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

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.

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

10.1101/763235 ◽

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.

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Targeted Gene Knockout Reveals a Role in Meiotic Recombination for ZHP-3, a Zip3-Related Protein in Caenorhabditis elegans

Molecular and Cellular Biology ◽

10.1128/mcb.24.18.7998-8006.2004 ◽

2004 ◽

Vol 24 (18) ◽

pp. 7998-8006 ◽

Cited By ~ 69

Author(s):

Verena Jantsch ◽

Pawel Pasierbek ◽

Michael M. Mueller ◽

Dieter Schweizer ◽

Michael Jantsch ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Meiotic Recombination ◽

Fluorescent Protein ◽

Gene Knockout ◽

Chiasma Formation ◽

Double Strand Breaks ◽

Dna Double Strand Breaks ◽

Strand Breaks ◽

C Elegans ◽

Reciprocal Recombination

ABSTRACT The meiotically expressed Zip3 protein is found conserved from Saccharomyces cerevisiae to humans. In baker's yeast, Zip3p has been implicated in synaptonemal complex (SC) formation, while little is known about the protein's function in multicellular organisms. We report here the successful targeted gene disruption of zhp-3 (K02B12.8), the ZIP3 homolog in the nematode Caenorhabditis elegans. Homozygous zhp-3 knockout worms show normal homologue pairing and SC formation. Also, the timing of appearance and the nuclear localization of the recombination protein Rad-51 seem normal in these animals, suggesting proper initiation of meiotic recombination by DNA double-strand breaks. However, the occurrence of univalents during diplotene indicates that C. elegans ZHP-3 protein is essential for reciprocal recombination between homologous chromosomes and thus chiasma formation. In the absence of ZHP-3, reciprocal recombination is abolished and double-strand breaks seem to be repaired via alternative pathways, leading to achiasmatic chromosomes and the occurrence of univalents during meiosis I. Green fluorescent protein-tagged C. elegans ZHP-3 forms lines between synapsed chromosomes and requires the SC for its proper localization.

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Robust designation of meiotic crossover sites by CDK-2 through phosphorylation of the MutSγ complex

10.1101/2021.08.31.458431 ◽

2021 ◽

Author(s):

Jocelyn Haversat ◽

Alexander Woglar ◽

Kayla Klatt ◽

Chantal C. Akerib ◽

Victoria Roberts ◽

...

Keyword(s):

Positive Feedback ◽

Feedback Mechanism ◽

Double Strand Breaks ◽

Cyclin Dependent Kinase ◽

Wild Type ◽

Strand Breaks ◽

Homologous Chromosomes ◽

C Elegans ◽

Meiotic Crossover ◽

Positive Feedback Mechanism

SUMMARYCrossover formation is essential for proper segregation of homologous chromosomes during meiosis. Here we show that C. elegans Cyclin-dependent kinase 2 (CDK-2) forms a complex with cyclin-like protein COSA-1 and supports crossover formation by promoting conversion of meiotic double-strand breaks (DSBs) into crossover-specific recombination intermediates. Further, we identify MutSγ component MSH-5 as a CDK-2 phosphorylation target. MSH-5 has a disordered C-terminal tail that contains 13 potential CDK phosphosites and is required to concentrate crossover-promoting proteins at recombination sites. Phosphorylation of the MSH-5 tail appears dispensable in a wild- type background, but when MutSγ activity is partially compromised, crossover formation and retention of CDK-2/COSA-1 at recombination sites are exquisitely sensitive to phosphosite loss. Our data support a model in which robustness of crossover designation reflects a positive feedback mechanism involving CDK-2-mediated phosphorylation and scaffold-like properties of the MSH-5 C-terminal tail, features that combine to promote full recruitment and activity of crossover-promoting complexes.

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Methylation of histone H3K23 blocks DNA damage in pericentric heterochromatin during meiosis

eLife ◽

10.7554/elife.02996 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 35

Author(s):

Romeo Papazyan ◽

Ekaterina Voronina ◽

Jessica R Chapman ◽

Teresa R Luperchio ◽

Tonya M Gilbert ◽

...

