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Cell Aug 2022Human cleavage-stage embryos frequently acquire chromosomal aneuploidies during mitosis due to unknown mechanisms. Here, we show that S phase at the 1-cell stage shows...
Human cleavage-stage embryos frequently acquire chromosomal aneuploidies during mitosis due to unknown mechanisms. Here, we show that S phase at the 1-cell stage shows replication fork stalling, low fork speed, and DNA synthesis extending into G2 phase. DNA damage foci consistent with collapsed replication forks, DSBs, and incomplete replication form in G2 in an ATR- and MRE11-dependent manner, followed by spontaneous chromosome breakage and segmental aneuploidies. Entry into mitosis with incomplete replication results in chromosome breakage, whole and segmental chromosome errors, micronucleation, chromosome fragmentation, and poor embryo quality. Sites of spontaneous chromosome breakage are concordant with sites of DNA synthesis in G2 phase, locating to gene-poor regions with long neural genes, which are transcriptionally silent at this stage of development. Thus, DNA replication stress in mammalian preimplantation embryos predisposes gene-poor regions to fragility, and in particular in the human embryo, to the formation of aneuploidies, impairing developmental potential.
Topics: Aneuploidy; Animals; Chromosome Breakage; Chromosome Segregation; DNA; DNA Replication; Embryonic Development; Humans; Mammals
PubMed: 35858625
DOI: 10.1016/j.cell.2022.06.028 -
Genes Mar 2019Satellite DNAs are now regarded as powerful and active contributors to genomic and chromosomal evolution. Paired with mobile transposable elements, these repetitive... (Review)
Review
Satellite DNAs are now regarded as powerful and active contributors to genomic and chromosomal evolution. Paired with mobile transposable elements, these repetitive sequences provide a dynamic mechanism through which novel karyotypic modifications and chromosomal rearrangements may occur. In this review, we discuss the regulatory activity of satellite DNA and their neighboring transposable elements in a chromosomal context with a particular emphasis on the integral role of both in centromere function. In addition, we discuss the varied mechanisms by which centromeric repeats have endured evolutionary processes, producing a novel, species-specific centromeric landscape despite sharing a ubiquitously conserved function. Finally, we highlight the role these repetitive elements play in the establishment and functionality of de novo centromeres and chromosomal breakpoints that underpin karyotypic variation. By emphasizing these unique activities of satellite DNAs and transposable elements, we hope to disparage the conventional exemplification of repetitive DNA in the historically-associated context of 'junk'.
Topics: Centromere; Chromosome Breakage; Chromosomes, Human; DNA Transposable Elements; DNA, Satellite; Evolution, Molecular; Humans; Species Specificity
PubMed: 30884847
DOI: 10.3390/genes10030223 -
Chromosoma Dec 2022The maintenance of genome integrity is ensured by proper chromosome inheritance during mitotic and meiotic cell divisions. The chromosomal counterpart responsible for...
The maintenance of genome integrity is ensured by proper chromosome inheritance during mitotic and meiotic cell divisions. The chromosomal counterpart responsible for chromosome segregation to daughter cells is the centromere, at which the spindle apparatus attaches through the kinetochore. Although all mammalian centromeres are primarily composed of megabase-long repetitive sequences, satellite-free human neocentromeres have been described. Neocentromeres and evolutionary new centromeres have revolutionized traditional knowledge about centromeres. Over the past 20 years, insights have been gained into their organization, but in spite of these advancements, the mechanisms underlying their formation and evolution are still unclear. Today, through modern and increasingly accessible genome editing and long-read sequencing techniques, research in this area is undergoing a sudden acceleration. In this article, we describe the primary sequence of a previously described human chromosome 3 neocentromere and observe its possible evolution and repair results after a chromosome breakage induced through CRISPR-Cas9 technologies. Our data represent an exciting advancement in the field of centromere/neocentromere evolution and chromosome stability.
Topics: Humans; Animals; CRISPR-Cas Systems; Centromere; Kinetochores; Chromosome Segregation; Chromosome Breakage; Mammals
PubMed: 35978051
DOI: 10.1007/s00412-022-00779-y -
Developmental Cell Nov 2012During oncogenesis, cells acquire multiple genetic alterations that confer essential tumor-specific traits, including immortalization, escape from antimitogenic... (Review)
Review
During oncogenesis, cells acquire multiple genetic alterations that confer essential tumor-specific traits, including immortalization, escape from antimitogenic signaling, neovascularization, invasiveness, and metastatic potential. In most instances, these alterations are thought to arise incrementally over years, if not decades. However, recent progress in sequencing cancer genomes has begun to challenge this paradigm, because a radically different phenomenon, termed chromothripsis, has been suggested to cause complex intra- and interchromosomal rearrangements on short timescales. In this Review, we review established pathways crucial for genome integrity and discuss how their dysfunction could precipitate widespread chromosome breakage and rearrangement in the course of malignancy.
