, Caner; van der Burg, Mirjam; Duprez, Laurence; Ferster, Alina; Gennery, Andrew R.; Gimelli, Giorgio; Reisli, Ismail; Schuetz, Catharina; Schulz, Ansgar; Smeets, Dominique F. C. M.; Sznajer, Yves; Wijmenga, Cisca; van Eggermond, Maria C.; van Ostaijen-ten Dam, Monique M.; Lankester, Arjan C.; van Tol, Maarten J. D.; van den Elsen, Peter J.; Weemaes, Corry M.; van der Maarel, Silvere M.

    2011-01-01

    Autosomal-recessive immunodeficiency, centromeric instability, and facial anomalies (ICE) syndrome is mainly characterized by recurrent, often fatal, respiratory and gastrointestinal infections. About 50% of patients carry mutations in the DNA methyltransferase 3B gene (DNMT3B) (ICF1). The remaining

  8. Is the time dimension of the cell cycle re-entry in AD regulated by centromere cohesion dynamics?

    Science.gov (United States)

    Bajić, Vladan P; Spremo-Potparević, Biljana; Zivković, Lada; Djelić, Ninoslav; Smith, Mark A

    2008-01-01

    Chromosomal involvement is a legitimate, yet not well understood, feature of Alzheimer disease (AD). Firstly, AD affects more women than men. Secondly, the amyloid-β protein precursor genetic mutations, responsible for a cohort of familial AD cases, reside on chromosome 21, the same chromosome responsible for the developmental disorder Down's syndrome. Thirdly, lymphocytes from AD patients display a novel chromosomal phenotype, namely premature centromere separation (PCS). Other documented morphological phenomena associated with AD include the occurrence of micronuclei, aneuploidy, binucleation, telomere instability, and cell cycle re-entry protein expression. Based on these events, here we present a novel hypothesis that the time dimension of cell cycle re-entry in AD is highly regulated by centromere cohesion dynamics. In view of the fact that neurons can re-enter the cell division cycle, our hypothesis predicts that alterations in the signaling pathway leading to premature cell death in neurons is a consequence of altered regulation of the separation of centromeres as a function of time. It is well known that centromeres in the metaphase-anaphase transition separate in a non-random, sequential order. This sequence has been shown to be deregulated in aging cells, various tumors, syndromes of chromosome instability, following certain chemical inductions, as well as in AD. Over time, premature chromosome separation is both a result of, and a driving force behind, further cohesion impairment, activation of cyclin dependent kinases, and mitotic catastrophe, a vicious circle resulting in cellular degeneration and death.

  9. A DNA polymerase alpha accessory protein, Mcl1, is required for propagation of centromere structures in fission yeast.

    Directory of Open Access Journals (Sweden)

    Toyoaki Natsume

    Full Text Available Specialized chromatin exists at centromeres and must be precisely transmitted during DNA replication. The mechanisms involved in the propagation of these structures remain elusive. Fission yeast centromeres are composed of two chromatin domains: the central CENP-A(Cnp1 kinetochore domain and flanking heterochromatin domains. Here we show that fission yeast Mcl1, a DNA polymerase alpha (Pol alpha accessory protein, is critical for maintenance of centromeric chromatin. In a screen for mutants that alleviate both central domain and outer repeat silencing, we isolated several cos mutants, of which cos1 is allelic to mcl1. The mcl1-101 mutation causes reduced CENP-A(Cnp1 in the central domain and an aberrant increase in histone acetylation in both domains. These phenotypes are also observed in a mutant of swi7(+, which encodes a catalytic subunit of Pol alpha. Mcl1 forms S-phase-specific nuclear foci, which colocalize with those of PCNA and Pol alpha. These results suggest that Mcl1 and Pol alpha are required for propagation of centromere chromatin structures during DNA replication.

  10. Variation in Copy Number of Ty3/Gypsy Centromeric Retrotransposons in the Genomes of Thinopyrum intermedium and Its Diploid Progenitors.

    Directory of Open Access Journals (Sweden)

    Mikhail G Divashuk

    Full Text Available Speciation and allopolyploidization in cereals may be accompanied by dramatic changes in abundance of centromeric repeated transposable elements. Here we demonstrate that the reverse transcriptase part of Ty3/gypsy centromeric retrotransposon (RT-CR is highly conservative in the segmental hexaploid Thinopyrum intermedium (JrJvsSt and its possible diploid progenitors Th. bessarabicum (Jb, Pseudoroegneria spicata (St and Dasypyrum villosum (V but the abundance of the repeats varied to a large extent. Fluorescence in situ hybridization (FISH showed hybridization signals in centromeric region of all chromosomes in the studied species, although the intensity of the signals drastically differed. In Th. intermedium, the strongest signal of RT-CR probe was detected on the chromosomes of Jv, intermediate on Jr and faint on Js and St subgenome suggesting different abundance of RT-CR on the individual chromosomes rather than the sequence specificity of RT-CRs of the subgenomes. RT-CR quantification using real-time PCR revealed that its content per genome in Th. bessarabicum is ~ 2 times and P. spicata is ~ 1,5 times higher than in genome of D. villosum. The possible burst of Ty3/gypsy centromeric retrotransposon in Th. intermedium during allopolyploidization and its role in proper mitotic and meiotic chromosome behavior in a nascent allopolyploid is discussed.

  11. Microsatellite-centromere mapping in Japanese scallop ( Patinopecten yessoensis) through half-tetrad analysis in gynogenetic diploid families

    Science.gov (United States)

    Li, Qi; Qi, Mingjun; Nie, Hongtao; Kong, Lingfeng; Yu, Hong

    2016-06-01

    Gene-centromere mapping is an essential prerequisite for understanding the composition and structure of genomes. Half-tetrad analysis is a powerful tool for mapping genes and understanding chromosomal behavior during meiosis. The Japanese scallop ( Patinopecten yessoensis), a cold-tolerant species inhabiting the northwestern Pacific coast, is a commercially important marine bivalve in Asian countries. In this study, inheritance of 32 informative microsatellite loci was examined in 70-h D-shaped larvae of three induced meiogynogenetic diploid families of P. yessoensis for centromere mapping using half-tetrad analysis. The ratio of gynogenetic diploids was proven to be 100%, 100% and 96% in the three families, respectively. Inheritance analysis in the control crosses showed that 51 of the 53 genotypic ratios observed were in accordance with Mendelian expectations at the 5% level after Bonferroni correction. Seven of the 32 microsatellite loci showed the existence of null alleles in control crosses. The second division segregation frequency ( y) of the microsatellite loci ranged from 0.07 to 0.85 with a mean of 0.38, suggesting the existence of positive interference after a single chiasma formation in some chromosomes in the scallop. Microsatellite-centromere distances ranged from 4 cM to 42 cM under the assumption of complete interference. Information on the positions of centromeres in relation to the microsatellite loci will represent a contribution towards the assembly of genetic maps in the commercially important scallop species.

  12. Effects of sense and antisense centromere/kinetochore complex protein-B (CENP-B) in cell cycle regulation

    Institute of Scientific and Technical Information of China (English)

    LUO Song; LIN Haiyan; QI Jianguo; WANG Yongchao

    2005-01-01

    This paper investigates the effects of sense and antisense centromere/kinetochore complex protein-B (CENP-B) in cell cycle regulation. Full-length cenpb cDNA was subcloned into pBI-EGFP eukaryotic expression vector in both sense and antisense orientation. HeLa-Tet-Off cells were transfected with sense or antisense cenpb vectors. Sense transfection of HeLa-Tet-Off cells resulted in the formation of a large centromere/kinetochore complex, and apoptosis of cells following several times of cell division. A stable antisense cenpb transfected cell line, named HACPB, was obtained. The centromere/kinetochore complex of HACPB cells became smaller than control HeLa-Tet-Off cells and scattered, and the expression of CENP-B was down-regulated. In addition, delayed cell cycle progression, inhibited malignant phenotype, restrained ability of tumor formation in nude mice, and delayed entry from G2/M phase into next G1 phase were observed in HACPB cells. Furthermore, the expression of cyclin-dependent kinases (CDKs), cyclins, and CDK inhibitors (CKIs) were modulated during different phases of the cell cycle. CENP-B is an essential protein for the maintenance of the structure and function of centromere/kinetochore complex, and plays important roles in cell cycle regulation.

  13. Cis-acting determinants affecting centromere function, sister-chromatid cohesion and reciprocal recombination during meiosis in Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Sears, D.D.; Hieter, P. [Johns Hopkins Univ. School of Medicine, Baltimore, MD (United States); Shero, J.H. [Johns Hopkins Univ. School of Medicine, Baltimore, MD (United States)]|[Baylor College of Medicine, Houston, TX (United States); Hegemann, J.H. [Justus Liebig Universitaet, Giessen (Germany)

    1995-03-01

    We have employed a system that utilizes homologous pairs of human DNA-derived yeast artificial chromosomes (YACs) as marker chromosomes to assess the specific role(s) of conserved centromere DNA elements (CDEI, CDEII, and CDEIII) in meiotic chromosome disjunction fidelity. Thirteen different centromere (CEN) mutations were tested for their effects on meiotic centromere function. YACs containing a wild-type CEN DNA sequence segregate with high fidelity in meiosis I (99% normal segregation) and in meiosis II (96% normal segregation). YACs containing a 31-bp deletion mutation in centromere DNA element II (CDEII{Delta}31) in either a heterocentric (mutant/wild type), homocentric (mutant/mutant) or monosomic (mutant/-) YAC pair configuration exhibited high levels (16-28%) of precocious sister-chromatid segregation (PSS) and increased levels (1-6%) of nondisjunction meiosis I (NDI). YACs containing this mutation also exhibit high levels (21%) of meiosis II nondisjunction. Interestingly, significant alterations in homolog recombination frequency were observed in the exceptional PSS class of tetrads, suggesting unusual interactions between prematurely separated sister chromatids and their homologous nonsister chromatids. We also have assessed the meiotic segregation effects of rare gene conversion events occurring at sites located immediately adjacent to or distantly from the centromere region. Proximal gene conversion events were associated with extremely high levels (60%) of meiosis I segregation errors (including both PSS and NDI), whereas distal events had no apparent effect. Taken together, our results indicate a critical role for CDEII in meiosis and underscore the importance of maintaining sister-chromatid cohesion for proper recombination in meiotic prophase and for proper disjunction in meiosis I. 49 refs., 4 figs., 5 tabs.

  14. Centromere pairing by a plasmid-encoded type I ParB protein

    DEFF Research Database (Denmark)

    Ringgaard, Simon; Löwe, Jan; Gerdes, Kenn

    2007-01-01

    over the nucleoid. ParB ribbon-helix-helix dimers bind cooperatively to direct repeats in parC1 and parC2. Using four different assays we obtain solid evidence that ParB can pair parC1- and parC2-encoding DNA fragments in vitro. Convincingly, electron microscopy revealed that ParB mediates binary...... pairing of parC fragments. In addition to binary complexes, ParB mediated the formation of higher order complexes consisting of several DNA fragments joined by ParB at centromere site parC. N-terminal truncated versions of ParB still possessing specific DNA binding activity were incompetent in pairing...

  15. Isolation and molecular characterization of a highly polymorphic centromeric tandem repeat in the family Falconidae.

    Science.gov (United States)

    Longmire, J L; Lewis, A K; Brown, N C; Buckingham, J M; Clark, L M; Jones, M D; Meincke, L J; Meyne, J; Ratliff, R L; Ray, F A

    1988-01-01

    An abundant tandem repeat has been cloned from genomic DNA of the merlin (Falco columbarius). The cloned sequence is 174 bp in length, and maps by in situ hybridization to the centromeric regions of several of the large chromosomes within the merlin karyotype. Complementary sequences have been identified within a variety of falcon species; these sequences are either absent or in very low copy number in the family Accipitridae. The cloned merlin repeat reveals highly polymorphic restriction patterns in the peregrine falcon (Falco peregrinus). These polymorphisms, which have been shown to be stably inherited within a family of captive peregrines, can be used to differentiate the Greenland and Argentina populations of this endangered raptor species.

  16. A hemicentric inversion in the maize line knobless Tama flint created two sites of centromeric elements and moved the kinetochore-forming region.

    Science.gov (United States)

    Lamb, Jonathan C; Meyer, Julie M; Birchler, James A

    2007-06-01

    A maize line, knobless Tama flint (KTF), was found to contain a version of chromosome 8 with two spatially distinct regions of centromeric elements, one at the original genetic position and the other at a novel location on the long arm. The new site of centromeric elements functions as the kinetochore-forming region resulting in a change of arm length ratio. Examination of fluorescence in situ hybridization markers on chromosome 8 revealed an inversion between the two centromere sites relative to standard maize lines, indicating that this chromosome 8 resulted from a hemicentric inversion with one breakpoint approximately 20 centi-McClintocks (cMc) on the long arm (20% of the total arm length from the centromere) and the other in the original cluster of centromere repeats. This inversion moved the kinetochore-forming region but left the remainder of the centromere repeats. In a hybrid between a standard line (Mo17) and KTF, both chromosome 8 homologues were completely synapsed at pachytene despite the inversion. Although the homologous centromeres were not paired, they were always correctly oriented at anaphase and migrated to opposite poles. Additionally, recombination on 8L was severely repressed in the hybrid.

  17. DNA Topoisomerase II Is a Determinant of the Tensile Properties of Yeast Centromeric Chromatin and the Tension Checkpoint

    OpenAIRE

    Warsi, Tariq H.; Navarro, Michelle S.; Bachant, Jeff

    2008-01-01

    Centromeric (CEN) chromatin is placed under mechanical tension and stretches as kinetochores biorient on the mitotic spindle. This deformation could conceivably provide a readout of biorientation to error correction mechanisms that monitor kinetochore–spindle interactions, but whether CEN chromatin acts in a tensiometer capacity is unresolved. Here, we report observations linking yeast Topoisomerase II (Top2) to both CEN mechanics and assessment of interkinetochore tension. First, in top2-4 a...

  18. Maximum-likelihood method identifies meiotic restitution mechanism from heterozygosity transmission of centromeric loci: application in citrus

    Science.gov (United States)

    Cuenca, José; Aleza, Pablo; Juárez, José; García-Lor, Andrés; Froelicher, Yann; Navarro, Luis; Ollitrault, Patrick

    2015-01-01

    Polyploidisation is a key source of diversification and speciation in plants. Most researchers consider sexual polyploidisation leading to unreduced gamete as its main origin. Unreduced gametes are useful in several crop breeding schemes. Their formation mechanism, i.e., First-Division Restitution (FDR) or Second-Division Restitution (SDR), greatly impacts the gametic and population structures and, therefore, the breeding efficiency. Previous methods to identify the underlying mechanism required the analysis of a large set of markers over large progeny. This work develops a new maximum-likelihood method to identify the unreduced gamete formation mechanism both at the population and individual levels using independent centromeric markers. Knowledge of marker-centromere distances greatly improves the statistical power of the comparison between the SDR and FDR hypotheses. Simulating data demonstrated the importance of selecting markers very close to the centromere to obtain significant conclusions at individual level. This new method was used to identify the meiotic restitution mechanism in nineteen mandarin genotypes used as female parents in triploid citrus breeding. SDR was identified for 85.3% of 543 triploid hybrids and FDR for 0.6%. No significant conclusions were obtained for 14.1% of the hybrids. At population level SDR was the predominant mechanisms for the 19 parental mandarins. PMID:25894579

  19. Maximum-likelihood method identifies meiotic restitution mechanism from heterozygosity transmission of centromeric loci: application in citrus.

    Science.gov (United States)

    Cuenca, José; Aleza, Pablo; Juárez, José; García-Lor, Andrés; Froelicher, Yann; Navarro, Luis; Ollitrault, Patrick

    2015-04-20

    Polyploidisation is a key source of diversification and speciation in plants. Most researchers consider sexual polyploidisation leading to unreduced gamete as its main origin. Unreduced gametes are useful in several crop breeding schemes. Their formation mechanism, i.e., First-Division Restitution (FDR) or Second-Division Restitution (SDR), greatly impacts the gametic and population structures and, therefore, the breeding efficiency. Previous methods to identify the underlying mechanism required the analysis of a large set of markers over large progeny. This work develops a new maximum-likelihood method to identify the unreduced gamete formation mechanism both at the population and individual levels using independent centromeric markers. Knowledge of marker-centromere distances greatly improves the statistical power of the comparison between the SDR and FDR hypotheses. Simulating data demonstrated the importance of selecting markers very close to the centromere to obtain significant conclusions at individual level. This new method was used to identify the meiotic restitution mechanism in nineteen mandarin genotypes used as female parents in triploid citrus breeding. SDR was identified for 85.3% of 543 triploid hybrids and FDR for 0.6%. No significant conclusions were obtained for 14.1% of the hybrids. At population level SDR was the predominant mechanisms for the 19 parental mandarins.

  20. Pregnancy-associated thrombotic thrombocytopenic purpura with anti-centromere antibody-positive Raynaud's syndrome.

    Science.gov (United States)

    Watanabe, Ryu; Shirai, Tsuyoshi; Tajima, Yumi; Ohguchi, Hiroto; Onishi, Yasushi; Fujii, Hiroshi; Takasawa, Naruhiko; Ishii, Tomonori; Harigae, Hideo

    2010-01-01

    Thrombotic thrombocytopenic purpura (TTP), scleroderma renal crisis (SRC), and hemolysis, elevated liver enzyme levels, and a low platelet count (HELLP) syndrome display common symptoms that include microangiopathic hemolytic anemia, thrombocytopenia, and renal failure. Therefore, it is important to distinguish between them because their treatments vary: however, the differential diagnosis is sometimes difficult. We report a 32-year-old woman who was referred to our department for further examination of microangiopathic hemolytic anemia, thrombocytopenia, and a slightly elevated serum creatinine level with anti-centromere antibody-positive Raynaud's syndrome in the early puerperal period. TTP, SRC, and HELLP syndrome were considered in the differential diagnosis, but the measurement of a disintegrin-like metalloprotease with thrombospondin type 1 motifs 13 (ADAMTS 13) activity and its inhibitor level led to the diagnosis of TTP. She was successfully treated by plasma exchange and high-dose prednisolone and angiotensin-converting enzyme inhibitor. If microangiopathic hemolytic anemia and thrombocytopenia are observed in perinatal women or patients with signs of systemic sclerosis, the measurement of ADAMTS13 activity and its inhibitor level are essential for diagnosis and therapeutic choice.

  1. Expression and prognostic relevance of centromere protein A in primary osteosarcoma.

    Science.gov (United States)

    Gu, Xiao-Min; Fu, Jie; Feng, Xiao-Jun; Huang, Xue; Wang, Shou-Mei; Chen, Xin-Feng; Zhu, Ming-Hua; Zhang, Shu-Hui

    2014-04-01

    Centromere protein A (CENP-A) is one of the fundamental components of the human active kinetochore and plays important roles in cell-cycle regulation, cell survival, and genetic stability. The aim of the present study was to explore the expression and prognostic significance of CENP-A in osteosarcoma. The results of real-time quantitative PCR and Western blotting analysis revealed an enhanced expression of CENP-A in osteosarcomas relative to adjacent non-tumorous bone tissues at both mRNA and protein levels. Immunohistochemically, 72 of the 123 osteosarcoma specimens (58.5%) had high expression of CENP-A. CENP-A overexpression was significantly correlated with tumor size (P=0.002), poor response to neoadjuvant chemotherapy (P=0.016), local recurrence/lung metastasis (P=0.001), high Ki-67 index (P=0.004), and P53 positivity (P=0.005). Median overall and recurrence-free survival time was significantly shorter in patients with high-CENP-A osteosarcomas than in those with low-CENP-A osteosarcomas. Multivariate analysis identified CENP-A as an independent poor prognostic factor for osteosarcoma. In conclusion, our results demonstrate that elevated CENP-A expression is significantly associated with osteosarcoma progression and has an independent prognostic value in predicting overall and recurrence-free survival for patients with osteosarcoma.

  2. The coordination of centromere replication, spindle formation, and kinetochore-microtubule interaction in budding yeast.

    Directory of Open Access Journals (Sweden)

    Hong Liu

    2008-11-01

    Full Text Available The kinetochore is a protein complex that assembles on centromeric DNA to mediate chromosome-microtubule interaction. Most eukaryotic cells form the spindle and establish kinetochore-microtubule interaction during mitosis, but budding yeast cells finish these processes in S-phase. It has long been noticed that the S-phase spindle in budding yeast is shorter than that in metaphase, but the biological significance of this short S-phase spindle structure remains unclear. We addressed this issue by using ask1-3, a temperature-sensitive kinetochore mutant that exhibits partially elongated spindles at permissive temperature in the presence of hydroxyurea (HU, a DNA synthesis inhibitor. After exposure to and removal of HU, ask1-3 cells show a delayed anaphase entry. This delay depends on the spindle checkpoint, which monitors kinetochore-microtubule interaction defects. Overproduction of microtubule-associated protein Ase1 or Cin8 also induces spindle elongation in HU-arrested cells. The spindle checkpoint-dependent anaphase entry delay is also observed after ASE1 or CIN8 overexpression in HU-arrested cells. Therefore, the shorter spindle in S-phase cells is likely to facilitate proper chromosome-microtubule interaction.

  3. Overexpression of centromere protein H is significantly associated with breast cancer progression and overall patient survival

    Institute of Scientific and Technical Information of China (English)

    Wen-Ting Liao; Yan Feng; Men-Lin Li; Guang-Lin Liu; Man-Zhi Li; Mu-Sheng Zeng; Li-Bing Song

    2011-01-01

    Breast cancer is one of the leading causes of cancer death worldwide.This study aimed to analyze the expression of centromere protein H (CENP-H) in breast cancer and to correlate it with clinicopathologic data,including patient survival.Using reverse transcription-polymerase chain reaction and Westem blotting to detect the expression of CENP-H in normal mammary epithelial cells,immortalized mammary epithelial cell lines,and breast cancer cell lines,we observed that the mRNA and protein levels of CENP-H were higher in breast cancer cell lines and in immortalized mammary epithelial cells than in normal mammary epithelial cells.We next examined CENP-H expression in 307 paraffin-embedded archived samples of clinicopathologically characterized breast cancer using immunohistochemistry,and detected high CENP-H expression in 134 (43.6%) samples.Statistical analysis showed that CENP-H expression was related with clinical stage (P = 0.001),T classification (P = 0.032),N classification (P =0.018),and Ki-67 (P<0.001).Patients with high CENP-H expression had short overall survival.Multivariate analysis showed that CENP-H expression was an independent prognostic indicator for patient survival.Our results suggest that CENP-H protein is a valuable marker of breast cancer progression and prognosis.

  4. The activity regulation of the mitotic centromere-associated kinesin by Polo-like kinase 1.

    Science.gov (United States)

    Ritter, Andreas; Sanhaji, Mourad; Steinhäuser, Kerstin; Roth, Susanne; Louwen, Frank; Yuan, Juping

    2015-03-30

    The mitotic centromere-associated kinesin (MCAK), a potent microtubule depolymerase, is involved in regulating microtubule dynamics. The activity and subcellular localization of MCAK are tightly regulated by key mitotic kinases, such as Polo-like kinase 1 (Plk1) by phosphorylating multiple residues in MCAK. Since Plk1 phosphorylates very often different residues of substrates at different stages, we have dissected individual phosphorylation of MCAK by Plk1 and characterized its function in more depth. We have recently shown that S621 in MCAK is the major phosphorylation site of Plk1, which is responsible for regulating MCAK's degradation by promoting the association of MCAK with APC/CCdc20. In the present study, we have addressed another two residues phosphorylated by Plk1, namely S632/S633 in the C-terminus of MCAK. Our data suggest that Plk1 phosphorylates S632/S633 and regulates its catalytic activity in mitosis. This phosphorylation is required for proper spindle assembly during early phases of mitosis. The subsequent dephosphorylation of S632/S633 might be necessary to timely align the chromosomes onto the metaphase plate. Therefore, our studies suggest new mechanisms by which Plk1 regulates MCAK: the degradation of MCAK is controlled by Plk1 phosphorylation on S621, whereas its activity is modulated by Plk1 phosphorylation on S632/S633 in mitosis.

  5. New tool for biological dosimetry: Reevaluation and automation of the gold standard method following telomere and centromere staining

    Energy Technology Data Exchange (ETDEWEB)

    M’kacher, Radhia [Laboratoire de Radiobiologie et Oncologie (LRO), Commissariat à l’Energie Atomique (CEA), Route du Panorama, 92265 Fontenay-aux-Roses (France); Maalouf, Elie E.L. [Laboratoire de Radiobiologie et Oncologie (LRO), Commissariat à l’Energie Atomique (CEA), Route du Panorama, 92265 Fontenay-aux-Roses (France); Laboratoire MIPS – Groupe TIIM3D, Université de Haute-Alsace, F-68093 Mulhouse (France); Ricoul, Michelle [Laboratoire de Radiobiologie et Oncologie (LRO), Commissariat à l’Energie Atomique (CEA), Route du Panorama, 92265 Fontenay-aux-Roses (France); Heidingsfelder, Leonhard [MetaSystems GmbH, Robert-Bosch-Str. 6, 68804 Altlussheim (Germany); Laplagne, Eric [Pole Concept, 61 Rue Erlanger, 75016 Paris (France); Cuceu, Corina; Hempel, William M. [Laboratoire de Radiobiologie et Oncologie (LRO), Commissariat à l’Energie Atomique (CEA), Route du Panorama, 92265 Fontenay-aux-Roses (France); Colicchio, Bruno; Dieterlen, Alain [Laboratoire MIPS – Groupe TIIM3D, Université de Haute-Alsace, F-68093 Mulhouse (France); Sabatier, Laure, E-mail: laure.sabatier@cea.fr [Laboratoire de Radiobiologie et Oncologie (LRO), Commissariat à l’Energie Atomique (CEA), Route du Panorama, 92265 Fontenay-aux-Roses (France)

    2014-12-15

    Graphical abstract: - Highlights: • We have applied telomere and centromere (TC) staining to the scoring of dicentrics. • TC staining renders the scoring of dicentrics more rapid and robust. • TC staining allows the scoring of not only dicentrics but all chromosomal anomalies. • TC staining has led to a reevaluation of the radiation dose–response curve. • TC staining allows automation of the scoring of chromosomal aberations. • Automated scoring of dicentrics after TC staining was as efficient as manual scoring. - Abstract: Purpose: The dicentric chromosome (dicentric) assay is the international gold-standard method for biological dosimetry and classification of genotoxic agents. The introduction of telomere and centromere (TC) staining offers the potential to render dicentric scoring more efficient and robust. In this study, we improved the detection of dicentrics and all unstable chromosomal aberrations (CA) leading to a significant reevaluation of the dose–effect curve and developed an automated approach following TC staining. Material and methods: Blood samples from 16 healthy donors were exposed to {sup 137}Cs at 8 doses from 0.1 to 6 Gy. CA were manually and automatically scored following uniform (Giemsa) or TC staining. The detection of centromeric regions and telomeric sequences using PNA probes allowed the detection of all unstable CA: dicentrics, centric and acentric rings, and all acentric fragments (with 2, 4 or no telomeres) leading to the precise quantification of estimated double strand breaks (DSB). Results: Manual scoring following TC staining revealed a significantly higher frequency of dicentrics (p < 10{sup −3}) (up to 30%) and estimated DSB (p < 10{sup −4}) compared to uniform staining due to improved detection of dicentrics with centromeres juxtaposed with other centromeres or telomeres. This improvement permitted the development of the software, TCScore, that detected 95% of manually scored dicentrics compared to 50% for

  6. Different patterns of evolution in the centromeric and telomeric regions of group A and B haplotypes of the human killer cell Ig-like receptor locus.

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    Chul-Woo Pyo

    Full Text Available The fast evolving human KIR gene family encodes variable lymphocyte receptors specific for polymorphic HLA class I determinants. Nucleotide sequences for 24 representative human KIR haplotypes were determined. With three previously defined haplotypes, this gave a set of 12 group A and 15 group B haplotypes for assessment of KIR variation. The seven gene-content haplotypes are all combinations of four centromeric and two telomeric motifs. 2DL5, 2DS5 and 2DS3 can be present in centromeric and telomeric locations. With one exception, haplotypes having identical gene content differed in their combinations of KIR alleles. Sequence diversity varied between haplotype groups and between centromeric and telomeric halves of the KIR locus. The most variable A haplotype genes are in the telomeric half, whereas the most variable genes characterizing B haplotypes are in the centromeric half. Of the highly polymorphic genes, only the 3DL3 framework gene exhibits a similar diversity when carried by A and B haplotypes. Phylogenetic analysis and divergence time estimates, point to the centromeric gene-content motifs that distinguish A and B haplotypes having emerged ~6 million years ago, contemporaneously with the separation of human and chimpanzee ancestors. In contrast, the telomeric motifs that distinguish A and B haplotypes emerged more recently, ~1.7 million years ago, before the emergence of Homo sapiens. Thus the centromeric and telomeric motifs that typify A and B haplotypes have likely been present throughout human evolution. The results suggest the common ancestor of A and B haplotypes combined a B-like centromeric region with an A-like telomeric region.

  7. A CENP-S/X complex assembles at the centromere in S and G2 phases of the human cell cycle.

    Science.gov (United States)

    Dornblut, Carsten; Quinn, Nadine; Monajambashi, Shamci; Prendergast, Lisa; van Vuuren, Chelly; Münch, Sandra; Deng, Wen; Leonhardt, Heinrich; Cardoso, M Cristina; Hoischen, Christian; Diekmann, Stephan; Sullivan, Kevin F

    2014-02-12

    The functional identity of centromeres arises from a set of specific nucleoprotein particle subunits of the centromeric chromatin fibre. These include CENP-A and histone H3 nucleosomes and a novel nucleosome-like complex of CENPs -T, -W, -S and -X. Fluorescence cross-correlation spectroscopy and Förster resonance energy transfer (FRET) revealed that human CENP-S and -X exist principally in complex in soluble form and retain proximity when assembled at centromeres. Conditional labelling experiments show that they both assemble de novo during S phase and G2, increasing approximately three- to fourfold in abundance at centromeres. Fluorescence recovery after photobleaching (FRAP) measurements documented steady-state exchange between soluble and assembled pools, with CENP-X exchanging approximately 10 times faster than CENP-S (t1/2 ∼ 10 min versus 120 min). CENP-S binding to sites of DNA damage was quite distinct, with a FRAP half-time of approximately 160 s. Fluorescent two-hybrid analysis identified CENP-T as a uniquely strong CENP-S binding protein and this association was confirmed by FRET, revealing a centromere-bound complex containing CENP-S, CENP-X and CENP-T in proximity to histone H3 but not CENP-A. We propose that deposition of the CENP-T/W/S/X particle reveals a kinetochore-specific chromatin assembly pathway that functions to switch centromeric chromatin to a mitosis-competent state after DNA replication. Centromeres shuttle between CENP-A-rich, replication-competent and H3-CENP-T/W/S/X-rich mitosis-competent compositions in the cell cycle.

  8. Heterochromatic genes undergo epigenetic changes and escape silencing in immunodeficiency, centromeric instability, facial anomalies (ICF syndrome.

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    Marie-Elisabeth Brun

    Full Text Available Immunodeficiency, Centromeric Instability, Facial Anomalies (ICF syndrome is a rare autosomal recessive disorder that is characterized by a marked immunodeficiency, severe hypomethylation of the classical satellites 2 and 3 associated with disruption of constitutive heterochromatin, and facial anomalies. Sixty percent of ICF patients have mutations in the DNMT3B (DNA methyltransferase 3B gene, encoding a de novo DNA methyltransferase. In the present study, we have shown that, in ICF lymphoblasts and peripheral blood, juxtacentromeric heterochromatic genes undergo dramatic changes in DNA methylation, indicating that they are bona fide targets of the DNMT3B protein. DNA methylation in heterochromatic genes dropped from about 80% in normal cells to approximately 30% in ICF cells. Hypomethylation was observed in five ICF patients and was associated with activation of these silent genes. Although DNA hypomethylation occurred in all the analyzed heterochromatic genes and in all the ICF patients, gene expression was restricted to some genes, every patient having his own group of activated genes. Histone modifications were preserved in ICF patients. Heterochromatic genes were associated with histone modifications that are typical of inactive chromatin: they had low acetylation on H3 and H4 histones and were slightly enriched in H3K9Me(3, both in ICF and controls. This was also the case for those heterochromatic genes that escaped silencing. This finding suggests that gene activation was not generalized to all the cells, but rather was restricted to a clonal cell population that may contribute to the phenotypic variability observed in ICF syndrome. A slight increase in H3K27 monomethylation was observed both in heterochromatin and active euchromatin in ICF patients; however, no correlation between this modification and activation of heterochromatic genes was found.

  9. Chromosome 17 centromere duplication and responsiveness to anthracycline-based neoadjuvant chemotherapy in breast cancer.

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    Tibau, Ariadna; López-Vilaró, Laura; Pérez-Olabarria, Maitane; Vázquez, Tania; Pons, Cristina; Gich, Ignasi; Alonso, Carmen; Ojeda, Belén; Ramón y Cajal, Teresa; Lerma, Enrique; Barnadas, Agustí; Escuin, Daniel

    2014-10-01

    Human epidermal growth factor receptor 2 (HER2) and topoisomerase II alpha (TOP2A) genes have been proposed as predictive biomarkers of sensitivity to anthracycline chemotherapy. Recently, chromosome 17 centromere enumeration probe (CEP17) duplication has also been associated with increased responsiveness to anthracyclines. However, reports are conflicting and none of these tumor markers can yet be considered a clinically reliable predictor of response to anthracyclines. We studied the association of TOP2A gene alterations, HER2 gene amplification, and CEP17 duplication with response to anthracycline-based neoadjuvant chemotherapy in 140 patients with operable or locally advanced breast cancer. HER2 was tested by fluorescence in situ hybridization and TOP2A and CEP17 by chromogenic in situ hybridization. Thirteen patients (9.3%) achieved pathologic complete response (pCR). HER2 amplification was present in 24 (17.5%) of the tumors. TOP2A amplification occurred in seven tumors (5.1%). CEP17 duplication was detected in 13 patients (9.5%). CEP17 duplication correlated with a higher rate of pCR [odds ratio (OR) 6.55, 95% confidence interval (95% CI) 1.25-34.29, P = .026], and analysis of TOP2A amplification showed a trend bordering on statistical significance (OR 6.97, 95% CI 0.96-50.12, P = .054). TOP2A amplification and CEP17 duplication combined were strongly associated with pCR (OR 6.71, 95% CI 1.66-27.01, P = .007). HER2 amplification did not correlate with pCR. Our results suggest that CEP17 duplication predicts pCR to primary anthracycline-based chemotherapy. CEP17 duplication, TOP2A amplifications, and HER2 amplifications were not associated with prognosis.

  10. 植物着丝粒结构及进化的研究进展%Research Progress on Structure and Evolution of Plant Centromeres

    Institute of Scientific and Technical Information of China (English)

    刘青

    2015-01-01

    植物着丝粒是染色体重要结构域,介导动粒装配。不同物种间着丝粒重复序列快速趋异进化,着丝粒功能保守,确保有丝分裂和减数分裂过程中染色体正确分离和准确传递。伴随染色质免疫共沉淀技术(Chromatin immunoprecipitation, ChIP)、ChIP与高密度芯片相结合技术(ChIP-chip)、ChIP与高通量测序相结合技术(ChIP-seq)的应用,植物着丝粒研究获得里程碑式进展:某些模式植物着丝粒DNA序列、蛋白质结构、功能获得大量新认识;着丝粒基本蛋白质组蛋白H3被用来界定着丝粒大小和边界;某些非着丝粒区域被激活为新着丝粒,在世代传递中保持稳定性。本文对植物着丝粒结构、功能、进化研究进行了综述,并探讨了植物着丝粒研究存在的问题。%The plant centromere is the most important chromosome domain mediating the assembly of kinetochore. The rapid divergent evolution of centromeric repeat sequences and function conservation of centromeres among different species ensure correct segregation and faithful transmission of chromosome in mitosis and meiosis. Along with the development of chromatin immunoprecipitation (ChIP), ChIP-chip, and ChIP-sequencing (ChIP-seq) technologies, three milestone discoveries have achieved in plant centromere research since the last 20 years, such as a lot of new knowledge on the structure, function, and evolution of centromeres from model plants, the fundamental kinetochore protein CENH3 used to delimiting the size and boundaries of centromere, the neocentromeres activated from non-centromeric regions stably transmitted to subsequent generations. The research progress on structure, function, and evolution of plant centromeres are reviewed and the remaining questions of plant centromere studies are discussed.

  11. Molecular and evolutionary characteristics of the fraction of human alpha satellite DNA associated with CENP-A at the centromeres of chromosomes 1, 5, 19, and 21

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    Roizès Gérard

    2010-03-01

    Full Text Available Abstract Background The mode of evolution of the highly homogeneous Higher-Order-Repeat-containing alpha satellite arrays is still subject to discussion. This is also true of the CENP-A associated repeats where the centromere is formed. Results In this paper, we show that the molecular mechanisms by which these arrays evolve are identical in multiple chromosomes: i accumulation of crossovers that homogenise and expand the arrays into different domains and subdomains that are mostly unshared between homologues and ii sporadic mutations and conversion events that simultaneously differentiate them from one another. Individual arrays are affected by these mechanisms to different extents that presumably increase with time. Repeats associated with CENP-A, where the centromere is formed, are subjected to the same evolutionary mechanisms, but constitute minor subsets that exhibit subtle sequence differences from those of the bulk repeats. While the DNA sequence per se is not essential for centromere localisation along an array, it appears that certain sequences can be selected against. On chromosomes 1 and 19, which are more affected by the above evolutionary mechanisms than are chromosomes 21 and 5, CENP-A associated repeats were also recovered from a second homogeneous array present on each chromosome. This could be a way for chromosomes to sustain mitosis and meiosis when the normal centromere locus is ineluctably undermined by the above mechanisms. Conclusion We discuss, in light of these observations, possible scenarios for the normal evolutionary fates of human centromeric regions.

  12. Differential repetitive DNA composition in the centromeric region of chromosomes of Amazonian lizard species in the family Teiidae.

    Science.gov (United States)

    Carvalho, Natalia D M; Carmo, Edson; Neves, Rogerio O; Schneider, Carlos Henrique; Gross, Maria Claudia

    2016-01-01

    Differences in heterochromatin distribution patterns and its composition were observed in Amazonian teiid species. Studies have shown repetitive DNA harbors heterochromatic blocks which are located in centromeric and telomeric regions in Ameiva ameiva (Linnaeus, 1758), Kentropyx calcarata (Spix, 1825), Kentropyx pelviceps (Cope, 1868), and Tupinambis teguixin (Linnaeus, 1758). In Cnemidophorus sp.1, repetitive DNA has multiple signals along all chromosomes. The aim of this study was to characterize moderately and highly repetitive DNA sequences by C ot1-DNA from Ameiva ameiva and Cnemidophorus sp.1 genomes through cloning and DNA sequencing, as well as mapping them chromosomally to better understand its organization and genome dynamics. The results of sequencing of DNA libraries obtained by C ot1-DNA showed that different microsatellites, transposons, retrotransposons, and some gene families also comprise the fraction of repetitive DNA in the teiid species. FISH using C ot1-DNA probes isolated from both Ameiva ameiva and Cnemidophorus sp.1 showed these sequences mainly located in heterochromatic centromeric, and telomeric regions in Ameiva ameiva, Kentropyx calcarata, Kentropyx pelviceps, and Tupinambis teguixin chromosomes, indicating they play structural and functional roles in the genome of these species. In Cnemidophorus sp.1, C ot1-DNA probe isolated from Ameiva ameiva had multiple interstitial signals on chromosomes, whereas mapping of C ot1-DNA isolated from the Ameiva ameiva and Cnemidophorus sp.1 highlighted centromeric regions of some chromosomes. Thus, the data obtained showed that many repetitive DNA classes are part of the genome of Ameiva ameiva, Cnemidophorus sp.1, Kentroyx calcarata, Kentropyx pelviceps, and Tupinambis teguixin, and these sequences are shared among the analyzed teiid species, but they were not always allocated at the same chromosome position.

  13. High Throughput Analyses of Budding Yeast ARSs Reveal New DNA Elements Capable of Conferring Centromere-Independent Plasmid Propagation.

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    Hoggard, Timothy; Liachko, Ivan; Burt, Cassaundra; Meikle, Troy; Jiang, Katherine; Craciun, Gheorghe; Dunham, Maitreya J; Fox, Catherine A

    2016-04-07

    The ability of plasmids to propagate in Saccharomyces cerevisiae has been instrumental in defining eukaryotic chromosomal control elements. Stable propagation demands both plasmid replication, which requires a chromosomal replication origin (i.e., an ARS), and plasmid distribution to dividing cells, which requires either a chromosomal centromere for segregation or a plasmid-partitioning element. While our knowledge of yeast ARSs and centromeres is relatively advanced, we know less about chromosomal regions that can function as plasmid partitioning elements. The Rap1 protein-binding site (RAP1) present in transcriptional silencers and telomeres of budding yeast is a known plasmid-partitioning element that functions to anchor a plasmid to the inner nuclear membrane (INM), which in turn facilitates plasmid distribution to daughter cells. This Rap1-dependent INM-anchoring also has an important chromosomal role in higher-order chromosomal structures that enhance transcriptional silencing and telomere stability. Thus, plasmid partitioning can reflect fundamental features of chromosome structure and biology, yet a systematic screen for plasmid partitioning elements has not been reported. Here, we couple deep sequencing with competitive growth experiments of a plasmid library containing thousands of short ARS fragments to identify new plasmid partitioning elements. Competitive growth experiments were performed with libraries that differed only in terms of the presence or absence of a centromere. Comparisons of the behavior of ARS fragments in the two experiments allowed us to identify sequences that were likely to drive plasmid partitioning. In addition to the silencer RAP1 site, we identified 74 new putative plasmid-partitioning motifs predicted to act as binding sites for DNA binding proteins enriched for roles in negative regulation of gene expression and G2/M-phase associated biology. These data expand our knowledge of chromosomal elements that may function in plasmid

  14. Human chromosome pellicle antibody recognizing centromere protein—C (CENP0C),the main component of the kinetochore

    Institute of Scientific and Technical Information of China (English)

    XIEYONG; ZUMEINI; 等

    1997-01-01

    Recently the antichromosome antisera from several sclerogerma patients have been found to recognize the pellicle of metaphase and anaphase chromosomes.In order to identify the pellicle components,we used these antichromosome antisera to screen a human embryonic cDNA library.The sequences of the positive clones are identical to the cDNA gene sequence of CENP-C (centromere protein C),a human centromere autoantigen.This result suggusts that CENP-C is a component of the pellicle of human metaphase and anaphase chromosomes.

  15. MHF1–2/CENP-S-X performs distinct roles in centromere metabolism and genetic recombination

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    Bhattacharjee, Sonali; Osman, Fekret; Feeney, Laura; Lorenz, Alexander; Bryer, Claire; Whitby, Matthew C.

    2013-01-01

    The histone-fold proteins Mhf1/CENP-S and Mhf2/CENP-X perform two important functions in vertebrate cells. First, they are components of the constitutive centromere-associated network, aiding kinetochore assembly and function. Second, they work with the FANCM DNA translocase to promote DNA repair. However, it has been unclear whether there is crosstalk between these roles. We show that Mhf1 and Mhf2 in fission yeast, as in vertebrates, serve a dual function, aiding DNA repair/recombination and localizing to centromeres to promote chromosome segregation. Importantly, these functions are distinct, with the former being dependent on their interaction with the FANCM orthologue Fml1 and the latter not. Together with Fml1, they play a second role in aiding chromosome segregation by processing sister chromatid junctions. However, a failure of this activity does not manifest dramatically increased levels of chromosome missegregation due to the Mus81–Eme1 endonuclease, which acts as a failsafe to resolve DNA junctions before the end of mitosis. PMID:24026537

  16. Functional analysis of phosphorylation of the mitotic centromere-associated kinesin by Aurora B kinase in human tumor cells.

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    Ritter, Andreas; Sanhaji, Mourad; Friemel, Alexandra; Roth, Susanne; Rolle, Udo; Louwen, Frank; Yuan, Juping

    2015-01-01

    Mitotic centromere-associated kinesin (MCAK) is the best characterized member of the kinesin-13 family and plays important roles in microtubule dynamics during mitosis. Its activity and subcellular localization is tightly regulated by an orchestra of mitotic kinases, such as Aurora B. It is well known that serine 196 of MCAK is the major phosphorylation site of Aurora B in Xenopus leavis extracts and that this phosphorylation regulates its catalytic activity and subcellular localization. In the current study, we have addressed the conserved phosphorylation site serine 192 in human MCAK to characterize its function in more depth in human cancer cells. Our data confirm that S192 is the major phosphorylation site of Aurora B in human MCAK and that this phosphorylation has crucial roles in regulating its catalytic activity and localization at the kinetochore/centromere region in mitosis. Interfering with this phosphorylation leads to a delayed progression through prometa- and metaphase associated with mitotic defects in chromosome alignment and segregation. We show further that MCAK is involved in directional migration and invasion of tumor cells, and interestingly, interference with the S192 phosphorylation affects this capability of MCAK. These data provide the first molecular explanation for clinical observation, where an overexpression of MCAK was associated with lymphatic invasion and lymph node metastasis in gastric and colorectal cancer patients.

  17. Analysis of premature centromere division (PCD) of the X chromosome in Alzheimer patients through the cell cycle.

    Science.gov (United States)

    Spremo-Potparevic, Biljana; Zivkovic, Lada; Djelic, Ninoslav; Bajic, Vladan

    2004-05-01

    Cytogenetic analysis of the X chromosome in phytohaemagglutinin stimulated peripheral blood lymphocytes was evaluated in 12 sporadic Alzheimer disease (AD) patients and in 11 healthy subjects. For chromosome analysis two methods were used: (1) standard analysis of G-banded metaphase chromosomes and; (2) fluorescent in situ hybridization (FISH) for the detection of the X chromosome centromeric region in interphase nuclei. Cytogenetic analysis revealed that the X chromosome expresses premature centromere division (PCD) in AD females in 10.53% of metaphase cells and in 15.22% of interphase nuclei. In AD men the percentages were 3.98 and 6.06%, respectively. X chromosome PCD in the female control group showed a percentage of 7.46% in metaphase cells and 9.35% in interphase nuclei and in male controls the percentages were 2.84% in metaphases and 5.54% in interphase nuclei. The results of FISH analysis showed that PCD could occur much earlier than metaphase of mitosis, i.e. in interphase of the cell cycle, immediately after replication. The FISH method can be used for PCD verification in all phases of the cell cycle in various disorders including AD.

  18. Toxoplasma gondii chromodomain protein 1 binds to heterochromatin and colocalises with centromeres and telomeres at the nuclear periphery.

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    Mathieu Gissot

    Full Text Available BACKGROUND: Apicomplexan parasites are responsible for some of the most deadly parasitic diseases afflicting humans, including malaria and toxoplasmosis. These obligate intracellular parasites exhibit a complex life cycle and a coordinated cell cycle-dependant expression program. Their cell division is a coordinated multistep process. How this complex mechanism is organised remains poorly understood. METHODS AND FINDINGS: In this study, we provide evidence for a link between heterochromatin, cell division and the compartmentalisation of the nucleus in Toxoplasma gondii. We characterised a T. gondii chromodomain containing protein (named TgChromo1 that specifically binds to heterochromatin. Using ChIP-on-chip on a genome-wide scale, we report TgChromo1 enrichment at the peri-centromeric chromatin. In addition, we demonstrate that TgChromo1 is cell-cycle regulated and co-localised with markers of the centrocone. Through the loci-specific FISH technique for T. gondii, we confirmed that TgChromo1 occupies the same nuclear localisation as the peri-centromeric sequences. CONCLUSION: We propose that TgChromo1 may play a role in the sequestration of chromosomes at the nuclear periphery and in the process of T. gondii cell division.

  19. Loss of maternal ATRX results in centromere instability and aneuploidy in the mammalian oocyte and pre-implantation embryo.

    Science.gov (United States)

    Baumann, Claudia; Viveiros, Maria M; De La Fuente, Rabindranath

    2010-09-23

    The α-thalassemia/mental retardation X-linked protein (ATRX) is a chromatin-remodeling factor known to regulate DNA methylation at repetitive sequences of the human genome. We have previously demonstrated that ATRX binds to pericentric heterochromatin domains in mouse oocytes at the metaphase II stage where it is involved in mediating chromosome alignment at the meiotic spindle. However, the role of ATRX in the functional differentiation of chromatin structure during meiosis is not known. To test ATRX function in the germ line, we developed an oocyte-specific transgenic RNAi knockdown mouse model. Our results demonstrate that ATRX is required for heterochromatin formation and maintenance of chromosome stability during meiosis. During prophase I arrest, ATRX is necessary to recruit the transcriptional regulator DAXX (death domain associated protein) to pericentric heterochromatin. At the metaphase II stage, transgenic ATRX-RNAi oocytes exhibit abnormal chromosome morphology associated with reduced phosphorylation of histone 3 at serine 10 as well as chromosome segregation defects leading to aneuploidy and severely reduced fertility. Notably, a large proportion of ATRX-depleted oocytes and 1-cell stage embryos exhibit chromosome fragments and centromeric DNA-containing micronuclei. Our results provide novel evidence indicating that ATRX is required for centromere stability and the epigenetic control of heterochromatin function during meiosis and the transition to the first mitosis.

  20. Loss of maternal ATRX results in centromere instability and aneuploidy in the mammalian oocyte and pre-implantation embryo.

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    Claudia Baumann

    2010-09-01

    Full Text Available The α-thalassemia/mental retardation X-linked protein (ATRX is a chromatin-remodeling factor known to regulate DNA methylation at repetitive sequences of the human genome. We have previously demonstrated that ATRX binds to pericentric heterochromatin domains in mouse oocytes at the metaphase II stage where it is involved in mediating chromosome alignment at the meiotic spindle. However, the role of ATRX in the functional differentiation of chromatin structure during meiosis is not known. To test ATRX function in the germ line, we developed an oocyte-specific transgenic RNAi knockdown mouse model. Our results demonstrate that ATRX is required for heterochromatin formation and maintenance of chromosome stability during meiosis. During prophase I arrest, ATRX is necessary to recruit the transcriptional regulator DAXX (death domain associated protein to pericentric heterochromatin. At the metaphase II stage, transgenic ATRX-RNAi oocytes exhibit abnormal chromosome morphology associated with reduced phosphorylation of histone 3 at serine 10 as well as chromosome segregation defects leading to aneuploidy and severely reduced fertility. Notably, a large proportion of ATRX-depleted oocytes and 1-cell stage embryos exhibit chromosome fragments and centromeric DNA-containing micronuclei. Our results provide novel evidence indicating that ATRX is required for centromere stability and the epigenetic control of heterochromatin function during meiosis and the transition to the first mitosis.

  1. Fluorescence In Situ Hybridization (FISH-Based Karyotyping Reveals Rapid Evolution of Centromeric and Subtelomeric Repeats in Common Bean (Phaseolus vulgaris and Relatives

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    Aiko Iwata-Otsubo

    2016-04-01

    Full Text Available Fluorescence in situ hybridization (FISH-based karyotyping is a powerful cytogenetics tool to study chromosome organization, behavior, and chromosome evolution. Here, we developed a FISH-based karyotyping system using a probe mixture comprised of centromeric and subtelomeric satellite repeats, 5S rDNA, and chromosome-specific BAC clones in common bean, which enables one to unambiguously distinguish all 11 chromosome pairs. Furthermore, we applied the karyotyping system to several wild relatives and landraces of common bean from two distinct gene pools, as well as other related Phaseolus species, to investigate repeat evolution in the genus Phaseolus. Comparison of karyotype maps within common bean indicates that chromosomal distribution of the centromeric and subtelomeric satellite repeats is stable, whereas the copy number of the repeats was variable, indicating rapid amplification/reduction of the repeats in specific genomic regions. In Phaseolus species that diverged approximately 2–4 million yr ago, copy numbers of centromeric repeats were largely reduced or diverged, and chromosomal distributions have changed, suggesting rapid evolution of centromeric repeats. We also detected variation in the distribution pattern of subtelomeric repeats in Phaseolus species. The FISH-based karyotyping system revealed that satellite repeats are actively and rapidly evolving, forming genomic features unique to individual common bean accessions and Phaseolus species.

  2. Schizosaccharomyces pombe centromere protein Mis19 links Mis16 and Mis18 to recruit CENP-A through interacting with NMD factors and the SWI/SNF complex.

    Science.gov (United States)

    Hayashi, Takeshi; Ebe, Masahiro; Nagao, Koji; Kokubu, Aya; Sajiki, Kenichi; Yanagida, Mitsuhiro

    2014-07-01

    CENP-A is a centromere-specific variant of histone H3 that is required for accurate chromosome segregation. The fission yeast Schizosaccharomyces pombe and mammalian Mis16 and Mis18 form a complex essential for CENP-A recruitment to centromeres. It is unclear, however, how the Mis16-Mis18 complex achieves this function. Here, we identified, by mass spectrometry, novel fission yeast centromere proteins Mis19 and Mis20 that directly interact with Mis16 and Mis18. Like Mis18, Mis19 and Mis20 are localized at the centromeres during interphase, but not in mitosis. Inactivation of Mis19 in a newly isolated temperature-sensitive mutant resulted in CENP-A delocalization and massive chromosome missegregation, whereas Mis20 was dispensable for proper chromosome segregation. Mis19 might be a bridge component for Mis16 and Mis18. We isolated extragenic suppressor mutants for temperature-sensitive mis18 and mis19 mutants and used whole-genome sequencing to determine the mutated sites. We identified two groups of loss-of-function suppressor mutations in non-sense-mediated mRNA decay factors (upf2 and ebs1), and in SWI/SNF chromatin-remodeling components (snf5, snf22 and sol1). Our results suggest that the Mis16-Mis18-Mis19-Mis20 CENP-A-recruiting complex, which is functional in the G1-S phase, may be counteracted by the SWI/SNF chromatin-remodeling complex and non-sense-mediated mRNA decay, which may prevent CENP-A deposition at the centromere.

  3. Statistical analysis of 3D images detects regular spatial distributions of centromeres and chromocenters in animal and plant nuclei.

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    Philippe Andrey

    Full Text Available In eukaryotes, the interphase nucleus is organized in morphologically and/or functionally distinct nuclear "compartments". Numerous studies highlight functional relationships between the spatial organization of the nucleus and gene regulation. This raises the question of whether nuclear organization principles exist and, if so, whether they are identical in the animal and plant kingdoms. We addressed this issue through the investigation of the three-dimensional distribution of the centromeres and chromocenters. We investigated five very diverse populations of interphase nuclei at different differentiation stages in their physiological environment, belonging to rabbit embryos at the 8-cell and blastocyst stages, differentiated rabbit mammary epithelial cells during lactation, and differentiated cells of Arabidopsis thaliana plantlets. We developed new tools based on the processing of confocal images and a new statistical approach based on G- and F- distance functions used in spatial statistics. Our original computational scheme takes into account both size and shape variability by comparing, for each nucleus, the observed distribution against a reference distribution estimated by Monte-Carlo sampling over the same nucleus. This implicit normalization allowed similar data processing and extraction of rules in the five differentiated nuclei populations of the three studied biological systems, despite differences in chromosome number, genome organization and heterochromatin content. We showed that centromeres/chromocenters form significantly more regularly spaced patterns than expected under a completely random situation, suggesting that repulsive constraints or spatial inhomogeneities underlay the spatial organization of heterochromatic compartments. The proposed technique should be useful for identifying further spatial features in a wide range of cell types.

  4. Escape from Mitotic Arrest: An Unexpected Connection Between Microtubule Dynamics and Epigenetic Regulation of Centromeric Chromatin in Schizosaccharomyces pombe.

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    George, Anuja A; Walworth, Nancy C

    2015-12-01

    Accurate chromosome segregation is necessary to ensure genomic integrity. Segregation depends on the proper functioning of the centromere, kinetochore, and mitotic spindle microtubules and is monitored by the spindle assembly checkpoint (SAC). In the fission yeast Schizosaccharomyces pombe, defects in Dis1, a microtubule-associated protein that influences microtubule dynamics, lead to mitotic arrest as a result of an active SAC and consequent failure to grow at low temperature. In a mutant dis1 background (dis1-288), loss of function of Msc1, a fission yeast homolog of the KDM5 family of proteins, suppresses the growth defect and promotes normal mitosis. Genetic analysis implicates a histone deacetylase (HDAC)-linked pathway in suppression because HDAC mutants clr6-1, clr3∆, and sir2∆, though not hos2∆, also promote normal mitosis in the dis1-288 mutant. Suppression of the dis phenotype through loss of msc1 function requires the spindle checkpoint protein Mad2 and is limited by the presence of the heterochromatin-associated HP1 protein homolog Swi6. We speculate that alterations in histone acetylation promote a centromeric chromatin environment that compensates for compromised dis1 function by allowing for successful kinetochore-microtubule interactions that can satisfy the SAC. In cells arrested in mitosis by mutation of dis1, loss of function of epigenetic determinants such as Msc1 or specific HDACs can promote cell survival. Because the KDM5 family of proteins has been implicated in human cancers, an appreciation of the potential role of this family of proteins in chromosome segregation is warranted.

  5. SGO1 maintains bovine meiotic and mitotic centromeric cohesions of sister chromatids and directly affects embryo development.

    Directory of Open Access Journals (Sweden)

    Feng-Xia Yin

    Full Text Available Shugoshin (SGO is a critical factor that enforces cohesion from segregation of paired sister chromatids during mitosis and meiosis. It has been studied mainly in invertebrates. Knowledge of SGO(s in a mammalian system has only been reported in the mouse and Hela cells. In this study, the functions of SGO1 in bovine oocytes during meiotic maturation, early embryonic development and somatic cell mitosis were investigated. The results showed that SGO1 was expressed from germinal vesicle (GV to the metaphase II stage. SGO1 accumulated on condensed and scattered chromosomes from pre-metaphase I to metaphase II. The over-expression of SGO1 did not interfere with the process of homologous chromosome separation, although once separated they were unable to move to the opposing spindle poles. This often resulted in the formation of oocytes with 60 replicated chromosomes. Depletion of SGO1 in GV oocytes affected chromosomal separation resulting in abnormal chromosome alignment at a significantly higher proportion than in control oocytes. Knockdown of SGO1 expression significantly decreased the embryonic developmental rate and quality. To further confirm the function(s of SGO1 during mitosis, bovine embryonic fibroblast cells were transfected with SGO1 siRNAs. SGO1 depletion induced the premature dissociation of chromosomal cohesion at the centromere and along the chromosome arm giving rise to abnormal appearing mitotic patterns. The results of this study infer that SGO1 is involved in the centromeric cohesion of sister chromatids and chromosomal movement towards the spindle poles. Depletion of SGO1 causes arrestment of cell division in meiosis and mitosis.

