Characteristics and homogeneity of N6-methylation in human genomes.

A novel DNA modification, N-6 methylated deoxyadenosine (m6dA), has recently been discovered in eukaryotic genomes. Despite its low abundance in eukaryotes, m6dA is implicated in human diseases such as cancer. It is therefore important to precisely identify and characterize m6dA in the human genome. Here, we identify m6dA sites at nucleotide level, in different human cells, genome wide. We compare m6dA features between distinct human cells and identify m6dA characteristics in human genomes. Our data demonstrates for the first time that despite low m6dA abundance, the m6dA mark does often occur consistently at the same genomic location within a given human cell type, demonstrating m6dA homogeneity. We further show, for the first time, higher levels of m6dA homogeneity within one chromosome. Most m6dA are found on a single chromosome from a diploid sample, suggesting inheritance. Our transcriptome analysis not only indicates that human genes with m6dA are associated with higher RNA transcript levels but identifies allele-specific gene transcripts showing haplotype-specific m6dA methylation, which are implicated in different biological functions. Our analyses demonstrate the precision and consistency by which the m6dA mark occurs within the human genome, suggesting that m6dA marks are precisely inherited in humans

Histone H3 lysine 9 trimethylation is required for suppressing the expression of an embryonically activated retrotransposon in Xenopus laevis.

Transposable elements in the genome are generally silenced in differentiated somatic cells. However, increasing evidence indicates that some of them are actively transcribed in early embryos and the proper regulation of retrotransposon expression is essential for normal development. Although their developmentally regulated expression has been shown, the mechanisms controlling retrotransposon expression in early embryos are still not well understood. Here, we observe a dynamic expression pattern of retrotransposons with three out of ten examined retrotransposons (1a11, λ-olt 2-1 and xretpos(L)) being transcribed solely during early embryonic development. We also identified a transcript that contains the long terminal repeat (LTR) of λ-olt 2-1 and shows a similar expression pattern to λ-olt 2-1 in early Xenopus embryos. All three retrotransposons are transcribed by RNA polymerase II. Although their expression levels decline during development, the LTRs are marked by histone H3 lysine 4 trimethylation. Furthermore, retrotransposons, especially λ-olt 2-1, are enriched with histone H3 lysine 9 trimethylation (H3K9me3) when their expression is repressed. Overexpression of lysine-specific demethylase 4d removes H3K9me3 marks from Xenopus embryos and inhibits the repression of λ-olt 2-1 after gastrulation. Thus, our study shows that H3K9me3 is important for silencing the developmentally regulated retrotransposon in Xenopus laevis.Gurdon laboratory is supported by grants from the Wellcome Trust (RG69899) and MRC to J.B.GThis is the final version of the article. It first appeared from Nature Publishing Group via http://dx.doi.org/10.1038/srep1423

A Unique Gene Regulatory Network Resets the Human Germline Epigenome for Development.

Resetting of the epigenome in human primordial germ cells (hPGCs) is critical for development. We show that the transcriptional program of hPGCs is distinct from that in mice, with co-expression of somatic specifiers and naive pluripotency genes TFCP2L1 and KLF4. This unique gene regulatory network, established by SOX17 and BLIMP1, drives comprehensive germline DNA demethylation by repressing DNA methylation pathways and activating TET-mediated hydroxymethylation. Base-resolution methylome analysis reveals progressive DNA demethylation to basal levels in week 5-7 in vivo hPGCs. Concurrently, hPGCs undergo chromatin reorganization, X reactivation, and imprint erasure. Despite global hypomethylation, evolutionarily young and potentially hazardous retroelements, like SVA, remain methylated. Remarkably, some loci associated with metabolic and neurological disorders are also resistant to DNA demethylation, revealing potential for transgenerational epigenetic inheritance that may have phenotypic consequences. We provide comprehensive insight on early human germline transcriptional network and epigenetic reprogramming that subsequently impacts human development and disease.W.C.C.T is supported by Croucher Foundation and Cambridge Trust. P.F.C.is a Wellcome Trust Senior Fellow in Clinical Science (101876/Z/13/Z), and a UK NIHR Senior Investigator with additional support from the Wellcome Trust Centre for Mitochondrial Research (096919Z/11/Z). M.A.S. is supported by HFSP and Wellcome Trust Investigator Award.This is the final version of the article. It first appeared from Elsevier via http://dx.doi.org/10.1016/j.cell.2015.04.05

Sperm is epigenetically programmed to regulate gene transcription in embryos.

