Genetic determinants of micronucleus formation in vivo.

Genomic instability arising from defective responses to DNA damage1 or mitotic chromosomal imbalances2 can lead to the sequestration of DNA in aberrant extranuclear structures called micronuclei (MN). Although MN are a hallmark of ageing and diseases associated with genomic instability, the catalogue of genetic players that regulate the generation of MN remains to be determined. Here we analyse 997 mouse mutant lines, revealing 145 genes whose loss significantly increases (n = 71) or decreases (n = 74) MN formation, including many genes whose orthologues are linked to human disease. We found that mice null for Dscc1, which showed the most significant increase in MN, also displayed a range of phenotypes characteristic of patients with cohesinopathy disorders. After validating the DSCC1-associated MN instability phenotype in human cells, we used genome-wide CRISPR-Cas9 screening to define synthetic lethal and synthetic rescue interactors. We found that the loss of SIRT1 can rescue phenotypes associated with DSCC1 loss in a manner paralleling restoration of protein acetylation of SMC3. Our study reveals factors involved in maintaining genomic stability and shows how this information can be used to identify mechanisms that are relevant to human disease biology1.

GDF15 linked to maternal risk of nausea and vomiting during pregnancy.

GDF15, a hormone acting on the brainstem, has been implicated in the nausea and vomiting of pregnancy, including its most severe form, hyperemesis gravidarum (HG), but a full mechanistic understanding is lacking1-4. Here we report that fetal production of GDF15 and maternal sensitivity to it both contribute substantially to the risk of HG. We confirmed that higher GDF15 levels in maternal blood are associated with vomiting in pregnancy and HG. Using mass spectrometry to detect a naturally labelled GDF15 variant, we demonstrate that the vast majority of GDF15 in the maternal plasma is derived from the feto-placental unit. By studying carriers of rare and common genetic variants, we found that low levels of GDF15 in the non-pregnant state increase the risk of developing HG. Conversely, women with ß-thalassaemia, a condition in which GDF15 levels are chronically high5, report very low levels of nausea and vomiting of pregnancy. In mice, the acute food intake response to a bolus of GDF15 is influenced bi-directionally by prior levels of circulating GDF15 in a manner suggesting that this system is susceptible to desensitization. Our findings support a putative causal role for fetally derived GDF15 in the nausea and vomiting of human pregnancy, with maternal sensitivity, at least partly determined by prepregnancy exposure to the hormone, being a major influence on its severity. They also suggest mechanism-based approaches to the treatment and prevention of HG.

MC3R links nutritional state to childhood growth and the timing of puberty.

The state of somatic energy stores in metazoans is communicated to the brain, which regulates key aspects of behaviour, growth, nutrient partitioning and development1. The central melanocortin system acts through melanocortin 4 receptor (MC4R) to control appetite, food intake and energy expenditure2. Here we present evidence that MC3R regulates the timing of sexual maturation, the rate of linear growth and the accrual of lean mass, which are all energy-sensitive processes. We found that humans who carry loss-of-function mutations in MC3R, including a rare homozygote individual, have a later onset of puberty. Consistent with previous findings in mice, they also had reduced linear growth, lean mass and circulating levels of IGF1. Mice lacking Mc3r had delayed sexual maturation and an insensitivity of reproductive cycle length to nutritional perturbation. The expression of Mc3r is enriched in hypothalamic neurons that control reproduction and growth, and expression increases during postnatal development in a manner that is consistent with a role in the regulation of sexual maturation. These findings suggest a bifurcating model of nutrient sensing by the central melanocortin pathway with signalling through MC4R controlling the acquisition and retention of calories, whereas signalling through MC3R primarily regulates the disposition of calories into growth, lean mass and the timing of sexual maturation.

A distinctive epigenetic ageing profile in human granulosa cells.

