Rad9/53BP1 promotes DNA repair via crossover recombination by limiting the Sgs1 and Mph1 helicases

The DNA damage checkpoint (DDC) is often robustly activated during the homologous recombination (HR) repair of DNA double strand breaks (DSBs). DDC activation controls several HR repair factors by phosphorylation, preventing premature segregation of entangled chromosomes formed during HR repair. The DDC mediator 53BP1/Rad9 limits the nucleolytic processing (resection) of a DSB, controlling the formation of the 3\u2032 single-stranded DNA (ssDNA) filament needed for recombination, from yeast to human. Here we show that Rad9 promotes stable annealing between the recombinogenic filament and the donor template in yeast, limiting strand rejection by the Sgs1 and Mph1 helicases. This regulation allows repair by long tract gene conversion, crossover recombination and break-induced replication (BIR), only after DDC activation. These findings shed light on how cells couple DDC with the choice and effectiveness of HR sub-pathways, with implications for genome instability and cancer

Slx4 and Rtt107 control checkpoint signalling and DNA resection at double-strand breaks

The DNA damage checkpoint pathway is activated in response to DNA lesions and replication stress to preserve genome integrity. However, hyper-activation of this surveillance system is detrimental to the cell, because it might prevent cell cycle re-start after repair, which may also lead to senescence. Here we show that the scaffold proteins Slx4 and Rtt107 limit checkpoint signalling at a persistent double-strand DNA break (DSB) and at uncapped telomeres. We found that Slx4 is recruited within a few kilobases of an irreparable DSB, through the interaction with Rtt107 and the multi-BRCT domain scaffold Dpb11. In the absence of Slx4 or Rtt107, Rad9 binding near the irreparable DSB is increased, leading to robust checkpoint signalling and slower nucleolytic degradation of the 5' strand. Importantly, in slx4\u394 sae2\u394 double mutant cells these phenotypes are exacerbated, causing a severe Rad9-dependent defect in DSB repair. Our study sheds new light on the molecular mechanism that coordinates the processing and repair of DSBs with DNA damage checkpoint signalling, preserving genome integrity

Mapping geographical inequalities in childhood diarrhoeal morbidity and mortality in low-income and middle-income countries, 2000–17 : analysis for the Global Burden of Disease Study 2017

Background Across low-income and middle-income countries (LMICs), one in ten deaths in children younger than 5 years is attributable to diarrhoea. The substantial between-country variation in both diarrhoea incidence and mortality is attributable to interventions that protect children, prevent infection, and treat disease. Identifying subnational regions with the highest burden and mapping associated risk factors can aid in reducing preventable childhood diarrhoea. Methods We used Bayesian model-based geostatistics and a geolocated dataset comprising 15 072 746 children younger than 5 years from 466 surveys in 94 LMICs, in combination with findings of the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2017, to estimate posterior distributions of diarrhoea prevalence, incidence, and mortality from 2000 to 2017. From these data, we estimated the burden of diarrhoea at varying subnational levels (termed units) by spatially aggregating draws, and we investigated the drivers of subnational patterns by creating aggregated risk factor estimates. Findings The greatest declines in diarrhoeal mortality were seen in south and southeast Asia and South America, where 54·0% (95% uncertainty interval [UI] 38·1–65·8), 17·4% (7·7–28·4), and 59·5% (34·2–86·9) of units, respectively, recorded decreases in deaths from diarrhoea greater than 10%. Although children in much of Africa remain at high risk of death due to diarrhoea, regions with the most deaths were outside Africa, with the highest mortality units located in Pakistan. Indonesia showed the greatest within-country geographical inequality; some regions had mortality rates nearly four times the average country rate. Reductions in mortality were correlated to improvements in water, sanitation, and hygiene (WASH) or reductions in child growth failure (CGF). Similarly, most high-risk areas had poor WASH, high CGF, or low oral rehydration therapy coverage. Interpretation By co-analysing geospatial trends in diarrhoeal burden and its key risk factors, we could assess candidate drivers of subnational death reduction. Further, by doing a counterfactual analysis of the remaining disease burden using key risk factors, we identified potential intervention strategies for vulnerable populations. In view of the demands for limited resources in LMICs, accurately quantifying the burden of diarrhoea and its drivers is important for precision public health

Mapping local patterns of childhood overweight and wasting in low- and middle-income countries between 2000 and 2017

A double burden of malnutrition occurs when individuals, household members or communities experience both undernutrition and overweight. Here, we show geospatial estimates of overweight and wasting prevalence among children under 5 years of age in 105 low- and middle-income countries (LMICs) from 2000 to 2017 and aggregate these to policy-relevant administrative units. Wasting decreased overall across LMICs between 2000 and 2017, from 8.4 (62.3 (55.1�70.8) million) to 6.4 (58.3 (47.6�70.7) million), but is predicted to remain above the World Health Organization�s Global Nutrition Target of <5 in over half of LMICs by 2025. Prevalence of overweight increased from 5.2 (30 (22.8�38.5) million) in 2000 to 6.0 (55.5 (44.8�67.9) million) children aged under 5 years in 2017. Areas most affected by double burden of malnutrition were located in Indonesia, Thailand, southeastern China, Botswana, Cameroon and central Nigeria. Our estimates provide a new perspective to researchers, policy makers and public health agencies in their efforts to address this global childhood syndemic. © 2020, The Author(s)

Author Correction: Mapping local patterns of childhood overweight and wasting in low- and middle-income countries between 2000 and 2017 (Nature Medicine, (2020), 26, 5, (750-759), 10.1038/s41591-020-0807-6)

