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Nature Mar 2017Human cells have twenty-three pairs of chromosomes. In cancer, however, genes can be amplified in chromosomes or in circular extrachromosomal DNA (ecDNA), although the...
Human cells have twenty-three pairs of chromosomes. In cancer, however, genes can be amplified in chromosomes or in circular extrachromosomal DNA (ecDNA), although the frequency and functional importance of ecDNA are not understood. We performed whole-genome sequencing, structural modelling and cytogenetic analyses of 17 different cancer types, including analysis of the structure and function of chromosomes during metaphase of 2,572 dividing cells, and developed a software package called ECdetect to conduct unbiased, integrated ecDNA detection and analysis. Here we show that ecDNA was found in nearly half of human cancers; its frequency varied by tumour type, but it was almost never found in normal cells. Driver oncogenes were amplified most commonly in ecDNA, thereby increasing transcript level. Mathematical modelling predicted that ecDNA amplification would increase oncogene copy number and intratumoural heterogeneity more effectively than chromosomal amplification. We validated these predictions by quantitative analyses of cancer samples. The results presented here suggest that ecDNA contributes to accelerated evolution in cancer.
Topics: Chromosomes, Human; Cytogenetic Analysis; DNA Copy Number Variations; DNA Mutational Analysis; Evolution, Molecular; Gene Amplification; Genetic Heterogeneity; Genome, Human; Humans; Metaphase; Models, Genetic; Neoplasms; Oncogenes; RNA, Messenger; RNA, Neoplasm; Reproducibility of Results; Software
PubMed: 28178237
DOI: 10.1038/nature21356 -
Reproductive Biology and Endocrinology... Jan 2019A central dogma of mammalian reproductive biology is that the size of the primordial follicle pool represents reproductive capacity in females. The assembly of the... (Review)
Review
A central dogma of mammalian reproductive biology is that the size of the primordial follicle pool represents reproductive capacity in females. The assembly of the primordial follicle starts after the primordial germ cells (PGCs)-derived oocyte releases from the synchronously dividing germline cysts. PGCs initiate meiosis during fetal development. However, after synapsis and recombination of homologous chromosomes, they arrest at the diplotene stage of the first meiotic prophase (MI). The diplotene-arrested oocyte, together with the surrounding of a single layer of flattened granulosa cells, forms a basic unit of the ovary, the primordial follicle. At the start of each estrous (animal) or menstrual cycle (human), in response to a surge of luteinizing hormone (LH) from the pituitary gland, a limited number of primordial follicles are triggered to develop into primary follicles, preantral follicles, antral follicles and reach to preovulatory follicle stage. During the transition from the preantral to antral stages, the enclosed oocyte gradually acquires the capacity to resume meiosis. Meiotic resumption from the prophase of MI is morphologically characterized by the dissolution of the oocyte nuclear envelope, which is generally termed the "germinal vesicle breakdown" (GVBD). Following GVBD and completion of MI, the oocyte enters meiosis II without an obvious S-phase and arrests at metaphase phase II (MII) until fertilization. The underlying mechanism of meiotic arrest has been widely explored in numerous studies. Many studies indicated that two cellular second messengers, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) play an essential role in maintaining oocyte meiotic arrest. This review will discuss how these two cyclic nucleotides regulate oocyte maturation by blocking or initiating meiotic processes, and to provide an insight in future research.
Topics: Animals; Cyclic AMP; Cyclic GMP; Female; Granulosa Cells; Humans; Mammals; Meiosis; Meiotic Prophase I; Metaphase; Models, Biological; Oocytes
PubMed: 30611263
DOI: 10.1186/s12958-018-0445-8 -
Cells Apr 2022A conserved feature of virtually all higher eukaryotes is that the centromeres are embedded in heterochromatin. Here we provide evidence that this tight association... (Review)
Review
A conserved feature of virtually all higher eukaryotes is that the centromeres are embedded in heterochromatin. Here we provide evidence that this tight association between pericentric heterochromatin and the centromere is essential for proper metaphase exit and progression into telophase. Analysis of chromosome rearrangements that separate pericentric heterochromatin and centromeres indicates that they must remain associated in order to balance Cohesin/DNA catenation-based binding forces and centromere-based pulling forces during the metaphase-anaphase transition. In addition, a centromere embedded in heterochromatin facilitates nuclear envelope assembly around the entire complement of segregating chromosomes. Because the nuclear envelope initially forms on pericentric heterochromatin, nuclear envelope formation proceeds from the pole, thus providing time for incorporation of lagging and trailing chromosome arms into the newly formed nucleus. Additional analysis of noncanonical mitoses provides further insights into the functional significance of the tight association between heterochromatin and centromeres.
Topics: Anaphase; Centromere; Heterochromatin; Metaphase; Mitosis
PubMed: 35406810
DOI: 10.3390/cells11071247 -
Cell Jul 2017High-order chromatin structure plays important roles in gene expression regulation. Knowledge of the dynamics of 3D chromatin structures during mammalian embryo...
