scholarly journals Chromatin accessibility dynamics reveal novel functional enhancers in C. elegans

2016 ◽  
Author(s):  
Aaron C. Daugherty ◽  
Robin Yeo ◽  
Jason D. Buenrostro ◽  
William J. Greenleaf ◽  
Anshul Kundaje ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Developmental Stages ◽  
Model Organism ◽  
High Sensitivity ◽  
Chromatin Accessibility ◽  
Chromatin Dynamics ◽  
C Elegans ◽  
Crucial Component ◽  
Organismal Development

AbstractChromatin accessibility, a crucial component of genome regulation, has primarily been studied in homogeneous and simple systems, such as isolated cell populations or early-development models. Whether chromatin accessibility can be assessed in complex, dynamic systems in vivo with high sensitivity remains largely unexplored. In this study, we use ATAC-seq to identify chromatin accessibility changes in a whole animal, the model organism C. elegans, from embryogenesis to adulthood. Chromatin accessibility changes between developmental stages are highly reproducible, recapitulate histone modification changes, and reveal key regulatory aspects of the epigenomic landscape throughout organismal development. We find that over 5,000 distal non-coding regions exhibit dynamic changes in chromatin accessibility between developmental stages, and could thereby represent putative enhancers. When tested in vivo, several of these putative enhancers indeed drive novel cell-type-and temporal-specific patterns of expression. Finally, by integrating transcription factor binding motifs in a machine learning framework, we identify EOR-1 as a unique transcription factor that may regulate chromatin dynamics during development. Our study provides a unique resource for C. elegans, a system in which the prevalence and importance of enhancers remains poorly characterized, and demonstrates the power of using whole organism chromatin accessibility to identify novel regulatory regions in complex systems.

scholarly journals Dynamic neurotransmitter specific transcription factor expression profiles during Drosophila development

2019 ◽  
Author(s):  
Alicia Estacio-Gómez ◽  
Amira Hassan ◽  
Emma Walmsley ◽  
Lily Wong Le ◽  
Tony D. Southall
Keyword(s):  
Transcription Factor ◽  
Transcription Factors ◽  
Developmental Stages ◽  
Wide Spectrum ◽  
Expression Profiles ◽  
Developmental Time ◽  
Adult Brain ◽  
Combinatorial Regulation ◽  
C Elegans

AbstractThe remarkable diversity of neurons in the nervous system is generated during development, when properties such as cell morphology, receptor profiles and neurotransmitter identities are specified. In order to gain a greater understanding of neurotransmitter specification we profiled the transcription state of cholinergic, GABAergic and glutamatergic neurons in vivo at three developmental time points. We identified 86 differentially expressed transcription factors that are uniquely enriched, or uniquely depleted, in a specific neurotransmitter type. Some transcription factors show a similar profile across development, others only show enrichment or depletion at specific developmental stages. Profiling of Acj6 (cholinergic enriched) and Ets65A (cholinergic depleted) binding sites in vivo reveals that they both directly bind the ChAT locus, in addition to a wide spectrum of other key neuronal differentiation genes. We also show that cholinergic enriched transcription factors are expressed in mostly non-overlapping populations in the adult brain, implying the absence of combinatorial regulation of neurotransmitter fate in this context. Furthermore, our data underlines that, similar to C. elegans, there are no simple transcription factor codes for neurotransmitter type specification.

scholarly journals Agrimonia procera Wallr. Extract Increases Stress Resistance and Prolongs Life Span in Caenorhabditis elegans via Transcription Factor DAF-16 (FoxO Orthologue)

Antioxidants ◽  
2018 ◽  
Vol 7 (12) ◽  
pp. 192 ◽  
Author(s):  
Christina Saier ◽  
Inge Gommlich ◽  
Volker Hiemann ◽  
Sabrina Baier ◽  
Karoline Koch ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Caenorhabditis Elegans ◽  
Life Span ◽  
Stress Resistance ◽  
Radical Scavenging Activity ◽  
Radical Scavenging ◽  
Model Organism ◽  
Amyloid Β ◽  
Ethanol Extract