  6. Lack of independent prognostic and predictive value of centromere 17 copy number changes in breast cancer patients with known HER2 and TOP2A status

    DEFF Research Database (Denmark)

    Nielsen, Kirsten Vang; Ejlertsen, Bent; Møller, Susanne

    2011-01-01

    The clinical benefit of anthracyclines has been connected to HER2 status, TOP2A status and centromere 17 copy numbers (CEN-17). Data from a clinical trial randomizing patients to anthracyclines was used to assess whether the number of CEN-17 in breast cancers may predict incremental responsiveness...... to anthracyclines besides what is obtained when used relatively to TOP2A and HER2. As cut sections of paraffin-embedded tissue are prone to truncation of nuclei, strict definition of ploidy levels is lacking. We therefore used normal breast tissue to assist define ploidy levels in cut sections. Fluorescence in situ...... hybridization (FISH) with centromere 17 (CEN-17) and TOP2A was performed on 120 normal breast specimens. The diploid CEN-17 copy number was reduced from the expected two signals in whole nuclei to an average of 1.68 signals per nucleus in cut sections of normal breast. Ploidy levels determined in normal breast...

  7. A stable acentric marker chromosome derived from distal 8p: Reactivation of a latent ancient centromere at 8p23.1?

    Energy Technology Data Exchange (ETDEWEB)

    Ohashi, H.; Wakui, K.; Ogawa, K. [Saitama Children`s Medical Ctr., Iwatsuki (Japan)] [and others

    1994-09-01

    Centromere is considered to be an essential chromosomal component for faithful segregation, and acentric chromosomes are unstable and lost through cell divisions. We report a novel marker chromosome that was acentric but stable through cell divisions. The patient was a 2-year-old girl with mental retardation, patent ductus arteriosus and mild dysmorphic features. G-banded chromosome analysis revealed an additional small marker chromosome in all 100 cells examined. Using the targeted chromosome-band painting method, the marker was found to originate from the distal region of 8p, and subsequent two color FISH analysis with cosmid probes around the region revealed the marker was a rearranged chromosome interpreted as 8pter{r_arrow}p23.1{r_arrow}8pter. No centromeric region was involved in the marker. By FISH, no {alpha}-satellite sequence was detected on the marker, while telomere sequence was detected at each end. Antikinetochore immunostaining using a serum from a patient with CREST syndrome showed a pair of signals on the marker, which indicated that a functional kinetochore was present on the marker, presumably at 8p23.1-corresponding region. The patient may provide evidence that an ancient centromere sequence exists at 8p23.1 and was reactivated through the chromosome rearrangement in the patient.

  8. Preferential recruitment of the maternal centromere-specific histone H3 (CENH3) in oat (Avena sativa L.) × pearl millet (Pennisetum glaucum L.) hybrid embryos.

    Science.gov (United States)

    Ishii, Takayoshi; Sunamura, Naohiro; Matsumoto, Ayaka; Eltayeb, Amin Elsadig; Tsujimoto, Hisashi

    2015-12-01

    Chromosome elimination occurs frequently in interspecific hybrids between distantly related species in Poaceae. However, chromosomes from both parents behave stably in a hybrid of female oat (Avena sativa L.) pollinated by pearl millet (Pennisetum glaucum L.). To analyze the chromosome behavior in this hybrid, we cloned the centromere-specific histone H3 (CENH3) genes of oat and pearl millet and produced a pearl millet-specific anti-CENH3 antibody. Application of this antibody together with a grass species common anti-CENH3 antibody revealed the dynamic CENH3 composition of the hybrid cells before and after fertilization. Despite co-expression of CENH3 genes encoded by oat and pearl millet, only an oat-type CENH3 was incorporated into the centromeres of both species in the hybrid embryo. Oat CENH3 enables a functional centromere in pearl millet chromosomes in an oat genetic background. Comparison of CENH3 genes among Poaceae species that show chromosome elimination in interspecific hybrids revealed that the loop 1 regions of oat and pearl millet CENH3 exhibit exceptionally high similarity.

  9. DNA topoisomerase II is a determinant of the tensile properties of yeast centromeric chromatin and the tension checkpoint.

    Science.gov (United States)

    Warsi, Tariq H; Navarro, Michelle S; Bachant, Jeff

    2008-10-01

    Centromeric (CEN) chromatin is placed under mechanical tension and stretches as kinetochores biorient on the mitotic spindle. This deformation could conceivably provide a readout of biorientation to error correction mechanisms that monitor kinetochore-spindle interactions, but whether CEN chromatin acts in a tensiometer capacity is unresolved. Here, we report observations linking yeast Topoisomerase II (Top2) to both CEN mechanics and assessment of interkinetochore tension. First, in top2-4 and sumoylation-resistant top2-SNM mutants CEN chromatin stretches extensively during biorientation, resulting in increased sister kinetochore separation and preanaphase spindle extension. Our data indicate increased CEN stretching corresponds with alterations to CEN topology induced in response to tension. Second, Top2 potentiates aspects of the tension checkpoint. Mutations affecting the Mtw1 kinetochore protein activate Ipl1 kinase to detach kinetochores and induce spindle checkpoint arrest. In mtw1top2-4 and mtw1top2-SNM mutants, however, kinetochores are resistant to detachment and checkpoint arrest is attenuated. For top2-SNM cells, CEN stretching and checkpoint attenuation occur even in the absence of catenation linking sister chromatids. In sum, Top2 seems to play a novel role in CEN compaction that is distinct from decatenation. Perturbations to this function may allow weakened kinetochores to stretch CENs in a manner that mimics tension or evades Ipl1 surveillance.

  10. A KIR B centromeric region present in Africans but not Europeans protects pregnant women from pre-eclampsia.

    Science.gov (United States)

    Nakimuli, Annettee; Chazara, Olympe; Hiby, Susan E; Farrell, Lydia; Tukwasibwe, Stephen; Jayaraman, Jyothi; Traherne, James A; Trowsdale, John; Colucci, Francesco; Lougee, Emma; Vaughan, Robert W; Elliott, Alison M; Byamugisha, Josaphat; Kaleebu, Pontiano; Mirembe, Florence; Nemat-Gorgani, Neda; Parham, Peter; Norman, Paul J; Moffett, Ashley

    2015-01-20

    In sub-Saharan Africans, maternal mortality is unacceptably high, with >400 deaths per 100,000 births compared with pre-eclampsia, a syndrome arising from defective placentation. Controlling placentation are maternal natural killer (NK) cells that use killer-cell immunoglobulin-like receptor (KIR) to recognize the fetal HLA-C molecules on invading trophoblast. We analyzed genetic polymorphisms of maternal KIR and fetal HLA-C in 484 normal and 254 pre-eclamptic pregnancies at Mulago Hospital, Kampala, Uganda. The combination of maternal KIR AA genotypes and fetal HLA-C alleles encoding the C2 epitope associates with pre-eclampsia [P = 0.0318, odds ratio (OR) = 1.49]. The KIR genes associated with protection are located in centromeric KIR B regions that are unique to sub-Saharan African populations and contain the KIR2DS5 and KIR2DL1 genes (P = 0.0095, OR = 0.59). By contrast, telomeric KIR B genes protect Europeans against pre-eclampsia. Thus, different KIR B regions protect sub-Saharan Africans and Europeans from pre-eclampsia, whereas in both populations, the KIR AA genotype is a risk factor for the syndrome. These results emphasize the importance of undertaking genetic studies of pregnancy disorders in African populations with the potential to provide biological insights not available from studies restricted to European populations.

  11. Condensin HEAT subunits required for DNA repair, kinetochore/centromere function and ploidy maintenance in fission yeast.

    Directory of Open Access Journals (Sweden)

    Xingya Xu

    Full Text Available Condensin, a central player in eukaryotic chromosomal dynamics, contains five evolutionarily-conserved subunits. Two SMC (structural maintenance of chromosomes subunits contain ATPase, hinge, and coiled-coil domains. One non-SMC subunit is similar to bacterial kleisin, and two other non-SMC subunits contain HEAT (similar to armadillo repeats. Here we report isolation and characterization of 21 fission yeast (Schizosaccharomyces pombe mutants for three non-SMC subunits, created using error-prone mutagenesis that resulted in single-amino acid substitutions. Beside condensation, segregation, and DNA repair defects, similar to those observed in previously isolated SMC and cnd2 mutants, novel phenotypes were observed for mutants of HEAT-repeats containing Cnd1 and Cnd3 subunits. cnd3-L269P is hypersensitive to the microtubule poison, thiabendazole, revealing defects in kinetochore/centromere and spindle assembly checkpoints. Three cnd1 and three cnd3 mutants increased cell size and doubled DNA content, thereby eliminating the haploid state. Five of these mutations reside in helix B of HEAT repeats. Two non-SMC condensin subunits, Cnd1 and Cnd3, are thus implicated in ploidy maintenance.

  12. High resolution mapping of Dense spike-ar (dsp.ar) to the genetic centromere of barley chromosome 7H.

    Science.gov (United States)

    Shahinnia, Fahimeh; Druka, Arnis; Franckowiak, Jerome; Morgante, Michele; Waugh, Robbie; Stein, Nils

    2012-02-01

    Spike density in barley is under the control of several major genes, as documented previously by genetic analysis of a number of morphological mutants. One such class of mutants affects the rachis internode length leading to dense or compact spikes and the underlying genes were designated dense spike (dsp). We previously delimited two introgressed genomic segments on chromosome 3H (21 SNP loci, 35.5 cM) and 7H (17 SNP loci, 20.34 cM) in BW265, a BC(7)F(3) nearly isogenic line (NIL) of cv. Bowman as potentially containing the dense spike mutant locus dsp.ar, by genotyping 1,536 single nucleotide polymorphism (SNP) markers in both BW265 and its recurrent parent. Here, the gene was allocated by high-resolution bi-parental mapping to a 0.37 cM interval between markers SC57808 (Hv_SPL14)-CAPSK06413 residing on the short and long arm at the genetic centromere of chromosome 7H, respectively. This region putatively contains more than 800 genes as deduced by comparison with the collinear regions of barley, rice, sorghum and Brachypodium, Classical map-based isolation of the gene dsp.ar thus will be complicated due to the infavorable relationship of genetic to physical distances at the target locus.

  13. Detection and Automated Scoring of Dicentric Chromosomes in Nonstimulated Lymphocyte Prematurely Condensed Chromosomes After Telomere and Centromere Staining

    Energy Technology Data Exchange (ETDEWEB)

    M' kacher, Radhia [Laboratoire de Radiobiologie et Oncologie, Commissariat à l' Energie Atomique, Fontenay-aux-Roses (France); El Maalouf, Elie [Laboratoire de Radiobiologie et Oncologie, Commissariat à l' Energie Atomique, Fontenay-aux-Roses (France); Laboratoire Modélisation Intelligence Processus Systèmes (MIPS)–Groupe TIIM3D, Université de Haute-Alsace, Mulhouse (France); Terzoudi, Georgia [Laboratory of Radiobiology & Biodosimetry, National Center for Scientific Research Demokritos, Athens (Greece); Ricoul, Michelle [Laboratoire de Radiobiologie et Oncologie, Commissariat à l' Energie Atomique, Fontenay-aux-Roses (France); Heidingsfelder, Leonhard [MetaSystems, Altlussheim (Germany); Karachristou, Ionna [Laboratory of Radiobiology & Biodosimetry, National Center for Scientific Research Demokritos, Athens (Greece); Laplagne, Eric [Pole Concept, Paris (France); Hempel, William M. [Laboratoire de Radiobiologie et Oncologie, Commissariat à l' Energie Atomique, Fontenay-aux-Roses (France); Colicchio, Bruno; Dieterlen, Alain [Laboratoire Modélisation Intelligence Processus Systèmes (MIPS)–Groupe TIIM3D, Université de Haute-Alsace, Mulhouse (France); Pantelias, Gabriel [Laboratory of Radiobiology & Biodosimetry, National Center for Scientific Research Demokritos, Athens (Greece); Sabatier, Laure, E-mail: laure.sabatier@cea.fr [Laboratoire de Radiobiologie et Oncologie, Commissariat à l' Energie Atomique, Fontenay-aux-Roses (France)

    2015-03-01

    Purpose: To combine telomere and centromere (TC) staining of premature chromosome condensation (PCC) fusions to identify dicentrics, centric rings, and acentric chromosomes, making possible the realization of a dose–response curve and automation of the process. Methods and Materials: Blood samples from healthy donors were exposed to {sup 60}Co irradiation at varying doses up to 8 Gy, followed by a repair period of 8 hours. Premature chromosome condensation fusions were carried out, and TC staining using peptide nucleic acid probes was performed. Chromosomal aberration (CA) scoring was carried out manually and automatically using PCC-TCScore software, developed in our laboratory. Results: We successfully optimized the hybridization conditions and image capture parameters, to increase the sensitivity and effectiveness of CA scoring. Dicentrics, centric rings, and acentric chromosomes were rapidly and accurately detected, leading to a linear-quadratic dose–response curve by manual scoring at up to 8 Gy. Using PCC-TCScore software for automatic scoring, we were able to detect 95% of dicentrics and centric rings. Conclusion: The introduction of TC staining to the PCC fusion technique has made possible the rapid scoring of unstable CAs, including dicentrics, with a level of accuracy and ease not previously possible. This new approach can be used for biological dosimetry in radiation emergency medicine, where the rapid and accurate detection of dicentrics is a high priority using automated scoring. Because there is no culture time, this new approach can also be used for the follow-up of patients treated by genotoxic therapy, creating the possibility to perform the estimation of induced chromosomal aberrations immediately after the blood draw.

  14. Anti-centromere antibody-seropositive Sjögren's syndrome differs from conventional subgroup in clinical and pathological study

    Directory of Open Access Journals (Sweden)

    Ida Hiroaki

    2010-07-01

    Full Text Available Abstract Background To clarify the clinicopathological characteristics of primary Sjögren's syndrome (pSS with anti-centromere antibody (ACA. Methods Characteristics of 14 patients of pSS with ACA were evaluated. All patients were anti-SS-A/Ro and SS-B/La antibodies negative (ACA+ group without sclerodactyly. The prevalence of Raynaud's phenomenon (RP, titer of IgG and focus score (FS in the minor salivary glands (MSGs were determined. Quantification analysis of Azan Mallory staining was performed to detect collagenous fiber. Forty eight patients in whom ACA was absent were chosen as the conventional (ACA- pSS group. Results Prevalence of ACA+ SS patients was 14 out of 129 (10.85% pSS patients. RP was observed in 61.5% of the patients with ACA. The level of IgG in the ACA+ group was significantly lower than that of the ACA- group (p = 0.018. Statistical difference was also found in the FS of MSGs from the ACA+ group (1.4 ± 1.0 as compared with the ACA- group (2.3 ± 1.6 (p = 0.035. In contrast, the amount of fibrous tissue was much higher in the ACA+ group (65052.2 ± 14520.6 μm2 versus 26251.3 ± 14249.8 μm2 (p = 1.3 × 10-12. Conclusions Low cellular infiltration but with an increase in fibrous tissues may explain the clinical feature of a high prevalence of RP and normal IgG concentration in ACA+ pSS.

  15. Molecular characterization and chromosomal distribution of a species-specific transcribed centromeric satellite repeat from the olive fruit fly, Bactrocera oleae.

    Directory of Open Access Journals (Sweden)

    Konstantina T Tsoumani

    Full Text Available Satellite repetitive sequences that accumulate in the heterochromatin consist a large fraction of a genome and due to their properties are suggested to be implicated in centromere function. Current knowledge of heterochromatic regions of Bactrocera oleae genome, the major pest of the olive tree, is practically nonexistent. In our effort to explore the repetitive DNA portion of B. oleae genome, a novel satellite sequence designated BoR300 was isolated and cloned. The present study describes the genomic organization, abundance and chromosomal distribution of BoR300 which is organized in tandem, forming arrays of 298 bp-long monomers. Sequence analysis showed an AT content of 60.4%, a CENP-B like-motif and a high curvature value based on predictive models. Comparative analysis among randomly selected monomers demonstrated a high degree of sequence homogeneity (88%-97% of BoR300 repeats, which are present at approximately 3,000 copies per haploid genome accounting for about 0.28% of the total genomic DNA, based on two independent qPCR approaches. In addition, expression of the repeat was also confirmed through RT-PCR, by which BoR300 transcripts were detected in both sexes. Fluorescence in situ hybridization (FISH of BoR300 on mitotic metaphases and polytene chromosomes revealed signals to the centromeres of two out of the six chromosomes which indicated a chromosome-specific centromeric localization. Moreover, BoR300 is not conserved in the closely related Bactrocera species tested and it is also absent in other dipterans, but it's rather restricted to the B. oleae genome. This feature of species-specificity attributed to BoR300 satellite makes it a good candidate as an identification probe of the insect among its relatives at early development stages.

  16. Advance in Research of Centromeres in the Meiotic Process%着丝粒及其在减数分裂中的作用研究进展

    Institute of Scientific and Technical Information of China (English)

    张峰; 林娟; 赵军

    2013-01-01

    Centromeres are the important functional elements of chromosomes in eukaryotes, their roles played in mitosis and meiosis processes have been paid more and more attentions. Though the chromosomes vary greatly in DNA sequences, their roles are conserved in all eukaryotic organisms, which make sure the faithful segregation of chromosomes and correct cell division. With the continuous development of molecular biology techniques, we have a deep understanding of the functions of centromeres. Here we review the function of centromeres during the meiotic progress.%着丝粒作为真核生物染色体的重要结构之一,是有丝分裂和减数分裂过程中重要的功能元件,其所起到的重要作用越来越受到人们的重视。在整个真核生物类群中,尽管不同物种之间着丝粒的DNA序列相差极大,但是其功能却是相当保守,这确保了着丝粒在调控细胞分裂和染色单体分离过程中能够正常行使其功能,除此之外,人们还发现着丝粒蛋白不仅在细胞的有丝分裂中起作用,还在减数分裂中起着重要的作用。

  17. The cell cycle timing of centromeric chromatin assembly in Drosophila meiosis is distinct from mitosis yet requires CAL1 and CENP-C.

    Science.gov (United States)

    Dunleavy, Elaine M; Beier, Nicole L; Gorgescu, Walter; Tang, Jonathan; Costes, Sylvain V; Karpen, Gary H

    2012-01-01

    CENP-A (CID in flies) is the histone H3 variant essential for centromere specification, kinetochore formation, and chromosome segregation during cell division. Recent studies have elucidated major cell cycle mechanisms and factors critical for CENP-A incorporation in mitosis, predominantly in cultured cells. However, we do not understand the roles, regulation, and cell cycle timing of CENP-A assembly in somatic tissues in multicellular organisms and in meiosis, the specialized cell division cycle that gives rise to haploid gametes. Here we investigate the timing and requirements for CID assembly in mitotic tissues and male and female meiosis in Drosophila melanogaster, using fixed and live imaging combined with genetic approaches. We find that CID assembly initiates at late telophase and continues during G1 phase in somatic tissues in the organism, later than the metaphase assembly observed in cultured cells. Furthermore, CID assembly occurs at two distinct cell cycle phases during male meiosis: prophase of meiosis I and after exit from meiosis II, in spermatids. CID assembly in prophase I is also conserved in female meiosis. Interestingly, we observe a novel decrease in CID levels after the end of meiosis I and before meiosis II, which correlates temporally with changes in kinetochore organization and orientation. We also demonstrate that CID is retained on mature sperm despite the gross chromatin remodeling that occurs during protamine exchange. Finally, we show that the centromere proteins CAL1 and CENP-C are both required for CID assembly in meiosis and normal progression through spermatogenesis. We conclude that the cell cycle timing of CID assembly in meiosis is different from mitosis and that the efficient propagation of CID through meiotic divisions and on sperm is likely to be important for centromere specification in the developing zygote.

  18. The cell cycle timing of centromeric chromatin assembly in Drosophila meiosis is distinct from mitosis yet requires CAL1 and CENP-C.

    Directory of Open Access Journals (Sweden)

    Elaine M Dunleavy

    Full Text Available CENP-A (CID in flies is the histone H3 variant essential for centromere specification, kinetochore formation, and chromosome segregation during cell division. Recent studies have elucidated major cell cycle mechanisms and factors critical for CENP-A incorporation in mitosis, predominantly in cultured cells. However, we do not understand the roles, regulation, and cell cycle timing of CENP-A assembly in somatic tissues in multicellular organisms and in meiosis, the specialized cell division cycle that gives rise to haploid gametes. Here we investigate the timing and requirements for CID assembly in mitotic tissues and male and female meiosis in Drosophila melanogaster, using fixed and live imaging combined with genetic approaches. We find that CID assembly initiates at late telophase and continues during G1 phase in somatic tissues in the organism, later than the metaphase assembly observed in cultured cells. Furthermore, CID assembly occurs at two distinct cell cycle phases during male meiosis: prophase of meiosis I and after exit from meiosis II, in spermatids. CID assembly in prophase I is also conserved in female meiosis. Interestingly, we observe a novel decrease in CID levels after the end of meiosis I and before meiosis II, which correlates temporally with changes in kinetochore organization and orientation. We also demonstrate that CID is retained on mature sperm despite the gross chromatin remodeling that occurs during protamine exchange. Finally, we show that the centromere proteins CAL1 and CENP-C are both required for CID assembly in meiosis and normal progression through spermatogenesis. We conclude that the cell cycle timing of CID assembly in meiosis is different from mitosis and that the efficient propagation of CID through meiotic divisions and on sperm is likely to be important for centromere specification in the developing zygote.

  19. Quantification, by solid-phase minisequencing, of the telomeric and centromeric copies of the survival motor neuron gene in families with spinal muscular atrophy

    DEFF Research Database (Denmark)

    Schwartz, M; Sørensen, N; Hansen, F J

    1997-01-01

    In an analysis of 30 families affected by spinal muscular atrophy (SMA) we have used the solid-phase minisequencing method to determine the ratio between the number of telomeric and centromeric copies of the survival motor neuron gene (SMN and cBCD541 respectively) on normal and SMA chromosomes...... be explained by an underrepresentation of the haplotype completely lacking SMN genes, which is expected to cause early embryonic death in homozygotes. This first report of a direct haplotype analysis of SMN and cBCD541 should help clarify the role of cBCD541 in the pathogenesis of SMA....

  20. DNA deformability changes of single base pair mutants within CDE binding sites in S. Cerevisiae centromere DNA correlate with measured chromosomal loss rates and CDE binding site symmetries

    Directory of Open Access Journals (Sweden)

    Marx Kenneth A

    2006-03-01

    Full Text Available Abstract Background The centromeres in yeast (S. cerevisiae are organized by short DNA sequences (125 bp on each chromosome consisting of 2 conserved elements: CDEI and CDEIII spaced by a CDEII region. CDEI and CDEIII are critical sequence specific protein binding sites necessary for correct centromere formation and following assembly with proteins, are positioned near each other on a specialized nucleosome. Hegemann et al. BioEssays 1993, 15: 451–460 reported single base DNA mutants within the critical CDEI and CDEIII binding sites on the centromere of chromosome 6 and quantitated centromere loss of function, which they measured as loss rates for the different chromosome 6 mutants during cell division. Olson et al. Proc Natl Acad Sci USA 1998, 95: 11163–11168 reported the use of protein-DNA crystallography data to produce a DNA dinucleotide protein deformability energetic scale (PD-scale that describes local DNA deformability by sequence specific binding proteins. We have used the PD-scale to investigate the DNA sequence dependence of the yeast chromosome 6 mutants' loss rate data. Each single base mutant changes 2 PD-scale values at that changed base position relative to the wild type. In this study, we have utilized these mutants to demonstrate a correlation between the change in DNA deformability of the CDEI and CDEIII core sites and the overall experimentally measured chromosome loss rates of the chromosome 6 mutants. Results In the CDE I and CDEIII core binding regions an increase in the magnitude of change in deformability of chromosome 6 single base mutants with respect to the wild type correlates to an increase in the measured chromosome loss rate. These correlations were found to be significant relative to 105 Monte Carlo randomizations of the dinucleotide PD-scale applied to the same calculation. A net loss of deformability also tends to increase the loss rate. Binding site position specific, 4 data-point correlations were also

  1. Actin-binding protein (ABP-280) filamin gene (FLN) maps telomeric to the color vision locus (R/GCP) and centromeric to G6PD in Xq28

    Energy Technology Data Exchange (ETDEWEB)

    Gorlin, J.B. (Brigham and Women' s Hospital, Boston, MA (United States) Dana-Farber Cancer Institute, Boston, MA (United States)); Henske, E.; Hartwig, J.H.; Kwiatkowski, D.J. (Brigham and Women' s Hospital, Boston, MA (United States)); Warren, S.T.; Kunst, C.B. (Emory Univ. School of Medicine, Atlanta, GA (United States)); D' Urso, M.; Palmieri, G. (International Institute of Genetics and Biophysics, Naples, (Italy)); Bruns, G. (Children' s Hospital, Boston, MA (United States))

    1993-08-01

    Actin-binding protein-280 (ABP-280) is a dimeric actin filament-crosslinking protein that promotes orthogonal branching of actin filaments and links actin filaments to membrane glycoproteins. The authors have mapped the ABP-280 filamin gene (FLN) to Xq28 by Southern blot analysis of somatic cell hybrid lines, by fluorescence in situ hybridization, and through identification of portions of the FLN gene within cosmids and YACs mapped to Xq28. The FLN gene is found within a 200-kb region centromeric to the G6PD locus and telomeric to DSX52 and the color vision locus. 23 refs., 2 figs.

  2. 植物着丝粒区串联重复序列的研究进展%Research Progress of Tandem Repetitive Sequence in the Centromere of Plant

    Institute of Scientific and Technical Information of China (English)

    郝薇薇; 周岩

    2013-01-01

      着丝粒是细胞染色体的重要结构组成,控制姊妹染色单体的结合、动粒的组装和纺锤丝的附着,确保真核生物细胞在有丝分裂和减数分裂过程中染色体的正常分离及遗传信息的稳定传递。植物着丝粒DNA序列主要由反转录转座子和串联重复序列构成。串联重复序列在着丝粒功能实现和基因组进化过程中起重要作用。随着测序技术的成熟,近年来对串联重复序列的研究取得了很大的进展。综述了植物串联重复序列结构、分析方法及在进化中的作用,以期为相关研究提供参考。%Centromeres are the important domains of chromosomes that are responsible for sister chromatid cohesion, kinetochore assembly and spindle attachment, and are essential for proper chromosome segregation during mitosis and meiosis. Satellite DNA and retrotransposons are the most abundant DNA elements found in plant centromere regions. Centromeric tandem repeat play an important role in the centromere function and genome evolution. The study of centromeric tandem repeats got great progress for the development of sequencing technology. This paper introduces the development of centromeric tandem repeat of plants.

  3. Transcriptional Response of Human Neurospheres to Helper-Dependent CAV-2 Vectors Involves the Modulation of DNA Damage Response, Microtubule and Centromere Gene Groups.

    Directory of Open Access Journals (Sweden)

    Stefania Piersanti

    Full Text Available Brain gene transfer using viral vectors will likely become a therapeutic option for several disorders. Helper-dependent (HD canine adenovirus type 2 vectors (CAV-2 are well suited for this goal. These vectors are poorly immunogenic, efficiently transduce neurons, are retrogradely transported to afferent structures in the brain and lead to long-term transgene expression. CAV-2 vectors are being exploited to unravel behavior, cognition, neural networks, axonal transport and therapy for orphan diseases. With the goal of better understanding and characterizing HD-CAV-2 for brain therapy, we analyzed the transcriptomic modulation induced by HD-CAV-2 in human differentiated neurospheres derived from midbrain progenitors. This 3D model system mimics several aspects of the dynamic nature of human brain. We found that differentiated neurospheres are readily transduced by HD-CAV-2 and that transduction generates two main transcriptional responses: a DNA damage response and alteration of centromeric and microtubule probes. Future investigations on the biochemistry of processes highlighted by probe modulations will help defining the implication of HD-CAV-2 and CAR receptor binding in enchaining these functional pathways. We suggest here that the modulation of DNA damage genes is related to viral DNA, while the alteration of centromeric and microtubule probes is possibly enchained by the interaction of the HD-CAV-2 fibre with CAR.

  4. Role of Ctf3 and COMA subcomplexes in meiosis: Implication in maintaining Cse4 at the centromere and numeric spindle poles.

    Science.gov (United States)

    Agarwal, Meenakshi; Mehta, Gunjan; Ghosh, Santanu K

    2015-03-01

    During mitosis and meiosis, kinetochore, a conserved multi-protein complex, connects microtubule with the centromere and promotes segregation of the chromosomes. In budding yeast, central kinetochore complex named Ctf19 has been implicated in various functions and is believed to be made up of three biochemically distinct subcomplexes: COMA, Ctf3 and Iml3-Chl4. In this study, we aimed to identify whether Ctf3 and COMA subcomplexes have any unshared function at the kinetochore. Our data suggests that both these subcomplexes may work as a single functional unit without any unique functions, which we tested. Analysis of severity of the defects in the mutants suggests that COMA is epistatic to Ctf3 subcomplex. Interestingly, we noticed that these subcomplexes affect the organization of mitotic and meiotic kinetochores with subtle differences and they promote maintenance of Cse4 at the centromeres specifically during meiosis which is similar to the role of Mis6 (Ctf3 homolog) in fission yeast during mitosis. Interestingly, analysis of ctf3Δ and ctf19Δ mutants revealed a novel role of Ctf19 complex in regulation of SPB cohesion and duplication in meiosis.

  5. Mis17 is a regulatory module of the Mis6-Mal2-Sim4 centromere complex that is required for the recruitment of CenH3/CENP-A in fission yeast.

    Directory of Open Access Journals (Sweden)

    Yoshiharu Shiroiwa

    Full Text Available BACKGROUND: The centromere is the chromosome domain on which the mitotic kinetochore forms for proper segregation. Deposition of the centromeric histone H3 (CenH3, CENP-A is vital for the formation of centromere-specific chromatin. The Mis6-Mal2-Sim4 complex of the fission yeast S. pombe is required for the recruitment of CenH3 (Cnp1, but its function remains obscure. METHODOLOGY/PRINCIPAL FINDINGS: Mass spectrometry was performed on the proteins precipitated with Mis6- and Mis17-FLAG. The results together with the previously identified Sim4- and Mal2-TAP precipitated proteins indicated that the complex contains 12 subunits, Mis6, Sim4, Mal2, Mis15, Mis17, Cnl2, Fta1-4, Fta6-7, nine of which have human centromeric protein (CENP counterparts. Domain dissection indicated that the carboxy-half of Mis17 is functional, while its amino-half is regulatory. Overproduction of the amino-half caused strong negative dominance, which led to massive chromosome missegregation and hypersensitivity to the histone deacetylase inhibitor TSA. Mis17 was hyperphosphorylated and overproduction-induced negative dominance was abolished in six kinase-deletion mutants, ssp2 (AMPK, ppk9 (AMPK, ppk15 (Yak1, ppk30 (Ark1, wis4 (Ssk2, and lsk1 (P-TEFb. CONCLUSIONS: Mis17 may be a regulatory module of the Mis6 complex. Negative dominance of the Mis17 fragment is exerted while the complex and CenH3 remain at the centromere, a result that differs from the mislocalization seen in the mis17-362 mutant. The known functions of the kinases suggest an unexpected link between Mis17 and control of the cortex actin, nutrition, and signal/transcription. Possible interpretations are discussed.

  6. Dicentric chromosome aberration analysis using giemsa and centromere specific fluorescence in-situ hybridization for biological dosimetry: An inter- and intra-laboratory comparison in Indian laboratories.

    Science.gov (United States)

    Bhavani, M; Tamizh Selvan, G; Kaur, Harpreet; Adhikari, J S; Vijayalakshmi, J; Venkatachalam, P; Chaudhury, N K

    2014-09-01

    To facilitate efficient handling of large samples, an attempt towards networking of laboratories in India for biological dosimetry was carried out. Human peripheral blood samples were exposed to (60)Co γ-radiation for ten different doses (0-5Gy) at a dose rate of 0.7 and 2Gy/min. The chromosomal aberrations (CA) were scored in Giemsa-stained and fluorescence in-situ hybridization with centromere-specific probes. No significant difference (p>0.05) was observed in the CA yield for given doses except 4 and 5Gy, between the laboratories, among the scorers and also staining methods adapted suggest the reliability and validates the inter-lab comparisons exercise for triage applications.

  7. JAR细胞染色体着丝粒点变异的研究%Study on centromeric dots variation of JAR cells

    Institute of Scientific and Technical Information of China (English)

    曹波

    2013-01-01

    目的:探讨染色体着丝粒点(Cd)的变异与JAR细胞染色体非整倍性畸变的关系。方法肿瘤细胞株传代后用常规方法制备染色体,用直接法制备胚胎绒毛组织染色体标本,采用Cd-核仁组织区(NOR)同步银染分析技术研究JAR细胞染色体Cd的变异。结果 JAR细胞染色体Cd缺失率为1.33%、Cd迟滞复制率为1.23%、小Cd率为1.73%、Cd-NOR融合率为0.62%,与健康人胚胎绒毛细胞染色体Cd相比较,JAR细胞染色体Cd缺失、Cd迟滞复制显著升高(P<0.0125),而小Cd、Cd-NOR融合其差异无统计学意义。结论 JAR细胞染色体非整倍性畸变可能主要源于Cd缺失、Cd迟滞复制。%Objective To study the relation between the centromeric dots (Cd)variation of JAR cells and the chromosome aneuploidy aberration .Methods Chromosome of tumor cells were prepared with general method ,and chromosome of normal embryonic villi were prepared straightly .A Cd-NOR in-phase argentum dye analysis tech-nique modified by this laboratory was used to study centromeric dots (Cd) variation of JAR cells .Results Fre-quency of Cd loss in JAR cells was 1.33% .Frequency of Cd duplication laggard was 1.23% .Frequency of little Cd was 1.73% .Frequency of Cd-NOR amalgamation was 0.62% .Frequency of Cd loss and Cd duplication lag-gard in JAR cells were significantly higher than those of normal embryonic villi cells (P<0.0125) .Frequency of little Cd and Cd-NOR amalgamation showed no difference between JAR cells and normal embryonic villi .Conclu-sion Cd loss and Cd duplication laggard might be related with aneuploidy aberration of JAR cells .

  8. HSV-1 genome subnuclear positioning and associations with host-cell PML-NBs and centromeres regulate LAT locus transcription during latency in neurons.

    Directory of Open Access Journals (Sweden)

    Frédéric Catez

    Full Text Available Major human pathologies are caused by nuclear replicative viruses establishing life-long latent infection in their host. During latency the genomes of these viruses are intimately interacting with the cell nucleus environment. A hallmark of herpes simplex virus type 1 (HSV-1 latency establishment is the shutdown of lytic genes expression and the concomitant induction of the latency associated (LAT transcripts. Although the setting up and the maintenance of the latent genetic program is most likely dependent on a subtle interplay between viral and nuclear factors, this remains uninvestigated. Combining the use of in situ fluorescent-based approaches and high-resolution microscopic analysis, we show that HSV-1 genomes adopt specific nuclear patterns in sensory neurons of latently infected mice (28 days post-inoculation, d.p.i.. Latent HSV-1 genomes display two major patterns, called "Single" and "Multiple", which associate with centromeres, and with promyelocytic leukemia nuclear bodies (PML-NBs as viral DNA-containing PML-NBs (DCP-NBs. 3D-image reconstruction of DCP-NBs shows that PML forms a shell around viral genomes and associated Daxx and ATRX, two PML partners within PML-NBs. During latency establishment (6 d.p.i., infected mouse TGs display, at the level of the whole TG and in individual cells, a substantial increase of PML amount consistent with the interferon-mediated antiviral role of PML. "Single" and "Multiple" patterns are reminiscent of low and high-viral genome copy-containing neurons. We show that LAT expression is significantly favored within the "Multiple" pattern, which underlines a heterogeneity of LAT expression dependent on the viral genome copy number, pattern acquisition, and association with nuclear domains. Infection of PML-knockout mice demonstrates that PML/PML-NBs are involved in virus nuclear pattern acquisition, and negatively regulate the expression of the LAT. This study demonstrates that nuclear domains including

  9. Mitosis Phase Enrichment with Identification of Mitotic Centromere-Associated Kinesin As a Therapeutic Target in Castration-Resistant Prostate Cancer

    Science.gov (United States)

    Sircar, Kanishka; Huang, Heng; Hu, Limei; Liu, Yuexin; Dhillon, Jasreman; Cogdell, David; Aprikian, Armen; Efstathiou, Eleni; Navone, Nora; Troncoso, Patricia; Zhang, Wei

    2012-01-01

    The recently described transcriptomic switch to a mitosis program in castration-resistant prostate cancer (CRPC) suggests that mitotic proteins may be rationally targeted at this lethal stage of the disease. In this study, we showed upregulation of the mitosis-phase at the protein level in our cohort of 51 clinical CRPC cases and found centrosomal aberrations to also occur preferentially in CRPC compared with untreated, high Gleason–grade hormone-sensitive prostate cancer (P<0.0001). Expression profiling of chemotherapy-resistant CRPC samples (n = 25) was performed, and the results were compared with data from primary chemotherapy-naïve CRPC (n = 10) and hormone-sensitive prostate cancer cases (n = 108). Our results showed enrichment of mitosis-phase genes and pathways, with progression to both castration-resistant and chemotherapy-resistant disease. The mitotic centromere-associated kinesin (MCAK) was identified as a novel mitosis-phase target in prostate cancer that was overexpressed in multiple CRPC gene-expression datasets. We found concordant gene expression of MCAK between our parent and murine CRPC xenograft pairs and increased MCAK protein expression with clinical progression of prostate cancer to a castration-resistant disease stage. Knockdown of MCAK arrested the growth of prostate cancer cells suggesting its utility as a potential therapeutic target. PMID:22363599

  10. Mitosis phase enrichment with identification of mitotic centromere-associated kinesin as a therapeutic target in castration-resistant prostate cancer.

    Directory of Open Access Journals (Sweden)

    Kanishka Sircar

    Full Text Available The recently described transcriptomic switch to a mitosis program in castration-resistant prostate cancer (CRPC suggests that mitotic proteins may be rationally targeted at this lethal stage of the disease. In this study, we showed upregulation of the mitosis-phase at the protein level in our cohort of 51 clinical CRPC cases and found centrosomal aberrations to also occur preferentially in CRPC compared with untreated, high Gleason-grade hormone-sensitive prostate cancer (P<0.0001. Expression profiling of chemotherapy-resistant CRPC samples (n = 25 was performed, and the results were compared with data from primary chemotherapy-naïve CRPC (n = 10 and hormone-sensitive prostate cancer cases (n = 108. Our results showed enrichment of mitosis-phase genes and pathways, with progression to both castration-resistant and chemotherapy-resistant disease. The mitotic centromere-associated kinesin (MCAK was identified as a novel mitosis-phase target in prostate cancer that was overexpressed in multiple CRPC gene-expression datasets. We found concordant gene expression of MCAK between our parent and murine CRPC xenograft pairs and increased MCAK protein expression with clinical progression of prostate cancer to a castration-resistant disease stage. Knockdown of MCAK arrested the growth of prostate cancer cells suggesting its utility as a potential therapeutic target.

  11. Multilocus half-tetrad analysis and centromere mapping in citrus: evidence of SDR mechanism for 2n megagametophyte production and partial chiasma interference in mandarin cv 'Fortune'.

    Science.gov (United States)

    Cuenca, J; Froelicher, Y; Aleza, P; Juárez, J; Navarro, L; Ollitrault, P

    2011-10-01

    The genetic structure of 2n gametes and, particularly, the parental heterozygosity restitution at each locus depends on the meiotic process by which they originated, with first-division restitution and second-division restitution (SDR) being the two major mechanisms. The origin of 2n gametes in citrus is still controversial, although sexual polyploidisation is widely used for triploid seedless cultivar development. In this study, we report the analysis of 2n gametes of mandarin cv 'Fortune' by genotyping 171 triploid hybrids with 35 simple sequence repeat markers. The microsatellite DNA allele counting-peak ratios method for allele-dosage evaluation proved highly efficient in segregating triploid progenies and allowed half-tetrad analysis (HTA) by inferring the 2n gamete allelic configuration. All 2n gametes arose from the female genitor. The observed maternal heterozygosity restitution varied between 10 and 82%, depending on the locus, thus SDR appears to be the mechanism underlying 2n gamete production in mandarin cv 'Fortune'. A new method to locate the centromere, based on the best fit between observed heterozygosity restitution within a linkage group and theoretical functions under either partial or no chiasmata interference hypotheses was successfully applied to linkage group II. The maximum value of heterozygosity restitution and the pattern of restitution along this linkage group would suggest there is partial chiasma interference. The implications of such a restitution mechanism for citrus breeding are discussed.

  12. Historical perspectives on the discovery and elucidation of autoantibodies to centromere proteins (CENP) and the emerging importance of antibodies to CENP-F.

    Science.gov (United States)

    Fritzler, Marvin J; Rattner, Jerome B; Luft, LeeAnne M; Edworthy, Steven M; Casiano, Carlos A; Peebles, Carol; Mahler, Michael

    2011-02-01

    Autoantibodies to the centromere proteins (CENP), which are major constituents of the primary constriction of metaphase chromosomes, were first described in 1980. In those seminal publications and 30 years of research that have followed, a number of CENP have been identified as autoantibody targets in human diseases. Historically, autoantibodies directed to CENP-A, -B and -C have been considered relatively specific biomarkers for limited cutaneous systemic sclerosis (lcSSc) or the calcinosis, Raynaud's phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasia (CREST) syndrome. These autoantibodies, found in up to 40% of SSc sera, can be identified by indirect immunofluorescence (IIF) on a variety of tissue culture cell lines as a discrete speckled staining pattern of both interphase nuclei and metaphase chromatin. Early in the investigation of anti-CENP, it became apparent that some autoantibodies had a similar IIF pattern wherein as cells entered into the cell cycle, speckled staining of the metaphase chromatin could be observed but, unlike conventional CENP staining, interphase nuclei were not stained. Subsequent studies identified one of the targets of these autoantibodies to be CENP-F, a kinesin binding protein essential for completion of the cell cycle. Early clinical studies found that, unlike antibodies to the earlier described CENP, lcSSc rarely expressed anti-CENP-F and approximately 50% of these patients had a malignancy. This review provides a historical perspective of CENP autoantibodies and focuses on an update of the information on CENP-F and their clinical associations.

  13. High resolution linkage disequilibrium and haplotype maps for the genes in the centromeric region of chromo- some 15 in Tibetans and comparisons with Han population

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Genetic variations and their functional implications have been one of the focuses in recent genome research. With the release of the HapMap by the International Consortium, and the availability of the ultra-high-volume genotyping platform, it will soon be possible to use genome-wide association approach to identify genetic variations responsible for complex traits/diseases. While the power of this approach is generally agreed, it is a debated issue as to how much population difference should be exploited, and how best it should be applied. To address this issue we have sequenced 7 genes in the centromeric region of chromosome 15, investigated their SNPs, SNP frequencies, tagSNPs, LD structures, and haplotypes in 50 Tibetan subjects, and compared them with those from the Han population. Genetic diversities between the two populations were also quantified. Our results show that the overall genetic variation between the two populations is very little, but there are differences, primarily in allele frequencies, which is a dominating factor for haplotypes and tagSNPs. In general Tibetans have longer LD and less diversity in the region studied. These data provide genetic evidence for the close relationship between the two populations, and support the idea that all populations are fundamentally the same, but also indicate population variations, particularly in allele frequency, should be taken into account in complex traits/ diseases analysis. Data obtained in this investigation not only help us understand the genome region, but also provide road maps for variation study in the genes/ region in Tibetan population.

  14. The Inner Centromere Protein (INCENP) Coil Is a Single α-Helix (SAH) Domain That Binds Directly to Microtubules and Is Important for Chromosome Passenger Complex (CPC) Localization and Function in Mitosis.

    Science.gov (United States)

    Samejima, Kumiko; Platani, Melpomeni; Wolny, Marcin; Ogawa, Hiromi; Vargiu, Giulia; Knight, Peter J; Peckham, Michelle; Earnshaw, William C

    2015-08-28

    The chromosome passenger complex (CPC) is a master regulator of mitosis. Inner centromere protein (INCENP) acts as a scaffold regulating CPC localization and activity. During early mitosis, the N-terminal region of INCENP forms a three-helix bundle with Survivin and Borealin, directing the CPC to the inner centromere where it plays essential roles in chromosome alignment and the spindle assembly checkpoint. The C-terminal IN box region of INCENP is responsible for binding and activating Aurora B kinase. The central region of INCENP has been proposed to comprise a coiled coil domain acting as a spacer between the N- and C-terminal domains that is involved in microtubule binding and regulation of the spindle checkpoint. Here we show that the central region (213 residues) of chicken INCENP is not a coiled coil but a ∼ 32-nm-long single α-helix (SAH) domain. The N-terminal half of this domain directly binds to microtubules in vitro. By analogy with previous studies of myosin 10, our data suggest that the INCENP SAH might stretch up to ∼ 80 nm under physiological forces. Thus, the INCENP SAH could act as a flexible "dog leash," allowing Aurora B to phosphorylate dynamic substrates localized in the outer kinetochore while at the same time being stably anchored to the heterochromatin of the inner centromere. Furthermore, by achieving this flexibility via an SAH domain, the CPC avoids a need for dimerization (required for coiled coil formation), which would greatly complicate regulation of the proximity-induced trans-phosphorylation that is critical for Aurora B activation.

  15. Meiosis-specific cohesin component, Stag3 is essential for maintaining centromere chromatid cohesion, and required for DNA repair and synapsis between homologous chromosomes.

    Science.gov (United States)

    Hopkins, Jessica; Hwang, Grace; Jacob, Justin; Sapp, Nicklas; Bedigian, Rick; Oka, Kazuhiro; Overbeek, Paul; Murray, Steve; Jordan, Philip W

    2014-07-01

    Cohesins are important for chromosome structure and chromosome segregation during mitosis and meiosis. Cohesins are composed of two structural maintenance of chromosomes (SMC1-SMC3) proteins that form a V-shaped heterodimer structure, which is bridged by a α-kleisin protein and a stromal antigen (STAG) protein. Previous studies in mouse have shown that there is one SMC1 protein (SMC1β), two α-kleisins (RAD21L and REC8) and one STAG protein (STAG3) that are meiosis-specific. During meiosis, homologous chromosomes must recombine with one another in the context of a tripartite structure known as the synaptonemal complex (SC). From interaction studies, it has been shown that there are at least four meiosis-specific forms of cohesin, which together with the mitotic cohesin complex, are lateral components of the SC. STAG3 is the only meiosis-specific subunit that is represented within all four meiosis-specific cohesin complexes. In Stag3 mutant germ cells, the protein level of other meiosis-specific cohesin subunits (SMC1β, RAD21L and REC8) is reduced, and their localization to chromosome axes is disrupted. In contrast, the mitotic cohesin complex remains intact and localizes robustly to the meiotic chromosome axes. The instability of meiosis-specific cohesins observed in Stag3 mutants results in aberrant DNA repair processes, and disruption of synapsis between homologous chromosomes. Furthermore, mutation of Stag3 results in perturbation of pericentromeric heterochromatin clustering, and disruption of centromere cohesion between sister chromatids during meiotic prophase. These defects result in early prophase I arrest and apoptosis in both male and female germ cells. The meiotic defects observed in Stag3 mutants are more severe when compared to single mutants for Smc1β, Rec8 and Rad21l, however they are not as severe as the Rec8, Rad21l double mutants. Taken together, our study demonstrates that STAG3 is required for the stability of all meiosis-specific cohesin

  16. Meiosis-specific cohesin component, Stag3 is essential for maintaining centromere chromatid cohesion, and required for DNA repair and synapsis between homologous chromosomes.

    Directory of Open Access Journals (Sweden)

    Jessica Hopkins

    2014-07-01

    Full Text Available Cohesins are important for chromosome structure and chromosome segregation during mitosis and meiosis. Cohesins are composed of two structural maintenance of chromosomes (SMC1-SMC3 proteins that form a V-shaped heterodimer structure, which is bridged by a α-kleisin protein and a stromal antigen (STAG protein. Previous studies in mouse have shown that there is one SMC1 protein (SMC1β, two α-kleisins (RAD21L and REC8 and one STAG protein (STAG3 that are meiosis-specific. During meiosis, homologous chromosomes must recombine with one another in the context of a tripartite structure known as the synaptonemal complex (SC. From interaction studies, it has been shown that there are at least four meiosis-specific forms of cohesin, which together with the mitotic cohesin complex, are lateral components of the SC. STAG3 is the only meiosis-specific subunit that is represented within all four meiosis-specific cohesin complexes. In Stag3 mutant germ cells, the protein level of other meiosis-specific cohesin subunits (SMC1β, RAD21L and REC8 is reduced, and their localization to chromosome axes is disrupted. In contrast, the mitotic cohesin complex remains intact and localizes robustly to the meiotic chromosome axes. The instability of meiosis-specific cohesins observed in Stag3 mutants results in aberrant DNA repair processes, and disruption of synapsis between homologous chromosomes. Furthermore, mutation of Stag3 results in perturbation of pericentromeric heterochromatin clustering, and disruption of centromere cohesion between sister chromatids during meiotic prophase. These defects result in early prophase I arrest and apoptosis in both male and female germ cells. The meiotic defects observed in Stag3 mutants are more severe when compared to single mutants for Smc1β, Rec8 and Rad21l, however they are not as severe as the Rec8, Rad21l double mutants. Taken together, our study demonstrates that STAG3 is required for the stability of all meiosis

  17. Highly species-specific centromeric repetitive DNA sequences in lizards: molecular cytogenetic characterization of a novel family of satellite DNA sequences isolated from the water monitor lizard (Varanus salvator macromaculatus, Platynota).

    Science.gov (United States)

    Chaiprasertsri, Nampech; Uno, Yoshinobu; Peyachoknagul, Surin; Prakhongcheep, Ornjira; Baicharoen, Sudarath; Charernsuk, Saranon; Nishida, Chizuko; Matsuda, Yoichi; Koga, Akihiko; Srikulnath, Kornsorn

    2013-01-01

    Two novel repetitive DNA sequences, VSAREP1 and VSAREP2, were isolated from the water monitor lizard (Varanus salvator macromaculatus, Platynota) and characterized using molecular cytogenetics. The respective lengths and guanine-cytosine (GC) contents of the sequences were 190 bp and 57.5% for VSAREP1 and 185 bp and 59.7% for VSAREP2, and both elements were tandemly arrayed as satellite DNA in the genome. VSAREP1 and VSAREP2 were each located at the C-positive heterochromatin in the pericentromeric region of chromosome 2q, the centromeric region of chromosome 5, and 3 pairs of microchromosomes. This suggests that genomic compartmentalization between macro- and microchromosomes might not have occurred in the centromeric repetitive sequences of V. salvator macromaculatus. These 2 sequences did only hybridize to genomic DNA of V. salvator macromaculatus, but no signal was observed even for other squamate reptiles, including Varanus exanthematicus, which is a closely related species of V. salvator macromaculatus. These results suggest that these sequences were differentiated rapidly or were specifically amplified in the V. salvator macromaculatus genome.

  18. Centromere cohesion: regulating the guardian

    Institute of Scientific and Technical Information of China (English)

    Lin Fang; Guowei Fang

    2007-01-01

    @@ Mitosis, a process in which duplicated chromosomes are segregated into two daughter cells, is the most spectacular event in cell cycle. In mitosis, cells undergo drastic rearrangement of cytoskeleton, lipid and chromosomes under amazingly strict temporal and spatial control so that genetic information is faithfully transmitted to daughter cells.

  19. Increased frequency of dicentric chromosomes in therapy-related MDS and AML compared to de novo disease is significantly related to previous treatment with alkylating agents and suggests a specific susceptibility to chromosome breakage at the centromere.

    Science.gov (United States)

    Andersen, M K; Pedersen-Bjergaard, J

    2000-01-01

    Dicentric chromosomes are observed in many malignant diseases including myelodysplasia (MDS) and acute myeloid leukemia (AML) and have often been observed in a subset of these diseases, namely therapy-related MDS (t-MDS) and AML (t-AML). Using fluorescence in situ hybridization (FISH) with centromere-specific probes, we investigated the frequency and type of dicentric chromosomes in 180 consecutive patients with t-MDS and t-AML and in 231 consecutive patients with de novo MDS and AML, whose karyotypes had been studied previously by conventional G-banding. Twenty-seven out of 180 patients with t-MDS or t-AML presented dicentric chromosomes compared to only seven out of 231 patients with de novo disease (P = 0.00003). A dic(1q;7p) was observed in 10 cases, a dic(5p;17q) was observed in six cases, whereas various isodicentric chromosomes were observed in six cases. Excluding these six cases with isodicentrics, all 25 patients with dicentric chromosomes had involvement of at least one of the chromosome arms 1q, 5p, or 7p resulting in monosomy for 5q or 7q, and/or trisomy for 1q. Patients with dicentric chromosomes presented significantly more often as t-MDS compared to patients without dicentrics (P = 0.046), and the presence of a dicentric chromosome was significantly related to previous therapy with alkylating agents (P = 0.026). Thus, only one out of 27 patients with a dicentric chromosome had not previously received an alkylating agent. A specific susceptibility to breakage at the centromere after exposure to alkylating agents is suggested and may explain the frequent loss of whole chromosomes, in particular chromosomes 5 and 7 in t-MDS and t-AML, if the breaks are not followed by rejoining. Leukemia (2000) 14, 105-111.

  20. Multilocus half-tetrad analysis and centromere mapping in citrus: evidence of SDR mechanism for 2n megagametophyte production and partial chiasma interference in mandarin cv ‘Fortune'

    Science.gov (United States)

    Cuenca, J; Froelicher, Y; Aleza, P; Juárez, J; Navarro, L; Ollitrault, P

    2011-01-01

    The genetic structure of 2n gametes and, particularly, the parental heterozygosity restitution at each locus depends on the meiotic process by which they originated, with first-division restitution and second-division restitution (SDR) being the two major mechanisms. The origin of 2n gametes in citrus is still controversial, although sexual polyploidisation is widely used for triploid seedless cultivar development. In this study, we report the analysis of 2n gametes of mandarin cv ‘Fortune' by genotyping 171 triploid hybrids with 35 simple sequence repeat markers. The microsatellite DNA allele counting-peak ratios method for allele-dosage evaluation proved highly efficient in segregating triploid progenies and allowed half-tetrad analysis (HTA) by inferring the 2n gamete allelic configuration. All 2n gametes arose from the female genitor. The observed maternal heterozygosity restitution varied between 10 and 82%, depending on the locus, thus SDR appears to be the mechanism underlying 2n gamete production in mandarin cv ‘Fortune'. A new method to locate the centromere, based on the best fit between observed heterozygosity restitution within a linkage group and theoretical functions under either partial or no chiasmata interference hypotheses was successfully applied to linkage group II. The maximum value of heterozygosity restitution and the pattern of restitution along this linkage group would suggest there is partial chiasma interference. The implications of such a restitution mechanism for citrus breeding are discussed. PMID:21587302

  1. Synthetic Studies on Centromere-Associated Protein-E (CENP-E) Inhibitors: 2. Application of Electrostatic Potential Map (EPM) and Structure-Based Modeling to Imidazo[1,2-a]pyridine Derivatives as Anti-Tumor Agents.

    Science.gov (United States)

    Hirayama, Takaharu; Okaniwa, Masanori; Banno, Hiroshi; Kakei, Hiroyuki; Ohashi, Akihiro; Iwai, Kenichi; Ohori, Momoko; Mori, Kouji; Gotou, Mika; Kawamoto, Tomohiro; Yokota, Akihiro; Ishikawa, Tomoyasu

    2015-10-22

    To develop centromere-associated protein-E (CENP-E) inhibitors for use as anticancer therapeutics, we designed novel imidazo[1,2-a]pyridines, utilizing previously discovered 5-bromo derivative 1a. By site-directed mutagenesis analysis, we confirmed the ligand binding site. A docking model revealed the structurally important molecular features for effective interaction with CENP-E and could explain the superiority of the inhibitor (S)-isomer in CENP-E inhibition vs the (R)-isomer based on the ligand conformation in the L5 loop region. Additionally, electrostatic potential map (EPM) analysis was employed as a ligand-based approach to optimize functional groups on the imidazo[1,2-a]pyridine scaffold. These efforts led to the identification of the 5-methoxy imidazo[1,2-a]pyridine derivative (+)-(S)-12, which showed potent CENP-E inhibition (IC50: 3.6 nM), cellular phosphorylated histone H3 (p-HH3) elevation (EC50: 180 nM), and growth inhibition (GI50: 130 nM) in HeLa cells. Furthermore, (+)-(S)-12 demonstrated antitumor activity (T/C: 40%, at 75 mg/kg) in a human colorectal cancer Colo205 xenograft model in mice.

  2. 水稻双着丝粒染色体有性繁殖过程中的遗传稳定性%Centromere inactivation in a dicentric rice chromosome during sexual reproduction

    Institute of Scientific and Technical Information of China (English)

    龚志云; 刘秀秀; 张明亮; 薛超; 于恒秀; 裔传灯; 顾铭洪

    2013-01-01

    T0135是2009年从中籼3037第11染色体短臂端三体(2n+11S)自交后代中发现的形态变异株.选用第11号染色体着丝粒区域的特异分子细胞学标记5S rDNA进行染色体荧光原位杂交(fluorescent in situ hybridization,FISH)分析,发现T0135是第11染色体短臂双着丝粒染色体.为了研究该双着丝粒染色体有性繁殖过程中的遗传稳定性,对T0135有性繁殖后代进行FISH分析,观察后代中染色体类型并进行统计,发现后代染色体类型与亲本类型相同的占94%,即双着丝粒染色体能进行正常的减数分裂,通过着丝粒特异组蛋白(centromere specific histone H3,CENH3)免疫检测确定在双着丝粒染色体中只有一个有CENH3信号,说明该双着丝粒材料中只有一个行使正常功能的着丝粒,另外一个着丝粒处于失活状态.

  3. 乳腺癌组织中着丝粒蛋白CENP-B的表达及意义%Expression and significance of centromere protein B (CENP-B) in breast cancer

    Institute of Scientific and Technical Information of China (English)

    廖雯婷; 翁桂香; 冶亚平; 丁彦青; 王曦

    2012-01-01

    Purpose To investigate the expression level and clinicopathologic significance of centromere protein B ( CENP-B ) in breast cancer. Methods CENP-B mRNA and protein expressions in 4 pairs of breast cancer biopsies and their matched adjacent tissues were detected by RT-PCR, Q-PCR and Western blot, respectively. Immunohistochemincal analysis was used to examine the expression of CENP-B protein expressions in 53 cases of paraffin-embedded breast cancer tissues. The correlation between CENP-B expression with clinicopathologic features was analyzed by SPSS 10. 0. Results CENP-B was positively detected in 90. 6% ( 48/53 ) paraffin-embedded breast cancer tissues, and it was strongly expressed in 52. 8( 28/53 ) samples. The CENP-B protein level was higher in cancer cells than that in adjacent non-cancerous areas. Statistical analysis displayed that strong expression of CENP-B was significantly associated with clinical stage and T classification, while not with age, lymph node involvement and distant metastasis. Conclusion CENP-B may serve as a valuable molecular marker for prediction of breast cancer development and progression.%目的 检测乳腺癌组织中着丝粒蛋白CENP-B的表达,探讨CENP-B的表达与乳腺癌患者临床病理特征的关系及意义.方法 分别用RT-PCR、Q-PCR和Western blot法检测4例乳腺癌活检组织及相应癌旁组织中CENP-B mRNA和蛋白表达水平的差异;采用免疫组化SP法检测CENP-B在53例乳腺癌石蜡组织切片中的表达,并采用SPSS 10.0软件分析其与乳腺癌的临床病理特征的关系.结果 CENP-B mRNA和蛋白在4例乳腺癌组织中的表达高于癌旁组织;53例乳腺癌组织中CENP-B阳性率为90.6%(48/53),强阳性率为52.8%(28/53),CENP-B在乳腺肿瘤组织中的表达水平高于癌旁组织.统计分析结果发现CENP-B强阳性与临床分期、T分期密切相关.结论 CENP-B可能成为评价乳腺癌发生、发展的重要生物学指标.

