Life Sciences Research for Lifelong Health

Publications

The Babraham Institute Publications database contains details of all publications resulting from our research groups and scientific services.

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Title / Authors / Details Open Access Download

Modeling Meets Metabolomics-The WormJam Consensus Model as Basis for Metabolic Studies in the Model Organism .
Witting M, Hastings J, Rodriguez N, Joshi CJ, Hattwell JPN, Ebert PR, van Weeghel M, Gao AW, Wakelam MJO, Houtkooper RH, Mains A, Le Novère N, Sadykoff S, Schroeder F, Lewis NE, Schirra HJ, Kaleta C, Casanueva O

Metabolism is one of the attributes of life and supplies energy and building blocks to organisms. Therefore, understanding metabolism is crucial for the understanding of complex biological phenomena. Despite having been in the focus of research for centuries, our picture of metabolism is still incomplete. Metabolomics, the systematic analysis of all small molecules in a biological system, aims to close this gap. In order to facilitate such investigations a blueprint of the metabolic network is required. Recently, several metabolic network reconstructions for the model organism have been published, each having unique features. We have established the WormJam Community to merge and reconcile these (and other unpublished models) into a single consensus metabolic reconstruction. In a series of workshops and annotation seminars this model was refined with manual correction of incorrect assignments, metabolite structure and identifier curation as well as addition of new pathways. The WormJam consensus metabolic reconstruction represents a rich data source not only for network-based approaches like flux balance analysis, but also for metabolomics, as it includes a database of metabolites present in , which can be used for annotation. Here we present the process of model merging, correction and curation and give a detailed overview of the model. In the future it is intended to expand the model toward different tissues and put special emphasizes on lipid metabolism and secondary metabolism including ascaroside metabolism in accordance to their central role in physiology.

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Frontiers in molecular biosciences, 5, 2296-889X, 96, 2018

PMID: 30488036

Trophoblast organoids as a model for maternal-fetal interactions during human placentation.
Turco MY, Gardner L, Kay RG, Hamilton RS, Prater M, Hollinshead MS, McWhinnie A, Esposito L, Fernando R, Skelton H, Reimann F, Gribble FM, Sharkey A, Marsh SGE, O'Rahilly S, Hemberger M, Burton GJ, Moffett A

The placenta is the extraembryonic organ that supports the fetus during intrauterine life. Although placental dysfunction results in major disorders of pregnancy with immediate and lifelong consequences for the mother and child, our knowledge of the human placenta is limited owing to a lack of functional experimental models. After implantation, the trophectoderm of the blastocyst rapidly proliferates and generates the trophoblast, the unique cell type of the placenta. In vivo, proliferative villous cytotrophoblast cells differentiate into two main sub-populations: syncytiotrophoblast, the multinucleated epithelium of the villi responsible for nutrient exchange and hormone production, and extravillous trophoblast cells, which anchor the placenta to the maternal decidua and transform the maternal spiral arteries. Here we describe the generation of long-term, genetically stable organoid cultures of trophoblast that can differentiate into both syncytiotrophoblast and extravillous trophoblast. We used human leukocyte antigen (HLA) typing to confirm that the organoids were derived from the fetus, and verified their identities against four trophoblast-specific criteria. The cultures organize into villous-like structures, and we detected the secretion of placental-specific peptides and hormones, including human chorionic gonadotropin (hCG), growth differentiation factor 15 (GDF15) and pregnancy-specific glycoprotein (PSG) by mass spectrometry. The organoids also differentiate into HLA-G extravillous trophoblast cells, which vigorously invade in three-dimensional cultures. Analysis of the methylome reveals that the organoids closely resemble normal first trimester placentas. This organoid model will be transformative for studying human placental development and for investigating trophoblast interactions with the local and systemic maternal environment.

