scholarly journals The CHK-2 antagonizing phosphatase PPM-1.D regulates meiotic entry via catalytic and non-catalytic activities

2021 ◽  
Author(s):  
Antoine Baudrimont ◽  
Dimitra Paouneskou ◽  
Ariz Mohammad ◽  
Raffael Lichtenberger ◽  
Joshua Blundon ◽  
...  
Keyword(s):  
Meiotic Prophase ◽  
Nuclear Periphery ◽  
Dna Double Strand Breaks ◽  
Timely Initiation ◽  
Regulatory Pathways ◽  
Strand Breaks ◽  
Catalytic Activities ◽  
C Elegans ◽  
Transcriptional Regulatory ◽  
Meiotic Entry

The transition from the stem cell/progenitor fate to meiosis is mediated by several redundant post-transcriptional regulatory pathways in C. elegans. Interfering with all three branches causes tumorous germlines. SCFPROM-1 comprises one branch and mediates a scheduled degradation step at entry into meiosis. prom-1 mutants show defects in timely initiation of events of meiotic prophase I, resulting in high rates of embryonic lethality. Here, we identify a crucial substrate for PROM-1, encoded by the phosphatase PPM-1.D/Wip1. We report that it antagonizes CHK-2 kinase, a key regulator for meiotic prophase initiation e.g., DNA double strand breaks, chromosome pairing and synaptonemal complex formation. We propose that PPM-1.D controls the amount of active CHK-2 by both catalytic and non-catalytic activities, where strikingly the non-catalytic regulation seems to be crucial at meiotic entry. PPM-1.D sequesters CHK-2 at the nuclear periphery and programmed SCFPROM-1 mediated degradation of PPM-1.D liberates the kinase and promotes meiotic entry.

Loss Of NIPA Leads To Defects In The Repair Of DNA Double Strand Breaks In Mice

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2488-2488
Author(s):  
Anna Lena Illert ◽  
Cristina Antinozzi ◽  
Hiroyuki Kawaguchi ◽  
Michal Kulinski ◽  
Christine Klitzing ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Meiotic Prophase ◽  
Conflicts Of Interest ◽  
Cyclin B1 ◽  
Double Strand Breaks ◽  
Conditional Knockout ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Chromosome Axis

Abstract Regulated oscillation of protein expression is an essential mechanism of cell cycle control. The SCF class of E3 ubiquitin ligases is involved in this process by targeting cell cycle regulatory proteins for degradation by the proteasome. We previously reported the cloning of NIPA (Nuclear Interaction Partner of ALK) in complex with constitutively active oncogenic fusions of ALK, which contributes to the development of lymphomas and sarcomas. Subsequently we characterized NIPA as a F-Box protein that defines an oscillating ubiquitin E3 ligase targeting nuclear cyclin B1 in interphase thus contributing to the timing of mitotic entry. Using a conditional knockout strategy we inactivated the gene encoding Nipa. Nipa-deficient animals are viable, but show a lower birth rate and a reduced body weight. Furthermore, Nipa-deficient males were sterile due to a block of spermatogenesis during meiotic prophase. Virtually no spermatocytes progress beyond a late-zygotene to early-pachytene stage and reach an aberrant stage, with synaptonemal complex disassembly and incomplete synapsis. Nipa-/- females are sub-fertile with an early and severe meiotic defect during embryogenesis with extensive apoptosis in early prophase (E13.5-E14.5). Here we report, that Nipa-/- meiocytes exhibit persistent cytological markers for DNA double strand break repair proteins (like DMC1, RAD51) in meiotic prophase with more than twice as many DMC1 foci as control animals. Kinetic analysis of the first wave of spermatogenesis showed increased DMC1/RAD51 foci in Nipa-/- cells as soon as early-pachynema cells appear (13-14 days post partum). Moreover, we show that Nipa deficiency does not lead to a defect in meiotic sex chromosome inactivation despite epithelial stage IV apoptosis. Nipa-deficient spermatocytes exhibit numerous abnormalities in staining of chromosome axis associated proteins (like SYCP3 and STAG3) indicating that chromosome axis defects were associated with compromised chromosome axis integrity leading to overt chromosome fragmentation. Further in vitro analyses with bleomycin treated MEFs displayed high pH2AX levels in cells lacking NIPA. Repair of DNA DSB seemed to be abolished in these cells as the pH2AX-level were sustained and still visible after 90 min of timecourse, where wildtype cells already repaired sides of DNA Damage. Consistent with these findings NIPA-deficient spleen cells showed compromised DNA Damage repair measured in a comet assay with a significantly longer olive tail moment in NIPA knockout cells under repair conditions. Taken together, the phenotype of Nipa-knockout mice is a definitive proof of the meiotic significance of NIPA and our results show a new, unsuspected role of NIPA in chromosome stability and the repair of DNA double strand breaks. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Actin’ between phase separated domains for heterochromatin repair