High-order chromatin structure plays important roles in gene expression regulation. Knowledge of the dynamics of 3D chromatin structures during mammalian embryo development remains limited. We report the 3D chromatin architecture of mouse gametes and early embryos using an optimized Hi-C method with low-cell samples. We find that mature oocytes at the metaphase II stage do not have topologically associated domains (TADs). In sperm, extra-long-range interactions (>4 Mb) and interchromosomal interactions occur frequently. The high-order structures of both the paternal and maternal genomes in zygotes and two-cell embryos are obscure but are gradually re-established through development. The establishment of the TAD structure requires DNA replication but not zygotic genome activation. Furthermore, unmethylated CpGs are enriched in A compartment, and methylation levels are decreased to a greater extent in A compartment than in B compartment in embryos. In summary, the global reprogramming of chromatin architecture occurs during early mammalian development.
Topics: Animals; Chromatin; CpG Islands; DNA Methylation; DNA Replication; Embryo, Mammalian; Embryonic Development; Epigenesis, Genetic; Female; Germ Cells; Male; Metaphase; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Oocytes; Spermatozoa; Zygote
PubMed: 28709003
DOI: 10.1016/j.cell.2017.06.029 -
Cytometry. Part a : the Journal of the... Apr 2021Flow cytometric analysis and sorting of plant mitotic chromosomes has been mastered by only a few laboratories worldwide. Yet, it has been contributing significantly to... (Review)
Review
Flow cytometric analysis and sorting of plant mitotic chromosomes has been mastered by only a few laboratories worldwide. Yet, it has been contributing significantly to progress in plant genetics, including the production of genome assemblies and the cloning of important genes. The dissection of complex genomes by flow sorting into the individual chromosomes that represent small parts of the genome reduces DNA sample complexity and streamlines projects relying on molecular and genomic techniques. Whereas flow cytometric analysis, that is, chromosome classification according to fluorescence and light scatter properties, is an integral part of any chromosome sorting project, it has rarely been used on its own due to lower resolution and sensitivity as compared to other cytogenetic methods. To perform chromosome analysis and sorting, commercially available electrostatic droplet sorters are suitable. However, in order to resolve and purify chromosomes of interest the instrument must offer high resolution of optical signals as well as stability during long runs. The challenge is thus not the instrumentation, but the adequate sample preparation. The sample must be a suspension of intact mitotic metaphase chromosomes and the protocol, which includes the induction of cell cycle synchrony, accumulation of dividing cells at metaphase, and release of undamaged chromosomes, is time consuming and laborious and needs to be performed very carefully. Moreover, in addition to fluorescent staining chromosomal DNA, the protocol may include specific labelling of DNA repeats to facilitate discrimination of particular chromosomes. This review introduces the applications of chromosome sorting in plants, and discusses in detail sample preparation, chromosome analysis and sorting to achieve the highest purity in flow-sorted fractions, and their suitability for downstream applications.
Topics: Cell Cycle; Chromosomes, Plant; Flow Cytometry; Metaphase; Plants
PubMed: 33615737
DOI: 10.1002/cyto.a.24324 -
Cells Feb 2020Oocyte maturation is a process that occurs in the ovaries, where an immature oocyte resumes meiosis to attain competence for normal fertilization after... (Review)
Review
Oocyte maturation is a process that occurs in the ovaries, where an immature oocyte resumes meiosis to attain competence for normal fertilization after ovulation/spawning. In starfish, the hormone 1-methyladenine binds to an unidentified receptor on the plasma membrane of oocytes, inducing a conformational change in the heterotrimeric GTP-binding protein α-subunit (Gα), so that the α-subunit binds GTP in exchange of GDP on the plasma membrane. The GTP-binding protein βγ-subunit (Gβγ) is released from Gα, and the released Gβγ activates phosphatidylinositol-3 kinase (PI3K), followed by the target of rapamycin kinase complex2 (TORC2) and 3-phosphoinositide-dependent protein kinase 1 (PDK1)-dependent phosphorylation of serum- and glucocorticoid-regulated kinase (SGK) of ovarian oocytes. Thereafter, SGK activates Na/H exchanger (NHE) to increase the intracellular pH (pHi) from ~6.7 to ~6.9. Moreover, SGK phosphorylates Cdc25 and Myt1, thereby inducing the de-phosphorylation and activation of cyclin B-Cdk1, causing germinal vesicle breakdown (GVBD). Both pHi increase and GVBD are required for spindle assembly at metaphase I, followed by MI arrest at pHi 6.9 until spawning. Due to MI arrest or SGK-dependent pHi control, spawned oocytes can be fertilized normally.