  4. Donor Killer Immunoglobulin-Like Receptor Profile Bx1 Imparts a Negative Effect and Centromeric B-Specific Gene Motifs Render a Positive Effect on Standard-Risk Acute Myeloid Leukemia/Myelodysplastic Syndrome Patient Survival after Unrelated Donor Hematopoietic Stem Cell Transplantation.

    Science.gov (United States)

    Bao, Xiaojing; Wang, Miao; Zhou, Huifen; Zhang, Huanhuan; Wu, Xiaojin; Yuan, Xiaoni; Li, Yang; Wu, Depei; He, Jun

    2016-02-01

    Donor killer immunoglobulin-like receptor (KIR) group B profiles (Bx) and homozygous of centromeric motif B (Cen-B/B) are the most preferable KIR gene content motifs for hematopoietic stem cell transplantation (HSCT). The risk of transplant from Bx1 donors and the benefit of the presence of Cen-B (regardless of number) were observed for standard-risk acute myeloid leukemia/myelodysplastic syndrome (AML/MDS) patients in this 4-year retrospective study. A total of 210 Chinese patients who underwent unrelated donor HSCT were investigated. Donor KIR profile Bx was associated with significantly improved overall survival (OS; P = .026) and relapse-free survival (RFS; P = .021) and reduced nonrelapse mortality (NRM; P = .017) in AML/MDS patients. A significantly lower survival rate was observed for transplants from Bx1 donors compared with Bx2, Bx3, and Bx4 donors for patients in first complete remission (n = 82; OS: P = .024; RFS: P = .021). Transplant from donors with Cen-B resulted in improved OS (HR = .256; 95% CI, .084 to .774; P = .016) and RFS (HR = .252; 95% CI, .084 to .758; P = .014) in AML/MDS patients at standard risk. However, this particular effect did not increase with a higher number of Cen-B motifs (cB/B versus cA/B; OS: P = .755; RFS: P = .768). No effect was observed on high-risk AML/MDS, acute lymphoblastic leukemia/non-Hodgkin lymphoma, and chronic myelogenous leukemia patients. Avoiding the selection of HSCT donors of KIR profile Bx1 is strongly advisable for standard-risk AML/MDS patients. The presence of the Cen-B motif rather than its number was more important in donor selection for the Chinese population.

  5. A gene-protein assay for human epidermal growth factor receptor 2 (HER2: brightfield tricolor visualization of HER2 protein, the HER2 gene, and chromosome 17 centromere (CEN17 in formalin-fixed, paraffin-embedded breast cancer tissue sections

    Directory of Open Access Journals (Sweden)

    Nitta Hiroaki

    2012-05-01

    Full Text Available Abstract Background The eligibility of breast cancer patients for human epidermal growth factor receptor 2 (HER2-directed therapies is determined by the HER2 gene amplification and/or HER2 protein overexpression status of the breast tumor as determined by in situ hybridization (ISH or immunohistochemistry (IHC, respectively. Our objective was to combine the US Food and Drug Administration (FDA-approved HER2 & chromosome 17 centromere (CEN17 brightfield ISH (BISH and HER2 IHC assays into a single automated HER2 gene-protein assay allowing simultaneous detection of all three targets in a single tissue section. Methods The HER2 gene-protein assay was optimized using formalin-fixed, paraffin-embedded (FFPE samples of the xenograft tumors MCF7 [HER2 negative (non-amplified gene, protein negative] and Calu-3 [HER2 positive (amplified gene, protein positive]. HER2 IHC was performed using a rabbit monoclonal anti-HER2 antibody (clone 4B5 and a conventional 3,3'-diaminobenzidine IHC detection. The HER2 & CEN17 BISH signals were visualized using horseradish peroxidase-based silver and alkaline phosphatase-based red detection systems, respectively with a cocktail of 2,4-dinitrophenyl-labeled HER2 and digoxigenin-labeled CEN17 probes. The performance of the gene-protein assay on tissue microarray slides containing 189 randomly selected FFPE clinical breast cancer tissue cores was compared to that of the separate HER2 IHC and HER2 & CEN17 BISH assays. Results HER2 protein detection was optimal when the HER2 IHC protocol was used before (rather than after the BISH protocol. The sequential use of HER2 IHC and HER2 & CEN17 BISH detection steps on FFPE xenograft tumor sections appropriately co-localized the HER2 protein, HER2 gene, and CEN17 signals after mitigating the silver background staining by using a naphthol phosphate-containing hybridization buffer for the hybridization step. The HER2 protein and HER2 gene status obtained using the multiplex HER2 gene

  6. Down-regulated centromere protein-Ⅰ arrests cell growth at G2/M phase in human embryo kidney cells%CENP-Ⅰ表达下调对HEK293细胞生长及染色体分离的影响

    Institute of Scientific and Technical Information of China (English)

    袁泰先; 蔡燕; 彭乙华; 翁亚光; 施琼; 刘子杰; 刘斌; 李素彦

    2009-01-01

    Objective To construct the RNA interference eukaryotic expression vector targeting human centromere protein-Ⅰ (CENP-Ⅰ) and to observe its effect on the growth of human embryo kidney 293 cells ( HEK 293). Methods The expression vectors of pGenesil-1/CENP-Ⅰ-siRNA-1, pGenesil-1/CENP-Ⅰ-siRNA-2 and pGenesil-1/CENP-Ⅰ-siRNA-3 were constructed by gene recombination and then were transfected into the HEK293 cells by liposome. The expressions of CENP-Ⅰ at the protein and mRNA levels were detected by West-em blotting and fluorescence quality PCR (FQ-PCR). The effective vector and the best transfection time were selected. The growth and the cell cycle of the transfected cells were assessed by MTT assay and flow cytometry. Giemsa was used to stain the transfected cells to calculate the mitotic index. Results Sequence-specific siR-NAs targeting CENP-Ⅰ significantly down-regulated the expression of CENP-Ⅰ in HEK293 cells. The recombinant plasmid of pGenesil-1/CENP-Ⅰ-siRNA-3 was the effective vector. After transfecting for 72 h the best inhibited efficiency was achieved. In CENP-Ⅰ-siRNA transfected cells, the rate of cell growth was decreased markedly. Cells at G2/M phase and the mitotic index were increased conspicuous compared with the cells transfected with the blank vector or untransfected. Conclusion Down-regulation of CENP-Ⅰ in HEK293 cells by sequence spe-cific siRNA delays the cell growth and postpones the cell division.%目的 构建CENP-Ⅰ特异性RNA干扰真核表达载体,体外观察抑制CENP-Ⅰ基因表达对HEK293细胞生长、细胞周期和染色体数目异常率的影响.方法 构建CENP-Ⅰ siRNA真核表达载体,脂质体法将pGenesil-1空载体和pGene-sil-1/CENP-I-siRNA-1、pGenesil-1/CENP-Ⅰ-siRNA-2、pGenesil-1/CENP-Ⅰ-siRNA-3真核表达载体分别导入人胚肾HEK293细胞.转染后24、48、72 h收集细胞用Western blot和FQ-PCR检测HEK293细胞内CENP-Ⅰ蛋白及mRNA水平的表达情况,以确定干扰效果及

  7. The Finnish lapphund retinal atrophy locus maps to the centromeric region of CFA9

    Directory of Open Access Journals (Sweden)

    Sargan David R

    2007-07-01

    Full Text Available Abstract Background Dogs have the second largest number of genetic diseases, after humans. Among the diseases present in dogs, progressive retinal atrophy has been reported in more than a hundred breeds. In some of them, the mutation has been identified and genetic tests have allowed the identification of carriers, thus enabling a drastic reduction in the incidence of the disease. The Finnish lapphund is a dog breed presenting late-onset progressive retinal atrophy for which the disease locus remains unknown. Results In this study we mapped the progressive retinal atrophy locus in the Finnish lapphund using a DNA pooling approach, assuming that all affected dogs within the breed share the same identical-by descent-mutation as the cause of the disease (genetic homogeneity. Autosomal recessive inheritance was also assumed, after ruling out, from pedigree analysis, dominant and X-linked inheritance. DNA from 12 Finnish lapphund cases was mixed in one pool, and DNA from 12 first-degree relatives of these cases was mixed to serve as the control pool. The 2 pools were tested with 133 microsatellite markers, 3 of which showed a shift towards homozygosity in the cases. Individual genotyping with these 3 markers confirmed homozygosity for the GALK1 microsatellite only (chromosome 9. Further individual genotyping with additional samples (4 cases and 59 controls confirmed the association between this marker and the disease locus (p Conclusion The locus for progressive rod-cone degeneration is known to be close to the GALK1 locus, on the telomeric region of chromosome 9, where the retinal atrophy locus of the Finnish lapphund has been mapped. This suggests that the disease in this breed, as well as in the Swedish lapphund, may correspond to progressive rod-cone degeneration. This would increase the number of known dog breeds having this particular form of progressive retinal atrophy.

  8. The Finnish lapphund retinal atrophy locus maps to the centromeric region of CFA9

    OpenAIRE

    Sargan David R; Wickström Kaisa; Aguirre-Hernández Jesús

    2007-01-01

    Abstract Background Dogs have the second largest number of genetic diseases, after humans. Among the diseases present in dogs, progressive retinal atrophy has been reported in more than a hundred breeds. In some of them, the mutation has been identified and genetic tests have allowed the identification of carriers, thus enabling a drastic reduction in the incidence of the disease. The Finnish lapphund is a dog breed presenting late-onset progressive retinal atrophy for which the disease locus...

  9. The centromeric/nucleolar chromatin protein ZFP-37 may function to specify neuronal nuclear domains

    NARCIS (Netherlands)

    E. Payen; T. Verkerk (Ton); D. Michalovich (David); S.D. Dreyer; A. Winterpacht; B. Lee (Brendan); C.I. de Zeeuw (Chris); N.J. Galjart (Niels); F.G. Grosveld (Frank)

    1998-01-01

    textabstractMurine ZFP-37 is a member of the large family of C2H2 type zinc finger proteins. It is characterized by a truncated NH2-terminal Kruppel-associated box and is thought to play a role in transcriptional regulation. During development Zfp-37 mRNA is most abunda

  10. Identification of Drosophila centromere associated proteins by quantitative affinity purification-mass spectrometry

    Directory of Open Access Journals (Sweden)

    Teresa K. Barth

    2015-09-01

    The data accompanying the manuscript on this approach (Barth et al., 2015, Proteomics 14:2167-78, DOI: 10.1002/pmic.201400052 has been deposited to the ProteomeXchange Consortium (http://www.proteomexchange.org via the PRIDE partner repository with the dataset identifier PXD000758.

  11. MYEOV : A candidate gene for DNA amplification events occurring centromeric to CCNDI in breast cancer

    NARCIS (Netherlands)

    Janssen, JWG; Cuny, M; Orsetti, B; Rodriguez, C; Valles, H; Bartram, CR; Schuuring, E; Theillet, C

    2002-01-01

    Rearrangements of chromosome 11q13 are frequently observed in human cancer. The 11q13 region harbors several chromosomal breakpoint clusters found in hematologic malignancies and exhibits frequent DNA amplification in carcinomas. DNA amplification patterns in breast tumors are consistent with the ex

  12. A human homologue to the yeast omnipotent suppressor 45 maps 100 kb centromeric to HLA-A

    Energy Technology Data Exchange (ETDEWEB)

    Chauvel, B.; Dorval, I.; Fergelot, P. [CNRS, Faculte de Medecine, Rennes (France)] [and others

    1995-04-01

    Idipathic hemochromatosis is a common autosomal recessive inherited disorder of iron metabolism. The molecular defect is unknown. However, the gene responsible for the disease (HFE) has been localized on the short arm of chromosome 6. It is closely linked to the HLA class I genes and possibly within a 350 kilobase (kb) region around the HLA-A locus. In order to identify candidate genes for hemochromatosis, we applied a cDNA selection technique to isolate transcribed sequences encoded on yeast artificial chromosomes (YAC). At first, we screened a cDNA library derived from normal human duodenal mucosa with the YAC B30 H3. This YAC contains a 320 kb DNA insert including the HLA-A gene and spanning 150 kb of the 350 kb zone where the hemochromatosis gene is in linkage disequilibrium with restriction fragment length polymorphism (RFLP) markers. Preparation of the cDNA library of duodenal mucosa in Lambda Zap II phage and library screening with YAC B30 were carried out as previously described. In this way, we isolated seven non-HLA-A cDNAs corresponding to seven new genomic sequences. These potential genes were named hemochromatosis candidate gene (HCG) and numbered I to VII. The survey of all these cDNAs and their corresponding genomic sequences is in progress. In this work, we are especially interested in one of the seven non-HLA class I cDNA clones, named clone 58. 12 refs., 2 figs.

  13. The nuclear lamina promotes telomere aggregation and centromere peripheral localization during senescence of human mesenchymal stem cells

    NARCIS (Netherlands)

    Raz, Vered; Vermolen, Bart J.; Garini, Yuval; Onderwater, Jos J.M.; Mommaas-Kienhuis, Mieke A.; Koster, Abraham J.; Young, Ian T.; Tanke, Hans; Dirks, Roeland W.

    2008-01-01

    Ex vivo, human mesenchymal stem cells (hMSCs) undergo spontaneous cellular senescence after a limited number of cell divisions. Intranuclear structures of the nuclear lamina were formed in senescent hMSCs, which are identified by the presence of Hayflick-senescence-associated factors. Notably, spati

  14. The maternal nucleolus plays a key role in centromere satellite maintenance during the oocyte to embryo transition.

    Science.gov (United States)

    Fulka, Helena; Langerova, Alena

    2014-04-01

    The oocyte (maternal) nucleolus is essential for early embryonic development and embryos originating from enucleolated oocytes arrest at the 2-cell stage. The reason for this is unclear. Surprisingly, RNA polymerase I activity in nucleolus-less mouse embryos, as manifested by pre-rRNA synthesis, and pre-rRNA processing are not affected, indicating an unusual role of the nucleolus. We report here that the maternal nucleolus is indispensable for the regulation of major and minor satellite repeats soon after fertilisation. During the first embryonic cell cycle, absence of the nucleolus causes a significant reduction in major and minor satellite DNA by 12% and 18%, respectively. The expression of satellite transcripts is also affected, being reduced by more than half. Moreover, extensive chromosome bridging of the major and minor satellite sequences was observed during the first mitosis. Finally, we show that the absence of the maternal nucleolus alters S-phase dynamics and causes abnormal deposition of the H3.3 histone chaperone DAXX in pronuclei of nucleolus-less zygotes.

  15. 探讨染色体非整倍体形成与CENP变化的关系%Relationship between the variations of centromere protein and aneuploid

    Institute of Scientific and Technical Information of China (English)

    阎惠娜; 张月莲; 郑梅玲; 化爱玲; 张桂林; 贺江梅; 王晶晶

    2013-01-01

    目的 探讨子代染色体非整倍体形成与着丝粒蛋白(CENP)表达及其双亲染色体CENP表达的关系.方法 以染色体非整倍体流产绒毛为研究组,以染色体正常绒毛为对照组.应用染色体G显带技术及免疫组化技术,对每例标本及其流产夫妇双方进行染色体核型分析及CENP检测.结果 ①染色体正常夫妇的流产绒毛的染色体非整倍体发生率为44.78%,在非整倍体中性染色体异常发生率最高,占40.00%,其次21-三体综合征,占23.33%,②两组流产绒毛染色体中均有CENP的表达,且研究组显著低于对照组(t=3.012,P<0.05).研究组非整倍体绒毛染色体中CENP含量与其父方外周血染色体的比较有显著性差异,前者显著低于后者(t=4.279,P<0.05),而与母方的差异无统计学意义(t=1.927,P>0.05).对照组正常绒毛与其父方外周血染色体CENP含量比较有显著性差异,前者CENP含量显著低于后者(t =3.008,P<0.05),而与母方的差异无统计学意义(t=1.234 P>0.05).结论 CENP含量降低可能是染色体着丝粒不分离造成非整倍体形成的原因之一,且CENP的含量与其母亲具有一定的同源性、相似性.%Objectives:In order to explore the relation between embryos of aneuploid occurrence and their parents,this paper investigated the variations of CENPs of them.Methods:there are two groups,the test group are chromosome of aneuploid embryos which were been diagnosed had stopped growth and the chromosome of their parents,the compare group are the embryos which are chromosome of normal and the chromosome of their parents,The peripheral blood lymphocyte chromosome specimens were made with the routine chromosomal technique,G-band were developed.useing the technique of immunhitehmisty to get the CENPs.all the cases have whole data of G-band technique.Results:① The rate of aneuploid chorion chromosome in study group was 44.78%,Among 30 cases with karyotype disorder,12 cases of sex chromosome take 40.00% of the cases.7 cases of 21-trisomy syndrome take 23.33% of the cases.②The test group and the compare group chromosome both have CENPS.And the CENPS of the test group was much lower than the compare group.t =3.012 P < 0.05).the CENPS of the test group was much lower then their paternal group (t =4.279 P <0.001),No significant difference between the test group and their maternal group.(t =1.927 P >0.05) ; The CENPS of the compare group was much lower then their paternal group (t =3.008 P < 0.05),No significant difference between the test group and their maternal group.(t =1.234 P > 0.05).Conclusions:Less CENPS are closely related to aneuploid occurrence,and contents of CENPS may come from the maternal group,in order to offer evidences of the genetics between descendants and their mothers

  16. Inherited variants in the inner centromere protein (INCENP) gene of the chromosomal passenger complex contribute to the susceptibility of ER-negative breast cancer

    Science.gov (United States)

    Kabisch, Maria; Lorenzo Bermejo, Justo; Dünnebier, Thomas; Ying, Shibo; Michailidou, Kyriaki; Bolla, Manjeet K.; Wang, Qin; Dennis, Joe; Shah, Mitul; Perkins, Barbara J.; Czene, Kamila; Darabi, Hatef; Eriksson, Mikael; Bojesen, Stig E.; Nordestgaard, Børge G.; Nielsen, Sune F.; Flyger, Henrik; Lambrechts, Diether; Neven, Patrick; Peeters, Stephanie; Weltens, Caroline; Couch, Fergus J.; Olson, Janet E.; Wang, Xianshu; Purrington, Kristen; Chang-Claude, Jenny; Rudolph, Anja; Seibold, Petra; Flesch-Janys, Dieter; Peto, Julian; dos-Santos-Silva, Isabel; Johnson, Nichola; Fletcher, Olivia; Nevanlinna, Heli; Muranen, Taru A.; Aittomäki, Kristiina; Blomqvist, Carl; Schmidt, Marjanka K.; Broeks, Annegien; Cornelissen, Sten; Hogervorst, Frans B.L.; Li, Jingmei; Brand, Judith S.; Humphreys, Keith; Guénel, Pascal; Truong, Thérèse; Menegaux, Florence; Sanchez, Marie; Burwinkel, Barbara; Marmé, Frederik; Yang, Rongxi; Bugert, Peter; González-Neira, Anna; Benitez, Javier; Pilar Zamora, M.; Arias Perez, Jose I.; Cox, Angela; Cross, Simon S.; Reed, Malcolm W.R.; Andrulis, Irene L.; Knight, Julia A.; Glendon, Gord; Tchatchou, Sandrine; Sawyer, Elinor J.; Tomlinson, Ian; Kerin, Michael J.; Miller, Nicola; Haiman, Christopher A.; Schumacher, Fredrick; Henderson, Brian E.; Le Marchand, Loic; Lindblom, Annika; Margolin, Sara; Hooning, Maartje J.; Hollestelle, Antoinette; Kriege, Mieke; Koppert, Linetta B.; Hopper, John L.; Southey, Melissa C.; Tsimiklis, Helen; Apicella, Carmel; Slettedahl, Seth; Toland, Amanda E.; Vachon, Celine; Yannoukakos, Drakoulis; Giles, Graham G.; Milne, Roger L.; McLean, Catriona; Fasching, Peter A.; Ruebner, Matthias; Ekici, Arif B.; Beckmann, Matthias W.; Brenner, Hermann; Dieffenbach, Aida K.; Arndt, Volker; Stegmaier, Christa; Ashworth, Alan; Orr, Nicholas; Schoemaker, Minouk J.; Swerdlow, Anthony; García-Closas, Montserrat; Figueroa, Jonine; Chanock, Stephen J.; Lissowska, Jolanta; Goldberg, Mark S.; Labrèche, France; Dumont, Martine; Winqvist, Robert; Pylkäs, Katri; Jukkola-Vuorinen, Arja; Grip, Mervi; Brauch, Hiltrud; Brüning, Thomas; Ko, Yon-Dschun; Radice, Paolo; Peterlongo, Paolo; Scuvera, Giulietta; Fortuzzi, Stefano; Bogdanova, Natalia; Dörk, Thilo; Mannermaa, Arto; Kataja, Vesa; Kosma, Veli-Matti; Hartikainen, Jaana M.; Devilee, Peter; Tollenaar, Robert A.E.M.; Seynaeve, Caroline; Van Asperen, Christi J.; Jakubowska, Anna; Lubinski, Jan; Jaworska-Bieniek, Katarzyna; Durda, Katarzyna; Zheng, Wei; Shrubsole, Martha J.; Cai, Qiuyin; Torres, Diana; Anton-Culver, Hoda; Kristensen, Vessela; Bacot, François; Tessier, Daniel C.; Vincent, Daniel; Luccarini, Craig; Baynes, Caroline; Ahmed, Shahana; Maranian, Mel; Simard, Jacques; Chenevix-Trench, Georgia; Hall, Per; Pharoah, Paul D.P.; Dunning, Alison M.; Easton, Douglas F.; Hamann, Ute

    2015-01-01

    The chromosomal passenger complex (CPC) plays a pivotal role in the regulation of cell division. Therefore, inherited CPC variability could influence tumor development. The present candidate gene approach investigates the relationship between single nucleotide polymorphisms (SNPs) in genes encoding key CPC components and breast cancer risk. Fifteen SNPs in four CPC genes (INCENP, AURKB, BIRC5 and CDCA8) were genotyped in 88 911 European women from 39 case-control studies of the Breast Cancer Association Consortium. Possible associations were investigated in fixed-effects meta-analyses. The synonymous SNP rs1675126 in exon 7 of INCENP was associated with overall breast cancer risk [per A allele odds ratio (OR) 0.95, 95% confidence interval (CI) 0.92–0.98, P = 0.007] and particularly with estrogen receptor (ER)-negative breast tumors (per A allele OR 0.89, 95% CI 0.83–0.95, P = 0.0005). SNPs not directly genotyped were imputed based on 1000 Genomes. The SNPs rs1047739 in the 3ʹ untranslated region and rs144045115 downstream of INCENP showed the strongest association signals for overall (per T allele OR 1.03, 95% CI 1.00–1.06, P = 0.0009) and ER-negative breast cancer risk (per A allele OR 1.06, 95% CI 1.02–1.10, P = 0.0002). Two genotyped SNPs in BIRC5 were associated with familial breast cancer risk (top SNP rs2071214: per G allele OR 1.12, 95% CI 1.04–1.21, P = 0.002). The data suggest that INCENP in the CPC pathway contributes to ER-negative breast cancer susceptibility in the European population. In spite of a modest contribution of CPC-inherited variants to the total burden of sporadic and familial breast cancer, their potential as novel targets for breast cancer treatment should be further investigated. PMID:25586992

  17. Inherited variants in the inner centromere protein (INCENP) gene of the chromosomal passenger complex contribute to the susceptibility of ER-negative breast cancer

    NARCIS (Netherlands)

    M. Kabisch (Maria); J.L. Bermejo (Justo Lorenzo); T. Dun̈nebier (Thomas); S. Ying (Shibo); K. Michailidou (Kyriaki); M.K. Bolla (Manjeet); Q. Wang (Qing); J. Dennis (Joe); M. Shah (Mitul); B. Perkins (Barbara); K. Czene (Kamila); H. Darabi (Hatef); M. Eriksson (Mikael); S.E. Bojesen (Stig); B.G. Nordestgaard (Børge); S.F. Nielsen (Sune); H. Flyger (Henrik); D. Lambrechts (Diether); P. Neven (Patrick); S.T.H. Peeters (Stephanie); C. Weltens (Caroline); F.J. Couch (Fergus); J.E. Olson (Janet); X. Wang (Xianshu); K. Purrington (Kristen); J. Chang-Claude (Jenny); A. Rudolph (Anja); P. Seibold (Petra); D. Flesch-Janys (Dieter); J. Peto (Julian); I. dos Santos Silva (Isabel); N. Johnson (Nichola); O. Fletcher (Olivia); H. Nevanlinna (Heli); T.A. Muranen (Taru); K. Aittomäki (Kristiina); C. Blomqvist (Carl); M.K. Schmidt (Marjanka); A. Broeks (Annegien); S. Cornelissen (Sten); F.B.L. Hogervorst (Frans); J. Li (Jingmei); J.S. Brand (Judith S.); M.K. Humphreys (Manjeet); P. Guénel (Pascal); T. Truong (Thérèse); F. Menegaux (Florence); M. Sanchez (Marie); B. Burwinkel (Barbara); F. Marme (Federick); R. Yang (Rongxi); P. Bugert (Peter); A. González-Neira (Anna); J. Benítez (Javier); M.P. Zamora (Pilar); J.I. Arias Pérez (José Ignacio); A. Cox (Angela); S.S. Cross (Simon); M.W.R. Reed (Malcolm); I.L. Andrulis (Irene); J.A. Knight (Julia); G. Glendon (Gord); S. Tchatchou (Sandrine); E.J. Sawyer (Elinor); I.P. Tomlinson (Ian); M. Kerin (Michael); N. Miller (Nicola); C.A. Haiman (Christopher); F.R. Schumacher (Fredrick); B.E. Henderson (Brian); L. Le Marchand (Loic); A. Lindblom (Annika); S. Margolin (Sara); M.J. Hooning (Maartje); A. Hollestelle (Antoinette); M. Kriege (Mieke); L.B. Koppert (Linetta); J. Hopper (John); M.C. Southey (Melissa); H. Tsimiklis (Helen); C. Apicella (Carmel); S. Slettedahl (Seth); A.E. Toland (Amanda); C. Vachon (Celine); D. Yannoukakos (Drakoulis); G.G. Giles (Graham); R.L. Milne (Roger); C.A. McLean (Catriona Ann); P.A. Fasching (Peter); M. Ruebner (Matthias); A.B. Ekici (Arif); M.W. Beckmann (Matthias); H. Brenner (Hermann); A.K. Dieffenbach (Aida Karina); V. Arndt (Volker); C. Stegmaier (Christa); A. Ashworth (Alan); N. Orr (Nick); M. Schoemaker (Minouk); A.J. Swerdlow (Anthony ); M. García-Closas (Montserrat); J.D. Figueroa (Jonine); S.J. Chanock (Stephen); J. Lissowska (Jolanta); M.S. Goldberg (Mark); F. Labrèche (France); M. Dumont (Martine); R. Winqvist (Robert); K. Pykäs (Katri); A. Jukkola-Vuorinen (Arja); M. Grip (Mervi); H. Brauch (Hiltrud); T. Brüning (Thomas); Y-D. Ko (Yon-Dschun); P. Radice (Paolo); P. Peterlongo (Paolo); G. Scuvera (Giulietta); S. Fortuzzi (S.); N.V. Bogdanova (Natalia); T. Dörk (Thilo); A. Mannermaa (Arto); V. Kataja (Vesa); V-M. Kosma (Veli-Matti); J.M. Hartikainen (J.); P. Devilee (Peter); R.A.M. Tollenaar (Robert A.M.); C.M. Seynaeve (Caroline); C.J. van Asperen (Christi); A. Jakubowska (Anna); J. Lubinski (Jan); K. Jaworska-Bieniek (Katarzyna); K. Durda (Katarzyna); W. Zheng (Wei); M. Shrubsole (Martha); Q. Cai (Qiuyin); D. Torres (Diana); H. Anton-Culver (Hoda); V. Kristensen (Vessela); F. Bacot (Francois); D.C. Tessier (Daniel C.); D. Vincent (Daniel); C. Luccarini (Craig); C. Baynes (Caroline); S. Ahmed (Shahana); M. Maranian (Melanie); J. Simard (Jacques); G. Chenevix-Trench (Georgia); P. Hall (Per); P.D.P. Pharoah (Paul); A.M. Dunning (Alison); D.F. Easton (Douglas); U. Hamann (Ute)

    2014-01-01

    textabstractThe chromosomal passenger complex (CPC) plays a pivotal role in the regulation of cell division. Therefore, inherited CPC variability could influence tumor development. The present candidate gene approach investigates the relationship between single nucleotide polymorphisms (SNPs) in gen

  18. Inherited variants in the inner centromere protein (INCENP) gene of the chromosomal passenger complex contribute to the susceptibility of ER-negative breast cancer

    DEFF Research Database (Denmark)

    Kabisch, Maria; Lorenzo Bermejo, Justo; Dünnebier, Thomas

    2015-01-01

    The chromosomal passenger complex (CPC) plays a pivotal role in the regulation of cell division. Therefore, inherited CPC variability could influence tumor development. The present candidate gene approach investigates the relationship between single nucleotide polymorphisms (SNPs) in genes encodi...

  19. CENP-A chromatin disassembly in stressed and senescent murine cells

    Science.gov (United States)

    Hédouin, Sabrine; Grillo, Giacomo; Ivkovic, Ivana; Velasco, Guillaume; Francastel, Claire

    2017-01-01

    Centromeres are chromosomal domains essential for genomic stability. We report here the remarkable transcriptional and epigenetic perturbations at murine centromeres in genotoxic stress conditions. A strong and selective transcriptional activation of centromeric repeats is detected within hours. This is followed by disorganization of centromeres with striking delocalization of nucleosomal CENP-A, the key determinant of centromere identity and function, in a mechanism requiring active transcription of centromeric repeats, the DNA Damage Response (DDR) effector ATM and chromatin remodelers/histone chaperones. In the absence of p53 checkpoint, activated transcription of centromeric repeats and CENP-A delocalization do not occur and cells accumulate micronuclei indicative of genomic instability. In addition, activated transcription and loss of centromeres identity are features of permanently arrested senescent cells with persistent DDR activation. Together, these findings bring out cooperation between DDR effectors and loss of centromere integrity as a safeguard mechanism to prevent genomic instability in context of persistent DNA damage signalling. PMID:28186195

  20. Heterogeneous clinical presentation in ICF syndrome: correlation with underlying gene defects.

    NARCIS (Netherlands)

    Weemaes, C.M.R.; Tol, M.J. van; Wang, J.; Ostaijen-ten Dam, M.M. van; Eggermond, M.C. van; Thijssen, P.E.; Aytekin, C.; Brunetti-Pierri, N.; Burg, M. van der; aham Davies, E. Gr; Ferster, A.; Furthner, D.; Gimelli, G.; Gennery, A.; Kloeckener-Gruissem, B.; Meyn, S.; Powell, C.; Reisli, I.; Schuetz, C.; Schulz, A.; Shugar, A.; Elsen, P.J. van den; Maarel, S.M. van der

    2013-01-01

    Immunodeficiency with centromeric instability and facial anomalies (ICF) syndrome is a primary immunodeficiency, predominantly characterized by agammaglobulinemia or hypoimmunoglobulinemia, centromere instability and facial anomalies. Mutations in two genes have been discovered to cause ICF syndrome

  1. A new stable human dicentric chromosome, tdic(4;21)(p16;q22), in a woman with first trimester abortion.

    OpenAIRE

    Wang, F.; Li, Y.

    1993-01-01

    A woman with first trimester abortion and a dicentric chromosome formed from a 4 and a 21 is described. The dicentric chromosome was stable and in the majority of cells the 21 centromere was active, while in a minority the chromosome 4 centromere was active. This shows that both centromeres were functional, but that only one functioned in any given cell. Suppression of the activity of one centromere might be the mechanism by which this dicentric chromosome achieved its stability. A dicentric ...

  2. Klinefelter综合征及双亲X染色体着丝粒区α-卫星DNA变异研究%X chromosome centromeric alpha satellite DNA variation in Klinefelter's syndrome patients and their parents

    Institute of Scientific and Technical Information of China (English)

    李恬; 翁亚光; 王应雄; 何俊琳; 刘学庆; 陈雪梅

    2005-01-01

    目的:本文从DNA分子水平对Klinifelter综合征患者及双亲X染色体着丝粒区域的α-卫星DNA变异进行研究,探讨Klinefelter综合征患者X染色体不分离形成的原因.方法:采用(representative sampling of multiple repetitive units,rep)PCR方法,通过设计适当的引物,对-卫星DNA多个拷贝串联重复序列同时进行扩增,检测患者、双亲和正常人重复序列中变异发生.结果:(1)扩增产物中分别出现1774bp、570bp(由缺失产生)片段和1410bp、583bp(由缺失产生)、220bp(由缺失产生)片段,(2)患者组570bp和583bp产生率高于正常组,x2统计有显著性意义(P<0.05).(3)患者与患者双亲比较差异不显著.结论:Klinefelter综合征患者及双亲的X染色体着丝粒区α-卫星DNA表现出较正常人具更多的变异.提示通过孕前或产前对双亲外周血X染色体着丝粒区域的α-卫星DNA变异分析,对Klinefelter综合征发生风险进行估计,为产前绒毛或羊水染色体检查提供重要的依据.

  3. Comparison of BAC FISH with specific telomeres and centromere probes and chromosome painting on detection of chromosome translocation induced by irradiation%BAC FISH与PAINT法检测辐射诱发染色体易位的比较

    Institute of Scientific and Technical Information of China (English)

    刘青杰; 陆雪; 封江彬; 王晓维; 陈德清

    2008-01-01

    Objective To compare the efficiency of BAC FISH established in our lab and conventional chromosome painting(PAINT)on detection of radiation-induced chromosome translocation.Methods Healthy human peripheral blood samples were irradiated with 0~5.0 Gy 60Co γ-rays.Then chromosome translocations in these samples were detected with BAC FISH and PAINT using chromosomes 1.2 and 4.The genome translocation rates were calculated with observed chromosome translocation rates,and the dose-response curve of two methods were established.Results The genome translocation rates induced by 0~5.0 Gy 60Co γ-rays detected by BAC FISH and PAINT were increased with absorbed doses.The observed translocation rates with BAC FISH were higher than that with PAINT at each dose level.The dose-response curve were Y=0.043 D2+0.0008D+0.0048 for BAC FISH and Y=0.043D2+0.006D+0.0027 for PAINT.Conclusions The translocation rate detected by BAC FISH was higher than that by PAINT,and the parameters β in dose-response curve equation were same by two methods.%目的 比较自行建立的BAC FISH方法和常规染色体涂染(PAINT)方法分析辐射诱发染色体易位有效性的不同.方法 对正常人外周血照射不同剂量(0~5.0 Gy)的60Co γ射线,用1、2和4号染色体特异性端粒和着丝粒探针BAC FISH及PAINT分析染色体易位,将观察到的染色体易位率换算为全基因组易位率,并建立两种方法分析辐射诱发的染色体易位率剂量-效应曲线.结果 用两种方法分析0~5.0 Gy 60Coγ射线诱发的全基因组易位率均随着吸收剂量的增加而增高;在相同的剂量点,BAC FISH染色体易位检出率高于PAINT方法.两种方法分析吸收剂量和全基因组易位率之间的剂量-效应曲线均为二次方程模式,分别为Y=0.043D2+0.0008D+0.0048(BAC FISH)和Y=0.043D2+0.006D+0.0027(PAINT).结论 自行建立的BAC FISH方法分析辐射诱发染色体易位检出率高于常规染色体涂染方法,两种方法建立的剂量-效应曲线方程的β系数相同.

  4. Isolation and characterization of centromere RCS1 homology sequence from Triticum boeoticum%野生一粒小麦着丝粒RCS1相关序列的克隆鉴定

    Institute of Scientific and Technical Information of China (English)

    陈凡国; 封德顺; 夏光敏

    2003-01-01

    用水稻着丝粒重复序列RCS1为探针,与3072个克隆进行菌落杂交,得到了32个阳性克隆,用RCS1与拟斯卑尔脱山羊草着丝粒重复序列Tcs250为探针进一步筛选,在32个RCS1相关的阳性克隆中任选10个克隆进行点杂交,分别有6个和 5个阳性克隆.为了克隆RCS1相关片段,依据RCS1的序列设计了三对引物,将引物3从上述阳性克隆中扩增的一个543 bp的片段克隆测序,发现与水稻RCS1部分片段达到约83%的同源,与大麦的反转座子(Ty3/gypsy)部分序列同源性达到了92%,与节节麦中着丝粒的整合酶基因部分序列同源性达到了96%,命名为TBRCS1.TBRCS1可能是野生一粒小麦着丝粒区的组成部分.

  5. Exact break point of a 50 kb deletion 8 kb centromeric of the HLA-A locus with HLA-A*24:02: the same deletion observed in other A*24 alleles and A*23:01 allele.

    Science.gov (United States)

    Mitsunaga, Shigeki; Okudaira, Yuko; Kunii, Nanae; Cui, Tailin; Hosomichi, Kazuyoshi; Oka, Akira; Suzuki, Yasuo; Homma, Yasuhiko; Sato, Shinji; Inoue, Ituro; Inoko, Hidetoshi

    2011-08-01

    In a structural aberration analysis of patients with arthritis mutilans, a 50 kb deletion near the HLA-A locus with HLA-A*24:02 allele was detected. It was previously reported that HLA-A*24:02 haplotype harbored a large-scale deletion telomeric of the HLA-A gene in healthy individuals. In order to confirm that the deletion are the same in patients with arthritis mutilans and in healthy individuals, and to identify the break point of this deletion, the boundary sequences across the deletion in A*24:02 was amplified by polymerase chain reaction (PCR) as a 3.7 kb genomic fragment and subjected to nucleotide sequence determination. A comparison of these genomic sequences with those of the non-A*24:02 haplotype revealed that the deleted genomic region spanning 50 kb was flanked by 3.7 kb repetitive element-rich segments homologous to each other on both sides in non-A*24. The nucleotide sequences of the PCR products were identical in patients with arthritis mutilans and in healthy individuals, revealing that the deletion linked to A*24:02 is irrelevant to the onset of arthritis mutilans. The deletion was detected in all other A*24 alleles so far examined but not in other HLA-A alleles, except A*23:01. This finding, along with the phylogenic tree of HLA-A alleles and the presence of the 3.7 kb highly homologous segments at the boundary of the deleted genomic region in A*03 and A*32, may suggest that this HLA-A*24:02-linked deletion was generated by homologous recombination within two 3.7 kb homologous segments situated 50 kb apart in the ancestral A*24 haplotype after divergence from the A*03 and A*32 haplotypes.

  6. Reconstruction of the kinetochore: a prelude to meiosis

    Directory of Open Access Journals (Sweden)

    Haraguchi Tokuko

    2007-06-01

    Full Text Available Abstract In eukaryotic organisms, chromosomes are spatially organized within the nucleus. Such nuclear architecture provides a physical framework for the genetic activities of chromosomes, and changes its functional organization as the cell moves through the phases of the cell cycle. The fission yeast Schizosaccharomyces pombe provides a striking example of nuclear reorganization during the transition from mitosis to meiosis. In this organism, centromeres remain clustered at the spindle-pole body (SPB; a centrosome-equivalent structure in fungi during mitotic interphase. In contrast, during meiotic prophase, centromeres dissociate from the SPB and telomeres cluster to the SPB. Recent studies revealed that this repositioning of chromosomes is regulated by mating pheromone signaling. Some centromere proteins disappear from the centromere in response to mating pheromone, leading to dissociation of centromeres from the SPB. Interestingly, mating pheromone signaling is also required for monopolar orientation of the kinetochore which is crucial for proper segregation of sister chromatids during meiosis. When meiosis is induced in the absence of mating pheromone signaling, aberrant chromosome behaviors are observed: the centromere proteins remain at the centromere; the centromere remains associated with the SPB; and sister chromatids segregate precociously in the first meiotic division. These aberrant chromosome behaviors are all normalized by activating the mating pheromone signaling pathway. Thus, action of mating pheromone on the centromere is important for coherent behavior of chromosomes in meiosis. Here we discuss repositioning and reconstruction of the centromere during the transition from mitosis to meiosis, and highlight its significance for proper progression of meiosis.

  7. Allelic imbalance and cytogenetic deletion of 1p in colorectal adenomas: a target region identified between DIS199 and DIS234

    DEFF Research Database (Denmark)

    Bomme, L; Heim, S; Bardi, G;

    1998-01-01

    centromere 1 signals were invariably found. In the cases hybridized with the 1p-telomeric probe, we found the same frequencies of telomeric and centromeric signals, in agreement with the interpretation that the deletions were interstitial. One of the 53 adenomas had genomic instability, seen as new alleles...

  8. [A simple method to correct genetic distance between linked genes and a correction of calculating data in tetrad analysis in Neurospora crassa].

    Science.gov (United States)

    Li, You-Yong; Zhao, Yuan-Zeng

    2003-05-01

    The present paper is dealing with a simple method to correct the distance of linked genes in tetrad analysis in Neurospora crassa. It is suggested that the data of 4-thread double crossing over should be added in two single crossing over respectively in centromere maping when calculating crossover value of the two genes locating across the centromere.

  9. Environmental lead exposure increases micronuclei in children

    DEFF Research Database (Denmark)

    Kapka, Lucyna; Baumgartner, Adolf; Siwińska, Ewa

    2007-01-01

    and control group (standard MN test: 2.96 +/- 2.36 versus 1.16 +/- 1.28; FISH technique: 3.57 +/- 3.02 versus 1.43 +/- 1.69, respectively). The frequencies of both centromere-positive (C+MN) and centromere-negative (C-MN) micronuclei were significantly increased in exposed children; however, the contribution...

  10. Yeast Interacting Proteins Database: YOR026W, YGR188C [Yeast Interacting Proteins Database

    Lifescience Database Archive (English)

    Full Text Available the presence of spindle damage, associates with centromere DNA via Skp1p Rows with this prey as prey (1) Row...ase in the presence of spindle damage, associates with centromere DNA via Skp1p R

  11. Expression of regulators of mitotic fidelity are associated with intercellular heterogeneity and chromosomal instability in primary breast cancer

    DEFF Research Database (Denmark)

    Roylance, Rebecca; Endesfelder, David; Jamal-Hanjani, Mariam

    2014-01-01

    , as determined by centromeric FISH and defined by modal centromere deviation, was analysed. Significantly poorer clinical outcome was observed in patients with high AURKA expression levels. Expression of SURVIVIN was elevated in ER-negative relative to ER-positive breast cancer. Both AURKA and SURVIVIN increased...

  12. Supernumerary ring chromosome 20 characterized by fluorescence in situ hybridization

    NARCIS (Netherlands)

    Van Langen, Irene M.; Otter, Mariëlle A.; Aronson, Daniël C.; Overweg-Plandsoen, W.C.G.; Hennekam, Raoul C.M.; Leschot, Nico J.; Hoovers, Jan M.N.

    1996-01-01

    We report on a boy with mild dysmorphic features and developmental delay, in whom karyotyping showed an additional minute ring chromosome in 60% of metaphases. Fluorescence in situ hybridization (FISH) with a centromere specific probe demonstrated that the ring chromosome contained the centromeric r

  13. Molecular distinction between true centric fission and pericentric duplication-fission

    NARCIS (Netherlands)

    Perry, J; Nouri, S; La, P; Daniel, A; Wu, ZH; Purvis-Smith, S; Northrop, E; Choo, KHA; Slater, HR

    2005-01-01

    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

  14. Test for Chemical Induction of Chromosome Aberrations in Cultured Chinese Hamster Ovary (CHO) Cells with and without Metabolic Activation, Test Article: 3-Nitro-1,2,4-Triazol-5-one (NTO)

    Science.gov (United States)

    2008-10-30

    3110 Dicentric - an asymmetrical exchange between two chromosomes resulting in a chromosome with two centromeres with or without an accompanying...chromatid union. Dicentric - an asymmetrical exchange between two chromosomes resulting in a chromosome with two centromeres with or without an...Test for Chemical fuduction of Chromosome Aberrations in Cultured Chinese Hamster Ovary (CHO) Cells With and Without Metabolic Activation Test

  15. Nonrandom distribution of interhomolog recombination events induced by breakage of a dicentric chromosome in Saccharomyces cerevisiae.

    Science.gov (United States)

    Song, Wei; Gawel, Malgorzata; Dominska, Margaret; Greenwell, Patricia W; Hazkani-Covo, Einat; Bloom, Kerry; Petes, Thomas D

    2013-05-01

    Dicentric chromosomes undergo breakage in mitosis, resulting in chromosome deletions, duplications, and translocations. In this study, we map chromosome break sites of dicentrics in Saccharomyces cerevisiae by a mitotic recombination assay. The assay uses a diploid strain in which one homolog has a conditional centromere in addition to a wild-type centromere, and the other homolog has only the wild-type centromere; the conditional centromere is inactive when cells are grown in galactose and is activated when the cells are switched to glucose. In addition, the two homologs are distinguishable by multiple single-nucleotide polymorphisms (SNPs). Under conditions in which the conditional centromere is activated, the functionally dicentric chromosome undergoes double-stranded DNA breaks (DSBs) that can be repaired by mitotic recombination with the homolog. Such recombination events often lead to loss of heterozygosity (LOH) of SNPs that are centromere distal to the crossover. Using a PCR-based assay, we determined the position of LOH in multiple independent recombination events to a resolution of ∼4 kb. This analysis shows that dicentric chromosomes have recombination breakpoints that are broadly distributed between the two centromeres, although there is a clustering of breakpoints within 10 kb of the conditional centromere.

  16. Karyomorphology of six taxa in Chrysanthemum sensu lato (Anthemideae) in Egypt and their genetic relationships by Giemsa C-banding

    Institute of Scientific and Technical Information of China (English)

    Magdy Hussein ABD EL-TWAB; Ahmad Mohammad M. MEKAWY; Mohammad Saad EL-KATATNY

    2012-01-01

    Giemsa C-banding was applied to the chromosome complements of six diploid species belonging to six genera in Chrysanthemum sensu lato (Anthemideae) distributed in Egypt.Four types of C-banding distribution were observed in the taxa as follows:(i) negative C-banding in Anacyclus monanthos (L.) Thell.; (ii) all bands in terminal regions in Achilleafragrantissima (Forssk.) Sch.Bip,which showed 32 bands on 18 chromosomes; (iii)all eight bands at centromeric regions on eight chromosomes in Matricaria recutita L.; and (iv) bands at terminal and centromeric regions in Brocchia cinerea Vis.(12 terminal and six centromeric bands on 12 chromosomes),Cotula barbata DC.(four terminal,six centromeric,and eight short arm bands on 16 chromosomes),and Glebionis coronaria (L.) Cass.ex Spach.(eight terminal on the short arms and four large bands in centromeric regions on 12chromosomes).

  17. Location of chromosomes in the nucleus of human mesenchymal stem cells.

    Science.gov (United States)

    Lavrov, A V; Voldgorn, Y I

    2011-08-01

    For evaluation of the spatial structure of chromatin in nuclei of mesenchymal SC we determined the position of centromeres and individual chromosomes in interphase nucleus of mesenchymal SC. More than 300 nuclei in 7 cultures of mesenchymal SC were analyzed. Centromeres of chromosomes 6, 8, and 11 lie at a longer (0.68, 0.67, 0.7), while centromere of chromosome 18 at a shorter radial distance (0.49). Homologues of each chromosome had different radial distances. No differences in radial distances of centromeres were detected between mesenchymal SC from the adipose tissue and BM. After passage 8, distal displacement of chromosome 6 centromere (from 0.66 to 0.72) was observed, which probably indicates aging or spontaneous differentiation of cells.

  18. Telomere disruption results in non-random formation of de novo dicentric chromosomes involving acrocentric human chromosomes.

    Directory of Open Access Journals (Sweden)

    Kaitlin M Stimpson

    2010-08-01

    Full Text Available Genome rearrangement often produces chromosomes with two centromeres (dicentrics that are inherently unstable because of bridge formation and breakage during cell division. However, mammalian dicentrics, and particularly those in humans, can be quite stable, usually because one centromere is functionally silenced. Molecular mechanisms of centromere inactivation are poorly understood since there are few systems to experimentally create dicentric human chromosomes. Here, we describe a human cell culture model that enriches for de novo dicentrics. We demonstrate that transient disruption of human telomere structure non-randomly produces dicentric fusions involving acrocentric chromosomes. The induced dicentrics vary in structure near fusion breakpoints and like naturally-occurring dicentrics, exhibit various inter-centromeric distances. Many functional dicentrics persist for months after formation. Even those with distantly spaced centromeres remain functionally dicentric for 20 cell generations. Other dicentrics within the population reflect centromere inactivation. In some cases, centromere inactivation occurs by an apparently epigenetic mechanism. In other dicentrics, the size of the alpha-satellite DNA array associated with CENP-A is reduced compared to the same array before dicentric formation. Extra-chromosomal fragments that contained CENP-A often appear in the same cells as dicentrics. Some of these fragments are derived from the same alpha-satellite DNA array as inactivated centromeres. Our results indicate that dicentric human chromosomes undergo alternative fates after formation. Many retain two active centromeres and are stable through multiple cell divisions. Others undergo centromere inactivation. This event occurs within a broad temporal window and can involve deletion of chromatin that marks the locus as a site for CENP-A maintenance/replenishment.

  19. Telomere disruption results in non-random formation of de novo dicentric chromosomes involving acrocentric human chromosomes.

    Science.gov (United States)

    Stimpson, Kaitlin M; Song, Ihn Young; Jauch, Anna; Holtgreve-Grez, Heidi; Hayden, Karen E; Bridger, Joanna M; Sullivan, Beth A

    2010-08-12

    Genome rearrangement often produces chromosomes with two centromeres (dicentrics) that are inherently unstable because of bridge formation and breakage during cell division. However, mammalian dicentrics, and particularly those in humans, can be quite stable, usually because one centromere is functionally silenced. Molecular mechanisms of centromere inactivation are poorly understood since there are few systems to experimentally create dicentric human chromosomes. Here, we describe a human cell culture model that enriches for de novo dicentrics. We demonstrate that transient disruption of human telomere structure non-randomly produces dicentric fusions involving acrocentric chromosomes. The induced dicentrics vary in structure near fusion breakpoints and like naturally-occurring dicentrics, exhibit various inter-centromeric distances. Many functional dicentrics persist for months after formation. Even those with distantly spaced centromeres remain functionally dicentric for 20 cell generations. Other dicentrics within the population reflect centromere inactivation. In some cases, centromere inactivation occurs by an apparently epigenetic mechanism. In other dicentrics, the size of the alpha-satellite DNA array associated with CENP-A is reduced compared to the same array before dicentric formation. Extra-chromosomal fragments that contained CENP-A often appear in the same cells as dicentrics. Some of these fragments are derived from the same alpha-satellite DNA array as inactivated centromeres. Our results indicate that dicentric human chromosomes undergo alternative fates after formation. Many retain two active centromeres and are stable through multiple cell divisions. Others undergo centromere inactivation. This event occurs within a broad temporal window and can involve deletion of chromatin that marks the locus as a site for CENP-A maintenance/replenishment.

  20. The requirement for the Dam1 complex is dependent upon the number of kinetochore proteins and microtubules.

    Science.gov (United States)

    Burrack, Laura S; Applen, Shelly E; Berman, Judith

    2011-05-24

    The Dam1 complex attaches the kinetochore to spindle microtubules and is a processivity factor in vitro. In Saccharomyces cerevisiae, which has point centromeres that attach to a single microtubule, deletion of any Dam1 complex member results in chromosome segregation failures and cell death. In Schizosaccharomyces pombe, which has epigenetically defined regional centromeres that each attach to 3-5 kinetochore microtubules, Dam1 complex homologs are not essential. To determine why the complex is essential in some organisms and not in others, we used Candida albicans, a multimorphic yeast with regional centromeres that attach to a single microtubule. Interestingly, the Dam1 complex was essential in C. albicans, suggesting that the number of microtubules per centromere is critical for its requirement. Importantly, by increasing CENP-A expression levels, more kinetochore proteins and microtubules were recruited to the centromeres, which remained fully functional. Furthermore, Dam1 complex members became less crucial for growth in cells with extra kinetochore proteins and microtubules. Thus, the requirement for the Dam1 complex is not due to the DNA-specific nature of point centromeres. Rather, the Dam1 complex is less critical when chromosomes have multiple kinetochore complexes and microtubules per centromere, implying that it functions as a processivity factor in vivo as well as in vitro.

  1. The PP2A inhibitor I2PP2A is essential for sister chromatid segregation in oocyte meiosis II.

    Science.gov (United States)

    Chambon, Jean-Philippe; Touati, Sandra A; Berneau, Stéphane; Cladière, Damien; Hebras, Céline; Groeme, Rachel; McDougall, Alex; Wassmann, Katja

    2013-03-18

    Haploid gametes are generated through two consecutive meiotic divisions, with the segregation of chromosome pairs in meiosis I and sister chromatids in meiosis II. Separase-mediated stepwise removal of cohesion, first from chromosome arms and later from the centromere region, is a prerequisite for maintaining sister chromatids together until their separation in meiosis II [1]. In all model organisms, centromeric cohesin is protected from separase-dependent removal in meiosis I through the activity of PP2A-B56 phosphatase, which is recruited to centromeres by shugoshin/MEI-S332 (Sgo) [2-5]. How this protection of centromeric cohesin is removed in meiosis II is not entirely clear; we find that all the PP2A subunits remain colocalized with the cohesin subunit Rec8 at the centromere of metaphase II chromosomes. Here, we show that sister chromatid separation in oocytes depends on a PP2A inhibitor, namely I2PP2A. I2PP2A colocalizes with the PP2A enzyme at centromeres at metaphase II, independently of bipolar attachment. When I2PP2A is depleted, sister chromatids fail to segregate during meiosis II. Our findings demonstrate that in oocytes I2PP2A is essential for faithful sister chromatid segregation by mediating deprotection of centromeric cohesin in meiosis II.

  2. Genome sequence, comparative analysis and population genetics of the domestic horse (Equus caballus)

    Science.gov (United States)

    Wade, CM; Giulotto, E; Sigurdsson, S; Zoli, M; Gnerre, S; Imsland, F; Lear, TL; Adelson, DL; Bailey, E; Bellone, RR; Blöcker, H; Distl, O; Edgar, RC; Garber, M; Leeb, T; Mauceli, E; MacLeod, JN; Penedo, MCT; Raison, JM; Sharpe, T; Vogel, J; Andersson, L; Antczak, DF; Biagi, T; Binns, MM; Chowdhary, BP; Coleman, SJ; Della Valle, G; Fryc, S; Guérin, G; Hasegawa, T; Hill, EW; Jurka, J; Kiialainen, A; Lindgren, G; Liu, J; Magnani, E; Mickelson, JR; Murray, J; Nergadze, SG; Onofrio, R; Pedroni, S; Piras, MF; Raudsepp, T; Rocchi, M; Røed, KH; Ryder, OA; Searle, S; Skow, L; Swinburne, JE; Syvänen, AC; Tozaki, T; Valberg, SJ; Vaudin, M; White, JR; Zody, MC; Lander, ES; Lindblad-Toh, K

    2013-01-01

    We report a high-quality draft sequence of the genome of the horse (Equus caballus). The genome is relatively repetitive, but has little segmental duplication. Chromosomes appear to have undergone few historical rearrangements – 48% of equine chromosomes show conserved synteny to a single human chromosome. Equine chromosome 11 is shown to have an evolutionary novel centromere devoid of centromeric satellite DNA, suggesting that centromeric function may arise prior to satellite repeat accumulation. Linkage disequilibrium, showing the influences of early domestication of large herds of female horses, is intermediate in length between dog and human, and there is long-range haplotype sharing among breeds. PMID:19892987

  3. ##%The effects of a heterochromatin polymorphism in chromosome 6 on premature ovarian failure

    Institute of Scientific and Technical Information of China (English)

    Halime Kk; Yunus Aydin; Ebru Erzurumluoglu; Muhsin zdemir; Hikmet Hassa; Sevilhan Artan

    2015-01-01

    Through cytogenetic analysis, heteromorphisms were detected in chromosomes 1,9,16 and Y and defined as non-phenotypic variations. The polymorphism in the centromeric heterochromatin region of chromosome 6 is a rare variant, and only five cases have been documented in the literature. In our study, we used cytogenetic and molecular techniques to detect an increase in the centromeric heterochromatin region in the short arm of both copies of homologous chromosome 6 in a premature ovarian failure (POF) case. The report of this case is important for determining the relationship between fertility and the frequency of rare variants of the centromeric heterochromatin region of chromosome 6 in the general population.

  4. Kinetochores coordinate pericentromeric cohesion and early DNA replication by Cdc7-Dbf4 kinase recruitment.

    Science.gov (United States)

    Natsume, Toyoaki; Müller, Carolin A; Katou, Yuki; Retkute, Renata; Gierliński, Marek; Araki, Hiroyuki; Blow, J Julian; Shirahige, Katsuhiko; Nieduszynski, Conrad A; Tanaka, Tomoyuki U

    2013-06-01

    Centromeres play several important roles in ensuring proper chromosome segregation. Not only do they promote kinetochore assembly for microtubule attachment, but they also support robust sister chromatid cohesion at pericentromeres and facilitate replication of centromeric DNA early in S phase. However, it is still elusive how centromeres orchestrate all these functions at the same site. Here, we show that the budding yeast Dbf4-dependent kinase (DDK) accumulates at kinetochores in telophase, facilitated by the Ctf19 kinetochore complex. This promptly recruits Sld3-Sld7 replication initiator proteins to pericentromeric replication origins so that they initiate replication early in S phase. Furthermore, DDK at kinetochores independently recruits the Scc2-Scc4 cohesin loader to centromeres in G1 phase. This enhances cohesin loading and facilitates robust pericentromeric cohesion in S phase. Thus, we have found the central mechanism by which kinetochores orchestrate early S phase DNA replication and robust sister chromatid cohesion at microtubule attachment sites.

  5. CENP-C directs a structural transition of CENP-A nucleosomes mainly through sliding of DNA gyres.

    Science.gov (United States)

    Falk, Samantha J; Lee, Jaehyoun; Sekulic, Nikolina; Sennett, Michael A; Lee, Tae-Hee; Black, Ben E

    2016-03-01

    The histone H3 variant CENP-A is incorporated into nucleosomes that mark centromere location. We have recently reported that CENP-A nucleosomes, compared with their H3 counterparts, confer an altered nucleosome shape. Here, using a single-molecule fluorescence resonance energy transfer (FRET) approach with recombinant human histones and centromere DNA, we found that the nucleosome shape change directed by CENP-A is dominated by lateral passing of two DNA gyres (gyre sliding). A nonhistone centromere protein, CENP-C, binds and reshapes the nucleosome, sliding the DNA gyres back to positions similar to those in canonical nucleosomes containing conventional histone H3. The model that we generated to explain the CENP-A-nucleosome transition provides an example of a shape change imposed by external binding proteins and has notable implications for understanding of the epigenetic basis of the faithful inheritance of centromere location on chromosomes.