2019 ◽  
Author(s):  
Chetan C Rawal ◽  
Christopher P Caridi ◽  
Irene Chiolo
Keyword(s):  
Homologous Recombination ◽  
Actin Filaments ◽  
Repetitive Sequences ◽  
Nuclear Periphery ◽  
Double Strand Breaks ◽  
Nuclear Actin ◽  
Dna Double Strand Breaks ◽  
Directed Motion ◽  
Strand Breaks ◽  
Increased Risk

DNA double-strand breaks (DSBs) are particularly challenging to repair in pericentromeric heterochromatin because of the increased risk of aberrant recombination in highly repetitive sequences. Recent studies have identified specialized mechanisms enabling ‘safe’ homologous recombination (HR) repair in heterochromatin. These include striking nuclear actin filaments (F-actin) and myosins that drive the directed motion of repair sites to the nuclear periphery for ‘safe' repair. Here, we summarize our current understanding of the mechanisms involved, and propose how they might operate in the context of a phase-separated environment.

scholarly journals Actin’ between phase separated domains for heterochromatin repair

2019 ◽  
Author(s):  
Chetan C Rawal ◽  
Christopher P Caridi ◽  
Irene Chiolo
Keyword(s):  
Homologous Recombination ◽  
Actin Filaments ◽  
Repetitive Sequences ◽  
Nuclear Periphery ◽  
Double Strand Breaks ◽  
Nuclear Actin ◽  
Dna Double Strand Breaks ◽  
Directed Motion ◽  
Strand Breaks ◽  
Increased Risk

DNA double-strand breaks (DSBs) are particularly challenging to repair in pericentromeric heterochromatin because of the increased risk of aberrant recombination in highly repetitive sequences. Recent studies have identified specialized mechanisms enabling ‘safe’ homologous recombination (HR) repair in heterochromatin. These include striking nuclear actin filaments (F-actin) and myosins that drive the directed motion of repair sites to the nuclear periphery for ‘safe' repair. Here, we summarize our current understanding of the mechanisms involved, and propose how they might operate in the context of a phase-separated environment.

scholarly journals Mechanism and Control of Meiotic DNA Double-Strand Break Formation in S. cerevisiae

2021 ◽  
Vol 9 ◽  
Author(s):  
Vikash Kumar Yadav ◽  
Corentin Claeys Bouuaert
Keyword(s):  
Strand Break ◽  
Current View ◽  
Dna Double Strand Breaks ◽  
Recombination Mechanism ◽  
Regulatory Pathways ◽  
Strand Breaks ◽  
Dna Double Strand Break ◽  
And Function ◽  
And Control

Developmentally programmed formation of DNA double-strand breaks (DSBs) by Spo11 initiates a recombination mechanism that promotes synapsis and the subsequent segregation of homologous chromosomes during meiosis. Although DSBs are induced to high levels in meiosis, their formation and repair are tightly regulated to minimize potentially dangerous consequences for genomic integrity. In S. cerevisiae, nine proteins participate with Spo11 in DSB formation, but their molecular functions have been challenging to define. Here, we describe our current view of the mechanism of meiotic DSB formation based on recent advances in the characterization of the structure and function of DSB proteins and discuss regulatory pathways in the light of recent models.

