Publications by authors named "Victor G Corces"

109 Publications

Principles of 3D compartmentalization of the human genome.

Cell Rep 2021 Jun;35(13):109330

Department of Human Genetics, Emory University School of Medicine, 615 Michael St., Atlanta, GA 30322, USA. Electronic address:

Chromatin is organized in the nucleus via CTCF loops and compartmental domains. Here, we compare different cell types to identify distinct paradigms of compartmental domain formation in human tissues. We identify and quantify compartmental forces correlated with histone modifications characteristic of transcriptional activity and previously underappreciated roles for distinct compartmental domains correlated with the presence of H3K27me3 and H3K9me3, respectively. We present a computer simulation model capable of predicting compartmental organization based on the biochemical characteristics of independent chromatin features. Using this model, we show that the underlying forces responsible for compartmental domain formation in human cells are conserved and that the diverse compartmentalization patterns seen across cell types are due to differences in chromatin features. We extend these findings to Drosophila to suggest that the same principles are at work beyond humans. These results offer mechanistic insights into the fundamental forces driving the 3D organization of the genome.
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http://dx.doi.org/10.1016/j.celrep.2021.109330DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8265014PMC
June 2021

Exposure to sevoflurane results in changes of transcription factor occupancy in sperm and inheritance of autism.

Biol Reprod 2021 May 12. Epub 2021 May 12.

Department of Human Genetics, Emory University School of Medicine, 615 Michael St, Atlanta, GA 30322, USA.

One in 54 children in the U.S. is diagnosed with Autism Spectrum Disorder (ASD). De novo germline and somatic mutations cannot account for all cases of ASD, suggesting that epigenetic alterations triggered by environmental exposures may be responsible for a subset of ASD cases. Human and animal studies have shown that exposure of the developing brain to general anesthetic (GA) agents can trigger neurodegeneration and neurobehavioral abnormalities but the effects of general anesthetics on the germ line have not been explored in detail. We exposed pregnant mice to sevoflurane during the time of embryonic development when the germ cells undergo epigenetic reprogramming and found that more than 38% of the directly exposed F1 animals exhibit impairments in anxiety and social interactions. Strikingly, 44-47% of the F2 and F3 animals, which were not directly exposed to sevoflurane, show the same behavioral problems. We performed ATAC-seq and identified more than 1200 differentially accessible sites in the sperm of F1 animals, 69 of which are also present in the sperm of F2 animals. These sites are located in regulatory regions of genes strongly associated with ASD, including Arid1b, Ntrk2, and Stmn2. These findings suggest that epimutations caused by exposing germ cells to sevoflurane can lead to ASD in the offspring, and this effect can be transmitted through the male germline inter and trans-generationally.
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http://dx.doi.org/10.1093/biolre/ioab097DOI Listing
May 2021

Replication timing maintains the global epigenetic state in human cells.

Science 2021 04 22;372(6540):371-378. Epub 2021 Apr 22.

Department of Biological Science, Florida State University, Tallahassee, FL 32306, USA.

The temporal order of DNA replication [replication timing (RT)] is correlated with chromatin modifications and three-dimensional genome architecture; however, causal links have not been established, largely because of an inability to manipulate the global RT program. We show that loss of RIF1 causes near-complete elimination of the RT program by increasing heterogeneity between individual cells. RT changes are coupled with widespread alterations in chromatin modifications and genome compartmentalization. Conditional depletion of RIF1 causes replication-dependent disruption of histone modifications and alterations in genome architecture. These effects were magnified with successive cycles of altered RT. These results support models in which the timing of chromatin replication and thus assembly plays a key role in maintaining the global epigenetic state.
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http://dx.doi.org/10.1126/science.aba5545DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8173839PMC
April 2021

Regulation of 3D chromatin organization by CTCF.

Curr Opin Genet Dev 2021 04 28;67:33-40. Epub 2020 Nov 28.

