Life Sciences Research for Lifelong Health

Publications wolf-reik

Title / Authors / Details Open Access Download

Voices in methods development.
Anikeeva P, Boyden E, Brangwynne C, Cissé II, Fiehn O, Fromme P, Gingras AC, Greene CS, Heard E, Hell SW, Hillman E, Jensen GJ, Karchin R, Kiessling LL, Kleinstiver BP, Knight R, Kukura P, Lancaster MA, Loman N, Looger L, Lundberg E, Luo Q, Miyawaki A, Myers EW, Nolan GP, Picotti P, Reik W, Sauer M, Shalek AK, Shendure J, Slavov N, Tanay A, Troyanskaya O, van Valen D, Wang HW, Yi C, Yin P, Zernicka-Goetz M, Zhuang X

Nature methods, 16, 1548-7105, 945-951, 2019

PMID: 31562479

Ageing affects DNA methylation drift and transcriptional cell-to-cell variability in mouse muscle stem cells.
Hernando-Herraez I, Evano B, Stubbs T, Commere PH, Jan Bonder M, Clark S, Andrews S, Tajbakhsh S, Reik W

Age-related tissue alterations have been associated with a decline in stem cell number and function. Although increased cell-to-cell variability in transcription or epigenetic marks has been proposed to be a major hallmark of ageing, little is known about the molecular diversity of stem cells during ageing. Here we present a single cell multi-omics study of mouse muscle stem cells, combining single-cell transcriptome and DNA methylome profiling. Aged cells show a global increase of uncoordinated transcriptional heterogeneity biased towards genes regulating cell-niche interactions. We find context-dependent alterations of DNA methylation in aged stem cells. Importantly, promoters with increased methylation heterogeneity are associated with increased transcriptional heterogeneity of the genes they drive. These results indicate that epigenetic drift, by accumulation of stochastic DNA methylation changes in promoters, is associated with the degradation of coherent transcriptional networks during stem cell ageing. Furthermore, our observations also shed light on the mechanisms underlying the DNA methylation clock.

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Nature communications, 10, 2041-1723, 4361, 2019

PMID: 31554804

Open Access

Distinct Molecular Trajectories Converge to Induce Naive Pluripotency.
Stuart HT, Stirparo GG, Lohoff T, Bates LE, Kinoshita M, Lim CY, Sousa EJ, Maskalenka K, Radzisheuskaya A, Malcolm AA, Alves MRP, Lloyd RL, Nestorowa S, Humphreys P, Mansfield W, Reik W, Bertone P, Nichols J, Göttgens B, Silva JCR

Understanding how cell identity transitions occur and whether there are multiple paths between the same beginning and end states are questions of wide interest. Here we show that acquisition of naive pluripotency can follow transcriptionally and mechanistically distinct routes. Starting from post-implantation epiblast stem cells (EpiSCs), one route advances through a mesodermal state prior to naive pluripotency induction, whereas another transiently resembles the early inner cell mass and correspondingly gains greater developmental potency. These routes utilize distinct signaling networks and transcription factors but subsequently converge on the same naive endpoint, showing surprising flexibility in mechanisms underlying identity transitions and suggesting that naive pluripotency is a multidimensional attractor state. These route differences are reconciled by precise expression of Oct4 as a unifying, essential, and sufficient feature. We propose that fine-tuned regulation of this "transition factor" underpins multidimensional access to naive pluripotency, offering a conceptual framework for understanding cell identity transitions.

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Cell stem cell, , 1875-9777, , 2019

PMID: 31422912

Screening for genes that accelerate the epigenetic aging clock in humans reveals a role for the H3K36 methyltransferase NSD1.
Martin-Herranz DE, Aref-Eshghi E, Bonder MJ, Stubbs TM, Choufani S, Weksberg R, Stegle O, Sadikovic B, Reik W, Thornton JM

Epigenetic clocks are mathematical models that predict the biological age of an individual using DNA methylation data and have emerged in the last few years as the most accurate biomarkers of the aging process. However, little is known about the molecular mechanisms that control the rate of such clocks. Here, we have examined the human epigenetic clock in patients with a variety of developmental disorders, harboring mutations in proteins of the epigenetic machinery.