scholarly journals ATR is required to complete meiotic recombination in mice

2017 ◽  
Author(s):  
Sarai Pacheco ◽  
Andros Maldonado-Linares ◽  
Marina Marcet-Ortega ◽  
Cristina Rojas ◽  
Ana Martínez-Marchal ◽  
...  
Keyword(s):  
Meiotic Recombination ◽  
Meiotic Prophase ◽  
Protein A ◽  
Homology Search ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Molecular Features ◽  
Chromosome Synapsis ◽  
Meiotic Dsbs

ABSTRACTPrecise execution of recombination during meiosis is essential for forming chromosomally balanced gametes. Meiotic recombination initiates with the formation and resection of DNA double-strand breaks (DSBs). Binding of replication protein A (RPA) at resected DSBs fosters association of RAD51 and DMC1, the primary effectors of homology search. It is well appreciated that cellular responses to meiotic DSBs are critical for efficient repair and quality control, but molecular features of these responses remain poorly understood, particularly in mammals. Here we provide evidence that the DNA damage response protein kinase ATR is crucial for meiotic recombination and completion of meiotic prophase in mice. Using a hypomorphic Atr mutation and pharmacological inhibition of ATR in vivo and in cultured spermatocytes, we show that ATR, through its effector kinase CHK1, promotes efficient RAD51 and DMC1 assembly at RPA-coated DSB sites and establishment of interhomolog connections during meiosis. Furthermore, our findings suggest that ATR promotes local accumulation of recombination markers on unsynapsed axes during meiotic prophase to favor homologous chromosome synapsis. These data reveal that ATR plays multiple roles in mammalian meiotic recombination.

scholarly journals The fate of irreparable DNA double-strand breaks and eroded telomeres at the nuclear periphery

Nucleus ◽  
2010 ◽  
Vol 1 (2) ◽  
pp. 158-161 ◽  
Author(s):  
Michael Lisby ◽  
Teresa Teixeira ◽  
Eric Gilson ◽  
Vincent Géli
Keyword(s):  
Nuclear Periphery ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks

scholarly journals Heterologous synapsis in C. elegans is regulated by meiotic double-strand breaks and crossovers

2021 ◽  
Author(s):  
Hanwenheng Liu ◽  
Spencer G. Gordon ◽  
Ofer Rog
Keyword(s):  
Nuclear Envelope ◽  
Synaptonemal Complex ◽  
Meiotic Prophase ◽  
Double Strand Breaks ◽  
Strand Breaks ◽  
C Elegans ◽  
Homologous Sequences ◽  
Almost All ◽  
Break Formation ◽  
Successful Production

AbstractAlignment of the parental chromosomes during meiotic prophase is key to the formation of genetic exchanges, or crossovers, and consequently to the successful production of gametes. In almost all studied organisms, alignment involves synapsis: the assembly of a conserved inter-chromosomal interface called the synaptonemal complex (SC). While the SC usually synapses homologous sequences, it can assemble between heterologous sequences. However, little is known about the regulation of heterologous synapsis. Here we study the dynamics of heterologous synapsis in the nematode C. elegans. We characterize two experimental scenarios: SC assembly onto a folded-back chromosome that cannot pair with its homologous partner; and synapsis of pseudo-homologs, a fusion chromosome partnering with an unfused chromosome half its size. We observed elevated levels of heterologous synapsis when the number of meiotic double-strand breaks or crossovers were reduced, indicating that the promiscuity of synapsis is regulated by break formation or repair. By manipulating the levels of breaks and crossovers, we infer both chromosome-specific and nucleus-wide regulation on heterologous synapsis. Finally, we identify differences between the two conditions, suggesting that attachment to the nuclear envelope plays a role in regulating heterologous synapsis.