  6. Delineation of Methyl-DNA Binding Protein Interactions in the Prostate Cancer Genome

    Science.gov (United States)

    2013-07-01

    2 133028422 A001 Uronyl  2-­‐ sulfotransferase 4 133034161 A002 4 133421447 A003 8 43092956 A004 centromere 8 43095150 A005 centromere 8 43096794...Associa.on  Sites  for  MeCP2  and  MBD1 Uronyl  2-­‐ sulfotransferase TABLE 2: Genes up-regulated in Stage 3 Prostate Cancer Probe ID Stage 3

  7. Cd branding studies in a homologous Robertsonian 13;13 translocation.

    Science.gov (United States)

    Romain, D R; Columbano-Green, L; Sullivan, J; Smythe, R H; Gebbie, O; Parfitt, R; Chapman, C

    1982-01-01

    A phenotypically normal female with a history of two miscarriages was found to have the karyotype 45,XX,t(13p:13p). C banding showed the translocation to have two regions of centromeric constitutive heterochromatin, silver staining showed an active NOR in 60% of the cells screened, and Cd banding studies showed a single Cd band with absence of the Cd band at the suppressed centromere. Images PMID:7120322

  8. ICBP90 Regulation of DNA Methylation, Histone Ubiquitination, and Tumor Suppressor Gene Expression in Breast Cancer Cells

    Science.gov (United States)

    2013-09-01

    the plant lines with HpaII, a methylation-senstive enzyme, then visualized the digestion pattern using southern blots against a 180-base pair...methylation at centromere repeats. Genomic DNA was digested with HpaII or MspI as indicated, then probed in southern blots with a 180- bp centromere...at the Center for Vertebrate Genomics (CVG) which discusses papers on cancer and is attended by members of the Nikitn and Weiss labs , which study

  9. Chromosome engineering: generation of mono- and dicentric isochromosomes in a somatic cell hybrid system.

    Science.gov (United States)

    Higgins, A W; Schueler, M G; Willard, H F

    1999-08-01

    The most common isochromosome found in humans involves the long arm of the X, i(Xq), and is associated with a subset of Turner syndrome cases. To study the formation and behavior of isochromosomes in a more tractable experimental system, we have developed a somatic cell hybrid model system that allows for the selection of mono- or dicentric isochromosomes involving the short arm of the X, i(Xp). Simultaneous positive and negative counterselection of a mouse/human somatic cell hybrid containing a human X chromosome, selecting for retention of the UBE1 locus in Xp but against the HPRT locus in Xq, results in a variety of abnormalities of the X chromosome involving deletions of Xq. We have generated 70 such "Pushmi-Pullyu" hybrids derived from seven independent X chromosomes. Cytogenetic analysis of these hybrids using fluorescence in situ hybridization showed i(Xp) chromosomes in approximately 19% of the hybrids. Southern blot and polymerase chain reaction analyses of the Pushmi-Pullyu hybrids revealed a distribution of breakpoints along Xq. The distance between the centromeres of the dicentric i(Xp)s generated ranged from approximately 2 Mb to approximately 20 Mb. To examine centromeric activity in these dicentric i(Xp)s, we used indirect immunofluorescence with antibodies to centromere protein E (CENP-E). CENP-E was detected at only one of the centromeres of a dicentric i(Xp) with approximately 2-3 Mb of Xq DNA. In contrast, CENP-E was detected at both centromeres of a dicentric i(Xp) with approximately 14 Mb of Xq DNA. Two other dicentric i(Xp) chromosomes were heterogeneous with respect to centromeric activity, suggesting that centromeric activity and chromosome stability of dicentric chromosomes may be more complicated than previously thought. The Pushmi-Pullyu model system presented in this study may provide a tool for examining the structure and function of mammalian centromeres.

  10. Test for Chemical Induction of Chromosome Aberrations in Cultured Chinese Hamster Ovary (CHO) Cells With and Without Metabolic Activation. Test Article: Dimethylamine-2-2ethyl azide (DMAZ)

    Science.gov (United States)

    2008-07-26

    chromosomes leading to four-armed configurations. This could be asymmetrical with formation of a dicentric and an acentric chromatid, ifunion is complete...chromatid union. Dicentric - an asymmetrical exchange between two chromosomes resulting in a chromosome with two centromeres with or without an...no sister chromatid union. ’ d - Dicentric - an asymmetrical exchange between two chromosomes resulting, r dm in a chromosome with two centromeres

  11. Misregulation of Scm3p/HJURP Causes Chromosome Instability in Saccharomyces cerevisiae and Human Cells

    OpenAIRE

    Prashant K. Mishra; Wei-Chun Au; John S. Choy; P Henning Kuich; Baker, Richard E.; Foltz, Daniel R.; Basrai, Munira A.

    2011-01-01

    The kinetochore (centromeric DNA and associated proteins) is a key determinant for high fidelity chromosome transmission. Evolutionarily conserved Scm3p is an essential component of centromeric chromatin and is required for assembly and function of kinetochores in humans, fission yeast, and budding yeast. Overexpression of HJURP, the mammalian homolog of budding yeast Scm3p, has been observed in lung and breast cancers and is associated with poor prognosis; however, the physiological relevanc...

  12. The CENP-T/-W complex is a binding partner of the histone chaperone FACT.

    Science.gov (United States)

    Prendergast, Lisa; Müller, Sebastian; Liu, Yiwei; Huang, Hongda; Dingli, Florent; Loew, Damarys; Vassias, Isabelle; Patel, Dinshaw J; Sullivan, Kevin F; Almouzni, Geneviève

    2016-06-01

    The CENP-T/-W histone fold complex, as an integral part of the inner kinetochore, is essential for building a proper kinetochore at the centromere in order to direct chromosome segregation during mitosis. Notably, CENP-T/-W is not inherited at centromeres, and new deposition is absolutely required at each cell cycle for kinetochore function. However, the mechanisms underlying this new deposition of CENP-T/-W at centromeres are unclear. Here, we found that CENP-T deposition at centromeres is uncoupled from DNA synthesis. We identified Spt16 and SSRP1, subunits of the H2A-H2B histone chaperone facilitates chromatin transcription (FACT), as CENP-W binding partners through a proteomic screen. We found that the C-terminal region of Spt16 binds specifically to the histone fold region of CENP-T/-W. Furthermore, depletion of Spt16 impairs CENP-T and CENP-W deposition at endogenous centromeres, and site-directed targeting of Spt16 alone is sufficient to ensure local de novo CENP-T accumulation. We propose a model in which the FACT chaperone stabilizes the soluble CENP-T/-W complex in the cell and promotes dynamics of exchange, enabling CENP-T/-W deposition at centromeres.

  13. c-Myc—Dependent Formation of Robertsonian Translocation Chromosomes in Mouse Cells

    Directory of Open Access Journals (Sweden)

    Amanda Guffei

    2007-07-01

    Full Text Available 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.

  14. Topography of Genetic Loci in Tissue Samples: Towards New Diagnostic Tool Using Interphase FISH and High-Resolution Image Analysis Techniques

    Directory of Open Access Journals (Sweden)

    I. Koutná

    2000-01-01

    Full Text Available Using single and dual colour fluorescence in situ hybridisation (FISH combined with image analysis techniques the topographic characteristics of genes and centromeres in nuclei of human colon tissue cells were investigated. The distributions of distances from the centre‐of‐nucleus to genes (centromeres and from genes to genes (centromeres to centromeres were studied in normal colon tissue cells found in the neighbourhood of tumour samples, in tumour cell line HT‐29 and in promyelocytic HL‐60 cell line for comparison. Our results show that the topography of genetic loci determined in 3D‐fixed cell tissue corresponds to that obtained for 2D‐fixed cells separated from the tissue. The distributions of the centre‐of‐nucleus to gene (centromere distances and gene to gene (centromere to centromere distances and their average values are different for various genetic loci but similar for normal colon tissue cells, HT‐29 colon tumour cell line and HL‐60 promyelocytic cell line. It suggests that the arrangement of genetic loci in cell nucleus is conserved in different types of human cells. The investigations of trisomic loci in HT‐29 cells revealed that the location of the third genetic element is not different from the location of two homologues in diploid cells. We have shown that the topographic parameters used in our experiments for different genetic elements are not tissue or tumour specific. In order to validate high‐resolution cytometry for oncology, further investigations should include more precise parameters reflecting the state of chromatin in the neighbourhood of critical oncogenes or tumour suppresser genes.

  15. Cloning of DNA sequences localized on proximal fluorescent chromosome bands by microdissection in Pinus densiflora Sieb. & Zucc.

    Science.gov (United States)

    Hizume, M; Shibata, F; Maruyama, Y; Kondo, T

    2001-09-01

    Japanese red pine, Pinus densiflora, has 2n=24 chromosomes, of which most carry chromomycin A3 (CMA) and 4',6-diamidino-2-phenylindole (DAPI) bands at their centromere-proximal regions. It was proposed that these regions contain highly repetitive DNA. The DNA localized in the proximal fluorescent bands was isolated and characterized. In P. densiflora, centromeric and neighboring segments of the somatic chromosomes were dissected with a manual micromanipulator. The centromeric DNA was amplified from the DNA contained in dissected centromeric segments by degenerate oligonucleotide primed-polymerase chain reaction (DOP-PCR) and a cloned DNA library was constructed. Thirty-one clones carrying highly repetitive DNA were selected by colony hybridization using Cot-1 DNA from this species as a probe, and their chromosomal localization was determined by fluorescent in situ hybridization (FISH). Clone PDCD501 was localized to the proximal CMA band of 20 chromosomes. This clone contained tandem repeats, comprising a 27 bp repeat unit, which was sufficient to provide the proximal FISH signal, with a 52.3% GC content. The repetitive sequence was named PCSR (proximal CMA band-specific repeat). Clone PDCD159 was 1700 bp in length, with a 61.7% AT content, and produced FISH signals at the proximal DAPI band of the remaining four chromosomes. Four clones hybridized strongly to the secondary constriction and gave weak signals at the centromeric region of several chromosomes. Clone PDCD537, one of the four clones, was homologous to the 26S rRNA gene. A PCR experiment using microdissected centromeric regions suggested that the centromeric region contains 18S and 26S rDNA. Another 24 clones hybridized to whole chromosome arms, with varying intensities and might represent dispersed repetitive DNA.

  16. Molecular cloning and sequence analysis of hamster CENP-A cDNA

    Science.gov (United States)

    Figueroa, Javier; Pendón, Carlos; Valdivia, Manuel M

    2002-01-01

    Background The centromere is a specialized locus that mediates chromosome movement during mitosis and meiosis. This chromosomal domain comprises a uniquely packaged form of heterochromatin that acts as a nucleus for the assembly of the kinetochore a trilaminar proteinaceous structure on the surface of each chromatid at the primary constriction. Kinetochores mediate interactions with the spindle fibers of the mitotic apparatus. Centromere protein A (CENP-A) is a histone H3-like protein specifically located to the inner plate of kinetochore at active centromeres. CENP-A works as a component of specialized nucleosomes at centromeres bound to arrays of repeat satellite DNA. Results We have cloned the hamster homologue of human and mouse CENP-A. The cDNA isolated was found to contain an open reading frame encoding a polypeptide consisting of 129 amino acid residues with a C-terminal histone fold domain highly homologous to those of CENP-A and H3 sequences previously released. However, significant sequence divergence was found at the N-terminal region of hamster CENP-A that is five and eleven residues shorter than those of mouse and human respectively. Further, a human serine 7 residue, a target site for Aurora B kinase phosphorylation involved in the mechanism of cytokinesis, was not found in the hamster protein. A human autoepitope at the N-terminal region of CENP-A described in autoinmune diseases is not conserved in the hamster protein. Conclusions We have cloned the hamster cDNA for the centromeric protein CENP-A. Significant differences on protein sequence were found at the N-terminal tail of hamster CENP-A in comparison with that of human and mouse. Our results show a high degree of evolutionary divergence of kinetochore CENP-A proteins in mammals. This is related to the high diverse nucleotide repeat sequences found at the centromere DNA among species and support a current centromere model for kinetochore function and structural plasticity. PMID:12019018

  17. Observações citológicas em Dysdercus cadeias de cromossômios em tecido somático de Dysdercus mendesi Bloete (Hemiptera-Pyrrhocoridae

    Directory of Open Access Journals (Sweden)

    Luiz O. T. Mendes

    1949-01-01

    Full Text Available The male of Dysdercus mendesi Bloete (1937 has 2n = 16 chromosomes : 14 autosomes and 2 sex-chromosomes. Details are presented on the morphology of these chromosomes of the somatic cells of embryonic tissue. The long pair of rod shaped chromosomes is the longest of the set, and they show a conspicuous sub-terminal constriction (supposed to be the centromere and five other smaller constrictions. In most of the other chromosomes sub-terminally localized constriction (supposed also to be the centromere can also be observed. Based on these observations it has been concluded that the chromosomes of D. mendesi are morphologically normal. Dicentric chromosomes in the somatic tissue of this species are absent and the hypothesis of existence of diffuse centromere is also excluded. All of the chromosomes appear to have a definitive localized centromere. Metaphase spermatogonial plates have been observed where all the chromosomes appear as short and thick rods, apparently attached end-to-end, by definite connections, and forming a ring-shaped chain. Sometimes one or two chromosomes were found inside a ring formed by the others, but always connected to one or two of the chromosomes of the ring. Chromosomes chains of varied shapes were also found in the metaphase plates. In anaphase the longitudinally split chromosomes of the rings attain a curved shape as they move to the poles, giving the impression that both their extremities are pulled to the poles. A new hypothesis is presented to explain why the normal chromosomes of D. mendesi, provided with one localized centromere, can attain the curved shape observed in the anaphase configurations. It is based on the existence of connections attaching the chromosomes by their extremities, from metaphase until anaphase, and giving origin to the chain configuration already mentioned. These chains of chromosomes behave in anaphase as one rather long chromosomes with several centromeres. The centromeres are pulled to

  18. CDK-regulated dimerization of M18BP1 on a Mis18 hexamer is necessary for CENP-A loading

    Science.gov (United States)

    Pan, Dongqing; Klare, Kerstin; Petrovic, Arsen; Take, Annika; Walstein, Kai; Singh, Priyanka; Rondelet, Arnaud; Bird, Alexander W; Musacchio, Andrea

    2017-01-01

    Centromeres are unique chromosomal loci that promote the assembly of kinetochores, macromolecular complexes that bind spindle microtubules during mitosis. In most organisms, centromeres lack defined genetic features. Rather, they are specified epigenetically by a centromere-specific histone H3 variant, CENP-A. The Mis18 complex, comprising the Mis18α:Mis18β subcomplex and M18BP1, is crucial for CENP-A homeostasis. It recruits the CENP-A-specific chaperone HJURP to centromeres and primes it for CENP-A loading. We report here that a specific arrangement of Yippee domains in a human Mis18α:Mis18β 4:2 hexamer binds two copies of M18BP1 through M18BP1’s 140 N-terminal residues. Phosphorylation by Cyclin-dependent kinase 1 (CDK1) at two conserved sites in this region destabilizes binding to Mis18α:Mis18β, limiting complex formation to the G1 phase of the cell cycle. Using an improved viral 2A peptide co-expression strategy, we demonstrate that CDK1 controls Mis18 complex recruitment to centromeres by regulating oligomerization of M18BP1 through the Mis18α:Mis18β scaffold. DOI: http://dx.doi.org/10.7554/eLife.23352.001 PMID:28059702

  19. Chromatin structure and ATRX function in mouse oocytes.

    Science.gov (United States)

    De La Fuente, Rabindranath; Baumann, Claudia; Viveiros, Maria M

    2012-01-01

    Differentiation of chromatin structure and function during oogenesis is essential to confer the mammalian oocyte with meiotic and developmental potential. Errors in chromosome segregation during female meiosis and subsequent transmission of an abnormal chromosome complement (aneuploidy) to the early conceptus are one of the leading causes of pregnancy loss in women. The chromatin remodeling protein ATRX (α-thalassemia mental retardation X-linked) has recently emerged as a critical factor involved in heterochromatin formation at mammalian centromeres during meiosis. In mammalian oocytes, ATRX binds to centromeric heterochromatin domains where it is required for accurate chromosome segregation. Loss of ATRX function induces abnormal meiotic chromosome morphology, reduces histone H3 phosphorylation, and promotes a high incidence of aneuploidy associated with severely reduced fertility. The presence of centromeric breaks during the transition to the first mitosis in the early embryo indicates that the role of ATRX in chromosome segregation is mediated through an epigenetic mechanism involving the maintenance of chromatin modifications associated with pericentric heterochromatin (PCH) formation and chromosome condensation. This is consistent with the existence of a potential molecular link between centromeric and PCH in the epigenetic control of centromere function and maintenance of chromosome stability in mammalian oocytes. Dissecting the molecular mechanisms of ATRX function during meiosis will have important clinical implications towards uncovering the epigenetic factors contributing to the onset of aneuploidy in the human oocyte.

  20. Pericentric satellite DNA and molecular phylogeny in Acomys (Rodentia).

    Science.gov (United States)

    Kunze, B; Traut, W; Garagna, S; Weichenhan, D; Redi, C A; Winking, H

    1999-01-01

    Satellite DNAs (stDNAs) of four Acomys species (spiny-mice), A. cahirinus, A. cineraceus, A. dimidiatus and A. russatus, belong to closely related sequence families. Monomer sizes range from 338 to 364 bp. Between-species sequence identity was from 81.0% to 97.2%. The molecular phylogeny of the sequences helps to clarify the taxonomy of this 'difficult' group. The A. dimidiatus genome contains about 60000 repeats. According to the restriction patterns, repeats are arranged in tandem. The stDNA maps to the centromeric heterochromatin of most autosomes, both acrocentric and metacentric, but appears to be absent in the centromeric region of Y chromosomes. A well-conserved centromere protein B (CENP-B) box is present in the stDNA of A. russatus while it is degenerated in the other species.

  1. Cytokinesis breaks dicentric chromosomes preferentially at pericentromeric regions and telomere fusions.

    Science.gov (United States)

    Lopez, Virginia; Barinova, Natalja; Onishi, Masayuki; Pobiega, Sabrina; Pringle, John R; Dubrana, Karine; Marcand, Stéphane

    2015-02-01

    Dicentric chromosomes are unstable products of erroneous DNA repair events that can lead to further genome rearrangements and extended gene copy number variations. During mitosis, they form anaphase bridges, resulting in chromosome breakage by an unknown mechanism. In budding yeast, dicentrics generated by telomere fusion break at the fusion, a process that restores the parental karyotype and protects cells from rare accidental telomere fusion. Here, we observed that dicentrics lacking telomere fusion preferentially break within a 25- to 30-kb-long region next to the centromeres. In all cases, dicentric breakage requires anaphase exit, ruling out stretching by the elongated mitotic spindle as the cause of breakage. Instead, breakage requires cytokinesis. In the presence of dicentrics, the cytokinetic septa pinch the nucleus, suggesting that dicentrics are severed after actomyosin ring contraction. At this time, centromeres and spindle pole bodies relocate to the bud neck, explaining how cytokinesis can sever dicentrics near centromeres.

  2. Physical maps and recombination frequency of six rice chromosomes.

    Science.gov (United States)

    Wu, Jianzhong; Mizuno, Hiroshi; Hayashi-Tsugane, Mika; Ito, Yukiyo; Chiden, Yoshino; Fujisawa, Masaki; Katagiri, Satoshi; Saji, Shoko; Yoshiki, Shoji; Karasawa, Wataru; Yoshihara, Rie; Hayashi, Akiko; Kobayashi, Harumi; Ito, Kazue; Hamada, Masao; Okamoto, Masako; Ikeno, Maiko; Ichikawa, Yoko; Katayose, Yuichi; Yano, Masahiro; Matsumoto, Takashi; Sasaki, Takuji

    2003-12-01

    We constructed physical maps of rice chromosomes 1, 2, and 6-9 with P1-derived artificial chromosome (PAC) and bacterial artificial chromosome (BAC) clones. These maps, with only 20 gaps, cover more than 97% of the predicted length of the six chromosomes. We submitted a total of 193 Mbp of non-overlapping sequences to public databases. We analyzed the DNA sequences of 1316 genetic markers and six centromere-specific repeats to facilitate characterization of chromosomal recombination frequency and of the genomic composition and structure of the centromeric regions. We found marked changes in the relative recombination rate along the length of each chromosome. Chromosomal recombination at the centromere core and surrounding regions on the six chromosomes was completely suppressed. These regions have a total physical length of about 23 Mbp, corresponding to 11.4% of the entire size of the six chromosomes. Chromosome 6 has the longest quiescent region, with about 5.6 Mbp, followed by chromosome 8, with quiescent region about half this size. Repetitive sequences accounted for at least 40% of the total genomic sequence on the partly sequenced centromeric region of chromosome 1. Rice CentO satellite DNA is arrayed in clusters and is closely associated with the presence of Centromeric Retrotransposon of Rice (CRR)- and RIce RetroElement 7 (RIRE7)-like retroelement sequences. We also detected relatively small coldspot regions outside the centromeric region; their repetitive content and gene density were similar to those of regions with normal recombination rates. Sequence analysis of these regions suggests that either the amount or the organization patterns of repetitive sequences may play a role in the inactivation of recombination.

  3. Molecular cloning and characterization of satellite DNA sequences from constitutive heterochromatin of the habu snake (Protobothrops flavoviridis, Viperidae) and the Burmese python (Python bivittatus, Pythonidae).

    Science.gov (United States)

    Matsubara, Kazumi; Uno, Yoshinobu; Srikulnath, Kornsorn; Seki, Risako; Nishida, Chizuko; Matsuda, Yoichi

    2015-12-01

    Highly repetitive DNA sequences of the centromeric heterochromatin provide valuable molecular cytogenetic markers for the investigation of genomic compartmentalization in the macrochromosomes and microchromosomes of sauropsids. Here, the relationship between centromeric heterochromatin and karyotype evolution was examined using cloned repetitive DNA sequences from two snake species, the habu snake (Protobothrops flavoviridis, Crotalinae, Viperidae) and Burmese python (Python bivittatus, Pythonidae). Three satellite DNA (stDNA) families were isolated from the heterochromatin of these snakes: 168-bp PFL-MspI from P. flavoviridis and 196-bp PBI-DdeI and 174-bp PBI-MspI from P. bivittatus. The PFL-MspI and PBI-DdeI sequences were localized to the centromeric regions of most chromosomes in the respective species, suggesting that the two sequences were the major components of the centromeric heterochromatin in these organisms. The PBI-MspI sequence was localized to the pericentromeric region of four chromosome pairs. The PFL-MspI and the PBI-DdeI sequences were conserved only in the genome of closely related species, Gloydius blomhoffii (Crotalinae) and Python molurus, respectively, although their locations on the chromosomes were slightly different. In contrast, the PBI-MspI sequence was also in the genomes of P. molurus and Boa constrictor (Boidae), and additionally localized to the centromeric regions of eight chromosome pairs in B. constrictor, suggesting that this sequence originated in the genome of a common ancestor of Pythonidae and Boidae, approximately 86 million years ago. The three stDNA sequences showed no genomic compartmentalization between the macrochromosomes and microchromosomes, suggesting that homogenization of the centromeric and/or pericentromeric stDNA sequences occurred in the macrochromosomes and microchromosomes of these snakes.

  4. Single-epitope recognition imaging of native chromatin

    Directory of Open Access Journals (Sweden)

    Wang Hongda

    2008-12-01

    Full Text Available Abstract Background Direct visualization of chromatin has the potential to provide important insights into epigenetic processes. In particular, atomic force microscopy (AFM can visualize single nucleosomes under physiological ionic conditions. However, AFM has mostly been applied to chromatin that has been reconstituted in vitro, and its potential as a tool for the dissection of native nucleosomes has not been explored. Recently we applied AFM to native Drosophila chromatin containing the centromere-specific histone 3 (CenH3, showing that it is greatly enriched in smaller particles. Taken together with biochemical analyses of CenH3 nucleosomes, we propose that centromeric nucleosomes are hemisomes, with one turn of DNA wrapped around a particle consisting of one molecule each of centromere-specific CenH3, H4, H2A and H2B. Results Here we apply a recognition mode of AFM imaging to directly identify CenH3 within histone core particles released from native centromeric chromatin. More than 90% of these particles were found to be tetrameric in height. The specificity of recognition was confirmed by blocking with a CenH3 peptide, and the strength of the interaction was quantified by force measurements. These results imply that the particles imaged by AFM are indeed mature CenH3-containing hemisomes. Conclusion Efficient and highly specific recognition of CenH3 in histone core particles isolated from native centromeric chromatin demonstrates that tetramers are the predominant form of centromeric nucleosomes in mature tetramers. Our findings provide proof of principle that this approach can yield insights into chromatin biology using direct and rapid detection of native nucleosomes in physiological salt concentrations.

  5. Modified C-band technique for the analysis of chromosome abnormalities in irradiated human lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Nakata, Akifumi; Akiyama, Miho; Yamada, Yuji [Biodosimetry Section, Department of Radiation Dosimetry, Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan); Yoshida, Mitsuaki A., E-mail: myoshida@cc.hirosaki-u.ac.jp [Biodosimetry Section, Department of Radiation Dosimetry, Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan)

    2011-10-15

    A modified C-band technique was developed in order to analyze more accurately dicentric, tricentric, and ring chromosomes in irradiated human peripheral lymphocytes. Instead of the original method relying on treatment with barium hydroxide Ba(OH){sub 2}, C-bands were obtained using a modified form of heat treatment in formamide followed with DAPI staining. This method was tentatively applied to the analysis of dicentric chromosomes in irradiated human lymphocytes to examine its availability. The frequency of dicentric chromosome was almost the same with conventional Giemsa staining and the modified C-band technique. In the analysis using Giemsa staining, it is relatively difficult to identify the centromere on the elongated chromosomes, over-condensed chromosomes, fragment, and acentric ring. However, the modified C-band method used in this study makes it easier to identify the centromere on such chromosomes than with the use of Giemsa staining alone. Thus, the modified C-band method may give more information about the location of the centromere. Therefore, this method may be available and more useful for biological dose estimation due to the analysis of the dicentric chromosome in human lymphocytes exposed to the radiation. Furthermore, this method is simpler and faster than the original C-band protocol and fluorescence in situ hybridization (FISH) method with the centromeric DNA probe. - Highlights: > The dicentric (dic) assay is the most effective for the radiation biodosimetry. > It is important to recognize the centromere of the dic. > We improved a C-band technique based on heat denaturation. > This technique enables the accurate detection of a centromere. > This method may be available and more useful for biological dose estimation.

  6. Identification of the origin of chromosomal aberrations by laser microdissection: Double minutes observed in two cases derive from different chromosomes

    Energy Technology Data Exchange (ETDEWEB)

    Rajcan-Separovic, E. [Univ. of Ottawa (Canada); Wang, H.S.; Janes, L. [Children`s Hospital of Eastern Ontario, Ottawa (Canada)

    1994-09-01

    Single copies of tiny chromosome fragments appearing as double minutes were observed in high frequency in amniotic fluid cultures of two mothers who underwent prenatal testing because of advanced age. In case 1, the minutes were C band and NOR negative, while in case 2 they were C band positive, NOR negative. In both cases centromeric DNA sequences were detected on double minutes by fluorescence in situ hybridization (FISH) using all human centromeric probe (ONCOR). We applied the laser microdissection method to diagnose the origin of double minutes. The diagnostic procedures consisted of microdissection of double minutes from single cells, polymerase chain reaction (PCR) amplification of the dissected DNA and subsequent FISH using PCR products as a probe pool on metaphase chromosomes from the patient`s amniocytes and fibroblasts from a karyotypically normal newborn. Using this strategy we observed strong FISH signals on double minutes and centromeres of a D and G group chromosome in case 1 amniocyytes, while in case 2 the signal was present on double minutes and a C group chromosome centromere. Hybridization of amniocyte chromosome spreads with centromeric {alpha}-satellite probes for the candidate chromosomes 13/21 and 14/22 (ONCOR) in case 1 revealed FISH signals on double minutes only with the 13/21 probe. In case 2, {alpha}-satellite probes for candidate chromosomes 10 and 12 (ONCOR) were used, and only the probe for chromosome 12 hybridized to double minutes. With the laser microdissection method we were thus able to diagnose and confirm that the double minutes observed in human amniocytes derived from centromeres of chromosomes 13/21 in case 1, and from centomeres of chromosome 12 in case 2. This demonstrates the utility of laser microdissection for identification of chromosomal abnormalities of unknown origin.

  7. Insight into F plasmid DNA segregation revealed by structures of SopB and SopB–DNA complexes

    OpenAIRE

    2010-01-01

    Accurate DNA segregation is essential for genome transmission. Segregation of the prototypical F plasmid requires the centromere-binding protein SopB, the NTPase SopA and the sopC centromere. SopB displays an intriguing range of DNA-binding properties essential for partition; it binds sopC to form a partition complex, which recruits SopA, and it also coats DNA to prevent non-specific SopA–DNA interactions, which inhibits SopA polymerization. To understand the myriad functions of SopB, we dete...

  8. Chromosomal localization of a tandemly repeated DNA sequence in Trifilium repens L.

    Institute of Scientific and Technical Information of China (English)

    ZHUJM; NWELLISON; 等

    1996-01-01

    A karyotype of Trifolium repens constructed from mitotic cells revealed 13 pairs of metacentric and 3 pairs of submetacentric chromosomes including a pair of satellites located at the end of the short arm of chromosome 16.C-bands were identified around the centromeric regions of 8 pairs of chromosomes.A 350 bp tandemly repeated DNAsequence from T.repens labelled with digoxygenin hybridized to the proximal centromeric regions of 12 chromosome pairs.Some correlation between the distribution of the repeat sequence and the distribution of C-banding was demonstrated.

  9. Regulation of localization and activity of the microtubule depolymerase MCAK.

    Science.gov (United States)

    Tanenbaum, Marvin E; Medema, René H; Akhmanova, Anna

    2011-03-01

    Mitotic Centromere Associated Kinesin (MCAK) is a potent microtubule depolymerizing and catastrophe-inducing factor, which uses the energy of ATP hydrolysis to destabilize microtubule ends. MCAK is localized to inner centromeres, kinetochores and spindle poles of mitotic cells, and is also present in the cytoplasm. Both in interphase and in mitosis, MCAK can specifically accumulate at the growing microtubule ends. Here we discuss the mechanisms, which modulate subcellular localization and activity of MCAK through the interaction with the End Binding (EB) proteins and phosphorylation.

  10. The fission yeast heterochromatin protein Rik1 is required for telomere clustering during meiosis

    DEFF Research Database (Denmark)

    Tuzon, Creighton T; Borgstrøm, Britta; Weilguny, Dietmar

    2004-01-01

    Telomeres share the ability to silence nearby transcription with heterochromatin, but the requirement of heterochromatin proteins for most telomere functions is unknown. The fission yeast Rik1 protein is required for heterochromatin formation at centromeres and the mating-type locus, as it recrui...... meiosis. However, Rik1 is dispensable for the protective roles of telomeres in preventing chromosome end-fusion. Thus, a Swi6-independent heterochromatin function distinct from that at centromeres and the mating-type locus operates at telomeres during sexual differentiation....

  11. Molecular characterization and association analysis of porcine PANE1 gene

    DEFF Research Database (Denmark)

    Huang, Honggang; Deng, Hong; Yang, Yiling

    2010-01-01

    The proliferation associated nuclear element 1 (PANE1) is a novel gene that is involved in immune response besides its primary role in centromere assembly. Different PANE1 transcripts show a distinct expression patterns in resting and activated CD19+ cells. In this study, we cloned and characteri......The proliferation associated nuclear element 1 (PANE1) is a novel gene that is involved in immune response besides its primary role in centromere assembly. Different PANE1 transcripts show a distinct expression patterns in resting and activated CD19+ cells. In this study, we cloned...

  12. A neocentromere on human chromosome 3 without detectable alpha-satellite DNA forms morphologically normal kinetochores

    DEFF Research Database (Denmark)

    Wandall, A; Tranebjaerg, L; Tommerup, Niels

    1998-01-01

    A neocentromere at 3q26 was observed in a father and his daughter on a chromosome 3 with deleted centromeric region. No alpha-satellite DNA was detectable at the 3q26 neocentromere, but it was weakly positive with anticentromere (CREST) antibodies. Electron microscopy showed that the neocentromere...... formed microtubule-associated kinetochores with normal morphology and of the same size as the kinetochores of other large chromosomes. The deleted centromere formed a small linear marker chromosome that reacted strongly with anticentromere antibodies, but showed reduced kinetochore size. The 3q26...

  13. A Specialized Nucleosome Has a “Point” to Make

    OpenAIRE

    Zhang, Weiguo; Mellone, Barbara G; Karpen, Gary H

    2007-01-01

    Three recent papers, including Mizuguchi et al. (2007) in this issue, show that the nonhistone protein Scm3 is required for the recruitment of the histone H3 variant Cse4 to centromeres in budding yeast. Scm3 forms a chromatin component with Cse4:histone H4 tetramers that appear to lack H2A/H2B histones. These studies provide key insights into the pathway that recruits Cse4 to centromeres and have important implications for other functions of chromatin.

  14. Overexpression of LSD1 contributes to human carcinogenesis through chromatin regulation in various cancers.

    Science.gov (United States)

    Hayami, Shinya; Kelly, John D; Cho, Hyun-Soo; Yoshimatsu, Masanori; Unoki, Motoko; Tsunoda, Tatsuhiko; Field, Helen I; Neal, David E; Yamaue, Hiroki; Ponder, Bruce A J; Nakamura, Yusuke; Hamamoto, Ryuji

    2011-02-01

    A number of histone demethylases have been identified and biochemically characterized, but the pathological roles of their dysfunction in human disease like cancer have not been well understood. Here, we demonstrate important roles of lysine-specific demethylase 1 (LSD1) in human carcinogenesis. Expression levels of LSD1 are significantly elevated in human bladder carcinomas compared with nonneoplastic bladder tissues (p human embryonic kidney fibroblast cells. Expression profile analysis showed that LSD1 could affect the expression of genes involved in various chromatin-modifying pathways such as chromatin remodeling at centromere, centromeric heterochromatin formation and chromatin assembly, indicating its essential roles in carcinogenesis through chromatin modification.

  15. DNA gains at 8q23.2

    DEFF Research Database (Denmark)

    da Silva Veiga, Luciana Caricati; Bérgamo, Nádia Aparecida; dos Reis, Patrícia Pintor

    2003-01-01

    Gains or amplifications involving chromosome arm 8q are one of the most recurrent chromosomal alterations in head and neck tumors. To characterize previously reported gains, we performed fluorescence in situ hybridization (FISH) using the sequences BAC RP1179E1 and 8-centromere PMJ 128 as probes....

  16. Why is chromosome segregation error in oocytes increased with maternal aging?

    Science.gov (United States)

    Wang, Zhen-Bo; Schatten, Heide; Sun, Qing-Yuan

    2011-10-01

    It is well documented that female fertility is decreased with advanced maternal age due to chromosome abnormality in oocytes. Increased chromosome missegregation is mainly caused by centromeric cohesion reduction. Other factors such as weakened homologous recombination, improper spindle organization, spindle assembly checkpoint (SAC) malfunction, chromatin epigenetic changes, and extra-oocyte factors may also cause chromosome errors.

  17. Sequence Classification: 893553 [

    Lifescience Database Archive (English)

    Full Text Available n involved in repression of transcription at the silent mating-type loci HML and HMR; recruitment to silent chromatin requires intera...ctions with Orc1p and with Sir4p, through a common Sir1p domain; binds to centromeric chromatin; Sir1p || http://www.ncbi.nlm.nih.gov/protein/6322954 ...

  18. Genetics and epigenetics of repeat derepression in human disease

    NARCIS (Netherlands)

    Thijssen, P.E.

    2016-01-01

    A large part of the human genome consists of repetitive DNA. In this thesis two human diseases have been studied in which deregulation of repetitive DNA is a central feature: facioscapulohumeral muscular dystrophy (FSHD) and immunodeficiency, centromere instability and facial anomalies (ICF) syndrom

  19. Recurrent unbalanced whole-arm t(1;10)(q10;p10) in myelodysplastic syndrome: a case report and literature review.

    Science.gov (United States)

    Odish, Omar Ferkad Faraj; Gotoh, Akihiko; Liu, Yi-Chang; Shoji, Nohoko; Kimura, Yukihiko; Kodama, Atsushi; Ohyashiki, Kazuma

    2007-01-15

    We report a patient with myelodysplastic syndrome (refractory anemia) showing the karyotype 46,XY,+1,der(1;10)(q10;p10), resulting in trisomy 1q and monosomy 10q abnormality. This finding suggests that either trisomy of 1q or centromeric connection between chromosomes 1 and 10, rather than the absence of 10q, might be essential toward neoplastic transformation.

  20. DNA Catenation Maintains Structure of Human Metaphase Chromosomes

    DEFF Research Database (Denmark)

    L. V. Bauer, David; Marie, Rodolphe; Rasmussen, Kristian Hagsted

    2012-01-01

    , and destroyed to complete sister chromatid disjunction. In addition to demonstrating the value of microfluidics as a tool for examining chromosome structure, these results lend support to certain models of DNA catenation organization and regulation: in particular, we conclude from our observation of centromere...

  1. Generating of rice OsCENH3-GFP transgenic plants and their genetic applications

    Institute of Scientific and Technical Information of China (English)

    YU HengXiu; WANG Xin; GONG ZhiYun; TANG Ding; GU MingHong; CHENG ZhuKuan

    2008-01-01

    In order to investigate rice functional centromeres, OsCENH3-GFP chimeric gene was constructed and transformed into the indica rice variety, Zhongxian 3037, mediated by Agrobacturium. The integration of the exogenous genes in the transgenic plants was confirmed by PCR and Southern blotting. The transgenic plants grow normally during their whole life time, just like Zhongxian 3037. No significant defects were detected in either mitosis or meiosis of the transgenic plants. The overlapping of GFP signals and anti-CENH3 foci in both mitotic and meiotic cells from To and T1 generation plants indicated that GFP had been successfully fused with CENH3, so the GFP signals can well represent the CENH3 locations on each chromosome. To evaluate the applicability of the transgenic plants to other genetic studies, fluorescence in situ hybridization (FISH) using rice centromeric tandem repetitive sequence CentO as the probe was conducted on the zygotene chromosomes of pollen mother cells (PMCs). It has been revealed that the GFP signals are overlapping with CentO FISH signals, showing that CentO is one of the key elements constituting rice functional centromeres. Immunofluorescent staining using anti-α-tublin antibody and anti-PAIR2 antibody on the chromosomes during mitosis and meiosis stages of the transgenic plants further reveals that OsCENH3-GFP transgenic plants can be widely used for studying rice molecular biology, especially for tagging functional centromeres in both living cells and tissues.

  2. AcEST: BP914944 [AcEST

    Lifescience Database Archive (English)

    Full Text Available ntromere-associated protein OS=Parascaris univalens Align length 142 Score (bit) 48.1 E-value 2.0e-05 Report...nd centromere-associated protein OS=Parascaris univalens GN=PUMA1 PE=2 SV=1 Lengt

  3. Cytogenetic Characterization of the TM4 Mouse Sertoli Cell Line. II. Chromosome Microdissection, FISH, Scanning Electron Microscopy, and Confocal Laser Scanning Microscopy.

    Science.gov (United States)

    Schmid, Michael; Guttenbach, Martina; Steinlein, Claus; Wanner, Gerhard; Houben, Andreas

    2015-01-01

    The chromosomes and interphase cell nuclei of the permanent mouse Sertoli cell line TM4 were examined by chromosome microdissection, FISH, scanning electron microscopy, and confocal laser scanning microscopy. The already known marker chromosomes m1-m5 were confirmed, and 2 new large marker chromosomes m6 and m7 were characterized. The minute heterochromatic marker chromosomes m4 and m5 were microdissected and their DNA amplified by DOP-PCR. FISH of this DNA probe on TM4 metaphase chromosomes demonstrated that the m4 and m5 marker chromosomes have derived from the centromeric regions of normal telocentric mouse chromosomes. Ectopic pairing of the m4 and m5 marker chromosomes with the centromeric region of any of the other chromosomes (centromeric associations) was apparent in ∼60% of the metaphases. Scanning electron microscopy revealed DNA-protein bridges connecting the centromeric regions of normal chromosomes and the associated m4 and m5 marker chromosomes. Interphase cell nuclei of TM4 Sertoli cells did not exhibit the characteristic morphology of Sertoli cells in the testes of adult mice as shown by fluorescence microscopy and confocal laser scanning microscopy.

  4. Not para-, not peri-, but centric inversion of chromosome 12

    DEFF Research Database (Denmark)

    Silahtaroglu, A N; Hacihanefioglu, S; Güven, G S;

    1998-01-01

    A 39 year old male with primary infertility was diagnosed as having Klinefelter syndrome by conventional cytogenetic analysis, which also showed an abnormal chromosome 12. Fluorescence in situ hybridisation (FISH) analysis of the aberrant chromosome using a 12 specific centromeric probe showed...

  5. The Karyotypes,C-banding Patterns and AgNORs of Epinephelus malabaricus

    Institute of Scientific and Technical Information of China (English)

    Zou Jixing(邹记兴); Hu Chaoqun; Xiang Wenzhou; Yu Qixing; Zhou Fei

    2004-01-01

    The chromosome specimens of Epinephelus malabaricus (Bloch & Schneider, 1801) are obtained from metaphase of kindney cell by vivi-injection of PHA and culture of colchicines, hypatoic-air drying technique, and then by studying their Giemsa stain, C-bands and AgNORs. The results are as follows: (1)E. malabaricus has a diploid chromosome number of 48 and its karyotype formula is 48t, NF=48, sex chromosome is not found. (2) There is a pair of chromosomes with secondary constriction near the centromere of chromosome t24. (3) 1~4 nucleoli appear in the nucleus of interphase, 55% nuclei has 1 nucleolus and only 2% for 4 nucleoli. (4) AgNORs appear in the chromosome t24 of 50% metaphase, sometimes in the chromosome t5, but not in other chromosomes. (5) The AgNORs polymorphisms are individually specific, 1~4 pairs of the number, and the frequency of 4 AgNORs are lowest. (6) The secondary constrictions and positive C-bands are coincident, close to the centromere of the chromosome, and mass constrictive heterochromatins appear in that region. (7) All the centromeres of chromosomes are darkly stained C-bands, and the whole arm of chromosome t24 and its centromere are same positive C-bands. (8) The evolutive regulation of the karyotype and the developing mechanism of AgNORs and C-bands are discussed.

  6. ZEBRAFISH CHROMOSOME-BANDING

    NARCIS (Netherlands)

    PIJNACKER, LP; FERWERDA, MA

    1995-01-01

    Banding techniques were carried out on metaphase chromosomes of zebrafish (Danio rerio) embryos. The karyotypes with the longest chromosomes consist of 12 metacentrics, 26 submetacentrics, and 12 subtelocentrics (2n = 50). All centromeres are C-band positive. Eight chromosomes have a pericentric C-b

  7. cDNA cloning, mRNA distribution and heterogeneity, chromosomal location, and RFLP analysis of human osteopontin (OPN)

    DEFF Research Database (Denmark)

    Young, M F; Kerr, J M; Termine, J D;

    1990-01-01

    on a region of 4q that is near the centromere. A high-frequency restriction fragment length polymorphism was evident in the DNA from 29 unrelated individuals using the enzyme BglII. Analysis of total genomic DNA by digestion with several restriction enzymes, Southern blotting, and hybridization with the human...

  8. Telomere Length as a Predictor of Aggressive Prostate Cancer

    Science.gov (United States)

    2008-11-01

    45. PMID: 19668150 Vander Griend DJ, Konishi Y, De Marzo AM, Isaacs JT, Meeker AK. Dual-label centromere and telomere FISH identifies human, rat ...K, Rimm EB, Stampfer MJ, Willett WC, Giovannucci E. Statin drugs and risk of advanced prostate cancer. J Natl Cancer Inst 2006;98:1819-25. PMID

  9. The ParB-parS Chromosome Segregation System Modulates Competence Development in Streptococcus pneumoniae

    NARCIS (Netherlands)

    Attaiech, Laetitia; Minnen, Anita; Kjos, Morten; Gruber, Stephan; Veening, Jan-Willem

    2015-01-01

    UNLABELLED: ParB proteins bind centromere-like DNA sequences called parS sites and are involved in plasmid and chromosome segregation in bacteria. We previously showed that the opportunistic human pathogen Streptococcus pneumoniae contains four parS sequences located close to the origin of replicati

  10. Test: Construction of genomic libraries.

    Science.gov (United States)

    Szeberényi, Jozséf

    2005-03-01

    Terms to be familiar with before you start to solve the test: genomic library, gel filtration, restriction endonuclease, plasmid, sticky ends, blunt ends, ligation, recombinant DNA, bacterium transformation, denaturation and renaturation of DNA, satellite DNA, telomere, centromere, unique and repetitive sequences.

  11. The split personality of CENP-A nucleosomes.

    Science.gov (United States)

    Westhorpe, Frederick G; Straight, Aaron F

    2012-07-20

    The composition and structure of centromeric nucleosomes, which contain the histone H3 variant CENP-A, is intensely debated. Two independent studies in this issue, in yeast and human cells, now suggest that CENP-A nucleosomes adopt different structures depending on the stage of the cell cycle.

  12. FISH and DAPI staining of the synaptonemal complex of the Nile tilapia (Oreochromis niloticus) allow orientation of the unpaired region of bivalent 1 observed during early pachytene.

    Science.gov (United States)

    Ocalewicz, Konrad; Mota-Velasco, Jose C; Campos-Ramos, Rafael; Penman, David J

    2009-01-01

    Bivalent 1 of the synaptonemal complex (SC) in XY male Oreochromis niloticus shows an unpaired terminal region in early pachytene. This appears to be related to recombination suppression around a sex determination locus. To allow more detailed analysis of this, and unpaired regions in the karyotype of other Oreochromis species, we developed techniques for FISH on SC preparations, combined with DAPI staining. DAPI staining identified presumptive centromeres in SC bivalents, which appeared to correspond to the positions observed in the mitotic karyotype (the kinetochores could be identified only sporadically in silver-stained EM SC images). Furthermore, two BAC clones containing Dmo (dmrt4) and OniY227 markers that hybridize to known positions in chromosome pair 1 in mitotic spreads (near the centromere, Flpter 0.25, and the putative sex-determination locus, Flpter 0.57, respectively) were used as FISH probes on SCs to verify that the presumptive centromere identified by DAPI staining was located in the expected position. Visualization of both the centromere and FISH signals on bivalent 1 allowed the unpaired region to be positioned at Flpter 0.80 to 1.00, demonstrating that the unpaired region is located in the distal part of the long arm(s). Finally, differences between mitotic and meiotic measurements are discussed.

  13. Ndc10 is a platform for inner kinetochore assembly in budding yeast

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Uhn-Soo; Harrison, Stephen C. (Harvard-Med)

    2012-01-10

    Kinetochores link centromeric DNA to spindle microtubules and ensure faithful chromosome segregation during mitosis. In point-centromere yeasts, the CBF3 complex Skp1-Ctf13-(Cep3){sub 2}-(Ndc10){sub 2} recognizes a conserved centromeric DNA element through contacts made by Cep3 and Ndc10. We describe here the five-domain organization of Kluyveromyces lactis Ndc10 and the structure at 2.8 {angstrom} resolution of domains I-II (residues 1-402) bound to DNA. The structure resembles tyrosine DNA recombinases, although it lacks both endonuclease and ligase activities. Structural and biochemical data demonstrate that each subunit of the Ndc10 dimer binds a separate fragment of DNA, suggesting that Ndc10 stabilizes a DNA loop at the centromere. We describe in vitro association experiments showing that specific domains of Ndc10 interact with each of the known inner-kinetochore proteins or protein complexes in budding yeast. We propose that Ndc10 provides a central platform for inner-kinetochore assembly.

  14. Looping in on Ndc80 - how does a protein loop at the kinetochore control chromosome segregation?

    DEFF Research Database (Denmark)

    Nilsson, Jakob

    2012-01-01

    Segregation of chromosomes during mitosis requires the interaction of dynamic microtubules with the kinetochore, a large protein structure established on the centromere region of sister chromatids. The core microtubule-binding activity of the kinetochore resides in the KMN network, an outer...

  15. Histone modifications: Cycling with chromosomal replication

    DEFF Research Database (Denmark)

    Thon, Genevieve

    2008-01-01

    Histone modifications tend to be lost during chromosome duplication. Several recent studies suggest that the RNA interference pathway becomes active during the weakened transcriptional repression occurring at centromeres in S phase, resulting in the re-establishment of histone modifications that ...

  16. Acquisition and processing of a conditional dicentric chromosome in Saccharomyces cerevisiae.

    OpenAIRE

    Hill, A.; Bloom, K.

    1989-01-01

    The introduction of a conditional centromere into chromosome III of Saccharomyces cerevisiae provided an opportunity to evaluate phenotypic and karyotypic consequences in cells harboring dicentric chromosomes upon entry into mitosis. A mitotic pause ensued, and monocentric derivatives of chromosome III were generated at a high frequency.

  17. Increase in dicentric chromosome formation after a single CT scan in adults.

    Science.gov (United States)

    Abe, Yu; Miura, Tomisato; Yoshida, Mitsuaki A; Ujiie, Risa; Kurosu, Yumiko; Kato, Nagisa; Katafuchi, Atsushi; Tsuyama, Naohiro; Ohba, Takashi; Inamasu, Tomoko; Shishido, Fumio; Noji, Hideyoshi; Ogawa, Kazuei; Yokouchi, Hiroshi; Kanazawa, Kenya; Ishida, Takashi; Muto, Satoshi; Ohsugi, Jun; Suzuki, Hiroyuki; Ishikawa, Tetsuo; Kamiya, Kenji; Sakai, Akira

    2015-09-09

    Excess risk of leukemia and brain tumors after CT scans in children has been reported. We performed dicentric chromosome assay (DCAs) before and after CT scan to assess effects of low-dose ionizing radiation on chromosomes. Peripheral blood (PB) lymphocytes were collected from 10 patients before and after a CT scan. DCA was performed by analyzing either 1,000 or 2,000 metaphases using both Giemsa staining and centromere-fluorescence in situ hybridization (Centromere-FISH). The increment of DIC formation was compared with effective radiation dose calculated using the computational dosimetry system, WAZA-ARI and dose length product (DLP) in a CT scan. Dicentric chromosome (DIC) formation increased significantly after a single CT scan, and increased DIC formation was found in all patients. A good correlation between the increment of DIC formation determined by analysis of 2,000 metaphases using Giemsa staining and those by 2,000 metaphases using Centromere-FISH was observed. However, no correlation was observed between the increment of DIC formation and the effective radiation dose. Therefore, these results suggest that chromosome cleavage may be induced by one CT scan, and we recommend 2,000 or more metaphases be analyzed in Giemsa staining or Centromere-FISH for DCAs in cases of low-dose radiation exposure.

  18. 45,X/46,XY Turner Syndrome: A Psu dic(y) can appear normal by G-banding

    Energy Technology Data Exchange (ETDEWEB)

    Higgins, J.V.; Gutai, J.; Shreeve, J.

    1994-09-01

    Mosaic Turner syndrome is a common finding with 50% of the individuals with Turner syndrome having a second cell line. In rare cases, the additional chromosome is reported to be a Y or variant of the Y chromosome. We report a female with Turner Syndrome, who has a mosaic pattern, with the Y chromosome by GTL-banding appearing entirely normal, about the length of chromosome 22. Quinicrine-banding revealed no heterochromatic region. The father of the girl was unavailable for analysis. C-banding resulted in the determination of two centromeric regions, one active at the primary constriction and the second was inactive. The Y centromeric probe (Oncor) showed both the active and inactive centromeres to be of Y origin. Painting using whole chromosome Y paint (Imagenetics) revealed only Y material. In cases of males and females where 45,X/46,X,dic(Y) has been reported, a mosaic pattern has resulted from both centromeres being active in early embryogenesis. In this case the resulting chromosome looks to be a normal Y in both shape and size but with a mosaic pattern. Clearly, G-banding is not always adequate to determine a dic(Y). This suggests that any 45,X/46,SY mosaic pattern in either males or females should be evaluated for a possible dicentric Y chromosome.

  19. GLO1 — EDRN Public Portal

    Science.gov (United States)

    From NCBI Gene: The enzyme encoded by this gene is responsible for the catalysis and formation of S-lactoyl-glutathione from methylglyoxal condensation and reduced glutatione. Glyoxalase I is linked to HLA and is localized to 6p21.3-p21.1, between HLA and the centromere. [provided by RefSeq, Jul 2008

  20. ICBP90 Regulation of DNA Methylation, Histone Ubiquitination, and Tumor Suppressor Gene Expression in Breast Cancer Cells

    Science.gov (United States)

    2012-07-01

    targeted for site- directed mutagenesis is indicated with a red X. liquid chromatography followed by tandem mass spectrometry. The two tags...type and VIM1 RING domain mutants will be examined via cytological techniques such as fluorescence in situ hybridization (FISH) against centromeric

  1. A dual inhibitory mechanism sufficient to maintain cell cycle restricted CENP-A assembly

    Science.gov (United States)

    Stankovic, Ana; Guo, Lucie Y.; Mata, João F.; Bodor, Dani L.; Cao, Xing-Jun; Bailey, Aaron O.; Shabanowitz, Jeffrey; Hunt, Donald F.; Garcia, Benjamin A.; Black, Ben E.; Jansen, Lars E.T

    2017-01-01

    Summary Chromatin featuring the H3 variant CENP-A at the centromere is critical for its mitotic function and epigenetic maintenance. Assembly of centromeric chromatin is restricted to G1 phase through inhibitory action of Cdk1/2 kinases in other phases of the cell cycle. Here, we identify the two key targets sufficient to maintain cell cycle control of CENP-A assembly. We uncovered a single phosphorylation site in the licensing factor M18BP1 and a cyclin A binding site in the CENP-A chaperone, HJURP, mediating specific inhibitory phosphorylation. Simultaneous expression of mutant proteins lacking these residues, results in complete uncoupling from the cell cycle. Consequently, CENP-A assembly is fully recapitulated under high Cdk activities, indistinguishable from G1 assembly. We find that Cdk-mediated inhibition is exerted by sequestering active factors away from the centromere. Finally, we show that displacement of M18BP1 from the centromere is critical for the assembly mechanism of CENP-A. PMID:28017591

  2. Mammalian satellite DNA: a speaking dumb.

    Science.gov (United States)

    Enukashvily, Natella I; Ponomartsev, Nikita V

    2013-01-01

    The tandemly organized highly repetitive satellite DNA is the main DNA component of centromeric/pericentromeric constitutive heterochromatin. For almost a century, it was considered as "junk DNA," only a small portion of which is used for kinetochore formation. The current review summarizes recent data about satellite DNA transcription. The possible functions of the transcripts are discussed.

  3. Genetic mapping in the Mexican axolotl, Ambystoma mexicanum.

    Science.gov (United States)

    Armstrong, J B

    1984-02-01

    In the Mexican axolotl, Ambystoma mexicanum, gynogenetic diploids can be produced by suppressing the release of the second polar body in eggs activated with irradiated sperm. If the female is heterozygous for a particular mutation, some of the progeny will be homozygous for the mutation. The proportion depends on the distance from the centromere and can be used to determine the gene--centromere (or gene-kinetochore) distance. The mapping function is based on the Neurospora tetrad mapping function. Several variations on this function, based on considerations of how coincidence varies with map distance, are considered. Three genes have been mapped: c at 5.9, t at 24.3, and m at 59.1 map units from their respective centromeres. Four other genes (a, ax, p, and the sex locus) appear to be distant from their centromeres but precise map distances cannot be determined. Based on these data, the total length of the genome has been estimated as at least 2600 map units.

  4. Assessing Telomere Length Using Surface Enhanced Raman Scattering

    Science.gov (United States)

    Zong, Shenfei; Wang, Zhuyuan; Chen, Hui; Cui, Yiping

    2014-11-01

    Telomere length can provide valuable insight into telomeres and telomerase related diseases, including cancer. Here, we present a brand-new optical telomere length measurement protocol using surface enhanced Raman scattering (SERS). In this protocol, two single strand DNA are used as SERS probes. They are labeled with two different Raman molecules and can specifically hybridize with telomeres and centromere, respectively. First, genome DNA is extracted from cells. Then the telomere and centromere SERS probes are added into the genome DNA. After hybridization with genome DNA, excess SERS probes are removed by magnetic capturing nanoparticles. Finally, the genome DNA with SERS probes attached is dropped onto a SERS substrate and subjected to SERS measurement. Longer telomeres result in more attached telomere probes, thus a stronger SERS signal. Consequently, SERS signal can be used as an indicator of telomere length. Centromere is used as the inner control. By calibrating the SERS intensity of telomere probe with that of the centromere probe, SERS based telomere measurement is realized. This protocol does not require polymerase chain reaction (PCR) or electrophoresis procedures, which greatly simplifies the detection process. We anticipate that this easy-operation and cost-effective protocol is a fine alternative for the assessment of telomere length.

  5. Surface Enhanced Raman Scattering Based in Situ Hybridization Strategy for Telomere Length Assessment.

    Science.gov (United States)

    Zong, Shenfei; Chen, Chen; Wang, Zhuyuan; Zhang, Yizhi; Cui, Yiping

    2016-02-23

    Assessing telomere length is of vital importance since telomere length is closely related with several fatal diseases such as atherosclerosis and cancer. Here, we present a strategy to assess/measure telomere length, that is, surface enhanced Raman scattering (SERS) based in situ hybridization (SISH). The SISH method uses two kinds of SERS nanoprobes to hybridize in situ with telomeres and centromeres, respectively. The telomere specific SERS nanoprobe is called the Telo-probe, while the centromere specific SERS nanoprobe is called the Centro-probe. They are composed of metal nanoparticles (NPs), Raman reporter molecules and specially designed DNA strands. With longer telomeres, more Telo-probes will hybridize with them, resulting in a stronger SERS signal. To exclude possible influence of the SERS intensity by external factors (such as the nanoprobe concentration, the cell number or different batches of nanoprobes), centromeres are used as the inner control, which can be recognized by Centro-probes. Telomere length is evaluated using a redefined telomere-to-centromere ratio (T/C ratio). The calculation method for T/C ratio in SISH method is more reliable than that in fluorescent in situ hybridization (FISH). In addition, unlike FISH method, the SISH method is insensitive to autofluorescence. Moreover, SISH method can be used to analyze single telomeres. These features make SISH an excellent alternative strategy for telomere length measurement.

  6. Meiotic abnormalities

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1993-12-31

    Chapter 19, describes meiotic abnormalities. These include nondisjunction of autosomes and sex chromosomes, genetic and environmental causes of nondisjunction, misdivision of the centromere, chromosomally abnormal human sperm, male infertility, parental age, and origin of diploid gametes. 57 refs., 2 figs., 1 tab.

  7. Separase Cleaves the N-Tail of the CENP-A Related Protein CPAR-1 at the Meiosis I Metaphase-Anaphase Transition in C. elegans.

