scholarly journals Single cell multi-omics profiling reveals a hierarchical epigenetic landscape during mammalian germ layer specification

2019 ◽  
Author(s):  
Ricard Argelaguet ◽  
Hisham Mohammed ◽  
Stephen J Clark ◽  
L Carine Stapel ◽  
Christel Krueger ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Cell Fate ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
List Type ◽  
Germ Layer ◽  
Epigenetic Landscape ◽  
Germ Layers ◽  
Cell Fate Decisions

AbstractFormation of the three primary germ layers during gastrulation is an essential step in the establishment of the vertebrate body plan. Recent studies employing single cell RNA-sequencing have identified major transcriptional changes associated with germ layer specification. Global epigenetic reprogramming accompanies these changes, but the role of the epigenome in regulating early cell fate choice remains unresolved, and the coordination between different epigenetic layers is unclear. Here we describe the first single cell triple-omics map of chromatin accessibility, DNA methylation and RNA expression during the exit from pluripotency and the onset of gastrulation in mouse embryos. We find dynamic dependencies between the different molecular layers, with evidence for distinct modes of epigenetic regulation. The initial exit from pluripotency coincides with the establishment of a global repressive epigenetic landscape, followed by the emergence of local lineage-specific epigenetic patterns during gastrulation. Notably, cells committed to mesoderm and endoderm undergo widespread coordinated epigenetic rearrangements, driven by loss of methylation in enhancer marks and a concomitant increase of chromatin accessibility. In striking contrast, the epigenetic landscape of ectodermal cells is already established in the early epiblast. Hence, regulatory elements associated with each germ layer are either epigenetically primed or epigenetically remodelled prior to overt cell fate decisions during gastrulation, providing the molecular logic for a hierarchical emergence of the primary germ layers.HighlightsFirst map of mouse gastrulation using comprehensive single cell triple-omic analysis.Exit from pluripotency is associated with a global repressive epigenetic landscape, driven by a sharp gain of DNA methylation and a gradual decrease of chromatin accessibility.DNA methylation and chromatin accessibility changes in enhancers, but not in promoters, are associated with germ layer formation.Mesoderm and endoderm enhancers become open and demethylated upon lineage commitment.Ectoderm enhancers are primed in the early epiblast and protected from the global repressive dynamics, supporting a default model of ectoderm commitment in vivo.

scholarly journals Inference of emergent spatio-temporal processes from single-cell sequencing reveals feedback between de novo DNA methylation and chromatin condensates

2021 ◽  
Author(s):  
Fabrizio Olmeda ◽  
Tim Lohoff ◽  
Stephen J Clark ◽  
Laura Benson ◽  
Felix Krüger ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Cell Fate ◽  
De Novo ◽  
Dimensional Space ◽  
Theoretical Physics ◽  
List Type ◽  
Cell Fate Decisions ◽  
Spatio Temporal

SummaryRecent breakthroughs in single-cell genomics allow probing molecular states of cells with unprecedented detail along the sequence of the DNA. Biological function relies, however, on emergent processes in the three-dimensional space of the nucleus, such as droplet formation through phase separation. Here, we use single-cell multi-omics sequencing to develop a theoretical framework to rigorously map epigenome profiling along the DNA sequence onto a description of the emergent spatial dynamics in the nucleus. Drawing on scNMT-seq multi-omics sequencing in vitro and in vivo we exemplify our approach in the context of exit from pluripotency and global de novo methylation of the genome. We show how DNA methylation patterns of the embryonic genome are established through the interplay between spatially correlated DNA methylation and topological changes to the DNA. This feedback leads to the predicted formation of 30-40nm sized condensates of methylated DNA and determines genome-scale DNA methylation rates. We verify these findings with orthogonal single cell multi-omics data that combine the methylome with HiC measurements. Notably, this scale of chromatin organization has recently been described by super-resolution microscopy. Using this framework, we identify local methylation correlations in gene bodies that precede transcriptional changes at the exit from pluripotency. Our work provides a general framework of how mechanistic insights into emergent processes underlying cell fate decisions can be gained by the combination of single-cell multi-omics and methods from theoretical physics that have not been applied in the context of genomics before.HighlightsWe develop methodology to infer collective spatio-temporal processes in the physical space of the nucleus from single-cell methylome sequencing experiments.We show that DNA methylation relies on a feedback between de novo methylation and nanoscale changes in DNA topology, leading to the formation of methylation condensates.Chromatin condensates at this scale have recently been described by high-resolution microscopy but have remained without mechanistic explanation.Using this framework, we identify changes in the distribution of DNA methylation marks in gene bodies that precede gene silencing at the exit from pluripotency.

