scholarly journals Dynamics of Cardiomyocyte Transcriptome and Chromatin Landscape Demarcates Key Events of Heart Development

2018 ◽  
Author(s):  
Michal Pawlak ◽  
Katarzyna Z. Kedzierska ◽  
Maciej Migdal ◽  
Karim Abu Nahia ◽  
Jordan A. Ramilowski ◽  
...  
Keyword(s):  
Heart Development ◽  
Regulatory Networks ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Proximal Promoter ◽  
Global Changes ◽  
Open Chromatin ◽  
Sequence Motifs ◽  
Loss Of Function ◽  
Heart Looping

ABSTRACTThe development of an organ involves dynamic regulation of gene transcription and complex multipathway interactions. To better understand transcriptional regulatory mechanism driving heart development and the consequences of its disruption, we isolated cardiomyocytes (CMs) from wild-type zebrafish embryos at 24, 48 and 72 hours post fertilization corresponding to heart looping, chamber formation and heart maturation, and from mutant lines carrying loss-of-function mutations in gata5, tbx5a and hand2, transcription factors (TFs) required for proper heart development. The integration of CM transcriptomics (RNA-seq) and genome-wide chromatin accessibility maps (ATAC-seq) unravelled dynamic regulatory networks driving crucial events of heart development. These networks contained key cardiac TFs including Gata5/6, Nkx2.5, Tbx5/20, and Hand2, and are associated with open chromatin regions enriched for DNA sequence motifs belonging to the family of the corresponding TFs. These networks were disrupted in cardiac TF mutants, indicating their importance in proper heart development. The most prominent gene expression changes, which correlated with chromatin accessibility modifications within their proximal promoter regions, occurred between heart looping and chamber formation, and were associated with metabolic and hematopoietic/cardiac switch during CM maturation. Furthermore, loss of function of cardiac TFs Gata5, Tbx5a, and Hand2 affected the cardiac regulatory networks and caused global changes in chromatin accessibility profile. Among regions with differential chromatin accessibility in mutants were highly conserved non-coding elements which represent putative cis regulatory elements with potential role in heart development and disease. Altogether, our results revealed the dynamic regulatory landscape at key stages of heart development and identified molecular drivers of heart morphogenesis.

scholarly journals Characterising open chromatin in chick embryos identifies cis-regulatory elements important for paraxial mesoderm formation and axis extension

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Gi Fay Mok ◽  
Leighton Folkes ◽  
Shannon A. Weldon ◽  
Eirini Maniou ◽  
Victor Martinez-Heredia ◽  
...  
Keyword(s):  
Regulatory Networks ◽  
Integrated Approach ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Paraxial Mesoderm ◽  
Open Chromatin ◽  
Regulatory Circuits ◽  
Axis Extension ◽  
Footprint Analysis

AbstractSomites arising from paraxial mesoderm are a hallmark of the segmented vertebrate body plan. They form sequentially during axis extension and generate musculoskeletal cell lineages. How paraxial mesoderm becomes regionalised along the axis and how this correlates with dynamic changes of chromatin accessibility and the transcriptome remains unknown. Here, we report a spatiotemporal series of ATAC-seq and RNA-seq along the chick embryonic axis. Footprint analysis shows differential coverage of binding sites for several key transcription factors, including CDX2, LEF1 and members of HOX clusters. Associating accessible chromatin with nearby expressed genes identifies cis-regulatory elements (CRE) for TCF15 and MEOX1. We determine their spatiotemporal activity and evolutionary conservation in Xenopus and human. Epigenome silencing of endogenous CREs disrupts TCF15 and MEOX1 gene expression and recapitulates phenotypic abnormalities of anterior–posterior axis extension. Our integrated approach allows dissection of paraxial mesoderm regulatory circuits in vivo and has implications for investigating gene regulatory networks.

scholarly journals Coupled single-cell CRISPR screening and epigenomic profiling reveals causal gene regulatory networks

2018 ◽  
Author(s):  
Adam J. Rubin ◽  
Kevin R. Parker ◽  
Ansuman T. Satpathy ◽  
Yanyan Qi ◽  
Beijing Wu ◽  
...  
Keyword(s):  
Gene Regulatory Networks ◽  
Regulatory Networks ◽  
Single Cells ◽  
Nucleosome Positioning ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Open Chromatin ◽  
General Strategy ◽  
List Type ◽  
Gene Regulatory