Keyword(s):

Posttranslational Modifications ◽

Model Organism ◽

Pericentric Heterochromatin ◽

Double Strand Breaks ◽

Dna Double Strand Breaks ◽

Strand Breaks ◽

C Elegans ◽

Evolutionarily Conserved ◽

Established Role

Despite the well-established role of heterochromatin in protecting chromosomal integrity during meiosis and mitosis, the contribution and extent of heterochromatic histone posttranslational modifications (PTMs) remain poorly defined. Here, we gained novel functional insight about heterochromatic PTMs by analyzing histone H3 purified from the heterochromatic germline micronucleus of the model organism Tetrahymena thermophila. Mass spectrometric sequencing of micronuclear H3 identified H3K23 trimethylation (H3K23me3), a previously uncharacterized PTM. H3K23me3 became particularly enriched during meiotic leptotene and zygotene in germline chromatin of Tetrahymena and C. elegans. Loss of H3K23me3 in Tetrahymena through deletion of the methyltransferase Ezl3p caused mislocalization of meiosis-induced DNA double-strand breaks (DSBs) to heterochromatin, and a decrease in progeny viability. These results show that an evolutionarily conserved developmental pathway regulates H3K23me3 during meiosis, and our studies in Tetrahymena suggest this pathway may function to protect heterochromatin from DSBs.

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Repair pathway choice for double-strand breaks

Essays in Biochemistry ◽

10.1042/ebc20200007 ◽

2020 ◽

Vol 64 (5) ◽

pp. 765-777 ◽

Cited By ~ 2

Author(s):

Yixi Xu ◽

Dongyi Xu

Keyword(s):

Cell Cycle ◽

Double Strand Breaks ◽

Homologous Region ◽

Gene Repair ◽

Repair Pathway ◽

Dna Double Strand Breaks ◽

Environmental Radiation ◽

Strand Breaks ◽

Dna End Resection ◽

End Resection

Abstract Deoxyribonucleic acid (DNA) is at a constant risk of damage from endogenous substances, environmental radiation, and chemical stressors. DNA double-strand breaks (DSBs) pose a significant threat to genomic integrity and cell survival. There are two major pathways for DSB repair: nonhomologous end-joining (NHEJ) and homologous recombination (HR). The extent of DNA end resection, which determines the length of the 3′ single-stranded DNA (ssDNA) overhang, is the primary factor that determines whether repair is carried out via NHEJ or HR. NHEJ, which does not require a 3′ ssDNA tail, occurs throughout the cell cycle. 53BP1 and the cofactors PTIP or RIF1-shieldin protect the broken DNA end, inhibit long-range end resection and thus promote NHEJ. In contrast, HR mainly occurs during the S/G2 phase and requires DNA end processing to create a 3′ tail that can invade a homologous region, ensuring faithful gene repair. BRCA1 and the cofactors CtIP, EXO1, BLM/DNA2, and the MRE11–RAD50–NBS1 (MRN) complex promote DNA end resection and thus HR. DNA resection is influenced by the cell cycle, the chromatin environment, and the complexity of the DNA end break. Herein, we summarize the key factors involved in repair pathway selection for DSBs and discuss recent related publications.

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Bestimmung der DNA-Schädigung in vitro

Nuklearmedizin ◽

10.1055/s-0038-1626526 ◽

2010 ◽

Vol 49 (S 01) ◽

pp. S64-S68

Author(s):

E. Dikomey

Keyword(s):

Dna Damage ◽

Cell Lines ◽

Chromosome Aberrations ◽

Genetic Factors ◽

Double Strand Breaks ◽

Strand Breaks ◽

Cell Inactivation ◽

Repair Mechanisms ◽

Break Repair

SummaryIonising irradiation acts primarily via induction of DNA damage, among which doublestrand breaks are the most important lesions. These lesions may lead to lethal chromosome aberrations, which are the main reason for cell inactivation. Double-strand breaks can be repaired by several different mechanisms. The regulation of these mechanisms appears be fairly different for normal and tumour cells. Among different cell lines capacity of doublestrand break repair varies by only few percents and is known to be determined mostly by genetic factors. Knowledge about doublestrand break repair mechanisms and their regulation is important for the optimal application of ionising irradiation in medicine.

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