Single cell chromatin accessibility reveals regulatory elements and developmental trajectories in the embryonic forebrain

2021 ◽  
Author(s):  
Christopher T. Rhodes ◽  
Apratim Mitra ◽  
Dongjin R. Lee ◽  
Daniel J. Lee ◽  
Yajun Zhang ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Fate ◽  
Developmental Trajectories ◽  
Transcript Abundance ◽  
Regulatory Elements ◽  
Brain Regions ◽  
Chromatin Accessibility ◽  
Embryonic Cells ◽  
Embryonic Mouse ◽  
Cell Fate Decisions

SUMMARYWhile epigenetic modifications are critical for cell state changes throughout development, a detailed characterization of chromatin accessibility during neurogenesis has not been explored. We collected single-cell chromatin accessibility profiles from four distinct neurogenic regions of the embryonic mouse forebrain using single nuclei ATAC-Seq (snATAC-Seq). We identified thousands of differentially accessible peaks, many restricted to distinct progenitor cell types or brain regions. Integrating snATAC-Seq and transcriptome data, we characterized changes of chromatin accessibility at enhancers and promoters that were tightly coupled to transcript abundance during neurodevelopment. Integrating chromatin accessibility profiles from embryonic and adult interneurons with the iPSYCH2012 dataset revealed several disease-associated polymorphisms overlapped with accessible regions in embryonic cells. These findings highlight a diverse chromatin landscape in embryonic neural progenitors, extensive coordination between chromatin accessibility and gene expression during neuron fate determination, and open the door for future studies to define critical enhancer-promoter interactions that direct cell fate decisions.

scholarly journals Genome-scale oscillations in DNA methylation during exit from pluripotency

2018 ◽  
Author(s):  
Steffen Rulands ◽  
Heather J Lee ◽  
Stephen J Clark ◽  
Christof Angermueller ◽  
Sébastien A Smallwood ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Epigenetic Mark ◽  
List Type ◽  
Dna Hypomethylation ◽  
Cell Fate Decisions ◽  
Oscillatory Dynamics ◽  
Genome Scale

SummaryPluripotency 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, coexpression 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 modelling, 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 fresh insights into the emergence of epigenetic heterogeneity during early embryo development, indicating that dynamic changes in DNA methylation might influence early cell fate decisions.HighlightsCo-expression of DNMT3s and TETs drive genome-scale oscillations of DNA methylationOscillation amplitude is greatest at a CpG density characteristic of enhancersCell synchronisation reveals oscillation period and link with primary transcriptsMultiomic single-cell profiling provides evidence for oscillatory dynamics in vivo

scholarly journals Lifelong single-cell profiling of cranial neural crest diversification

2021 ◽  
Author(s):  
Peter Fabian ◽  
Kuo-Chang Tseng ◽  
Mathi Thiruppathy ◽  
Claire Arata ◽  
Hung-Jhen Chen ◽  
...  
Keyword(s):  
Neural Crest ◽  
Single Cell ◽  
Cell Fate ◽  
Intrinsic Property ◽  
Chromatin Accessibility ◽  
Specific Cell ◽  
List Type ◽  
Cranial Neural Crest ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

AbstractThe cranial neural crest generates a huge diversity of derivatives, including the bulk of connective and skeletal tissues of the vertebrate head. How neural crest cells acquire such extraordinary lineage potential remains unresolved. By integrating single-cell transcriptome and chromatin accessibility profiles of cranial neural crest-derived cells across the zebrafish lifetime, we observe region-specific establishment of enhancer accessibility for distinct fates. Neural crest-derived cells rapidly diversify into specialized progenitors, including multipotent skeletal progenitors, stromal cells with a regenerative signature, fibroblasts with a unique metabolic signature linked to skeletal integrity, and gill-specific progenitors generating cell types for respiration. By retrogradely mapping the emergence of lineage-specific chromatin accessibility, we identify a wealth of candidate lineage-priming factors, including a Gata3 regulatory circuit for respiratory cell fates. Rather than multilineage potential being an intrinsic property of cranial neural crest, our findings support progressive and region-specific chromatin remodeling underlying acquisition of diverse neural crest lineage potential.HighlightsSingle-cell transcriptome and chromatin atlas of cranial neural crestProgressive emergence of region-specific cell fate competencyChromatin accessibility mapping identifies candidate lineage regulatorsGata3 function linked to gill-specific respiratory programGraphical Abstract

scholarly journals Genetic control of pluripotency epigenome determines differentiation bias in mouse embryonic stem cells

2021 ◽  
Author(s):  
Candice Byers ◽  
Catrina Spruce ◽  
Haley J. Fortin ◽  
Anne Czechanski ◽  
Steven C. Munger ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cell Fate ◽  
Genetic Regulation ◽  
Embryonic Stem ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Embryoid Bodies ◽  
Cell Fate Decisions