SummaryHere we present Perturb-ATAC, a method which combines multiplexed CRISPR interference or knockout with genome-wide chromatin accessibility profiling in single cells, based on the simultaneous detection of CRISPR guide RNAs and open chromatin sites by assay of transposase-accessible chromatin with sequencing (ATAC-seq). We applied Perturb-ATAC to transcription factors (TFs), chromatin-modifying factors, and noncoding RNAs (ncRNAs) in ∼4,300 single cells, encompassing more than 63 unique genotype-phenotype relationships. Perturb-ATAC in human B lymphocytes uncovered regulators of chromatin accessibility, TF occupancy, and nucleosome positioning, and identified a hierarchical organization of TFs that govern B cell state, variation, and disease-associatedcis-regulatory elements. Perturb-ATAC in primary human epidermal cells revealed three sequential modules ofcis-elements that specify keratinocyte fate, orchestrated by the TFs JUNB, KLF4, ZNF750, CEBPA, and EHF. Combinatorial deletion of all pairs of these TFs uncovered their epistatic relationships and highlighted genomic co-localization as a basis for synergistic interactions. Thus, Perturb-ATAC is a powerful and general strategy to dissect gene regulatory networks in development and disease.HighlightsA new method for simultaneous measurement of CRISPR perturbations and chromatin state in single cells.Perturb-ATAC reveals regulatory factors that controlcis-element accessibility,trans-factor occupancy, and nucleosome positioning.Perturb-ATAC reveals regulatory modules of coordinatedtrans-factor activity in B lymphoblasts.Keratinocyte differentiation is orchestrated by synergistic activities of co-binding TFs oncis-elements.

scholarly journals EPCO-31. EPIGENOMIC INTRATUMORAL HETEROGENEITY OF GLIOBLASTOMA IN THREE-DIMENSIONAL SPACE

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii76-ii76
Author(s):  
Radhika Mathur ◽  
Sriranga Iyyanki ◽  
Stephanie Hilz ◽  
Chibo Hong ◽  
Joanna Phillips ◽  
...  
Keyword(s):  
Spatial Organization ◽  
Dimensional Space ◽  
Three Dimensional ◽  
Spatial Location ◽  
Therapy Resistance ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Intratumoral Heterogeneity ◽  
Tumor Evolution ◽  
Open Chromatin

Abstract Treatment failure in glioblastoma is often attributed to intratumoral heterogeneity (ITH), which fosters tumor evolution and generation of therapy-resistant clones. While ITH in glioblastoma has been well-characterized at the genomic and transcriptomic levels, the extent of ITH at the epigenomic level and its biological and clinical significance are not well understood. In collaboration with neurosurgeons, neuropathologists, and biomedical imaging experts, we have established a novel topographical approach towards characterizing epigenomic ITH in three-dimensional (3-D) space. We utilize pre-operative MRI scans to define tumor volume and then utilize 3-D surgical neuro-navigation to intra-operatively acquire 10+ samples representing maximal anatomical diversity. The precise spatial location of each sample is mapped by 3-D coordinates, enabling tumors to be visualized in 360-degrees and providing unprecedented insight into their spatial organization and patterning. For each sample, we conduct assay for transposase-accessible chromatin using sequencing (ATAC-Seq), which provides information on the genomic locations of open chromatin, DNA-binding proteins, and individual nucleosomes at nucleotide resolution. We additionally conduct whole-exome sequencing and RNA sequencing for each spatially mapped sample. Integrative analysis of these datasets reveals distinct patterns of chromatin accessibility within glioblastoma tumors, as well as their associations with genetically defined clonal expansions. Our analysis further reveals how differences in chromatin accessibility within tumors reflect underlying transcription factor activity at gene regulatory elements, including both promoters and enhancers, and drive expression of particular gene expression sets, including neuronal and immune programs. Collectively, this work provides the most comprehensive characterization of epigenomic ITH to date, establishing its importance for driving tumor evolution and therapy resistance in glioblastoma. As a resource for further investigation, we have provided our datasets on an interactive data sharing platform – The 3D Glioma Atlas – that enables 360-degree visualization of both genomic and epigenomic ITH.

scholarly journals ATRT-07. DEFINING LOST AND GAINED TRANSCRIPTIONAL REGULATORY NETWORKS IN ATYPICAL TERATOID RHABDOID TUMOR