Topics: Adenine; Animals; Female; Fertilization; GTP-Binding Protein beta Subunits; GTP-Binding Protein gamma Subunits; Hydrogen-Ion Concentration; Immediate-Early Proteins; Metaphase; Oocytes; Oogenesis; Ovary; Phosphorylation; Protein Serine-Threonine Kinases; Receptors, G-Protein-Coupled; Starfish
PubMed: 32092921
DOI: 10.3390/cells9020476 -
STAR Protocols Mar 2021Whole planarian chromosome squash allows researchers to qualitatively analyze chromosome integrity. Treatment with colchicine is used to halt dividing cells within...
Whole planarian chromosome squash allows researchers to qualitatively analyze chromosome integrity. Treatment with colchicine is used to halt dividing cells within metaphase and does not require amputation or tissue puncturing. In combination with acetic-orcein, a stain-fixative for chromosomes, this strategy is suitable for animals with friable tissues caused by drug treatment, radiation, and RNA interference phenotypes. The whole planarian squash method presented here is a minimally invasive procedure that facilitates simultaneous analysis of chromosomal integrity in control and experimental animals. For complete details on the use and execution of this protocol, please refer to Peiris et al. (2016).
Topics: Animals; Chromosomes; Metaphase; Planarians; RNA Interference
PubMed: 33490976
DOI: 10.1016/j.xpro.2020.100257 -
Blood Dec 2022
Topics: Humans; Venous Thromboembolism; Multiple Myeloma; Cytogenetics; Metaphase
PubMed: 36480221
DOI: 10.1182/blood.2022017517 -
ELife May 2022Proliferating cells undergo metabolic changes in synchrony with cell cycle progression and cell division. Mitochondria provide fuel, metabolites, and ATP during...
Proliferating cells undergo metabolic changes in synchrony with cell cycle progression and cell division. Mitochondria provide fuel, metabolites, and ATP during different phases of the cell cycle, however it is not completely understood how mitochondrial function and the cell cycle are coordinated. CLUH (clustered mitochondria homolog) is a post-transcriptional regulator of mRNAs encoding mitochondrial proteins involved in oxidative phosphorylation and several metabolic pathways. Here, we show a role of CLUH in regulating the expression of astrin, which is involved in metaphase to anaphase progression, centrosome integrity, and mTORC1 inhibition. We find that CLUH binds both the mRNA and its product astrin, and controls the synthesis and the stability of the full-length astrin-1 isoform. We show that CLUH interacts with astrin-1 specifically during interphase. Astrin-depleted cells show mTORC1 hyperactivation and enhanced anabolism. On the other hand, cells lacking CLUH show decreased astrin levels and increased mTORC1 signaling, but cannot sustain anaplerotic and anabolic pathways. In absence of CLUH, cells fail to grow during G1, and progress faster through the cell cycle, indicating dysregulated matching of growth, metabolism, and cell cycling. Our data reveal a role of CLUH in coupling growth signaling pathways and mitochondrial metabolism with cell cycle progression.
Topics: Alcian Blue; Cell Cycle; Mechanistic Target of Rapamycin Complex 1; Metaphase; Mitochondria; Mitochondrial Proteins; Phenazines; Phenothiazines; RNA, Messenger; Resorcinols
PubMed: 35559794
DOI: 10.7554/eLife.74552 -
EMBO Reports May 2023Microspherule protein 1 (Mcrs1) is a component of the nonspecific lethal (NSL) complex and the chromatin remodeling INO80 complex, which participates in transcriptional...
Microspherule protein 1 (Mcrs1) is a component of the nonspecific lethal (NSL) complex and the chromatin remodeling INO80 complex, which participates in transcriptional regulation during mitosis. Here, we investigate the roles of Mcrs1 during female meiosis in mice. We demonstrate that Mcrs1 is a novel regulator of the meiotic G2/M transition and spindle assembly in mouse oocytes. Mcrs1 is present in the nucleus and associates with spindle poles and chromosomes of oocytes during meiosis I. Depletion of Mcrs1 alters HDAC2-mediated H4K16ac, H3K4me2, and H3K9me2 levels in nonsurrounded nucleolus (NSN)-type oocytes, and reduces CDK1 activity and cyclin B1 accumulation, leading to G2/M transition delay. Furthermore, Mcrs1 depletion results in abnormal spindle assembly due to reduced Aurora kinase (Aurka and Aurkc) and Kif2A activities, suggesting that Mcrs1 also plays a transcription-independent role in regulation of metaphase I oocytes. Taken together, our results demonstrate that the transcription factor Mcrs1 has important roles in cell cycle regulation and spindle assembly in mouse oocyte meiosis.
Topics: Female; Mice; Animals; Spindle Apparatus; Meiosis; Metaphase; Oocytes; Cell Cycle Checkpoints; Repressor Proteins; Kinesins; RNA-Binding Proteins
PubMed: 36951681
DOI: 10.15252/embr.202256273