AbstractGenetically diverse pluripotent stem cells (PSCs) display varied, heritable responses to differentiation cues in the culture environment. By harnessing these disparities through derivation of embryonic stem cells (ESCs) from the BXD mouse genetic reference panel, along with C57BL/6J (B6) and DBA/2J (D2) parental strains, we demonstrate genetically determined biases in lineage commitment and identify major regulators of the pluripotency epigenome. Upon transition to formative pluripotency using epiblast-like cells (EpiLCs), B6 quickly dissolves naïve networks adopting gene expression modules indicative of neuroectoderm lineages; whereas D2 retains aspects of naïve pluripotency with little bias in differentiation. Genetic mapping identifies 6 major trans-acting loci co-regulating chromatin accessibility and gene expression in ESCs and EpiLCs, indicating a common regulatory system impacting cell state transition. These loci distally modulate occupancy of pluripotency factors, including TRIM28, P300, and POU5F1, at hundreds of regulatory elements. One trans-acting locus on Chr 12 primarily impacts chromatin accessibility in ESCs; while in EpiLCs the same locus subsequently influences gene expression, suggesting early chromatin priming. Consequently, the distal gene targets of this locus are enriched for neurogenesis genes and were more highly expressed when cells carried B6 haplotypes at this Chr 12 locus, supporting genetic regulation of biases in cell fate. Spontaneous formation of embryoid bodies validated this with B6 showing a propensity towards neuroectoderm differentiation and D2 towards definitive endoderm, confirming the fundamental importance of genetic variation influencing cell fate decisions.

scholarly journals Chromatin accessibility dynamics and single cell RNA-Seq reveal new regulators of regeneration in neural progenitors

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Anneke Dixie Kakebeen ◽  
Alexander Daniel Chitsazan ◽  
Madison Corinne Williams ◽  
Lauren M Saunders ◽  
Andrea Elizabeth Wills
Keyword(s):  
Single Cell ◽  
Cell Fate ◽  
Neural Progenitor Cells ◽  
Cell Types ◽  
Chromatin Accessibility ◽  
Rna Seq ◽  
Tail Regeneration ◽  
Cell Fate Decisions ◽  
Transcriptional Regulatory ◽  
Regulatory Dynamics

Vertebrate appendage regeneration requires precisely coordinated remodeling of the transcriptional landscape to enable the growth and differentiation of new tissue, a process executed over multiple days and across dozens of cell types. The heterogeneity of tissues and temporally-sensitive fate decisions involved has made it difficult to articulate the gene regulatory programs enabling regeneration of individual cell types. To better understand how a regenerative program is fulfilled by neural progenitor cells (NPCs) of the spinal cord, we analyzed pax6-expressing NPCs isolated from regenerating Xenopus tropicalis tails. By intersecting chromatin accessibility data with single-cell transcriptomics, we find that NPCs place an early priority on neuronal differentiation. Late in regeneration, the priority returns to proliferation. Our analyses identify Pbx3 and Meis1 as critical regulators of tail regeneration and axon organization. Overall, we use transcriptional regulatory dynamics to present a new model for cell fate decisions and their regulators in NPCs during regeneration.

scholarly journals Sex reversal following deletion of a single distal enhancer of Sox9

Science ◽  
2018 ◽  
Vol 360 (6396) ◽  
pp. 1469-1473 ◽  
Author(s):  
Nitzan Gonen ◽  
Chris R. Futtner ◽  
Sophie Wood ◽  
S. Alexandra Garcia-Moreno ◽  
Isabella M. Salamone ◽  
...  
Keyword(s):  
Cell Fate ◽  
Transcriptional Start Site ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Sex Reversal ◽  
Distal Enhancer ◽  
Gene Desert ◽  
Cell Fate Decisions ◽  
Xy Sex Reversal

Cell fate decisions require appropriate regulation of key genes. Sox9, a direct target of SRY, is pivotal in mammalian sex determination. In vivo high-throughput chromatin accessibility techniques, transgenic assays, and genome editing revealed several novel gonadal regulatory elements in the 2-megabase gene desert upstream of Sox9. Although others are redundant, enhancer 13 (Enh13), a 557–base pair element located 565 kilobases 5′ from the transcriptional start site, is essential to initiate mouse testis development; its deletion results in XY females with Sox9 transcript levels equivalent to those in XX gonads. Our data are consistent with the time-sensitive activity of SRY and indicate a strict order of enhancer usage. Enh13 is conserved and embedded within a 32.5-kilobase region whose deletion in humans is associated with XY sex reversal, suggesting that it is also critical in humans.

scholarly journals Epigenetic dynamics shaping melanophore and iridophore cell fate in zebrafish