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Nature, , 1476-4687, , 2018

PMID: 30487605

Hepatic gene body hypermethylation is a shared epigenetic signature of murine longevity.
Hahn O, Stubbs TM, Reik W, Grönke S, Beyer A, Partridge L

Dietary, pharmacological and genetic interventions can extend health- and lifespan in diverse mammalian species. DNA methylation has been implicated in mediating the beneficial effects of these interventions; methylation patterns deteriorate during ageing, and this is prevented by lifespan-extending interventions. However, whether these interventions also actively shape the epigenome, and whether such epigenetic reprogramming contributes to improved health at old age, remains underexplored. We analysed published, whole-genome, BS-seq data sets from mouse liver to explore DNA methylation patterns in aged mice in response to three lifespan-extending interventions: dietary restriction (DR), reduced TOR signaling (rapamycin), and reduced growth (Ames dwarf mice). Dwarf mice show enhanced DNA hypermethylation in the body of key genes in lipid biosynthesis, cell proliferation and somatotropic signaling, which strongly correlates with the pattern of transcriptional repression. Remarkably, DR causes a similar hypermethylation in lipid biosynthesis genes, while rapamycin treatment increases methylation signatures in genes coding for growth factor and growth hormone receptors. Shared changes of DNA methylation were restricted to hypermethylated regions, and they were not merely a consequence of slowed ageing, thus suggesting an active mechanism driving their formation. By comparing the overlap in ageing-independent hypermethylated patterns between all three interventions, we identified four regions, which, independent of genetic background or gender, may serve as novel biomarkers for longevity-extending interventions. In summary, we identified gene body hypermethylation as a novel and partly conserved signature of lifespan-extending interventions in mouse, highlighting epigenetic reprogramming as a possible intervention to improve health at old age.

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PLoS genetics, 14, 1553-7404, e1007766, 2018

PMID: 30462643

Open Access

Single cell transcriptome analysis of human, marmoset and mouse embryos reveals common and divergent features of preimplantation development.
Boroviak T, Stirparo GG, Dietmann S, Hernando-Herraez I, Mohammed H, Reik W, Smith A, Sasaki E, Nichols J, Bertone P

The mouse embryo is the canonical model for mammalian preimplantation development. Recent advances in single cell profiling allow detailed analysis of embryogenesis in other eutherian species, including human, to distinguish conserved from divergent regulatory programs and signalling pathways in the rodent paradigm. Here, we identify and compare transcriptional features of human, marmoset and mouse embryos by single cell RNA-seq. Zygotic genome activation correlates with the presence of polycomb repressive complexes in all three species, while ribosome biogenesis emerges as a predominant attribute in primate embryos, supporting prolonged translation of maternally deposited RNAs. We find that transposable element expression signatures are species, stage and lineage specific. The pluripotency network in the primate epiblast lacks certain regulators that are operative in mouse, but encompasses WNT components and genes associated with trophoblast specification. Sequential activation of GATA6, SOX17 and GATA4 markers of primitive endoderm identity is conserved in primates. Unexpectedly, OTX2 is also associated with primitive endoderm specification in human and non-human primate blastocysts. Our cross-species analysis demarcates both conserved and primate-specific features of preimplantation development, and underscores the molecular adaptability of early mammalian embryogenesis.

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Development (Cambridge, England), 145, 1477-9129, , 2018

PMID: 30413530

Open Access

5-Formylcytosine organizes nucleosomes and forms Schiff base interactions with histones in mouse embryonic stem cells.
Raiber EA, Portella G, Martínez Cuesta S, Hardisty R, Murat P, Li Z, Iurlaro M, Dean W, Spindel J, Beraldi D, Liu Z, Dawson MA, Reik W, Balasubramanian S

Nucleosomes are the basic unit of chromatin that help the packaging of genetic material while controlling access to the genetic information. The underlying DNA sequence, together with transcription-associated proteins and chromatin remodelling complexes, are important factors that influence the organization of nucleosomes. Here, we show that the naturally occurring DNA modification, 5-formylcytosine (5fC) is linked to tissue-specific nucleosome organization. Our study reveals that 5fC is associated with increased nucleosome occupancy in vitro and in vivo. We demonstrate that 5fC-associated nucleosomes at enhancers in the mammalian hindbrain and heart are linked to elevated gene expression. Our study also reveals the formation of a reversible-covalent Schiff base linkage between lysines of histone proteins and 5fC within nucleosomes in a cellular environment. We define their specific genomic loci in mouse embryonic stem cells and look into the biological consequences of these DNA-histone Schiff base sites. Collectively, our findings show that 5fC is a determinant of nucleosome organization and plays a role in establishing distinct regulatory regions that control transcription.

