Budding yeast ATM/ATR control meiotic double-strand break (DSB) levels by down-regulating Rec114, an essential component of the DSB-machinery

An essential feature of meiosis is Spo11 catalysis of programmed DNA double strand breaks (DSBs). Evidence suggests that the number of DSBs generated per meiosis is genetically determined and that this ability to maintain a pre-determined DSB level, or "DSB homeostasis", might be a property of the meiotic program. Here, we present direct evidence that Rec114, an evolutionarily conserved essential component of the meiotic DSB-machinery, interacts with DSB hotspot DNA, and that Tel1 and Mec1, the budding yeast ATM and ATR, respectively, down-regulate Rec114 upon meiotic DSB formation through phosphorylation. Mimicking constitutive phosphorylation reduces the interaction between Rec114 and DSB hotspot DNA, resulting in a reduction and/or delay in DSB formation. Conversely, a non-phosphorylatable rec114 allele confers a genome-wide increase in both DSB levels and in the interaction between Rec114 and the DSB hotspot DNA. These observations strongly suggest that Tel1 and/or Mec1 phosphorylation of Rec114 following Spo11 catalysis down-regulates DSB formation by limiting the interaction between Rec114 and DSB hotspots. We also present evidence that Ndt80, a meiosis specific transcription factor, contributes to Rec114 degradation, consistent with its requirement for complete cessation of DSB formation. Loss of Rec114 foci from chromatin is associated with homolog synapsis but independent of Ndt80 or Tel1/Mec1 phosphorylation. Taken together, we present evidence for three independent ways of regulating Rec114 activity, which likely contribute to meiotic DSBs-homeostasis in maintaining genetically determined levels of breaks

Physical properties of Centaur (60558) 174P/Echeclus from stellar occultations

The Centaur (60558) Echeclus was discovered on March 03, 2000, orbiting between the orbits of Jupiter and Uranus. After exhibiting frequent outbursts, it also received a comet designation, 174P. If the ejected material can be a source of debris to form additional structures, studying the surroundings of an active body like Echeclus can provide clues about the formation scenarios of rings, jets, or dusty shells around small bodies. Stellar occultation is a handy technique for this kind of investigation, as it can, from Earth-based observations, detect small structures with low opacity around these objects. Stellar occultation by Echeclus was predicted and observed in 2019, 2020, and 2021. We obtain upper detection limits of rings with widths larger than 0.5 km and optical depth of $\tau$ = 0.02. These values are smaller than those of Chariklo's main ring; in other words, a Chariklo-like ring would have been detected. The occultation observed in 2020 provided two positive chords used to derive the triaxial dimensions of Echeclus based on a 3D model and pole orientation available in the literature. We obtained $a = 37.0\pm0.6$ km, $b = 28.4 \pm 0.5$ km, and $c= 24.9 \pm 0.4$ km, resulting in an area-equivalent radius of $30.0 \pm 0.5$ km. Using the projected limb at the occultation epoch and the available absolute magnitude ($\rm{H}_{\rm{v}} = 9.971 \pm 0.031$), we calculate an albedo of $p_{\rm{v}} = 0.050 \pm 0.003$. Constraints on the object's density and internal friction are also proposed.Comment: Corrected and typeset versio

Budding Yeast Pch2, a Widely Conserved Meiotic Protein, Is Involved in the Initiation of Meiotic Recombination

Budding yeast Pch2 protein is a widely conserved meiosis-specific protein whose role is implicated in the control of formation and displacement of meiotic crossover events. In contrast to previous studies where the function of Pch2 was implicated in the steps after meiotic double-strand breaks (DSBs) are formed, we present evidence that Pch2 is involved in meiotic DSB formation, the initiation step of meiotic recombination. The reduction of DSB formation caused by the pch2 mutation is most prominent in the sae2 mutant background, whereas the impact remains mild in the rad51 dmc1 double mutant background. The DSB reduction is further pronounced when pch2 is combined with a hypomorphic allele of SPO11. Interestingly, the level of DSB reduction is highly variable between chromosomes, with minimal impact on small chromosomes VI and III. We propose a model in which Pch2 ensures efficient formation of meiotic DSBs which is necessary for igniting the subsequent meiotic checkpoint responses that lead to proper differentiation of meiotic recombinants

FANCD2-associated nuclease 1 partially compensates for the lack of Exonuclease 1 in mismatch repair

Germline mutations in the mismatch repair (MMR) genes MSH2, MSH6, MLH1 and PMS2 are linked to cancer of the colon and other organs, characterised by microsatellite instability and a large increase in mutation frequency. Unexpectedly, mutations in EXO1, encoding the only exonuclease genetically implicated in MMR, are not linked to familial cancer and cause a substantially weaker mutator phenotype. This difference could be explained if eukaryotic cells possessed additional exonucleases redundant with EXO1. Analysis of the MLH1 interactome identified FANCD2-associated nuclease 1 (FAN1), a novel enzyme with biochemical properties resembling EXO1. We now show that FAN1 efficiently substitutes for EXO1 in MMR assays and that this functional complementation is modulated by its interaction with MLH1. FAN1 also contributes towards MMR in vivo: cells lacking both EXO1 and FAN1 have a MMR defect and display resistance to N-methyl-N-nitrosourea (MNU) and 6-thioguanine (TG). Moreover, FAN1 loss amplifies the mutational profile of EXO1-deficient cells, implying that the two nucleases act redundantly in the same antimutagenic pathway. However, the increased drug resistance and mutator phenotype of FAN1/EXO1-deficient cells are less prominent than those seen in cells lacking MSH6 or MLH1. Eukaryotic cells thus apparently possess additional mechanisms that compensate for the loss of EXO1

Participation of TDP1 in the repair of formaldehyde-induced DNA-protein cross- links in chicken DT40 cells

Proteins are covalently trapped on DNA to form DNA-protein cross-links (DPCs) when cells are exposed to DNA-damaging agents. Aldehyde compounds produce common types of DPCs that contain proteins in an undisrupted DNA strand. Tyrosyl-DNA phosphodiesterase 1 (TDP1) repairs topoisomerase 1 (TOPO1) that is trapped at the 3’-end of DNA. In the pres- ent study, we examined the contribution of TDP1 to the repair of formaldehyde-induced DPCs using a reverse genetic strategy with chicken DT40 cells. The results obtained showed that cells deficient in TDP1 were sensitive to formaldehyde. The removal of formal- dehyde-induced DPCs was slower in tdp1-deficient cells than in wild type cells. We also found that formaldehyde did not produce trapped TOPO1, indicating that trapped TOPO1 was not a primary cytotoxic DNA lesion that was generated by formaldehyde and repaired by TDP1. The formaldehyde treatment resulted in the accumulation of chromosomal break- ages that were more prominent in tdp1-deficient cells than in wild type cells. Therefore, TDP1 plays a critical role in the repair of formaldehyde-induced DPCs that are distinct from trapped TOPO1