    Directory of Open Access Journals (Sweden)

    Joost Monen

    Full Text Available Centromeres are defined epigenetically in the majority of eukaryotes by the presence of chromatin containing the centromeric histone H3 variant CENP-A. Most species have a single gene encoding a centromeric histone variant whereas C. elegans has two: HCP-3 (also known as CeCENP-A and CPAR-1. Prior RNAi replacement experiments showed that HCP-3 is the functionally dominant isoform, consistent with CPAR-1 not being detectable in embryos. GFP::CPAR-1 is loaded onto meiotic chromosomes in diakinesis and is enriched on bivalents until meiosis I. Here we show that GFP::CPAR-1 signal loss from chromosomes precisely coincides with homolog segregation during anaphase I. This loss of GFP::CPAR-1 signal reflects proteolytic cleavage between GFP and the histone fold of CPAR-1, as CPAR-1::GFP, in which GFP is fused to the C-terminus of CPAR-1, does not exhibit any loss of GFP signal. A focused candidate screen implicated separase, the protease that initiates anaphase by cleaving the kleisin subunit of cohesin, in this cleavage reaction. Examination of the N-terminal tail sequence of CPAR-1 revealed a putative separase cleavage site and mutation of the signature residues in this site eliminated the cleavage reaction, as visualized by retention of GFP::CPAR-1 signal on separating homologous chromosomes at the metaphase-anaphase transition of meiosis I. Neither cleaved nor uncleavable CPAR-1 were centromere-localized in mitosis and instead localized throughout chromatin, indicating that centromere activity has not been retained in CPAR-1. Although the functions of CPAR-1 and of its separase-dependent cleavage remain to be elucidated, this effort reveals a new substrate of separase and provides an in vivo biosensor to monitor separase activity at the onset of meiosis I anaphase.

  8. Are ribosomal DNA clusters rearrangement hotspots? A case study in the genus Mus (Rodentia, Muridae

    Directory of Open Access Journals (Sweden)

    Douzery Emmanuel JP

    2011-05-01

    Full Text Available Abstract Background Recent advances in comparative genomics have considerably improved our knowledge of the evolution of mammalian karyotype architecture. One of the breakthroughs was the preferential localization of evolutionary breakpoints in regions enriched in repetitive sequences (segmental duplications, telomeres and centromeres. In this context, we investigated the contribution of ribosomal genes to genome reshuffling since they are generally located in pericentromeric or subtelomeric regions, and form repeat clusters on different chromosomes. The target model was the genus Mus which exhibits a high rate of karyotypic change, a large fraction of which involves centromeres. Results The chromosomal distribution of rDNA clusters was determined by in situ hybridization of mouse probes in 19 species. Using a molecular-based reference tree, the phylogenetic distribution of clusters within the genus was reconstructed, and the temporal association between rDNA clusters, breakpoints and centromeres was tested by maximum likelihood analyses. Our results highlighted the following features of rDNA cluster dynamics in the genus Mus: i rDNA clusters showed extensive diversity in number between species and an almost exclusive pericentromeric location, ii a strong association between rDNA sites and centromeres was retrieved which may be related to their shared constraint of concerted evolution, iii 24% of the observed breakpoints mapped near an rDNA cluster, and iv a substantial rate of rDNA cluster change (insertion, deletion also occurred in the absence of chromosomal rearrangements. Conclusions This study on the dynamics of rDNA clusters within the genus Mus has revealed a strong evolutionary relationship between rDNA clusters and centromeres. Both of these genomic structures coincide with breakpoints in the genus Mus, suggesting that the accumulation of a large number of repeats in the centromeric region may contribute to the high level of chromosome

  9. Adaptive changes in the kinetochore architecture facilitate proper spindle assembly.

    Science.gov (United States)

    Magidson, Valentin; Paul, Raja; Yang, Nachen; Ault, Jeffrey G; O'Connell, Christopher B; Tikhonenko, Irina; McEwen, Bruce F; Mogilner, Alex; Khodjakov, Alexey

    2015-09-01

    Mitotic spindle formation relies on the stochastic capture of microtubules at kinetochores. Kinetochore architecture affects the efficiency and fidelity of this process with large kinetochores expected to accelerate assembly at the expense of accuracy, and smaller kinetochores to suppress errors at the expense of efficiency. We demonstrate that on mitotic entry, kinetochores in cultured human cells form large crescents that subsequently compact into discrete structures on opposite sides of the centromere. This compaction occurs only after the formation of end-on microtubule attachments. Live-cell microscopy reveals that centromere rotation mediated by lateral kinetochore-microtubule interactions precedes the formation of end-on attachments and kinetochore compaction. Computational analyses of kinetochore expansion-compaction in the context of lateral interactions correctly predict experimentally observed spindle assembly times with reasonable error rates. The computational model suggests that larger kinetochores reduce both errors and assembly times, which can explain the robustness of spindle assembly and the functional significance of enlarged kinetochores.

  10. Two siblings with immunodeficiency, facial abnormalities and chromosomal instability without mutation in DNMT3B gene but liability towards malignancy; a new chromatin disorder delineation?

    Directory of Open Access Journals (Sweden)

    Neitzel Heidemarie

    2010-03-01

    Full Text Available Abstract Background ICF syndrome (standing for Immunodeficiency, Centromere instability and Facial anomalies syndrome is a very rare autosomal recessive immune disorder caused by mutations of the gene de novo DNA-methyltransferase 3B (DNMT3B. However, in the literature similar clinical cases without such mutations are reported, as well. Results We report on a family in which the unrelated spouses had two female siblings sharing similar phenotypic features resembling ICF-syndrome, i.e. congenital abnormalities, immunodeficiency, developmental delay and high level of chromosomal instability, including high frequency of centromeric/pericentromeric rearrangements and breaks, chromosomal fragments despiralization or pulverization. However, mutations in DNMT3B could not be detected. Conclusion The discovery of a new so-called 'chromatin disorder' is suggested. Clinical, molecular genetic and cytogenetic characteristics are reported and compared to other 'chromatin disorders'.

  11. Comparison of Two Alternative Dominant Selectable Markers for Wine Yeast Transformation

    Science.gov (United States)

    Cebollero, Eduardo; Gonzalez, Ramon

    2004-01-01

    Genetic improvement of industrial yeast strains is restricted by the availability of selectable transformation markers. Antibiotic resistance markers have to be avoided for public health reasons, while auxotrophy markers are generally not useful for wine yeast strain transformation because most industrial Saccharomyces cerevisiae strains are prototrophic. For this work, we performed a comparative study of the usefulness of two alternative dominant selectable markers in both episomic and centromeric plasmids. Even though the selection for sulfite resistance conferred by FZF1-4 resulted in a larger number of transformants for a laboratory strain, the p-fluoro-dl-phenylalanine resistance conferred by ARO4-OFP resulted in a more suitable selection marker for all industrial strains tested. Both episomic and centromeric constructions carrying this marker resulted in transformation frequencies close to or above 103 transformants per μg of DNA for the three wine yeast strains tested. PMID:15574895

  12. Cytogenetic analysis in two scallops (Bivalvia: Pectinidae) by PRINS and PI banding

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Cytogenetic analysis was performed for the bay scallop (Argopecten irradians Lamarck 1819) and the Japanese scallop (Patinopecten yessoensis Jay 1857) by primed in situ labeling (PRINS) and propidium iodide (PI) banding techniques. The PRINS analysis revealed that major rRNA genes were clustered in two loci on the telomeric regions of the short arms on two acrocentric chromosome pairs in A. irradians and on two submetacentric pairs in P. yessoensis. The histone H3 gene sites differed in number and location between these two species. The C-band-like patterns revealed by PI staining varied considerably between these two species. A. irradians displayed terminal bands at long arms on all chromosomes, centromeric bands on some pairs and interstitial bands on five pairs. P. yessoensis exhibited only centromeric bands on all chromosomes. These results would contribute to the better understanding of karyotype evolution in A. irradians and P. yessoensis.

  13. Systematic hybrid LOH: a new method to reduce false positives and negatives during screening of yeast gene deletion libraries

    DEFF Research Database (Denmark)

    Alvaro, D.; Sunjevaric, I.; Reid, R. J.;

    2006-01-01

    We have developed a new method, systematic hybrid loss of heterozygosity, to facilitate genomic screens utilizing the yeast gene deletion library. Screening is performed using hybrid diploid strains produced through mating the library haploids with strains from a different genetic background......, to minimize the contribution of unpredicted recessive genetic factors present in the individual library strains. We utilize a set of strains where each contains a conditional centromere construct on one of the 16 yeast chromosomes that allows the destabilization and selectable loss of that chromosome. After...... mating a library gene deletion haploid to such a conditional centromere strain, which corresponds to the chromosome carrying the gene deletion, loss of heterozygosity (LOH) at the gene deletion locus can be generated in these otherwise hybrid diploids. The use of hybrid diploid strains permits...

  14. The molecular characterization of maize B chromosome specific AFLPs

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The origin and evolution of B chromosomes could be explained by the specific DNA sequence on them.But the specific sequences known were quite limited. To investigate maize B chromosome sqicific DNA sequeces, maize genomes with and without B chromosomes were analyzed by AFLP. Only 5 markers were found specific to genomes with B chromosomes among about 2000 AFLP markers. Southern hybridization and sequence analysis revealed that only the sequence of M8-2D was a B chromosome specific sequence.This sequence contained the telomeric repeat unit AGGGTTT conserved in plant chromosome telomeres.In addition, the sequence of M8-2D shared low homology to clones from maize chromosome 4 centromere as well. M8-2D were localized to B chromosome centromeric and telomeric regions.

  15. A highly conserved pericentromeric domain in human and gorilla chromosomes.

    Science.gov (United States)

    Pita, M; Gosálvez, J; Gosálvez, A; Nieddu, M; López-Fernández, C; Mezzanotte, R

    2009-01-01

    Significant similarity between human and gorilla genomes has been found in all chromosome arms, but not in centromeres, using whole-comparative genomic hybridization (W-CGH). In human chromosomes, centromeric regions, generally containing highly repetitive DNAs, are characterized by the presence of specific human DNA sequences and an absence of homology with gorilla DNA sequences. The only exception is the pericentromeric area of human chromosome 9, which, in addition to a large block of human DNA, also contains a region of homology with gorilla DNA sequences; the localization of these sequences coincides with that of human satellite III. Since highly repetitive DNAs are known for their high mutation frequency, we hypothesized that the chromosome 9 pericentromeric DNA conserved in human chromosomes and deriving from the gorilla genome may thus play some important functional role.

  16. In vitro kinetochore assembly

    Science.gov (United States)

    Miell, Matthew D D; Straight, Aaron F

    2016-01-01

    The kinetochore is the primary site of interaction between chromosomes and microtubules of the mitotic spindle during chromosome segregation. The kinetochore is a complex of more than 100 proteins that transiently assemble during mitosis at a single defined region on each chromosome, known as the centromere. Kinetochore assembly and activity must be tightly regulated to ensure proper microtubule interaction and faithful chromosome segregation because perturbation of kinetochores often results in aneuploidy and cell lethality. As such, cell free and reconstituted systems to analyze kinetochore formation and function are invaluable in probing the biochemical activities of kinetochores. In vitro approaches to studying kinetochores have enabled the manipulation of kinetochore protein structure, function, interactions and regulation that are not possible in cells. Here we outline a cell-free approach for the assembly of centromeres and recruitment of functional kinetochores that enables their manipulation and analysis. PMID:27193846

  17. The characteristics of the spindle fiber attachments(SFAs) enriched fraction from mouse nuclei

    Institute of Scientific and Technical Information of China (English)

    NIZUMEI; NINGYI; 等

    1993-01-01

    The characteristics of the particulate mouse centromere enriched fraction from isolated nuclei obtained in our laboratory were investigated by indirect immunofluorescence,test of the activity of microtubule organizing center(MTOC),SDS-PAGE,and fluorescence in situ hybridization.Most of the particles of the fraction are complexes of DNA and kinetochore proteins and show MTOC activity.The DNA isolated from the fraction can hybridize with DNA in the regions of the primary constrictions of all chromosomes of ascions of the primary constrictions of all chromosomes of ascites cells.The kinetochore proteins isolated from the fraction are mainly those with molecular weight of 55 KD and 59 KD.Results suggested that the fraction obtained is a centromere enriched nuclear fraction as indicated in our previous report.

  18. Plant minichromosomes.

    Science.gov (United States)

    Birchler, James A; Graham, Nathaniel D; Swyers, Nathan C; Cody, Jon P; McCaw, Morgan E

    2016-02-01

    Plant minichromosomes have the potential for stacking multiple traits on a separate entity from the remainder of the genome. Transgenes carried on an independent chromosome would facilitate conferring many new properties to plants and using minichromosomes as genetic tools. The favored method for producing plant minichromosomes is telomere-mediated chromosomal truncation because the epigenetic nature of centromere function prevents using centromere sequences to confer the ability to organize a kinetochore when reintroduced into plant cells. Because haploid induction procedures are not always complete in eliminating one parental genome, chromosomes from the inducer lines are often present in plants that are otherwise haploid. This fact suggests that minichromosomes could be combined with doubled haploid breeding to transfer stacked traits more easily to multiple lines and to use minichromosomes for massive scale genome editing.

  19. Sister chromatid segregation in meiosis II: deprotection through phosphorylation.

    Science.gov (United States)

    Wassmann, Katja

    2013-05-01

    Meiotic divisions (meiosis I and II) are specialized cell divisions to generate haploid gametes. The first meiotic division with the separation of chromosomes is named reductional division. The second division, which takes place immediately after meiosis I without intervening S-phase, is equational, with the separation of sister chromatids, similar to mitosis. This meiotic segregation pattern requires the two-step removal of the cohesin complex holding sister chromatids together: cohesin is removed from chromosome arms that have been subjected to homologous recombination in meiosis I and from the centromere region in meiosis II. Cohesin in the centromere region is protected from removal in meiosis I, but this protection has to be removed--deprotected--for sister chromatid segregation in meiosis II. Whereas the mechanisms of cohesin protection are quite well understood, the mechanisms of deprotection have been largely unknown until recently. In this review I summarize our current knowledge on cohesin deprotection.

  20. Polo kinase Cdc5 is a central regulator of meiosis I.

    Science.gov (United States)

    Attner, Michelle A; Miller, Matthew P; Ee, Ly-sha; Elkin, Sheryl K; Amon, Angelika

    2013-08-27

    During meiosis, two consecutive rounds of chromosome segregation yield four haploid gametes from one diploid cell. The Polo kinase Cdc5 is required for meiotic progression, but how Cdc5 coordinates multiple cell-cycle events during meiosis I is not understood. Here we show that CDC5-dependent phosphorylation of Rec8, a subunit of the cohesin complex that links sister chromatids, is required for efficient cohesin removal from chromosome arms, which is a prerequisite for meiosis I chromosome segregation. CDC5 also establishes conditions for centromeric cohesin removal during meiosis II by promoting the degradation of Spo13, a protein that protects centromeric cohesin during meiosis I. Despite CDC5's central role in meiosis I, the protein kinase is dispensable during meiosis II and does not even phosphorylate its meiosis I targets during the second meiotic division. We conclude that Cdc5 has evolved into a master regulator of the unique meiosis I chromosome segregation pattern.

  1. The Aurora B kinase in chromosome biorientation and spindle checkpoint signalling

    Directory of Open Access Journals (Sweden)

    Veronica eKrenn

    2015-10-01

    Full Text Available Aurora B, a member of the Aurora family of serine/threonine protein kinases, is a key player in chromosome segregation. As part of a macromolecular complex known as the chromosome passenger complex, Aurora B concentrates early during mitosis in the proximity of centromeres and kinetochores, the sites of attachment of chromosomes to spindle microtubules. There, it contributes to a number of processes that impart fidelity to cell division, including kinetochore stabilization, kinetochore-microtubule attachment, and the regulation of a surveillance mechanism named the spindle assembly checkpoint. In the regulation of these processes, Aurora B is the fulcrum of a remarkably complex network of interactions that feed back on its localization and activation state. In this review we discuss the multiple roles of Aurora B during mitosis, focusing in particular on its role at centromeres and kinetochores. Many details of the network of interactions at these locations remain poorly understood, and we focus here on several crucial outstanding questions.

  2. Characterization of a highly repeated DNA sequence family in five species of the genus Eulemur.

    Science.gov (United States)

    Ventura, M; Boniotto, M; Cardone, M F; Fulizio, L; Archidiacono, N; Rocchi, M; Crovella, S

    2001-09-19

    The karyotypes of Eulemur species exhibit a high degree of variation, as a consequence of the Robertsonian fusion and/or centromere fission. Centromeric and pericentromeric heterochromatin of eulemurs is constituted by highly repeated DNA sequences (including some telomeric TTAGGG repeats) which have so far been investigated and used for the study of the systematic relationships of the different species of the genus Eulemur. In our study, we have cloned a set of repetitive pericentromeric sequences of five Eulemur species: E. fulvus fulvus (EFU), E. mongoz (EMO), E. macaco (EMA), E. rubriventer (ERU), and E. coronatus (ECO). We have characterized these clones by sequence comparison and by comparative fluorescence in situ hybridization analysis in EMA and EFU. Our results showed a high degree of sequence similarity among Eulemur species, indicating a strong conservation, within the five species, of these pericentromeric highly repeated DNA sequences.

  3. [Rapid dicentric assay of human blood lymphocytes after exposure to low doses of ionizing radiation].

    Science.gov (United States)

    Repin, M V; Repina, L A

    2011-01-01

    The probability of losses of different chromosome aberrations during the dicentric chromosome assay of metaphase cells with incomplete sets of chromosome centromeres was estimated using a mathematical model for low doses of ionizing radiation. A dicentric assay of human blood lymphocytes without determination of the total amount of chromosome centromeres in cells without chromosome aberrations (rapid dicentric assay) has been proposed. The rapid dicentric analysis allows to register chromosome aberrations in full compliance with the conventional classification. The experimental data have shown no statistically significant difference between the frequencies of dicentric chromosomes detected by rapid and classical dicentric chromosome assays of human lymphocytes exposed to 0.5 Gy of 60Co gamma-rays. The rate of the rapid dicentric assay was almost twice as high as that of the classical dicentric assay.

  4. Homologies between human and marmoset (Callithrix jacchus) chromosomes revealed by comparative chromosome painting

    Energy Technology Data Exchange (ETDEWEB)

    Sherlock, J.K.; Griffin, D.K.; Delhanty, J.D.A.; Parrington, J.M. [Univ. College London (United Kingdom)

    1996-04-15

    Regions of DNA homology between human and marmoset (Callithrix jacchus) chromosomes have been demonstrated using fluorescence in situ hybridization. All 24 chromosome paints and two centromere repeat sequences from Homo sapiens (HSA) have been annealed to previously G-banded metaphase spreads of Callithrix jacchus. All human paint probes, except Y, successfully hybridized to marmoset chromosomes. Fifteen of them hybridized to one region only, seven to two regions, and paint 1 to three regions. Homologies proposed from previous banding comparisons have been confirmed for HSA 2, 4-6, 10-12, 18, 19, 21, and X and partially confirmed for HSA 1 and 3, but were not in agreement for HSA 14 and 17. Human centromere repeat sequences for X and 18 did not hybridize to marmoset chromosomes. Because, at present, there is the confusing situation of several different numbering systems for marmoset chromosomes, we propose a new simpler nomenclature based on descending order of chromosome size. 25 refs., 3 figs.

  5. Relationship of Extreme Chromosomal Instability with Long-term Survival in a Retrospective Analysis of Primary Breast Cancer

    DEFF Research Database (Denmark)

    Roylance, Rebecca; Endesfelder, David; Gorman, Patricia

    2011-01-01

    methods, such as centromeric FISH, aimed at determining the variation around the modal number of two or more chromosomes within individual tumor nuclei. Here, we document the frequency of tumor CIN by dual centromeric FISH analysis in a retrospective primary breast cancer cohort of 246 patients...... with survival outcome. Results: There was increased CIN and clonal eterogeneity in ER-negative compared with ER-positive breast cancer. Consistent with a negative impact of CIN on cellular fitness, extreme CIN in ER-negative breast cancer was an independent variable associated with improved long-term survival......Background: Chromosomal instability (CIN) is thought to be associated with poor prognosis in solid tumors; however, evidence from preclinical and mouse tumor models suggest that CIN may paradoxically enhance or impair cancer cell fitness. Breast cancer prognostic expression signature sets, which...

  6. Donkey genome and insight into the imprinting of fast karyotype evolution.

    Science.gov (United States)

    Huang, Jinlong; Zhao, Yiping; Bai, Dongyi; Shiraigol, Wunierfu; Li, Bei; Yang, Lihua; Wu, Jing; Bao, Wuyundalai; Ren, Xiujuan; Jin, Burenqiqige; Zhao, Qinan; Li, Anaer; Bao, Sarula; Bao, Wuyingga; Xing, Zhencun; An, Aoruga; Gao, Yahan; Wei, Ruiyuan; Bao, Yirugeletu; Bao, Taoketao; Han, Haige; Bai, Haitang; Bao, Yanqing; Zhang, Yuhong; Daidiikhuu, Dorjsuren; Zhao, Wenjing; Liu, Shuyun; Ding, Jinmei; Ye, Weixing; Ding, Fangmei; Sun, Zikui; Shi, Yixiang; Zhang, Yan; Meng, He; Dugarjaviin, Manglai

    2015-09-16

    The donkey, like the horse, is a promising model for exploring karyotypic instability. We report the de novo whole-genome assemblies of the donkey and the Asiatic wild ass. Our results reflect the distinct characteristics of donkeys, including more effective energy metabolism and better immunity than horses. The donkey shows a steady demographic trajectory. We detected abundant satellite sequences in some inactive centromere regions but not in neocentromere regions, while ribosomal RNAs frequently emerged in neocentromere regions but not in the obsolete centromere regions. Expanded miRNA families and five newly discovered miRNA target genes involved in meiosis may be associated with fast karyotype evolution. APC/C, controlling sister chromatid segregation, cytokinesis, and the establishment of the G1 cell cycle phase were identified by analysis of miRNA targets and rapidly evolving genes.

  7. Chromosome analysis of arsenic affected cattle

    Directory of Open Access Journals (Sweden)

    S. Shekhar

    2014-10-01

    Full Text Available Aim: The aim was to study the chromosome analysis of arsenic affected cattle. Materials and Methods: 27 female cattle (21 arsenic affected and 6 normal were selected for cytogenetical study. The blood samples were collected, incubated, and cultured using appropriate media and specific methods. The samples were analyzed for chromosome number and morphology, relative length of the chromosome, arm ratio, and centromere index of X chromosome and chromosomal abnormalities in arsenic affected cattle to that of normal ones. Results: The diploid number of metaphase chromosomes in arsenic affected cattle as well as in normal cattle were all 2n=60, 58 being autosomes and 2 being sex chromosomes. From the centromeric position, karyotyping studies revealed that all the 29 pair of autosomes was found to be acrocentric or telocentric, and the sex chromosomes (XX were submetacentric in both normal and arsenic affected cattle. The relative length of all the autosome pairs and sex chrosomosome pair was found to be higher in normal than that of arsenic affected cattle. The mean arm ratio of X-chromosome was higher in normal than that of arsenic affected cattle, but it is reverse in case of centromere index value of X-chromosome. There was no significant difference of arm ratio and centromere index of X-chromosomes between arsenic affected and normal cattle. No chromosomal abnormalities were found in arsenic affected cattle. Conclusion: The chromosome analysis of arsenic affected cattle in West Bengal reported for the first time in this present study which may serve as a guideline for future studies in other species. These reference values will also help in comparison of cytological studies of arsenic affected cattle to that of various toxicants.

  8. New Insights into Nested Long Terminal Repeat Retrotransposons in Brassica Species

    Institute of Scientific and Technical Information of China (English)

    Lijuan Wei; Meili Xiao; Zeshan An; Bi Ma; Annaliese S.Mason; Wei Qian; Jiana Li

    2013-01-01

    Long terminal repeat (LTR) retrotransposons,one of the foremost types of transposons,continually change or modify gene function and reorganize the genome through bursts of dramatic proliferation.Many LTR-TEs preferentially insert within other LTR-TEs,but the cause and evolutionary significance of these nested LTR-TEs are not well understood.In this study,a total of 1.52 Gb of Brassica sequence containing 2020 bacterial artificial chromosomes (BACs) was scanned,and six bacterial artificial chromosome (BAC) clones with extremely nested LTR-TEs (LTR-TEs density:7.24/kb)were selected for further analysis.The majority of the LTR-TEs in four of the six BACs were found to be derived from the rapid proliferation of retrotransposons originating within the BAC regions,with only a few LTR-TEs originating from the proliferation and insertion of retrotransposons from outside the BAC regions approximately 5-23 Mya.LTR-TEs also preferably inserted into TA-rich repeat regions.Gene prediction by Genescan identified 207 genes in the 0.84 Mb of total BAC sequences.Only a few genes (3/207) could be matched to the Brassica expressed sequence tag (EST) database,indicating that most genes were inactive after retrotransposon insertion.Five of the six BACs were putatively centromeric.Hence,nested LTR-TEs in centromere regions are rapidly duplicated,repeatedly inserted,and act to suppress activity of genes and to reshuffle the structure of the centromeric sequences.Our results suggest that LTR-TEs burst and proliferate on a local scale to create nested LTR-TE regions,and that these nested LTR-TEs play a role in the formation of centromeres.

  9. EST Table: FY014444 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available FY014444 rbmov11f03 11/12/09 GO hit GO:0000775(chromosome, centromeric region)|GO:0...003677(DNA binding) 11/11/04 n.h 11/11/04 n.h 11/11/04 n.h 11/11/04 n.h 11/11/04 n.h 11/11/04 n.h FY014444 bmov ...

  10. Role of DNMT3B in the regulation of early neural and neural crest specifiers

    OpenAIRE

    Martins-Taylor, Kristen; Schroeder, Diane I.; LaSalle, Janine M.; Lalande, Marc; Xu, Ren-He

    2012-01-01

    The de novo DNA methyltransferase DNMT3B functions in establishing DNA methylation patterns during development. DNMT3B missense mutations cause immunodeficiency, centromere instability and facial anomalies (ICF) syndrome. The restriction of Dnmt3b expression to neural progenitor cells, as well as the mild cognitive defects observed in ICF patients, suggests that DNMT3B may play an important role in early neurogenesis. We performed RNAi knockdown of DNMT3B in human embryonic stem cells (hESCs)...

  11. Characters of DNA Constitution in the Rye B Chromosome

    Institute of Scientific and Technical Information of China (English)

    Hong Long; Zhong-Xia Qi; Xiao-Ming Sun; Cheng-Bin Chen; Xiu-Lan Li; Wen-Qin Song; Rui-Yang Chen

    2008-01-01

    We have used chromosome microdissection and microcloning to construct a DNA library of the entire B chromosome (B) of rye. New rye B-specific sequences have been screened from this pool, blasted with other sequences and analyzed to elucidate the characters of DNA constitution and the possible pathway of the origin of the rye B chromosome. We report the discovery of a new sequence that is specific to the rye B centromere.

  12. Giemsa C-banding of Barley Chromosomes. I: Banding Pattern Polymorphism

    DEFF Research Database (Denmark)

    Linde-Laursen, Ib

    1978-01-01

    Twenty barley (Hordeum vulgare) lines studied had a common basic chromosome banding pattern. Most bands ranged from medium to very small in size. The most conspicuous banding occurred at or near the centromeres, in the proximal, intercalary parts of most chromosome arms and beside the secondary c...... 7. Seventeen differently banded karyotypes were found. Some banding pattern polymorphisms can be used in cytological and cytogenetic studies....

  13. SMC is recruited to oriC by ParB and promotes chromosome segregation in Streptococcus pneumoniae

    OpenAIRE

    Minnen, Anita; Attaiech, Laetitia; Thon, Maria; Gruber, Stephan; Veening, Jan-Willem

    2011-01-01

    Segregation of replicated chromosomes is an essential process in all organisms. How bacteria, such as the oval-shaped human pathogen Streptococcus pneumoniae, efficiently segregate their chromosomes is poorly understood. Here we show that the pneumococcal homologue of the DNA-binding protein ParB recruits S. pneumoniae condensin (SMC) to centromere-like DNA sequences (parS) that are located near the origin of replication, in a similar fashion as was shown for the rod-shaped model bacterium Ba...

  14. Isochromosome 13 in a patient with childhood-onset schizophrenia, ADHD, and motor tic disorder

    Directory of Open Access Journals (Sweden)

    Graw Sharon L

    2012-01-01

    Full Text Available Abstract Background A small percentage of all cases of schizophrenia have a childhood onset. The impact on the individual and family can be devastating. We report the results of genetic analyses from a patient with onset of visual hallucinations at 5 years, and a subsequent diagnosis at 9 years of schizophrenia, attention deficit hyperactivity disorder (ADHD with hyperactivity and impulsivity, and chronic motor tic disorder. Results Karyotypic analysis found 45,XX,i(13(q10 in all cells examined. Alpha satellite FISH of isochromosome 13 revealed a large unsplit centromeric region, interpreted as two centromeres separated by minimal or undetectable short-arm material or as a single monocentric centromere, indicating that the isochromosome likely formed post-zygotically by a short arm U-type or centromeric exchange. Characterization of chromosome 13 simple tandem repeats and Affymetrix whole-genome 6.0 SNP array hybridization found homozygosity for all markers, and the presence of only a single paternal allele in informative markers, consistent with an isodisomic isochromosome of paternal origin. Analysis of two chromosome 13 schizophrenia candidate genes, D-amino acid oxidase activator (DAOA and 5-hydroxytryptamine (serotonin receptor 2A (5-HTR2A, failed to identify non-synonymous coding mutations but did identify homozygous risk polymorphisms. Conclusions We report a female patient with childhood-onset schizophrenia, ADHD, and motor tic disorder associated with an isodisomic isochromosome 13 of paternal origin and a 45,XX,i(13(q10q10 karyotype. We examined two potential mechanisms to explain chromosome 13 involvement in the patient's pathology, including reduction to homozygosity of a paternal mutation and reduction to homozygosity of a paternal copy number variation, but were unable to identify any overtly pathogenic abnormality. Future studies may consider whether epigenetic mechanisms resulting from uniparental disomy (UPD and the lack of

  15. Protocols for Pachytene Chromosome and DNA Fiber FISH in Cotton

    Institute of Scientific and Technical Information of China (English)

    PENG Ren-hai; LING Jian; WANG Kun-bo; WANG Chun-ying; SONG Guo-li; LIU Fang

    2008-01-01

    @@ Fluorescence in situ hybridization (FISH) has become the most important technique in plant molecular cytogenetics research.FISH-based physical mapping provides a valuable complementary approach in genome sequeneing to measure the physical distances between adjacent BAC contigs and delineating the structure and DNA composition of genomic regions of centromere and telomere.The accuracy and precision of the FISH-base physical maps depend on the resolution power of FISH.

  16. Review fifteen years of search for strong nucleosomes.

    Science.gov (United States)

    Trifonov, Edward N; Nibhani, Reshma

    2015-08-01

    Don Crothers, Mikael Kubista, Jon Widom, and their teams have been first to look for strong nucleosomes, in a bid to reveal the nucleosome positioning pattern(s) carried by the nucleosome DNA sequences. They were first to demonstrate that the nucleosome stability correlates with 10-11 base sequence periodicity, and that the strong nucleosomes localize preferentially in centromeres. This review describes these findings and their connection to recent discovery of the strong nucleosomes (SNs) with visibly periodic nucleosome DNA sequences.

  17. Automating dicentric chromosome detection from cytogenetic biodosimetry data.

    Science.gov (United States)

    Rogan, Peter K; Li, Yanxin; Wickramasinghe, Asanka; Subasinghe, Akila; Caminsky, Natasha; Khan, Wahab; Samarabandu, Jagath; Wilkins, Ruth; Flegal, Farrah; Knoll, Joan H

    2014-06-01

    We present a prototype software system with sufficient capacity and speed to estimate radiation exposures in a mass casualty event by counting dicentric chromosomes (DCs) in metaphase cells from many individuals. Top-ranked metaphase cell images are segmented by classifying and defining chromosomes with an active contour gradient vector field (GVF) and by determining centromere locations along the centreline. The centreline is extracted by discrete curve evolution (DCE) skeleton branch pruning and curve interpolation. Centromere detection minimises the global width and DAPI-staining intensity profiles along the centreline. A second centromere is identified by reapplying this procedure after masking the first. Dicentrics can be identified from features that capture width and intensity profile characteristics as well as local shape features of the object contour at candidate pixel locations. The correct location of the centromere is also refined in chromosomes with sister chromatid separation. The overall algorithm has both high sensitivity (85 %) and specificity (94 %). Results are independent of the shape and structure of chromosomes in different cells, or the laboratory preparation protocol followed. The prototype software was recoded in C++/OpenCV; image processing was accelerated by data and task parallelisation with Message Passaging Interface and Intel Threading Building Blocks and an asynchronous non-blocking I/O strategy. Relative to a serial process, metaphase ranking, GVF and DCE are, respectively, 100 and 300-fold faster on an 8-core desktop and 64-core cluster computers. The software was then ported to a 1024-core supercomputer, which processed 200 metaphase images each from 1025 specimens in 1.4 h.

  18. Systemic sclerosis with portal hypertensive ascites responded to corticosteroid treatment

    Institute of Scientific and Technical Information of China (English)

    LENG Xiao-mei; SUN Xue-feng; ZHANG Xuan; ZHANG Wen; LI Meng-tao; ZENG Xiao-feng

    2012-01-01

    We describe a case of systemic sclerosis (SSc) complicated with portal hypertensive ascites which did not improve with diuretics and ascitic drainage.When corticosteroid added,her ascites diminished dramatically.Though portal hypertension can be imputed to other causes,such as polycystic liver in this case,it can occur in limited SSc with positive anti-centromere antibody and respond to corticosteroid treatment.

  19. Functional Characterization of CENP-A Post-Translational Modifications in Chromosome Segregation

    Science.gov (United States)

    2014-07-01

    be conducted in year 2 and 3 of this proposal to deduce the relevance of this increased CENP-A methylation during the beginning of mitosis . To... mitosis and accurately segregate chromosomes. Overexpression of CENP-A leads to its mislocalisation and missegregation of chromosomes9. Similarly loss...centromeric chromatin requires exit from mitosis . The Journal of cell biology. 2007;176(6):795-805. 9. Tomonaga T, Matsushita K, Yamaguchi S, Oohashi T

  20. Retrospective study of trisomy 18 in chorionic villi with fluorescent in situ hybridization on archival direct preparations

    OpenAIRE

    Van Opstal, Diane; Berg, Cardi; Jahoda, M.; Brandenburg, Helen; Los, F.J.; in 't Veld, Peter

    1995-01-01

    textabstractTrisomy 18 in direct chorionic villus preparations needs further investigation since the chromosome abnormality may be confined to the placenta and may not represent the actual fetal karyotype. We performed, retrospectively, fluorescent in situ hybridization (FISH) with the chromosome 18 centromere probe (L1.84) on interphase nuclei of destained slides of all cases of full trisomy 18 (n=22) and mosaic trisomy 18 (n=8) detected among 7600 first-trimester chorionic villus samples du...

  1. c-Myc-dependent formation of robertsonian translocation chromosomes in mouse cells

    OpenAIRE

    Amanda Guffei; Zelda Lichtensztejn; Amanda Gonçlves {ptdos} Santos Silva; Louis, Sherif F; Andrea Caporali; Sabine Mai

    2007-01-01

    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 joini...

  2. c-Myc-Dependent Formation of Robertsonian Translocation Chromosomes in Mouse Cells12

    OpenAIRE

    Guffei, Amanda; Lichtensztejn, Zelda; Gonçalves dos Santos Silva, Amanda; Louis, Sherif F; Caporali, Andrea; Mai, Sabine

    2007-01-01

    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...

  3. Evolutionarily different alphoid repeat DNA on homologous chromosomes in human and chimpanzee.

    OpenAIRE

    Jørgensen, A L; Laursen, H B; Jones, C; Bak, A L

    1992-01-01

    Centromeric alphoid DNA in primates represents a class of evolving repeat DNA. In humans, chromosomes 13 and 21 share one subfamily of alphoid DNA while chromosomes 14 and 22 share another subfamily. We show that similar pairwise homogenizations occur in the chimpanzee (Pan troglodytes), where chromosomes 14 and 22, homologous to human chromosomes 13 and 21, share one partially homogenized alphoid DNA subfamily and chromosomes 15 and 23, homologous to human chromosomes 14 and 22, share anothe...

  4. Mutagenicity Evaluation of Ammonium Picrate in the Ames Salmonella/Microsome Plate Test. Segment Report,

    Science.gov (United States)

    1979-02-01

    Dicentric : A chromosome containing two centromeres. > = Greater than 10 aberrations: A cell which contains more than 10 aberrations. *Scored as "AP" on...Chromatid Deletion Ring Dicentric Chromosome Break Trans locati on Tr radial Quadri radial Translocation I Cong 1ex Rearrangenlent PROTOCOL 1...frequency and dose: SCE/ chromosome = 0.0006 D + 0.303, where D = dose in Pg/ ml, and the correlation factor (r) = 0.88. Therefore, ammonium picrate

  5. Genotoxicity Assessment of Mixed Oligomers of Chlorotrifluoroethylene using a Battery of In Vitro and In Vivo/In Vitro Assays

    Science.gov (United States)

    1990-11-01

    exchange among more than two chromosomes or fragments which is the result of several breaks. D Dicentric : An exchange between two chromosomes which results...in a chromosome with two centromeres. This is often associated with an acentric fragment in which case it is classified as DF. DF Dicentric with... Chromosome Aberration Assay, the BALB/c- 3T3 Cell Transformation Assay an in In Vivo/lIn Vitro Unscheduled DNA Synthesis Assay and S-Phase Synthesis Assay

  6. In situ hybridization analysis of isodicentric X-chromosomes with short arm fusion

    DEFF Research Database (Denmark)

    Koch, J E; Kølvraa, S; Hertz, Jens Michael;

    1990-01-01

    We present here an alternative approach to the study of mosaic cell lines containing dicentric chromosomes. The approach is based on chromosome-specific non-radioactive in situ hybridization with centromere (alpha satellite DNA) probes. The hybridization analysis may be used as an alternative...... it for the analysis of two cases of isodicentric X-chromosomes. The approach is expected to be generally applicable, so that it may be applied to the scoring of other types of chromosomal mosaicism as well....

  7. Development of Anti-Cancer Therapeutics That Modulate the RAD51-BRCA2 Complex

    Science.gov (United States)

    2005-03-01

    and other (including dicentric and ring chromosomes ). Figure 4. Fluorescence In situ hybridization with probes for centro- centromeric probe. An...composite image; the telomeric probe is green. Chromosomes arrow). Acentric and dicentric chromosomes or minichromosomes were counterstained with DAPI...minichromosomes are actually chromosome frag- MICE AND CELLS DELETED FOR BRCA2 EXON 27 32S ments. In addition to acentrics, dicentric minichromo- matid is

  8. Induced dicentric chromosome formation promotes genomic rearrangements and tumorigenesis

    OpenAIRE

    Gascoigne, Karen E; Cheeseman, Iain M.

    2013-01-01

    Chromosomal rearrangements can radically alter gene products and their function, driving tumor formation or progression. However, the molecular origins and evolution of such rearrangements are varied and poorly understood, with cancer cells often containing multiple, complex rearrangements. One mechanism that can lead to genomic rearrangements is the formation of a “dicentric” chromosome containing two functional centromeres. Indeed, such dicentric chromosomes have been observed in cancer cel...

  9. Test for Chemical Induction of Chromosome Aberration in Cultured Chinese Hamster Ovary (CHO) Cells With and Without Metabolic Activation. Test Article: N,N,N’,N’-tetramethyl Ethanediamine (TMEDA)

    Science.gov (United States)

    2008-06-13

    union. d Dicentric - an asymmetrical exchange between two chromosomes resulting in a chromosome with two centromeres with or without an accompanying...Test for Chemical Induction of Chromosome Aberrations in Cultured Chinese Hamster Ovary (CHO) Cells With and Without Metabolic Activation Test...number. 1. REPORT DATE 26 JUN 2008 2. REPORT TYPE 3. DATES COVERED 4. TITLE AND SUBTITLE Test for Chemical Induction of Chromosome

  10. Molecular Cytogenetic Characterization of a Non-Robertsonian Dicentric Chromosome 14;19 Identified in a Girl with Short Stature and Amenorrhea

    OpenAIRE

    Dutta, Usha R.; Vijaya Kumar Pidugu; Ashwin Dalal

    2012-01-01

    We report a 16-year-old girl who presented with short stature and amenorrhea. Initially the cytogenetic analysis showed the presence of a mosaic non-Robertsonian dicentric chromosome involving chromosomes 14 and 19. Subsequent molecular cytogenetic analysis by fluorescence in situ hybridization (FISH) using whole chromosome paints, centromeric probes, as well as gene specific probes confirmed the dicentric nature of the derivative chromosome and indicated that the rearrangement involved the s...

  11. The Role of the Low Molecular Weight (LMW) Isoforms of Cyclin E in Breast Cancer Tumorigenesis

    Science.gov (United States)

    2007-06-01

    clones include: 1) dicentric chromosomes , chromosomes which contain two copies of the same centromere, 2) ring chromosomes , which results from 2...cells also had more metaphases with chromosome fragments and dicentric chromosomes than the metaphases of EL or empty vector overexpressing cells...fragments or dicentric chromosomes . Overall our results show that the full-length cyclin E does generate genomic instability in 76NE6 cells, however, T1

  12. DRDC Ottawa Participation in the SILENE Accident Dosimetry Intercomparison Exercise. June 10-21, 2002

    Science.gov (United States)

    2002-11-01

    The expected background of chromosomal aberrations in a normal (unexposed population) is generally accepted to be in the order of 1 dicentric per 1000...acentric terminal or interstitial chromosome fragment of varying size. When it is formed independently of a dicentric or centric ring chromosome aberration...variation). dicentric chromosome aberration resulting from annealing of the centromeric pieces of two broken chromosomes (accompanied by an acentric

  13. The genetics of folate metabolism and maternal risk of birth of a child with Down syndrome and associated congenital heart defects

    Directory of Open Access Journals (Sweden)

    Fabio eCoppedè

    2015-06-01

    Full Text Available Almost 15 years ago it was hypothesized that polymorphisms of genes encoding enzymes involved in folate metabolism could lead to aberrant methylation of peri-centromeric regions of chromosome 21, favoring its abnormal segregation during maternal meiosis. Subsequently, more than 50 small case-control studies investigated whether or not maternal polymorphisms of folate pathway genes could be risk factors for the birth of a child with Down syndrome (DS, yielding conflicting and inconclusive results. However, recent meta-analyses of those studies suggest that at least three of those polymorphisms, namely MTHFR 677C>T, MTRR 66A>G, and RFC1 80G>A, are likely to act as maternal risk factors for the birth of a child with trisomy 21, revealing also complex gene-nutrient interactions. A large-cohort study also revealed that lack of maternal folic acid supplementation at peri-conception resulted in increased risk for a DS birth due to errors occurred at maternal meiosis II in the aging oocyte, and it was shown that the methylation status of chromosome 21 peri-centromeric regions could favor recombination errors during meiosis leading to its malsegregation. In this regard, two recent case-control studies revealed association of maternal polymorphisms or haplotypes of the DNMT3B gene, coding for an enzyme required for the regulation of DNA methylation at centromeric and peri-centromeric regions of human chromosomes, with risk of having a birth with DS. Furthermore, congenital heart defects (CHD are found in almost a half of DS births, and increasing evidence points to a possible contribution of lack of folic acid supplementation at peri-conception, maternal polymorphisms of folate pathway genes, and resulting epigenetic modifications of several genes, at the basis of their occurrence. This review summarizes available case-control studies and literature meta-analyses in order to provide a critical and up to date overview of what we currently know in this

  14. Brassica juncea Lines with Substituted Chimeric GFP-CENH3 Give Haploid and Aneuploid Progenies on Crossing with Other Lines

    OpenAIRE

    Watts, Anshul; Singh, Sunil K.; Bhadouria, Jyoti; Naresh, Vasupalli; Bishoyi, Ashok K.; K.A. Geetha; Chamola, Rohit; Pattanayak, Debasis; Bhat, Shripad R.

    2017-01-01

    Haploids and doubled haploids are invaluable for basic genetic studies and in crop improvement. A novel method of haploid induction through genetic engineering of the Centromere Histone Protein gene, CENH3, has been demonstrated in Arabidopsis. The present study was undertaken to develop haploid inducer (HI) lines of Brassica juncea based on the principles elaborated in Arabidopsis. B. juncea was found to carry three copies of CENH3 which generated five different transcripts, of which three t...

  15. Effect of Chromosome Tethering on Nuclear Organization in Yeast

    OpenAIRE

    Barış Avşaroğlu; Gabriel Bronk; Susannah Gordon-Messer; Jungoh Ham; Debra A Bressan; Haber, James E; Jane Kondev

    2014-01-01

    Interphase chromosomes in Saccharomyces cerevisiae are tethered to the nuclear envelope at their telomeres and to the spindle pole body (SPB) at their centromeres. Using a polymer model of yeast chromosomes that includes these interactions, we show theoretically that telomere attachment to the nuclear envelope is a major determinant of gene positioning within the nucleus only for genes within 10 kb of the telomeres. We test this prediction by measuring the distance between the SPB and the sil...

  16. Recruiting a microtubule-binding complex to DNA directs chromosome segregation in budding yeast

    OpenAIRE

    Murray, Andrew W.; Lacefield, Soni; Lau, Tsz Cham Derek

    2009-01-01

    Accurate chromosome segregation depends on the kinetochore, the complex of proteins that link microtubules to centromeric DNA1. The budding yeast kinetochore consists of more than 80 proteins assembled on a 125bp region of DNA1. We studied the assembly and function of kinetochore components by fusing individual kinetochore proteins to the lactose repressor (LacI) and testing their ability to improve the segregation of a plasmid carrying tandem repeats of the lactose operator (LacO). Targeting...

  17. Genome-Wide Synthetic Lethal Screens Identify an Interaction Between the Nuclear Envelope Protein, Apq12p, and the Kinetochore in Saccharomyces cerevisiae

    OpenAIRE

    Montpetit, Ben; Thorne, Ken; Barrett, Irene; Andrews, Kim; Jadusingh, Ravi; Hieter, Phil; Measday, Vivien

    2005-01-01

    The maintenance of genome stability is a fundamental requirement for normal cell cycle progression. The budding yeast Saccharomyces cerevisiae is an excellent model to study chromosome maintenance due to its well-defined centromere and kinetochore, the region of the chromosome and associated protein complex, respectively, that link chromosomes to microtubules. To identify genes that are linked to chromosome stability, we performed genome-wide synthetic lethal screens using a series of novel t...

  18. Cold Spring Harbor symposia on quantitative biology. Volume XLVII, Part 1. Structures of DNA

    Energy Technology Data Exchange (ETDEWEB)

    1983-01-01

    The proceedings for the 47th Annual Cold Spring Harbor Symposia on Quantitative Biology are presented. This symposium focused on the Structure of DNA. Topics presented covered research in the handedness of DNA, conformational analysis, chemically modified DNA, chemical synthesis of DNA, DNA-protein interactions, DNA within nucleosomes, DNA methylation, DNA replication, gyrases and topoisomerases, recombining and mutating DNA, transcription of DNA and its regulation, the organization of genes along DNA, repetitive DNA and pseudogenes, and origins of replication, centromeres, and teleomeres.

  19. A comparison of the chromosome G-banding pattern in two Sorex species, S. satunini and S. araneus (Mammalia, Insectivora

    Directory of Open Access Journals (Sweden)

    Yuri Borisov

    2012-08-01

    Full Text Available The G-banded karyotype of S. satunini was compared with the karyotype of Sorex araneus. Extensive homology was revealed. The major chromosomal rearrangements involved in the evolutionary divergence of these species have been identified as centric fusions and centromeric shifts. From the known palaeontological age of S. satunini it is obvious that the vast chromosomal polymorphism of the S. araneus group originated during the middle Pleistocene.

  20. Chromosome complement, C-banding, Ag-NOR and replication banding in the zebrafish Danio rerio.

    Science.gov (United States)

    Daga, R R; Thode, G; Amores, A

    1996-01-01

    The chromosome complement of Danio rerio was investigated by Giemsa staining and C-banding, Ag-NORs and replication banding. The diploid number of this species is 2n = 50 and the arm number (NF) = 100. Constitutive heterochromatin was located at the centromeric position of all chromosome pairs. Nucleolus organizer regions appeared in the terminal position of the long arms of chromosomes 1, 2 and 8. Replication banding pattern allowed the identification of each chromosome pair.

  1. Aurora B prevents chromosome arm separation defects by promoting telomere dispersion and disjunction

    OpenAIRE

    Reyes, Céline; Serrurier, Céline; Gauthier, Tiphaine; Gachet, Yannick; Tournier, Sylvie

    2015-01-01

    The segregation of centromeres and telomeres at mitosis is coordinated at multiple levels to prevent the formation of aneuploid cells, a phenotype frequently observed in cancer. Mitotic instability arises from chromosome segregation defects, giving rise to chromatin bridges at anaphase. Most of these defects are corrected before anaphase onset by a mechanism involving Aurora B kinase, a key regulator of mitosis in a wide range of organisms. Here, we describe a new role for Aurora B in telomer...

  2. Epigenetics and Triplet-Repeat Neurological Diseases

    OpenAIRE

    Nageshwaran, Sathiji; Festenstein, Richard

    2015-01-01

    The term “junk DNA” has been reconsidered following the delineation of the functional significance of repetitive DNA regions. Typically associated with centromeres and telomeres, DNA repeats are found in nearly all organisms throughout their genomes. Repetitive regions are frequently heterochromatinized resulting in silencing of intrinsic and nearby genes. However, this is not a uniform rule, with several genes known to require such an environment to permit transcription. Repetitive regions f...

  3. Epigenetics and triplet repeat neurological diseases

    OpenAIRE

    Sathiji eNageshwaran; Richard eFestenstein

    2015-01-01

    The term ‘junk DNA’ has been reconsidered following the delineation of the functional significance of repetitive DNA regions. Typically associated with centromeres and telomeres, DNA repeats are found in nearly all organisms throughout their genomes. Repetitive regions are frequently heterchromatinised resulting in silencing of intrinsic and nearby genes. However, this is not a uniform rule, with several genes known to require such an environment to permit transcription. Repetitive regions fr...

  4. Dual-resolution modeling demonstrates greater conformational heterogeneity of CENP-A/H4 dimer than that of H3/H4

    Science.gov (United States)

    Zhao, Haiqing

    Centromere protein A (CENP-A) is a centromere-specific H3 histone variant and shares only about 50% amino acid sequence identity with the canonical H3 protein. CENP-A is required for packaging the centromere and for the proper separation of chromosomes during mitosis. Despite their discrete functions, previously reported crystal structures of the CENP-A/H4 and H3/H4 dimers reveal surprising similarity. In this work, we characterize the structure and dynamics of CENP-A/H4 and H3/H4 dimers with a dual-resolution approach, using both all-atom and coarse-grained (CG) molecular dynamics (MD) simulations. Interestingly, the histone dimer containing CENP-A is more structurally variable than the canonical H3 dimer. Furthermore, our calculations revealed significant conformational distinctions between the interface profiles of CENP-A/H4 and H3/H4. In addition, the presence of the CENP-A-specific chaperone HJURP dramatically reduced the conformational heterogeneity of CENP-A/H4. Overall, these results are in general agreement with the available experimental data and provide new dynamic insights into the mechanisms underpinning the chaperone-mediated assembly of CENP-A nucleosomes in vivo.

  5. Studies on the mitotic chromosome scaffold of Allium sativum

    Institute of Scientific and Technical Information of China (English)

    ZHAOJIAN; SHAOBOJIN; 等

    1995-01-01

    An argentophilic structure is present in the metaphase chromosomes of garlic(Allium sativum),Cytochemical studies indicate that the main component of the structure is non-histone proteins(NHPs).The results of light and electron microscopic observations reveal that the chromosme NHP scaffold is a network which is composed of fibres and granules and distributed throughout the chromosomes.In the NHP network,there are many condensed regions that are connected by redlatively looser regions.The distribution of the condensed regions varies in individual chromosomes.In some of the chromosomes the condensed regions are lognitudinally situsted in the central part of a chromatid while in others these regions appear as coillike transverse bands.At early metaphase.scaffolds of the sister chromatids of a chromosome are linked to each other in the centromeric region,meanwhile,they are connected by scafold materials along the whole length of the chromosome.At late metaphase,however,the connective scaffold materials between the two sister chromatids disappear gradually and the chromatids begin to separate from one another at their ends.but the chromatids are linked together in the centromeric region until anaphase.This connection seems to be related to the special structure of the NHP scaffold formed in the centromeric region.The morphological features and dynamic changes of the chromosome scaffold are discussed.

  6. Timeless links replication termination to mitotic kinase activation.

    Science.gov (United States)

    Dheekollu, Jayaraju; Wiedmer, Andreas; Hayden, James; Speicher, David; Gotter, Anthony L; Yen, Tim; Lieberman, Paul M

    2011-05-06

    The mechanisms that coordinate the termination of DNA replication with progression through mitosis are not completely understood. The human Timeless protein (Tim) associates with S phase replication checkpoint proteins Claspin and Tipin, and plays an important role in maintaining replication fork stability at physical barriers, like centromeres, telomeres and ribosomal DNA repeats, as well as at termination sites. We show here that human Tim can be isolated in a complex with mitotic entry kinases CDK1, Auroras A and B, and Polo-like kinase (Plk1). Plk1 bound Tim directly and colocalized with Tim at a subset of mitotic structures in M phase. Tim depletion caused multiple mitotic defects, including the loss of sister-chromatid cohesion, loss of mitotic spindle architecture, and a failure to exit mitosis. Tim depletion caused a delay in mitotic kinase activity in vivo and in vitro, as well as a reduction in global histone H3 S10 phosphorylation during G2/M phase. Tim was also required for the recruitment of Plk1 to centromeric DNA and formation of catenated DNA structures at human centromere alpha satellite repeats. Taken together, these findings suggest that Tim coordinates mitotic kinase activation with termination of DNA replication.

  7. Timeless links replication termination to mitotic kinase activation.

    Directory of Open Access Journals (Sweden)

    Jayaraju Dheekollu

    Full Text Available The mechanisms that coordinate the termination of DNA replication with progression through mitosis are not completely understood. The human Timeless protein (Tim associates with S phase replication checkpoint proteins Claspin and Tipin, and plays an important role in maintaining replication fork stability at physical barriers, like centromeres, telomeres and ribosomal DNA repeats, as well as at termination sites. We show here that human Tim can be isolated in a complex with mitotic entry kinases CDK1, Auroras A and B, and Polo-like kinase (Plk1. Plk1 bound Tim directly and colocalized with Tim at a subset of mitotic structures in M phase. Tim depletion caused multiple mitotic defects, including the loss of sister-chromatid cohesion, loss of mitotic spindle architecture, and a failure to exit mitosis. Tim depletion caused a delay in mitotic kinase activity in vivo and in vitro, as well as a reduction in global histone H3 S10 phosphorylation during G2/M phase. Tim was also required for the recruitment of Plk1 to centromeric DNA and formation of catenated DNA structures at human centromere alpha satellite repeats. Taken together, these findings suggest that Tim coordinates mitotic kinase activation with termination of DNA replication.

  8. Localization of latency-associated nuclear antigen (LANA) on mitotic chromosomes

    Energy Technology Data Exchange (ETDEWEB)

    Rahayu, Retno; Ohsaki, Eriko [Division of Virology, Department of Microbiology and Immunology, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871 (Japan); Omori, Hiroko [Central Instrumentation Laboratory Research Institute for Microbial Diseases (BIKEN), Osaka University, Osaka 565-0871 (Japan); Ueda, Keiji, E-mail: kueda@virus.med.osaka-u.ac.jp [Division of Virology, Department of Microbiology and Immunology, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871 (Japan)

    2016-09-15

    In latent infection of Kaposi's sarcoma-associated herpesvirus (KSHV), viral gene expression is extremely limited and copy numbers of viral genomes remain constant. Latency-associated nuclear antigen (LANA) is known to have a role in maintaining viral genome copy numbers in growing cells. Several studies have shown that LANA is localized in particular regions on mitotic chromosomes, such as centromeres/pericentromeres. We independently examined the distinct localization of LANA on mitotic chromosomes during mitosis, using super-resolution laser confocal microscopy and correlative fluorescence microscopy–electron microscopy (FM-EM) analyses. We found that the majority of LANA were not localized at particular regions such as telomeres/peritelomeres, centromeres/pericentromeres, and cohesion sites, but at the bodies of condensed chromosomes. Thus, LANA may undergo various interactions with the host factors on the condensed chromosomes in order to tether the viral genome to mitotic chromosomes and realize faithful viral genome segregation during cell division. - Highlights: • This is the first report showing LANA dots on mitotic chromosomes by fluorescent microscopy followed by electron microscopy. • LANA dots localized randomly on condensed chromosomes other than centromere/pericentromere and telomere/peritelomre. • Cellular mitotic checkpoint should not be always involved in the segregation of KSHV genomes in the latency.

  9. A spindle-like apparatus guides bacterial chromosome segregation.

    Science.gov (United States)

    Ptacin, Jerod L; Lee, Steven F; Garner, Ethan C; Toro, Esteban; Eckart, Michael; Comolli, Luis R; Moerner, W E; Shapiro, Lucy

    2010-08-01

    Until recently, a dedicated mitotic apparatus that segregates newly replicated chromosomes into daughter cells was believed to be unique to eukaryotic cells. Here we demonstrate that the bacterium Caulobacter crescentus segregates its chromosome using a partitioning (Par) apparatus that has surprising similarities to eukaryotic spindles. We show that the C. crescentus ATPase ParA forms linear polymers in vitro and assembles into a narrow linear structure in vivo. The centromere-binding protein ParB binds to and destabilizes ParA structures in vitro. We propose that this ParB-stimulated ParA depolymerization activity moves the centromere to the opposite cell pole through a burnt bridge Brownian ratchet mechanism. Finally, we identify the pole-specific TipN protein as a new component of the Par system that is required to maintain the directionality of DNA transfer towards the new cell pole. Our results elucidate a bacterial chromosome segregation mechanism that features basic operating principles similar to eukaryotic mitotic machines, including a multivalent protein complex at the centromere that stimulates the dynamic disassembly of polymers to move chromosomes into daughter compartments.