Emory University School of Medicine, Department of Human Genetics, 615 Michael Street, Atlanta, GA 30322, USA. Electronic address:

Studies of nuclear architecture using chromosome conformation capture methods have provided a detailed view of how chromatin folds in the 3D nuclear space. New variants of this technology now afford unprecedented resolution and allow the identification of ever smaller folding domains that offer new insights into the mechanisms by which this organization is established and maintained. Here we review recent results in this rapidly evolving field with an emphasis on CTCF function, with the goal of gaining a mechanistic understanding of the principles by which chromatin is folded in the eukaryotic nucleus.
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http://dx.doi.org/10.1016/j.gde.2020.10.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8084898PMC
April 2021

Haspin kinase modulates nuclear architecture and Polycomb-dependent gene silencing.

PLoS Genet 2020 08 4;16(8):e1008962. Epub 2020 Aug 4.

Institut de Biologia Molecular de Barcelona, IBMB-CSIC, Barcelona, Spain.

Haspin, a highly conserved kinase in eukaryotes, has been shown to be responsible for phosphorylation of histone H3 at threonine 3 (H3T3ph) during mitosis, in mammals and yeast. Here we report that haspin is the kinase that phosphorylates H3T3 in Drosophila melanogaster and it is involved in sister chromatid cohesion during mitosis. Our data reveal that haspin also phosphorylates H3T3 in interphase. H3T3ph localizes in broad silenced domains at heterochromatin and lamin-enriched euchromatic regions. Loss of haspin compromises insulator activity in enhancer-blocking assays and triggers a decrease in nuclear size that is accompanied by changes in nuclear envelope morphology. We show that haspin is a suppressor of position-effect variegation involved in heterochromatin organization. Our results also demonstrate that haspin is necessary for pairing-sensitive silencing and it is required for robust Polycomb-dependent homeotic gene silencing. Haspin associates with the cohesin complex in interphase, mediates Pds5 binding to chromatin and cooperates with Pds5-cohesin to modify Polycomb-dependent homeotic transformations. Therefore, this study uncovers an unanticipated role for haspin kinase in genome organization of interphase cells and demonstrates that haspin is required for homeotic gene regulation.
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http://dx.doi.org/10.1371/journal.pgen.1008962DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7428214PMC
August 2020

Protection from DNA re-methylation by transcription factors in primordial germ cells and pre-implantation embryos can explain trans-generational epigenetic inheritance.

Genome Biol 2020 05 18;21(1):118. Epub 2020 May 18.

Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, 30322, USA.

Background: A growing body of evidence suggests that certain epiphenotypes can be passed across generations via both the male and female germlines of mammals. These observations have been difficult to explain owing to a global loss of the majority of known epigenetic marks present in parental chromosomes during primordial germ cell development and after fertilization.

Results: By integrating previously published BS-seq, DNase-seq, ATAC-seq, and RNA-seq data collected during multiple stages of primordial germ cell and pre-implantation development, we find that the methylation status of the majority of CpGs genome-wide is restored after global de-methylation, despite the fact that global CpG methylation drops to 10% in primordial germ cells and 20% in the inner cell mass of the blastocyst. We estimate the proportion of such CpGs with preserved methylation status to be 78%. Further, we find that CpGs at sites bound by transcription factors during the global re-methylation phases of germline and embryonic development remain hypomethylated across all developmental stages observed. On the other hand, CpGs at sites not bound by transcription factors during the global re-methylation phase have high methylation levels prior to global de-methylation, become de-methylated during global de-methylation, and then become re-methylated.

Conclusions: The results suggest that transcription factors can act as carriers of epigenetic information during germ cell and pre-implantation development by ensuring that the methylation status of CpGs is maintained. These findings provide the basis for a mechanistic description of trans-generational inheritance of epigenetic information in mammals.
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http://dx.doi.org/10.1186/s13059-020-02036-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236515PMC
May 2020

Analysis of Hi-C data using SIP effectively identifies loops in organisms from to mammals.

Genome Res 2020 03 3;30(3):447-458. Epub 2020 Mar 3.

Department of Human Genetics, Emory University School of Medicine, Atlanta, Georgia 30322, USA.