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Nature chemistry, , 1755-4349, , 2018

PMID: 30349137

Regulation of Placental Development and Its Impact on Fetal Growth-New Insights From Mouse Models.
Woods L, Perez-Garcia V, Hemberger M

The placenta is the chief regulator of nutrient supply to the growing embryo during gestation. As such, adequate placental function is instrumental for developmental progression throughout intrauterine development. One of the most common complications during pregnancy is insufficient growth of the fetus, a problem termed intrauterine growth restriction (IUGR) that is most frequently rooted in a malfunctional placenta. Together with conventional gene targeting approaches, recent advances in screening mouse mutants for placental defects, combined with the ability to rapidly induce mutations and by CRISPR-Cas9 technology, has provided new insights into the contribution of the genome to normal placental development. Most importantly, these data have demonstrated that far more genes are required for normal placentation than previously appreciated. Here, we provide a summary of common types of placental defects in established mouse mutants, which will help us gain a better understanding of the genes impacting on human placentation. Based on a recent mouse mutant screen, we then provide examples on how these data can be mined to identify novel molecular hubs that may be critical for placental development. Given the close association between placental defects and abnormal cardiovascular and brain development, these functional nodes may also shed light onto the etiology of birth defects that co-occur with placental malformations. Taken together, recent insights into the regulation of mouse placental development have opened up new avenues for research that will promote the study of human pregnancy conditions, notably those based on defects in placentation that underlie the most common pregnancy pathologies such as IUGR and pre-eclampsia.

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Frontiers in endocrinology, 9, 1664-2392, 570, 2018

PMID: 30319550

Open Access

Transgenerational transmission of hedonic behaviors and metabolic phenotypes induced by maternal overnutrition.
Sarker G, Berrens R, von Arx J, Pelczar P, Reik W, Wolfrum C, Peleg-Raibstein D

Maternal overnutrition has been associated with increased susceptibility to develop obesity and neurological disorders later in life. Most epidemiological as well as experimental studies have focused on the metabolic consequences across generations following an early developmental nutritional insult. Recently, it has been shown that maternal high-fat diet (HFD) affects third-generation female body mass via the paternal lineage. We showed here that the offspring born to HFD ancestors displayed addictive-like behaviors as well as obesity and insulin resistance up to the third generation in the absence of any further exposure to HFD. These findings, implicate that the male germ line is a major player in transferring phenotypic traits. These behavioral and physiological alterations were paralleled by reduced striatal dopamine levels and increased dopamine 2 receptor density. Interestingly, by the third generation a clear gender segregation emerged, where females showed addictive-like behaviors while male HFD offspring showed an obesogenic phenotype. However, methylome profiling of F1 and F2 sperm revealed no significant difference between the offspring groups, suggesting that the sperm methylome might not be the major carrier for the transmission of the phenotypes observed in our mouse model. Together, our study for the first time demonstrates that maternal HFD insult causes sustained alterations of the mesolimbic dopaminergic system suggestive of a predisposition to develop obesity and addictive-like behaviors across multiple generations.

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Translational psychiatry, 8, 2158-3188, 195, 2018

PMID: 30315171

Open Access

Divergent wiring of repressive and active chromatin interactions between mouse embryonic and trophoblast lineages.
Schoenfelder S, Mifsud B, Senner CE, Todd CD, Chrysanthou S, Darbo E, Hemberger M, Branco MR

The establishment of the embryonic and trophoblast lineages is a developmental decision underpinned by dramatic differences in the epigenetic landscape of the two compartments. However, it remains unknown how epigenetic information and transcription factor networks map to the 3D arrangement of the genome, which in turn may mediate transcriptional divergence between the two cell lineages. Here, we perform promoter capture Hi-C experiments in mouse trophoblast (TSC) and embryonic (ESC) stem cells to understand how chromatin conformation relates to cell-specific transcriptional programmes. We find that key TSC genes that are kept repressed in ESCs exhibit interactions between H3K27me3-marked regions in ESCs that depend on Polycomb repressive complex 1. Interactions that are prominent in TSCs are enriched for enhancer-gene contacts involving key TSC transcription factors, as well as TET1, which helps to maintain the expression of TSC-relevant genes. Our work shows that the first developmental cell fate decision results in distinct chromatin conformation patterns establishing lineage-specific contexts involving both repressive and active interactions.

