Shortening Of Chromosomes Research Articles

Spontaneous deletions and reciprocal translocations in Saccharomyces cerevisiae: influence of ploidy

Studying spontaneous chromosomal rearrangements throws light on the rules underlying the genome reshaping events occurring in eukaryotic cells, which are part of the evolutionary process. In Saccharomyces cerevisiae, translocation and deletion processes have been frequently described in haploids, but little is known so far about these processes at the diploid level. Here we investigated the nature and the frequency of the chromosomal rearrangements occurring at this ploidy level. Using a positive selection screen based on a particular mutated allele of the URA2 gene, spontaneous diploid revertants were selected and analysed. Surprisingly, the diploid state was found to be correlated with a decrease in chromosome rearrangement frequency, along with an increase in the complexity of the rearrangements occurring in the target gene. The presence of short DNA tandem repeat sequences seems to be a key requirement for deletion and reciprocal translocation processes to occur in diploids. After discussing the differences between the haploid and diploid levels, some mechanisms possibly involved in chromosome shortening and arm exchange are suggested.

A different approach to telomere analysis with ddPRINS in chronic lymphocytic leukemia

Telomeric sequences, located at the very end of the chromosomes, compensate for the chromosomal shortening as it happens after each round of cell division. Telomeric sequences influence the progress of cellular senescence and cancer progression. It has been reported that telomeres are shortened in acute leukemias where the cell turnover is high. B-cell chronic lymphocytic leukemia (CLL) is a particularly interesting haematological malignancy in regard to telomere dynamics because most of the malignant cells in CLL are mitotically inactive. In this study, we analysed the telomere length in patients with B-cell CLL in a comparison with the control group by using ddPRINS technique. Twenty patients with CLL and four healthy donors as a control group were included. We found short telomeres and no detectable telomeric repeats at the sites of chromosome fusion. We hypothesise that the telomeric erosion in CLL may reflect the dominance of malignant cells with an abnormally long life span. These cells may have encountered many antigenic stimulants in the past and hence underwent multiple clonal expansions. Our findings imply that shortened telomeres in CLL may be reflecting the “history” of the disease and serve as an independent prognostic factor.

Crossing over is coupled to late meiotic prophase bivalent differentiation through asymmetric disassembly of the SC

Homologous chromosome pairs (bivalents) undergo restructuring during meiotic prophase to convert a configuration that promotes crossover recombination into one that promotes bipolar spindle attachment and localized cohesion loss. We have imaged remodeling of meiotic chromosome structures after pachytene exit in Caenorhabditis elegans. Chromosome shortening during diplonema is accompanied by coiling of chromosome axes and highly asymmetric departure of synaptonemal complex (SC) central region proteins SYP-1 and SYP-2, which diminish over most of the length of each desynapsing bivalent while becoming concentrated on axis segments distal to the single emerging chiasma. This and other manifestations of asymmetry along chromosomes are lost in synapsis-proficient crossover-defective mutants, which often retain SYP-1,2 along the full lengths of coiled diplotene axes. Moreover, a γ-irradiation treatment that restores crossovers in the spo-11 mutant also restores asymmetry of SYP-1 localization. We propose that crossovers or crossover precursors serve as symmetry-breaking events that promote differentiation of subregions of the bivalent by triggering asymmetric disassembly of the SC.

Distinct functions of condensin I and II in mitotic chromosome assembly

Condensin is a protein complex associated with mitotic chromosomes that has been implicated in chromosome condensation. In vertebrates, two types of condensin complexes have recently been identified, called condensin I and II. Here, we show that in mammalian cells condensin II associates with chromatin in prophase, in contrast to condensin I which is cytoplasmic and can thus interact with chromosomes only after nuclear envelope breakdown. RNA interference experiments in conjunction with imaging of live and fixed cells revealed that condensin II is required for chromosome condensation in early prophase, whereas condensin I appears to be dispensable at this stage. By contrast, condensin I is required for the complete dissociation of cohesin from chromosome arms, for chromosome shortening and for normal timing of progression through prometaphase and metaphase, whereas normal condensin II levels are dispensable for these processes. After depletion of both condensin complexes, the onset of chromosome condensation is delayed until the end of prophase, but is then initiated rapidly before nuclear envelope breakdown. These results reveal that condensin II and I associate with chromosomes sequentially and have distinct functions in mitotic chromosome assembly.