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Hyo Sik Jang ◽  
Yujie Chen ◽  
Jiaxin Ge ◽  
Alicia N. Wilkening ◽  
Yiran Hou ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Cell Differentiation ◽  
Neural Crest ◽  
Cell Fate ◽  
Pigment Cell ◽  
Chromatin Accessibility ◽  
Pigment Cells ◽  
Epigenetic Landscape ◽  
Cell Fate Decisions ◽  
Neural Crest Differentiation

Abstract Background Zebrafish pigment cell differentiation provides an attractive model for studying cell fate progression as a neural crest progenitor engenders diverse cell types, including two morphologically distinct pigment cells: black melanophores and reflective iridophores. Nontrivial classical genetic and transcriptomic approaches have revealed essential molecular mechanisms and gene regulatory circuits that drive neural crest-derived cell fate decisions. However, how the epigenetic landscape contributes to pigment cell differentiation, especially in the context of iridophore cell fate, is poorly understood. Results We chart the global changes in the epigenetic landscape, including DNA methylation and chromatin accessibility, during neural crest differentiation into melanophores and iridophores to identify epigenetic determinants shaping cell type-specific gene expression. Motif enrichment in the epigenetically dynamic regions reveals putative transcription factors that might be responsible for driving pigment cell identity. Through this effort, in the relatively uncharacterized iridophores, we validate alx4a as a necessary and sufficient transcription factor for iridophore differentiation and present evidence on alx4a’s potential regulatory role in guanine synthesis pathway. Conclusions Pigment cell fate is marked by substantial DNA demethylation events coupled with dynamic chromatin accessibility to potentiate gene regulation through cis-regulatory control. Here, we provide a multi-omic resource for neural crest differentiation into melanophores and iridophores. This work led to the discovery and validation of iridophore-specific alx4a transcription factor.

scholarly journals Single cell epigenomic atlas of the developing human brain and organoids

2019 ◽  
Author(s):  
Ryan S. Ziffra ◽  
Chang N. Kim ◽  
Amy Wilfert ◽  
Tychele N. Turner ◽  
Maximilian Haeussler ◽  
...  
Keyword(s):  
Human Brain ◽  
Single Cell ◽  
Cell Fate ◽  
Cell Types ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Chromatin State ◽  
Cell Type ◽  
Fate Specification ◽  
Cell Type Specific

AbstractDynamic changes in chromatin accessibility coincide with important aspects of neuronal differentiation, such as fate specification and arealization and confer cell type-specific associations to neurodevelopmental disorders. However, studies of the epigenomic landscape of the developing human brain have yet to be performed at single-cell resolution. Here, we profiled chromatin accessibility of >75,000 cells from eight distinct areas of developing human forebrain using single cell ATAC-seq (scATACseq). We identified thousands of loci that undergo extensive cell type-specific changes in accessibility during corticogenesis. Chromatin state profiling also reveals novel distinctions between neural progenitor cells from different cortical areas not seen in transcriptomic profiles and suggests a role for retinoic acid signaling in cortical arealization. Comparison of the cell type-specific chromatin landscape of cerebral organoids to primary developing cortex found that organoids establish broad cell type-specific enhancer accessibility patterns similar to the developing cortex, but lack many putative regulatory elements identified in homologous primary cell types. Together, our results reveal the important contribution of chromatin state to the emerging patterns of cell type diversity and cell fate specification and provide a blueprint for evaluating the fidelity and robustness of cerebral organoids as a model for cortical development.

scholarly journals The cis-regulatory dynamics of embryonic development at single cell resolution

2017 ◽  
Author(s):  
Darren A. Cusanovich ◽  
James P. Reddington ◽  
David A. Garfield ◽  
Riza Daza ◽  
Raquel Marco-Ferreres ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Fate ◽  
Cell Types ◽  
Chromatin Accessibility ◽  
Open Chromatin ◽  
Germ Layer ◽  
Regulatory Changes ◽  
Single Cell Profiling ◽  
Transgenic Embryos ◽  
Cell Trajectories

ABSTRACTSingle cell measurements of gene expression are providing new insights into lineage commitment, yet the regulatory changes underlying individual cell trajectories remain elusive. Here, we profiled chromatin accessibility in over 20,000 single nuclei across multiple stages of Drosophila embryogenesis. Our data reveal heterogeneity in the regulatory landscape prior to gastrulation that reflects anatomical position, a feature that aligns with future cell fate. During mid embryogenesis, tissue granularity emerges such that cell types can be inferred by their chromatin accessibility, while maintaining a signature of their germ layer of origin. We identify over 30,000 distal elements with tissue-specific accessibility. Using transgenic embryos, we tested the germ layer specificity of a subset of predicted enhancers, achieving near-perfect accuracy. Overall, these data demonstrate the power of shotgun single cell profiling of embryos to resolve dynamic changes in open chromatin during development, and to uncover the cis-regulatory programs of germ layers and cell types.

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