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Nature communications, 9, 2041-1723, 4189, 2018

PMID: 30305613

Open Access

Tri-methylation of histone H3 lysine 4 facilitates gene expression in ageing cells.
Cruz C, Della Rosa M, Krueger C, Gao Q, Horkai D, King M, Field L, Houseley J

Transcription of protein coding genes is accompanied by recruitment of COMPASS to promoter-proximal chromatin, which methylates histone H3 lysine 4 (H3K4) to form H3K4me1, H3K4me2 and H3K4me3. Here, we determine the importance of COMPASS in maintaining gene expression across lifespan in budding yeast. We find that COMPASS mutations reduce replicative lifespan and cause expression defects in almost 500 genes. Although H3K4 methylation is reported to act primarily in gene repression, particularly in yeast, repressive functions are progressively lost with age while hundreds of genes become dependent on H3K4me3 for full expression. Basal and inducible expression of these genes is also impaired in young cells lacking COMPASS components Swd1 or Spp1. Gene induction during ageing is associated with increasing promoter H3K4me3, but H3K4me3 also accumulates in non-promoter regions and the ribosomal DNA. Our results provide clear evidence that H3K4me3 is required to maintain normal expression of many genes across organismal lifespan.

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eLife, 7, 2050-084X, , 2018

PMID: 30274593

Open Access

Genome organization and chromatin analysis identify transcriptional downregulation of insulin-like growth factor signaling as a hallmark of aging in developing B cells.
Koohy H, Bolland DJ, Matheson LS, Schoenfelder S, Stellato C, Dimond A, Várnai C, Chovanec P, Chessa T, Denizot J, Manzano Garcia R, Wingett SW, Freire-Pritchett P, Nagano T, Hawkins P, Stephens L, Elderkin S, Spivakov M, Fraser P, Corcoran AE, Varga-Weisz PD

Aging is characterized by loss of function of the adaptive immune system, but the underlying causes are poorly understood. To assess the molecular effects of aging on B cell development, we profiled gene expression and chromatin features genome-wide, including histone modifications and chromosome conformation, in bone marrow pro-B and pre-B cells from young and aged mice.

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Genome biology, 19, 1474-760X, 126, 2018

PMID: 30180872

Open Access

Genome-Scale Oscillations in DNA Methylation during Exit from Pluripotency.
Rulands S, Lee HJ, Clark SJ, Angermueller C, Smallwood SA, Krueger F, Mohammed H, Dean W, Nichols J, Rugg-Gunn P, Kelsey G, Stegle O, Simons BD, Reik W

Pluripotency is accompanied by the erasure of parental epigenetic memory, with naïve pluripotent cells exhibiting global DNA hypomethylation both in vitro and in vivo. Exit from pluripotency and priming for differentiation into somatic lineages is associated with genome-wide de novo DNA methylation. We show that during this phase, co-expression of enzymes required for DNA methylation turnover, DNMT3s and TETs, promotes cell-to-cell variability in this epigenetic mark. Using a combination of single-cell sequencing and quantitative biophysical modeling, we show that this variability is associated with coherent, genome-scale oscillations in DNA methylation with an amplitude dependent on CpG density. Analysis of parallel single-cell transcriptional and epigenetic profiling provides evidence for oscillatory dynamics both in vitro and in vivo. These observations provide insights into the emergence of epigenetic heterogeneity during early embryo development, indicating that dynamic changes in DNA methylation might influence early cell fate decisions.

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Cell systems, , 2405-4712, , 2018

PMID: 30031774

Open Access

Promoter Capture Hi-C: High-resolution, Genome-wide Profiling of Promoter Interactions.
Schoenfelder S, Javierre BM, Furlan-Magaril M, Wingett SW, Fraser P