Agrimonia procera is a pharmacologically interesting plant which is proposed to protect against various diseases due to its high amount of phytochemicals, e.g., polyphenols. However, in spite of the amount of postulated health benefits, studies concerning the mechanistic effects of Agrimonia procera are limited. Using the nematode Caenorhabditis elegans, we were able to show that an ethanol extract of Agrimonia procera herba (eAE) mediates strong antioxidative effects in the nematode: Beside a strong radical-scavenging activity, eAE reduces accumulation of reactive oxygen species (ROS) accumulation and protects against paraquat-induced oxidative stress. The extract does not protect against amyloid-β-mediated toxicity, but efficiently increases the life span (up to 12.7%), as well as the resistance to thermal stress (prolongation of survival up to 22%), of this model organism. Using nematodes deficient in the forkhead box O (FoxO)-orthologue DAF-16, we were able to demonstrate that beneficial effects of eAE on stress resistance and life span were mediated via this transcription factor. We showed antioxidative, stress-reducing, and life-prolonging effects of eAE in vivo and were able to demonstrate a molecular mechanism of this extract. These results may be important for identifying further molecular targets of eAE in humans.

scholarly journals An atlas of dynamic chromatin landscapes in mouse fetal development

Nature ◽  
2020 ◽  
Vol 583 (7818) ◽  
pp. 744-751 ◽  
Author(s):  
David U. Gorkin ◽  
Iros Barozzi ◽  
Yuan Zhao ◽  
Yanxiao Zhang ◽  
Hui Huang ◽  
...  
Keyword(s):  
Fetal Development ◽  
Target Genes ◽  
Developmental Stages ◽  
Chromatin Accessibility ◽  
Chromatin State ◽  
Mammalian Development ◽  
Mouse Fetus ◽  
Chromatin Dynamics ◽  
Mouse Tissues ◽  
Genomic Resource

AbstractThe Encyclopedia of DNA Elements (ENCODE) project has established a genomic resource for mammalian development, profiling a diverse panel of mouse tissues at 8 developmental stages from 10.5 days after conception until birth, including transcriptomes, methylomes and chromatin states. Here we systematically examined the state and accessibility of chromatin in the developing mouse fetus. In total we performed 1,128 chromatin immunoprecipitation with sequencing (ChIP–seq) assays for histone modifications and 132 assay for transposase-accessible chromatin using sequencing (ATAC–seq) assays for chromatin accessibility across 72 distinct tissue-stages. We used integrative analysis to develop a unified set of chromatin state annotations, infer the identities of dynamic enhancers and key transcriptional regulators, and characterize the relationship between chromatin state and accessibility during developmental gene regulation. We also leveraged these data to link enhancers to putative target genes and demonstrate tissue-specific enrichments of sequence variants associated with disease in humans. The mouse ENCODE data sets provide a compendium of resources for biomedical researchers and achieve, to our knowledge, the most comprehensive view of chromatin dynamics during mammalian fetal development to date.

scholarly journals C. elegans TFIIH subunit GTF-2H5/TTDA is a non-essential transcription factor indispensable for DNA repair

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Karen L. Thijssen ◽  
Melanie van der Woude ◽  
Carlota Davó-Martínez ◽  
Dick H. W. Dekkers ◽  
Mariangela Sabatella ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Nucleotide Excision Repair ◽  
Developmental Disorder ◽  
Excision Repair ◽  
C Elegans ◽  
Nucleotide Excision ◽  
Or Gene ◽  
Essential Transcription Factor

AbstractThe 10-subunit TFIIH complex is vital to transcription and nucleotide excision repair. Hereditary mutations in its smallest subunit, TTDA/GTF2H5, cause a photosensitive form of the rare developmental disorder trichothiodystrophy. Some trichothiodystrophy features are thought to be caused by subtle transcription or gene expression defects. TTDA/GTF2H5 knockout mice are not viable, making it difficult to investigate TTDA/GTF2H5 in vivo function. Here we show that deficiency of C. elegans TTDA ortholog GTF-2H5 is, however, compatible with life, in contrast to depletion of other TFIIH subunits. GTF-2H5 promotes TFIIH stability in multiple tissues and is indispensable for nucleotide excision repair, in which it facilitates recruitment of TFIIH to DNA damage. Strikingly, when transcription is challenged, gtf-2H5 embryos die due to the intrinsic TFIIH fragility in absence of GTF-2H5. These results support the idea that TTDA/GTF2H5 mutations cause transcription impairment underlying trichothiodystrophy and establish C. elegans as model for studying pathogenesis of this disease.

