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Developmental Biology Feb 2021The early stages of development involve complex sequences of morphological changes that are both reproducible from embryo to embryo and often robust to environmental...
The early stages of development involve complex sequences of morphological changes that are both reproducible from embryo to embryo and often robust to environmental variability. To investigate the relationship between reproducibility and robustness we examined cell cycle progression in early Drosophila embryos at different temperatures. Our experiments show that while the subdivision of cell cycle steps is conserved across a wide range of temperatures (5-35 °C), the relative duration of individual steps varies with temperature. We find that the transition into prometaphase is delayed at lower temperatures relative to other cell cycle events, arguing that it has a different mechanism of regulation. Using an in vivo biosensor, we quantified the ratio of activities of the major mitotic kinase, Cdk1 and one of the major mitotic phosphatases PP1. Comparing activation profile with cell cycle transition times at different temperatures indicates that in early fly embryos activation of Cdk1 drives entry into prometaphase but is not required for earlier cell cycle events. In fact, chromosome condensation can still occur when Cdk1 activity is inhibited pharmacologically. These results demonstrate that different kinases are rate-limiting for different steps of mitosis, arguing that robust inter-regulation may be needed for rapid and ordered mitosis.
Topics: Animals; CDC2 Protein Kinase; Cell Cycle; Cell Cycle Checkpoints; Cyclin B; Drosophila Proteins; Drosophila melanogaster; Embryo, Nonmammalian; Enzyme Activation; Metaphase; Mitosis; Prometaphase; Prophase; Protein Phosphatase 1; Temperature
PubMed: 33278404
DOI: 10.1016/j.ydbio.2020.11.010 -
Genes & Development Dec 2020Proper segregation during meiosis requires that homologs be connected by the combination of crossovers and sister chromatid cohesion. To generate crossovers, numerous... (Review)
Review
Proper segregation during meiosis requires that homologs be connected by the combination of crossovers and sister chromatid cohesion. To generate crossovers, numerous double-strand breaks (DSBs) are introduced throughout the genome by the conserved Spo11 endonuclease. DSB formation and its repair are then highly regulated to ensure that homologous chromosomes contain at least one crossover and no DSBs remain prior to meiosis I segregation. The synaptonemal complex (SC) is a meiosis-specific structure formed between homologous chromosomes during prophase that promotes DSB formation and biases repair of DSBs to homologs over sister chromatids. Synapsis occurs when a particular recombination pathway is successful in establishing stable interhomolog connections. In this issue of , Mu and colleagues (pp. 1605-1618) show that SC formation between individual chromosomes provides the feedback to down-regulate Spo11 activity, thereby revealing an additional function for the SC.
Topics: Chromatids; DNA Breaks, Double-Stranded; Homologous Recombination; Meiosis; Synaptonemal Complex
PubMed: 33262143
DOI: 10.1101/gad.345488.120 -
Genes Mar 2022To generate gametes, sexually reproducing organisms need to achieve a reduction in ploidy, via meiosis. Several mechanisms are set in place to ensure proper reductional... (Review)
Review
To generate gametes, sexually reproducing organisms need to achieve a reduction in ploidy, via meiosis. Several mechanisms are set in place to ensure proper reductional chromosome segregation at the first meiotic division (MI), including chromosome remodeling during late prophase I. Chromosome remodeling after crossover formation involves changes in chromosome condensation and restructuring, resulting in a compact bivalent, with sister kinetochores oriented to opposite poles, whose structure is crucial for localized loss of cohesion and accurate chromosome segregation. Here, we review the general processes involved in late prophase I chromosome remodeling, their regulation, and the strategies devised by different organisms to produce bivalents with configurations that promote accurate segregation.
Topics: Chromosome Segregation; Kinetochores; Meiosis; Meiotic Prophase I
PubMed: 35328099
DOI: 10.3390/genes13030546 -
ELife Jan 2023Meiotic chromosome segregation relies on synapsis and crossover (CO) recombination between homologous chromosomes. These processes require multiple steps that are...
Meiotic chromosome segregation relies on synapsis and crossover (CO) recombination between homologous chromosomes. These processes require multiple steps that are coordinated by the meiotic cell cycle and monitored by surveillance mechanisms. In diverse species, failures in chromosome synapsis can trigger a cell cycle delay and/or lead to apoptosis. How this key step in 'homolog engagement' is sensed and transduced by meiotic cells is unknown. Here we report that in , recruitment of the Polo-like kinase PLK-2 to the synaptonemal complex triggers phosphorylation and inactivation of CHK-2, an early meiotic kinase required for pairing, synapsis, and double-strand break (DSB) induction. Inactivation of CHK-2 terminates DSB formation and enables CO designation and cell cycle progression. These findings illuminate how meiotic cells ensure CO formation and accurate chromosome segregation.