The three-dimensional organization of the genome is linked to its function. For example, regulatory elements such as transcriptional enhancers control the spatio-temporal expression of their target genes through physical contact, often bridging considerable (in some cases hundreds of kilobases) genomic distances and bypassing nearby genes. The human genome harbors an estimated one million enhancers, the vast majority of which have unknown gene targets. Assigning distal regulatory regions to their target genes is thus crucial to understand gene expression control. We developed Promoter Capture Hi-C (PCHi-C) to enable the genome-wide detection of distal promoter-interacting regions (PIRs), for all promoters in a single experiment. In PCHi-C, highly complex Hi-C libraries are specifically enriched for promoter sequences through in-solution hybrid selection with thousands of biotinylated RNA baits complementary to the ends of all promoter-containing restriction fragments. The aim is to then pull-down promoter sequences and their frequent interaction partners such as enhancers and other potential regulatory elements. After high-throughput paired-end sequencing, a statistical test is applied to each promoter-ligated restriction fragment to identify significant PIRs at the restriction fragment level. We have used PCHi-C to generate an atlas of long-range promoter interactions in dozens of human and mouse cell types. These promoter interactome maps have contributed to a greater understanding of mammalian gene expression control by assigning putative regulatory regions to their target genes and revealing preferential spatial promoter-promoter interaction networks. This information also has high relevance to understanding human genetic disease and the identification of potential disease genes, by linking non-coding disease-associated sequence variants in or near control sequences to their target genes.

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Journal of visualized experiments : JoVE, , 1940-087X, , 2018

PMID: 30010637

Open Access

Epigenetic regulation in development: is the mouse a good model for the human?
Hanna CW, Demond H, Kelsey G

Over the past few years, advances in molecular technologies have allowed unprecedented mapping of epigenetic modifications in gametes and during early embryonic development. This work is allowing a detailed genomic analysis, which for the first time can answer long-standing questions about epigenetic regulation and reprogramming, and highlights differences between mouse and human, the implications of which are only beginning to be explored.

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Human reproduction update, , 1460-2369, , 2018

PMID: 29992283

Cold-induced epigenetic programming of the sperm enhances brown adipose tissue activity in the offspring.
Sun W, Dong H, Becker AS, Dapito DH, Modica S, Grandl G, Opitz L, Efthymiou V, Straub LG, Sarker G, Balaz M, Balazova L, Perdikari A, Kiehlmann E, Bacanovic S, Zellweger C, Peleg-Raibstein D, Pelczar P, Reik W, Burger IA, von Meyenn F, Wolfrum C

Recent research has focused on environmental effects that control tissue functionality and systemic metabolism. However, whether such stimuli affect human thermogenesis and body mass index (BMI) has not been explored. Here we show retrospectively that the presence of brown adipose tissue (BAT) and the season of conception are linked to BMI in humans. In mice, we demonstrate that cold exposure (CE) of males, but not females, before mating results in improved systemic metabolism and protection from diet-induced obesity of the male offspring. Integrated analyses of the DNA methylome and RNA sequencing of the sperm from male mice revealed several clusters of co-regulated differentially methylated regions (DMRs) and differentially expressed genes (DEGs), suggesting that the improved metabolic health of the offspring was due to enhanced BAT formation and increased neurogenesis. The conclusions are supported by cell-autonomous studies in the offspring that demonstrate an enhanced capacity to form mature active brown adipocytes, improved neuronal density and more norepinephrine release in BAT in response to cold stimulation. Taken together, our results indicate that in humans and in mice, seasonal or experimental CE induces an epigenetic programming of the sperm such that the offspring harbor hyperactive BAT and an improved adaptation to overnutrition and hypothermia.

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Nature medicine, , 1546-170X, , 2018

PMID: 29988127

Promoter interactome of human embryonic stem cell-derived cardiomyocytes connects GWAS regions to cardiac gene networks.
Choy MK, Javierre BM, Williams SG, Bar.oss SL, Liu Y, Wingett SW, Akbarov A, Wallace C, Freire-Pritchett P, Rugg-Gunn PJ, Spivakov M, Fraser P, Keavney BD

Long-range chromosomal interactions bring distal regulatory elements and promoters together to regulate gene expression in biological processes. By performing promoter capture Hi-C (PCHi-C) on human embryonic stem cell-derived cardiomyocytes (hESC-CMs), we show that such promoter interactions are a key mechanism by which enhancers contact their target genes after hESC-CM differentiation from hESCs. We also show that the promoter interactome of hESC-CMs is associated with expression quantitative trait loci (eQTLs) in cardiac left ventricular tissue; captures the dynamic process of genome reorganisation after hESC-CM differentiation; overlaps genome-wide association study (GWAS) regions associated with heart rate; and identifies new candidate genes in such regions. These findings indicate that regulatory elements in hESC-CMs identified by our approach control gene expression involved in ventricular conduction and rhythm of the heart. The study of promoter interactions in other hESC-derived cell types may be of utility in functional investigation of GWAS-associated regions.