A different approach to telomere analysis with ddPRINS in chronic lymphocytic leukemia

Telomeric sequences, located at the very end of the chromosomes, compensate for the chromosomal shortening as it happens after each round of cell division. Telomeric sequences influence the progress of cellular senescence and cancer progression. It has been reported that telomeres are shortened in acute leukemias where the cell turnover is high. B-cell chronic lymphocytic leukemia (CLL) is a particularly interesting haematological malignancy in regard to telomere dynamics because most of the malignant cells in CLL are mitotically inactive. In this study, we analysed the telomere length in patients with B-cell CLL in a comparison with the control group by using ddPRINS technique. Twenty patients with CLL and four healthy donors as a control group were included. We found short telomeres and no detectable telomeric repeats at the sites of chromosome fusion. We hypothesise that the telomeric erosion in CLL may reflect the dominance of malignant cells with an abnormally long life span. These cells may have encountered many antigenic stimulants in the past and hence underwent multiple clonal expansions. Our findings imply that shortened telomeres in CLL may be reflecting the history of the disease and serve as an independent prognostic factor.

Identification of telomerase activity in gametocytes of Plasmodium falciparum

Telomerase, a specialized cellular reverse transcriptase, compensates the chromosome shortening during the replication of most eukaryotic cells and contributes to cellular immortalization in cell culture (in vitro) and cancerous cell (in vivo). In the present study, the telomerase activity in the gametocytes of Plasmodium falciparum was investigated. Here, we report for the first time, the presence of telomerase activity in the gametocytes of P. falciparum using P. falciparum telomere repeat amplification protocol (Pf-TRAP) assay and Southern blot hybridization. Telomerase inhibitors such as 7-deaza-dGTP and AZT-TP, when used with the cytoplasmic extract of gametocytes in the Pf-TRAP assay, efficiently inhibit the product, which confirms the presence of telomerase in the gametocytes. The presence of telomerase activity in the laboratory adapted local (eastern India) isolates of P. falciparum indicates that telomerase might be the major player in chromosomal end protection during replication. The finding suggests that telomerase can be a potent target for the transmission blocking vaccine and drugs for combating malaria caused by P. falciparum.

Telomerase activity in the subventricular zone of adult mice

The subventricular zone (SVZ) is the most active site for the production of new neurons in the adult mouse brain. Neural stem cells in the adult SVZ give rise to neuroblasts that travel via the rostral migratory stream (RMS) to the olfactory bulb, where they differentiate into interneurons. The enzyme telomerase has been identified in other population of stem cells and is necessary for the synthesis of telomeric DNA to prevent chromosomal shortening, end-to-end fusions, and apoptosis during successive rounds of cell division. However, previous studies have failed to detect telomerase in the adult mammalian brain. Here we demonstrate that telomerase is expressed by all brain regions shortly after birth, but becomes restricted to the SVZ and olfactory bulb in the adult mouse brain. Cultures of neural precursor cells or of migratory neuroblasts purified from the SVZ were each found to possess telomerase activity. After elimination of migrating neuroblasts and immature precursor cells in vivo by treatment with cytosine-β- d-arabinofuranoside (Ara-C), telomerase activity was still detectable in the remaining SVZ, which includes a population of neural stem cells. Following withdrawal of Ara-C, telomerase activity subsequently increased with a time course that parallels regeneration of the SVZ network and RMS. Finally, intracranial surgery alone, which has previously been shown to increase the number of cells in the SVZ, produced higher telomerase levels in the SVZ. We conclude that telomerase is active in neural precursor cells of the adult mouse and suggest that its regulation is an important parameter for cellular proliferation to occur in the mammalian brain.

E2F-1 represses transcription of the human telomerase reverse transcriptase gene.

The ends of human chromosomes (telomeres) lose up to 200 bp of DNA per cell division. Chromosomal shortening ultimately leads to senescence and death in normal cells. Many human carcinoma lines are immortal in vitro, suggesting that these cells have a mechanism for maintaining the ends of their chromosomes. Telomerase is a ribonucleoprotein complex that synthesizes telomeric DNA onto chromosomes using its RNA component as template. Telomerase activity is found in most tumor cells, but is absent from normal cells. Little is known about how normal human cells repress telomerase (hTERT) gene expression. Mice carrying an E2F-1 null mutation develop a variety of malignant tumors, suggesting that this transcription factor has a tumor suppressor function. To determine mechanisms by which E2F-1 suppresses tumor formation, we examined the role of this transcription factor in regulation of the hTERT promoter in human cells. We identified two putative E2F-1-binding sites proximal to the transcriptional start site of the hTERT promoter. Mutation of these sites produced dramatic increases in promoter activity. Overexpression of E2F-1 but not a mutant E2F-1 repressed hTERT promoter activity in reporter gene assays. This repression was abolished by mutation of the E2F-1-binding sites in the hTERT promoter. Human cancer cell lines stably overexpressing E2F-1 exhibited decreased hTERT mRNA expression and telomerase activity. We conclude that E2F-1 has an atypical function as a transcriptional repressor of the hTERT gene in human cells.