Chromatin loops are a major component of 3D nuclear organization, visually apparent as intense point-to-point interactions in Hi-C maps. Identification of these loops is a critical part of most Hi-C analyses. However, current methods often miss visually evident CTCF loops in Hi-C data sets from mammals, and they completely fail to identify high intensity loops in other organisms. We present SIP, Significant Interaction Peak caller, and SIPMeta, which are platform independent programs to identify and characterize these loops in a time- and memory-efficient manner. We show that SIP is resistant to noise and sequencing depth, and can be used to detect loops that were previously missed in human cells as well as loops in other organisms. SIPMeta corrects for a common visualization artifact by accounting for Manhattan distance to create average plots of Hi-C and HiChIP data. We then demonstrate that the use of SIP and SIPMeta can lead to biological insights by characterizing the contribution of several transcription factors to CTCF loop stability in human cells. We also annotate loops associated with the SMC component of the dosage compensation complex (DCC) in and demonstrate that loop anchors represent bidirectional blocks for symmetrical loop extrusion. This is in contrast to the asymmetrical extrusion until unidirectional blockage by CTCF that is presumed to occur in mammals. Using HiChIP and multiway ligation events, we then show that DCC loops form a network of strong interactions that may contribute to X Chromosome-wide condensation in hermaphrodites.
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http://dx.doi.org/10.1101/gr.257832.119DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7111518PMC
March 2020

Mini-review: Epigenetic mechanisms that promote transgenerational actions of endocrine disrupting chemicals: Applications to behavioral neuroendocrinology.

Horm Behav 2020 03 22;119:104677. Epub 2020 Jan 22.

Center for Human Health and the Environment, North Carolina State University, Raleigh, NC 27695, United States of America. Electronic address:

It is our hope this mini-review will stimulate discussion and new research. Here we briefly examine the literature on transgenerational actions of endocrine disrupting chemicals (EDCs) on brain and behavior and their underlying epigenetic mechanisms including: DNA methylation, histone modifications, and non-coding RNAs. We stress that epigenetic modifications need to be examined in a synergistic manner, as they act together in situ on chromatin to change transcription. Next we highlight recent work from one of our laboratories (VGC). The data provide new evidence that the sperm genome is poised for transcription. In developing sperm, gene enhancers and promoters are accessible for transcription and these activating motifs are also found in preimplantation embryos. Thus, DNA modifications associated with transcription factors during fertilization, in primordial germ cells (PGCs), and/or during germ cell maturation may be passed to offspring. We discuss the implications of this model to EDC exposures and speculate on whether natural variation in hormone levels during fertilization and PGC migration may impart transgenerational effects on brain and behavior. Lastly we discuss how this mechanism could apply to neural sexual differentiation.
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http://dx.doi.org/10.1016/j.yhbeh.2020.104677DOI Listing
March 2020

Widespread long-range cis-regulatory elements in the maize genome.

Nat Plants 2019 12 18;5(12):1237-1249. Epub 2019 Nov 18.

Department of Plant Biology, University of Georgia, Athens, GA, USA.

Genetic mapping studies on crops suggest that agronomic traits can be controlled by gene-distal intergenic loci. Despite the biological importance and the potential agronomic utility of these loci, they remain virtually uncharacterized in all crop species to date. Here, we provide genetic, epigenomic and functional molecular evidence to support the widespread existence of gene-distal (hereafter, distal) loci that act as long-range transcriptional cis-regulatory elements (CREs) in the maize genome. Such loci are enriched for euchromatic features that suggest their regulatory functions. Chromatin loops link together putative CREs with genes and recapitulate genetic interactions. Putative CREs also display elevated transcriptional enhancer activities, as measured by self-transcribing active regulatory region sequencing. These results provide functional support for the widespread existence of CREs that act over large genomic distances to control gene expression.
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http://dx.doi.org/10.1038/s41477-019-0547-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6904520PMC
December 2019

Chromatin accessibility and transcription dynamics during in vitro astrocyte differentiation of Huntington's Disease Monkey pluripotent stem cells.

Epigenetics Chromatin 2019 11 13;12(1):67. Epub 2019 Nov 13.