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Nature communications, 9, 2041-1723, 2526, 2018

PMID: 29955040

Defective germline reprogramming rewires the spermatogonial transcriptome.
Vasiliauskaitė L, Berrens RV, Ivanova I, Carrieri C, Reik W, Enright AJ, O'Carroll D

Defective germline reprogramming in Piwil4 (Miwi2)- and Dnmt3l-deficient mice results in the failure to reestablish transposon silencing, meiotic arrest and progressive loss of spermatogonia. Here we sought to understand the molecular basis for this spermatogonial dysfunction. Through a combination of imaging, conditional genetics and transcriptome analysis, we demonstrate that germ cell elimination in the respective mutants arises as a result of defective de novo genome methylation during reprogramming rather than because of a function for the respective factors within spermatogonia. In both Miwi2 and Dnmt3l spermatogonia, the intracisternal-A particle (IAP) family of endogenous retroviruses is derepressed, but, in contrast to meiotic cells, DNA damage is not observed. Instead, we find that unmethylated IAP promoters rewire the spermatogonial transcriptome by driving expression of neighboring genes. Finally, spermatogonial numbers, proliferation and differentiation are altered in Miwi2 and Dnmt3l mice. In summary, defective reprogramming deregulates the spermatogonial transcriptome and may underlie spermatogonial dysfunction.

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Nature structural & molecular biology, 25, 1545-9985, 394-404, 2018

PMID: 29728652

Dynamics of the epigenetic landscape during the maternal-to-zygotic transition.
Eckersley-Maslin MA, Alda-Catalinas C, Reik W

A remarkable epigenetic remodelling process occurs shortly after fertilization, which restores totipotency to the zygote. This involves global DNA demethylation, chromatin remodelling, genome spatial reorganization and substantial transcriptional changes. Key to these changes is the transition from the maternal environment of the oocyte to an embryonic-driven developmental expression programme, a process termed the maternal-to-zygotic transition (MZT). Zygotic genome activation occurs predominantly at the two-cell stage in mice and the eight-cell stage in humans, yet the dynamics of its control are still mostly obscure. In recent years, partly due to single-cell and low-cell number epigenomic studies, our understanding of the epigenetic and chromatin landscape of preimplantation development has improved considerably. In this Review, we discuss the latest advances in the study of the MZT, focusing on DNA methylation, histone post-translational modifications, local chromatin structure and higher-order genome organization. We also discuss key mechanistic studies that investigate the mode of action of chromatin regulators, transcription factors and non-coding RNAs during preimplantation development. Finally, we highlight areas requiring additional research, as well as new technological advances that could assist in eventually completing our understanding of the MZT.

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Nature reviews. Molecular cell biology, , 1471-0080, , 2018

PMID: 29686419

Correction: Epigenetic resetting of human pluripotency (doi:10.1242/dev.146811).
Guo G, von Meyenn F, Rostovskaya M, Clarke J, Dietmann S, Baker D, Sahakyan A, Myers S, Bertone P, Reik W, Plath K, Smith A

Development (Cambridge, England), 145, 1477-9129, , 2018

PMID: 29669738

Open Access

A Critical Role of TET1/2 Proteins in Cell-Cycle Progression of Trophoblast Stem Cells.
Chrysanthou S, Senner CE, Woods L, Fineberg E, Okkenhaug H, Burge S, Perez-Garcia V, Hemberger M

The ten-eleven translocation (TET) proteins are well known for their role in maintaining naive pluripotency of embryonic stem cells. Here, we demonstrate that, jointly, TET1 and TET2 also safeguard the self-renewal potential of trophoblast stem cells (TSCs) and have partially redundant roles in maintaining the epithelial integrity of TSCs. For the more abundantly expressed TET1, we show that this is achieved by binding to critical epithelial genes, notably E-cadherin, which becomes hyper-methylated and downregulated in the absence of TET1. The epithelial-to-mesenchymal transition phenotype of mutant TSCs is accompanied by centrosome duplication and separation defects. Moreover, we identify a role of TET1 in maintaining cyclin B1 stability, thereby acting as facilitator of mitotic cell-cycle progression. As a result, Tet1/2 mutant TSCs are prone to undergo endoreduplicative cell cycles leading to the formation of polyploid trophoblast giant cells. Taken together, our data reveal essential functions of TET proteins in the trophoblast lineage.