STUDY QUESTION: Does women's age affect the DNA methylation (DNAm) profile differently in mural granulosa cells (MGCs) from other somatic cells? SUMMARY ANSWER: Accumulation of epimutations by age and a higher number of age-related differentially methylated regions (DMR) in MGCs were found compared to leukocytes from the same woman, suggesting that the MGCs have a distinctive epigenetic profile. WHAT IS KNOWN ALREADY: The mechanisms underlying the decline in women's fertility from the mid-30s remain to be fully elucidated. The DNAm age of many healthy tissues changes predictably with and follows chronological age, but DNAm age in some reproductive tissues has been shown to depart from chronological age (older: endometrium; younger: cumulus cells, spermatozoa). STUDY DESIGN, SIZE, DURATION: This study is a multicenter cohort study based on retrospective analysis of prospectively collected data and material derived from healthy women undergoing IVF or ICSI treatment following ovarian stimulation with antagonist protocol. One hundred and nineteen women were included from September 2016 to June 2018 from four clinics in Denmark and Sweden. PARTICIPANTS/MATERIALS, SETTING, METHODS: Blood samples were obtained from 118 healthy women with varying ovarian reserve status. MGCs were collected from 63 of the 119 women by isolation from pooled follicles immediately after oocyte retrieval. DNA from leukocytes and MGCs was extracted and analysed with a genome-wide methylation array. Data from the methylation array were processed using the ENmix package. Subsequently, DNAm age was calculated using established and tailored age predictors and DMRs were analysed with the DMRcate package. MAIN RESULTS AND ROLE OF CHANCE: Using established age predictors, DNAm age in MGCs was found to be considerable younger and constant (average: 2.7 years) compared to chronological age (average: 33.9 years). A Granulosa Cell clock able to predict the age of both MGCs (average: 32.4 years) and leukocytes (average: 38.8 years) was successfully developed. MGCs differed from leukocytes in having a higher number of epimutations (P = 0.003) but predicted telomere lengths unaffected by age (Pearson's correlation coefficient = -0.1, P = 0.47). DMRs associated with age (age-DMRs) were identified in MGCs (n = 335) and in leukocytes (n = 1) with a significant enrichment in MGCs for genes involved in RNA processing (45 genes, P = 3.96 × 10-08) and gene expression (152 genes, P = 2.3 × 10-06). The top age-DMRs included the metastable epiallele VTRNA2-1, the DNAm regulator ZFP57 and the anti-Müllerian hormone (AMH) gene. The apparent discordance between different epigenetic measures of age in MGCs suggests that they reflect difference stages in the MGC life cycle. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: No gene expression data were available to associate with the epigenetic findings. The MGCs are collected during ovarian stimulation, which may influence DNAm; however, no correlation between FSH dose and number of epimutations was found. WIDER IMPLICATIONS OF THE FINDINGS: Our findings underline that the somatic compartment of the follicle follows a different methylation trajectory with age than other somatic cells. The higher number of epimutations and age-DMRs in MGCs suggest that their function is affected by age. STUDY FUNDING/COMPETING INTEREST(S): This project is part of ReproUnion collaborative study, co-financed by the European Union, Interreg V ÖKS, the Danish National Research Foundation and the European Research Council. The authors declare no conflict of interest.

Genome-wide association and epidemiological analyses reveal common genetic origins between uterine leiomyomata and endometriosis.

Uterine leiomyomata (UL) are the most common neoplasms of the female reproductive tract and primary cause for hysterectomy, leading to considerable morbidity and high economic burden. Here we conduct a GWAS meta-analysis in 35,474 cases and 267,505 female controls of European ancestry, identifying eight novel genome-wide significant (P < 5 × 10-8) loci, in addition to confirming 21 previously reported loci, including multiple independent signals at 10 loci. Phenotypic stratification of UL by heavy menstrual bleeding in 3409 cases and 199,171 female controls reveals genome-wide significant associations at three of the 29 UL loci: 5p15.33 (TERT), 5q35.2 (FGFR4) and 11q22.3 (ATM). Four loci identified in the meta-analysis are also associated with endometriosis risk; an epidemiological meta-analysis across 402,868 women suggests at least a doubling of risk for UL diagnosis among those with a history of endometriosis. These findings increase our understanding of genetic contribution and biology underlying UL development, and suggest overlapping genetic origins with endometriosis.

Voice break in boys-temporal relations with other pubertal milestones and likely causal effects of BMI.