Division of Neuropharmacology and Neurologic Diseases, Yerkes National Primate Research Center, Atlanta, GA, 30322, USA.

Background: Huntington's Disease (HD) is a fatal neurodegenerative disorder caused by a CAG repeat expansion, resulting in a mutant huntingtin protein. While it is now clear that astrocytes are affected by HD and significantly contribute to neuronal dysfunction and pathogenesis, the alterations in the transcriptional and epigenetic profiles in HD astrocytes have yet to be characterized. Here, we examine global transcription and chromatin accessibility dynamics during in vitro astrocyte differentiation in a transgenic non-human primate model of HD.

Results: We found global changes in accessibility and transcription across different stages of HD pluripotent stem cell differentiation, with distinct trends first observed in neural progenitor cells (NPCs), once cells have committed to a neural lineage. Transcription of p53 signaling and cell cycle pathway genes was highly impacted during differentiation, with depletion in HD NPCs and upregulation in HD astrocytes. E2F target genes also displayed this inverse expression pattern, and strong associations between E2F target gene expression and accessibility at nearby putative enhancers were observed.

Conclusions: The results suggest that chromatin accessibility and transcription are altered throughout in vitro HD astrocyte differentiation and provide evidence that E2F dysregulation contributes to aberrant cell-cycle re-entry and apoptosis throughout the progression from NPCs to astrocytes.
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http://dx.doi.org/10.1186/s13072-019-0313-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6852955PMC
November 2019

Immediate and deferred epigenomic signatures of in vivo neuronal activation in mouse hippocampus.

Nat Neurosci 2019 10 9;22(10):1718-1730. Epub 2019 Sep 9.

Instituto de Neurociencias, Universidad Miguel Hernández-Consejo Superior de Investigaciones Científicas, Sant Joan d'Alacant, Alicante, Spain.

Activity-driven transcription plays an important role in many brain processes, including those underlying memory and epilepsy. Here we combine genetic tagging of nuclei and ribosomes with RNA sequencing, chromatin immunoprecipitation with sequencing, assay for transposase-accessible chromatin using sequencing and Hi-C to investigate transcriptional and chromatin changes occurring in mouse hippocampal excitatory neurons at different time points after synchronous activation during seizure and sparse activation by novel context exploration. The transcriptional burst is associated with an increase in chromatin accessibility of activity-regulated genes and enhancers, de novo binding of activity-regulated transcription factors, augmented promoter-enhancer interactions and the formation of gene loops that bring together the transcription start site and transcription termination site of induced genes and may sustain the fast reloading of RNA polymerase complexes. Some chromatin occupancy changes and interactions, particularly those driven by AP1, remain long after neuronal activation and could underlie the changes in neuronal responsiveness and circuit connectivity observed in these neuroplasticity paradigms, perhaps thereby contributing to metaplasticity in the adult brain.
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http://dx.doi.org/10.1038/s41593-019-0476-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6875776PMC
October 2019

Ecdysone-Induced 3D Chromatin Reorganization Involves Active Enhancers Bound by Pipsqueak and Polycomb.

Cell Rep 2019 09;28(10):2715-2727.e5

Instituto de Neurociencias, Consejo Superior de Investigaciones Científicas-Universidad Miguel Hernández (CSIC-UMH), 03550 Sant Joan, Alicante, Spain. Electronic address:

Evidence suggests that Polycomb (Pc) is present at chromatin loop anchors in Drosophila. Pc is recruited to DNA through interactions with the GAGA binding factors GAF and Pipsqueak (Psq). Using HiChIP in Drosophila cells, we find that the psq gene, which has diverse roles in development and tumorigenesis, encodes distinct isoforms with unanticipated roles in genome 3D architecture. The BR-C, ttk, and bab domain (BTB)-containing Psq isoform (Psq) colocalizes genome-wide with known architectural proteins. Conversely, Psq lacking the BTB domain (Psq) is consistently found at Pc loop anchors and at active enhancers, including those that respond to the hormone ecdysone. After stimulation by this hormone, chromatin 3D organization is altered to connect promoters and ecdysone-responsive enhancers bound by Psq. Our findings link Psq variants lacking the BTB domain to Pc-bound active enhancers, thus shedding light into their molecular function in chromatin changes underlying the response to hormone stimulus.
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http://dx.doi.org/10.1016/j.celrep.2019.07.096DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6754745PMC
September 2019

Seeing Is Believing: ORCA Allows Visualization of Three-Dimensional Genome Organization at Single-Cell Resolution.

