scholarly journals An Efficient Chromatin Immunoprecipitation (ChIP) Protocol for Studying Histone Modifications in Peach Reproductive Tissues

2021 ◽  
Author(s):  
Monica Canton ◽  
Silvia Farinati ◽  
Cristian Forestan ◽  
Justin Joseph ◽  
Claudio Bonghi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Histone Modifications ◽  
Chromatin Immunoprecipitation ◽  
Histone H3 ◽  
Fruit Trees ◽  
Fruit Growth ◽  
Bud Dormancy ◽  
Chromatin States ◽  
Reproductive Tissues ◽  
The Impact

Abstract BackgroundPerennial fruit trees display a perennial growth behaviour characterized by an annual cycling between growth and dormancy, with complex physiological features. Rosaceae fruit trees represent excellent models for studying not only the fruit growth/patterning, but also the progression of the reproductive cycle depending upon the impact of climate conditions. In addition, the current development of high‐throughput technologies is starting to have an important impact on Rosaceae tree research for investigating genome structure and function as well as (epi)genetic mechanisms involved in important developmental and environmental response processes during fruit tree growth. Among the epigenetic mechanisms, chromatin remodelling mediated by both histone modifications and other chromatin-related processes play a crucial role in gene modulation, controlling gene expression process. A very useful technique to investigate the chromatin states in plants and their dynamics is chromatin immunoprecipitation (ChIP), generally applied for studies on chromatin states and enrichment in post transcriptional modifications (PTMs) of histone proteins. Results Because peach is a model in Rosaceae family for studies in bud formation, dormancy and organ differentiation for climacteric fruits, in our work, we primarily established specific protocols for chromatin extraction and immunoprecipitation in reproductive tissues of peach Prunus persica. Subsequently focused our investigations on the role of two chromatin marks, namely trimethylation of histone H3 at lysine in position 4 (H3K4me3) and trimethylation of histone H3 at lysine 27 (H3K27me3) on modulating specific gene expression. Bud dormancy and fruit growth were investigated in a nectarine genotype called Fantasia as a model system. ConclusionsHere we presented general strategies to systematically optimize ChIP protocols for buds and mesocarp tissues and analyzed the correlation between gene expression and chromatin mark enrichment/depletion. Confirming like histone modifications are implicated in regulating bud dormancy progression and the core ripening genes.

scholarly journals Chromatin Profiling of the Repetitive and Nonrepetitive Genomes of the Human Fungal Pathogen Candida albicans

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Robert Jordan Price ◽  
Esther Weindling ◽  
Judith Berman ◽  
Alessia Buscaino
Keyword(s):  
Gene Expression ◽  
Candida Albicans ◽  
Histone Modifications ◽  
Chromatin Structure ◽  
Fungal Pathogen ◽  
Histone H3 ◽  
Content Type ◽  
Chromatin States ◽  
Genome Wide ◽  
Chromatin Profiling