For a long time, it has been assumed that the only role of sperm at fertilization is to introduce the male genome into the egg. Recently, ideas have emerged that the epigenetic state of the sperm nucleus could influence transcription in the embryo. However, conflicting reports have challenged the existence of epigenetic marks on sperm genes, and there are no functional tests supporting the role of sperm epigenetic marking on embryonic gene expression. Here, we show that sperm is epigenetically programmed to regulate embryonic gene expression. By comparing the development of sperm- and spermatid-derived frog embryos, we show that the programming of sperm for successful development relates to its ability to regulate transcription of a set of developmentally important genes. During spermatid maturation into sperm, these genes lose H3K4me2/3 and retain H3K27me3 marks. Experimental removal of these epigenetic marks at fertilization de-regulates gene expression in the resulting embryos in a paternal chromatin-dependent manner. This demonstrates that epigenetic instructions delivered by the sperm at fertilization are required for correct regulation of gene expression in the future embryos. The epigenetic mechanisms of developmental programming revealed here are likely to relate to the mechanisms involved in transgenerational transmission of acquired traits. Understanding how parental experience can influence development of the progeny has broad potential for improving human health.We thank: T. Jenuwein and N. Shukeir for anti-H3K27me3 antibody; A. Bannister, J. Ahringer and E. Miska for comments on the manuscript; Gurdon group members for reading the manuscript; The International Xenopus laevis Genome Project Consortium (the Harland, Rokhsar, Taira labs and others) for providing unpublished genome and gene annotation information. M.T. is supported by WT089613 and by MR/K011022/1. V.G. and P.Z. are funded by AICR 10-0908. A.S. is supported by MR/K011022/1. K.M. is a Research Fellow at Wolfson College and is supported by the Herchel Smith Postdoctoral Fellowship. E.M.M. is supported by National Institutes of Health, National Science Foundation, Cancer Prevention Research Institute of Texas, and the Welch Foundation (F1515). J.J. and J.B.G. are supported by WT101050/Z/13/Z. S.E. acknowledges Boehringer Ingelheim Fond fellowship. A.H.F.M.P. is supported by the Swiss National Science Foundation (31003A_125386) and the Novartis Research Foundation. All members of the Gurdon Institute acknowledge the core support provided by CRUK C6946/A14492 and WT092096.This is the final version of the article. It first appeared from Cold Spring Harbor Laboratory Press via https://doi.org/10.1101/gr.201541.11

Mechanical cell competition kills cells via induction of lethal p53 levels.

Cell competition is a quality control mechanism that eliminates unfit cells. How cells compete is poorly understood, but it is generally accepted that molecular exchange between cells signals elimination of unfit cells. Here we report an orthogonal mechanism of cell competition, whereby cells compete through mechanical insults. We show that MDCK cells silenced for the polarity gene scribble (scrib(KD)) are hypersensitive to compaction, that interaction with wild-type cells causes their compaction and that crowding is sufficient for scrib(KD) cell elimination. Importantly, we show that elevation of the tumour suppressor p53 is necessary and sufficient for crowding hypersensitivity. Compaction, via activation of Rho-associated kinase (ROCK) and the stress kinase p38, leads to further p53 elevation, causing cell death. Thus, in addition to molecules, cells use mechanical means to compete. Given the involvement of p53, compaction hypersensitivity may be widespread among damaged cells and offers an additional route to eliminate unfit cells.This work was supported by a Cancer Research UK Programme Grant (EP and LW A12460), a Royal Society University Research fellowship to EP (UF0905080), a Wellcome Trust PhD studentship to I.K, a Cambridge Cancer Centre PhD studentship to MG and Core grant funding from the Wellcome Trust (092096) and CRUK (C6946/A14492).This is the final version of the article. It first appeared from Nature Publishing Group via https://doi.org/10.1038/ncomms1137

RNF43/ZNRF3 loss predisposes to hepatocellular-carcinoma by impairing liver regeneration and altering the liver lipid metabolic ground-state.

RNF43/ZNRF3 negatively regulate WNT signalling. Both genes are mutated in several types of cancers, however, their contribution to liver disease is unknown. Here we describe that hepatocyte-specific loss of Rnf43/Znrf3 results in steatohepatitis and in increase in unsaturated lipids, in the absence of dietary fat supplementation. Upon injury, Rnf43/Znrf3 deletion results in defective hepatocyte regeneration and liver cancer, caused by an imbalance between differentiation/proliferation. Using hepatocyte-, hepatoblast- and ductal cell-derived organoids we demonstrate that the differentiation defects and lipid alterations are, in part, cell-autonomous. Interestingly, ZNRF3 mutant liver cancer patients present poorer prognosis, altered hepatic lipid metabolism and steatohepatitis/NASH signatures. Our results imply that RNF43/ZNRF3 predispose to liver cancer by controlling the proliferative/differentiation and lipid metabolic state of hepatocytes. Both mechanisms combined facilitate the progression towards malignancy. Our findings might aid on the management of those RNF43/ZNRF3 mutated individuals at risk of developing fatty liver and/or liver cancer