Rb and E2F-1 regulate telomerase activity in human cancer cells

The ends of human chromosomes (telomeres) lose up to 200 bp of DNA per cell division. Chromosomal shortening ultimately leads to senescence and death in normal cells. Many human carcinoma lines are immortal in vitro, suggesting that these cells have a mechanism for maintaining the ends of their chromosomes. Telomerase is a ribonucleoprotein complex that synthesizes telomeric DNA onto chromosomes using its RNA component as a template. Recent studies have shown that inactivation of the retinoblastoma gene product pRb and the cyclin dependent kinase inhibitor p16 INK4A is required for telomerase activity in epithelial cells. We have demonstrated previously that restoration of functional retinoblastoma (Rb) expression is sufficient to downregulate telomerase activity in carcinoma cells. To determine mechanisms by which Rb regulates telomerase expression, we examined the effects of cyclin dependent kinase (cdk) mediated Rb inactivation and the release of E2F-1 on telomerase activity in human carcinoma cells. Overexpression of cdk2 and cdk4 but not a dominant negative cdk2 rescued Rb mediated downregulation of telomerase activity. Overexpression of the cdk regulatory subunit cyclin D1 also rescued telomerase downregulation and p16 expression alone was sufficient to ablate activity. E2F-1 overexpression was sufficient to rescue Rb mediated reduction of telomerase activity, but an E2F-1 mutant defective in DNA and Rb binding activities failed to produce this effect. Tumor tissue from E2F-1 −/− mice was negative for telomerase activity, indicating a key regulatory role for this transcription factor.

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

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

Intact functional domains of the retinoblastoma gene product (pRb) are required for downregulation of telomerase activity

The ends of human chromosomes (telomeres) consist of tandem repeats of the sequence TTAGGG. Telomeres lose up to 200 base pairs of DNA per cell division due to the inability of DNA polymerase to completely replicate the chromosomal ends. Chromosomal shortening ultimately leads to senescence and cell death in normal cells. However, some immortal cells do not lose telomeric sequence during DNA replication. Many human carcinoma lines are immortal in vitro, suggesting that these cells have a mechanism for maintaining the ends of their chromosomes. Telomerase is a ribonucleoprotein complex that synthesizes telomeric DNA onto chromosomes using its RNA component as a template. To elucidate potential mechanisms for telomerase regulation, we tested human squamous cell carcinoma lines (SCCs) for telomerase activity. All SCC lines expressed high levels of telomerase activity. Synchronization in specific cell cycle phases caused marked reduction in telomerase activity in G 0 and S, but not in G 1 or M. Reduction in telomerase activity correlated with induction of Rb protein in these phases. Overexpression of full length Rb resulted in significant downregulation of telomerase activity. However, expression of an Rb N-terminal oligomerization domain deletion construct, a C-terminal DNA binding domain deletion construct, or a pocket domain mutant failed to downregulate telomerase activity. We concluded that functionally intact Rb was required for cell cycle-dependent downregulation of telomerase activity in SCC lines.

Plasmodium falciparum telomerase: de novo telomere addition to telomeric and nontelomeric sequences and role in chromosome healing.

Telomerase, a specialized cellular reverse transcriptase, compensates for chromosome shortening during the proliferation of most eucaryotic cells and contributes to cellular immortalization. The mechanism used by the single-celled protozoan malaria parasite Plasmodium falciparum to complete the replication of its linear chromosomes is currently unknown. In this study, telomerase activity has for the first time been identified in cell extracts of P. falciparum. The de novo synthesis of highly variable telomere repeats to the 3' end of DNA oligonucleotide primers by plasmodial telomerase is demonstrated. Permutated telomeric DNA primers are extended by the addition of the next correct base. In addition to elongating preexisting telomere sequences, P. falciparum telomerase can also add telomere repeats onto nontelomeric 3' ends. The sequence GGGTT was the predominant initial DNA sequence added to the nontelomeric 3' ends in vitro. Poly(C) at the 3' end of the oligonucleotide significantly alters the precision of the new telomerase added repeats. The efficiency of nontelomeric primer elongation was dependent on the presence of a G-rich cassette upstream of the 3' terminus. Oligonucleotide primers based on natural P. falciparum chromosome breakpoints are efficiently used as telomerase substrates. These results imply that P. falciparum telomerase contributes to chromosome maintenance and to de novo telomere formation on broken chromosomes. Reverse transcriptase inhibitors such as dideoxy GTP efficiently inhibit P. falciparum telomerase activity in vitro. These data point to malaria telomerase as a new target for the development of drugs that could induce parasite cell senescence.