Topics: Animals; Apoptosis; Chromosomal Instability; Chromosome Aberrations; Chromosome Breakage; DNA Repair; DNA Replication; Gene Rearrangement; Humans; Mitosis; Models, Genetic; Mutation; Neoplasms; Telomere Shortening
PubMed: 23153487
DOI: 10.1016/j.devcel.2012.10.010 -
Genetics May 2023Chromosome breakage plays an important role in the evolution of karyotypes and can produce deleterious effects within a single individual, such as aneuploidy or cancer....
Chromosome breakage plays an important role in the evolution of karyotypes and can produce deleterious effects within a single individual, such as aneuploidy or cancer. Forces that influence how and where chromosomes break are not fully understood. In humans, breakage tends to occur in conserved hotspots called common fragile sites (CFS), especially during replication stress. By following the fate of dicentric chromosomes in Drosophila melanogaster, we find that breakage under tension also tends to occur in specific hotspots. Our experimental approach was to induce sister chromatid exchange in a ring chromosome to generate a dicentric chromosome with a double chromatid bridge. In the following cell division, the dicentric bridges may break. We analyzed the breakage patterns of 3 different ring-X chromosomes. These chromosomes differ by the amount and quality of heterochromatin they carry as well as their genealogical history. For all 3 chromosomes, breakage occurs preferentially in several hotspots. Surprisingly, we found that the hotspot locations are not conserved between the 3 chromosomes: each displays a unique array of breakage hotspots. The lack of hotspot conservation, along with a lack of response to aphidicolin, suggests that these breakage sites are not entirely analogous to CFS and may reveal new mechanisms of chromosome fragility. Additionally, the frequency of dicentric breakage and the durability of each chromosome's spindle attachment vary significantly between the 3 chromosomes and are correlated with the origin of the centromere and the amount of pericentric heterochromatin. We suggest that different centromere strengths could account for this.
Topics: Animals; Humans; Chromosome Breakage; Drosophila melanogaster; Heterochromatin; Centromere; X Chromosome
PubMed: 37010100
DOI: 10.1093/genetics/iyad052 -
Molecular Microbiology Jun 2006DNA double-strand breaks (DSBs) are among the most deleterious types of damage that can occur in the genome of eukaryotic cells because failure to repair them can lead... (Review)
Review
DNA double-strand breaks (DSBs) are among the most deleterious types of damage that can occur in the genome of eukaryotic cells because failure to repair them can lead to loss of genetic information and chromosome rearrangements. DSBs can arise by failures in DNA replication and by exposure to environmental factors, such as ionizing radiations and radiomimetic chemicals. Moreover, they might arise when telomeres undergo extensive erosion, leading to the activation of the DNA damage response pathways and the onset of apoptosis and/or senescence. Importantly, DSBs can also form in a programmed manner during development. For example, meiotic recombination and rearrangement of the immunoglobulin genes in lymphocytes require the generation of site- or region-specific DSBs through the action of specific endonucleases. Efficient DSB repair is crucial in safeguarding genome integrity, whose maintenance in the face of DSBs involves branched signalling networks that switch on DNA damage checkpoints, activate DNA repair, induce chromatin reorganization and modulate numerous cellular processes. Not surprisingly, defects in these networks result in a variety of diseases ranging from severe genetic disorders to cancer predisposition and accelerated ageing.
Topics: Animals; Cell Cycle; Chromosome Breakage; DNA Breaks, Double-Stranded; DNA Repair; Histones; Humans; Models, Genetic; Recombination, Genetic; Signal Transduction
PubMed: 16689788
DOI: 10.1111/j.1365-2958.2006.05186.x -
The EMBO Journal Sep 2023DNA single-strand breaks (SSBs) disrupt DNA replication and induce chromosome breakage. However, whether SSBs induce chromosome breakage when present behind replication...
DNA single-strand breaks (SSBs) disrupt DNA replication and induce chromosome breakage. However, whether SSBs induce chromosome breakage when present behind replication forks or ahead of replication forks is unclear. To address this question, we exploited an exquisite sensitivity of SSB repair-defective human cells lacking PARP activity or XRCC1 to the thymidine analogue 5-chloro-2'-deoxyuridine (CldU). We show that incubation with CldU in these cells results in chromosome breakage, sister chromatid exchange, and cytotoxicity by a mechanism that depends on the S phase activity of uracil DNA glycosylase (UNG). Importantly, we show that CldU incorporation in one cell cycle is cytotoxic only during the following cell cycle, when it is present in template DNA. In agreement with this, while UNG induces SSBs both in nascent strands behind replication forks and in template strands ahead of replication forks, only the latter trigger fork collapse and chromosome breakage. Finally, we show that BRCA-defective cells are hypersensitive to CldU, either alone and/or in combination with PARP inhibitor, suggesting that CldU may have clinical utility.
Topics: Humans; Poly(ADP-ribose) Polymerase Inhibitors; DNA-Binding Proteins; Chromosome Breakage; DNA Repair; DNA Replication; DNA; Antineoplastic Agents; X-ray Repair Cross Complementing Protein 1
PubMed: 37492888
DOI: 10.15252/embj.2022113190 -
The Journal of Cell Biology Nov 2023In response to chromatin bridges, the abscission checkpoint delays completion of cytokinesis to prevent chromosome breakage or tetraploidization. Here, we show that...