2021 ◽  
Vol 23 (Supplement_1) ◽  
pp. i2-i2
Author(s):  
Cody Nesvick ◽  
Liang Zhang ◽  
Alex Wixom ◽  
Feda Hamdan ◽  
Steven Johnsen ◽  
...  
Keyword(s):  
Cell Lines ◽  
Regulatory Networks ◽  
Regulatory Elements ◽  
Rhabdoid Tumor ◽  
Allelic Loss ◽  
Global Changes ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Transcriptional Regulatory Networks ◽  
Transcriptional Regulatory ◽  
Atypical Teratoid

Abstract Atypical teratoid rhabdoid tumor (ATRT) is a central nervous system cancer of infancy and early childhood that may occur anywhere along the neuraxis and is associated with a high rate of mortality. While contemporary multimodal therapeutic approaches have significantly improved overall survival, targeted therapy remains elusive, and treatment is often associated with significant morbidity. ATRT is unique in its genomic stability, with the only recurrent genetic abnormality being bi-allelic loss of the SMARCB1 gene, which encodes a core subunit of the BAF chromatin remodeling complex. The epigenetic mechanisms by which SMARCB1 loss leads to tumorigenesis are not yet well-defined and addressing this gap in understanding is necessary for creating efficacious, targeted therapeutics. To better understand the epigenetic features gained and lost in ATRT, we re-expressed SMARCB1 in a library of patient-derived and established ATRT cell lines of multiple molecular subtypes. SMARCB1 restoration significantly reduced or eliminated the proliferative and clonogenic capacity of each cell line. We performed assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-Seq) and RNA sequencing (RNA-Seq) to define putative transcriptional regulatory networks that are gained and lost in ATRT. SMARCB1 restoration was associated with global changes in chromatin openness consistent with the creation of new regulatory elements throughout the genome, and these were associated with induction of a diverse developmental transcriptional signature. Motif enrichment analysis of regions with increased accessibility defined a small but consistent number of centrally enriched transcription factor motifs across cell lines indicative of putative pioneer factors whose functions may be lost in ATRT. Pertinent chromatin immunoprecipitation with sequencing (ChIP-Seq) data will be discussed in the context of lost and gained transcriptional regulatory networks in ATRT and normal cellular development.

scholarly journals Enhanced TF binding site maps improve regulatory networks learned from accessible chromatin data

2019 ◽  
Author(s):  
Shubhada R. Kulkarni ◽  
D. Marc Jones ◽  
Klaas Vandepoele
Keyword(s):  
Binding Site ◽  
Dna Sequences ◽  
Regulatory Networks ◽  
Transcriptional Control ◽  
Chromatin Accessibility ◽  
Systematic Evaluation ◽  
Open Chromatin ◽  
Ensemble Approach ◽  
Complete Cell ◽  
Regulatory Dna

ABSTRACTDetermining where transcription factors (TF) bind in genomes provides insights into which transcriptional programs are active across organs, tissue types, and environmental conditions. Recent advances in high-throughput profiling of regulatory DNA have yielded large amounts of information about chromatin accessibility. Interpreting the functional significance of these datasets requires knowledge of which regulators are likely to bind these regions. This can be achieved by using information about TF binding preferences, or motifs, to identify TF binding events that are likely to be functional. Although different approaches exist to map motifs to DNA sequences, a systematic evaluation of these tools in plants is missing. Here we compare four motif mapping tools widely used in the Arabidopsis research community and evaluate their performance using chromatin immunoprecipitation datasets for 40 TFs. Downstream gene regulatory network (GRN) reconstruction was found to be sensitive to the motif mapper used. We further show that the low recall of FIMO, one of the most frequently used motif mapping tools, can be overcome by using an Ensemble approach, which combines results from different mapping tools. Several examples are provided demonstrating how the Ensemble approach extends our view on transcriptional control for TFs active in different biological processes. Finally, a new protocol is presented to efficiently derive more complete cell type-specific GRNs through the integrative analysis of open chromatin regions, known binding site information, and expression datasets.

scholarly journals Chromatin accessibility and regulatory vocabulary across indicine cattle tissues

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Pâmela A. Alexandre ◽  
Marina Naval-Sánchez ◽  
Moira Menzies ◽  
Loan T. Nguyen ◽  
Laercio R. Porto-Neto ◽  
...  
Keyword(s):  
Motif Discovery ◽  
Target Genes ◽  
Developmental Stages ◽  
Bos Taurus ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Model Organisms ◽  
Open Chromatin ◽  
Health And Disease ◽  
Collaborative Efforts