Topics: Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Checkpoint Kinase 2; Chromosome Pairing; Meiosis; Synaptonemal Complex
PubMed: 36700544
DOI: 10.7554/eLife.84492 -
The Journal of Cell Biology Feb 2021Meiosis creates genetic diversity by recombination and segregation of chromosomes. The synaptonemal complex assembles during meiotic prophase I and assists faithful...
Meiosis creates genetic diversity by recombination and segregation of chromosomes. The synaptonemal complex assembles during meiotic prophase I and assists faithful exchanges between homologous chromosomes, but how its assembly/disassembly is regulated remains to be understood. Here, we report how two major posttranslational modifications, phosphorylation and ubiquitination, cooperate to promote synaptonemal complex assembly. We found that the ubiquitin ligase complex SCF is important for assembly and maintenance of the synaptonemal complex in Drosophila female meiosis. This function of SCF is mediated by two substrate-recognizing F-box proteins, Slmb/βTrcp and Fbxo42. SCF-Fbxo42 down-regulates the phosphatase subunit PP2A-B56, which is important for synaptonemal complex assembly and maintenance.
Topics: Animals; DNA Breaks, Double-Stranded; Down-Regulation; Drosophila Proteins; Drosophila melanogaster; F-Box Proteins; Meiosis; Protein Phosphatase 2; Recombination, Genetic; SKP Cullin F-Box Protein Ligases; Synaptonemal Complex
PubMed: 33382409
DOI: 10.1083/jcb.202009167 -
Journal of Cell Science Feb 2022Appropriate DNA double-strand break (DSB) and crossover distributions are required for proper meiotic chromosome segregation. Schizosaccharomyces pombe linear element...
Appropriate DNA double-strand break (DSB) and crossover distributions are required for proper meiotic chromosome segregation. Schizosaccharomyces pombe linear element proteins (LinEs) determine DSB hotspots; LinE-bound hotspots form three-dimensional clusters over ∼200 kb chromosomal regions. Here, we investigated LinE configurations and distributions in live cells using super-resolution fluorescence microscopy. We found LinEs form two chromosomal structures, dot-like and linear structures, in both zygotic and azygotic meiosis. Dot-like LinE structures appeared around the time of meiotic DNA replication, underwent dotty-to-linear-to-dotty configurational transitions and disassembled before the first meiotic division. DSB formation and repair did not detectably influence LinE structure formation but failure of DSB formation delayed disassembly. Recombination-deficient LinE missense mutants formed dot-like, but not linear, LinE structures. Our quantitative study reveals a transient form of LinE structures and suggests a novel role for LinE proteins in regulating meiotic events, such as DSB repair. We discuss the relationship of LinEs and the synaptonemal complex in other species. This article has an associated First Person interview with the first author of the paper.
Topics: DNA; DNA Breaks, Double-Stranded; Humans; Meiosis; Schizosaccharomyces; Schizosaccharomyces pombe Proteins; Synaptonemal Complex
PubMed: 35028663
DOI: 10.1242/jcs.259061 -
Nature Communications Mar 2023During meiotic prophase I, spermatocytes must balance transcriptional activation with homologous recombination and chromosome synapsis, biological processes requiring...
During meiotic prophase I, spermatocytes must balance transcriptional activation with homologous recombination and chromosome synapsis, biological processes requiring extensive changes to chromatin state. We explored the interplay between chromatin accessibility and transcription through prophase I of mammalian meiosis by measuring genome-wide patterns of chromatin accessibility, nascent transcription, and processed mRNA. We find that Pol II is loaded on chromatin and maintained in a paused state early during prophase I. In later stages, paused Pol II is released in a coordinated transcriptional burst mediated by the transcription factors A-MYB and BRDT, resulting in ~3-fold increase in transcription. Transcriptional activity is temporally and spatially segregated from key steps of meiotic recombination: double strand breaks show evidence of chromatin accessibility earlier during prophase I and at distinct loci from those undergoing transcriptional activation, despite shared chromatin marks. Our findings reveal mechanisms underlying chromatin specialization in either transcription or recombination in meiotic cells.
