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

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.

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

10.1101/519207 ◽

2019 ◽

Cited By ~ 5

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.

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

10.1101/2021.01.15.426861 ◽

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.

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

eLife ◽

10.7554/elife.52648 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 2

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.

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Sex reversal following deletion of a single distal enhancer of Sox9

Science ◽

10.1126/science.aas9408 ◽

2018 ◽

Vol 360 (6396) ◽

pp. 1469-1473 ◽

Cited By ~ 72

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.

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Reversed graph embedding resolves complex single-cell developmental trajectories

10.1101/110668 ◽

2017 ◽

Cited By ~ 36

Author(s):

Xiaojie Qiu ◽

Qi Mao ◽

Ying Tang ◽

Li Wang ◽

Raghav Chawla ◽

...

Keyword(s):

Single Cell ◽

Cell Fate ◽

Developmental Trajectories ◽

Single Cells ◽

Graph Embedding ◽

Developmental Trajectory ◽

Loss Of Function ◽

Cell Fate Decisions ◽

Principal Graph ◽

Cell Trajectories

AbstractOrganizing single cells along a developmental trajectory has emerged as a powerful tool for understanding how gene regulation governs cell fate decisions. However, learning the structure of complex single-cell trajectories with two or more branches remains a challenging computational problem. We present Monocle 2, which uses reversed graph embedding to reconstruct single-cell trajectories in a fully unsupervised manner. Monocle 2 learns an explicit principal graph to describe the data, greatly improving the robustness and accuracy of its trajectories compared to other algorithms. Monocle 2 uncovered a new, alternative cell fate in what we previously reported to be a linear trajectory for differentiating myoblasts. We also reconstruct branched trajectories for two studies of blood development, and show that loss of function mutations in key lineage transcription factors diverts cells to alternative branches on the a trajectory. Monocle 2 is thus a powerful tool for analyzing cell fate decisions with single-cell genomics.

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Single-cell epigenomic identification of inherited risk loci in Alzheimer’s and Parkinson’s disease

10.1101/2020.01.06.896159 ◽

2020 ◽

Cited By ~ 9

Author(s):

M. Ryan Corces ◽

Anna Shcherbina ◽

Soumya Kundu ◽

Michael J. Gloudemans ◽

Laure Frésard ◽

...

Keyword(s):

Parkinson’S Disease ◽

Parkinson's Disease ◽

Single Cell ◽

Association Studies ◽

Regulatory Elements ◽

Brain Regions ◽

Chromatin Accessibility ◽

Adult Brain ◽

Genome Wide Association Studies ◽

Nucleotide Polymorphisms

ABSTRACTGenome-wide association studies (GWAS) have identified thousands of variants associated with disease phenotypes. However, the majority of these variants do not alter coding sequences, making it difficult to assign their function. To this end, we present a multi-omic epigenetic atlas of the adult human brain through profiling of the chromatin accessibility landscapes and three-dimensional chromatin interactions of seven brain regions across a cohort of 39 cognitively healthy individuals. Single-cell chromatin accessibility profiling of 70,631 cells from six of these brain regions identifies 24 distinct cell clusters and 359,022 cell type-specific regulatory elements, capturing the regulatory diversity of the adult brain. We develop a machine learning classifier to integrate this multi-omic framework and predict dozens of functional single nucleotide polymorphisms (SNPs), nominating gene and cellular targets for previously orphaned GWAS loci. These predictions both inform well-studied disease-relevant genes, such as BIN1 in microglia for Alzheimer’s disease (AD) and reveal novel gene-disease associations, such as STAB1 in microglia and MAL in oligodendrocytes for Parkinson’s disease (PD). Moreover, we dissect the complex inverted haplotype of the MAPT (encoding tau) PD risk locus, identifying ectopic enhancer-gene contacts in neurons that increase MAPT expression and may mediate this disease association. This work greatly expands our understanding of inherited variation in AD and PD and provides a roadmap for the epigenomic dissection of noncoding regulatory variation in disease.