Abstract Background Spatiotemporal changes in the chromatin accessibility landscape are essential to cell differentiation, development, health, and disease. The quest of identifying regulatory elements in open chromatin regions across different tissues and developmental stages is led by large international collaborative efforts mostly focusing on model organisms, such as ENCODE. Recently, the Functional Annotation of Animal Genomes (FAANG) has been established to unravel the regulatory elements in non-model organisms, including cattle. Now, we can transition from prediction to validation by experimentally identifying the regulatory elements in tropical indicine cattle. The identification of regulatory elements, their annotation and comparison with the taurine counterpart, holds high promise to link regulatory regions to adaptability traits and improve animal productivity and welfare. Results We generate open chromatin profiles for liver, muscle, and hypothalamus of indicine cattle through ATAC-seq. Using robust methods for motif discovery, motif enrichment and transcription factor binding sites, we identify potential master regulators of the epigenomic profile in these three tissues, namely HNF4, MEF2, and SOX factors, respectively. Integration with transcriptomic data allows us to confirm some of their target genes. Finally, by comparing our results with Bos taurus data we identify potential indicine-specific open chromatin regions and overlaps with indicine selective sweeps. Conclusions Our findings provide insights into the identification and analysis of regulatory elements in non-model organisms, the evolution of regulatory elements within two cattle subspecies as well as having an immediate impact on the animal genetics community in particular for a relevant productive species such as tropical cattle.

scholarly journals Functional Inference of Gene Regulation using Single-Cell Multi-Omics

2021 ◽  
Author(s):  
Vinay K Kartha ◽  
Fabiana M Duarte ◽  
Yan Hu ◽  
Sai Ma ◽  
Jennifer G Chew ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Cell ◽  
Regulatory Networks ◽  
Immunological Response ◽  
Cell Types ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Human Blood Cells ◽  
Functional Inference ◽  
Omic Data

Cells require coordinated control over gene expression when responding to environmental stimuli. Here, we apply scATAC-seq and scRNA-seq in resting and stimulated human blood cells. Collectively, we generate ~91,000 single-cell profiles, allowing us to probe the cis -regulatory landscape of immunological response across cell types, stimuli and time. Advancing tools to integrate multi-omic data, we develop FigR - a framework to computationally pair scATAC-seq with scRNA-seq cells, connect distal cis -regulatory elements to genes, and infer gene regulatory networks (GRNs) to identify candidate TF regulators. Utilizing these paired multi-omic data, we define Domains of Regulatory Chromatin (DORCs) of immune stimulation and find that cells alter chromatin accessibility prior to production of gene expression at time scales of minutes. Further, the construction of the stimulation GRN elucidates TF activity at disease-associated DORCs. Overall, FigR enables the elucidation of regulatory interactions across single-cell data, providing new opportunities to understand the function of cells within tissues.

scholarly journals The SEEL Motif and Members of the MYB-related REVEILLE Transcription Factor Family are Important for the Expression of LORELEI in the Synergid Cells of the Arabidopsis Female Gametophyte

2021 ◽  
Author(s):  
Jennifer A. Noble ◽  
Alex Seddon ◽  
Sahra Uygun ◽  
Steven E. Smith ◽  
Shin-Han Shiu ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Pollen Tube ◽  
Female Gametophyte ◽  
Regulatory Networks ◽  
Clock Genes ◽  
Regulatory Element ◽  
Regulatory Elements ◽  
Loss Of Function ◽  
Seed Formation ◽  
Synergid Cell

Synergid cells in the micropylar end of the female gametophyte are required for critical cell-cell signaling interactions between the pollen tube and the ovule that precede double fertilization and seed formation in flowering plants. LORELEI (LRE) encodes a GPI-anchored protein that is expressed primarily in the synergid cells, and together with FERONIA, a receptor-like kinase, it controls pollen tube reception by the receptive synergid cell. Still, how LRE expression is controlled in synergid cells remains poorly characterized. We identified candidate cis-regulatory elements enriched in LRE and other synergid cell-expressed genes. One of the candidate motifs (TAATATCT) in the LRE promoter was an uncharacterized variant of the Evening Element motif that we named as the Short Evening Element-like (SEEL) motif. Deletion or point mutations in the SEEL motif of the LRE promoter resulted in decreased reporter expression in synergid cells, demonstrating that the SEEL motif is important for expression of LRE in synergid cells. Additionally, we found that LRE expression is decreased in the loss of function mutants of REVEILLE (RVE) transcription factors, which are clock genes known to bind the SEEL and other closely related motifs. We propose that RVE transcription factors regulate LRE expression in synergid cells by binding to the SEEL motif in the LRE promoter. Identification of a cis-regulatory element and transcription factors involved in the expression of LRE will serve as a foundation to characterize the gene regulatory networks in synergid cells and investigate the potential connection between circadian rhythm and fertilization.