ABSTRACT Eukaryotic genomes are packaged into chromatin structures that play pivotal roles in regulating all DNA-associated processes. Histone posttranslational modifications modulate chromatin structure and function, leading to rapid regulation of gene expression and genome stability, key steps in environmental adaptation. Candida albicans, a prevalent fungal pathogen in humans, can rapidly adapt and thrive in diverse host niches. The contribution of chromatin to C. albicans biology is largely unexplored. Here, we generated the first comprehensive chromatin profile of histone modifications (histone H3 trimethylated on lysine 4 [H3K4me3], histone H3 acetylated on lysine 9 [H3K9Ac], acetylated lysine 16 on histone H4 [H4K16Ac], and γH2A) across the C. albicans genome and investigated its relationship to gene expression by harnessing genome-wide sequencing approaches. We demonstrated that gene-rich nonrepetitive regions are packaged into canonical euchromatin in association with histone modifications that mirror their transcriptional activity. In contrast, repetitive regions are assembled into distinct chromatin states; subtelomeric regions and the ribosomal DNA (rDNA) locus are assembled into heterochromatin, while major repeat sequences and transposons are packaged in chromatin that bears features of euchromatin and heterochromatin. Genome-wide mapping of γH2A, a marker of genome instability, identified potential recombination-prone genomic loci. Finally, we present the first quantitative chromatin profiling in C. albicans to delineate the role of the chromatin modifiers Sir2 and Set1 in controlling chromatin structure and gene expression. This report presents the first genome-wide chromatin profiling of histone modifications associated with the C. albicans genome. These epigenomic maps provide an invaluable resource to understand the contribution of chromatin to C. albicans biology and identify aspects of C. albicans chromatin organization that differ from that of other yeasts. IMPORTANCE The fungus Candida albicans is an opportunistic pathogen that normally lives on the human body without causing any harm. However, C. albicans is also a dangerous pathogen responsible for millions of infections annually. C. albicans is such a successful pathogen because it can adapt to and thrive in different environments. Chemical modifications of chromatin, the structure that packages DNA into cells, can allow environmental adaptation by regulating gene expression and genome organization. Surprisingly, the contribution of chromatin modification to C. albicans biology is still largely unknown. For the first time, we analyzed C. albicans chromatin modifications on a genome-wide basis. We demonstrate that specific chromatin states are associated with distinct regions of the C. albicans genome and identify the roles of the chromatin modifiers Sir2 and Set1 in shaping C. albicans chromatin and gene expression.

scholarly journals In planta chromatin immunoprecipitation in Zymoseptoria tritici reveals chromatin-based regulation of putative effector gene expression

2019 ◽  
Author(s):  
Jessica L. Soyer ◽  
Jonathan Grandaubert ◽  
Janine Haueisen ◽  
Klaas Schotanus ◽  
Eva H. Stukenbrock
Keyword(s):  
Gene Expression ◽  
Histone Modifications ◽  
Chromatin Immunoprecipitation ◽  
High Throughput Sequencing ◽  
Histone H3 ◽  
In Planta ◽  
Zymoseptoria Tritici ◽  
Effector Gene ◽  
Effector Genes

SummaryDuring infection, pathogens secrete effectors, key elements of pathogenesis. In several phytopathogenic fungi, synchronous waves of effector genes are expressed during plant infection to manipulate and silence plant defenses. In Zymoseptoria tritici, causing septoria leaf blotch of wheat, at least two waves of effector genes are expressed, during the asymptomatic phase and at the switch to necrotrophy. The underlying factors responsible for the fine-tuned regulation of effector gene expression in this pathogen are unknown. Previously, a detailed map of the chromatin structure in vitro of Z. tritici was generated by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) targeting histone modifications typical for euchromatin (di-methylation of the lysine 4 of the histone H3, H3K4me2) or heterochromatin (tri-methylation of the lysine 9 and 27 of the histone H3, H3K9me3 and H3K27me3). Based on the hypothesis that changes in the histone modifications contribute to the transcriptional control of pathogenicity-related genes, we tested whether different sets of genes are associated with different histone modifications in vitro. We correlated the in vitro histone maps with in planta transcriptome data and show that genes located in heterochromatic domains in vitro are highly up-regulated at the switch toward necrotrophy. We combined our integrated analyses of genomic, transcriptomic and epigenomic data with ChIP-qPCR in planta and thereby provide further evidence for the involvement of histone modifications in the transcriptional dynamic of putative pathogenicity-related genes of Z. tritici.

scholarly journals Binding of an X-Specific Condensin Correlates with a Reduction in Active Histone Modifications at Gene Regulatory Elements

Genetics ◽  
2019 ◽  
Vol 212 (3) ◽  
pp. 729-742 ◽  
Author(s):  
Lena Annika Street ◽  
Ana Karina Morao ◽  
Lara Heermans Winterkorn ◽  
Chen-Yu Jiao ◽  
Sarah Elizabeth Albritton ◽  
...  
Keyword(s):  
Gene Expression ◽  
Histone Modifications ◽  
Chromatin Immunoprecipitation ◽  
Protein Complexes ◽  
Regulatory Elements ◽  
Evolutionarily Conserved ◽  
Chromatin Immunoprecipitation Sequencing ◽  
Gene Regulatory Elements ◽  
Gene Regulatory ◽  
Regulatory Sites