Biochemistry 2019 08 12;58(33):3477-3479. Epub 2019 Aug 12.

Department of Biology , Emory University , 1510 Clifton Road Northeast , Atlanta , Georgia 30322 , United States.

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http://dx.doi.org/10.1021/acs.biochem.9b00611DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6702087PMC
August 2019

Rapid Irreversible Transcriptional Reprogramming in Human Stem Cells Accompanied by Discordance between Replication Timing and Chromatin Compartment.

Stem Cell Reports 2019 07 20;13(1):193-206. Epub 2019 Jun 20.

Department of Biological Science, Florida State University, 319 Stadium Drive, Tallahassee, FL 32306, USA. Electronic address:

The temporal order of DNA replication is regulated during development and is highly correlated with gene expression, histone modifications and 3D genome architecture. We tracked changes in replication timing, gene expression, and chromatin conformation capture (Hi-C) A/B compartments over the first two cell cycles during differentiation of human embryonic stem cells to definitive endoderm. Remarkably, transcriptional programs were irreversibly reprogrammed within the first cell cycle and were largely but not universally coordinated with replication timing changes. Moreover, changes in A/B compartment and several histone modifications that normally correlate strongly with replication timing showed weak correlation during the early cell cycles of differentiation but showed increased alignment in later differentiation stages and in terminally differentiated cell lines. Thus, epigenetic cell fate transitions during early differentiation can occur despite dynamic and discordant changes in otherwise highly correlated genomic properties.
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http://dx.doi.org/10.1016/j.stemcr.2019.05.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6627004PMC
July 2019

Maintenance of CTCF- and Transcription Factor-Mediated Interactions from the Gametes to the Early Mouse Embryo.

Mol Cell 2019 07 2;75(1):154-171.e5. Epub 2019 May 2.

Department of Biology, Emory University, 1510 Clifton Rd. NE, Atlanta, GA 30322, USA; Genetics and Molecular Biology Program, Graduate Division of Biological and Biomedical Sciences, 1462 Clifton Rd., Atlanta, GA 30322, USA. Electronic address:

The epigenetic information present in mammalian gametes and whether it is transmitted to the progeny are relatively unknown. We find that many promoters in mouse sperm are occupied by RNA polymerase II (Pol II) and Mediator. The same promoters are accessible in GV and MII oocytes and preimplantation embryos. Sperm distal ATAC-seq sites containing motifs for various transcription factors are conserved in monkeys and humans. ChIP-seq analyses confirm that Foxa1, ERα, and AR occupy distal enhancers in sperm. Accessible sperm enhancers containing H3.3 and H2A.Z are also accessible in oocytes and preimplantation embryos. Furthermore, their interactions with promoters in the gametes persist during early development. Sperm- or oocyte-specific interactions mediated by CTCF and cohesin are only present in the paternal or maternal chromosomes, respectively, in the zygote and 2-cell stages. These interactions converge in both chromosomes by the 8-cell stage. Thus, mammalian gametes contain complex patterns of 3D interactions that can be transmitted to the zygote after fertilization.
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http://dx.doi.org/10.1016/j.molcel.2019.04.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6625867PMC
July 2019

Condensin II Counteracts Cohesin and RNA Polymerase II in the Establishment of 3D Chromatin Organization.