Outcomes and Complications With Off-Label Use of Drug-Eluting Stents Results From the STENT (Strategic Transcatheter Evaluation of New Therapies) Group

ObjectivesThis study evaluates outcomes and complications in patients treated with drug-eluting stents (DES) for “off-label” indications.BackgroundDrug-eluting stents have been effective in randomized trials, but their safety and efficacy for off-label indications has not been well studied.MethodsThe STENT (Strategic Transcatheter Evaluation of New Therapies) Registry is the largest multicenter U.S. registry evaluating outcomes of DES. Off-label indications included ostial, left main, long, bifurcation, and in-stent restenotic lesions, saphenous vein grafts, chronic total occlusions, small or large vessels, multilesion or multivessel percutaneous coronary interventions, and ST-segment elevation myocardial infarction. Outcomes were adjusted using Cox proportional hazards regression and propensity analyses.ResultsDrug-eluting stents were used in an off-label manner in 59% of patients. The patients who received off-label treatment were more often male, had a higher incidence of prior infarction and bypass surgery, and lower ejection fractions. Off-label versus “on-label” use of DES was associated with higher rates of death, myocardial infarction, target vessel revascularization, major adverse cardiac events, and stent thrombosis at 9 months and 2 years. Off-label use of DES compared with off-label use of bare-metal stents (BMS) had lower rates of death, myocardial infarction, target vessel revascularization, and major adverse cardiac events at 9 months and 2 years and lower rates of stent thrombosis at 9 months.ConclusionsOff-label use of DES is associated with higher event rates compared with on-label use of DES, which is consistent with a higher risk clinical and lesion profile. However, event rates with off-label use of DES are lower compared with off-label use of BMS. Pending results from randomized trials, our data support the use of DES for off-label indications in selected patients

To what extent is behaviour a problem in English schools?:Exploring the scale and prevalence of deficits in classroom climate

The working atmosphere in the classroom is an important variable in the process of education in schools, with several studies suggesting that classroom climate is an important influence on pupil attainment. There are wide differences in the extent to which classroom climate is considered to be a problem in English schools. Some ‘official’ reports suggest that behaviour in schools is ‘satisfactory or better’ in the vast majority of schools; other sources have pointed to behaviour being a serious and widespread problem. The paper details four studies conducted over the past decade which aimed to explore these disparities. The aim of the research was to gain a more accurate insight into the extent to which deficits in classroom climate limit educational attainment and equality of educational opportunity in English schools. The findings question the suggestion that behaviour is satisfactory or better in 99.7% of English schools and the concluding section suggests ways in which deficits in classroom climate might be addressed. Although the study is limited to classrooms in England, OECD studies suggest that deficits in the working atmosphere in classrooms occur in many countries. The study therefore has potential relevance for education systems in other countries

MS AHI1 genetic risk promotes IFNγ+ CD4+ T cells

Objective: To study the influence of the Abelson helper integration site 1 (AHI1) locus associated with MS susceptibility on CD4+ T cell function. Methods: We characterized the chromatin state of T cells in the MS-associated AHI1 linkage disequilibrium (LD) block. The expression and the role of the AHI1 variant were examined in T cells from genotyped healthy subjects who were recruited from the PhenoGenetic Project, and the function of AHI1 was explored using T cells from Ahi1 knockout mice. Results: Chromatin state analysis reveals that the LD block containing rs4896153, which is robustly associated with MS susceptibility (odds ratio 1.15, p = 1.65 × 10−13), overlaps with strong enhancer regions that are present in human naive and memory CD4+ T cells. Relative to the rs4896153A protective allele, the rs4896153T susceptibility allele is associated with decreased AHI1 mRNA expression, specifically in naive CD4+ T cells (p = 1.73 × 10−74, n = 213), and we replicate this effect in an independent set of subjects (p = 2.5 × 10−9, n = 32). Functional studies then showed that the rs4896153T risk variant and the subsequent decreased AHI1 expression were associated with reduced CD4+ T cell proliferation and a specific differentiation into interferon gamma (IFNγ)–positive T cells when compared with the protective rs4896153A allele. This T cell phenotype was also observed in murine CD4+ T cells with genetic deletion of Ahi1. Conclusions: Our findings suggest that the effect of the AHI1 genetic risk for MS is mediated, in part, by enhancing the development of proinflammatory IFNγ+ T cells that have previously been implicated in MS and its mouse models