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Genome biology, 20, 1474-760X, 146, 2019

PMID: 31409373

Open Access

Establishment of porcine and human expanded potential stem cells.
Gao X, Nowak-Imialek M, Chen X, Chen D, Herrmann D, Ruan D, Chen ACH, Eckersley-Maslin MA, Ahmad S, Lee YL, Kobayashi T, Ryan D, Zhong J, Zhu J, Wu J, Lan G, Petkov S, Yang J, Antunes L, Campos LS, Fu B, Wang S, Yong Y, Wang X, Xue SG, Ge L, Liu Z, Huang Y, Nie T, Li P, Wu D, Pei D, Zhang Y, Lu L, Yang F, Kimber SJ, Reik W, Zou X, Shang Z, Lai L, Surani A, Tam PPL, Ahmed A, Yeung WSB, Teichmann SA, Niemann H, Liu P

We recently derived mouse expanded potential stem cells (EPSCs) from individual blastomeres by inhibiting the critical molecular pathways that predispose their differentiation. EPSCs had enriched molecular signatures of blastomeres and possessed developmental potency for all embryonic and extra-embryonic cell lineages. Here, we report the derivation of porcine EPSCs, which express key pluripotency genes, are genetically stable, permit genome editing, differentiate to derivatives of the three germ layers in chimeras and produce primordial germ cell-like cells in vitro. Under similar conditions, human embryonic stem cells and induced pluripotent stem cells can be converted, or somatic cells directly reprogrammed, to EPSCs that display the molecular and functional attributes reminiscent of porcine EPSCs. Importantly, trophoblast stem-cell-like cells can be generated from both human and porcine EPSCs. Our pathway-inhibition paradigm thus opens an avenue for generating mammalian pluripotent stem cells, and EPSCs present a unique cellular platform for translational research in biotechnology and regenerative medicine.

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Nature cell biology, 21, 1476-4679, 687-699, 2019

PMID: 31160711

TET3 prevents terminal differentiation of adult NSCs by a non-catalytic action at Snrpn.
Montalbán-Loro R, Lozano-Ureña A, Ito M, Krueger C, Reik W, Ferguson-Smith AC, Ferrón SR

Ten-eleven-translocation (TET) proteins catalyze DNA hydroxylation, playing an important role in demethylation of DNA in mammals. Remarkably, although hydroxymethylation levels are high in the mouse brain, the potential role of TET proteins in adult neurogenesis is unknown. We show here that a non-catalytic action of TET3 is essentially required for the maintenance of the neural stem cell (NSC) pool in the adult subventricular zone (SVZ) niche by preventing premature differentiation of NSCs into non-neurogenic astrocytes. This occurs through direct binding of TET3 to the paternal transcribed allele of the imprinted gene Small nuclear ribonucleoprotein-associated polypeptide N (Snrpn), contributing to transcriptional repression of the gene. The study also identifies BMP2 as an effector of the astrocytic terminal differentiation mediated by SNRPN. Our work describes a novel mechanism of control of an imprinted gene in the regulation of adult neurogenesis through an unconventional role of TET3.

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Nature communications, 10, 2041-1723, 1726, 2019

PMID: 30979904

Open Access

Correction to: 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

Following publication of the original article [1], it was reported that the incorrect "Additional file 3" was published. The correct additional file is given below.