STUDY QUESTION: How is timing of voice break related to other male pubertal milestones as well as to BMI? SUMMARY ANSWER: We provide a comprehensive temporal analysis of male pubertal milestones, including reproductive hormone dynamics, confirm voice break as a late milestone of male puberty and report a likely causal relationship between higher BMI and earlier age at voice break in men. WHAT IS KNOWN ALREADY: Voice break represents a late pubertal milestone and recalled age at voice break is frequently used in epidemiological studies as a measure of puberty. In contrast, clinical studies use mainly testicular enlargement and/or genital tanner stage as the marker of pubertal onset. However, neither correlation of pubertal milestones nor reproductive hormone dynamics have been assessed in detail previously. Further, although BMI and puberty timing are known to be closely linked, cause and effect between these traits are not known. STUDY DESIGN, SIZE, DURATION: The study included a population-based mixed cross-sectional and longitudinal cohort (2006-2014, COPENHAGEN Puberty Study) of 730 healthy Danish boys. Data for 55 871 male research participants from the 23andMe study were obtained, including genome-wide single nucleotide polymorphism data and age at voice break. PARTICIPANTS/MATERIALS, SETTING, METHODS: We performed a detailed evaluation of pubertal milestones and reproductive hormone levels (study population 1). A Mendelian randomization (MR) approach was used to determine the likely causal link between BMI and timing of voice break (study population 2). MAIN RESULTS AND THE ROLE OF CHANCE: Voice break occurred at mean age 13.6 (95% CI: 13.5-13.8) years. At voice break, mean (95% CI) testosterone levels, LH levels and bi-testicular volume were 10.9 (10.0-11.7) nmol/L, 2.4 (2.2-2.5) IU/L and 24 (23-25) mL, respectively. Voice break correlated moderately strongly with timing of male pubertal milestones, including testicular enlargement, gonadarche, pubarche, sweat odor, axillary hair growth and testosterone above limit of detection (r2 range: 0.43-0.61). Timing of all milestones was negatively associated with age-specific BMI (all P ≤ 0.001). MR analyses inferred likely causal effects of higher BMI on earlier voice break in males (-0.35 years/approximate SD, P < 0.001). LIMITATIONS, REASONS FOR CAUTION: Participation rate of the population-based cohort was 25%. Further, boys that were followed longitudinally were examined approximately every 6 months limiting the time resolution of pubertal milestones. Using adult BMI as exposure instead of prepubertal BMI in the MR analysis and the known inaccuracies of the testosterone immunoassay at low testosterone levels may be further limitations. WIDER IMPLICATIONS OF THE FINDINGS: We provide valuable normative data on the temporal relation of male pubertal milestones. Further, the likely causal relationship between BMI and puberty timing highlights the importance of preventing obesity in childhood. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by Danish Agency for Science, Technology and Innovation (09-067 180); Danish Ministry of the Environment, CeHoS (MST-621-00 065); Capital Region of Denmark (R129-A3966); Ministry of Higher Education and Science (DFF-1331-00 113); Innovation Fund Denmark (InnovationsFonden, 14-2013-4); The International Center for Research and Research Training in Endocrine Disrupting Effects of Male Reproduction and Child Health. B.H., F.R.D., J.R.B.P. and K.K.O. are supported by the Medical Research Council (MC_UU_12015/2). The 23andMe study is supported by the National Human Genome Research Institute of the National Institutes of Health (R44HG006981). Members of the 23andMe Research Team are employees of 23andMe, Inc. and hold stock or stock options in 23andMe. TRIAL REGISTRATION NUMBER: NCT01411527.

Circulating beta-carotene levels and type 2 diabetes-cause or effect?

AIMS/HYPOTHESIS: Circulating beta-carotene levels are inversely associated with risk of type 2 diabetes, but the causal direction of this association is not certain. In this study we used a Mendelian randomisation approach to provide evidence for or against the causal role of the antioxidant vitamin beta-carotene in type 2 diabetes. METHODS: We used a common polymorphism (rs6564851) near the BCMO1 gene, which is strongly associated with circulating beta-carotene levels (p = 2 x 10(-24)), with each G allele associated with a 0.27 standard deviation increase in levels. We used data from the InCHIANTI and Uppsala Longitudinal Study of Adult Men (ULSAM) studies to estimate the association between beta-carotene levels and type 2 diabetes. We next used a triangulation approach to estimate the expected effect of rs6564851 on type 2 diabetes risk and compared this with the observed effect using data from 4549 type 2 diabetes patients and 5579 controls from the Diabetes Genetics Replication And Meta-analysis (DIAGRAM) Consortium. RESULTS: A 0.27 standard deviation increase in beta-carotene levels was associated with an OR of 0.90 (95% CI 0.86-0.95) for type 2 diabetes in the InCHIANTI study. This association was similar to that of the ULSAM study (OR 0.90 [0.84-0.97]). In contrast, there was no association between rs6564851 and type 2 diabetes (OR 0.98 [0.93-1.04], p = 0.58); this effect size was also smaller than that expected, given the known associations between rs6564851 and beta-carotene levels, and the associations between beta-carotene levels and type 2 diabetes. CONCLUSIONS/INTERPRETATION: Our findings in this Mendelian randomisation study are in keeping with randomised controlled trials suggesting that beta-carotene is not causally protective against type 2 diabetes.