  10. Use of Repetitive Sequences for Molecular and Cytogenetic Characterization of Avena Species from Portugal

    Science.gov (United States)

    Tomás, Diana; Rodrigues, Joana; Varela, Ana; Veloso, Maria Manuela; Viegas, Wanda; Silva, Manuela

    2016-01-01

    Genomic diversity of Portuguese accessions of Avena species—diploid A. strigosa and hexaploids A. sativa and A. sterilis—was evaluated through molecular and cytological analysis of 45S rDNA, and other repetitive sequences previously studied in cereal species—rye subtelomeric sequence (pSc200) and cereal centromeric sequence (CCS1). Additionally, retrotransposons and microsatellites targeting methodologies—IRAP (inter-retrotransposon amplified polymorphism) and REMAP (retrotransposon-microsatellite amplified polymorphism)—were performed. A very high homology was detected for ribosomal internal transcribed sequences (ITS1 and ITS2) between the species analyzed, although nucleolar organizing regions (NOR) fluorescent in situ hybridization (FISH) analysis revealed distinct number of Nor loci between diploid and hexaploid species. Moreover, morphological diversity, evidenced by FISH signals with different sizes, was observed between distinct accessions within each species. pSc200 sequences were for the first time isolated from Avena species but proven to be highly similar in all genotypes analyzed. The use of primers designed for CCS1 unraveled a sequence homologous to the Ty3/gypsy retrotransposon Cereba, that was mapped to centromeric regions of diploid and hexaploid species, being however restricted to the more related A and D haplomes. Retrotransposon-based methodologies disclosed species- and accessions-specific bands essential for the accurate discrimination of all genotypes studied. Centromeric, IRAP and REMAP profiles therefore allowed accurate assessment of inter and intraspecific variability, demonstrating the potential of these molecular markers on future oat breeding programs. PMID:26861283

  11. Cloning and comparative mapping of a human chromosome 4-specific alpha satellite DNA sequence.

    Science.gov (United States)

    D'Aiuto, L; Antonacci, R; Marzella, R; Archidiacono, N; Rocchi, M

    1993-11-01

    We have isolated and characterized two human alphoid DNA clones: p4n1/4 and pZ4.1. Clone p4n1/4 identifies specifically the centromeric region of chromosome 4; pZ4.1 recognizes a subset of alphoid DNA shared by chromosomes 4 and 9. The specificity was determined using fluorescence in situ hybridization experiments on metaphase spreads and Southern blotting analysis of human-hamster somatic cell hybrids. The genomic organization of both subsets was also investigated. Comparative mapping on chimpanzee and gorilla chromosomes was performed. p4n1/4 hybridizes to chimpanzee chromosomes 11 and 13, homologs of human chromosomes 9 and 2q, respectively. On gorilla metaphase spreads, p4n1/4 hybridizes exclusively to the centromeric region of chromosome 19, partially homologous to human chromosome 17. No hybridization signal was detected on chromosome 3 of both chimpanzee and gorilla, in both species homolog of human chromosome 4. Identical comparative mapping results were obtained using pZ4.1 probe, although the latter recognizes an alphoid subset distinct from the one recognized by p4n1/4. The implications of these results in the evolution of centromeric regions of primate chromosomes are discussed.

  12. Cloning and comparative mapping of a human chromosome 4-specific alpha satellite DNA sequence

    Energy Technology Data Exchange (ETDEWEB)

    D' Aiuto, L.; Marzella, R.; Archidiacono, N.; Rocchi, M. (Universita di Bari (Italy)); Antonacci, R. (Instituto Anatomia Umana Normale, Modena (Italy))

    1993-11-01

    The authors have isolated and characterized two human alphoid DNA clones: p4n1/4 and pZ4.1. Clone p4n1/4 identifies specifically the centromeric region of chromosome 4; pZ4.1 recognizes a subset of alphoid DNA shared by chromosomes 4 and 9. The specificity was determined using fluorescence in situ hybridization experiments on metaphase spreads and Southern blotting analysis of human-hamster somatic cell hybrids. The genomic organization of both subsets was also investigated. Comparative mapping on chimpanzee and gorilla chromosomes was performed. p4n1/4 hybridizes to chimpanzee chromosomes 11 and 13, homologs of human chromosomes 9 and 2q, respectively. On gorilla metaphase spreads, p4n1/4 hybridizes exclusively to the centromeric region of chromosome 19, partially homologous to human chromosome 17. No hybridization signal was detected on chromosome 3 of both chimpanzee and gorilla, in both species homolog of human chromosome 4. Identical comparative mapping results were obtained using pZ4.1 probe, although the latter recognizes an alphoid subset distinct from the one recognized by p4n1/4. The implications of these results in the evolution of centromeric regions of primate chromosomes are discussed. 33 refs., 4 figs.

  13. Haspin has Multiple Functions in the Plant Cell Division Regulatory Network.

    Science.gov (United States)

    Kozgunova, Elena; Suzuki, Takamasa; Ito, Masaki; Higashiyama, Tetsuya; Kurihara, Daisuke

    2016-04-01

    Progression of cell division is controlled by various mitotic kinases. In animal cells, phosphorylation of histone H3 at Thr3 by the kinase Haspin (haploid germ cell-specific nuclear protein kinase) promotes centromeric Aurora B localization to regulate chromosome segregation. However, less is known about the function of Haspin in regulatory networks in plant cells. Here, we show that inhibition of Haspin with 5-iodotubercidin (5-ITu) in Bright Yellow-2 (BY-2) cells delayed chromosome alignment. Haspin inhibition also prevented the centromeric localization of Aurora3 kinase (AUR3) and disrupted its function. This suggested that Haspin plays a role in the specific positioning of AUR3 on chromosomes in plant cells, a function conserved in animals. The results also indicated that Haspin and AUR3 are involved in the same pathway, which regulates chromosome alignment during prometaphase/metaphase. Remarkably, Haspin inhibition by 5-ITu also led to a severe cytokinesis defect, resulting in binuclear cells with a partially formed cell plate. The 5-ITu treatment did not affect microtubules, AUR1/2 or the NACK-PQR pathway; however, it did alter the distribution of actin filaments on the cell plate. Together, these results suggested that Haspin has several functions in regulating cell division in plant cells: in the localization of AUR3 on centromeres and in regulating late cell plate expansion during cytokinesis.

  14. Synergistic Control of Kinetochore Protein Levels by Psh1 and Ubr2.

    Directory of Open Access Journals (Sweden)

    Eva Herrero

    2016-02-01

    Full Text Available The accurate segregation of chromosomes during cell division is achieved by attachment of chromosomes to the mitotic spindle via the kinetochore, a large multi-protein complex that assembles on centromeres. The budding yeast kinetochore comprises more than 60 different proteins. Although the structure and function of many of these proteins has been investigated, we have little understanding of the steady state regulation of kinetochores. The primary model of kinetochore homeostasis suggests that kinetochores assemble hierarchically from the centromeric DNA via the inclusion of a centromere-specific histone into chromatin. We tested this model by trying to perturb kinetochore protein levels by overexpressing an outer kinetochore gene, MTW1. This increase in protein failed to change protein recruitment, consistent with the hierarchical assembly model. However, we find that deletion of Psh1, a key ubiquitin ligase that is known to restrict inner kinetochore protein loading, does not increase levels of outer kinetochore proteins, thus breaking the normal kinetochore stoichiometry. This perturbation leads to chromosome segregation defects, which can be partially suppressed by mutation of Ubr2, a second ubiquitin ligase that normally restricts protein levels at the outer kinetochore. Together these data show that Psh1 and Ubr2 synergistically control the amount of proteins at the kinetochore.

  15. The nucleoporin Mlp2 is involved in chromosomal distribution during mitosis in trypanosomatids.

    Science.gov (United States)

    Morelle, Christelle; Sterkers, Yvon; Crobu, Lucien; MBang-Benet, Diane-Ethna; Kuk, Nada; Portalès, Pierre; Bastien, Patrick; Pagès, Michel; Lachaud, Laurence

    2015-04-30

    Nucleoporins are evolutionary conserved proteins mainly involved in the constitution of the nuclear pores and trafficking between the nucleus and cytoplasm, but are also increasingly viewed as main actors in chromatin dynamics and intra-nuclear mitotic events. Here, we determined the cellular localization of the nucleoporin Mlp2 in the 'divergent' eukaryotes Leishmania major and Trypanosoma brucei. In both protozoa, Mlp2 displayed an atypical localization for a nucleoporin, essentially intranuclear, and preferentially in the periphery of the nucleolus during interphase; moreover, it relocated at the mitotic spindle poles during mitosis. In T. brucei, where most centromeres have been identified, TbMlp2 was found adjacent to the centromeric sequences, as well as to a recently described unconventional kinetochore protein, in the periphery of the nucleolus, during interphase and from the end of anaphase onwards. TbMlp2 and the centromeres/kinetochores exhibited a differential migration towards the poles during mitosis. RNAi knockdown of TbMlp2 disrupted the mitotic distribution of chromosomes, leading to a surprisingly well-tolerated aneuploidy. In addition, diploidy was restored in a complementation assay where LmMlp2, the orthologue of TbMlp2 in Leishmania, was expressed in TbMlp2-RNAi-knockdown parasites. Taken together, our results demonstrate that Mlp2 is involved in the distribution of chromosomes during mitosis in trypanosomatids.

  16. An integrated molecular cytogenetic map of Cucumis sativus L. chromosome 2

    Directory of Open Access Journals (Sweden)

    Huang Sanwen

    2011-01-01

    Full Text Available Abstract Background Integration of molecular, genetic and cytological maps is still a challenge for most plant species. Recent progress in molecular and cytogenetic studies created a basis for developing integrated maps in cucumber (Cucumis sativus L.. Results In this study, eleven fosmid clones and three plasmids containing 45S rDNA, the centromeric satellite repeat Type III and the pericentriomeric repeat CsRP1 sequences respectively were hybridized to cucumber metaphase chromosomes to assign their cytological location on chromosome 2. Moreover, an integrated molecular cytogenetic map of cucumber chromosomes 2 was constructed by fluorescence in situ hybridization (FISH mapping of 11 fosmid clones together with the cucumber centromere-specific Type III sequence on meiotic pachytene chromosomes. The cytogenetic map was fully integrated with genetic linkage map since each fosmid clone was anchored by a genetically mapped simple sequence repeat marker (SSR. The relationship between the genetic and physical distances along chromosome was analyzed. Conclusions Recombination was not evenly distributed along the physical length of chromosome 2. Suppression of recombination was found in centromeric and pericentromeric regions. Our results also indicated that the molecular markers composing the linkage map for chromosome 2 provided excellent coverage of the chromosome.

  17. Use of Repetitive Sequences for Molecular and Cytogenetic Characterization of Avena Species from Portugal.

    Science.gov (United States)

    Tomás, Diana; Rodrigues, Joana; Varela, Ana; Veloso, Maria Manuela; Viegas, Wanda; Silva, Manuela

    2016-02-04

    Genomic diversity of Portuguese accessions of Avena species--diploid A. strigosa and hexaploids A. sativa and A. sterilis--was evaluated through molecular and cytological analysis of 45S rDNA, and other repetitive sequences previously studied in cereal species--rye subtelomeric sequence (pSc200) and cereal centromeric sequence (CCS1). Additionally, retrotransposons and microsatellites targeting methodologies--IRAP (inter-retrotransposon amplified polymorphism) and REMAP (retrotransposon-microsatellite amplified polymorphism)--were performed. A very high homology was detected for ribosomal internal transcribed sequences (ITS1 and ITS2) between the species analyzed, although nucleolar organizing regions (NOR) fluorescent in situ hybridization (FISH) analysis revealed distinct number of Nor loci between diploid and hexaploid species. Moreover, morphological diversity, evidenced by FISH signals with different sizes, was observed between distinct accessions within each species. pSc200 sequences were for the first time isolated from Avena species but proven to be highly similar in all genotypes analyzed. The use of primers designed for CCS1 unraveled a sequence homologous to the Ty3/gypsy retrotransposon Cereba, that was mapped to centromeric regions of diploid and hexaploid species, being however restricted to the more related A and D haplomes. Retrotransposon-based methodologies disclosed species- and accessions-specific bands essential for the accurate discrimination of all genotypes studied. Centromeric, IRAP and REMAP profiles therefore allowed accurate assessment of inter and intraspecific variability, demonstrating the potential of these molecular markers on future oat breeding programs.

  18. RNAi pathway participates in chromosome segregation in mammalian cells.

    Science.gov (United States)

    Huang, Chuan; Wang, Xiaolin; Liu, Xu; Cao, Shuhuan; Shan, Ge

    2015-01-01

    The RNAi machinery is a mighty regulator in a myriad of life events. Despite lines of evidence that small RNAs and components of the RNAi pathway may be associated with structure and behavior of mitotic chromosomes in diverse organisms, a direct role of the RNAi pathway in mammalian mitotic chromosome segregation remains elusive. Here we report that Dicer and AGO2, two central components of the mammalian RNAi pathway, participate in the chromosome segregation. Knockdown of Dicer or AGO2 results in a higher incidence of chromosome lagging, and this effect is independent from microRNAs as examined with DGCR8 knockout cells. Further investigation has revealed that α-satellite RNA, a noncoding RNA derived from centromeric repeat region, is managed by AGO2 under the guidance of endogenous small interference RNAs (ASAT siRNAs) generated by Dicer. Furthermore, the slicer activity of AGO2 is essential for the chromosome segregation. Level and distribution of chromosome-associated α-satellite RNA have crucial regulatory effect on the localization of centromeric proteins such as centromere protein C1 (CENPC1). With these results, we also provide a paradigm in which the RNAi pathway participates in vital cellular events through the maintenance of level and distribution of noncoding RNAs in cells.

  19. Engineered minichromosomes in plants.

    Science.gov (United States)

    Birchler, James A

    2015-02-01

    Engineered minichromosomes have been produced in several plant species via telomere-mediated chromosomal truncation. This approach bypasses the complications of the epigenetic nature of centromere function in plants, which has to date precluded the production of minichromosomes by the re-introduction of centromere sequences to a plant cell. Genes to be added to a cleaved chromosome are joined together with telomere repeats on one side. When these constructs are introduced into plant cells, the genes are ligated to the broken chromosomes but the telomere repeats will catalyze the formation of a telomere on the other end cutting the chromosome at that point. Telomere-mediated chromosomal truncation is sufficiently efficient that very small chromosomes can be generated consisting of basically the endogenous centromere and the added transgenes. The added transgenes provide a platform onto which it should be possible to assemble a synthetic chromosome to specification. Combining engineered minichromosomes with doubled haploid breeding should greatly expedite the transfer of transgenes to new lines and to test the interaction of transgenes in new background genotypes. Potential basic and applied applications of synthetic chromosomes are discussed.

  20. Functional characterization of kinetochore protein, Ctf19 in meiosis I: an implication of differential impact of Ctf19 on the assembly of mitotic and meiotic kinetochores in Saccharomyces cerevisiae.

    Science.gov (United States)

    Mehta, Gunjan D; Agarwal, Meenakshi; Ghosh, Santanu K

    2014-03-01

    Meiosis is a specialized cell division process through which chromosome numbers are reduced by half for the generation of gametes. Kinetochore, a multiprotein complex that connects centromeres to microtubules, plays essential role in chromosome segregation. Ctf19 is the key central kinetochore protein that recruits all the other non-essential proteins of the Ctf19 complex in budding yeast. Earlier studies have shown the role of Ctf19 complex in enrichment of cohesin around the centromeres both during mitosis and meiosis, leading to sister chromatid cohesion and meiosis II disjunction. Here we show that Ctf19 is also essential for the proper execution of the meiosis I specific unique events, such as non-homologous centromere coupling, homologue pairing, chiasmata resolution and proper orientation of homologues and sister chromatids with respect to the spindle poles. Additionally, this investigation reveals that proper kinetochore function is required for faithful chromosome condensation in meiosis. Finally, this study suggests that absence of Ctf19 affects the integrity of meiotic kinetochore differently than that of the mitotic kinetochore. Consequently, absence of Ctf19 leads to gross chromosome missegregation during meiosis as compared with mitosis. Hence, this study reports for the first time the differential impact of a non-essential kinetochore protein on the mitotic and meiotic kinetochore ensembles and hence chromosome segregation.

  1. Repetitive DNAs highlight the role of chromosomal fusions in the karyotype evolution of Dascyllus species (Pomacentridae, Perciformes).

    Science.gov (United States)

    Getlekha, Nuntaporn; Molina, Wagner Franco; de Bello Cioffi, Marcelo; Yano, Cassia Fernanda; Maneechot, Nuntiya; Bertollo, Luiz Antonio Carlos; Supiwong, Weerayuth; Tanomtong, Alongklod

    2016-04-01

    The Dascyllus genus consists of 11 species spread over vast regions of the Indo-Pacific, showing remarkable reductions in the diploid chromosome numbers (2n). The present study analyzed the karyotypes and other chromosomal characteristics of D. trimaculatus (2n = 48; 2st + 46a; NF = 50), D. carneus (2n = 48; 2st + 46a; NF = 50) and D. aruanus (2n = 30; 18m + 2st + 10a; NF = 50) from the Thailand Gulf (Pacific Ocean) and D. melanurus (2n = 48; 2st + 46a; NF = 50) from the Andaman Sea (Indian Ocean), employing conventional cytogenetic analyses and the chromosomal mapping of repetitive DNAs, using 18S and 5S rDNA, telomeric sequences and (CA)15, (GA)15, and (CAA)10 microsatellites as probes. The C-positive heterochromatin was found in the centromeric regions of most chromosomal pairs and 18S rDNA phenotypes were single in all species. However, in D. aruanus (2n = 30), which harbors nine metacentric pairs; the 5S rDNA sites were located in the centromeric region of the shortest one. The mapping of the telomeric sequences in D. aruanus revealed the presence of interstitial telomeric sites (ITS) in the centromeric region of four metacentric pairs, with one of these pairs also displaying an additional ITS in the long arms. Distinct chromosomal markers confirmed the reduction of the 2n by chromosomal fusions, highlighting the precise characterization of these rearrangements by the cytogenetic mapping of the repetitive DNAs.

  2. The nuclear position of pericentromeric DNA of chromosome 11 appears to be random in G0 and non-random in G1 human lymphocytes.

    Science.gov (United States)

    Hulspas, R; Houtsmuller, A B; Krijtenburg, P J; Bauman, J G; Nanninga, N

    1994-07-01

    The nuclear topography of pericentromeric DNA of chromosome 11 was analyzed in G0 (nonstimulated) and G1 [phytohemagglutinin (PHA) stimulated] human lymphocytes by confocal microscopy. In addition to the nuclear center, the centrosome was used as a second point of reference in the three-dimensional (3D) analysis. Pericentromeric DNA of chromosome 11 and the centrosome were labeled using a combination of fluorescent in situ hybridization (FISH) and immunofluorescence. To preserve the 3D morphology of the cells, these techniques were performed on whole cells in suspension. Three-dimensional images of the cells were analyzed with a recently developed 3D software program (Interactive Measurement of Axes and Positioning in 3 Dimensions). The distribution of the chromosome 11 centromeres appeared to be random during the G0 stage but clearly non-random during the G1 stage, when the nuclear center was used as a reference point. Further statistical analysis of the G1 cells revealed that the centromeres were randomly distributed in a shell underlying the nuclear membrane. A topographical relationship between the centrosome and the centromeres appeared to be absent during the G0 and G1 stages of the cell cycle.

  3. Human artificial chromosome assembly by transposon-based retrofitting of genomic BACs with synthetic alpha-satellite arrays.

    Science.gov (United States)

    Basu, Joydeep; Willard, Huntington F; Stromberg, Gregory

    2007-01-01

    The development of methodologies for the rapid assembly of synthetic alpha-satellite arrays recapitulating the higher-order periodic organization of native human centromeres permits the systematic investigation of the significance of primary sequence and sequence organization in centromere function. Synthetic arrays with defined mutations affecting sequence and/or organization may be evaluated in a de novo human artificial chromosome assay. This unit describes strategies for the assembly of custom built alpha-satellite arrays containing any desired mutation as well as strategies for the construction and manipulation of alpha satellite-based transposons. Transposons permit the rapid and reliable retrofitting of any genomic bacterial artificial chromosome (BAC) with synthetic alpha-satellite arrays and other functional components, thereby facilitating conversion into BAC-based human artificial chromosome vectors. These techniques permit identification and optimization of the critical parameters underlying the unique ability of alpha-satellite DNA to facilitate de novo centromere assembly, and they will establish the foundation for the next generation of human artificial chromosome vectors.

  4. A simple and rapid fluorescence in situ hybridization microwave protocol for reliable dicentric chromosome analysis.

    Science.gov (United States)

    Cartwright, Ian M; Genet, Matthew D; Kato, Takamitsu A

    2013-03-01

    Fluorescence in situhybridization (FISH) is an extremely effective and sensitive approach to analyzing chromosome aberrations. Until recently, this procedure has taken multiple days to complete. The introduction of telomeric and centromeric peptide nucleic acid (PNA) probes has reduced the procedure's duration to several hours, but the protocols still call for a high temperature (80-90°C) step followed by 1-3 h of hybridization. The newest method to speed up the FISH protocol is the use of a microwave to shorten the heating element to less than a minute; however this protocol still calls for a 1-h hybridization period. We have utilized PNA centromere/telomere probes in conjunction with a microwave oven to show telomere and centromere staining in as little as 30 s. We have optimized the hybridization conditions to increase the sensitivity and effectiveness of the new protocol and can effectively stain chromosomes in 2 min and 30 s of incubation. We have found that our new approach to FISH produces extremely clear and distinct signals. Radiation-induced dicentric formation in mouse and human fibroblast cells was analyzed by two individual scorers and the observed dicentrics matched very well.

  5. Characterization of the atypical karyotype of the black-winged kite Elanus caeruleus (Falconiformes: Accipitridae) by means of classical and molecular cytogenetic techniques.

    Science.gov (United States)

    Bed'Hom, Bertrand; Coullin, Philippe; Guillier-Gencik, Zuzana; Moulin, Sibyle; Bernheim, Alain; Volobouev, Vitaly

    2003-01-01

    The karyotype of the black-winged kite (Elanus caeruleus), a small diurnal raptor living in Africa, Asia and southern Europe, was studied with classical (G-, C-, R-banding, and Ag-NOR staining) and molecular cytogenetic methods, including primed in-situ labelling (PRINS) and fluorescence in-situ hybridization (FISH) with telomeric (TTAGGG) and centromeric DNA repeats. The study revealed that the genome size, measured by flow cytometry (3.1 pg), is in the normal avian range. However, the black-winged kite karyotype is particularly unusual among birds in having a moderate diploid number of 68 chromosomes, and containing only one pair of dot-shaped microchromosomes. Moreover, the macrochromosomes are medium-sized, with the Z and W gonosomes being clearly the largest in the set. C-banding shows that constitutive heterochromatin is located at the centromeric regions of all chromosomes, and that two pairs of small acrocentrics and the pair of microchromosomes are almost entirely heterochromatic and G-band negative. The distribution pattern of a centromeric repeated DNA sequence, as demonstrated by PRINS, follows that of C-heterochromatin. The localization of telomeric sequences by FISH and PRINS reveals many strong telomeric signals but no extratelomeric signal was observed. The atypical organization of the karyotype of the black-winged kite is considered in the context of the modes of karyotypic evolution in birds.

  6. ATRX contributes to epigenetic asymmetry and silencing of major satellite transcripts in the maternal genome of the mouse embryo.

    Science.gov (United States)

    De La Fuente, Rabindranath; Baumann, Claudia; Viveiros, Maria M

    2015-05-15

    A striking proportion of human cleavage-stage embryos exhibit chromosome instability (CIN). Notably, until now, no experimental model has been described to determine the origin and mechanisms of complex chromosomal rearrangements. Here, we examined mouse embryos deficient for the chromatin remodeling protein ATRX to determine the cellular mechanisms activated in response to CIN. We demonstrate that ATRX is required for silencing of major satellite transcripts in the maternal genome, where it confers epigenetic asymmetry to pericentric heterochromatin during the transition to the first mitosis. This stage is also characterized by a striking kinetochore size asymmetry established by differences in CENP-C protein between the parental genomes. Loss of ATRX results in increased centromeric mitotic recombination, a high frequency of sister chromatid exchanges and double strand DNA breaks, indicating the formation of mitotic recombination break points. ATRX-deficient embryos exhibit a twofold increase in transcripts for aurora kinase B, the centromeric cohesin ESCO2, DNMT1, the ubiquitin-ligase (DZIP3) and the histone methyl transferase (EHMT1). Thus, loss of ATRX activates a pathway that integrates epigenetic modifications and DNA repair in response to chromosome breaks. These results reveal the cellular response of the cleavage-stage embryo to CIN and uncover a mechanism by which centromeric fission induces the formation of large-scale chromosomal rearrangements. Our results have important implications to determine the epigenetic origins of CIN that lead to congenital birth defects and early pregnancy loss, as well as the mechanisms involved in the oocyte to embryo transition.

  7. Neocentric X-chromosome in a girl with Turner-like syndrome

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    Hemmat Morteza

    2012-06-01

    Full Text Available Abstract Background Neocentromeres are rare human chromosomal aberrations in which a new centromere has formed in a previously non-centromeric location. We report the finding of a structurally abnormal X chromosome with a neocentromere in a 15-year-old girl with clinical features suggestive of Turner syndrome, including short stature and primary amenorrhea. Result G-banded chromosome analysis revealed a mosaic female karyotype involving two abnormal cell lines. One cell line (84% of analyzed metaphases had a structurally abnormal X chromosome (duplication of the long arm and deletion of the short arm and a normal X chromosome. The other cell line (16% of cells exhibited monosomy X. C-banding studies were negative for the abnormal X chromosome. FISH analysis revealed lack of hybridization of the abnormal X chromosome with both the X centromere-specific probe and the “all human centromeres” probe, a pattern consistent with lack of the X chromosome endogenous centromere. A FISH study using an XIST gene probe revealed the presence of two XIST genes, one on each long arm of the iso(Xq, required for inactivation of the abnormal X chromosome. R-banding also demonstrated inactivation of the abnormal X chromosome. An assay for centromeric protein C (CENP-C was positive on both the normal and the abnormal X chromosomes. The position of CENP-C in the abnormal X chromosome defined a neocentromere, which explains its mitotic stability. The karyotype is thus designated as 46,X,neo(X(qter- > q12::q12- > q21.2- > neo- > q21.2- > qter[42]/45,X[8], which is consistent with stigmata of Turner syndrome. The mother of this patient has a normal karyotype; however, the father was not available for study. Conclusion To our knowledge, this is the first case of mosaic Turner syndrome involving an analphoid iso(Xq chromosome with a proven neocentromere among 90 previously described cases with a proven neocentromere.

  8. Analysis of genetic map of N. crassa by probability method and relative motion method%概率法及相对运动法对链孢霉的连锁遗传作图分析

    Institute of Scientific and Technical Information of China (English)

    庞振凌

    2012-01-01

    针对遗传学教材关于链孢霉的连锁遗传分析中的三个问题:(1)两对等位基因是否连锁;(2)是否位于着丝粒同侧;(3)着丝粒与nic重组率、nic与ade间的重组率、着丝粒与ade间的重组率三者间不存在等式关系.对于(1)、(2)两个问题分别采用概率法给予解答,对于(3)运用相对运动的观点给了三者各自的计算方法及三者间等式关系,即着丝粒与nic重组率+nic与ade间重组率=着丝粒与ade间的重组率.%This article on the "genetics" materials on c N. crassa chain genetic analysis of three issues : ( 1 ) whether two pairs of allele chain or not, (2) are they located in centromere of the same side, (3) reorganization rate among centromere and the nic and ade-rate are not existent equality. The two issues of ( 1 ) and (2), we answered by probability method ,but as for (3) relative motion method of the three respective perspectives to the calculation method and three-equation, reorganization rate namely: centromere and nic + ade and nic = centro- mere and ade.

  9. A cullin E3 ubiquitin ligase complex associates with Rik1 and the Clr4 histone H3-K9 methyltransferase and is required for RNAi-mediated heterochromatin formation.

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    Hong, Eun-Jin Erica; Villén, Judit; Gerace, Erica L; Gygi, Steven P; Moazed, Danesh

    2005-01-01

    The assembly of heterochromatin in fission yeast and metazoans requires histone H3-lysine 9 (-K9) methylation by the conserved Clr4/Suv39h methyltransferase. In fission yeast, H3-K9 methylation requires components of the RNAi machinery and is initiated by the RNA-Induced Transcriptional Silencing (RITS) complex. Here we report the purification of a novel complex that associates with the Clr4 methyltransferase, termed the CLRC (CLr4-Rik1-Cul4) complex. By affinity purification of the Clr4-associated protein Rik1, we show that, in addition to Clr4, Rik1 is associated with the fission yeast E3 ubiquitin ligase Cullin4 (Cul4, encoded by cul4(+)), the ubiquitin-like protein, Ned8, and two previously uncharacterized proteins, designated Cmc1 and Cmc2. In addition, the complex contains substochiometric amounts of histones H2B and H4, and the 14-3-3 protein, Rad24. Deletion of cul4(+), cmc1(+), cmc2(+) and rad24(+) results in a complete loss of silencing of a ura4(+) reporter gene inserted within centromeric DNA repeats or the silent mating type locus. Each of the above deletions also results in accumulation of noncoding RNAs transcribed from centromeric repeats and telomeric DNA regions, and a corresponding loss of small RNAs that are homologous to centromeric repeats, suggesting a defect in the processing of noncoding RNA to small RNA. Based on these results, we propose that the components of the Clr4-Rik1-Cul4 complex act concertedly at an early step in heterochromatin formation.

  10. Distinct Roles of Meiosis-Specific Cohesin Complexes in Mammalian Spermatogenesis

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    Biswas, Uddipta; Hempel, Kai; Llano, Elena; Pendas, Alberto; Jessberger, Rolf

    2016-01-01

    Mammalian meiocytes feature four meiosis-specific cohesin proteins in addition to ubiquitous ones, but the roles of the individual cohesin complexes are incompletely understood. To decipher the functions of the two meiosis-specific kleisins, REC8 or RAD21L, together with the only meiosis-specific SMC protein SMC1β, we generated Smc1β-/-Rec8-/- and Smc1β-/-Rad21L-/- mouse mutants. Analysis of spermatocyte chromosomes revealed that besides SMC1β complexes, SMC1α/RAD21 and to a small extent SMC1α/REC8 contribute to chromosome axis length. Removal of SMC1β and RAD21L almost completely abolishes all chromosome axes. The sex chromosomes do not pair in single or double mutants, and autosomal synapsis is impaired in all mutants. Super resolution microscopy revealed synapsis-associated SYCP1 aberrantly deposited between sister chromatids and on single chromatids in Smc1β-/-Rad21L-/- cells. All mutants show telomere length reduction and structural disruptions, while wild-type telomeres feature a circular TRF2 structure reminiscent of t-loops. There is no loss of centromeric cohesion in both double mutants at leptonema/early zygonema, indicating that, at least in the mutant backgrounds, an SMC1α/RAD21 complex provides centromeric cohesion at this early stage. Thus, in early prophase I the most prominent roles of the meiosis-specific cohesins are in axis-related features such as axis length, synapsis and telomere integrity rather than centromeric cohesion. PMID:27792785

  11. A strategy for constructing aneuploid yeast strains by transient nondisjunction of a target chromosome

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    Peck Anders T

    2009-07-01

    Full Text Available Abstract Background Most methods for constructing aneuploid yeast strains that have gained a specific chromosome rely on spontaneous failures of cell division fidelity. In Saccharomyces cerevisiae, extra chromosomes can be obtained when errors in meiosis or mitosis lead to nondisjunction, or when nuclear breakdown occurs in heterokaryons. We describe a strategy for constructing N+1 disomes that does not require such spontaneous failures. The method combines two well-characterized genetic tools: a conditional centromere that transiently blocks disjunction of one specific chromosome, and a duplication marker assay that identifies disomes among daughter cells. To test the strategy, we targeted chromosomes III, IV, and VI for duplication. Results The centromere of each chromosome was replaced by a centromere that can be blocked by growth in galactose, and ura3::HIS3, a duplication marker. Transient exposure to galactose induced the appearance of colonies carrying duplicated markers for chromosomes III or IV, but not VI. Microarray-based comparative genomic hybridization (CGH confirmed that disomic strains carrying extra chromosome III or IV were generated. Chromosome VI contains several genes that are known to be deleterious when overexpressed, including the beta-tubulin gene TUB2. To test whether a tubulin stoichiometry imbalance is necessary for the apparent lethality caused by an extra chromosome VI, we supplied the parent strain with extra copies of the alpha-tubulin gene TUB1, then induced nondisjunction. Galactose-dependent chromosome VI disomes were produced, as revealed by CGH. Some chromosome VI disomes also carried extra, unselected copies of additional chromosomes. Conclusion This method causes efficient nondisjunction of a targeted chromosome and allows resulting disomic cells to be identified and maintained. We used the method to test the role of tubulin imbalance in the apparent lethality of disomic chromosome VI. Our results indicate

  12. Comparative Study of Domoic Acid and Okadaic Acid Induced - Chromosomal Abnormalities in the CACO-2 Cell Line

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    Edmond E. Creppy

    2006-03-01

    Full Text Available Okadaic Acid (OA the major diarrheic shellfish poisoning (DSP toxin is known as a tumor promoter and seems likely implicated in the genesis of digestive cancer. Little is known regarding genotoxicity and carcinogenicity of Domoic Acid (DA, the major Amnesic Shellfish Poisoning (ASP toxin. Both OA and DA occur in seafood and are of human health concerns. Micronuclei (MN arise from abnormalities in nuclear division during mitosis due to a failure of the mitotic spindle or by complex chromosomal configurations that pose problems during anaphase. In order to evaluate the ability of okadaic acid (OA and domoic acid (DA to induce DNA damage we performed the micronucleus assay using the Caco-2 cell line. To discriminate between a clastogenic or aneugenic effect of OA and DA, the micronucleus assay was conducted by cytokinesis-block micronucleus assay using cytochalasin B with Giemsa staining and/or acridine orange staining, in parallel to fluorescence in situ hybridization (FISH using a concentrated human pan-centromeric chromosome paint probe. Our results showed that OA and DA significantly increased the frequency of MN in Caco-2 cells. The MN caused by OA are found in mononucleated cells and binucleated cells, whereas those caused by DA are mainly in binucleated cells. The results of FISH analysis showed that OA induced centromere-positive micronuclei and DA increased the percentage of MN without a centromeric signal. In conclusion, both OA and DA bear mutagenic potential as revealed in Caco-2 cells by induction of MN formation. Moreover, OA induced whole chromosome loss suggesting a specific aneugenic potential, whereas DA seems simply clastogenic. At present, one cannot rule out possible DNA damage of intestinal cells if concentrations studied are reached in vivo, since this may happen with concentrations of toxins just below regulatory limits in case of frequent consumption of contaminated shell fishes.

  13. Distribution of 5S and 45S rDNA sites in plants with holokinetic chromosomes and the "chromosome field" hypothesis.

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    Sousa, A; Barros e Silva, A E; Cuadrado, A; Loarce, Y; Alves, M V; Guerra, M

    2011-08-01

    Secondary constrictions or 45S rDNA sites are commonly reported to be located mainly in the terminal regions of the chromosomes. This distribution has been assumed to be related to the existence of a "chromosome field" lying between the centromere and the telomere, an area in which certain cytogenetic events may predominantly occur. If this hypothesis is true this distribution should not be observed in holokinetic chromosomes, as they do not have a localized centromere. In order to evaluate this hypothesis, a comparative study was made of the distributions of 5S and 45S rDNA sites using fluorescence in situ hybridization in representatives of the genera Eleocharis, Diplacrum, Fimbristylis, Kyllinga and Rhynchospora, all of which belong to the family Cyperaceae. The numbers of sites per diploid chromosome complement varied from 2 to ∼10 for 5S rDNA, and from 2 to ∼45 for 45S rDNA. All of the 11 species analyzed had terminally located 45S rDNA sites on the chromosomes whereas the 5S rDNA sites also generally had terminal distributions, except for the Rhynchospora species, where their position was almost always interstitial. These results, together with other previously published data, suggest that the variation in the number and position of the rDNA sites in species with holokinetic chromosomes is non-random and similar to that reported for species with monocentric chromosomes. Therefore, the predominant terminal position of the 45S rDNA sites does not appear to be influenced by the centromere-telomere polarization as suggested by the "chromosome field" hypothesis. Additionally, the hybridization of 5S and 45S rDNA sites provides interesting markers to distinguish several chromosomes on the rather symmetrical karyotypes of Cyperaceae.

  14. Mammalian ChlR1 has a role in heterochromatin organization

    Energy Technology Data Exchange (ETDEWEB)

    Inoue, Akira; Hyle, Judith [Department of Tumor Cell Biology, St. Jude Children' s Research Hospital, Memphis, TN (United States); Lechner, Mark S. [Faculty of Health Sciences, Simon Fraser University, Burnaby, BC (Canada); Lahti, Jill M., E-mail: jill.lahti@stjude.org [Department of Tumor Cell Biology, St. Jude Children' s Research Hospital, Memphis, TN (United States); Department of Molecular Sciences, University of Tennessee Health Science Center, Memphis, TN (United States)

    2011-10-15

    The ChlR1 DNA helicase, encoded by DDX11 gene, which is responsible for Warsaw breakage syndrome (WABS), has a role in sister-chromatid cohesion. In this study, we show that human ChlR1 deficient cells exhibit abnormal heterochromatin organization. While constitutive heterochromatin is discretely localized at perinuclear and perinucleolar regions in control HeLa cells, ChlR1-depleted cells showed dispersed localization of constitutive heterochromatin accompanied by disrupted centromere clustering. Cells isolated from Ddx11{sup -/-} embryos also exhibited diffuse localization of centromeres and heterochromatin foci. Similar abnormalities were found in HeLa cells depleted of combinations of HP1{alpha} and HP1{beta}. Immunofluorescence and chromatin immunoprecipitation showed a decreased level of HP1{alpha} at pericentric regions in ChlR1-depleted cells. Trimethyl-histone H3 at lysine 9 (H3K9-me3) was also modestly decreased at pericentric sequences. The abnormality in pericentric heterochromatin was further supported by decreased DNA methylation within major satellite repeats of Ddx11{sup -/-} embryos. Furthermore, micrococcal nuclease (MNase) assay revealed a decreased chromatin density at the telomeres. These data suggest that in addition to a role in sister-chromatid cohesion, ChlR1 is also involved in the proper formation of heterochromatin, which in turn contributes to global nuclear organization and pleiotropic effects. -- Highlights: {yields} New role for ChlR1 (DDX11), a cohesinopathy gene, in heterochromatin organization. {yields} Loss of ChlR1 altered heterochromatin localization and centromere clustering. {yields} Reduced ChlR1 levels also reduced HP1{alpha} and H3K9-me3 binding to pericentric DNA. {yields} Decreased DNA methylation was found in pericentric repeats of Ddx11{sup -/-} embryos. {yields} These findings will aid in understanding the pathogenesis of Warsaw breakage syndrome.

  15. Stable complex formation of CENP-B with the CENP-A nucleosome.

    Science.gov (United States)

    Fujita, Risa; Otake, Koichiro; Arimura, Yasuhiro; Horikoshi, Naoki; Miya, Yuta; Shiga, Tatsuya; Osakabe, Akihisa; Tachiwana, Hiroaki; Ohzeki, Jun-ichirou; Larionov, Vladimir; Masumoto, Hiroshi; Kurumizaka, Hitoshi

    2015-05-26

    CENP-A and CENP-B are major components of centromeric chromatin. CENP-A is the histone H3 variant, which forms the centromere-specific nucleosome. CENP-B specifically binds to the CENP-B box DNA sequence on the centromere-specific repetitive DNA. In the present study, we found that the CENP-A nucleosome more stably retains human CENP-B than the H3.1 nucleosome in vitro. Specifically, CENP-B forms a stable complex with the CENP-A nucleosome, when the CENP-B box sequence is located at the proximal edge of the nucleosome. Surprisingly, the CENP-B binding was weaker when the CENP-B box sequence was located in the distal linker region of the nucleosome. This difference in CENP-B binding, depending on the CENP-B box location, was not observed with the H3.1 nucleosome. Consistently, we found that the DNA-binding domain of CENP-B specifically interacted with the CENP-A-H4 complex, but not with the H3.1-H4 complex, in vitro. These results suggested that CENP-B forms a more stable complex with the CENP-A nucleosome through specific interactions with CENP-A, if the CENP-B box is located proximal to the CENP-A nucleosome. Our in vivo assay also revealed that CENP-B binding in the vicinity of the CENP-A nucleosome substantially stabilizes the CENP-A nucleosome on alphoid DNA in human cells.

  16. H3 Thr3 phosphorylation is crucial for meiotic resumption and anaphase onset in oocyte meiosis.

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    Wang, Qian; Wei, Haojie; Du, Juan; Cao, Yan; Zhang, Nana; Liu, Xiaoyun; Liu, Xiaoyu; Chen, Dandan; Ma, Wei

    2016-01-01

    Haspin-catalyzed histone H3 threonine 3 (Thr3) phosphorylation facilitates chromosomal passenger complex (CPC) docking at centromeres, regulating indirectly chromosome behavior during somatic mitosis. It is not fully known about the expression and function of H3 with phosphorylated Thr3 (H3T3-P) during meiosis in oocytes. In this study, we investigated the expression and sub-cellular distribution of H3T3-P, as well as its function in mouse oocytes during meiotic division. Western blot analysis revealed that H3T3-P expression was only detected after germinal vesicle breakdown (GVBD), and gradually increased to peak level at metaphase I (MI), but sharply decreased at metaphase II (MII). Immunofluorescence showed H3T3-P was only brightly labeled on chromosomes after GVBD, with relatively high concentration across the whole chromosome axis from pro-metaphase I (pro-MI) to MI. Specially, H3T3-P distribution was exclusively limited to the local space between sister centromeres at MII stage. Haspin inhibitor, 5-iodotubercidin (5-ITu), dose- and time-dependently blocked H3T3-P expression in mouse oocytes. H3T3-P inhibition delayed the resumption of meiosis (GVBD) and chromatin condensation. Moreover, the loss of H3T3-P speeded up the meiotic transition to MII of pro-MI oocytes in spite of the presence of non-aligned chromosomes, even reversed MI-arrest induced with Nocodazole. The inhibition of H3T3-P expression distinguishably damaged MAD1 recruitment on centromeres, which indicates the spindle assembly checkpoint was impaired in function, logically explaining the premature onset of anaphase I. Therefore, Haspin-catalyzed histone H3 phosphorylation is essential for chromatin condensation and the following timely transition from meiosis I to meiosis II in mouse oocytes during meiotic division.

  17. Karyotype variability in tropical maize sister inbred lines and hybrids compared with KYS standard line.

    Science.gov (United States)

    Mondin, Mateus; Santos-Serejo, Janay A; Bertäo, Mônica R; Laborda, Prianda; Pizzaia, Daniel; Aguiar-Perecin, Margarida L R

    2014-01-01

    Maize karyotype variability has been extensively investigated. The identification of maize somatic and pachytene chromosomes has improved with the development of fluorescence in situ hybridization (FISH) using tandemly repeated DNA sequences as probes. We identified the somatic chromosomes of sister inbred lines that were derived from a tropical flint maize population (Jac Duro [JD]), and hybrids between them, using FISH probes for the 180-bp knob repeat, centromeric satellite (CentC), centromeric satellite 4 (Cent4), subtelomeric clone 4-12-1, 5S ribosomal DNA and nucleolus organizing region DNA sequences. The observations were integrated with data based on C-banded mitotic metaphases and conventional analysis of pachytene chromosomes. Heterochromatic knobs visible at pachynema were coincident with C-bands and 180-bp FISH signals on somatic chromosomes, and most of them were large. Variation in the presence of some knobs was observed among lines. Small 180-bp knob signals were invariant on the short arms of chromosomes 1, 6, and 9. The subtelomeric 4-12-1 signal was also invariant and useful for identifying some chromosomes. The centromere location of chromosomes 2 and 4 differed from previous reports on standard maize lines. Somatic chromosomes of a JD line and the commonly used KYS line were compared by FISH in a hybrid of these lines. The pairing behavior of chromosomes 2 and 4 at pachytene stage in this hybrid was investigated using FISH with chromosome-specific probes. The homologues were fully synapsed, including the 5S rDNA and CentC sites on chromosome 2, and Cent4 and subtelomeric 4-12-1 sites on chromosome 4. This suggests that homologous chromosomes could pair through differential degrees of chromatin packaging in homologous arms differing in size. The results contribute to current knowledge of maize global diversity and also raise questions concerning the meiotic pairing of homologous chromosomes possibly differing in their amounts of repetitive DNA.

  18. Dicentric chromosomes and 20q11.2 amplification in MDS/AML with apparent monosomy 20.

    Science.gov (United States)

    Mackinnon, R N; Campbell, L J

    2007-01-01

    FISH analysis of 41 previously karyotyped cases of MDS and AML with apparent monosomy of chromosome 20 revealed a variety of dicentric abnormalities involving chromosome 20. These usually, but not always, involved a breakpoint in the long arm of chromosome 20 and loss of the common deleted region at 20q12. Not one case of true monosomy 20 was confirmed. We found evidence for dicentric chromosome formation in 21 of 24 unbalanced translocations containing chromosome 20 and that were studied in more detail. Subsequent loss of one of the centromeres had occurred in eight of these 24 cases, and was more frequent than centromere inactivation as a means of resolving the inherent instability of a dicentric chromosome. In the three cases with dicentric chromosomes from which proximal 20q had been excised along with the 20 centromere, the excised segment was retained, and in two of these it was amplified. Proximal 20q was clearly retained in all but three cases, and present in three or more copies in 17 of 41 cases. The retention and amplification of proximal 20q provides support for the hypothesis that there is an oncogene located in this region of 20q that is activated in cases of MDS/AML with del(20q). Apparent monosomy 20 in MDS/AML should be treated as evidence of unidentified chromosome 20 abnormalities, and familiarity with the typical G-banded morphology of these derivatives can help with their identification. The reported incidence of dicentric chromosomes is clearly an under-estimate but is increasing in myeloid disorders as more cases are studied with methods allowing their detection.

  19. Regulation of Budding Yeast CENP-A levels Prevents Misincorporation at Promoter Nucleosomes and Transcriptional Defects.

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    Erica M Hildebrand

    2016-03-01

    Full Text Available The exclusive localization of the histone H3 variant CENP-A to centromeres is essential for accurate chromosome segregation. Ubiquitin-mediated proteolysis helps to ensure that CENP-A does not mislocalize to euchromatin, which can lead to genomic instability. Consistent with this, overexpression of the budding yeast CENP-A(Cse4 is lethal in cells lacking Psh1, the E3 ubiquitin ligase that targets CENP-A(Cse4 for degradation. To identify additional mechanisms that prevent CENP-A(Cse4 misincorporation and lethality, we analyzed the genome-wide mislocalization pattern of overexpressed CENP-A(Cse4 in the presence and absence of Psh1 by chromatin immunoprecipitation followed by high throughput sequencing. We found that ectopic CENP-A(Cse4 is enriched at promoters that contain histone H2A.Z(Htz1 nucleosomes, but that H2A.Z(Htz1 is not required for CENP-A(Cse4 mislocalization. Instead, the INO80 complex, which removes H2A.Z(Htz1 from nucleosomes, promotes the ectopic deposition of CENP-A(Cse4. Transcriptional profiling revealed gene expression changes in the psh1Δ cells overexpressing CENP-A(Cse4. The down-regulated genes are enriched for CENP-A(Cse4 mislocalization to promoters, while the up-regulated genes correlate with those that are also transcriptionally up-regulated in an htz1Δ strain. Together, these data show that regulating centromeric nucleosome localization is not only critical for maintaining centromere function, but also for ensuring accurate promoter function and transcriptional regulation.

  20. Regulation of Budding Yeast CENP-A levels Prevents Misincorporation at Promoter Nucleosomes and Transcriptional Defects.

    Science.gov (United States)

    Hildebrand, Erica M; Biggins, Sue

    2016-03-01

    The exclusive localization of the histone H3 variant CENP-A to centromeres is essential for accurate chromosome segregation. Ubiquitin-mediated proteolysis helps to ensure that CENP-A does not mislocalize to euchromatin, which can lead to genomic instability. Consistent with this, overexpression of the budding yeast CENP-A(Cse4) is lethal in cells lacking Psh1, the E3 ubiquitin ligase that targets CENP-A(Cse4) for degradation. To identify additional mechanisms that prevent CENP-A(Cse4) misincorporation and lethality, we analyzed the genome-wide mislocalization pattern of overexpressed CENP-A(Cse4) in the presence and absence of Psh1 by chromatin immunoprecipitation followed by high throughput sequencing. We found that ectopic CENP-A(Cse4) is enriched at promoters that contain histone H2A.Z(Htz1) nucleosomes, but that H2A.Z(Htz1) is not required for CENP-A(Cse4) mislocalization. Instead, the INO80 complex, which removes H2A.Z(Htz1) from nucleosomes, promotes the ectopic deposition of CENP-A(Cse4). Transcriptional profiling revealed gene expression changes in the psh1Δ cells overexpressing CENP-A(Cse4). The down-regulated genes are enriched for CENP-A(Cse4) mislocalization to promoters, while the up-regulated genes correlate with those that are also transcriptionally up-regulated in an htz1Δ strain. Together, these data show that regulating centromeric nucleosome localization is not only critical for maintaining centromere function, but also for ensuring accurate promoter function and transcriptional regulation.

  1. LINE retrotransposon RNA is an essential structural and functional epigenetic component of a core neocentromeric chromatin.

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    Anderly C Chueh

    2009-01-01

    Full Text Available We have previously identified and characterized the phenomenon of ectopic human centromeres, known as neocentromeres. Human neocentromeres form epigenetically at euchromatic chromosomal sites and are structurally and functionally similar to normal human centromeres. Recent studies have indicated that neocentromere formation provides a major mechanism for centromere repositioning, karyotype evolution, and speciation. Using a marker chromosome mardel(10 containing a neocentromere formed at the normal chromosomal 10q25 region, we have previously mapped a 330-kb CENP-A-binding domain and described an increased prevalence of L1 retrotransposons in the underlying DNA sequences of the CENP-A-binding clusters. Here, we investigated the potential role of the L1 retrotransposons in the regulation of neocentromere activity. Determination of the transcriptional activity of a panel of full-length L1s (FL-L1s across a 6-Mb region spanning the 10q25 neocentromere chromatin identified one of the FL-L1 retrotransposons, designated FL-L1b and residing centrally within the CENP-A-binding clusters, to be transcriptionally active. We demonstrated the direct incorporation of the FL-L1b RNA transcripts into the CENP-A-associated chromatin. RNAi-mediated knockdown of the FL-L1b RNA transcripts led to a reduction in CENP-A binding and an impaired mitotic function of the 10q25 neocentromere. These results indicate that LINE retrotransposon RNA is a previously undescribed essential structural and functional component of the neocentromeric chromatin and that retrotransposable elements may serve as a critical epigenetic determinant in the chromatin remodelling events leading to neocentromere formation.

  2. Using microsatellites to understand the physical distribution of recombination on soybean chromosomes.

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    Alina Ott

    Full Text Available Soybean is a major crop that is an important source of oil and proteins. A number of genetic linkage maps have been developed in soybean. Specifically, hundreds of simple sequence repeat (SSR markers have been developed and mapped. Recent sequencing of the soybean genome resulted in the generation of vast amounts of genetic information. The objectives of this investigation were to use SSR markers in developing a connection between genetic and physical maps and to determine the physical distribution of recombination on soybean chromosomes. A total of 2,188 SSRs were used for sequence-based physical localization on soybean chromosomes. Linkage information was used from different maps to create an integrated genetic map. Comparison of the integrated genetic linkage maps and sequence based physical maps revealed that the distal 25% of each chromosome was the most marker-dense, containing an average of 47.4% of the SSR markers and 50.2% of the genes. The proximal 25% of each chromosome contained only 7.4% of the markers and 6.7% of the genes. At the whole genome level, the marker density and gene density showed a high correlation (R(2 of 0.64 and 0.83, respectively with the physical distance from the centromere. Recombination followed a similar pattern with comparisons indicating that recombination is high in telomeric regions, though the correlation between crossover frequency and distance from the centromeres is low (R(2 = 0.21. Most of the centromeric regions were low in recombination. The crossover frequency for the entire soybean genome was 7.2%, with extremes much higher and lower than average. The number of recombination hotspots varied from 1 to 12 per chromosome. A high correlation of 0.83 between the distribution of SSR markers and genes suggested close association of SSRs with genes. The knowledge of distribution of recombination on chromosomes may be applied in characterizing and targeting genes.

  3. A stable hybrid containing haploid genomes of two obligate diploid Candida species.

    Science.gov (United States)

    Chakraborty, Uttara; Mohamed, Aiyaz; Kakade, Pallavi; Mugasimangalam, Raja C; Sadhale, Parag P; Sanyal, Kaustuv

    2013-08-01

    Candida albicans and Candida dubliniensis are diploid, predominantly asexual human-pathogenic yeasts. In this study, we constructed tetraploid (4n) strains of C. albicans of the same or different lineages by spheroplast fusion. Induction of chromosome loss in the tetraploid C. albicans generated diploid or near-diploid progeny strains but did not produce any haploid progeny. We also constructed stable heterotetraploid somatic hybrid strains (2n + 2n) of C. albicans and C. dubliniensis by spheroplast fusion. Heterodiploid (n + n) progeny hybrids were obtained after inducing chromosome loss in a stable heterotetraploid hybrid. To identify a subset of hybrid heterodiploid progeny strains carrying at least one copy of all chromosomes of both species, unique centromere sequences of various chromosomes of each species were used as markers in PCR analysis. The reduction of chromosome content was confirmed by a comparative genome hybridization (CGH) assay. The hybrid strains were found to be stably propagated. Chromatin immunoprecipitation (ChIP) assays with antibodies against centromere-specific histones (C. albicans Cse4/C. dubliniensis Cse4) revealed that the centromere identity of chromosomes of each species is maintained in the hybrid genomes of the heterotetraploid and heterodiploid strains. Thus, our results suggest that the diploid genome content is not obligatory for the survival of either C. albicans or C. dubliniensis. In keeping with the recent discovery of the existence of haploid C. albicans strains, the heterodiploid strains of our study can be excellent tools for further species-specific genome elimination, yielding true haploid progeny of C. albicans or C. dubliniensis in future.

  4. Location of a High-Lysine Gene and the DDT-Resistance Gene on Barley Chromosome 7

    DEFF Research Database (Denmark)

    Jensen, J.

    1979-01-01

    mutants, nos 1508.18, and 19. Linkage studies with translocations locate the Lys3 locus in the centromere region ofchromosome 7. A linkage study involving the loci lys3 and ddt (resistance to DDT) together with the marker locifi (fragile stem), s(short rachilla hairs), and r (smooth awn) show...... that the order of the five loci on chromosome 7 from the long to the short chromosome arm is Y, s,fi, lys3, ddt. The distance from locus I to locus ddt is about 100 centimorgans....

  5. Unique small RNA signatures uncovered in the tammar wallaby genome

    Directory of Open Access Journals (Sweden)

    Lindsay James

    2012-10-01

    Full Text Available Abstract Background Small RNAs have proven to be essential regulatory molecules encoded within eukaryotic genomes. These short RNAs participate in a diverse array of cellular processes including gene regulation, chromatin dynamics and genome defense. The tammar wallaby, a marsupial mammal, is a powerful comparative model for studying the evolution of regulatory networks. As part of the genome sequencing initiative for the tammar, we have explored the evolution of each of the major classes of mammalian small RNAs in an Australian marsupial for the first time, including the first genome-scale analysis of the newest class of small RNAs, centromere repeat associated short interacting RNAs (crasiRNAs. Results Using next generation sequencing, we have characterized the major classes of small RNAs, micro (mi RNAs, piwi interacting (pi RNAs, and the centromere repeat associated short interacting (crasi RNAs in the tammar. We examined each of these small RNA classes with respect to the newly assembled tammar wallaby genome for gene and repeat features, salient features that define their canonical sequences, and the constitution of both highly conserved and species-specific members. Using a combination of miRNA hairpin predictions and co-mapping with miRBase entries, we identified a highly conserved cluster of miRNA genes on the X chromosome in the tammar and a total of 94 other predicted miRNA producing genes. Mapping all miRNAs to the tammar genome and comparing target genes among tammar, mouse and human, we identified 163 conserved target genes. An additional nine genes were identified in tammar that do not have an orthologous miRNA target in human and likely represent novel miRNA-regulated genes in the tammar. A survey of the tammar gonadal piRNAs shows that these small RNAs are enriched in retroelements and carry members from both marsupial and tammar-specific repeat classes. Lastly, this study includes the first in-depth analyses of the newly

  6. [Comparative molecular cytogenetic characterization of partial wheat-wheatgrass hybrids].

    Science.gov (United States)

    Krupin, P Yu; Divashuk, M G; Belov, V I; Glukhova, L I; Aleksandrov, O S; Karlov, G I

    2011-04-01

    The chromosomal composition of the Zernokormovaya 169, Istra 1, Ostankinskaya, and Otrastayushchaya 38 cultivars of octoploid partial wheat-wheatgrass hybrids was studied using genomic in situ hybridization (GISH). Differentiation of wheatgrass chromosomes by the distribution of the GISH signal along the chromosome was revealed. The wheatgrass chromosomes of the hybrid cultivars studied in the work differed in the type of differentiation, centromeric index, and absolute size. The cytogenetic distinctions of these chromosomes revealed by us can be used in making crosses and in studying the transmission through gametes of additional wheatgrass chromosomes.

  7. Partition of the Linear Plasmid N15: Interactions of N15 Partition Functions with the sop Locus of the F Plasmid

    OpenAIRE

    1999-01-01

    A locus close to one end of the linear N15 prophage closely resembles the sop operon which governs partition of the F plasmid; the promoter region contains similar operator sites, and the two putative gene products have extensive amino acid identity with the SopA and -B proteins of F. Our aim was to ascertain whether the N15 sop homologue functions in partition, to identify the centromere site, and to examine possible interchangeability of function with the F Sop system. When expressed at a m...

  8. Reference: 446 [Arabidopsis Phenome Database[Archive

    Lifescience Database Archive (English)

    Full Text Available rk E et al. 2006 Nov. Plant Physiol. 142(3):1004-13. Arabidopsis (Arabidopsis thaliana) QUARTET (QRT) genes are require...d for pollen separation during normal floral development. In qrt mutants, the four products of microsporogenesis re...main fused and pollen grains are released as tetrads. In Arabid...opsis, tetrad analysis in qrt mutants has been used to map all five centromeres, easily distinguish sporophy...tic from gametophytic mutations, and accurately assess crossover interference. Using a combination of forward and re

  9. Cloning of a New Gene/s in Chromosome 17p3.2-p13.1 that Control Apoptosis

    Science.gov (United States)

    2005-04-01

    Burk, F.G. Barr, B.S. Emanuel, Evidence for a 17p tumor related locus distinct from p53 in pediatric primitive neuroectodermal tumors . Cancer Res., 52...previous work of our laboratory shown LOH in the transformed cell line BP1E [18]. The tumor suppressor gene TP53 is located in l7p13.1 at 1.5cM centromeric...profiling 1 has been suggested as a tumor suppressor gene in breast cancer cells (69). By RT-PCR, no differences were found in TP53 and PFN1 expression

  10. Structures of actin-like ParM filaments show architecture of plasmid-segregating spindles

    OpenAIRE

    Bharat, Tanmay A. M.; Murshudov, Garib N.; Sachse, Carsten; Löwe, Jan

    2015-01-01

    Active segregation of E. coli low-copy number plasmid R1 involves formation of a bipolar spindle made of left-handed double-helical actin-like ParM filaments 1-6 . ParR links the filaments with centromeric parC plasmid DNA, while facilitating the addition of subunits to ParM filaments 3,7-9 . Growing ParMRC spindles push sister plasmids to the cell poles 9,10 . Here, using modern electron cryomicroscopy methods we have investigated the structures and arrangements of ParM filaments in vitro an...

  11. Preferential accumulation of sex and Bs chromosomes in biarmed karyotypes by meiotic drive and rates of chromosomal changes in fishes.

