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 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 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 Targeted chromatin conformation analysis identifies novel distal neural enhancers of ZEB2 in pluripotent stem cell differentiation

2020 ◽  
Vol 29 (15) ◽  
pp. 2535-2550
Author(s):  
Judith C Birkhoff ◽  
Rutger W W Brouwer ◽  
Petros Kolovos ◽  
Anne L Korporaal ◽  
Ana Bermejo-Santos ◽  
...  
Keyword(s):  
Cell Fate ◽  
Stem Cell Differentiation ◽  
Cell Types ◽  
Regulatory Elements ◽  
Specific Cell ◽  
Mrna Levels ◽  
Gene Desert ◽  
Protein Coding ◽  
Cell Fate Decisions ◽  
In Cis

Abstract The transcription factor zinc finger E-box binding protein 2 (ZEB2) controls embryonic and adult cell fate decisions and cellular maturation in many stem/progenitor cell types. Defects in these processes in specific cell types underlie several aspects of Mowat–Wilson syndrome (MOWS), which is caused by ZEB2 haplo-insufficiency. Human ZEB2, like mouse Zeb2, is located on chromosome 2 downstream of a ±3.5 Mb-long gene-desert, lacking any protein-coding gene. Using temporal targeted chromatin capture (T2C), we show major chromatin structural changes based on mapping in-cis proximities between the ZEB2 promoter and this gene desert during neural differentiation of human-induced pluripotent stem cells, including at early neuroprogenitor cell (NPC)/rosette state, where ZEB2 mRNA levels increase significantly. Combining T2C with histone-3 acetylation mapping, we identified three novel candidate enhancers about 500 kb upstream of the ZEB2 transcription start site. Functional luciferase-based assays in heterologous cells and NPCs reveal co-operation between these three enhancers. This study is the first to document in-cis Regulatory Elements located in ZEB2’s gene desert. The results further show the usability of T2C for future studies of ZEB2 REs in differentiation and maturation of multiple cell types and the molecular characterization of newly identified MOWS patients that lack mutations in ZEB2 protein-coding exons.

scholarly journals SUMO maintains the chromatin environment of human induced pluripotent stem cells

2020 ◽  
Author(s):  
Barbara Mojsa ◽  
Michael H. Tatham ◽  
Lindsay Davidson ◽  
Magda Liczmanska ◽  
Jane E. Wright ◽  
...  
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Cell Fate ◽  
Pluripotent Stem Cells ◽  
Transcriptional Start Site ◽  
Chromatin Accessibility ◽  
Covalent Attachment ◽  
Cell Fate Decisions ◽  
Sumo Modification ◽  
Induced Pluripotent

AbstractPluripotent stem cells represent a powerful system to identify the mechanisms governing cell fate decisions during early mammalian development. Covalent attachment of the Small Ubiquitin Like Modifier (SUMO) to proteins has emerged as an important factor in stem cell maintenance. Here we show that SUMO is required to maintain stem cells in their pluripotent state and identify many chromatin-associated proteins as bona fide SUMO substrates in human induced pluripotent stem cells (hiPSCs). Loss of SUMO increases chromatin accessibility and expression of long non-coding RNAs and human endogenous retroviral elements, indicating a role for the SUMO modification of SETDB1 and a large TRIM28 centric network of zinc finger proteins in silencing of these elements. While most protein coding genes are unaffected, the Preferentially Expressed Antigen of Melanoma (PRAME) gene locus becomes more accessible and transcription is dramatically increased after inhibition of SUMO modification. When PRAME is silent, a peak of SUMO over the transcriptional start site overlaps with ChIP-seq peaks for cohesin, RNA pol II, CTCF and ZNF143, with the latter two heavily modified by SUMO. These associations suggest that silencing of the PRAME gene is maintained by the influence of SUMO on higher order chromatin structure. Our data indicate that SUMO modification plays an important role in hiPSCs by repressing genes that disrupt pluripotency networks or drive differentiation.

scholarly journals Cis-regulatory similarities in the zebrafish and human pancreas uncover potential disease-related enhancers

2020 ◽  
Author(s):  
R. Bordeira-Carriço ◽  
J. Teixeira ◽  
M. Duque ◽  
M. Galhardo ◽  
D. Ribeiro ◽  
...  
Keyword(s):  
Gene Expression ◽  
Regulatory Elements ◽  
Suppressor Gene ◽  
Chromatin Accessibility ◽  
Tumour Suppressor Gene ◽  
Tumour Suppressor ◽  
Regulatory Sequences ◽  
Human Pancreas ◽  
Distal Enhancer

Introductory paragraphThe pancreas is a central organ for human diseases that have a dramatic societal burden, such as pancreatic cancer and diabetes1,2. Non-coding cis-regulatory elements (CREs) of DNA control gene expression3,4, being required for proper pancreas function. Most disease-associated alleles5,6 are non-coding, often overlapping with CREs5, suggesting that alterations in these regulatory sequences contribute to human pancreatic diseases by impairing gene expression. However, functional testing of CREs in vivo is not fully explored. Here we analysed histone modifications, transcription, chromatin accessibility and interactions, to identify zebrafish pancreas CREs and their human functional equivalents, uncovering disease-associated sequences across species. We found a human pancreatic enhancer whose deletion impairs the tumour suppressor gene ARID1A expression, conferring a potential tumour suppressor role to this non-coding sequence. Additionally, we identified a zebrafish ptf1a distal enhancer which deletion generates pancreatic agenesis, demonstrating the causality of this condition in humans7 and the interspecies functional equivalency of enhancers.