scholarly journals Improved single-cell ATAC-seq reveals chromatin dynamics of in vitro corticogenesis

2019 ◽  
Author(s):  
Ryan M. Mulqueen ◽  
Brooke A. DeRosa ◽  
Casey A. Thornton ◽  
Zeynep Sayar ◽  
Kristof A. Torkenczy ◽  
...  
Keyword(s):  
Single Cell ◽  
Gene Networks ◽  
Regulatory Gene ◽  
Regulatory Elements ◽  
Chromatin Accessibility ◽  
Chromatin Dynamics ◽  
Cell Library ◽  
Fate Decision

AbstractDevelopment is a complex process that requires the precise modulation of regulatory gene networks controlled through dynamic changes in the epigenome. Single-cell-omic technologies provide an avenue for understanding the mechanisms of these processes by capturing the progression of epigenetic cell states during the course of cellular differentiation using in vitro or in vivo models1. However, current single-cell epigenomic methods are limited in the information garnered per individual cell, which in turn limits their ability to measure chromatin dynamics and state shifts. Single-cell combinatorial indexing (sci-) has been applied as a strategy for identifying single-cell-omic originating libraries and removes the necessity of single-cell, single-compartment chemistry2. Here, we report an improved sci-assay for transposase accessible chromatin by sequencing (ATAC-seq), which utilizes the small molecule inhibitor Pitstop 2™ (scip-ATAC-seq)3. We demonstrate that these improvements, which theoretically could be applied to any in situ transposition method for single-cell library preparation, significantly increase the ability of transposase to enter the nucleus and generate highly complex single-cell libraries, without altering biological signal. We applied sci-ATAC-seq and scip-ATAC-seq to characterize the chromatin dynamics of developing forebrain-like organoids, an in vitro model of human corticogenesis4. Using these data, we characterized novel putative regulatory elements, compared the epigenome of the organoid model to human cortex data, generated a high-resolution pseudotemporal map of chromatin accessibility through differentiation, and measured epigenomic changes coinciding with a neurogenic fate decision point. Finally, we combined transcription factor motif accessibility with gene activity (GA) scores to directly observe the dynamics of complex regulatory programs that regulate neurogenesis through developmental pseudotime. Overall, scip-ATAC-seq increases information content per cell and bolsters the potential for future single-cell studies into complex developmental processes.

scholarly journals A fast and reliable method for monitoring genomic instability in the model organism Caenorhabditis elegans

2021 ◽  
Author(s):  
Merle Marie Nicolai ◽  
Barbara Witt ◽  
Andrea Hartwig ◽  
Tanja Schwerdtle ◽  
Julia Bornhorst
Keyword(s):  
Caenorhabditis Elegans ◽  
Animal Experiments ◽  
Model Organism ◽  
Animal Testing ◽  
C Elegans ◽  
Testing Strategies ◽  
Genotoxic Agents ◽  
Genotoxicity Testing

AbstractThe identification of genotoxic agents and their potential for genotoxic alterations in an organism is crucial for risk assessment and approval procedures of the chemical and pharmaceutical industry. Classically, testing strategies for DNA or chromosomal damage focus on in vitro and in vivo (mainly rodent) investigations. In cell culture systems, the alkaline unwinding (AU) assay is one of the well-established methods for detecting the percentage of double-stranded DNA (dsDNA). By establishing a reliable lysis protocol, and further optimization of the AU assay for the model organism Caenorhabditis elegans (C. elegans), we provided a new tool for genotoxicity testing in the niche between in vitro and rodent experiments. The method is intended to complement existing testing strategies by a multicellular organism, which allows higher predictability of genotoxic potential compared to in vitro cell line or bacterial investigations, before utilizing in vivo (rodent) investigations. This also allows working within the 3R concept (reduction, refinement, and replacement of animal experiments), by reducing and possibly replacing animal testing. Validation with known genotoxic agents (bleomycin (BLM) and tert-butyl hydroperoxide (tBOOH)) proved the method to be meaningful, reproducible, and feasible for high-throughput genotoxicity testing, and especially preliminary screening.