In response to chromatin bridges, the abscission checkpoint delays completion of cytokinesis to prevent chromosome breakage or tetraploidization. Here, we show that spontaneous or replication stress-induced chromatin bridges exhibit "knots" of catenated and overtwisted DNA next to the midbody. Topoisomerase IIα (Top2α) forms abortive Top2-DNA cleavage complexes (Top2ccs) on DNA knots; furthermore, impaired Top2α-DNA cleavage activity correlates with chromatin bridge breakage in cytokinesis. Proteasomal degradation of Top2ccs is required for Rad17 localization to Top2-generated double-strand DNA ends on DNA knots; in turn, Rad17 promotes local recruitment of the MRN complex and downstream ATM-Chk2-INCENP signaling to delay abscission and prevent chromatin breakage. In contrast, dicentric chromosomes that do not exhibit knotted DNA fail to activate the abscission checkpoint in human cells. These findings are the first to describe a mechanism by which the abscission checkpoint detects chromatin bridges, through generation of abortive Top2ccs on DNA knots, to preserve genome integrity.
Topics: Humans; Cell Cycle Proteins; Cell Nucleus; Chromatin; Chromosome Breakage; Cytokinesis; DNA; Cell Cycle Checkpoints; DNA Topoisomerases, Type II
PubMed: 37638884
DOI: 10.1083/jcb.202303123 -
Nature Reviews. Cancer Jul 2017Ever since initial suggestions that instability at common fragile sites (CFSs) could be responsible for chromosome rearrangements in cancers, CFSs and associated genes... (Review)
Review
Ever since initial suggestions that instability at common fragile sites (CFSs) could be responsible for chromosome rearrangements in cancers, CFSs and associated genes have been the subject of numerous studies, leading to questions and controversies about their role and importance in cancer. It is now clear that CFSs are not frequently involved in translocations or other cancer-associated recurrent gross chromosome rearrangements. However, recent studies have provided new insights into the mechanisms of CFS instability, their effect on genome instability, and their role in generating focal copy number alterations that affect the genomic landscape of many cancers.
Topics: Anaphase; Animals; Chromosomal Instability; Chromosome Breakage; Chromosome Fragile Sites; DNA Breaks, Double-Stranded; DNA Copy Number Variations; DNA Replication; Gene Rearrangement; Humans; Metaphase; Neoplasms; Oncogenes
PubMed: 28740117
DOI: 10.1038/nrc.2017.52 -
Journal of Biomedical Science Nov 2021RAD51-dependent homologous recombination (HR) is one of the most important pathways for repairing DNA double-strand breaks (DSBs), and its regulation is crucial to...
BACKGROUND
RAD51-dependent homologous recombination (HR) is one of the most important pathways for repairing DNA double-strand breaks (DSBs), and its regulation is crucial to maintain genome integrity. Elp1 gene encodes IKAP/ELP1, a core subunit of the Elongator complex, which has been implicated in translational regulation. However, how ELP1 contributes to genome maintenance is unclear.
METHODS
To investigate the function of Elp1, Elp1-deficient mouse embryonic fibroblasts (MEFs) were generated. Metaphase chromosome spreading, immunofluorescence, and comet assays were used to access chromosome abnormalities and DSB formation. Functional roles of Elp1 in MEFs were evaluated by cell viability, colony forming capacity, and apoptosis assays. HR-dependent DNA repair was assessed by reporter assay, immunofluorescence, and western blot. Polysome profiling was used to evaluate translational efficiency. Differentially expressed proteins and signaling pathways were identified using a label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomics approach.
RESULTS
Here, we report that Elp1 depletion enhanced genomic instability, manifested as chromosome breakage and genotoxic stress-induced genomic DNA fragmentation upon ionizing radiation (IR) exposure. Elp1-deficient cells were hypersensitive to DNA damage and exhibited impaired cell proliferation and defective HR repair. Moreover, Elp1 depletion reduced the formation of IR-induced RAD51 foci and decreased RAD51 protein levels. Polysome profiling analysis revealed that ELP1 regulated RAD51 expression by promoting its translation in response to DNA damage. Notably, the requirement for ELP1 in DSB repair could be partially rescued in Elp1-deficient cells by reintroducing RAD51, suggesting that Elp1-mediated HR-directed repair of DSBs is RAD51-dependent. Finally, using proteome analyses, we identified several proteins involved in cancer pathways and DNA damage responses as being differentially expressed upon Elp1 depletion.
CONCLUSIONS
Our study uncovered a molecular mechanism underlying Elp1-mediated regulation of HR activity and provides a novel link between translational regulation and genome stability.
Topics: Animals; Chromosome Breakage; DNA Damage; Fibroblasts; Genomic Instability; Intracellular Signaling Peptides and Proteins; Mice; Protein Biosynthesis; Rad51 Recombinase; Recombinational DNA Repair
PubMed: 34819065
DOI: 10.1186/s12929-021-00773-z