Long G Tails at Both Ends of Human Chromosomes Suggest a C Strand Degradation Mechanism for Telomere Shortening

The chromosomes of lower eukaryotes have short telomeric 3′ extensions. Using a primer-extension/nick-translation technique and nondenaturing hybridization, we find long 3′ G-rich tails at human chromosome ends in mortal primary fibroblasts, umbilical vein endothelial cells, and leukocytes, as well as in immortalized fibroblasts. For all cells tested, >80% of the telomeres have long G-rich overhangs, averaging 130–210 bases in length, in disagreement with the conventional model for incomplete lagging-strand replication, which predicts overhangs on 50% of the chromosome ends. The observed G tails must exist during most of the cell cycle and probably result from degradation of both chromosome ends. The average lengths of the G tails are quantitatively consistent with the observed rates of human chromosome shortening.

Coumarin and its 4- and 7- Substituted Derivatives as Retardants of Mitoses in Allium Root Promeristem

Coumarin and five derivatives (4-hydroxycoumarin, 7-hydroxycoumarin, 7-methylcoumarin, 7-methoxycoumarin, and 7-methoxy-4-methylcoumarin) were investigated for their potential as antimitotic agents. Coumarin and 4-hydroxycoumarin completely inhibited mitosis yet did not induce chromosomal aberrations in Allium sativum promeristem after 24 h treatment, and when postincubated in water did not exhibit mitotic recovery. 7-hydroxycoumarin also strongly inhibited cell proliferation (77% mitotic inhibition after 24 h). Sharp declines in mitotic activity of A. sativum promeristem were noted for 7-methylcoumarin, 7-methoxycoumarin, and 7-methoxy-4-methylcoumarin (83%, 90%, and 64% mitotic inhibition, respectively) in 24 h incubations. Chromosomal aberrations associated with 7-hydroxycoumarin treatment included shortening and thickening of chromosomes, postponed chromatid separation, anaphase and telophase bridges, and irregularly shaped nuclei. Disruption of normal chromosomal structure was also observed after exp...

Effect of Coumarin and its Derivatives on Mitosis and Ultrastructure of Meristematic Cells

AbstractUsing root tips of Allium cepa as a model system, coumarin, 7-hydroxycoumarin, 4-hydroxycoumarin, psoralen, and xanthotoxin inhibited mitosis. 7-Hydroxycoumarin caused the most marked changes in karyokinesis, which led to chromatin condensation and shortening of chromosomes, as well as disorganisation of microtubules. Other coumarins influenced mostly cytokinesis, causing aberrations in fragmoplast formation and displacement of the middle lamella. The cell wall between sister cells was aberrant, with telophase bridges often occurring late in the post-telophase stage. In the cytoplasm, changes were noted in the mitochondrial structure, ER arrangement and separation of parts of the cytoplasm, in which autolysis was observed. After treatment with 7-hydroxycoumarin the ER became hypertrophic, with cisternae of the rough ER still narrow and arranged in complexes of parallel sheets. After treatment with the psoralens the ER dilated and fragmented into small vesicles. Swollen and dividing megamitochondri...

Coumarins as Antimitotics

Coumarin, Two Of Its Hydroxy Derivatives (Umbelliferone and 4-Hydroxycoumarin), and Two Furano-Coumarins (Psoralen and Xanthotoxin) Were Investigated For Their Potential Use As Mitodepressants. In Sublethal Concentrations These Compounds Stopped Mitosis (Umbelliferone Inhibited 95%) In 24 H Incubations. All Phases Of Mitosis Were Affected In The Same Proportions, Except Umbelliferone, Which Prolonged Prophase and Blocked Metaphase. In Postincubation, The Meristematic Cells Kept In Coumarin Solution Did Not Recover. Postincubation Of Umbelliferone and 4-Hydroxycoumarin Resulted In Mitoses Reaching 40% and 120% Of The Control, Respectively. Furanocoumarins Showed A Few Mitoses In 24 H Postincubation, But After 72 H No Mitoses Occurred. Chromosomal Aberrations Were Not Observed After Coumarin and 4-Hydroxycoumarin Treatment. Umbelliferone Caused Shortening and Thickening Of Chromosomes, Caused Postponement Of Chromatid Separation, Occurrence Of Sticky Ends Leading To Anaphase, and Telophase Bridges. Similar Types Of Aberration, But More Frequent, Were Observed For Furanocoumarins, Which Additionally Showed Severe Aberrations Of Fragmoplast Formation Leading To The Occurrence Of Binucleate Cells, Micronuclei, Some Tetraploid Nuclei With Irregular Surfaces, and Fragmoplasts Located In Distant Areas Of The Cell.