scholarly journals Detecting changes in the Caenorhabditis elegans intestinal environment using an engineered bacterial biosensor

2019 ◽  
Author(s):  
Jack W. Rutter ◽  
Tanel Ozdemir ◽  
Leonor M. Quintaneiro ◽  
Geraint Thomas ◽  
Filipe Cabreiro ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Rational Design ◽  
Model Organism ◽  
Bacterial Biosensor ◽  
Intestinal Environment ◽  
C Elegans ◽  
Ideal Candidate ◽  
Diagnostic Applications

AbstractCaenorhabditis elegans has become a key model organism within biology. In particular, the transparent gut, rapid growing time and ability to create a defined gut microbiota make it an ideal candidate organism for understanding and engineering the host microbiota. Here we present the development of an experimental model which can be used to characterise whole-cell bacterial biosensors in vivo. A dual-plasmid sensor system responding to isopropyl β-D-1-thiogalactopyranoside was developed and fully characterised in vitro. Subsequently, we show the sensor was capable of detecting and reporting on changes in the intestinal environment of C. elegans after introducing exogenous inducer into the environment. The protocols presented here may be used for aiding the rational design of engineered bacterial circuits, primarily for diagnostic applications. In addition, the model system may serve to reduce the use of current animal models and aid in the exploration of complex questions within general nematode and host-microbe biology.

scholarly journals Profiling of chromatin accessibility identifies transcription factor binding sites across the genome of Aspergillus species

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Lianggang Huang ◽  
Xuejie Li ◽  
Liangbo Dong ◽  
Bin Wang ◽  
Li Pan
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Filamentous Fungi ◽  
Binding Sites ◽  
Transcription Factor Binding ◽  
Chromatin Accessibility ◽  
Culture Conditions ◽  
Factor Binding ◽  
Tn5 Transposase

Abstract Background The identification of open chromatin regions and transcription factor binding sites (TFBs) is an important step in understanding the regulation of gene expression in diverse species. ATAC-seq is a technique used for such purpose by providing high-resolution measurements of chromatin accessibility revealed through integration of Tn5 transposase. However, the existence of cell walls in filamentous fungi and associated difficulty in purifying nuclei have precluded the routine application of this technique, leading to a lack of experimentally determined and computationally inferred data on the identity of genome-wide cis-regulatory elements (CREs) and TFBs. In this study, we constructed an ATAC-seq platform suitable for filamentous fungi and generated ATAC-seq libraries of Aspergillus niger and Aspergillus oryzae grown under a variety of conditions. Results We applied the ATAC-seq assay for filamentous fungi to delineate the syntenic orthologue and differentially changed chromatin accessibility regions among different Aspergillus species, during different culture conditions, and among specific TF-deleted strains. The syntenic orthologues of accessible regions were responsible for the conservative functions across Aspergillus species, while regions differentially changed between culture conditions and TFs mutants drove differential gene expression programs. Importantly, we suggest criteria to determine TFBs through the analysis of unbalanced cleavage of distinct TF-bound DNA strands by Tn5 transposase. Based on this criterion, we constructed data libraries of the in vivo genomic footprint of A. niger under distinct conditions, and generated a database of novel transcription factor binding motifs through comparison of footprints in TF-deleted strains. Furthermore, we validated the novel TFBs in vivo through an artificial synthetic minimal promoter system. Conclusions We characterized the chromatin accessibility regions of filamentous fungi species, and identified a complete TFBs map by ATAC-seq, which provides valuable data for future analyses of transcriptional regulation in filamentous fungi.