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Single cell epigenomic atlas of the developing human brain and organoids

10.1101/2019.12.30.891549 ◽

2019 ◽

Cited By ~ 5

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.

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Single-cell chromatin and gene-regulatory dynamics of mouse nephron progenitors

10.1101/2021.10.13.464099 ◽

2021 ◽

Author(s):

Sylvia Hilliard ◽

Giovane Tortelote ◽

Hongbing Liu ◽

Chao-Hui Chen ◽

Samir S El-Dahr

Keyword(s):

Transcription Factors ◽

Single Cell ◽

Cell Fate ◽

Cell Types ◽

Regulatory Elements ◽

Chromatin Accessibility ◽

Bulk Analysis ◽

Disease States ◽

Differentiated Cells ◽

Nephron Progenitors

Background: Cis-regulatory elements (CREs), such as enhancers and promoters, and their cognate transcription factors play a central role in cell fate specification. Bulk analysis of CREs has provided insights into gene regulation in nephron progenitor cells (NPCs). However, the cellular resolution required to unravel the dynamic changes in regulatory elements associated with cell fate choices remains to be defined. Methods: We integrated single-cell chromatin accessibility (scATAC-seq) and gene expression (scRNA-seq) in embryonic E16.5 (self-renewing) and postnatal P2 (primed) mouse Six2GFP NPCs. This analysis revealed NPC diversity and identified candidate CREs. To validate these findings and gain additional insights into more differentiated cell types, we performed a multiome analysis of E16.5 and P2 kidneys. Results: CRE accessibility recovered the diverse states of NPCs and precursors of differentiated cells. Single-cell types such as podocytes, proximal and distal precursors are marked by differentially accessible CREs. Domains of regulatory chromatin as defined by rich CRE-gene associations identified NPC fate-determining transcription factors (TF). Likewise, key TF expression correlates well with its regulon activity. Young NPCs exhibited enrichment in accessible motifs for bHLH, homeobox, and Forkhead TFs, while older NPCs were enriched in AP-1, HNF1, and HNF4 motif activity. A subset of Forkhead factors exhibiting high chromatin activity in podocyte precursors. Conclusion: Defining the regulatory landscape of nephrogenesis at single-cell resolution informs the basic mechanisms of nephrogenesis and provides a foundation for future studies in disease states characterized by abnormal nephrogenesis.

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The Roles of Chromatin Accessibility in Regulating the Candida albicans White-Opaque Phenotypic Switch

Journal of Fungi ◽

10.3390/jof7010037 ◽

2021 ◽

Vol 7 (1) ◽

pp. 37

Author(s):

Mohammad N. Qasim ◽

Ashley Valle Arevalo ◽

Clarissa J. Nobile ◽

Aaron D. Hernday

Keyword(s):

Candida Albicans ◽

Cell Fate ◽

Cell Types ◽

Chromatin Accessibility ◽

Phenotypic Switching ◽

Phenotypic Switch ◽

Chromatin Remodelers ◽

Environmental Signals ◽

Cell Fate Decisions ◽

Simple Model System

Candida albicans, a diploid polymorphic fungus, has evolved a unique heritable epigenetic program that enables reversible phenotypic switching between two cell types, referred to as “white” and “opaque”. These cell types are established and maintained by distinct transcriptional programs that lead to differences in metabolic preferences, mating competencies, cellular morphologies, responses to environmental signals, interactions with the host innate immune system, and expression of approximately 20% of genes in the genome. Transcription factors (defined as sequence specific DNA-binding proteins) that regulate the establishment and heritable maintenance of the white and opaque cell types have been a primary focus of investigation in the field; however, other factors that impact chromatin accessibility, such as histone modifying enzymes, chromatin remodelers, and histone chaperone complexes, also modulate the dynamics of the white-opaque switch and have been much less studied to date. Overall, the white-opaque switch represents an attractive and relatively “simple” model system for understanding the logic and regulatory mechanisms by which heritable cell fate decisions are determined in higher eukaryotes. Here we review recent discoveries on the roles of chromatin accessibility in regulating the C. albicans white-opaque phenotypic switch.

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