Chromosome length and DNA loop size during early embryonic development of Xenopus laevis

The looped organization of the eukaryotic genome mediated by a skeletal framework of non-histone proteins is conserved throughout the cell cycle. The radial loop/scaffold model envisages that the higher order architecture of metaphase chromosomes relies on an axial structure around which looped DNA domains are radially arranged through stable attachment sites. In this light we investigated the relationship between the looped organization and overall morphology of chromosomes. In developing Xenopus laevis embryos at gastrulation, the bulk of the loops associated with histone-depleted nuclei exhibit a significant size increase, as visualized by fluorescence microscopy of the fully extended DNA halo surrounding high salt treated, ethidium bromide stained nuclei. This implies a reduction in the number of looped domains anchored to the supporting nucleoskeletal structure. The cytological analysis of metaphase plates from acetic acid fixed whole embryos, carried out in the absence of drugs inducing chromosome condensation, reveals a progressive thickening and shortening of metaphase chromosomes during development. We interpret these findings as a strong indication that the size and number of DNA loops influence the thickness and length of the chromosomes, respectively. The quantitative analysis of chromosome length distributions at different developmental stages suggests that the shortening is timed differently in different embryonic cells.

Possible involvement of nuclei in cadmium-induced modifications of cultured cells.

Reduced metaphase number and shortening of metaphase chromosomes were detected in McCoy cells exposed to 100 microM CdSO4 (maximal exposure time: 7 h). One hour exposure to 109Cd was enough to label the cell nucleus. This possibly suggests an early nuclear involvement in Cd-induced cell damage.

Changes of Chromosome Length and Constitutive Heterochromatin in Association with Cell Division during Early Development of Cynops pyrrhogaster embryo: (chromatin condensation/constitutive heterochromatin/cell division/ neural differentiation/amphibian embryo).

The length of chromosomes in the presumptive ectoderms of Cynops embryos was measured at nine successive cell divisions from the 6th (cleavage stage) to the 14th (gastrula stage). Up to the 10th cell division (cleavage stage) the chromosome length remained constant. At the 11th cell division the chromosomes began to shorten and continued to shorten at every cell division up to the 14th cell division. The degree of shortening and the mode of variation in length corresponded to the respective developmental stages of cleavage, blastula and gastrula. During those periods when chromosomes became shortened, some fine C-bands of the paracentromeric region found in earlier stages fused together. The chromatin of interphase nuclei also showed considerable changes during chromosome shortening. Besides the size reduction of interphase nuclei, the number of C-band granules in an interphase nucleus decreased in parallel with chromosome shortening and fusion of C-bands in mitotic chromosome.

Extreme chromosome shortening resembling fragmentation induced by 2,3-dimethyl p-benzoquinone in human lymphocytes.

Environmental MutagenesisVolume 5, Issue 6 p. 923-927 Brief Communication Extreme chromosome shortening resembling fragmentation induced by 2,3-dimethyl p-benzoquinone in human lymphocytes Máximo E. Drets, Corresponding Author Máximo E. Drets Department of Medical Genetics, University of Wisconsin, MadisonDivision of Human Cytogenetics and Quantitative Microscopy, Instituto de Investigaciones Biológicas Clemente Estable, Avda. Italia 3318, Montevideo, UruguaySearch for more papers by this author Máximo E. Drets, Corresponding Author Máximo E. Drets Department of Medical Genetics, University of Wisconsin, MadisonDivision of Human Cytogenetics and Quantitative Microscopy, Instituto de Investigaciones Biológicas Clemente Estable, Avda. Italia 3318, Montevideo, UruguaySearch for more papers by this author First published: 1983 https://doi.org/10.1002/em.2860050615Citations: 3AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume5, Issue61983Pages 923-927 RelatedInformation