    Science.gov (United States)

    Molina, Wagner F; Martinez, Pablo A; Bertollo, Luiz A C; Bidau, Claudio J

    2014-12-01

    Mechanisms of accumulation based on typical centromeric drive or of chromosomes carrying pericentric inversions are adjusted to the general karyotype differentiation in the principal Actinopterygii orders. Here, we show that meiotic drive in fish is also supported by preferential establishment of sex chromosome systems and B chromosomes in orders with predominantly bi-brachial chromosomes. The mosaic of trends acting at an infra-familiar level in fish could be explained as the interaction of the directional process of meiotic drive as background, modulated on a smaller scale by adaptive factors or specific karyotypic properties of each group, as proposed for the orthoselection model.

  12. An infant with a mosaic 45,X/46,X,psu dic(Y) (pter {r_arrow} q11.2::q11.2 {r_arrow}pter) karyotype and mixed gonadal dysgenesis studied for extent of mosaicism in the gonads

    Energy Technology Data Exchange (ETDEWEB)

    Reddy, K.S.; Sulcova, V. [Corning Nichols Inst., San Juan Capistrano, CA (United States); Ho, C.K.; Conner, E.D.; Khurana, A. [Medical Center of Central Georgia, Macon, GA (United States)

    1996-12-30

    An infant with mixed gonadal dysgenesis was found to have a 45,X/46,X,psu dic(Y) karyotype. A low level (8%) of mosaicism for the dic(Y) cell line was observed in peripheral blood lymphocytes and skin fibroblasts. The dicentric nature of the Y chromosome became apparent in fluorescence in situ hybridization studies. The presence of Y centromeric sequences was demonstrated in the paraffin-embedded testis and streak ovary sections. The ratio of Y-positive cells was higher in the testis than in the streak ovary. 7 refs., 2 figs.

  13. Comparative mapping of a gorilla-derived alpha satellite DNA clone on great ape and human chromosomes.

    Science.gov (United States)

    Baldini, A; Miller, D A; Shridhar, V; Rocchi, M; Miller, O J; Ward, D C

    1991-11-01

    We have isolated an alpha satellite DNA clone, pG3.9, from gorilla DNA. Fluorescence in situ hybridization on banded chromosomes under high stringency conditions revealed that pG3.9 identifies homologous sequences at the centromeric region of ten gorilla chromosomes, and, with few exceptions, also recognizes the homologous chromosomes in human. A pG3.9-like alphoid DNA is present on a larger number of orangutan chromosomes, but, in contrast, is present on only two chromosomes in the chimpanzee. These results show that the chromosomal subsets of related alpha satellite DNA sequences may undergo different patterns of evolution.

  14. pTAR-Encoded Proteins in Plasmid Partitioning

    OpenAIRE

    Kalnin, Kirill; Stegalkina, Svetlana; Yarmolinsky, Michael

    2000-01-01

    Partition cassettes, essential for the segregational stability of low-copy-number bacterial plasmids, typically encode two autoregulated proteins and an adjacent cis-acting centromere analog to which one or perhaps both proteins bind. The diminutive partition region of pTAR of Agrobacterium spp. was reported to be exceptional, encoding only a single protein, ParA (D. R. Gallie and C. I. Kado, J. Mol. Biol. 193:465–478, 1987). However, resequencing of the region revealed two small downstream g...

  15. Holocentric plant meiosis: first sisters, then homologues.

    Science.gov (United States)

    Heckmann, Stefan; Schubert, Veit; Houben, Andreas

    2014-01-01

    Meiosis is a crucial process of sexual reproduction by forming haploid gametes from diploid precursor cells. It involves 2 subsequent divisions (meiosis I and meiosis II) after one initial round of DNA replication. Homologous monocentric chromosomes are separated during the first and sister chromatids during the second meiotic division. The faithful segregation of monocentric chromosomes is realized by mono-orientation of fused sister kinetochores at metaphase I and by bi-orientation of sister kinetochores at metaphase II. Conventionally this depends on a 2-step loss of cohesion, along chromosome arms during meiosis I and at sister centromeres during meiosis II.

  16. Structural analysis of the ParR/parC plasmid partition complex

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Ringgaard, Simon; Mercogliano, Christopher P

    2007-01-01

    Accurate DNA partition at cell division is vital to all living organisms. In bacteria, this process can involve partition loci, which are found on both chromosomes and plasmids. The initial step in Escherichia coli plasmid R1 partition involves the formation of a partition complex between the DNA...... and biochemical experiments support a structural arrangement in which the centromere-like parC DNA is wrapped around a ParR protein scaffold. This structure holds implications for how ParM polymerization drives active DNA transport during plasmid partition....

  17. Dynamics of the mitochondrial network during mitosis.

    Science.gov (United States)

    Kanfer, Gil; Kornmann, Benoît

    2016-04-15

    During mitosis, cells undergo massive deformation and reorganization, impacting on all cellular structures. Mitochondria, in particular, are highly dynamic organelles, which constantly undergo events of fission, fusion and cytoskeleton-based transport. This plasticity ensures the proper distribution of the metabolism, and the proper inheritance of functional organelles. During cell cycle, mitochondria undergo dramatic changes in distribution. In this review, we focus on the dynamic events that target mitochondria during mitosis. We describe how the cell-cycle-dependent microtubule-associated protein centromeric protein F (Cenp-F) is recruited to mitochondria by the mitochondrial Rho GTPase (Miro) to promote mitochondrial transport and re-distribution following cell division.

  18. Complete or partial trisomy 3 in gastro-intestinal MALT lymphomas co-occurs with aberrations at 18q21 and correlates with advanced disease stage: A study on 25 cases

    Institute of Scientific and Technical Information of China (English)

    Jens Krugmann; Alexandar Tzankov; Stephan Dirnhofer; Falko Fend; Dominik Wolf; Reiner Siebert; Pensiri Probst; Martin Erdel

    2005-01-01

    @@ TO THE EDITOR Taji et al.[1] have reported in their study on 13 patients with gastric mucosa-associated lymphoid tissue (MALT) lymphomas an aggressive tumor course in trisomy 3 positive cases. The authors analyzed only stage I patients with classical low-grade marginal zone lymphoma of the MALT type and detected the trisomy 3 using an alphasatellite DNA probe directed to the centromere. Their data support the observation that trisomy 3 is the most frequent cytogenetic aberration in MALT lymphomas[2,3].

  19. Weeding out the genes: the Arabidopsis genome project.

    Science.gov (United States)

    Martienssen, R A

    2000-05-01

    The Arabidopsis genome sequence is scheduled for completion at the end of this year (December 2000). It will be the first higher plant genome to be sequenced, and will allow a detailed comparison with bacterial, yeast and animal genomes. Already, two of the five chromosomes have been sequenced, and we have had our first glimpse of higher eukaryotic centromeres, and the structure of heterochromatin. The implications for understanding plant gene function, genome structure and genome organization are profound. In this review, the lessons learned for future genome projects are reviewed as well as a summary of the initial findings in Arabidopsis.

  20. Novel karyotype in the Ullrich-Turner syndrome - 45,X/46,X,r(X)/46,X,dic(X) - investigated with fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Robson, L.; Jackson, J.; Cowell, C.; Sillence, D.; Smith, A. [Children`s Hospital, Camperdown (Australia)

    1994-04-15

    A 10-year-old girl with Ullrich-Turner syndrome was found to have the novel karyotype 45,X/46,X,r(X)(p11q11)/46,X,dic(X)(p11). Fluorescence in situ hybridization (FISH) with the {alpha} satellite X centromere probe established the origin of the small ring chromosome. Scanning a large number of cells by interphase FISH showed that the dicentric (X) was the least prevalent cell line. The common breakpoint of Xp11 suggests a sequence of errors as the mechanism whereby these 3 distinct cell lines have arisen. 11 refs., 4 figs., 1 tab.

  1. Mitotic and meiotic chromosomes of a southern Brazilian population of Boophilus microplus (Acari, Ixodidae

    Directory of Open Access Journals (Sweden)

    Rosane Nunes Garcia

    Full Text Available Using conventional staining with acetic orcein and C-banding techniques it was investigated constitutive heterochromatin chromosomal polymorphisms and the mitotic and the meiotic behavior of male and female chromosomes of Boophilus microplus (Canestrini, 1887. Some differences were detected in the population of southern Brazil as compared to the data of other authors for populations in other latitudes. The differences being mainly concerned with the distribution of constitutive centromeric heterochromatin and variation in the length of heterochromatic blocks in the pericentromeric regions of some chromosome pairs.

  2. The role of HP1β on genomic stability and cellular senescence

    OpenAIRE

    2010-01-01

    Heterochromatin protein 1 (HP1) proteins are fundamental units of heterochromatin packaging that are enriched at the centromeres and telomeres of nearly all eukaryotic chromosomes. HP1 homologues are found in a variety of organisms and are involved in the establishment and maintenance of higher-order chromatin structures by specifically recognizing and binding to (tri- and di-) methylated lysine 9 on histone H3. In mammals, there are three HP1 homologues termed HP1α (Cbx5), HP1β (Cbx1), and H...

  3. A New Approach to Dissect Nuclear Organization: TALE-Mediated Genome Visualization (TGV).

    Science.gov (United States)

    Miyanari, Yusuke

    2016-01-01

    Spatiotemporal organization of chromatin within the nucleus has so far remained elusive. Live visualization of nuclear remodeling could be a promising approach to understand its functional relevance in genome functions and mechanisms regulating genome architecture. Recent technological advances in live imaging of chromosomes begun to explore the biological roles of the movement of the chromatin within the nucleus. Here I describe a new technique, called TALE-mediated genome visualization (TGV), which allows us to visualize endogenous repetitive sequence including centromeric, pericentromeric, and telomeric repeats in living cells.

  4. Molecular cytogenetic mapping of Cucumis sativus and C. melo using highly repetitive DNA sequences.

    Science.gov (United States)

    Koo, Dal-Hoe; Nam, Young-Woo; Choi, Doil; Bang, Jae-Wook; de Jong, Hans; Hur, Yoonkang

    2010-04-01

    Chromosomes often serve as one of the most important molecular aspects of studying the evolution of species. Indeed, most of the crucial mutations that led to differentiation of species during the evolution have occurred at the chromosomal level. Furthermore, the analysis of pachytene chromosomes appears to be an invaluable tool for the study of evolution due to its effectiveness in chromosome identification and precise physical gene mapping. By applying fluorescence in situ hybridization of 45S rDNA and CsCent1 probes to cucumber pachytene chromosomes, here, we demonstrate that cucumber chromosomes 1 and 2 may have evolved from fusions of ancestral karyotype with chromosome number n = 12. This conclusion is further supported by the centromeric sequence similarity between cucumber and melon, which suggests that these sequences evolved from a common ancestor. It may be after or during speciation that these sequences were specifically amplified, after which they diverged and specific sequence variants were homogenized. Additionally, a structural change on the centromeric region of cucumber chromosome 4 was revealed by fiber-FISH using the mitochondrial-related repetitive sequences, BAC-E38 and CsCent1. These showed the former sequences being integrated into the latter in multiple regions. The data presented here are useful resources for comparative genomics and cytogenetics of Cucumis and, in particular, the ongoing genome sequencing project of cucumber.

  5. Clinical, cytogenetic and dual-color FISH studies on five cases of myelodysplastic syndrome or acute myeloid leukemia patients with 1;7 translocation

    Institute of Scientific and Technical Information of China (English)

    申咏梅; 薛永权; 李建勇; 潘金兰; 吴亚芳

    2003-01-01

    Objective To study the clinical and cytogenetic characteristics of four patients with myel odysplastic syndrome (MDS) and one with acute myeloid leukemia experiencing t(1; 7).Methods Five patients seen in our hospital from 1992 to 2001 were diagnosed as MDS and acute myelocytic leukemia (AML) according to the French-American-British (FAB) criteria. Chromosomes were prepared using the direct method as well as 24-hou r unstimulated cultures of fresh heparinized bone marrow for each subject, while R-banding was used to analyze karyotypes. Dual-color fluorescence in situ hy bridization (FISH) using SpectrumRed and SpectrumGreen directly labeled chromoso me 1-specific α-satellite DNA probe (red) and chromosome 7- specific α-sat ellite DNA probe (green) was performed for three cases. Results Of the five patients, three had 1;7 translocation due to along history of expos ure to benzene. In three cases, dual-color FISH resulted in three red signals and two green ones, in which one red signal adjoining one green signal in 27.6% , 84% and 18.5% metaphases, respectively. Conclusions Exposure to benzene may be the cause for Chinese MDS and AML patients with t(1;7 ) translocation. The result of dual-color FISH convincingly confirmed that the centromere of the derivative chromosome 7p/1q resulting from 1;7 translocation was made up of centromeres from both chromosomes 1 and 7.

  6. KIF20A regulates porcine oocyte maturation and early embryo development.

    Directory of Open Access Journals (Sweden)

    Yu Zhang

    Full Text Available KIF20A (Kinesin-like family member 20A, also called mitotic kinesin-like proteins 2 (MKLP2, is a mammalian mitotic kinesin-like motor protein of the Kinesin superfamily proteins (KIFs, which was originally involved in Golgi apparatus dynamics and thought to essential for cell cycle regulation during successful cytokinesis. In the present study, we investigated whether KIF20A has roles on porcine oocyte meiotic maturation and subsequent early embryo development. By immunofluorescence staining, KIF20A was found to exhibit a dynamic localization pattern during meiosis. KIF20A was restricted to centromeres after germinal vesicle breakdown (GVBD, transferred to the midbody at telophase I (TI, and again associated with centromeres at metaphase II (MII. Inhibition of endogenous KIF20A via a specific inhibitor, Paprotrain, resulted in failure of polar body extrusion. Further cell cycle analysis showed that the percentage of oocytes that arrested at early metaphase I (MI stage increased after KIF20A activity inhibition; however, the proportion of oocytes at anaphase/telophase I (ATI and MII stages decreased significantly. Our results also showed that KIF20A inhibition did not affect spindle morphology. In addition, KIF20A was localized at the nucleus of early embryos, and KIF20A inhibition resulted in failure of early parthenogenetic embryo development. These results demonstrated that KIF20A is critical for porcine oocyte meiotic maturation and subsequent early embryo development.

  7. Chromosomal loci associated with endosperm hardness in a malting barley cross.

    Science.gov (United States)

    Walker, Cassandra K; Panozzo, J F; Ford, R; Eckermann, P; Moody, D; Lehmensiek, A; Appels, R

    2011-01-01

    A breeding objective for the malting barley industry is to produce lines with softer, plumper grain containing moderate protein content (9-12%) as they are more likely to imbibe water readily and contain more starch per grain, which in turn produces higher levels of malt extract. In a malting barley mapping population, 'Arapiles' × 'Franklin', the most significant and robust quantitative trait locus (QTL) for endosperm hardness was observed on the short arm of chromosome 1H, across three environments over two growing seasons. This accounted for 22.6% (Horsham 2000), 26.8% (Esperance 2001), and 12.0% (Tarranyurk 2001) of the genetic variance and significantly increased endosperm hardness by 2.06-3.03 SKCS hardness units. Interestingly, Arapiles and Franklin do not vary in Ha locus alleles. Therefore, this region, near the centromere on chromosome 1H, may be of great importance when aiming to manipulate endosperm hardness and malting quality. Interestingly, this region, close to the centromere on chromosome 1H, in our study, aligns with the region of the genome that includes the HvCslF9 and the HvGlb1 genes. Potentially, one or both of these genes could be considered to be candidate genes that influence endosperm hardness in the barley grain. Additional QTLs for endosperm hardness were detected on chromosomes 2H, 3H, 6H and 7H, confirming that the hardness trait in barley is complex and multigenic, similar to many malting quality traits of interest.

  8. Telomeric associated sequences of Drosophila recruit polycomb-group proteins in vivo and can induce pairing-sensitive repression.

    Science.gov (United States)

    Boivin, Antoine; Gally, Christelle; Netter, Sophie; Anxolabéhère, Dominique; Ronsseray, Stéphane

    2003-05-01

    In Drosophila, relocation of a euchromatic gene near centromeric or telomeric heterochromatin often leads to its mosaic silencing. Nevertheless, modifiers of centromeric silencing do not affect telomeric silencing, suggesting that each location requires specific factors. Previous studies suggest that a subset of Polycomb-group (PcG) proteins could be responsible for telomeric silencing. Here, we present the effect on telomeric silencing of 50 mutant alleles of the PcG genes and of their counteracting trithorax-group genes. Several combinations of two mutated PcG genes impair telomeric silencing synergistically, revealing that some of these genes are required for telomeric silencing. In situ hybridization and immunostaining experiments on polytene chromosomes revealed a strict correlation between the presence of PcG proteins and that of heterochromatic telomeric associated sequences (TASs), suggesting that TASs and PcG complexes could be associated at telomeres. Furthermore, lines harboring a transgene containing an X-linked TAS subunit and the mini-white reporter gene can exhibit pairing-sensitive repression of the white gene in an orientation-dependent manner. Finally, an additional binding site for PcG proteins was detected at the insertion site of this type of transgene. Taken together, these results demonstrate that PcG proteins bind TASs in vivo and may be major players in Drosophila telomeric position effect (TPE).

  9. ICIS and Aurora B coregulate the microtubule depolymerase Kif2a.

    Science.gov (United States)

    Knowlton, Anne L; Vorozhko, Valeriya V; Lan, Weijie; Gorbsky, Gary J; Stukenberg, P Todd

    2009-05-12

    Kinesins in the mitotic spindle play major roles in determining spindle shape, size, and bipolarity, although specific regulation of these kinesins at distinct locations on the spindle is poorly understood. So that the forces that are required for spindle bipolarity are balanced, microtubule-depolymerizing kinesins are tightly regulated. Aurora B kinase phosphorylates the neck regions of the kinesin-13 family microtubule depolymerases Kif2a and mitotic centromere-associated kinesin (MCAK) and inhibits their depolymerase activities. How they are reactivated and how this is controlled independently on different kinetochore fibers is unknown. We show that inner centromere Kin-I stimulator (ICIS), which stimulates the related depolymerase MCAK, can reactivate Kif2a after Aurora B inhibition. When antibodies that block the ability of ICIS to activate Kif2a are injected into cells, monopolar spindles are generated. This phenotype is rescued by coinjection of anti-Nuf2 antibodies. We have performed a structure-function analysis of the ICIS protein and find that the N terminus of ICIS binds Aurora B and its regulators INCENP and TD60, whereas a central region binds MCAK, Kif2a, and microtubules, suggesting a scaffold function for ICIS. These data argue that ICIS and the chromosomal passenger complex (CPC) regulate Kif2a depolymerase activity.

  10. The architecture of chicken chromosome territories changes during differentiation

    Directory of Open Access Journals (Sweden)

    Stadler Sonja

    2004-11-01

    Full Text Available Abstract Background Between cell divisions the chromatin fiber of each chromosome is restricted to a subvolume of the interphase cell nucleus called chromosome territory. The internal organization of these chromosome territories is still largely unknown. Results We compared the large-scale chromatin structure of chromosome territories between several hematopoietic chicken cell types at various differentiation stages. Chromosome territories were labeled by fluorescence in situ hybridization in structurally preserved nuclei, recorded by confocal microscopy and evaluated visually and by quantitative image analysis. Chromosome territories in multipotent myeloid precursor cells appeared homogeneously stained and compact. The inactive lysozyme gene as well as the centromere of the lysozyme gene harboring chromosome located to the interior of the chromosome territory. In further differentiated cell types such as myeloblasts, macrophages and erythroblasts chromosome territories appeared increasingly diffuse, disaggregating to separable substructures. The lysozyme gene, which is gradually activated during the differentiation to activated macrophages, as well as the centromere were relocated increasingly to more external positions. Conclusions Our results reveal a cell type specific constitution of chromosome territories. The data suggest that a repositioning of chromosomal loci during differentiation may be a consequence of general changes in chromosome territory morphology, not necessarily related to transcriptional changes.

  11. A Three-Dimensional Model of the Yeast Genome

    Science.gov (United States)

    Noble, William; Duan, Zhi-Jun; Andronescu, Mirela; Schutz, Kevin; McIlwain, Sean; Kim, Yoo Jung; Lee, Choli; Shendure, Jay; Fields, Stanley; Blau, C. Anthony

    Layered on top of information conveyed by DNA sequence and chromatin are higher order structures that encompass portions of chromosomes, entire chromosomes, and even whole genomes. Interphase chromosomes are not positioned randomly within the nucleus, but instead adopt preferred conformations. Disparate DNA elements co-localize into functionally defined aggregates or factories for transcription and DNA replication. In budding yeast, Drosophila and many other eukaryotes, chromosomes adopt a Rabl configuration, with arms extending from centromeres adjacent to the spindle pole body to telomeres that abut the nuclear envelope. Nonetheless, the topologies and spatial relationships of chromosomes remain poorly understood. Here we developed a method to globally capture intra- and inter-chromosomal interactions, and applied it to generate a map at kilobase resolution of the haploid genome of Saccharomyces cerevisiae. The map recapitulates known features of genome organization, thereby validating the method, and identifies new features. Extensive regional and higher order folding of individual chromosomes is observed. Chromosome XII exhibits a striking conformation that implicates the nucleolus as a formidable barrier to interaction between DNA sequences at either end. Inter-chromosomal contacts are anchored by centromeres and include interactions among transfer RNA genes, among origins of early DNA replication and among sites where chromosomal breakpoints occur. Finally, we constructed a three-dimensional model of the yeast genome. Our findings provide a glimpse of the interface between the form and function of a eukaryotic genome.

  12. Characterization of a chromosome-specific chimpanzee alpha satellite subset: Evolutionary relationship to subsets on human chromosomes

    Energy Technology Data Exchange (ETDEWEB)

    Warburton, P.E.; Gosden, J.; Lawson, D. [Western General Hospital, Edinburgh (United Kingdom)] [and others

    1996-04-15

    Alpha satellite DNA is a tandemly repeated DNA family found at the centromeres of all primate chromosomes examined. The fundamental repeat units of alpha satellite DNA are diverged 169- to 172-bp monomers, often found to be organized in chromosome-specific higher-order repeat units. The chromosomes of human (Homo sapiens (HSA)), chimpanzee (Pan troglodytes (PTR) and Pan paniscus), and gorilla (Gorilla gorilla) share a remarkable similarity and synteny. It is of interest to ask if alpha satellite arrays at centromeres of homologous chromosomes between these species are closely related (evolving in an orthologous manner) or if the evolutionary processes that homogenize and spread these arrays within and between chromosomes result in nonorthologous evolution of arrays. By using PCR primers specific for human chromosome 17-specific alpha satellite DNA, we have amplified, cloned, and characterized a chromosome-specific subset from the PTR chimpanzee genome. Hybridization both on Southern blots and in situ as well as sequence analysis show that this subset is most closely related, as expected, to sequences on HSA 17. However, in situ hybridization reveals that this subset is not found on the homologous chromosome in chimpanzee (PTR 19), but instead on PTR 12, which is homologous to HSA 2p. 40 refs., 3 figs.

  13. First description of the karyotype and localization of major and minor ribosomal genes in Rhoadsia altipinna Fowler, 1911 (Characiformes, Characidae from Ecuador

    Directory of Open Access Journals (Sweden)

    Omar Sánchez-Romero

    2015-06-01

    Full Text Available Karyotypic features of Rhoadsia altipinna Fowler, 1911 from Ecuador were investigated by examining metaphase chromosomes through Giemsa staining, C-banding, Ag-NOR, and two-color-fluorescence in situ hybridization (FISH for mapping of 18S and 5S ribosomal genes. The species exhibit a karyotype with 2n = 50, composed of 10 metacentric, 26 submetacentric and 14 subtelocentric elements, with a fundamental number FN=86 and is characterized by the presence of a larger metacentric pair (number 1, which is about 2/3 longer than the average length of the rest of the metacentric series. Sex chromosomes were not observed. Heterochromatin is identifiable on 44 chromosomes, distributed in paracentromeric position near the centromere. The first metacentric pair presents two well-defined heterochromatic blocks in paracentromeric position, near the centromere. Impregnation with silver nitrate showed a single pair of Ag-positive NORs localized at terminal regions of the short arms of the subtelocentric chromosome pair number 12. FISH assay confirmed these localization of NORs and revealed that minor rDNA clusters occur interstitially on the larger metacentric pair number 1. Comparison of results here reported with those available on other Characidae permit to hypothesize that the presence of a very large metacentric pair might represent a unique and derived condition that characterize one of four major lineages molecularly identified in this family.

  14. First description of the karyotype and localization of major and minor ribosomal genes in Rhoadsiaaltipinna Fowler, 1911 (Characiformes, Characidae) from Ecuador.

    Science.gov (United States)

    Sánchez-Romero, Omar; Abad, César Quezada; Cordero, Patricio Quizhpe; de Sene, Viviani França; Nirchio, Mauro; Oliveira, Claudio

    2015-01-01

    Karyotypic features of Rhoadsiaaltipinna Fowler, 1911 from Ecuador were investigated by examining metaphase chromosomes through Giemsa staining, C-banding, Ag-NOR, and two-color-fluorescence in situ hybridization (FISH) for mapping of 18S and 5S ribosomal genes. The species exhibit a karyotype with 2n = 50, composed of 10 metacentric, 26 submetacentric and 14 subtelocentric elements, with a fundamental number FN=86 and is characterized by the presence of a larger metacentric pair (number 1), which is about 2/3 longer than the average length of the rest of the metacentric series. Sex chromosomes were not observed. Heterochromatin is identifiable on 44 chromosomes, distributed in paracentromeric position near the centromere. The first metacentric pair presents two well-defined heterochromatic blocks in paracentromeric position, near the centromere. Impregnation with silver nitrate showed a single pair of Ag-positive NORs localized at terminal regions of the short arms of the subtelocentric chromosome pair number 12. FISH assay confirmed these localization of NORs and revealed that minor rDNA clusters occur interstitially on the larger metacentric pair number 1. Comparison of results here reported with those available on other Characidae permit to hypothesize that the presence of a very large metacentric pair might represent a unique and derived condition that characterize one of four major lineages molecularly identified in this family.

  15. Cytogenetic and molecular analysis of a ring (21) in a patient with partial trisomy 21 and megakaryocytic leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Palmer, C.G.; Bull, M.; Breitfeld, P. [Indiana Univ., Indianapolis, IN (United States)] [and others

    1994-09-01

    We describe a patient with an asymmetrical double ring 21 in mosaic form, 45,XX, -21/46,XX, -21, +r(21) (q22.11{yields}p11.2::q11.1{yields}q22.3), who has limited manifestations of Down`s syndrome and who developed acute myelofibrosis and megakaryocytic leukemia (AMKL), M7, a hematologic disorder particularly common in Down`s syndrome patients. In situ hybridization studies (gene dosage and DNA polymorphism analysis) show that the ring chromosome carries a duplicated region which extends from D21S258 on the centromeric side and includes marker D21S1245 on the telomeric side, but does not include the region from PFKL and ITGB2 through the telomere. FISH studies indicate two sizes of ring 21 in the patient. The origin of the supernumerary chromosome 21 in the proband was paternal; furthermore, the r(21) was probably formed post-zygotically. Included in the duplicated segment are the candidate genes for leukemia AML-1, ETS2 and ERG. If disomic homozygosity is important in the development of AMKL in the proband, the gene responsible maps either between the centromere and D21S11 or between D21S1239 and D21S1245.

  16. The Dnmt3b splice variant is specifically expressed in in vitro-manipulated blastocysts and their derivative ES cells.

    Science.gov (United States)

    Horii, Takuro; Suetake, Isao; Yanagisawa, Eikichi; Morita, Sumiyo; Kimura, Mika; Nagao, Yasumitsu; Imai, Hiroshi; Tajima, Shoji; Hatada, Izuho

    2011-10-01

    Manipulation of preimplantation embryos in vitro, such as in vitro fertilization (IVF), in vitro culture (IVC), intracytoplasmic sperm injection (ICSI), somatic cell nuclear transfer (SCNT) and other assisted reproduction technologies (ART), has contributed to the development of infertility treatment and new animal reproduction methods. However, such embryos often exhibit abnormal DNA methylation patterns in imprinted genes and centromeric satellite repeats. These DNA methylation patterns are established and maintained by three DNA methyltransferases: Dnmt1, Dnmt3a and Dnmt3b. Dnmt3b is responsible for the creation of methylation patterns during the early stage of embryogenesis and consists of many alternative splice variants that affect methylation activity; nevertheless, the roles of these variants have not yet been identified. In this study, we found an alternatively spliced variant of Dnmt3b lacking exon 6 (Dnmt3bΔ6) that is specific to mouse IVC embryos. Dnmt3bΔ6 also showed prominent expression in embryonic stem (ES) cells derived from in vitro manipulated embryos. Interestingly, IVC blastocysts were hypomethylated in centromeric satellite repeat regions that could be susceptible to methylation by Dnmt3b. In vitro methylation activity assays showed that Dnmt3bΔ6 had lower activity than normal Dnmt3b. Our findings suggest that Dnmt3bΔ6 could induce a hypomethylation status especially in in vitro manipulated embryos.

  17. Vesicourethral reflux-induced renal failure in a patient with ICF syndrome due to a novel DNMT3B mutation.

    Science.gov (United States)

    Kutluğ, Seyhan; Ogur, Gönül; Yilmaz, Aysegül; Thijssen, Peter E; Abur, Ummet; Yildiran, Alisan

    2016-12-01

    ICF syndrome is a primary immunodeficiency disease characterized by hypo- or agammaglobulinemia, centromeric instability mainly on chromosomes 1, 9, and 16 and facial anomalies. ICF syndrome presents with frequent respiratory tract infections in infancy. A 20-month-old female patient was referred to our clinic due to frequent lower respiratory tract infections. ICF syndrome was considered because of comorbidity of hypogammaglobulinemia, facial anomalies, and neuromotor growth retardation. Metaphase chromosome analysis revealed centromeric instability on chromosomes 1, 9, and 16 and through Sanger a previously unreported homozygous missense mutation (c.1805T>C; [p.V602A]) was identified in the DNMT3B, confirming ICF1. The patient was found to have a breakdown in renal function 1 year later; the urinary system was examined and bilateral vesicoureteral reflux was found, warranting the need for dialysis in time. This report expands the mutation spectrum of ICF1 and is the first to describe bilateral vesicoureteral reflux accompanying ICF syndrome. © 2016 Wiley Periodicals, Inc.

  18. Drosophila CENP-A mutations cause a BubR1-dependent early mitotic delay without normal localization of kinetochore components.

    Directory of Open Access Journals (Sweden)

    Michael D Blower

    2006-07-01

    Full Text Available The centromere/kinetochore complex plays an essential role in cell and organismal viability by ensuring chromosome movements during mitosis and meiosis. The kinetochore also mediates the spindle attachment checkpoint (SAC, which delays anaphase initiation until all chromosomes have achieved bipolar attachment of kinetochores to the mitotic spindle. CENP-A proteins are centromere-specific chromatin components that provide both a structural and a functional foundation for kinetochore formation. Here we show that cells in Drosophila embryos homozygous for null mutations in CENP-A (CID display an early mitotic delay. This mitotic delay is not suppressed by inactivation of the DNA damage checkpoint and is unlikely to be the result of DNA damage. Surprisingly, mutation of the SAC component BUBR1 partially suppresses this mitotic delay. Furthermore, cid mutants retain an intact SAC response to spindle disruption despite the inability of many kinetochore proteins, including SAC components, to target to kinetochores. We propose that SAC components are able to monitor spindle assembly and inhibit cell cycle progression in the absence of sustained kinetochore localization.

  19. Extraordinary molecular evolution in the PRDM9 fertility gene.

    Directory of Open Access Journals (Sweden)

    James H Thomas

    Full Text Available Recent work indicates that allelic incompatibility in the mouse PRDM9 (Meisetz gene can cause hybrid male sterility, contributing to genetic isolation and potentially speciation. The only phenotype of mouse PRDM9 knockouts is a meiosis I block that causes sterility in both sexes. The PRDM9 gene encodes a protein with histone H3(K4 trimethyltransferase activity, a KRAB domain, and a DNA-binding domain consisting of multiple tandem C2H2 zinc finger (ZF domains. We have analyzed human coding polymorphism and interspecies evolutionary changes in the PRDM9 gene. The ZF domains of PRDM9 are evolving very rapidly, with compelling evidence of positive selection in primates. Positively selected amino acids are predominantly those known to make nucleotide specific contacts in C2H2 zinc fingers. These results suggest that PRDM9 is subject to recurrent selection to change DNA-binding specificity. The human PRDM9 protein is highly polymorphic in its ZF domains and nearly all polymorphisms affect the same nucleotide contact residues that are subject to positive selection. ZF domain nucleotide sequences are strongly homogenized within species, indicating that interfinger recombination contributes to their evolution. PRDM9 has previously been assumed to be a transcription factor required to induce meiosis specific genes, a role that is inconsistent with its molecular evolution. We suggest instead that PRDM9 is involved in some aspect of centromere segregation conflict and that rapidly evolving centromeric DNA drives changes in PRDM9 DNA-binding domains.

  20. Karyotype analysis and physical mapping of 45S rDNA in eight species of Sophora,Robinia,and Amorpha

    Institute of Scientific and Technical Information of China (English)

    LIU Bo; CHEN Chengbin; LI Xiulan; QI Liwang; HAN Suying

    2006-01-01

    The karyotype analysis and physical locations of 45S rDNA were carried out by means of fluorescence in situ hybridization in three species,and two forms of Sophora,two species of Robina,and one species of Amorpha.S.japonica L.,S.japonica L.f.oligophylla Franch.,S.japonica L.f.pendula Loud.,and S.xanthantha C.Y.Ma.are all tetraploids with 2n=28.There were four 45S rDNA sites in pericentromeric regions of two Pairs of chromosomes in each of them.S.rubriflora Tsoong.is a triploid with 2n=21,and three sites were located in each satellite of group 5 chromosomes.In R.pseudoacacia L.(2n=2x=22),we examined four intensive signals in telomeric regions of two pairs of satellite chromosomes.In R.hispida L.(2n=2x=30),there were four other signals in centromeric regions besides those like in R.pseudoacacia.Amorpha fruticosa L.has most chromosomes(2n=40)among the eight materials,however,there were only six 45S rDNA loci and they laid in centromeric regions,and satellites of three pairs of chromosomes.45S rDNA is a valuable chromosomal landmark in karyotype analysis.The distribution and genomic organization Of rDNA in the three genera were also discussed.

  1. Isolation and identification of the three rice monotelosomics

    Institute of Scientific and Technical Information of China (English)

    YU Hengxiu; GONG Zhiyun; SU Yan; GU Minghong

    2005-01-01

    In order to obtain rice monotelosomic, the progeny of 24 telotrisomics, derived from an indica rice variety, Zhongxian 3037, were screened. The variants that differed morphologically from the diploids and the original primary trisomics as well as the telotrisomics were collected for cytological identification. The variants with 24 chromosomes were selected according to the prometaphase chromosomes. From these variants, three monotelosomics with one chromosome arm deletion in each were verified by fluorescence in situ hybridization (FISH) using a rice centromeric BAC clone of 17p22 as a marker probe. The three monotelosomics were derived from telotrisomic 1S, 4L and 11L, respectively. Further identification was conducted on the prometaphase or pachytene chromosomes of the three variants, which were probed with the same centromeric BAC clone together with the corresponding chromosome arm specific makers, a0059H02 (on the short arm of chromosome 1), a0034E24 (on the long arm of chromosome 4), and a0071H11 (on the long arm of chromosome 11). The results indicated that the telocentric chromosomes in the three monotelosomics were derived from their respective corresponding telotrisomics. According to the telocentric chromosomes of the variants, they were monotelosomic 1S (one long arm of chromosome 1 was lost), monotelosomic 4L (one short arm of chromosome 4 was lost) and monotelosomic 11L (one short arm of chromosome 11 was lost), respectively.

  2. Cytogenetic diversity of simple sequences repeats in morphotypes of Brassica rapa ssp. chinensis

    Directory of Open Access Journals (Sweden)

    Jinshuang Zheng

    2016-07-01

    Full Text Available A significant fraction of the nuclear DNA of all eukaryotes is occupied by simple sequence repeats (SSRs. Although thesis sequences have sparked great interest as a means of studying genetic variation, linkage mapping and evolution, little attention had been paid to the chromosomal distribution and cytogenetic diversity of these sequences. This paper report the long-range organization of all possible classes of mono-, di- and tri-nucleotide SSRs in Brassica rapa. Fluorescence in situ hybridization (FISH was used to characterize the cytogenetic diversity of SSRs among morphotypes of B. rapa ssp. chinensis. The proportion of different SSR motifs varied among morphtypes of B. rapa, with trinucleotide SSRs more prevalent in the genome of B. rapa ssp. chinensis. The chromosomal characterizations of mono-, di- and tri-nucleotide repeats have been acquired. The data has revealed the non-random and motif-dependent chromosome distribution of SSRs in different morphtypes, and allowed the relative variability characterized by SSRs amount and similar chromosomal distribution in centromeric/peri-centromeric heterochromatin. The differences of SSRs in the abundance and distribution indicated the driving force of SSRs in relationship with the evolution of B. rapa species. The results provided a comprehensive view on the SSR sequence distribution and evolution for comparison among morphtypes B. rapa ssp. chinensis.

  3. A theory explaining the abnormality in 45,X/46,XY mosaicism with non-fluorescent Y chromosome. presentation of three cases.

    Science.gov (United States)

    Kaluzewski, B; Jokinen, A; Hortling, H; de la Chapelle, A

    1978-03-01

    Three patients with male habitus, short stature and testicular differentiation are described. All had mos 45,X/46,XY, the ratio of the two stemlines varying between the patients and between different tissues. The Y chromosome was abnormal, lacking the brilliant QFQ fluorescence and dark CGB staining characteristic of the distal part of the normal Y. Detailed banding studies suggested that the short arm and proximal part of the long arm were normal, while the distal part of the long arm was molecularly or otherwise altered, resulting in abnormal staining properties. Two of the patients were tested for H-Y antigen and found to be positive. These data and those collected from the literature are compatible with a model in which the primary lesion in X/XY mosaicism is a molecular alteration in the reiterated Y-specific DNA sequences (and possibly neighbouring sequences) of a 46,XY zygote resulting in the frequent mitotic loss of the Y and the emergence of a 45,X line. Provided the testis-determining gene(s) near the centromere are normal, testes are formed and the patient is H-Y antigen-positive. The extent of male or female differentiation depends in part on the prevalence, time of occurence, and distribution of the 45,X line and possibly in part on the alteration of other genes involved in sex differentiation and located on Yq further from the centromere.

  4. Characterization by fluorescence and electron microscopy in situ hybridization of a double Y isochromosome

    Energy Technology Data Exchange (ETDEWEB)

    Fetni, R.; Lemieux, N.; Richer, C.L. [Universite de Montreal, Quebec (Canada)] [and others

    1996-06-14

    A patient with mixed gonadal dysgenesis and Y isochromosomes I(Y) is described. Lymphocyte cultures from peripheral blood contained a high proportion of 45,X cells and several other cell lines with two different marker chromosomes (mars). These markers had either a monocentric (mar1) or a dicentric appearance (mar2). Following high-resolution GTG, RBG, QFQ, and CBG bandings, five cell lines were identified; 45,X/46,X, + mar1/46,X, + mar2/47,X, + mar1x2/47,X + mar2x2. The percentages were 66/6/26/1/1%, respectively. Chromosome banding analyses were insufficient for characterization of the markers. In situ hybridization of specific probes for the Y centromere and its short arm showed, both in fluorescence and electron microscopy (ENT), two different Y rearrangements. Mar1 is an isochromosome for the short arm i(Yp) and mar2 is a dicentric which was shown by EM to be a double isochromosome Yp, inv dup i(Yp). The breakpoint producing mar1 is within the centromere and the one producing mar2 is within one of the short arms of the Y isochromosome. The findings of different cell populations in peripheral blood lymphocytes indicate the postzygotic instability of this i(Yp). 24 refs., 3 figs., 1 tab.

  5. Essential roles of Snf21, a Swi2/Snf2 family chromatin remodeler, in fission yeast mitosis.

    Science.gov (United States)

    Yamada, Kentaro; Hirota, Kouji; Mizuno, Ken-Ichi; Shibata, Takehiko; Ohta, Kunihiro

    2008-10-01

    ATP-dependent chromatin remodelers (ADCRs) convert local chromatin structure into both transcriptional active and repressive state. Recent studies have revealed that ADCRs play diverse regulatory roles in chromosomal events such as DNA repair and recombination. Here we have newly identified a fission yeast gene encoding a Swi2/Snf2 family ADCR. The amino acid sequence of this gene, snf21(+), implies that Snf21 is a fission yeast orthologue of the budding yeast Sth1, the catalytic core of the RSC chromatin remodeling complex. The snf21(+) gene product is a nuclear protein essential to cell viability: the null mutant cells stop growing after several rounds of cell divisions. A temperature sensitive allele of snf21(+), snf21-36 exhibits at non-permissive temperature (34 degrees C) a cell cycle arrest at G2-M phase and defects in chromosome segregation, thereby causing cell elongation, lack of cell growth, and death of some cell population. snf21-36 shows thiabendazole (TBZ) sensitivity even at permissive temperature (25 degrees C). The TBZ sensitivity becomes severer as snf21-36 is combined with the deletion of a centromere-localized Mad2 spindle checkpoint protein. The cell cycle arrest phenotype at 34 degrees C cannot be rescued by the mad2(+) deletion, although it is substantially alleviated at 30 degrees C in mad2Delta. These data suggest that Snf21 plays an essential role in mitosis possibly functioning in centromeric chromatin.

  6. A Combined Global and Local Approach to Elucidate Spatial Organization of the Mycobacterial ParB-parS Partition

    Energy Technology Data Exchange (ETDEWEB)

    Chaudhuri, Barnali [University of Buffalo, The State University of New York; Gupta, Sayan [Case Western Reserve University; Urban, Volker S [ORNL; Chance, Mark [Case Western Reserve University; D' Mello, Rhijuta [Case Western Reserve University; Smith, Lauren [University of Buffalo, The State University of New York; Lyons, Kelly [University of Buffalo, The State University of New York; Gee, Jessica [University of Buffalo, The State University of New York

    2010-01-01

    Combining diverse sets of data at global (size, shape) and local (residue) scales is an emerging trend for elucidating the organization and function of the cellular assemblies. We used such a strategy, combining data from X-ray and neutron scattering with H/D-contrast variation and X-ray footprinting with mass spectrometry, to elucidate the spatial organization of the ParB-parS assembly from Mycobacterium tuberculosis. The ParB-parS participates in plasmid and chromosome segregation and condensation in predivisional bacterial cells. ParB polymerizes around the parS centromere(s) to form a higher-order assembly that serves to recruit cyto-skeletal ParA ATPases and SMC proteins for chromosome segregation. A hybrid model of the ParB-parS was built by combining and correlating computational models with experiment-derived information about size, shape, position of the symmetry axis within the shape, internal topology, DNA-protein interface, exposed surface patches, and prior knowledge. This first view of the ParB-parS leads us to propose how ParB spread on the chromosome to form a larger assembly.

  7. A Combined Global and Local Approach to Elucidate Spatial Organization of the Mycobacterial ParB-parS Partition Assembly

    Energy Technology Data Exchange (ETDEWEB)

    B Chaudhuri; S Gupta; V Urban; M Chance; R DMello; L Smith; K Lyons; J Gee

    2011-12-31

    Combining diverse sets of data at global (size, shape) and local (residue) scales is an emerging trend for elucidating the organization and function of the cellular assemblies. We used such a strategy, combining data from X-ray and neutron scattering with H/D-contrast variation and X-ray footprinting with mass spectrometry, to elucidate the spatial organization of the ParB-parS assembly from Mycobacterium tuberculosis. The ParB-parS participates in plasmid and chromosome segregation and condensation in predivisional bacterial cells. ParB polymerizes around the parS centromere(s) to form a higher-order assembly that serves to recruit cyto-skeletal ParA ATPases and SMC proteins for chromosome segregation. A hybrid model of the ParB-parS was built by combining and correlating computational models with experiment-derived information about size, shape, position of the symmetry axis within the shape, internal topology, DNA-protein interface, exposed surface patches, and prior knowledge. This first view of the ParB-parS leads us to propose how ParB spread on the chromosome to form a larger assembly.

  8. An SMC ATPase mutant disrupts chromosome segregation in Caulobacter.

    Science.gov (United States)

    Schwartz, Monica A; Shapiro, Lucy

    2011-12-01

    Accurate replication and segregation of the bacterial genome are essential for cell cycle progression. We have identified a single amino acid substitution in the Caulobacter structural maintenance of chromosomes (SMC) protein that disrupts chromosome segregation and cell division. The E1076Q point mutation in the SMC ATPase domain caused a dominant-negative phenotype in which DNA replication was able to proceed, but duplicated parS centromeres, normally found at opposite cell poles, remained at one pole. The cellular positions of other chromosomal loci were in the wild-type order relative to the parS centromere, but chromosomes remained unsegregated and appeared to be stacked upon one another. Purified SMC-E1076Q was deficient in ATP hydrolysis and exhibited abnormally stable binding to DNA. We propose that SMC spuriously links the duplicated chromosome immediately after passage of the replication fork. In wild-type cells, ATP hydrolysis opens the SMC dimer, freeing one chromosome to segregate to the opposite pole. The loss of ATP hydrolysis causes the SMC-E1076Q dimer to remain bound to both chromosomes, inhibiting segregation.

  9. Automixis in Artemia: solving a century-old controversy.

    Science.gov (United States)

    Nougué, O; Rode, N O; Jabbour-Zahab, R; Ségard, A; Chevin, L-M; Haag, C R; Lenormand, T

    2015-12-01

    Parthenogenesis (reproduction through unfertilized eggs) encompasses a variety of reproduction modes with (automixis) or without (apomixis) meiosis. Different modes of automixis have very different genetic and evolutionary consequences but can be particularly difficult to tease apart. In this study, we propose a new method to discriminate different types of automixis from population-level genetic data. We apply this method to diploid Artemia parthenogenetica, a crustacean whose reproductive mode remains controversial despite a century of intensive cytogenetic observations. We focus on A. parthenogenetica from two western Mediterranean populations. We show that they are diploid and that markers remain heterozygous in cultures maintained up to ~36 generations in the laboratory. Moreover, parallel patterns of population-wide heterozygosity levels between the two natural populations strongly support the conclusion that diploid A. parthenogenetica reproduce by automictic parthenogenesis with central fusion and low, but nonzero recombination. This settles a century-old controversy on Artemia, and, more generally, suggests that many automictic organisms harbour steep within-chromosome gradients of heterozygosity due to a transition from clonal transmission in centromere-proximal regions to a form of inbreeding similar to self-fertilization in centromere-distal regions. Such systems therefore offer a new avenue for contrasting the genomic consequences of asexuality and inbreeding.

  10. Unbiased K-mer Analysis Reveals Changes in Copy Number of Highly Repetitive Sequences During Maize Domestication and Improvement

    Science.gov (United States)

    Liu, Sanzhen; Zheng, Jun; Migeon, Pierre; Ren, Jie; Hu, Ying; He, Cheng; Liu, Hongjun; Fu, Junjie; White, Frank F.; Toomajian, Christopher; Wang, Guoying

    2017-01-01

    The major component of complex genomes is repetitive elements, which remain recalcitrant to characterization. Using maize as a model system, we analyzed whole genome shotgun (WGS) sequences for the two maize inbred lines B73 and Mo17 using k-mer analysis to quantify the differences between the two genomes. Significant differences were identified in highly repetitive sequences, including centromere, 45S ribosomal DNA (rDNA), knob, and telomere repeats. Genotype specific 45S rDNA sequences were discovered. The B73 and Mo17 polymorphic k-mers were used to examine allele-specific expression of 45S rDNA in the hybrids. Although Mo17 contains higher copy number than B73, equivalent levels of overall 45S rDNA expression indicates that transcriptional or post-transcriptional regulation mechanisms operate for the 45S rDNA in the hybrids. Using WGS sequences of B73xMo17 doubled haploids, genomic locations showing differential repetitive contents were genetically mapped, which displayed different organization of highly repetitive sequences in the two genomes. In an analysis of WGS sequences of HapMap2 lines, including maize wild progenitor, landraces, and improved lines, decreases and increases in abundance of additional sets of k-mers associated with centromere, 45S rDNA, knob, and retrotransposons were found among groups, revealing global evolutionary trends of genomic repeats during maize domestication and improvement. PMID:28186206

  11. Linkage relationships of 19 enzyme Loci in maize.

    Science.gov (United States)

    Goodman, M M; Stuber, C W; Newton, K; Weissinger, H H

    1980-11-01

    Linkage relationships of 19 enzyme loci have been examined. The chromosomal locations of eight of these loci are formally reported for the first time in this paper. These localizations should assist in the construction of additional useful chromosome marker stocks, especially since several of these enzyme loci lie in regions that were previously poorly mapped. Six loci are on the long arm of chromosome 1. The arrangement is (centromere)-Mdh4-mmm-Pgm1-Adh1-Phi-Gdh1, with about 46% recombination between Mdh4 and Gdh1.-Linkage studies with a2 and pr have resulted in the localization of four enzyme genes to chromosome 5 with arrangement Pgm2-Mdh5-Got3-a2-(centromere)-pr-Got2. Pgm2 lies approximately 35 map units distal to a2 in a previously unmapped region of the short arm of 5, beyond ameiotic.-Approximately 23% recombination was observed between Mdh4 and Pgm1 on chromosome 1, while 17% recombination occurred between Mdh5 and Pgm2 on chromosome 5. Similarly, linkages between Idh1 and Mdh1, about 22 map units apart on chromosome 8, and between Mdh2 and Idh2, less than 5 map units apart on chromosome 6, were observed. Thus, segments of chromosomes 1 and 5 and segments of 6 and 8 may represent duplications on nonhomologous chromosomes.

  12. Cytogenetic analyses using C-banding and DAPI/CMA3 staining of four populations of the maize weevil Sitophiluszeamais Motschulsky, 1855 (Coleoptera, Curculionidae).

    Science.gov (United States)

    da Silva, Alexandra A; Braga, Lucas S; Guedes, Raul Narciso C; Tavares, Mara G

    2015-01-01

    Cytogenetic data avalaible for the maize weevil Sitophiluszeamais Motschulsky, 1855 (Coleoptera: Curculionidae), one of the most destructive pests of stored cereal grains, are controversial. Earlier studies focused on single populations and emphasized chromosome number and sex determination system. In this paper, the karyotypes of four populations of Sitophiluszeamais were characterized by conventional staining, C-banding and sequential staining with the fluorochromes chromomycin-A3/4-6-diamidino-2-phenylindole (CMA3/DAPI). The analyses of metaphases obtained from the cerebral ganglia of last instar larvae and the testes of adults showed that the species had 2n = 22 chromosomes, with 10 autosomal pairs and a sex chromosome pair (XX in females and Xyp in males). Chromosome number, however, ranged from 2n = 22 to 26 due to the presence of 0-4 supernumerary chromosomes in individuals from the populations of Viçosa, Unai and Porto Alegre. With the exception of the Y chromosome, which was dot-like, all other chromosomes of this species were metacentric, including the supernumeraries. The heterochromatin was present in the centromeric regions of all autosomes and in the centromere of the X chromosome. The B chromosomes were partially or totally heterochromatic, and the Y chromosome was euchromatic. The heterochromatic regions were labeled with C-banding and DAPI, which showed that they were rich in AT base pairs.

  13. Cytogenetic analyses using C-banding and DAPI/CMA3 staining of four populations of the maize weevil Sitophilus zeamais Motschulsky, 1855 (Coleoptera, Curculionidae

    Directory of Open Access Journals (Sweden)

    Alexandra A. da Silva

    2015-03-01

    Full Text Available Cytogenetic data avalaible for the maize weevil Sitophilus zeamais Motschulsky, 1855 (Coleoptera: Curculionidae, one of the most destructive pests of stored cereal grains, are controversial. Earlier studies focused on single populations and emphasized chromosome number and sex determination system. In this paper, the karyotypes of four populations of S. zeamais were characterized by conventional staining, C-banding and sequential staining with the fluorochromes chromomycin-A3/4-6-diamidino-2-phenylindole (CMA3/DAPI. The analyses of metaphases obtained from the cerebral ganglia of last instar larvae and the testes of adults showed that the species had 2n = 22 chromosomes, with 10 autosomal pairs and a sex chromosome pair (XX in females and Xyp in males. Chromosome number, however, ranged from 2n = 22 to 26 due to the presence of 0–4 supernumerary chromosomes in individuals from the populations of Viçosa, Unai and Porto Alegre. With the exception of the Y chromosome, which was dot-like, all other chromosomes of this species were metacentric, including the supernumeraries. The heterochromatin was present in the centromeric regions of all autosomes and in the centromere of the X chromosome. The B chromosomes were partially or totally heterochromatic, and the Y chromosome was euchromatic. The heterochromatic regions were labeled with C-banding and DAPI, which showed that they were rich in AT base pairs.

  14. Genome-wide analysis of tandem repeats in Tribolium castaneum genome reveals abundant and highly dynamic tandem repeat families with satellite DNA features in euchromatic chromosomal arms.

    Science.gov (United States)

    Pavlek, Martina; Gelfand, Yevgeniy; Plohl, Miroslav; Meštrović, Nevenka

    2015-12-01

    Although satellite DNAs are well-explored components of heterochromatin and centromeres, little is known about emergence, dispersal and possible impact of comparably structured tandem repeats (TRs) on the genome-wide scale. Our bioinformatics analysis of assembled Tribolium castaneum genome disclosed significant contribution of TRs in euchromatic chromosomal arms and clear predominance of satellite DNA-typical 170 bp monomers in arrays of ≥5 repeats. By applying different experimental approaches, we revealed that the nine most prominent TR families Cast1-Cast9 extracted from the assembly comprise ∼4.3% of the entire genome and reside almost exclusively in euchromatic regions. Among them, seven families that build ∼3.9% of the genome are based on ∼170 and ∼340 bp long monomers. Results of phylogenetic analyses of 2500 monomers originating from these families show high-sequence dynamics, evident by extensive exchanges between arrays on non-homologous chromosomes. In addition, our analysis shows that concerted evolution acts more efficiently on longer than on shorter arrays. Efficient genome-wide distribution of nine TR families implies the role of transposition only in expansion of the most dispersed family, and involvement of other mechanisms is anticipated. Despite similarities in sequence features, FISH experiments indicate high-level compartmentalization of centromeric and euchromatic tandem repeats.

  15. Mitosin/CENP-F is a conserved kinetochore protein subjected to cytoplasmic dynein-mediated poleward transport

    Institute of Scientific and Technical Information of China (English)

    ZHEN YE YANG; JING GUO; NING LI; MIN QIAN; SHENG NIAN WANG; XUE LIANG ZHU

    2003-01-01

    Mitosin/CENP-F is a human nuclear protein transiently associated with the outer kinetochore plate in M phase and is involved in M phase progression. LEK1 and CMF1, which are its murine and chicken orthologs, however, are implicated in muscle differentiation and reportedly not distributed at kinetochores.We therefore conducted several assays to clarify this issue. The typical centromere staining patterns were observed in mitotic cells from both human primary culture and murine, canine, and mink cell lines. A C-terminal portion of LEK1 also conferred centromere localization. Our analysis further suggests conserved kinetochore localization of mammalian mitosin orthologs. Moreover, mitosin was associated preferentially with kinetochores of unaligned chromosomes. It was also constantly transported from kinetochores to spindle poles by cytoplasmic dynein. These properties resemble those of other kinetochore proteins important for the spindle checkpoint, thus implying a role of mitosin in this checkpoint. Therefore, mitosin family may serve as multifunctional proteins involved in both mitosis and differentiation.

  16. The genome of black raspberry (Rubus occidentalis).

    Science.gov (United States)

    VanBuren, Robert; Bryant, Doug; Bushakra, Jill M; Vining, Kelly J; Edger, Patrick P; Rowley, Erik R; Priest, Henry D; Michael, Todd P; Lyons, Eric; Filichkin, Sergei A; Dossett, Michael; Finn, Chad E; Bassil, Nahla V; Mockler, Todd C

    2016-09-01

    Black raspberry (Rubus occidentalis) is an important specialty fruit crop in the US Pacific Northwest that can hybridize with the globally commercialized red raspberry (R. idaeus). Here we report a 243 Mb draft genome of black raspberry that will serve as a useful reference for the Rosaceae and Rubus fruit crops (raspberry, blackberry, and their hybrids). The black raspberry genome is largely collinear to the diploid woodland strawberry (Fragaria vesca) with a conserved karyotype and few notable structural rearrangements. Centromeric satellite repeats are widely dispersed across the black raspberry genome, in contrast to the tight association with the centromere observed in most plants. Among the 28 005 predicted protein-coding genes, we identified 290 very recent small-scale gene duplicates enriched for sugar metabolism, fruit development, and anthocyanin related genes which may be related to key agronomic traits during black raspberry domestication. This contrasts patterns of recent duplications in the wild woodland strawberry F. vesca, which show no patterns of enrichment, suggesting gene duplications contributed to domestication traits. Expression profiles from a fruit ripening series and roots exposed to Verticillium dahliae shed insight into fruit development and disease response, respectively. The resources presented here will expedite the development of improved black and red raspberry, blackberry and other Rubus cultivars.

  17. rDNA mapping, heterochromatin characterization and AT/GC content of Agapanthus africanus (L. Hoffmanns (Agapanthaceae

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    ARYANE C. REIS

    2016-01-01

    Full Text Available ABSTRACT Agapanthus (Agapanthaceae has 10 species described. However, most taxonomists differ respect to this number because the great phenotypic plasticity of the species. The cytogenetic has been an important tool to aid the plant taxon identification, and to date, all taxa of Agapanthus L'Héritier studied cytologically, presented 2n = 30. Although the species possess large chromosomes, the group is karyologically little explored. This work aimed to increase the cytogenetic knowledge of Agapanthus africanus (L. Hoffmanns by utilization of chromosome banding techniques with DAPI / CMA3 and Fluorescent in situ Hybridization (FISH. In addition, flow cytometry was used for determination of DNA content and the percentage of AT / GC nitrogenous bases. Plants studied showed 2n = 30 chromosomes, ranging from 4.34 - 8.55 µm, with the karyotype formulae (KF = 10m + 5sm. Through FISH, one 45S rDNA signal was observed proximally to centromere of the chromosome 7, while for 5S rDNA sites we observed one signal proximally to centromere of chromosome 9. The 2C DNA content estimated for the species was 2C = 24.4 with 59% of AT and 41% of GC. Our data allowed important upgrade for biology and cytotaxonomy of Agapanthus africanus (L. Hoffmanns.

  18. Karyotypic evolution in squamate reptiles: comparative gene mapping revealed highly conserved linkage homology between the butterfly lizard (Leiolepis reevesii rubritaeniata, Agamidae, Lacertilia) and the Japanese four-striped rat snake (Elaphe quadrivirgata, Colubridae, Serpentes).

