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Damaged replication forks activate poly(ADP-ribose) polymerase 1 (PARP1), which catalyses poly(ADP-ribose) (PAR) formation; however, how PARP1 or poly(ADP-ribosyl)ation is involved in the S-phase checkpoint is unknown. Here we show that PAR, supplied by PARP1, interacts with Chk1 via a novel PAR-binding regulatory (PbR) motif in Chk1, independent of ATR and its activity. iPOND studies reveal that Chk1 associates readily with the unperturbed replication fork and that PAR is required for efficient retention of Chk1 and phosphorylated Chk1 at the fork. A PbR mutation, which disrupts PAR binding, but not the interaction with its partners Claspin or BRCA1, impairs Chk1 and the S-phase checkpoint activation, and mirrors Chk1 knockdown-induced hypersensitivity to fork poisoning. We find that long chains, but not short chains, of PAR stimulate Chk1 kinase activity. Collectively, we disclose a previously unrecognized mechanism of the S-phase checkpoint by PAR metabolism that modulates Chk1 activity at the replication fork.

Oxysterol nuclear receptors liver X receptor (LXR)alpha and LXRbeta are known to regulate lipid homeostasis in cells exposed to high amounts of cholesterol and/or fatty acids. In order to elucidate the specific and redundant roles of the LXRs in the testis, we explored the reproductive phenotypes of mice deficient of LXRalpha, LXRbeta, and both, of which only the lxralpha;beta-/- mice are infertile by 5 months of age. We demonstrate that LXRalpha-deficient mice had lower levels of testicular testosterone that correlated with a higher apoptotic rate of the germ cells. LXRbeta-deficient mice showed increased lipid accumulation in the Sertoli cells and a lower proliferation rate of the germ cells. In lxralpha;beta-/- mice, fatty acid metabolism was affected through a decrease of srebp1c and increase in scd1 mRNA expression. The retinoid acid signaling pathway was also altered in lxralpha;beta-/- mice, with a higher accumulation of all-trans retinoid receptor alpha, all-trans retinoid receptor beta, and retinoic aldehyde dehydrogenase-2 mRNA. Combination of these alterations might explain the deleterious phenotype of infertility observed only in lxralpha;beta-/- mice, even though lipid homeostasis seemed to be first altered. Wild-type mice treated with a specific LXR agonist showed an increase of testosterone production involving both LXR isoforms. Altogether, these data identify new roles of each LXR, collaborating to maintain both integrity and functions of the testis.

Epidemiological and experimental studies have shown that the peri-conception period, pregnancy, and infancy are windows of particular sensibility to environmental clues which influence lifelong trajectories across health and disease. Nutrition, stress, and toxins induce epigenetic marks that control long-term gene expression patterns and can be transmitted transgenerationally. Chronic diseases of adulthood such as hypertension, diabetes, and obesity thus have early, developmental origins in the perinatal period. The early epigenome, in interaction with other actors such as the microbiome, add powerful layers of diversity to the biological predisposition generated by the genome. Such “programming” is a normal, adaptive component of development, including in normal pregnancies and births. However, perinatal disease, either maternal (such as pre-eclampsia, ges­tational diabetes, or inflammatory disease) or fetal, and neonatal diseases (such as intrauterine growth restriction and preterm birth) are major conditions of altered programming, translated into an increased risk for chronic disease in these patients when they reach adulthood. Early prevention, optimal perinatal nutrition, and specific follow-up measures are key factors in the early preservation of long-term health.

BackgroundTesticular germ cell tumours are the most frequent cancer of young men with an increasing incidence all over the world. Pathogenesis and reasons of this increase remain unknown but epidemiological and clinical data have suggested that fetal exposure to environmental endocrine disruptors (EEDs) with estrogenic effects, could participate to testicular germ cell carcinogenesis. However, these EEDs (like bisphenol A) are often weak ligands for classical nuclear estrogen receptors. Several research groups recently showed that the non classical membrane G-protein coupled estrogen receptor (GPER/GPR30) mediates the effects of estrogens and several xenoestrogens through rapid non genomic activation of signal transduction pathways in various human estrogen dependent cancer cells (breast, ovary, endometrium). The aim of this study was to demonstrate that GPER was overexpressed in testicular tumours and was able to trigger JKT-1 seminoma cell proliferation.ResultsWe report here for the first time a complete morphological and functional characterization of GPER in normal and malignant human testicular germ cells. In normal adult human testes, GPER was expressed by somatic (Sertoli cells) and germ cells (spermatogonia and spermatocytes). GPER was exclusively overexpressed in seminomas, the most frequent testicular germ cell cancer, localized at the cell membrane and triggered a proliferative effect on JKT-1 cells in vitro, which was completely abolished by G15 (a GPER selective antagonist) and by siRNA invalidation.ConclusionThese results demonstrate that GPER is expressed by human normal adult testicular germ cells, specifically overexpressed in seminoma tumours and able to trigger seminoma cell proliferation in vitro. It should therefore be considered rather than classical ERs when xeno-estrogens or other endocrine disruptors are assessed in testicular germ cell cancers. It may also represent a prognosis marker and/or a therapeutic target for seminomas.

Abstract:Individuals born after intrauterine growth restriction (IUGR) have an increased risk of perinatal morbidity/mortality, and those who survive face long-term consequences such as cardiovascular-related diseases, including systemic hypertension, atherosclerosis, coronary heart disease, and chronic kidney disease. In addition to the demonstrated long-term effects of decreased nephron endowment and hyperactivity of the hypothalamic-pituitary-adrenal axis, individuals born after IUGR also exhibit early alterations in vascular structure and function, which have been identified as key factors of the development of cardiovascular-related diseases. The endothelium plays a Cambridge University Press Developmental Origins of Health and Disease -For Peer Review

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Different studies have pointed out that developmental exposure to environmental endocrine disruptors can induce long-term testicular germ cell death probably through epigenetic mechanisms. By using a model of early neonatal post-natal day (PND) 1 to 5 exposure of male rats to a xenoestrogen, estradiol benzoate (EB), we investigated the role of microRNA and DNA methyltransferases (DNMT) on the developmental effects of EB on the adult germ cell death process. Neonatal exposure to EB induced adult germ cell apoptosis together with a dose-dependent increase in miR-29a, miR-29b, and miR-29c expression. Increased miR-29 expression resulted in a decrease in DNMT1, DNMT3a, and DNMT3b and antiapoptotic myeloid cell leukemia sequence 1 (Mcl-1) protein levels as shown in 1) germ cells of adult rats exposed neonatally to EB and 2) in spermatogonial GC-1 transfected with miR-29. The DNMT decrease was associated with a concomitant increase in transcript levels of DNA methylation target genes, such as L1td1-1 ORF1 and ORF2, Cdkn2a, and Gstp1, in correlation with their pattern of methylation. Finally, GC-1 cell lines transfection with miR-29a, miR-29b, or miR-29c undergo apoptosis evidenced by Annexin-V expression. Together, the increased miR-29 with a subsequent reduction in DNMT and Mcl-1 protein levels may represent a basis of explanation for the adult expression of the germ cell apoptosis phenotype. These observations suggest that the increased expression of the "apoptomir" miR-29 family represents the upstream mechanism identified until now that is involved in adult germ cell apoptosis induced by a neonatal hormonal disruption.