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Stem cell reports, , 2213-6711, , 2018

PMID: 29576538

Open Access

Identifying Human Naïve Pluripotent Stem Cells - Evaluating State-Specific Reporter Lines and Cell-Surface Markers.
Collier AJ, Rugg-Gunn PJ

Recent reports that human pluripotent stem cells can be captured in a spectrum of states with variable properties has prompted a re-evaluation of how pluripotency is acquired and stabilised. The latest additions to the stem cell hierarchy open up opportunities for understanding human development, reprogramming, and cell state transitions more generally. Many of the new cell lines have been collectively termed 'naïve' human pluripotent stem cells to distinguish them from the conventional 'primed' cells. Here, several transcriptional and epigenetic hallmarks of human pluripotent states in the recently described cell lines are reviewed and evaluated. Methods to derive and identify human naïve pluripotent stem cells are also discussed, with a focus on the uses and future developments of state-specific reporter cell lines and cell-surface proteins. Finally, opportunities and uncertainties in naïve stem cell biology are highlighted, and the current limitations of human naïve pluripotent stem cells considered, particularly in the context of differentiation.

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BioEssays : news and reviews in molecular, cellular and developmental biology, , 1521-1878, e1700239, 2018

PMID: 29574793

Open Access

Comparison of whole-genome bisulfite sequencing library preparation strategies identifies sources of biases affecting DNA methylation data.
Olova N, Krueger F, Andrews S, Oxley D, Berrens RV, Branco MR, Reik W

Whole-genome bisulfite sequencing (WGBS) is becoming an increasingly accessible technique, used widely for both fundamental and disease-oriented research. Library preparation methods benefit from a variety of available kits, polymerases and bisulfite conversion protocols. Although some steps in the procedure, such as PCR amplification, are known to introduce biases, a systematic evaluation of biases in WGBS strategies is missing.

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Genome biology, 19, 1474-760X, 33, 2018

PMID: 29544553

Open Access

Integrin α2 marks a niche of trophoblast progenitor cells in first trimester human placenta
Lee CQE, Turco M, Gardner L, Simons B, Hemberger M, Moffett A

During pregnancy the trophoblast cells of the placenta are the only fetal cells in direct contact with maternal blood and decidua. Their functions include transport of nutrients and oxygen, secretion of pregnancy hormones, remodelling the uterine arteries, and communicating with maternal cells. Despite the importance of trophoblast cells in placental development and successful pregnancy, little is known about the identity, location and differentiation of human trophoblast progenitors. We identify a proliferative trophoblast niche at the base of the cytotrophoblast cell columns in first trimester placentas that is characterised by integrin α2 (ITGA2) expression. Pulse-chase experiments with 5-Iodo-2'-deoxyuridine (IdU) imply that these cells can contribute to both villous (VCT) and extravillous (EVT) lineages. These proliferating trophoblast cells can be isolated using ITGA2 as a marker by flow cytometry and express genes from both VCT and EVT. Microarray expression analysis shows that ITAG2cells display a unique transcriptional signature including NOTCH signalling and a combination of epithelial and mesenchymal characteristics. ITGA2 thus marks a niche allowing the study of pure populations of trophoblast progenitor cells.

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Development (Cambridge, England), , 1477-9129, , 2018

PMID: 29540503

Open Access

Placentation defects are highly prevalent in embryonic lethal mouse mutants.
Perez-Garcia V, Fineberg E, Wilson R, Murray A, Mazzeo CI, Tudor C, Sienerth A, White JK, Tuck E, Ryder EJ, Gleeson D, Siragher E, Wardle-Jones H, Staudt N, Wali N, Collins J, Geyer S, Busch-Nentwich EM, Galli A, Smith JC, Robertson E, Adams DJ, Weninger WJ, Mohun T, Hemberger M