Cell Rep 2019 03;26(11):2890-2903.e3

Department of Biology, Emory University, 1510 Clifton Road NE, Atlanta, GA 30322, USA. Electronic address:

Interaction domains in Drosophila chromosomes form by segregation of active and inactive chromatin in the absence of CTCF loops, but the role of transcription versus other architectural proteins in chromatin organization is unclear. Here, we find that positioning of RNAPII via transcription elongation is essential in the formation of gene loops, which in turn interact to form compartmental domains. Inhibition of transcription elongation or depletion of cohesin decreases gene looping and formation of active compartmental domains. In contrast, depletion of condensin II, which also localizes to active chromatin, causes increased gene looping, formation of compartmental domains, and stronger intra-chromosomal compartmental interactions. Condensin II has a similar role in maintaining inter-chromosomal interactions responsible for pairing between homologous chromosomes, whereas inhibition of transcription elongation or cohesin depletion has little effect on homolog pairing. The results suggest distinct roles for cohesin and condensin II in the establishment of 3D nuclear organization in Drosophila.
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http://dx.doi.org/10.1016/j.celrep.2019.01.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6424357PMC
March 2019

Pach-ing It in: The Peculiar Organization of Mammalian Pachytene Chromosomes.

Mol Cell 2019 02;73(3):392-394

Department of Biology, Emory University, 1510 Clifton Road NE, Atlanta, GA 30322, USA. Electronic address:

In this issue of Molecular Cell, Wang et al. (2019) use Hi-C to visualize at high resolution the complex reprogramming of chromatin architecture during spermatogenesis in rhesus monkeys and mice. They find that pachytene spermatocytes have a unique chromosome organization that may result from the presence of the synaptonemal complex and transcription-associated proteins.
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http://dx.doi.org/10.1016/j.molcel.2019.01.030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6701473PMC
February 2019

Chromatin changes in Anopheles gambiae induced by Plasmodium falciparum infection.

Epigenetics Chromatin 2019 01 7;12(1). Epub 2019 Jan 7.

Estación Biológica de Doñana (EBD), Consejo Superior de Investigaciones Científicas, 41092, Seville, Spain.

Background: Infection by the human malaria parasite leads to important changes in mosquito phenotypic traits related to vector competence. However, we still lack a clear understanding of the underlying mechanisms and, in particular, of the epigenetic basis for these changes. We have examined genome-wide distribution maps of H3K27ac, H3K9ac, H3K9me3 and H3K4me3 by ChIP-seq and the transcriptome by RNA-seq, of midguts from Anopheles gambiae mosquitoes blood-fed uninfected and infected with natural isolates of the human malaria parasite Plasmodium falciparum in Burkina Faso.

Results: We report 15,916 regions containing differential histone modification enrichment between infected and uninfected, of which 8339 locate at promoters and/or intersect with genes. The functional annotation of these regions allowed us to identify infection-responsive genes showing differential enrichment in various histone modifications, such as CLIP proteases, antimicrobial peptides-encoding genes, and genes related to melanization responses and the complement system. Further, the motif analysis of regions differentially enriched in various histone modifications predicts binding sites that might be involved in the cis-regulation of these regions, such as Deaf1, Pangolin and Dorsal transcription factors (TFs). Some of these TFs are known to regulate immunity gene expression in Drosophila and are involved in the Notch and JAK/STAT signaling pathways.

Conclusions: The analysis of malaria infection-induced chromatin changes in mosquitoes is important not only to identify regulatory elements and genes underlying mosquito responses to P. falciparum infection, but also for possible applications to the genetic manipulation of mosquitoes and to other mosquito-borne systems.
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http://dx.doi.org/10.1186/s13072-018-0250-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6322293PMC
January 2019

Resolution of the DNA methylation state of single CpG dyads using in silico strand annealing and WGBS data.

Nat Protoc 2019 01;14(1):202-216

Department of Biology, Emory University, Atlanta, GA, USA.