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Genome biology, 20, 1474-760X, 43, 2019

PMID: 30795792

Open Access

A single-cell molecular map of mouse gastrulation and early organogenesis.
Pijuan-Sala B, Griffiths JA, Guibentif C, Hiscock TW, Jawaid W, Calero-Nieto FJ, Mulas C, Ibarra-Soria X, Tyser RCV, Ho DLL, Reik W, Srinivas S, Simons BD, Nichols J, Marioni JC, Göttgens B

Across the animal kingdom, gastrulation represents a key developmental event during which embryonic pluripotent cells diversify into lineage-specific precursors that will generate the adult organism. Here we report the transcriptional profiles of 116,312 single cells from mouse embryos collected at nine sequential time points ranging from 6.5 to 8.5 days post-fertilization. We construct a molecular map of cellular differentiation from pluripotency towards all major embryonic lineages, and explore the complex events involved in the convergence of visceral and primitive streak-derived endoderm. Furthermore, we use single-cell profiling to show that Tal1 chimeric embryos display defects in early mesoderm diversification, and we thus demonstrate how combining temporal and transcriptional information can illuminate gene function. Together, this comprehensive delineation of mammalian cell differentiation trajectories in vivo represents a baseline for understanding the effects of gene mutations during development, as well as a roadmap for the optimization of in vitro differentiation protocols for regenerative medicine.

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Nature, , 1476-4687, , 2019

PMID: 30787436

Open Access

Combined single-cell profiling of expression and DNA methylation reveals splicing regulation and heterogeneity.
Linker SM, Urban L, Clark SJ, Chhatriwala M, Amatya S, McCarthy DJ, Ebersberger I, Vallier L, Reik W, Stegle O, Bonder MJ

Alternative splicing is a key regulatory mechanism in eukaryotic cells and increases the effective number of functionally distinct gene products. Using bulk RNA sequencing, splicing variation has been studied across human tissues and in genetically diverse populations. This has identified disease-relevant splicing events, as well as associations between splicing and genomic features, including sequence composition and conservation. However, variability in splicing between single cells from the same tissue or cell type and its determinants remains poorly understood.

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Genome biology, 20, 1474-760X, 30, 2019

PMID: 30744673

Open Access

Dppa2 and Dppa4 directly regulate the Dux-driven zygotic transcriptional program.
Eckersley-Maslin M, Alda-Catalinas C, Blotenburg M, Kreibich E, Krueger C, Reik W

The molecular regulation of zygotic genome activation (ZGA) in mammals remains an exciting area of research. Primed mouse embryonic stem cells contain a rare subset of "2C-like" cells that are epigenetically and transcriptionally similar to the two-cell embryo and thus represent an in vitro approximation for studying ZGA transcription regulation. Recently, the transcription factor Dux, expressed in the minor wave of ZGA, was described to activate many downstream ZGA transcripts. However, it remains unknown what upstream maternal factors initiate ZGA in either a Dux-dependent or Dux-independent manner. Here we performed a candidate-based overexpression screen, identifying, among others, developmental pluripotency-associated 2 (Dppa2) and Dppa4 as positive regulators of 2C-like cells and transcription of ZGA genes. In the germline, promoter DNA demethylation coincides with expression of Dppa2 and Dppa4, which remain expressed until embryonic day 7.5 (E7.5), when their promoters are remethylated. Furthermore, Dppa2 and Dppa4 are also expressed during induced pluripotent stem cell (iPSC) reprogramming at the time that 2C-like transcription transiently peaks. Through a combination of overexpression, knockdown, knockout, and rescue experiments together with transcriptional analyses, we show that Dppa2 and Dppa4 directly regulate the 2C-like cell population and associated transcripts, including Dux and the Zscan4 cluster. Importantly, we teased apart the molecular hierarchy in which the 2C-like transcriptional program is initiated and stabilized. Dppa2 and Dppa4 require Dux to initiate 2C-like transcription, suggesting that they act upstream by directly regulating Dux. Supporting this, ChIP-seq (chromatin immunoprecipitation [ChIP] combined with high-throughput sequencing) analysis revealed that Dppa2 and Dppa4 bind to the Dux promoter and gene body and drive its expression. Zscan4c is also able to induce 2C-like cells in wild-type cells but, in contrast to Dux, can no longer do so in Dppa2/4 double-knockout cells, suggesting that it may act to stabilize rather than drive the transcriptional network. Our findings suggest a model in which Dppa2/4 binding to the Dux promoter leads to Dux up-regulation and activation of the 2C-like transcriptional program, which is subsequently reinforced by Zscan4c.