An amendment to this paper has been published and can be accessed via a link at the top of the paper. © 2020, The Author(s)

Author Correction: Mapping local patterns of childhood overweight and wasting in low- and middle-income countries between 2000 and 2017 (Nature Medicine, (2020), 26, 5, (750-759), 10.1038/s41591-020-0807-6)

An amendment to this paper has been published and can be accessed via a link at the top of the paper. © 2020, The Author(s)

Cdc5 kinase activity links cell cycle regulation with genome stability

In response to DNA damage, all eukaryotic organisms activate a surveillance mechanism, called DNA damage checkpoint (DDC), to arrest cell cycle progression and facilitate DNA repair. Several factors are physically recruited to the damaged sites, and specific kinases phosphorylate multiple targets leading to checkpoint activation. Studies both in yeast and mammals have involved the Polo-like kinase 1 (PLK1; Cdc5 in yeast) in turning off the DNA damage checkpoint and promoting the cell cycle re-start after DNA damage mediated checkpoint arrest. To further characterize the Cdc5 pathway in preserving genome stability, we investigated the cdc5-T238A mutation, which abolishes the in trans-phosphorylation by an unknown kinase in the activation loop of the kinase domain. Firstly, we found that the Cdc5-T238A protein variant has a lower kinase activity by in vitro assay without affecting cell cycle dependent protein stability. Also, we found that the cdc5-T238A cells have a mild G2/M delay in unperturbed conditions. Of importance, cdc5-T238A cells show severe defects in DDC inactivation and cell cycle restart after one persistent DNA double strand break (DSB) and uncapped telomeres. Interestingly, this checkpoint adaptation defect is associated to a significant delay in localization of Cdc5-T238A protein variant at spindle pole bodies. Furthermore, genetic analysis indicates that cdc5-T238A cells have prolonged activation of multiple mitotic checkpoint factors, such as Bfa1, Mad2 and Cdh1, after one irreparable DSB. Finally, we found that cdc5-T238A cells have a significant increase in chromosome loss and gross chromosomal rearrangement rates, which are features of genome instability. In particular, genetic and biochemical analysis indicate that the cdc5-T238A cells have compromised activity of the Mus81-Mms4 pathway, which is involved in the resolution of recombination intermediates during stressful replication and in response to DNA damage. Further characterization of this and other cdc5 alleles in yeast promises to be informative to elucidate the functional role of PLK1 in preserving genome stability in mammals, and to develop novel cancer therapy approaches

Sen1/SETX helicase limits DNA:RNA hybrids at DNA double strand breaks and determines faithful repair through end joining and homologous recombination

The repair of DNA double strand breaks (DSB) through non-homologous end joining (NHEJ) or homologous recombination (HR) is a finely regulated process. Genetic and molecular impedances in either of the pathways affect the fate of the DSB repair and can lead to extended chromosome rearrangements, triggering inheritable genetic instability and tumorigenesis. Recent findings indicated that formation of DNA:RNA hybrids at DSB sites might interfere with DSB repair and DNA damage response. Here we characterized the role of budding yeast DNA:RNA helicase Sen1, orthologous of the human Senataxin/SETX, in DSB repair. Chromatin immunoprecipitation analyses showed that Sen1 is recruited to one HO-induced DSB, contributing to remove DNA:RNA hybrids nearby the lesion. Along with the increased DNA:RNA hybrids, the binding of Rpb3, a subunit of the RNA polymerase II complex, is also prolonged in Sen1-depleted cells. Remarkably, the persistent DNA:RNA hybrids cause abrupt Mre11 and Dna2 dependent resection of the DSB, in the absence of Sen1. By specific genetic backgrounds, we found that Sen1 depletion leads to faster ectopic recombination repair of a DSB and, surprisingly, to elevated NHEJ and chromosome translocations. In line with the increased NHEJ, KU binding at the DSB is also prolonged in Sen1-depleted cells. In summary, our data suggest molecular mechanism through which Sen1 removes DNA:RNA hybrids at DSB in order to prevent non-canonical resection initiation, also limiting KU persistence at DSB and dangerous NHEJ events. We believed that these results in yeast might contribute to explain molecularly pathologic phenotypes, recently described in human cells depleted of Senataxin

Senataxin Ortholog Sen1 Limits DNA:RNA Hybrid Accumulation at DNA Double-Strand Breaks to Control End Resection and Repair Fidelity

Rawal et al. identify a non-canonical mechanism of DSB processing in yeast cells lacking the Senataxin ortholog Sen1 helicase, which strictly depends on DNA:RNA hybrid formation at the break and is associated with unfaithful DSB repair and genome instability. An important but still enigmatic function of DNA:RNA hybrids is their role in DNA double-strand break (DSB) repair. Here, we show that Sen1, the budding yeast ortholog of the human helicase Senataxin, is recruited at an HO endonuclease-induced DSB and limits the local accumulation of DNA:RNA hybrids. In the absence of Sen1, hybrid accumulation proximal to the DSB promotes increased binding of the Ku70-80 (KU) complex at the break site, mutagenic non-homologous end joining (NHEJ), micro-homology-mediated end joining (MMEJ), and chromosome translocations. We also show that homology-directed recombination (HDR) by gene conversion is mostly proficient in sen1 mutants after single DSB. However, in the absence of Sen1, DNA:RNA hybrids, Mre11, and Dna2 initiate resection through a non-canonical mechanism. We propose that this resection mechanism through local DNA:RNA hybrids acts as a backup to prime HDR when canonical pathways are altered, but at the expense of genome integrity