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

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.

scholarly journals Defective Calmodulin-Mediated Nuclear Transport of the Sex-Determining Region of the Y Chromosome (SRY) in XY Sex Reversal

2005 ◽  
Vol 19 (7) ◽  
pp. 1884-1892 ◽  
Author(s):  
Helena Sim ◽  
Kieran Rimmer ◽  
Sabine Kelly ◽  
Louisa M. Ludbrook ◽  
Andrew H. A. Clayton ◽  
...  
Keyword(s):  
Y Chromosome ◽  
Nuclear Import ◽  
High Mobility ◽  
Sex Reversal ◽  
High Mobility Group ◽  
Missense Mutations ◽  
Nuclear Localization Signals ◽  
Xy Sex Reversal

Abstract The sex-determining region of the Y chromosome (SRY) plays a key role in human sex determination, as mutations in SRY can cause XY sex reversal. Although some SRY missense mutations affect DNA binding and bending activities, it is unclear how others contribute to disease. The high mobility group domain of SRY has two nuclear localization signals (NLS). Sex-reversing mutations in the NLSs affect nuclear import in some patients, associated with defective importin-β binding to the C-terminal NLS (c-NLS), whereas in others, importin-β recognition is normal, suggesting the existence of an importin-β-independent nuclear import pathway. The SRY N-terminal NLS (n-NLS) binds calmodulin (CaM) in vitro, and here we show that this protein interaction is reduced in vivo by calmidazolium, a CaM antagonist. In calmidazolium-treated cells, the dramatic reduction in nuclear entry of SRY and an SRY-c-NLS mutant was not observed for two SRY-n-NLS mutants. Fluorescence spectroscopy studies reveal an unusual conformation of SRY.CaM complexes formed by the two n-NLS mutants. Thus, CaM may be involved directly in SRY nuclear import during gonadal development, and disruption of SRY.CaM recognition could underlie XY sex reversal. Given that the CaM-binding region of SRY is well-conserved among high mobility group box proteins, CaM-dependent nuclear import may underlie additional disease states.

scholarly journals The LINC complex transmits integrin-dependent tension to the nuclear lamina and represses epidermal differentiation

2020 ◽  
Author(s):  
Emma Carley ◽  
Rachel K. Stewart ◽  
Abigail Zieman ◽  
Iman Jalilian ◽  
Diane. E. King ◽  
...  
Keyword(s):  
Cell Fate ◽  
Control Cell ◽  
Nuclear Lamina ◽  
Epidermal Differentiation ◽  
Keratinocyte Differentiation ◽  
Linc Complex ◽  
Force Transmission ◽  
Cell Fate Decisions

AbstractWhile the mechanisms by which chemical signals control cell fate have been well studied, how mechanical inputs impact cell fate decisions are not well understood. Here, using the well-defined system of keratinocyte differentiation in the skin, we examine whether and how direct force transmission to the nucleus regulates epidermal cell fate. Using a molecular biosensor, we find that tension on the nucleus through Linker of Nucleoskeleton and Cytoskeleton (LINC) complexes requires integrin engagement in undifferentiated epidermal stem cells, and is released during differentiation concomitant with decreased tension on A-type lamins. LINC complex ablation in mice reveals that LINC complexes are required to repress epidermal differentiation in vivo and in vitro and influence accessibility of epidermal differentiation genes, suggesting that force transduction from engaged integrins to the nucleus plays a role in maintaining keratinocyte progenitors. This work reveals a direct mechanotransduction pathway capable of relaying adhesion-specific signals to regulate cell fate.

scholarly journals Integrative epigenomic and high-throughput functional enhancer profiling reveals determinants of enhancer heterogeneity in gastric cancer

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Taotao Sheng ◽  
Shamaine Wei Ting Ho ◽  
Wen Fong Ooi ◽  
Chang Xu ◽  
Manjie Xing ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Histone Modification ◽  
Cell Fate ◽  
Copy Number ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Specific Gene ◽  
Functional Assay ◽  
Enhancer Activity

Abstract Background Enhancers are distal cis-regulatory elements required for cell-specific gene expression and cell fate determination. In cancer, enhancer variation has been proposed as a major cause of inter-patient heterogeneity—however, most predicted enhancer regions remain to be functionally tested. Methods We analyzed 132 epigenomic histone modification profiles of 18 primary gastric cancer (GC) samples, 18 normal gastric tissues, and 28 GC cell lines using Nano-ChIP-seq technology. We applied Capture-based Self-Transcribing Active Regulatory Region sequencing (CapSTARR-seq) to assess functional enhancer activity. An Activity-by-contact (ABC) model was employed to explore the effects of histone acetylation and CapSTARR-seq levels on enhancer-promoter interactions. Results We report a comprehensive catalog of 75,730 recurrent predicted enhancers, the majority of which are GC-associated in vivo (> 50,000) and associated with lower somatic mutation rates inferred by whole-genome sequencing. Applying CapSTARR-seq to the enhancer catalog, we observed significant correlations between CapSTARR-seq functional activity and H3K27ac/H3K4me1 levels. Super-enhancer regions exhibited increased CapSTARR-seq signals compared to regular enhancers, even when decoupled from native chromatin contexture. We show that combining histone modification and CapSTARR-seq functional enhancer data improves the prediction of enhancer-promoter interactions and pinpointing of germline single nucleotide polymorphisms (SNPs), somatic copy number alterations (SCNAs), and trans-acting TFs involved in GC expression. We identified cancer-relevant genes (ING1, ARL4C) whose expression between patients is influenced by enhancer differences in genomic copy number and germline SNPs, and HNF4α as a master trans-acting factor associated with GC enhancer heterogeneity. Conclusions Our results indicate that combining histone modification and functional assay data may provide a more accurate metric to assess enhancer activity than either platform individually, providing insights into the relative contribution of genetic (cis) and regulatory (trans) mechanisms to GC enhancer functional heterogeneity.

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