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Genes & development, 33, 1549-5477, 194-208, 2019

PMID: 30692203

Open Access

Transcriptional Heterogeneity in Naive and Primed Human Pluripotent Stem Cells at Single-Cell Resolution.
Messmer T, von Meyenn F, Savino A, Santos F, Mohammed H, Lun ATL, Marioni JC, Reik W

Conventional human embryonic stem cells are considered to be primed pluripotent but can be induced to enter a naive state. However, the transcriptional features associated with naive and primed pluripotency are still not fully understood. Here we used single-cell RNA sequencing to characterize the differences between these conditions. We observed that both naive and primed populations were mostly homogeneous with no clear lineage-related structure and identified an intermediate subpopulation of naive cells with primed-like expression. We found that the naive-primed pluripotency axis is preserved across species, although the timing of the transition to a primed state is species specific. We also identified markers for distinguishing human naive and primed pluripotency as well as strong co-regulatory relationships between lineage markers and epigenetic regulators that were exclusive to naive cells. Our data provide valuable insights into the transcriptional landscape of human pluripotency at a cellular and genome-wide resolution.

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Cell reports, 26, 2211-1247, 815-824.e4, 2019

PMID: 30673604

Open Access

The non-canonical SMC protein SmcHD1 antagonises TAD formation and compartmentalisation on the inactive X chromosome.
Gdula MR, Nesterova TB, Pintacuda G, Godwin J, Zhan Y, Ozadam H, McClellan M, Moralli D, Krueger F, Green CM, Reik W, Kriaucionis S, Heard E, Dekker J, Brockdorff N

The inactive X chromosome (Xi) in female mammals adopts an atypical higher-order chromatin structure, manifested as a global loss of local topologically associated domains (TADs), A/B compartments and formation of two mega-domains. Here we demonstrate that the non-canonical SMC family protein, SmcHD1, which is important for gene silencing on Xi, contributes to this unique chromosome architecture. Specifically, allelic mapping of the transcriptome and epigenome in SmcHD1 mutant cells reveals the appearance of sub-megabase domains defined by gene activation, CpG hypermethylation and depletion of Polycomb-mediated H3K27me3. These domains, which correlate with sites of SmcHD1 enrichment on Xi in wild-type cells, additionally adopt features of active X chromosome higher-order chromosome architecture, including A/B compartments and partial restoration of TAD boundaries. Xi chromosome architecture changes also occurred following SmcHD1 knockout in a somatic cell model, but in this case, independent of Xi gene derepression. We conclude that SmcHD1 is a key factor in defining the unique chromosome architecture of Xi.

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Nature communications, 10, 2041-1723, 30, 2019

PMID: 30604745

Open Access

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

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

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

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

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

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

scNMT-seq enables joint profiling of chromatin accessibility DNA methylation and transcription in single cells.
Clark SJ, Argelaguet R, Kapourani CA, Stubbs TM, Lee HJ, Alda-Catalinas C, Krueger F, Sanguinetti G, Kelsey G, Marioni JC, Stegle O, Reik W

Parallel single-cell sequencing protocols represent powerful methods for investigating regulatory relationships, including epigenome-transcriptome interactions. Here, we report a single-cell method for parallel chromatin accessibility, DNA methylation and transcriptome profiling. scNMT-seq (single-cell nucleosome, methylation and transcription sequencing) uses a GpC methyltransferase to label open chromatin followed by bisulfite and RNA sequencing. We validate scNMT-seq by applying it to differentiating mouse embryonic stem cells, finding links between all three molecular layers and revealing dynamic coupling between epigenomic layers during differentiation.