    Science.gov (United States)

    Srikulnath, Kornsorn; Nishida, Chizuko; Matsubara, Kazumi; Uno, Yoshinobu; Thongpan, Amara; Suputtitada, Saowanee; Apisitwanich, Somsak; Matsuda, Yoichi

    2009-01-01

    The butterfly lizard (Leiolepis reevesii rubritaeniata) has the diploid chromosome number of 2n = 36, comprising two distinctive components, macrochromosomes and microchromosomes. To clarify the conserved linkage homology between lizard and snake chromosomes and to delineate the process of karyotypic evolution in Squamata, we constructed a cytogenetic map of L. reevesii rubritaeniata with 54 functional genes and compared it with that of the Japanese four-striped rat snake (E. quadrivirgata, 2n = 36). Six pairs of the lizard macrochromosomes were homologous to eight pairs of the snake macrochromosomes. The lizard chromosomes 1, 2, 4, and 6 corresponded to the snake chromosomes 1, 2, 3, and Z, respectively. LRE3p and LRE3q showed the homology with EQU5 and EQU4, respectively, and LRE5p and LRE5q corresponded to EQU7 and EQU6, respectively. These results suggest that the genetic linkages have been highly conserved between the two species and that their karyotypic difference might be caused by the telomere-to-telomere fusion events followed by inactivation of one of two centromeres on the derived dicentric chromosomes in the lineage of L. reevesii rubritaeniata or the centric fission events of the bi-armed macrochromosomes and subsequent centromere repositioning in the lineage of E. quadrivirgata. The homology with L. reevesii rubritaeniata microchromosomes were also identified in the distal regions of EQU1p and 1q, indicating the occurrence of telomere-to-telomere fusions of microchromosomes to the p and q arms of EQU1.

  19. The Organization of Repetitive DNA in the Genomes of Amazonian Lizard Species in the Family Teiidae.

    Science.gov (United States)

    Carvalho, Natalia D M; Pinheiro, Vanessa S S; Carmo, Edson J; Goll, Leonardo G; Schneider, Carlos H; Gross, Maria C

    2015-01-01

    Repetitive DNA is the largest fraction of the eukaryote genome and comprises tandem and dispersed sequences. It presents variations in relation to its composition, number of copies, distribution, dynamics, and genome organization, and participates in the evolutionary diversification of different vertebrate species. Repetitive sequences are usually located in the heterochromatin of centromeric and telomeric regions of chromosomes, contributing to chromosomal structures. Therefore, the aim of this study was to physically map repetitive DNA sequences (5S rDNA, telomeric sequences, tropomyosin gene 1, and retroelements Rex1 and SINE) of mitotic chromosomes of Amazonian species of teiids (Ameiva ameiva, Cnemidophorus sp. 1, Kentropyx calcarata, Kentropyx pelviceps, and Tupinambis teguixin) to understand their genome organization and karyotype evolution. The mapping of repetitive sequences revealed a distinct pattern in Cnemidophorus sp. 1, whereas the other species showed all sequences interspersed in the heterochromatic region. Physical mapping of the tropomyosin 1 gene was performed for the first time in lizards and showed that in addition to being functional, this gene has a structural function similar to the mapped repetitive elements as it is located preferentially in centromeric regions and termini of chromosomes.

  20. The mouse salivary androgen-binding protein (ABP) gene cluster on chromosomes 7: characterization and evolutionary relationships.

    Science.gov (United States)

    Laukaitis, Christina M; Dlouhy, Stephen R; Karn, Robert C

    2003-10-01

    Mouse salivary androgen-binding protein (ABP) is a pair of dimers, composed of an alpha subunit disulfide bridged to either a beta or a gamma subunit. It has been proposed that each subunit is encoded by a distinct gene: Abpa, Abpb, and Abpg for the alpha, beta, and gamma subunits, respectively. We report here the structures and sequences of the genes that encode these three subunits. Each gene has three exons separated by two introns. Mouse salivary ABP is a member of the secretoglobin family, and we compare the structure of the three ABP subunit genes to those of 18 other mammalian secretoglobins. We map the three genes as a gene cluster located 10 cM from the centromere of Chromosome (Chr) 7 and show that Abpa is the closest of the three to the gene for glucose phosphate isomerase (GPI) and that Abpg is the closest to the centromere, with Abpb mapping between them. Abpa is oriented in the opposite direction to Abpb and Abpg, with its 5' end directed toward their 5' ends. We compare the location of these genes with other secretoglobin genes in the mouse genome and with the known locations of secretoglobin genes in the human genome and present evidence that strong positive selection has driven the divergence of the coding regions of Abpb and Abpg since the putative duplication event that created them.

  1. HER-2/neu oncogene amplification and chromosome 17 aneusomy in endometrial carcinoma: correlation with oncoprotein expression and conventional pathological parameters.

    Science.gov (United States)

    Cianciulli, A M; Guadagni, F; Marzano, R; Benevolo, M; Merola, R; Giannarelli, D; Marandino, F; Vocaturo, G; Mariani, L; Mottolese, M

    2003-06-01

    The objective of the present study was to evaluate the correlation between HER-2 gene amplification and HER-2 protein overexpression in endometrial carcinoma using fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC). We also analyzed chromosome 17 aneusomy and the association between these biological parameters and conventional clinicopathological variables. FISH analysis was performed on 73 selected paraffin-embedded sections from endometrial carcinomas which previously had HER-2 status determined immunohistochemically using monoclonal antibodies (MoAb) 300G9 and CB11. Using a ratio of more than two oncogene signals/centromere to indicate amplification, a total of 42 out of the 73 endometrial tumors included in this study resulted positive by FISH where as protein overexpression was identified in 29 out of 73 with a concordance rate of 74.3%. However, when the mean signals/centromere per nucleus increased (ratio > 4 2 4 < or = 5 when we grouped the amplified cases on the basis of HER-2:CEP17 ratio. In conclusion, molecular characteristics provide objective data that may be useful in predicting prognosis in patients with endometrial cancer.

  2. Species-specific and mating type-specific DNA regions adjacent to mating type idiomorphs in the genus Neurospora.

    Science.gov (United States)

    Randall, T A; Metzenberg, R L

    1995-09-01

    Mating type idiomorphs control mating and subsequent sexual development in Neurospora crassa and were previously shown to be well conserved in other Neurospora species. The centromere-proximal flanks of the A and a idiomorphs, but not the distal flanks from representative heterothallic, pseudohomothallic, and homothallic Neurospora species contain apparent species-specific and/or mating type-specific sequences adjacent to the well-conserved idiomorphs. The variable flank is bordered by regions that are highly homologous in all species. The sequence of approximately 1 kb immediately flanking the conserved idiomorphs of each species was determined. Sequence identity between species ranged from 20% (essentially unrelated) to > 90%. By contrast, the mt-A1 gene shows 88-98% identity. Sequence and hybridization data also show that the centromere-proximal flanks are very different between the two mating types for N. intermedia, N. discreta, and N. tetrasperma, but not for N. sitophila and N. crassa. The data suggest a close evolutionary relationship between several of the species; this is suppported by phylogenetic analysis of their respective mt-A1 genes. The origin of the variable regions adjacent to the evolutionarily conserved mating type idiomorphs is unknown.

  3. Mechanism of DNA Segregation in Prokaryotes: Replicon Pairing by parC of Plasmid R1

    Science.gov (United States)

    Jensen, Rasmus Bugge; Lurz, Rudi; Gerdes, Kenn

    1998-07-01

    Prokaryotic chromosomes and plasmids encode partitioning systems that are required for DNA segregation at cell division. The systems are thought to be functionally analogous to eukaryotic centromeres and to play a general role in DNA segregation. The parA system of plasmid R1 encodes two proteins ParM and ParR, and a cis-acting centromere-like site denoted parC. The ParR protein binds to parC in vivo and in vitro. The ParM protein is an ATPase that interacts with ParR specifically bound to parC. Using electron microscopy, we show here that parC mediates efficient pairing of plasmid molecules. The pairing requires binding of ParR to parC and is stimulated by the ParM ATPase. The ParM mediated stimulation of plasmid pairing is dependent on ATP hydrolysis by ParM. Using a ligation kinetics assay, we find that ParR stimulates ligation of parC-containing DNA fragments. The rate-of-ligation was increased by wild type ParM protein but not by mutant ParM protein deficient in the ATPase activity. Thus, two independent assays show that parC mediates pairing of plasmid molecules in vitro. These results are consistent with the proposal that replicon pairing is part of the mechanism of DNA segregation in prokaryotes.

  4. Tetrasomy 13q31.1qter due to an inverted duplicated neocentric marker chromosome in a fetus with multiple malformations.

    Science.gov (United States)

    Haddad, Véronique; Aboura, Azzedine; Tosca, Lucie; Guediche, Narjes; Mas, Anne-Elisabeth; L'Herminé, Aurore Coulomb; Druart, Luc; Picone, Olivier; Brisset, Sophie; Tachdjian, Gérard

    2012-04-01

    Small supernumerary marker chromosome (sSMC) lacking alpha satellite DNA or endogenous centromere regions are rare and contain fully functional centromeres, called neocentromeres. We report on a woman with a 14-week gestation pregnancy with a cystic hygroma and cerebellar hypoplasia at ultrasound examination. Cytogenetic studies showed a karyotype 47,XY,+mar dn. This sSMC was observed in chorionic villi, lung, and muscle tissue. Array Comparative Genomic Hybridization showed a gain from 13q31.1 to 13qter region. Fluorescent in situ hybridization with pan alpha satellite probe and probes specific for chromosome 13 showed a marker corresponding to an inversion duplication of the 13q distal chromosomal region without alpha satellite DNA sequence, suggesting the presence of a neocentromere. Examination of the fetus showed dysmorphic features, cystic cervical hygroma, postaxial polydactyly of the right hand and left foot with short fingers, malrotation of the gut, and a micropenis with hypospadias. Genotype-phenotype correlation in tetrasomy 13q is discussed according to the four 13q chromosomal breakpoints reported (13q32, 13q31, 13q21, 13q14) for chromosome 13 supernumerary markers.

  5. Cytogenetic, molecular and testicular tissue studies in an infertile 45,X male carrying an unbalanced (Y;22) translocation: case report.

    Science.gov (United States)

    Brisset, S; Izard, V; Misrahi, M; Aboura, A; Madoux, S; Ferlicot, S; Schoevaert, D; Soufir, J C; Frydman, R; Tachdjian, G

    2005-08-01

    (Y;autosome) translocations have been reported in association with male infertility. Different mechanisms have been suggested to explain the male infertility, such as deletion of the azoospermic factor (AZF) on the long arm of the Y chromosome, or meiosis impairment. We describe a new case with a de novo unbalanced translocation t(Y;22) and discuss the genotype-phenotype correlation. A 36 year old male with azoospermia was found to have a mosaic 45,X/46,X, + mar karyotype. Fluorescence in situ hybridization (FISH) showed the presence of a derivative Y chromosome containing the short arm, the centromere and a small proximal part of the long-arm euchromatin of the Y chromosome and the long arm of chromosome 22. The unstable small marker chromosome included the short arm and the centromere of chromosome 22. This unbalanced translocation t(Y;22)(q11.2;q11.1) generated the loss of the long arm of the Y chromosome involving a large part of AZFb, AZFc and Yq heterochromatin regions. Testicular tissue analyses showed sperm in the wet preparation. Our case shows the importance of documenting (Y;autosome) translocations with molecular and testicular tissue analyses.

  6. Comprehensive cytological characterization of the Gossypium hirsutum genome based on the development of a set of chromosome cytological markers

    Directory of Open Access Journals (Sweden)

    Wenbo Shan

    2016-08-01

    Full Text Available Cotton is the world's most important natural fiber crop. It is also a model system for studying polyploidization, genomic organization, and genome-size variation. Integrating the cytological characterization of cotton with its genetic map will be essential for understanding its genome structure and evolution, as well as for performing further genetic-map based mapping and cloning. In this study, we isolated a complete set of bacterial artificial chromosome clones anchored to each of the 52 chromosome arms of the tetraploid cotton Gossypium hirsutum. Combining these with telomere and centromere markers, we constructed a standard karyotype for the G. hirsutum inbred line TM-1. We dissected the chromosome arm localizations of the 45S and 5S rDNA and suggest a centromere repositioning event in the homoeologous chromosomes AT09 and DT09. By integrating a systematic karyotype analysis with the genetic linkage map, we observed different genome sizes and chromosomal structures between the subgenomes of the tetraploid cotton and those of its diploid ancestors. Using evidence of conserved coding sequences, we suggest that the different evolutionary paths of non-coding retrotransposons account for most of the variation in size between the subgenomes of tetraploid cotton and its diploid ancestors. These results provide insights into the cotton genome and will facilitate further genome studies in G. hirsutum.

  7. Compaction and transport properties of newly replicated Caulobacter crescentus DNA.

    Science.gov (United States)

    Hong, Sun-Hae; McAdams, Harley H

    2011-12-01

    Upon initiating replication of the Caulobacter chromosome, one copy of the parS centromere remains at the stalked pole; the other moves to the distal pole. We identified the segregation dynamics and compaction characteristics of newly replicated Caulobacter DNA during transport (highly variable from cell to cell) using time-lapse fluorescence microscopy. The parS centromere and a length (also highly variable) of parS proximal DNA on each arm of the chromosome are segregated with the same relatively slow transport pattern as the parS locus. Newly replicated DNA further than about 100 kb from parS segregates with a different and faster pattern, while loci at 48 kb from parS segregate with the slow pattern in some cells and the fast pattern in others. The observed parS-proximal DNA compaction characteristics have scaling properties that suggest the DNA is branched. HU2-deletion strains exhibited a reduced compaction phenotype except near the parS site where only the ΔHU1ΔHU2 double mutant had a compaction phenotype. The chromosome shows speed-dependent extension during translocation suggesting the DNA polymer is under tension. While DNA segregation is highly reliable and succeeds in virtually all wild-type cells, the high degree of cell to cell variation in the segregation process is noteworthy.

  8. pTAR-encoded proteins in plasmid partitioning.

    Science.gov (United States)

    Kalnin, K; Stegalkina, S; Yarmolinsky, M

    2000-04-01

    Partition cassettes, essential for the segregational stability of low-copy-number bacterial plasmids, typically encode two autoregulated proteins and an adjacent cis-acting centromere analog to which one or perhaps both proteins bind. The diminutive partition region of pTAR of Agrobacterium spp. was reported to be exceptional, encoding only a single protein, ParA (D. R. Gallie and C. I. Kado, J. Mol. Biol. 193:465-478, 1987). However, resequencing of the region revealed two small downstream genes, parB and orf-84, of which only parB was found to be essential for partitioning in A. tumefaciens. Purified ParA exhibited a weak ATPase activity that was modestly increased by nonspecific DNA. ParB bound in vitro to repeated sequences present in a region, parS, that possesses centromere and operator functions and within which we identified the primary transcription start site by primer extension. In certain respects the Par proteins behave normally in the foreign host Escherichia coli. In E. coli, as in A. tumefaciens, ParB repressed the partition operon; ParA, inactive alone, augmented this repression. Functional similarities between the partition system of pTAR and those of other plasmids and bacteria are prominent, despite differences in size, organization, and amino acid sequence.

  9. Locational diversity of alpha satellite DNA and intergeneric hybridization aspects in the Nomascus and Hylobates genera of small apes.

    Directory of Open Access Journals (Sweden)

    Sudarath Baicharoen

    Full Text Available Recently, we discovered that alpha satellite DNA has unique and genus-specific localizations on the chromosomes of small apes. This study describes the details of alpha satellite localization in the genera Nomascus and Hylobates and explores their usefulness in distinguishing parental genome sets in hybrids between these genera. Fluorescence in situ hybridization was used to establish diagnostic criteria of alpha satellite DNA markers in discriminating small ape genomes. In particular we established the genus specificity of alpha satellite distribution in three species of light-cheeked gibbons (Nomascus leucogenys, N. siki, and N. gabriellae in comparison to that of Hylobates lar. Then we determined the localization of alpha satellite DNA in a hybrid individual which resulted from a cross between these two genera. In Nomascus the alpha satellite DNA blocks were located at the centromere, telomere, and four interstitial regions. In Hylobates detectable amounts of alpha satellite DNA were seen only at centromeric regions. The differences in alpha satellite DNA locations between Nomascus and Hylobates allowed us to easily distinguish the parental chromosomal sets in the genome of intergeneric hybrid individuals found in Thai and Japanese zoos. Our study illustrates how molecular cytogenetic markers can serve as diagnostic tools to identify the origin of individuals. These molecular tools can aid zoos, captive breeding programs and conservation efforts in managing small apes species. Discovering more information on alpha satellite distribution is also an opportunity to examine phylogenetic and evolutionary questions that are still controversial in small apes.

  10. Locational diversity of alpha satellite DNA and intergeneric hybridization aspects in the Nomascus and Hylobates genera of small apes.

    Science.gov (United States)

    Baicharoen, Sudarath; Miyabe-Nishiwaki, Takako; Arsaithamkul, Visit; Hirai, Yuriko; Duangsa-ard, Kwanruen; Siriaroonrat, Boripat; Domae, Hiroshi; Srikulnath, Kornsorn; Koga, Akihiko; Hirai, Hirohisa

    2014-01-01

    Recently, we discovered that alpha satellite DNA has unique and genus-specific localizations on the chromosomes of small apes. This study describes the details of alpha satellite localization in the genera Nomascus and Hylobates and explores their usefulness in distinguishing parental genome sets in hybrids between these genera. Fluorescence in situ hybridization was used to establish diagnostic criteria of alpha satellite DNA markers in discriminating small ape genomes. In particular we established the genus specificity of alpha satellite distribution in three species of light-cheeked gibbons (Nomascus leucogenys, N. siki, and N. gabriellae) in comparison to that of Hylobates lar. Then we determined the localization of alpha satellite DNA in a hybrid individual which resulted from a cross between these two genera. In Nomascus the alpha satellite DNA blocks were located at the centromere, telomere, and four interstitial regions. In Hylobates detectable amounts of alpha satellite DNA were seen only at centromeric regions. The differences in alpha satellite DNA locations between Nomascus and Hylobates allowed us to easily distinguish the parental chromosomal sets in the genome of intergeneric hybrid individuals found in Thai and Japanese zoos. Our study illustrates how molecular cytogenetic markers can serve as diagnostic tools to identify the origin of individuals. These molecular tools can aid zoos, captive breeding programs and conservation efforts in managing small apes species. Discovering more information on alpha satellite distribution is also an opportunity to examine phylogenetic and evolutionary questions that are still controversial in small apes.

  11. Split hand/foot malformation genetics supports the chromosome 7 copy segregation mechanism for human limb development

    Science.gov (United States)

    Klar, Amar J. S.

    2016-01-01

    Genetic aberrations of several unlinked loci cause human congenital split hand/foot malformation (SHFM) development. Mutations of the DLX5 (distal-less) transcription factor-encoding gene in chromosome 7 cause SHFM through haploinsufficiency, but the vast majority of cases result from heterozygous chromosomal aberrations of the region without mutating the DLX5 gene. To resolve this paradox, we invoke a chromosomal epigenetic mechanism for limb development. It is composed of a monochromatid gene expression phenomenon that we discovered in two fission yeasts with the selective chromosome copy segregation phenomenon that we discovered in mouse cells. Accordingly, one daughter cell inherits both expressed DLX5 copies while the other daughter inherits both epigenetically silenced ones from a single deterministic cell of the developing limb. Thus, differentiated daughter cells after further proliferation will correspondingly produce proximal/distal-limb tissues. Published results of a Chr. 7 translocation with a centromere-proximal breakpoint situated over 41 million bases away from the DLX locus, centromeric and DLX5-region inversions have satisfied key genetic and developmental biology predictions of the mechanism. Further genetic tests of the mechanism are proposed. We propose that the DNA double helical structure itself causes the development of sister cells' gene regulation asymmetry. We also argue against the conventionally invoked morphogen model of development. This article is part of the themed issue ‘Provocative questions in left–right asymmetry’. PMID:27821526

  12. Quantitative trait loci (QTL mapping for growth traits on bovine chromosome 14

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    Marcelo Miyata

    2007-03-01

    Full Text Available Quantitative trait loci (QTL mapping in livestock allows the identification of genes that determine the genetic variation affecting traits of economic interest. We analyzed the birth weight and weight at 60 days QTL segregating on bovine chromosome BTA14 in a F2 resource population using genotypes produced from seven microsatellite markers. Phenotypes were derived from 346 F2 progeny produced from crossing Bos indicus Gyr x Holstein Bos taurus F1 parents. Interval analysis to detect QTL for birth weight revealed the presence of a QTL (p < 0.05 at 1 centimorgan (cM from the centromere with an additive effect of 1.210 ± 0.438 kg. Interval analysis for weight at 60 days revealed the presence of a QTL (p < 0.05 at 0 cM from the centromere with an additive effect of 2.122 ± 0.735 kg. The region to which the QTL were assigned is described in the literature as responsible for some growth traits, milk yield, milk composition, fat deposition and has also been related to reproductive traits such as daughter pregnancy rate and ovulation rate. The effects of the QTL described on other traits were not investigated.

  13. Cytogenetic studies in six species of Scinax (Anura, Hylidae clade Scinax ruber from northern and northeastern Brazil

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    Lídia Nogueira

    2015-06-01

    Full Text Available Scinax species are still underrepresented in cytogenetic studies, mainly with respect to populations from northeastern and northern Brazil. In this study, we provide new chromosomal information on Scinax boesemani, S. camposseabrai, S. garbei, S. pachycrus, S. trilineatus and S. x-signatus, all belonging to clade S. ruber. They were collected at two locations in the Caatinga biome (northeastern Brazil and at one in the Amazon (northern Brazil biomes. Chromosomes were analyzed by conventional staining, C-banding, Ag-NOR staining, and fluorochrome staining. All species shared a modal diploid value of 2n = 24 and fundamental arm number (FN of 48. Moreover, both chromosomal size and morphology were similar to other species in this Scinax clade. C-banding revealed centromeric heterochromatin in all species, along with terminal species-specific C-bands in some species. Active nucleolar organizer regions (Ag-NORs were identified at 11q in most species, except for S. boesemani and S. garbei (Ag-NORs at interstitial region of 8q. Differing from most anurans, GC-rich regions were not restricted to NORs, but also coincident with some centromeric and terminal C-bands. These data contribute to the cytotaxonomy of Scinax by providing chromosomal markers and demonstrating the occurrence of microstructural rearrangements and inversions on chromosomal evolution of Scinax.

  14. Ipl1/Aurora-B is necessary for kinetochore restructuring in meiosis I in Saccharomyces cerevisiae.

    Science.gov (United States)

    Meyer, Régis E; Chuong, Hoa H; Hild, Marrett; Hansen, Christina L; Kinter, Michael; Dawson, Dean S

    2015-09-01

    In mitosis, the centromeres of sister chromosomes are pulled toward opposite poles of the spindle. In meiosis I, the opposite is true: the sister centromeres move together to the same pole, and the homologous chromosomes are pulled apart. This change in segregation patterns demands that between the final mitosis preceding meiosis and the first meiotic division, the kinetochores must be restructured. In budding yeast, unlike mammals, kinetochores are largely stable throughout the mitotic cycle. In contrast, previous work with budding and fission yeast showed that some outer kinetochore proteins are lost in early meiosis. We use quantitative mass spectrometry methods and imaging approaches to explore the kinetochore restructuring process that occurs in meiosis I in budding yeast. The Ndc80 outer kinetochore complex, but not other subcomplexes, is shed upon meiotic entry. This shedding is regulated by the conserved protein kinase Ipl1/Aurora-B and promotes the subsequent assembly of a kinetochore that will confer meiosis-specific segregation patterns on the chromosome.

  15. Meikin is a conserved regulator of meiosis-I-specific kinetochore function.

    Science.gov (United States)

    Kim, Jihye; Ishiguro, Kei-ichiro; Nambu, Aya; Akiyoshi, Bungo; Yokobayashi, Shihori; Kagami, Ayano; Ishiguro, Tadashi; Pendas, Alberto M; Takeda, Naoki; Sakakibara, Yogo; Kitajima, Tomoya S; Tanno, Yuji; Sakuno, Takeshi; Watanabe, Yoshinori

    2015-01-22

    The kinetochore is the crucial apparatus regulating chromosome segregation in mitosis and meiosis. Particularly in meiosis I, unlike in mitosis, sister kinetochores are captured by microtubules emanating from the same spindle pole (mono-orientation) and centromeric cohesion mediated by cohesin is protected in the following anaphase. Although meiotic kinetochore factors have been identified only in budding and fission yeasts, these molecules and their functions are thought to have diverged earlier. Therefore, a conserved mechanism for meiotic kinetochore regulation remains elusive. Here we have identified in mouse a meiosis-specific kinetochore factor that we termed MEIKIN, which functions in meiosis I but not in meiosis II or mitosis. MEIKIN plays a crucial role in both mono-orientation and centromeric cohesion protection, partly by stabilizing the localization of the cohesin protector shugoshin. These functions are mediated mainly by the activity of Polo-like kinase PLK1, which is enriched to kinetochores in a MEIKIN-dependent manner. Our integrative analysis indicates that the long-awaited key regulator of meiotic kinetochore function is Meikin, which is conserved from yeasts to humans.

  16. The cohesion stabilizer sororin favors DNA repair and chromosome segregation during mouse oocyte meiosis.

    Science.gov (United States)

    Huang, Chun-Jie; Yuan, Yi-Feng; Wu, Di; Khan, Faheem Ahmed; Jiao, Xiao-Fei; Huo, Li-Jun

    2017-03-01

    Maintenance and timely termination of cohesion on chromosomes ensures accurate chromosome segregation to guard against aneuploidy in mammalian oocytes and subsequent chromosomally abnormal pregnancies. Sororin, a cohesion stabilizer whose relevance in antagonizing the anti-cohesive property of Wings-apart like protein (Wapl), has been characterized in mitosis; however, the role of Sororin remains unclear during mammalian oocyte meiosis. Here, we show that Sororin is required for DNA damage repair and cohesion maintenance on chromosomes, and consequently, for mouse oocyte meiotic program. Sororin is constantly expressed throughout meiosis and accumulates on chromatins at germinal vesicle (GV) stage/G2 phase. It localizes onto centromeres from germinal vesicle breakdown (GVBD) to metaphase II stage. Inactivation of Sororin compromises the GVBD and first polar body extrusion (PBE). Furthermore, Sororin inactivation induces DNA damage indicated by positive γH2AX foci in GV oocytes and precocious chromatin segregation in MII oocytes. Finally, our data indicate that PlK1 and MPF dissociate Sororin from chromosome arms without affecting its centromeric localization. Our results define Sororin as a determinant during mouse oocyte meiotic maturation by favoring DNA damage repair and chromosome separation, and thereby, maintaining the genome stability and generating haploid gametes.

  17. Role of Securin, Separase and Cohesins in female meiosis and polar body formation in Drosophila.

    Science.gov (United States)

    Guo, Zhihao; Batiha, Osamah; Bourouh, Mohammed; Fifield, Eric; Swan, Andrew

    2016-02-01

    Chromosome segregation in meiosis is controlled by a conserved pathway that culminates in Separase-mediated cleavage of the α-kleisin Rec8, leading to dissolution of cohesin rings. Drosophila has no gene encoding Rec8, and the absence of a known Separase target raises the question of whether Separase and its regulator Securin (Pim in Drosophila) are important in Drosophila meiosis. Here, we investigate the role of Securin, Separase and the cohesin complex in female meiosis using fluorescence in situ hybridization against centromeric and arm-specific sequences to monitor cohesion. We show that Securin destruction and Separase activity are required for timely release of arm cohesion in anaphase I and centromere-proximal cohesion in anaphase II. They are also required for release of arm cohesion on polar body chromosomes. Cohesion on polar body chromosomes depends on the cohesin components SMC3 and the mitotic α-kleisin Rad21 (also called Vtd in Drosophila). We provide cytological evidence that SMC3 is required for arm cohesion in female meiosis, whereas Rad21, in agreement with recent findings, is not. We conclude that in Drosophila meiosis, cohesion is regulated by a conserved Securin-Separase pathway that targets a diverged Separase target, possibly within the cohesin complex.

  18. 辐射诱导染色体畸变的快速FISH方法的建立%Establishment of a rapid FISH method to analyze chromosome aberrations induced by Establishment of a rapid FISH method to analyze chromosome aberrations induced by irradiation irradiation

    Institute of Scientific and Technical Information of China (English)

    李爽; 陆雪; 赵骅; 封江彬; 刘青杰

    2014-01-01

    目的:建立一种检测辐射诱导染色体畸变的快速荧光原位杂交(FISH)方法。方法:两步简并引物PCR法扩增人α-卫60星DNA,制备荧光基团直接标记的泛着丝粒探针;对 Coγ射线照射后的人外周血淋巴细胞染色体标本进行FISH分析,荧光显微镜下检测着丝粒及染色体形态。结果:直接标记泛着丝粒探针杂交后显示所有染色体着丝粒均有较强的信号;应用制备的FISH探针检测到γ射线照射诱导的双着丝粒、着丝粒环和易位染色体畸变,37℃杂交5-12 h后经过简单的洗涤即可在荧光显微镜下检测到较好的信号。结论:本研究制备的直接标记泛着丝粒探针,可用于FISH快速检测辐射诱导的染色体双着丝粒体、着丝粒环和易位畸变。目的:建立一种检测辐射诱导染色体畸变的快速荧光原位杂交(FISH)方法。方法:两步简并引物PCR法扩增人α-卫60星DNA,制备荧光基团直接标记的泛着丝粒探针;对 Coγ射线照射后的人外周血淋巴细胞染色体标本进行FISH分析,荧光显微镜下检测着丝粒及染色体形态。结果:直接标记泛着丝粒探针杂交后显示所有染色体着丝粒均有较强的信号;应用制备的FISH探针检测到γ射线照射诱导的双着丝粒、着丝粒环和易位染色体畸变,37℃杂交5-12 h后经过简单的洗涤即可在荧光显微镜下检测到较好的信号。结论:本研究制备的直接标记泛着丝粒探针,可用于FISH快速检测辐射诱导的染色体双着丝粒体、着丝粒环和易位畸变。%OBJECTIVE: To establish a rapid fluorescence in situ hybridization (FISH) method and to analyze the chromosome aberrations induced by irradiation.METHODS:Human centromeric alpha satellite DNA was amplified and directly labeled with Cy3-dUTP or Fluorescein-12-dUTP by degenerate oligonucleotide priming-PCR (DOP-PCR) to prepare two pan-centromeric probes. Chromosome

  19. Genetic modifiers of Hb E/β0 thalassemia identified by a two-stage genome-wide association study

    Directory of Open Access Journals (Sweden)

    Winichagoon Pranee

    2010-03-01

    Full Text Available Abstract Background Patients with Hb E/β0 thalassemia display remarkable variability in disease severity. To identify genetic modifiers influencing disease severity, we conducted a two-stage genome scan in groups of 207 mild and 305 severe unrelated patients from Thailand with Hb E/β0 thalassemia and normal α-globin genes. Methods First, we estimated and compared the allele frequencies of approximately 110,000 gene-based single nucleotide polymorphisms (SNPs in pooled DNAs from different severity groups. The 756 SNPs that showed reproducible allelic differences at P Results After adjustment for age, gender and geographic region, logistic regression models showed 50 SNPs significantly associated with disease severity (P P = 2.6 × 10-13. Seven SNPs in two distinct LD blocks within a region centromeric to the β-globin gene cluster that contains many olfactory receptor genes were also associated with disease severity; rs3886223 had the strongest association (OR = 3.03, P = 3.7 × 10-11. Several previously unreported SNPs were also significantly associated with disease severity. Conclusions These results suggest that there may be an additional regulatory region centromeric to the β-globin gene cluster that affects disease severity by modulating fetal hemoglobin expression.

  20. MAPK-activated protein kinase 2 is required for mouse meiotic spindle assembly and kinetochore-microtubule attachment.

    Directory of Open Access Journals (Sweden)

    Ju Yuan

    Full Text Available MAPK-activated protein kinase 2 (MK2, a direct substrate of p38 MAPK, plays key roles in multiple physiological functions in mitosis. Here, we show for the first time the unique distribution pattern of MK2 in meiosis. Phospho-MK2 was localized on bipolar spindle minus ends and along the interstitial axes of homologous chromosomes extending over centromere regions and arm regions at metaphase of first meiosis (MI stage in mouse oocytes. At metaphase of second meiosis (MII stage, p-MK2 was localized on the bipolar spindle minus ends and at the inner centromere region of sister chromatids as dots. Knockdown or inhibition of MK2 resulted in spindle defects. Spindles were surrounded by irregular nondisjunction chromosomes, which were arranged in an amphitelic or syntelic/monotelic manner, or chromosomes detached from the spindles. Kinetochore-microtubule attachments were impaired in MK2-deficient oocytes because spindle microtubules became unstable in response to cold treatment. In addition, homologous chromosome segregation and meiosis progression were inhibited in these oocytes. Our data suggest that MK2 may be essential for functional meiotic bipolar spindle formation, chromosome segregation and proper kinetochore-microtubule attachments.

  1. Physical mapping of the retinoid X receptor B gene in mouse and human

    Energy Technology Data Exchange (ETDEWEB)

    Nagata, T.; Kitagawa, K.; Taketo, M. [Banyu Tsukuba Research Institute, Tsukuba (Japan); Weiss, E.H. [Ludwig-Maximilians-Univ., Munich (Germany); Abe, K. [Kumamoto Univ. School of Medicine, Kumamoto (Japan); Ando, A.; Yara-Kikuti, Y.; Inoko, H. [Tokai Univ. School of Medicine, Isehara (Japan); Seldin, M.F. [Duke Univ. Medical Center, Durham, NC (United States); Ozato, K. [National Institutes of Health, Bethesda, MD (United States)

    1995-01-11

    Retinoid X receptors (RXRs) are zinc finger-containing nuclear transcription factors. They belong to the nuclear receptor superfamily that contains retinoid receptors, vitamin D receptors, thyroid hormone receptors, and steroid hormone receptors as well as the so-called orphan receptors. We previously mapped all three RXR genes on mouse chromosomes, using a panel of Mus spretus-Mus musculus interspecific backcross mice: namely, the RXRA-gene (Rxra) on Chr 2 near the centromere, the RXRB gene (Rxrb) on Chr 17 in the H2 region, and the RXRG gene (Rxrg) on distal Chr 1. Using cosmid clones that cover the major histocompatibility complex (MHC) region, we determined the precise physical map positions of the gene encoding mouse and human RXRB, respectively. The mouse gene (Rxrb) maps between H2-Ke4 and H2-Ke5: namely, immediately telomeric to H2-Ke4 which encodes a histidine-rich transmembrane protein, and 12 kilobases centromeric to H2-Ke5 which is expressed in lymphoid tissues, Rxrb and H2-Ke4 are transcribed into opposite directions from a CpG-rich promoter of about 250 base pairs. This gene organization is well conserved also in the human genome at the HLA-DP subregion of Chr 6p, underscoring the strong conservation of the gene organization in the MHC region between the two mammals. 54 refs., 4 figs.

  2. A new and unusual reciprocal translocation in cattle: rcp(11;25)(q11;q14-21).

    Science.gov (United States)

    Perucatti, A; Genualdo, V; Iannuzzi, A; De Lorenzi, L; Matassino, D; Parma, P; Di Berardino, D; Iannuzzi, L; Di Meo, G P

    2011-01-01

    A new and unusual reciprocal translocation was detected in a heifer of the Agerolese cattle breed during a routine cytogenetic screening carried out on 13 animals (2 males and 11 females) kept at the ConSDABI Conservation Center in Benevento (Southern Italy). The 13 animals investigated had a normal karyotype except for a 1-year-old female, which carried one autosome smaller than the smallest normal bovine autosomes. This small autosome showed very little C-banding in comparison to the other autosomes, while another medium-sized autosome showed 2 distinct and prominent C-bands. RBA-banding and karyotype analysis revealed that these 2 chromosomes were the result of a reciprocal translocation between chromosomes 11 and 25. FISH analysis with BAC142G06 mapping to the proximal (subcentromeric) region of both BTA25 and der11, BAC513H08 (ELN) mapping to BTA25q22dist and der25, and BAC533C11 mapping to the proximal region of BTA11 and der11 confirmed the localization of the breakpoints on band q11 (centromere) of chromosome 11 and q14-21 of chromosome 25. Ag-NOR and sequential RBA/Ag-NOR techniques detected the presence of NORs on both BTA11 and BTA25 and both der11 and der25. To our knowledge, this is the first report of a reciprocal translocation event in cattle with the breakpoint located in the centromeric region.

  3. Cytogenetical anchoring of sheep linkage map and syntenic groups using a sheep BAC library

    Directory of Open Access Journals (Sweden)

    Cribiu Edmond-Paul

    2000-07-01

    Full Text Available Abstract In order to simultaneously integrate linkage and syntenic groups to the ovine chromosomal map, a sheep bacterial artificial chromosome (BAC library was screened with previously assigned microsatellites using a sheep-hamster hybrid panel and genetic linkage. Thirty-three BACs were obtained, fluorescently labelled and hybridised on sheep-goat hybrid metaphases (2n = 57. This study allowed us, (i, to anchor all linkage groups on sheep chromosomes, (ii, to give information on the probable position of the centromere on the linkage map for the centromeric chromosomes, (iii, to contradict the previous orientation of the ovine × linkage group by the mapping of BMS1008 on OARXq38. Concerning our somatic cell hybrid panel, this study resulted in the assignment of all the previously unassigned groups to ovine chromosomes and a complete characterisation of the hybrid panel. In addition, since hybridisations were performed on a sheep-goat hybrid, new marker/anchoring points were added to the caprine cytogenetic map.

  4. History of chromosome rearrangements reflects the spatial organization of yeast chromosomes.

    Science.gov (United States)

    Khrameeva, Ekaterina E; Fudenberg, Geoffrey; Gelfand, Mikhail S; Mirny, Leonid A

    2016-04-01

    Three-dimensional (3D) organization of genomes affects critical cellular processes such as transcription, replication, and deoxyribo nucleic acid (DNA) repair. While previous studies have investigated the natural role, the 3D organization plays in limiting a possible set of genomic rearrangements following DNA repair, the influence of specific organizational principles on this process, particularly over longer evolutionary time scales, remains relatively unexplored. In budding yeast S.cerevisiae, chromosomes are organized into a Rabl-like configuration, with clustered centromeres and telomeres tethered to the nuclear periphery. Hi-C data for S.cerevisiae show that a consequence of this Rabl-like organization is that regions equally distant from centromeres are more frequently in contact with each other, between arms of both the same and different chromosomes. Here, we detect rearrangement events in Saccharomyces species using an automatic approach, and observe increased rearrangement frequency between regions with higher contact frequencies. Together, our results underscore how specific principles of 3D chromosomal organization can influence evolutionary events.

  5. Suppression of genome instability in pRB-deficient cells by enhancement of chromosome cohesion.

    Science.gov (United States)

    Manning, Amity L; Yazinski, Stephanie A; Nicolay, Brandon; Bryll, Alysia; Zou, Lee; Dyson, Nicholas J

    2014-03-20

    Chromosome instability (CIN), a common feature of solid tumors, promotes tumor evolution and increases drug resistance during therapy. We previously demonstrated that loss of the retinoblastoma protein (pRB) tumor suppressor causes changes in centromere structure and generates CIN. However, the mechanism and significance of this change was unclear. Here, we show that defects in cohesion are key to the pRB loss phenotype. pRB loss alters H4K20 methylation, a prerequisite for efficient establishment of cohesion at centromeres. Changes in cohesin regulation are evident during S phase, where they compromise replication and increase DNA damage. Ultimately, such changes compromise mitotic fidelity following pRB loss. Remarkably, increasing cohesion suppressed all of these phenotypes and dramatically reduced CIN in cancer cells lacking functional pRB. These data explain how loss of pRB undermines genomic integrity. Given the frequent functional inactivation of pRB in cancer, conditions that increase cohesion may provide a general strategy to suppress CIN.

  6. Three Supernumerary Marker Chromosomes in a Patient with Developmental Delay, Mental Retardation, and Dysmorphic Features

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    Jie Hu

    2011-01-01

    Full Text Available We characterized three supernumerary marker chromosomes (SMCs simultaneously present in a 2-year- and 10-month-old male patient with mental retardation and dysmorphic features. Peripheral blood chromosome analysis revealed two to three SMCs in 25/26 cells analyzed. The remaining one cell had one SMC. Microarray comparative genomic hybridization (aCGH showed mosaicism for gains of 5q35.3, 15q11.2q13.3, and 18p11.21q11.1 regions. All three gains contain multiple OMIM genes. FISH studies indicated that one of the SMCs is a dicentric ring 15 with two copies of the 15q11.2q13.3 region including SNRPN/UBE3A and two copies of the 5q35.3 region. One of the der(18s contains the 18 centromere and 18p11.2 regions, while the other der(18 has a signal for the 18 centromere only. The phenotype of the patient is compared with that of patients with tetrasomy 15q11.2q13.3, trisomy 5q35.3, and trisomy 18p11.2. Our study demonstrates that aCGH and FISH analyses are powerful tools, which complement the conventional cytogenetic analysis for the identification of SMCs.

  7. Trisomy 1 and 8 occur frequently in hepatocellular carcinoma but not in liver cell adenoma and focal nodular hyperplasia. A fluorescence in situ hybridization study.

    Science.gov (United States)

    Nasarek, A; Werner, M; Nolte, M; Klempnauer, J; Georgii, A

    1995-01-01

    Conventional cytogenetic studies revealed gains and structural aberrations of chromosome 1 to be the most consistent chromosomal aberrations in hepatocellular carcinoma (HCC). We investigated touch preparations of eight HCC, five cholangiocellular carcinomas (CCC), five liver cell adenomas (LCA), four focal nodular hyperplasias (FNH) as well as nine specimens of normal liver tissue using fluorescence in situ hybridization (FISH) with centromere specific probes for chromosomes 1 and 8. Polysomies of chromosome 1, especially trisomy 1, were found in five of eight HCC and four of five CCC but in no normal liver tissue or benign tumour. Only three of seven cases of HCC revealed trisomy 8 whereas the five benign liver tumours and all normal liver tissues examined had disomy 8. Our results confirm conventional cytogenetic findings in terms of chromosome 1 aberrations in HCC although they are not specific for these types of malignant liver tumours. Since alpha-satellite probes were used in our study, only gains or losses including the centromeric regions of the chromosomes 1 and 8 could be detected. Nevertheless, our findings suggest that FISH may help in the differential diagnosis of malignant versus benign neoplasms of the liver.

  8. A propagating ATPase gradient drives transport of surface-confined cellular cargo.

    Science.gov (United States)

    Vecchiarelli, Anthony G; Neuman, Keir C; Mizuuchi, Kiyoshi

    2014-04-01

    The faithful segregation of duplicated genetic material into daughter cells is critical to all organisms. In many bacteria, the segregation of chromosomes involves transport of "centromere-like" loci over the main body of the chromosome, the nucleoid, mediated by a two-protein partition system: a nonspecific DNA-binding ATPase, ParA, and an ATPase stimulator, ParB, which binds to the centromere-like loci. These systems have previously been proposed to function through a filament-based mechanism, analogous to actin- or microtubule-based movement. Here, we reconstituted the F-plasmid partition system using a DNA-carpeted flow cell as an artificial nucleoid surface and magnetic beads coated with plasmid partition complexes as surface-confined cargo. This minimal system recapitulated directed cargo motion driven by a surface ATPase gradient that propagated with the cargo. The dynamics are consistent with a diffusion-ratchet model, whereby the cargo dynamically establishes, and interacts with, a concentration gradient of the ATPase. A chemophoresis force ensues as the cargo perpetually chases the ATPase gradient, allowing the cargo to essentially "surf" the nucleoid on a continuously traveling wave of the ATPase. Demonstration of this non-filament-based motility mechanism in a biological context establishes a distinct class of motor system used for the transport and positioning of large cellular cargo.

  9. Dicentric chromosome stretching during anaphase reveals roles of Sir2/Ku in chromatin compaction in budding yeast.

    Science.gov (United States)

    Thrower, D A; Bloom, K

    2001-09-01

    We have used mitotic spindle forces to examine the role of Sir2 and Ku in chromatin compaction. Escherichia coli lac operator DNA was placed between two centromeres on a conditional dicentric chromosome in budding yeast cells and made visible by expression of a lac repressor-green fluorescent fusion protein. Centromeres on the same chromatid of a dicentric chromosome attach to opposite poles approximately 50% of the time, resulting in chromosome bridges during anaphase. In cells deleted for yKU70, yKU80, or SIR2, a 10-kb region of the dicentric chromosome stretched along the spindle axis to a length of 6 microm during anaphase. On spindle disassembly, stretched chromatin recoiled to the bud neck and was partitioned to mother and daughter cells after cytokinesis and cell separation. Chromatin immunoprecipitation revealed that Sir2 localizes to the lacO region in response to activation of the dicentric chromosome. These findings indicate that Ku and Sir proteins are required for proper chromatin compaction within regions of a chromosome experiencing tension or DNA damage. The association of Sir2 with the affected region suggests a direct role in this process, which may include the formation of heterochromatic DNA.

  10. Nuclear oscillations and nuclear filament formation accompany single-strand annealing repair of a dicentric chromosome in Saccharomyces cerevisiae.

    Science.gov (United States)

    Thrower, Douglas A; Stemple, Jennifer; Yeh, Elaine; Bloom, Kerry

    2003-02-01

    Dicentric chromosomes undergo breakage during mitosis as a result of the attachment of two centromeres on one sister chromatid to opposite spindle poles. Studies utilizing a conditional dicentric chromosome III in Saccharomyces cerevisiae have shown that dicentric chromosome repair occurs primarily by deletion of one centromere via a RAD52-dependent recombination pathway. We report that dicentric chromosome resolution requires RAD1, a gene involved in the single-strand annealing DNA repair pathway. We additionally show that single-strand annealing repair of a dicentric chromosome can occur in the absence of RAD52. RAD52-independent repair requires the adaptation-defective cdc5-ad allele of the yeast polo kinase and the DNA damage checkpoint gene RAD9. Dicentric chromosome breakage in cdc5-ad rad52 mutant cells is associated with a prolonged mitotic arrest, during which nuclei undergo microtubule-dependent oscillations, accompanied by dynamic changes in nuclear morphology. We further demonstrate that the frequency of spontaneous direct repeat recombination is suppressed in yeast cells treated with benomyl, a drug that perturbs microtubules. Our findings indicate that microtubule-dependent processes facilitate recombination.

  11. The Dicentric Chromosome dic(20;22) Is a Recurrent Abnormality in Myelodysplastic Syndromes and Is a Product of Telomere Fusion.

    Science.gov (United States)

    MacKinnon, Ruth N; Duivenvoorden, Hendrika M; Campbell, Lynda J; Wall, Meaghan

    2017-03-04

    We describe a recurrent dicentric chromosome formed by telomere fusion between chromosome 20 and chromosome 22 in 4 cases of myelodysplastic syndromes (MDS) or acute myeloid leukaemia (AML). In particular, the presence of residual telomere sequences at the site of translocation in 3 of the 4 cases makes a compelling case for telomere fusion. This is the first description of a recurrent telomere fusion event in any malignant condition. The 20q subtelomeric region was retained in all 4 examples despite deletion of the 20q12 region closer to the centromere. The original dicentric chromosome in all 4 cases contained nucleolus organiser region material from the short arm of chromosome 22 and had also undergone secondary rearrangements that produced amplification of the common gained region on 20q. We propose that the sequence of events producing this chromosome abnormality is: degradation of the telomeres, formation of an unstable dicentric chromosome by 20q and 22p telomere fusion, breakage-fusion-bridge cycles causing copy number aberration between the centromeres, selection of cells with 20q12 deletion, and further selection of cells with 20q11.2 gain. The last 2 steps are driver events responsible for the abnormal chromosomes found in the malignant cells. Finding recurrent patterns in the complex genome reorganisation events that characterise poor-prognosis, complex-karyotype AML and MDS will help us understand the mechanisms and oncogenic driver mutations in these poorly understood malignancies.

  12. Acentric chromosome ends are prone to fusion with functional chromosome ends through a homology-directed rearrangement.

    Science.gov (United States)

    Ohno, Yuko; Ogiyama, Yuki; Kubota, Yoshino; Kubo, Takuya; Ishii, Kojiro

    2016-01-08

    The centromeres of many eukaryotic chromosomes are established epigenetically on potentially variable tandem repeats; hence, these chromosomes are at risk of being acentric. We reported previously that artificially created acentric chromosomes in the fission yeast Schizosaccharomyces pombe can be rescued by end-to-end fusion with functional chromosomes. Here, we show that most acentric/functional chromosome fusion events in S. pombe cells harbouring an acentric chromosome I differed from the non-homologous end-joining-mediated rearrangements that result in deleterious dicentric fusions in normal cells, and were elicited by a previously unidentified homologous recombination (HR) event between chromosome end-associated sequences. The subtelomere repeats associated with the non-fusogenic ends were also destabilized in the surviving cells, suggesting a causal link between general subtelomere destabilization and acentric/functional chromosome fusion. A mutational analysis indicated that a non-canonical HR pathway was involved in the rearrangement. These findings are indicative of a latent mechanism that conditionally induces general subtelomere instability, presumably in the face of accidental centromere loss events, resulting in rescue of the fatal acentric chromosomes by interchromosomal HR.

  13. Alteration of Terminal Heterochromatin and Chromosome Rearrangements in Derivatives of Wheat-Rye Hybrids

    Institute of Scientific and Technical Information of China (English)

    Shulan Fu; Zhenling Lv; Xiang Guo; Xiangqi Zhang; Fangpu Han

    2013-01-01

    Wheat-rye addition and substitution lines and their self progenies revealed variations in telomeric heterochromatin and centromeres.Furthermore,a mitotically unstable dicentric chromosome and stable multicentric chromosomes were observed in the progeny of a Chinese Spring-Imperial rye 3R addition line.An unstable multicentric chromosome was found in the progeny of a 6R/6D substitution line.Drastic variation of terminal heterochromatin including movement and disappearance of terminal heterochromatin occurred in the progeny of wheatrye addition line 3R,and the 5RS ditelosomic addition line.Highly stable minichromosomes were observed in the progeny of a monosomic 4R addition line,a ditelosomic 5RS addition line and a 6R/6D substitution line.Minichromosomes,with and without the FISH signals for telomeric DNA (TTTAGGG)n,derived from a monosomic 4R addition line are stable and transmissible to the next generation.The results indicated that centromeres and terminal heterochromatin can be profoundly altered in wheat-rye hybrid derivatives.

  14. Foundations of identifying individual chromosomes by imaging flow cytometry with applications in radiation biodosimetry.

    Science.gov (United States)

    Beaton-Green, Lindsay A; Rodrigues, Matthew A; Lachapelle, Sylvie; Wilkins, Ruth C

    2017-01-01

    Biodosimetry is an important tool for triage in the case of large-scale radiological or nuclear emergencies, but traditional microscope-based methods can be tedious and prone to scorer fatigue. While the dicentric chromosome assay (DCA) has been adapted for use in triage situations, it is still time-consuming to create and score slides. Recent adaptations of traditional biodosimetry assays to imaging flow cytometry (IFC) methods have dramatically increased throughput. Additionally, recent improvements in image analysis algorithms in the IFC software have resulted in improved specificity for spot counting of small events. In the IFC method for the dicentric chromosome analysis (FDCA), lymphocytes isolated from whole blood samples are cultured with PHA and Colcemid. After incubation, lymphocytes are treated with a hypotonic solution and chromosomes are isolated in suspension, labelled with a centromere marker and stained for DNA content with DRAQ5. Stained individual chromosomes are analyzed on the ImageStream®(X) (EMD-Millipore, Billerica, MA) and mono- and dicentric chromosome populations are identified and enumerated using advanced image processing techniques. Both the preparation of the isolated chromosome suspensions as well as the image analysis methods were fine-tuned in order to optimize the FDCA. In this paper we describe the method to identify and score centromeres in individual chromosomes by IFC and show that the FDCA method may further improve throughput for triage biodosimetry in the case of large-scale radiological or nuclear emergencies.

  15. Cytogenetic analysis of five species of the subfamily Corydoradinae (Teleostei: Siluriformes: Callichthyidae

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    Cristiane Kioko Shimabukuro-Dias

    2004-01-01

    Full Text Available In the present study, five callichthyid species belonging to the subfamily Corydoradinae were karyotyped: three species of Aspidoras and two of Corydoras. The three species of Aspidoras had the same diploid number, 2n = 46 chromosomes, similar karyotypic formulae, with most chromosomes metacentric or submetacentric, single interstitial Ag-NORs and C-band positive segments mainly found in the centromeric position. The comparative analysis of cytogenetic data available for the genus Aspidoras and other species of Corydoradinae suggest that several events of centric fusion occurred in the origin of the species of Aspidoras. The two analyzed species of Corydoras showed high diploid numbers, 2n = 74 in C. sodalis and 2n = 90 in C. britskii. While C. sodalis exhibited single Ag-NORs and terminal, interstitial and centromeric C-band positive segments in almost all chromosomes, C. britskii showed multiple Ag-NORs and a small number of C-band positive segments found in the terminal position in one acrocentric (A pair and in the interstitial position in one subtelocentric (ST pair. The occurrence of high diploid numbers and many ST and A chromosomes are uncommon among the Corydoradinae, suggesting the occurrence of a high number of chromosome rearrangements, mainly centric fissions, in the origin of the Corydoras species studied.

  16. A satellite explosion in the genome of holocentric nematodes.

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    Juan A Subirana

    Full Text Available Centromere sequences in the genome are associated with the formation of kinetochores, where spindle microtubules grow in mitosis. Centromere sequences usually have long tandem repeats (satellites. In holocentric nematodes it is not clear how kinetochores are formed during mitosis; they are distributed throughout the chromosomes. For this reason it appeared of interest to study the satellites in nematodes in order to determine if they offer any clue on how kinetochores are assembled in these species. We have studied the satellites in the genome of six nematode species. We found that the presence of satellites depends on whether the nematode chromosomes are holocentric or monocentric. It turns out that holocentric nematodes are unique because they have a large number of satellites scattered throughout their genome. Their number, length and composition are different in each species: they apparently have very little evolutionary conservation. In contrast, no scattered satellites are found in the monocentric nematode Trichinella spiralis. It appears that the absence/presence of scattered satellites in the genome distinguishes monocentric from holocentric nematodes. We conclude that the presence of satellites is related to the holocentric nature of the chromosomes of most nematodes. Satellites may stabilize a higher order structure of chromatin and facilitate the formation of kinetochores. We also present a new program, SATFIND, which is suited to find satellite sequences.

  17. Cytogenetic analysis of two locariid species (Teleostei, Siluriformes from Iguatemi River (Parana River drainage in Brazil

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    Carlos Alexandre Fernandes

    2015-03-01

    Full Text Available Fishes of the Loricariidae family, known as “cascudos”, constitute an endemic group in Neotropical freshwaters. In this study, were cytogenetically examined two species of Loricariidae (Pterygoplichthys anisitsi Eigenmann & Kennedy, 1903 and Farlowella amazonum (Günther, 1864 belonging to Hypostominae and Loricariinae subfamilies respectively from Iguatemi River. Our study provide the first description regarding C-band and fluorochromic analysis in F. amazonum. In Farlowella amazonum, diploid number was 58 chromosomes, with single Ag-NOR and heterochromatic blocks in centromeric regions of some chromosomes and large subtelomeric blocks were evidenced on the long arm of the pair 27, being this region CMA3+/DAPI-. The Pterygoplichthys anisitsi showed diploid number equal 52 chromosomes, with single Ag-NOR and heterochromatic blocks in centromeric and telomeric regions of some chromosomes and conspicuous large telomeric blocks on the long arm of the pair 10, being this region CMA3+/DAPI-. The results show that karyotype formula is nonconservative in P. anisitsi and F. amazonum.

  18. [Homologue pairing: initiation sites and effects on crossing over and chromosome disjunction in Drosophila melanogaster].

    Science.gov (United States)

    Chubykin, V L

    1996-01-01

    The role of homologue pairing and chromocentral association of chromosomes in recombination and segregation during cell division is discussed. Peculiarities of mitotic and meiotic chromosome pairing in Drosophila males and females are considered. On the basis of our own and published data, the presence and localization of sites of homologue pairing initiation in euchromatin are substantiated. The effects of transfer of initiation sites along a chromosome (exemplified by inversions) on chromosome pairing (asynapsis), crossing over (intrachromosomal, interchromosomal, and centromeric effects), and segregation are discussed. To record the effects of pairing sites on crossing over, a method of comparing crossing-over frequencies in an inverted region with those in a region of the same size and position with regard to the centromere on cytological maps was proposed. Chromosomes orient toward opposite division poles during paracentromeric heterochromatin pairing. This occurs after successful euchromatin pairing, during which the chromocentral circular structure is reorganized. If heterochromatin pairing is disrupted because of structural or locus mutations, nonexchange bivalents segregate randomly. In this case, chromosome coordination may occur due to proximal chiasmata or chromocentral associations between homologues.

  19. Mechanisms of plasmid segregation: have multicopy plasmids been overlooked?

    Science.gov (United States)

    Million-Weaver, Samuel; Camps, Manel

    2014-09-01

    Plasmids are self-replicating pieces of DNA typically bearing non-essential genes. Given that plasmids represent a metabolic burden to the host, mechanisms ensuring plasmid transmission to daughter cells are critical for their stable maintenance in the population. Here we review these mechanisms, focusing on two active partition strategies common to low-copy plasmids: par systems type I and type II. Both involve three components: an adaptor protein, a motor protein, and a centromere, which is a sequence area in the plasmid that is recognized by the adaptor protein. The centromere-bound adaptor nucleates polymerization of the motor, leading to filament formation, which can pull plasmids apart (par I) or push them towards opposite poles of the cell (par II). No such active partition mechanisms are known to occur in high copy number plasmids. In this case, vertical transmission is generally considered stochastic, due to the random distribution of plasmids in the cytoplasm. We discuss conceptual and experimental lines of evidence questioning the random distribution model and posit the existence of a mechanism for segregation in high copy number plasmids that moves plasmids to cell poles to facilitate transmission to daughter cells. This mechanism would involve chromosomally-encoded proteins and the plasmid origin of replication. Modulation of this proposed mechanism of segregation could provide new ways to enhance plasmid stability in the context of recombinant gene expression, which is limiting for large-scale protein production and for bioremediation.

  20. Partial tetrasomy 12pter-12p12.3 in a girl with Pallister-Killian syndrome: extraordinary finding of an analphoid, inverted duplicated marker.

    Science.gov (United States)

    Dufke, A; Walczak, C; Liehr, T; Starke, H; Trifonov, V; Rubtsov, N; Schöning, M; Enders, H; Eggermann, T

    2001-08-01

    Cytogenetic analysis in a girl with multiple congenital anomalies indicating Pallister-Killian syndrome (PKS) showed a supernumerary marker chromosome in 1/76 lymphocytes and 34/75 fibroblast metaphases. GTG-banding pattern was consistent with the chromosomal region 12pter-12q11. While fluorescence-in-situ hybridisation (FISH) with a whole chromosome 12 painting probe confirmed the origin of the marker, a chromosome 12 specific alpha-satellite probe did not hybridise to it. FISH analysis with a specific subtelomeric probe 12p showed hybridisation to both ends of the marker chromosome. High-resolution multicolour-banding (MCB) studies revealed the marker to be a der(12)(pter-->p12.3::p12.3-->pter). Summarising the FISH information, we defined the marker as an inverted duplication of 12pter-12p12.3 leading to partial tetrasomy of chromosome 12p. In skin fibroblasts, cultured at the patient's age of 1 year and 9 years, the marker chromosome was found in similar frequencies, even after several culture passages. Therefore, we consider the marker to have a functional centromere although it lacks detectable centromeric alpha-satellite sequences. To the best of our knowledge, this is the first proven analphoid marker of chromosome 12. Molecular genetic studies indicated that this marker is of paternal origin. The finding of partial tetrasomy 12pter-12p12.3 in our PKS patient allows to narrow down the critical region for PKS.