Condensins are evolutionarily conserved protein complexes that are required for chromosome segregation during cell division and genome organization during interphase. In Caenorhabditis elegans, a specialized condensin, which forms the core of the dosage compensation complex (DCC), binds to and represses X chromosome transcription. Here, we analyzed DCC localization and the effect of DCC depletion on histone modifications, transcription factor binding, and gene expression using chromatin immunoprecipitation sequencing and mRNA sequencing. Across the X, the DCC accumulates at accessible gene regulatory sites in active chromatin and not heterochromatin. The DCC is required for reducing the levels of activating histone modifications, including H3K4me3 and H3K27ac, but not repressive modification H3K9me3. In X-to-autosome fusion chromosomes, DCC spreading into the autosomal sequences locally reduces gene expression, thus establishing a direct link between DCC binding and repression. Together, our results indicate that DCC-mediated transcription repression is associated with a reduction in the activity of X chromosomal gene regulatory elements.

scholarly journals Tying Metabolic Branches With Histone Tails Using Systems Biology

2019 ◽  
Vol 12 ◽  
pp. 251686571986968
Author(s):  
Sriram Chandrasekaran
Keyword(s):  
Gene Expression ◽  
Systems Biology ◽  
Histone Modifications ◽  
Metabolic Pathways ◽  
Cellular Mechanism ◽  
Histone Tails ◽  
Metabolic Fluxes ◽  
Adenosyl Methionine ◽  
The Impact ◽  
Influence Gene Expression

Histone modifications represent an innate cellular mechanism to link nutritional status to gene expression. Metabolites such as acetyl-CoA and S-adenosyl methionine influence gene expression by serving as substrates for modification of histones. Yet, we lack a predictive model for determining histone modification levels based on cellular metabolic state. The numerous metabolic pathways that intersect with histone marks makes it highly challenging to understand their interdependencies. Here, we highlight new systems biology tools to unravel the impact of nutritional cues and metabolic fluxes on histone modifications.

scholarly journals Characterization of histone modifications associated with DNA damage repair genes upon exposure to gamma rays in Arabidopsis seedlings

2016 ◽  
Vol 57 (6) ◽  
pp. 646-654 ◽  
Author(s):  
Suvendu Mondal ◽  
Young Sam Go ◽  
Seung Sik Lee ◽  
Byung Yeoup Chung ◽  
Jin-Hong Kim
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Gamma Irradiation ◽  
Histone Modifications ◽  
Gamma Rays ◽  
Regulation Of Gene Expression ◽  
Chromatin Modification ◽  
Histone H3 ◽  
Marker Genes ◽  
Transcription Start Sites

Abstract Dynamic histone modifications play an important role in controlling gene expression in response to various environmental cues. This mechanism of regulation of gene expression is important for sessile organisms, like land plants. We have previously reported consistent upregulation of various marker genes in response to gamma rays at various post-irradiation times. In the present study, we performed various chromatin modification analyses at selected loci using the standard chromatin immunoprecipitation procedure, and demonstrate that upregulation of these genes is associated with histone H3 lysine 4 tri-methylation (H3K4me3) at the gene body or transcription start sites of these loci. Further, at specific AtAgo2 loci, both H3K4me3 and histone H3 lysine 9 acetylation (H3K9ac) are important in controlling gene expression in response to gamma irradiation. There was no change in DNA methylation in these selected loci. We conclude that specific histone modification such as H3K4me3 and H3K9ac may be more important in activating gene expression in these selected loci in response to gamma irradiation than a change in DNA methylation.

scholarly journals Deciphering Histone Modifications in Rice by Chromatin Immunoprecipitation (ChIP): Applications to Study the Impact of Stress Imposition

10.5772/66424 ◽  
2017 ◽  
Author(s):  
Liliana J. Ferreira ◽  
Sebastião Ravasco ◽  
Duarte D. Figueiredo ◽  
Christoph Peterhänsel ◽  
Nelson J.M. Saibo ◽  
...  
Keyword(s):  
Histone Modifications ◽  
Chromatin Immunoprecipitation ◽  
The Impact

scholarly journals Epigenetic inheritance uncoupled from sequence-specific recruitment