scholarly journals NHJ-1 regulates canonical non-homologous end joining in Caenorhabditis elegans

2019 ◽  
Author(s):  
Aleksandar Vujin ◽  
Steven J. Jones ◽  
Monique Zetka
Keyword(s):  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Dapi Staining ◽  
Strand Breaks ◽  
C Elegans ◽  
Meiotic Dsbs ◽  
Ring Complex ◽  
Non Homologous End Joining

AbstractCanonical non-homologous end joining (cNHEJ) is a near-universally conserved pathway for the repair of DNA double-strand breaks (DSBs). While the cNHEJ pathway encompasses more than a dozen factors in vertebrates and is similarly complex in other eukaryotes, in the nematode C. elegans the entire known cNHEJ toolkit consists of two proteins that comprise the Ku ring complex, cku-70 and cku-80, and the terminal ligase lig-4. Here, we report the discovery of nhj-1 as the fourth cNHEJ factor in C. elegans. Observing a difference in the phenotypic response to ionizing radiation (IR) between two lines of the wild type N2 strain, we mapped the locus causative of IR-sensitivity to a candidate on chromosome V. Using CRISPR-Cas9 mutagenesis, we show that disrupting the nhj-1 sequence induces IR-sensitivity in an IR-resistant background. Double mutants of nhj-1 and the cNHEJ factors lig-4 or cku-80 do not exhibit additive IR-sensitivity, arguing that nhj-1 is a member of the cNHEJ pathway. Furthermore, like the loss of lig-4, the loss of nhj-1 in the com-1 genetic background, in which meiotic DSBs are repaired by cNHEJ instead of homologous recombination, increased the number of DAPI-staining bodies in diakinesis, consistent with increased chromosome fragmentation in the absence of cNHEJ repair. Finally, we show that NHJ-1 localizes to many somatic nuclei in the L1 larva, but not the primordial germline, which is in accord with a role in the predominantly somatically active cNHEJ. Although nhj-1 shares no sequence homology with other known eukaryotic cNHEJ factors and is taxonomically restricted to the Rhadbitid family, its discovery underscores the evolutionary plasticity of even highly conserved pathways, and may represent a springboard for further characterization of cNHEJ in C. elegans.

scholarly journals Downregulation of Barley Regulator of Telomere Elongation Helicase 1 Alters the Distribution of Meiotic Crossovers

2021 ◽  
Vol 12 ◽  
Author(s):  
Abdellah Barakate ◽  
Mikel Arrieta ◽  
Malcolm Macaulay ◽  
Sebastian Vivera ◽  
Diane Davidson ◽  
...  
Keyword(s):  
Snp Array ◽  
Reproductive Organs ◽  
Dna Helicase ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Telomere Elongation ◽  
C Elegans ◽  
Genome Wide ◽  
Gene Structure And Expression ◽  
And Control

Programmed meiotic DNA double-strand breaks (DSBs), necessary for proper chromosomal segregation and viable gamete formation, are repaired by homologous recombination (HR) as crossovers (COs) or non-crossovers (NCOs). The mechanisms regulating the number and distribution of COs are still poorly understood. The regulator of telomere elongation helicase 1 (RTEL1) DNA helicase was previously shown to enforce the number of meiotic COs in Caenorhabditis elegans but its function in plants has been studied only in the vegetative phase. Here, we characterised barley RTEL1 gene structure and expression using RNA-seq data previously obtained from vegetative and reproductive organs and tissues. Using RNAi, we downregulated RTEL1 expression specifically in reproductive tissues and analysed its impact on recombination using a barley 50k iSelect SNP Array. Unlike in C. elegans, in a population segregating for RTEL1 downregulated by RNAi, high resolution genome-wide genetic analysis revealed a significant increase of COs at distal chromosomal regions of barley without a change in their total number. Our data reveal the important role of RTEL1 helicase in plant meiosis and control of recombination.

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