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Nature communications, 9, 2041-1723, 781, 2018

PMID: 29472610

Science Forum: The Human Cell Atlas.
Regev A, Teichmann SA, Lander ES, Amit I, Benoist C, Birney E, Bodenmiller B, Campbell PJ, Carninci P, Clatworthy M, Clevers H, Deplancke B, Dunham I, Eberwine J, Eils R, Enard W, Farmer A, Fugger L, Göttgens B, Hacohen N, Haniffa M, Hemberg M, Kim SK, Klenerman P, Kriegstein A, Lein E, Linnarsson S, Lundberg E, Lundeberg J, Majumder P, Marioni JC, Merad M, Mhlanga M, Nawijn M, Netea M, Nolan G, Pe'er D, Phillipakis A, Ponting CP, Quake SR, Reik W, Rozenblatt-Rosen O, Sanes JR, Satija R, Schumacher TN, Shalek AK, Shapiro E, Sharma P, Shin JW, Stegle O, Stratton MR, Stubbington MJT, Theis FJ, Uhlen M, van Oudenaarden A, Wagner A, Watt FM, Weissman JS, Wold BJ, Xavier RJ, Yosef N,

The recent advent of methods for high-throughput single-cell molecular profiling has catalyzed a growing sense in the scientific community that the time is ripe to complete the 150-year-old effort to identify all cell types in the human body. The Human Cell Atlas Project is an international collaborative effort that aims to define all human cell types in terms of distinctive molecular profiles (such as gene expression profiles) and to connect this information with classical cellular descriptions (such as location and morphology). An open comprehensive reference map of the molecular state of cells in healthy human tissues would propel the systematic study of physiological states, developmental trajectories, regulatory circuitry and interactions of cells, and also provide a framework for understanding cellular dysregulation in human disease. Here we describe the idea, its potential utility, early proofs-of-concept, and some design considerations for the Human Cell Atlas, including a commitment to open data, code, and community.

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eLife, 6, 2050-084X, , 2017

PMID: 29206104

Open Access

Coupling shRNA screens with single-cell RNA-seq identifies a dual role for mTOR in reprogramming-induced senescence.
Aarts M, Georgilis A, Beniazza M, Beolchi P, Banito A, Carroll T, Kulisic M, Kaemena DF, Dharmalingam G, Martin N, Reik W, Zuber J, Kaji K, Chandra T, Gil J

Expression of the transcription factors OCT4, SOX2, KLF4, and cMYC (OSKM) reprograms somatic cells into induced pluripotent stem cells (iPSCs). Reprogramming is a slow and inefficient process, suggesting the presence of safeguarding mechanisms that counteract cell fate conversion. One such mechanism is senescence. To identify modulators of reprogramming-induced senescence, we performed a genome-wide shRNA screen in primary human fibroblasts expressing OSKM. In the screen, we identified novel mediators of OSKM-induced senescence and validated previously implicated genes such as CDKN1A We developed an innovative approach that integrates single-cell RNA sequencing (scRNA-seq) with the shRNA screen to investigate the mechanism of action of the identified candidates. Our data unveiled regulation of senescence as a novel way by which mechanistic target of rapamycin (mTOR) influences reprogramming. On one hand, mTOR inhibition blunts the induction of cyclin-dependent kinase (CDK) inhibitors (CDKIs), including p16(INK4a), p21(CIP1), and p15(INK4b), preventing OSKM-induced senescence. On the other hand, inhibition of mTOR blunts the senescence-associated secretory phenotype (SASP), which itself favors reprogramming. These contrasting actions contribute to explain the complex effect that mTOR has on reprogramming. Overall, our study highlights the advantage of combining functional screens with scRNA-seq to accelerate the discovery of pathways controlling complex phenotypes.

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Genes & development, , 1549-5477, , 2017

PMID: 29138277

Open Access