Science ◽  
2014 ◽  
Vol 348 (6230) ◽  
pp. 1258699 ◽  
Author(s):  
Kaushik Ragunathan ◽  
Gloria Jih ◽  
Danesh Moazed
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Histone Modifications ◽  
Posttranslational Modifications ◽  
Histone H3 ◽  
Epigenetic Inheritance ◽  
Meiotic Cell ◽  
Epigenetic Information ◽  
Cell Divisions ◽  
Jmjc Domain

Changes in histone posttranslational modifications are associated with epigenetic states that define distinct patterns of gene expression. It remains unclear whether epigenetic information can be transmitted through histone modifications independently of specific DNA sequence, DNA methylation, or RNA interference. Here we show that, in the fission yeast Schizosaccharomyces pombe, ectopically induced domains of histone H3 lysine 9 methylation (H3K9me), a conserved marker of heterochromatin, are inherited through several mitotic and meiotic cell divisions after removal of the sequence-specific initiator. The putative JmjC domain H3K9 demethylase, Epe1, and the chromodomain of the H3K9 methyltransferase, Clr4/Suv39h, play opposing roles in maintaining silent H3K9me domains. These results demonstrate how a direct “read-write” mechanism involving Clr4 propagates histone modifications and allows histones to act as carriers of epigenetic information.

scholarly journals Drosophila Paf1 Modulates Chromatin Structure at Actively Transcribed Genes

2006 ◽  
Vol 26 (1) ◽  
pp. 250-260 ◽  
Author(s):  
Karen Adelman ◽  
Wenxiang Wei ◽  
M. Behfar Ardehali ◽  
Janis Werner ◽  
Bing Zhu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase Ii ◽  
Chromatin Structure ◽  
Chromatin Immunoprecipitation ◽  
Promoter Region ◽  
Associated Factors ◽  
Histone H3 ◽  
Pol Ii ◽  
Stable Part ◽  
Paf1 Complex

ABSTRACT The Paf1 complex in yeast has been reported to influence a multitude of steps in gene expression through interactions with RNA polymerase II (Pol II) and chromatin-modifying complexes; however, it is unclear which of these many activities are primary functions of Paf1 and are conserved in metazoans. We have identified and characterized the Drosophila homologs of three subunits of the yeast Paf1 complex and found striking differences between the yeast and Drosophila Paf1 complexes. We demonstrate that although Drosophila Paf1, Rtf1, and Cdc73 colocalize broadly with actively transcribing, phosphorylated Pol II, and all are recruited to activated heat shock genes with similar kinetics; Rtf1 does not appear to be a stable part of the Drosophila Paf1 complex. RNA interference (RNAi)-mediated depletion of Paf1 or Rtf1 leads to defects in induction of Hsp70 RNA, but tandem RNAi-chromatin immunoprecipitation assays show that loss of neither Paf1 nor Rtf1 alters the density or distribution of phosphorylated Pol II on the active Hsp70 gene. However, depletion of Paf1 reduces trimethylation of histone H3 at lysine 4 in the Hsp70 promoter region and significantly decreases the recruitment of chromatin-associated factors Spt6 and FACT, suggesting that Paf1 may manifest its effects on transcription through modulating chromatin structure.

scholarly journals Changes in Histone Modification and DNA Methylation of the StAR and Cyp19a1 Promoter Regions in Granulosa Cells Undergoing Luteinization during Ovulation In Rats

Endocrinology ◽  
2013 ◽  
Vol 154 (1) ◽  
pp. 458-470 ◽  
Author(s):  
Lifa Lee ◽  
Hiromi Asada ◽  
Fumie Kizuka ◽  
Isao Tamura ◽  
Ryo Maekawa ◽  
...  
Keyword(s):  
Gene Expression ◽  
Histone Modifications ◽  
Granulosa Cells ◽  
Chromatin Immunoprecipitation ◽  
Binding Protein ◽  
Mrna Levels ◽  
Chromatin Immunoprecipitation Assay ◽  
Immunoprecipitation Assay ◽  
Rapid Changes