Topics: Animals; Male; Chromatin; Chromosomes; Gene Expression Regulation; Mammals; Meiosis; RNA Polymerase II; Spermatocytes; Proto-Oncogene Proteins; Trans-Activators; Nuclear Proteins
PubMed: 36990976
DOI: 10.1038/s41467-023-37408-w -
Molecular Cell Feb 2022We have used a combination of chemical genetics, chromatin proteomics, and imaging to map the earliest chromatin transactions during vertebrate cell entry into mitosis....
We have used a combination of chemical genetics, chromatin proteomics, and imaging to map the earliest chromatin transactions during vertebrate cell entry into mitosis. Chicken DT40 CDK1 cells undergo synchronous mitotic entry within 15 min following release from a 1NM-PP1-induced arrest in late G. In addition to changes in chromatin association with nuclear pores and the nuclear envelope, earliest prophase is dominated by changes in the association of ribonucleoproteins with chromatin, particularly in the nucleolus, where pre-rRNA processing factors leave chromatin significantly before RNA polymerase I. Nuclear envelope barrier function is lost early in prophase, and cytoplasmic proteins begin to accumulate on the chromatin. As a result, outer kinetochore assembly appears complete by nuclear envelope breakdown (NEBD). Most interphase chromatin proteins remain associated with chromatin until NEBD, after which their levels drop sharply. An interactive proteomic map of chromatin transactions during mitotic entry is available as a resource at https://mitoChEP.bio.ed.ac.uk.
Topics: Animals; CDC2 Protein Kinase; Cell Line, Tumor; Chickens; Chromatin; Chromatin Assembly and Disassembly; Chromosomes; DNA; Lamin Type B; Lymphoma, B-Cell; Nuclear Proteins; Prophase; Protein Binding; Proteome; Proteomics; RNA, Ribosomal; Time Factors
PubMed: 35090599
DOI: 10.1016/j.molcel.2021.12.039 -
MicroPublication Biology 2022Human retinal pigment epithelium RPE-1 cells are immortalized diploid wild-type cells. RPE-1 is increasingly used for studies of spindle assembly dynamics and chromosome...
Human retinal pigment epithelium RPE-1 cells are immortalized diploid wild-type cells. RPE-1 is increasingly used for studies of spindle assembly dynamics and chromosome segregation. Here, we imaged living RPE-1 cells using the spinning disk confocal microscope and report their complete spindle assembly dynamic parameters. Live-cell experiments enabled ascribing precise timing of function of the kinesin-5 Eg5 and kinesin-14 HSET throughout different phases of mitosis. Eg5 functions at prophase and metaphase, to assemble and maintain spindle bipolarity, respectively. Eg5 inhibition results in spindle collapse during prophase and metaphase, resulting in monoastral/monopolar spindles. HSET functions throughout mitosis to maintain spindle length. HSET degradation results in shorter spindles through all phases of mitosis. Double-inhibition of Eg5 and HSET produces only monoastral/monopolar spindles, indicating that Eg5 and HSET may not be antagonistic in wild-type RPE-1 cells, contrary to previous studies using cancer cells. In the context of spindle assembly, our results highlight potential important differences between RPE-1 and other cancer-derived cell lines.
PubMed: 36004005
DOI: 10.17912/micropub.biology.000623 -
Movement Disorders : Official Journal... Oct 2019While current effective therapies are available for the symptomatic control of PD, treatments to halt the progressive neurodegeneration still do not exist. Loss of... (Review)
Review
While current effective therapies are available for the symptomatic control of PD, treatments to halt the progressive neurodegeneration still do not exist. Loss of dopamine neurons in the SNc and dopamine terminals in the striatum drive the motor features of PD. Multiple lines of research point to several pathways which may contribute to dopaminergic neurodegeneration. These pathways include extensive axonal arborization, mitochondrial dysfunction, dopamine's biochemical properties, abnormal protein accumulation of α-synuclein, defective autophagy and lysosomal degradation, and synaptic impairment. Thus, understanding the essential features and mechanisms of dopaminergic neuronal vulnerability is a major scientific challenge and highlights an outstanding need for fostering effective therapies against neurodegeneration in PD. This article, which arose from the Movement Disorders 2018 Conference, discusses and reviews the possible mechanisms underlying neuronal vulnerability and potential therapeutic approaches in PD. © 2019 International Parkinson and Movement Disorder Society.
Topics: Animals; Axons; Chromosome Pairing; Dopaminergic Neurons; Humans; Parkinson Disease; Parkinsonian Disorders; Presynaptic Terminals
PubMed: 31483900
DOI: 10.1002/mds.27823