scholarly journals PyBoost: A parallelized Python implementation of 2D boosting with hierarchies

2017 ◽  
Author(s):  
Peyton G. Greenside ◽  
Nadine Hussami ◽  
Jessica Chang ◽  
Anshul Kundaje
Keyword(s):  
Gene Expression ◽  
Regulatory Networks ◽  
Stem Cell Differentiation ◽  
Cell Types ◽  
Chromatin Accessibility ◽  
Sequence Motifs ◽  
Specific Sequence ◽  
Tail Regeneration ◽  
Perturbation Data ◽  
Time Courses

AbstractMotivation:Gene expression is controlled by networks of transcription factors that bind specific sequence motifs in regulatory DNA elements such as promoters and enhancers. GeneClass is a boosting-based algorithm that learns gene regulatory networks from complementary paired feature sets such as transcription factor expression levels and binding motifs across conditions. This algorithm can be used to predict functional genomics measures of cell state, such as gene expression and chromatin accessibility, in different cellular conditions. We present a parallelized, Python-based implementation of GeneClass, called PyBoost, along with a novel hierarchical implementation of the algorithm, called HiBoost. HiBoost allows regulatory logic to be constrained to a hierarchical group of conditions or cell types. The software can be used to dissect differentiation cascades, time courses or other perturbation data that naturally form a hierarchy or trajectory. We demonstrate the application of PyBoost and HiBoost to learn regulators of tadpole tail regeneration and hematopoeitic stem cell differentiation and validate learned regulators through an inducible CRISPR system.Availability:The implementation is publicly available here:https://github.com/kundajelab/boosting2D/.

scholarly journals Characterization and Activity Analyses of the FLOWERING LOCUS T Promoter in Gossypium Hirsutum

2019 ◽  
Vol 20 (19) ◽  
pp. 4769 ◽  
Author(s):  
Na Sang ◽  
Darun Cai ◽  
Chao Li ◽  
Yuqiang Sun ◽  
Xianzhong Huang
Keyword(s):  
Gossypium Hirsutum ◽  
Regulatory Elements ◽  
Proximal Promoter ◽  
Activity Levels ◽  
Loss Of Function ◽  
Flowering Locus T ◽  
Promoter Regions ◽  
Core Motif ◽  
Late Flowering ◽  
Flowering Locus

Flowering transition is a crucial development process in cotton (Gossypium hirsutum L.), and the flowering time is closely correlated with the timing of FLOWERING LOCUS T (FT) expression. However, the mechanism underlying the coordination of various cis-regulatory elements in the FT promoter of cotton has not been determined. In this study, a 5.9-kb promoter of FT was identified from cotton. A bioinformatics analysis showed that multiple insertion–deletion sites existed in the 5.9-kb promoter. Different expression levels of a reporter gene, and the induction by sequential deletions in GhFT promoter, demonstrated that 1.8-kb of the GhFT promoter was stronger than 4.2-, 4.8-, and 5.9-kb promoter fragments. The binding sites of the CONSTANS (CO) and NUCLEAR FACTOR Y transcription factors were located within the 1.0-kb sequence upstream of the FT transcription start site. A large number of repeat segments were identified in proximal promoter regions (−1.1 to −1.4 kb). A complementation analysis of deletion constructs between 1.0 and 1.8 kb of G. hirsutum, Gossypium arboretum, and Gossypium raimondii FT promoters revealed that the 1.0-kb fragment significantly rescued the late-flowering phenotype of the Arabidopsis FT loss-of-function mutant ft-10, whereas the 1.8-kb promoter only slightly rescued the late-flowering phenotype. Furthermore, the conserved CORE motif in the cotton FT promoter is an atypical TGTG(N2-3)ATG, but the number of arbitrary bases between TGTG and ATG is uncertain. Thus, the proximal FT promoter region might play an important role affecting the activity levels of FT promoters in cotton flowering.

Sign in / Sign up

Export Citation Format

Share Document