The ovulatory LH surge induces rapid up-regulation of steroidogenic acute regulatory (StAR) protein and rapid down-regulation of aromatase (Cyp19a1) in granulosa cells (GCs) undergoing luteinization during ovulation. This study investigated in vivo whether epigenetic mechanisms including histone modifications are involved in the rapid changes of StAR and Cyp19a1 gene expression. GCs were obtained from rats treated with equine chorionic gonadotropin (CG) before (0 h) and after human (h)CG injection. StAR mRNA levels rapidly increased after hCG injection, reached a peak at 4 h, and then remained higher compared with 0 h until 12 h. Cyp19a1 mRNA levels gradually decreased after hCG injection and reached their lowest level at 12 h. A chromatin immunoprecipitation assay revealed that levels of histone-H4 acetylation (Ac-H4) and trimethylation of histone-H3 lysine-4 (H3K4me3) increased whereas H3K9me3 and H3K27me3 decreased in the StAR promoter after hCG injection. On the other hand, the levels of Ac-H3 and -H4 and H3K4me3 decreased, and H3K27me3 increased in the Cyp19a1 promoter after hCG injection. Chromatin condensation, which was analyzed using deoxyribonuclease I, decreased in the StAR promoter and increased in the Cyp19a1 promoter after hCG injection. A chromatin immunoprecipitation assay also showed that binding activities of CAATT/enhancer-binding protein β to the StAR promoter increased and binding activities of phosphorylated-cAMP response element binding protein to the Cyp19a1 promoter decreased after hCG injection. These results provide in vivo evidence that histone modifications are involved in the rapid changes of StAR and Cyp19a1 gene expression by altering chromatin structure of the promoters in GCs undergoing luteinization during ovulation.

Chromatin dynamics associated with sexual differentiation in a UV sex determination system

2020 ◽  
Author(s):  
Josselin Gueno ◽  
Simon Bourdareau ◽  
Guillaume Cossard ◽  
Olivier Godfroy ◽  
Agnieszka Lipinska ◽  
...  
Keyword(s):  
Gene Expression ◽  
Sex Determination ◽  
Histone Modifications ◽  
Sex Chromosomes ◽  
Sexual Differentiation ◽  
Chromatin State ◽  
State Transitions ◽  
Male And Female ◽  
Chromatin States ◽  
Males And Females

SummaryIn many eukaryotes, such as dioicous mosses and many algae, sex is determined by UV sex chromosomes and is expressed during the haploid phase of the life cycle. In these species, the male and female developmental programs are initiated by the presence of the U- or V-specific regions of the sex chromosomes but, as in XY and ZW systems, phenotypic differentiation is largely driven by autosomal sex-biased gene expression. The mechanisms underlying sex-biased transcription in XY, ZW or UV sexual systems currently remain elusive. Here, we set out to understand the extent and nature of epigenomic changes associated with sexual differentiation in the brown alga Ectocarpus, which has a well described UV system. Five histone modifications, H3K4me3, H3K27Ac, H3K9Ac, H3K36me3, H4K20me3, were quantified in near-isogenic male and female lines, leading to the identification of 13 different chromatin states across the Ectocarpus genome that showed different patterns of enrichment at transcribed, silent, housekeeping or narrowly-expressed genes. Chromatin states were strongly correlated with levels of gene expression indicating a relationship between the assayed marks and gene transcription. The relative proportion of each chromatin state across the genome remained stable in males and females, but a subset of genes exhibited different chromatin states in the two sexes. In particular, males and females displayed distinct patterns of histone modifications at sex-biased genes, indicating that chromatin state transitions occur preferentially at genes involved in sex-specific pathways. Finally, our results reveal a unique chromatin landscape of the U and V sex chromosomes compared to autosomes. Taken together, our observations reveal a role for histone modifications in sex determination and sexual differentiation in a UV sexual system, and suggest that the mechanisms of epigenetic regulation of genes on the UV sex chromosomes may differ from those operating on autosomal genes.

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