Fly-on-a-Chip: Microfluidics for Drosophila melanogaster Studies

2019 ◽  
Vol 11 (12) ◽  
pp. 425-443 ◽  
Author(s):  
Alireza Zabihihesari ◽  
Arthur J Hilliker ◽  
Pouya Rezai
Keyword(s):  
Drosophila Melanogaster ◽  
Developmental Stages ◽  
Model Organism ◽  
Fruit Fly ◽  
In Vitro Assays ◽  
Behavioral Studies ◽  
And Behavior ◽  
Manual Manipulation

Abstract The fruit fly or Drosophila melanogaster has been used as a promising model organism in genetics, developmental and behavioral studies as well as in the fields of neuroscience, pharmacology, and toxicology. Not only all the developmental stages of Drosophila, including embryonic, larval, and adulthood stages, have been used in experimental in vivo biology, but also the organs, tissues, and cells extracted from this model have found applications in in vitro assays. However, the manual manipulation, cellular investigation and behavioral phenotyping techniques utilized in conventional Drosophila-based in vivo and in vitro assays are mostly time-consuming, labor-intensive, and low in throughput. Moreover, stimulation of the organism with external biological, chemical, or physical signals requires precision in signal delivery, while quantification of neural and behavioral phenotypes necessitates optical and physical accessibility to Drosophila. Recently, microfluidic and lab-on-a-chip devices have emerged as powerful tools to overcome these challenges. This review paper demonstrates the role of microfluidic technology in Drosophila studies with a focus on both in vivo and in vitro investigations. The reviewed microfluidic devices are categorized based on their applications to various stages of Drosophila development. We have emphasized technologies that were utilized for tissue- and behavior-based investigations. Furthermore, the challenges and future directions in Drosophila-on-a-chip research, and its integration with other advanced technologies, will be discussed.

scholarly journals Reevaluation of the role of LIP-1 as an ERK/MPK-1 dual specificity phosphatase in the C. elegans germline

2022 ◽  
Vol 119 (3) ◽  
pp. e2113649119
Author(s):  
Debabrata Das ◽  
Jacob Seemann ◽  
David Greenstein ◽  
Tim Schedl ◽  
Swathi Arur
Keyword(s):  
Failure To Rescue ◽  
Gain Of Function ◽  
Erk Activation ◽  
C Elegans ◽  
Cellular Processes ◽  
Dual Specificity ◽  
Extracellular Signal ◽  
Dual Specificity Phosphatases ◽  
Organismal Development

The fidelity of a signaling pathway depends on its tight regulation in space and time. Extracellular signal-regulated kinase (ERK) controls wide-ranging cellular processes to promote organismal development and tissue homeostasis. ERK activation depends on a reversible dual phosphorylation on the TEY motif in its active site by ERK kinase (MEK) and dephosphorylation by DUSPs (dual specificity phosphatases). LIP-1, a DUSP6/7 homolog, was proposed to function as an ERK (MPK-1) DUSP in the Caenorhabditis elegans germline primarily because of its phenotype, which morphologically mimics that of a RAS/let-60 gain-of-function mutant (i.e., small oocyte phenotype). Our investigations, however, reveal that loss of lip-1 does not lead to an increase in MPK-1 activity in vivo. Instead, we show that loss of lip-1 leads to 1) a decrease in MPK-1 phosphorylation, 2) lower MPK-1 substrate phosphorylation, 3) phenocopy of mpk-1 reduction-of-function (rather than gain-of-function) allele, and 4) a failure to rescue mpk-1–dependent germline or fertility defects. Moreover, using diverse genetic mutants, we show that the small oocyte phenotype does not correlate with increased ectopic MPK-1 activity and that ectopic increase in MPK-1 phosphorylation does not necessarily result in a small oocyte phenotype. Together, these data demonstrate that LIP-1 does not function as an MPK-1 DUSP in the C. elegans germline. Our results caution against overinterpretation of the mechanistic underpinnings of orthologous phenotypes, since they may be a result of independent mechanisms, and provide a framework for characterizing the distinct molecular targets through which LIP-1 may mediate its several germline functions.

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