Large-scale phenotyping efforts have demonstrated that approximately 25-30% of mouse gene knockouts cause intrauterine lethality. Analysis of these mutants has largely focused on the embryo and not the placenta, despite the crucial role of this extraembryonic organ for developmental progression. Here we screened 103 embryonic lethal and sub-viable mouse knockout lines from the Deciphering the Mechanisms of Developmental Disorders program for placental phenotypes. We found that 68% of knockout lines that are lethal at or after mid-gestation exhibited placental dysmorphologies. Early lethality (embryonic days 9.5-14.5) is almost always associated with severe placental malformations. Placental defects correlate strongly with abnormal brain, heart and vascular development. Analysis of mutant trophoblast stem cells and conditional knockouts suggests that a considerable number of factors that cause embryonic lethality when ablated have primary gene function in trophoblast cells. Our data highlight the hugely under-appreciated importance of placental defects in contributing to abnormal embryo development and suggest key molecular nodes that govern placenta formation.

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Nature, , 1476-4687, , 2018

PMID: 29539633

Long-Range Enhancer Interactions Are Prevalent in Mouse Embryonic Stem Cells and Are Reorganized upon Pluripotent State Transition.
Novo CL, Javierre BM, Cairns J, Segonds-Pichon A, Wingett SW, Freire-Pritchett P, Furlan-Magaril M, Schoenfelder S, Fraser P, Rugg-Gunn PJ

Transcriptional enhancers, including super-enhancers (SEs), form physical interactions with promoters to regulate cell-type-specific gene expression. SEs are characterized by high transcription factor occupancy and large domains of active chromatin, and they are commonly assigned to target promoters using computational predictions. How promoter-SE interactions change upon cell state transitions, and whether transcription factors maintain SE interactions, have not been reported. Here, we used promoter-capture Hi-C to identify promoters that interact with SEs in mouse embryonic stem cells (ESCs). We found that SEs form complex, spatial networks in which individual SEs contact multiple promoters, and a rewiring of promoter-SE interactions occurs between pluripotent states. We also show that long-range promoter-SE interactions are more prevalent in ESCs than in epiblast stem cells (EpiSCs) or Nanog-deficient ESCs. We conclude that SEs form cell-type-specific interaction networks that are partly dependent on core transcription factors, thereby providing insights into the gene regulatory organization of pluripotent cells.

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Cell reports, 22, 2211-1247, 2615-2627, 2018

PMID: 29514091

Open Access

Molecular profiling of aged neural progenitors identifies Dbx2 as a candidate regulator of age-associated neurogenic decline.
Lupo G, Nisi PS, Esteve P, Paul YL, Novo CL, Sidders B, Khan MA, Biagioni S, Liu HK, Bovolenta P, Cacci E, Rugg-Gunn PJ

Adult neurogenesis declines with aging due to the depletion and functional impairment of neural stem/progenitor cells (NSPCs). An improved understanding of the underlying mechanisms that drive age-associated neurogenic deficiency could lead to the development of strategies to alleviate cognitive impairment and facilitate neuroregeneration. An essential step towards this aim is to investigate the molecular changes that occur in NSPC aging on a genomewide scale. In this study, we compare the transcriptional, histone methylation and DNA methylation signatures of NSPCs derived from the subventricular zone (SVZ) of young adult (3 months old) and aged (18 months old) mice. Surprisingly, the transcriptional and epigenomic profiles of SVZ-derived NSPCs are largely unchanged in aged cells. Despite the global similarities, we detect robust age-dependent changes at several hundred genes and regulatory elements, thereby identifying putative regulators of neurogenic decline. Within this list, the homeobox gene Dbx2 is upregulated in vitro and in vivo, and its promoter region has altered histone and DNA methylation levels, in aged NSPCs. Using functional in vitro assays, we show that elevated Dbx2 expression in young adult NSPCs promotes age-related phenotypes, including the reduced proliferation of NSPC cultures and the altered transcript levels of age-associated regulators of NSPC proliferation and differentiation. Depleting Dbx2 in aged NSPCs caused the reverse gene expression changes. Taken together, these results provide new insights into the molecular programmes that are affected during mouse NSPC aging, and uncover a new functional role for Dbx2 in promoting age-related neurogenic decline.

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Aging cell, , 1474-9726, , 2018

PMID: 29504228

Open Access