Whole-genome bisulfite sequencing (WGBS) has been widely used to quantify cytosine DNA methylation frequency in an expanding array of cell and tissue types. Because of the denaturing conditions used, this method ultimately leads to the measurement of methylation frequencies at single cytosines. Hence, the methylation frequency of CpG dyads (two complementary CG dinucleotides) can be only indirectly inferred by overlaying the methylation frequency of two cytosines measured independently. Furthermore, hemi-methylated CpGs (hemiCpGs) have not been previously analyzed in WGBS studies. We recently developed in silico strand annealing (iSA), a bioinformatics method applicable to WGBS data, to resolve the methylation status of CpG dyads into unmethylated, hemi-methylated, and methylated. HemiCpGs account for 4-20% of the DNA methylome in different cell types, and some can be inherited across cell divisions, suggesting a role as a stable epigenetic mark. Therefore, it is important to resolve hemiCpGs from fully methylated CpGs in WGBS studies. This protocol describes step-by-step commands to accomplish this task, including dividing alignments by strand, pairing alignments between strands, and extracting single-fragment methylation calls. The versatility of iSA enables its application downstream of other WGBS-related methods such as nasBS-seq (nascent DNA bisulfite sequencing), ChIP-BS-seq (ChIP followed by bisulfite sequencing), TAB-seq, oxBS-seq, and fCAB-seq. iSA is also tunable for analyzing the methylation status of cytosines in any sequence context. We exemplify this flexibility by uncovering the single-fragment non-CpG methylome. This protocol provides enough details for users with little experience in bioinformatic analysis and takes 2-7 h.
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http://dx.doi.org/10.1038/s41596-018-0090-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6311134PMC
January 2019

Engineering 3D genome organization.

Cell Res 2019 01;29(1):1-3

Department of Biology, Emory University, Atlanta, GA, 30322, USA.

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http://dx.doi.org/10.1038/s41422-018-0116-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6318305PMC
January 2019

Organizational principles of 3D genome architecture.

Nat Rev Genet 2018 12;19(12):789-800

Department of Biology, Emory University, Atlanta, GA, USA.

Studies of 3D chromatin organization have suggested that chromosomes are hierarchically organized into large compartments composed of smaller domains called topologically associating domains (TADs). Recent evidence suggests that compartments are smaller than previously thought and that the transcriptional or chromatin state is responsible for interactions leading to the formation of small compartmental domains in all organisms. In vertebrates, CTCF forms loop domains, probably via an extrusion process involving cohesin. CTCF loops cooperate with compartmental domains to establish the 3D organization of the genome. The continuous extrusion of the chromatin fibre by cohesin may also be responsible for the establishment of enhancer-promoter interactions and stochastic aspects of the transcription process. These observations suggest that the 3D organization of the genome is an emergent property of chromatin and its components, and thus may not be only a determinant but also a consequence of its function.
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http://dx.doi.org/10.1038/s41576-018-0060-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6312108PMC
December 2018

A tethered-inchworm model of SMC DNA translocation.

Nat Struct Mol Biol 2018 10 24;25(10):906-910. Epub 2018 Sep 24.

Department of Biology, Emory University, Atlanta, GA, USA.

The DNA loop extrusion model is a provocative new concept explaining the formation of chromatin loops that revolutionizes understanding of genome organization. Central to this model is the structural maintenance of chromosomes (SMC) protein family, which is now thought to function as a DNA motor. In this Perspective, we review and reinterpret the current knowledge of SMC structure and function and propose a novel mechanism for SMC motor activity.
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http://dx.doi.org/10.1038/s41594-018-0135-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6311135PMC
October 2018

Architectural Proteins and Pluripotency Factors Cooperate to Orchestrate the Transcriptional Response of hESCs to Temperature Stress.

Mol Cell 2018 09 16;71(6):940-955.e7. Epub 2018 Aug 16.

Department of Biology, Emory University, Atlanta, GA 30322, USA. Electronic address:

Cells respond to temperature stress via up- and downregulation of hundreds of genes. This process is thought to be regulated by the heat shock factor HSF1, which controls the release of RNAPII from promoter-proximal pausing. Here, we analyze the events taking place in hESCs upstream of RNAPII release. We find that temperature stress results in the activation or decommissioning of thousands of enhancers. This process involves alterations in the occupancy of transcription factors HSF1, AP-1, NANOG, KLF4, and OCT4 accompanied by nucleosome remodeling by BRG1 and changes in H3K27ac. Furthermore, redistribution of RAD21 and CTCF results in the formation and disassembly of interactions mediated by these two proteins. These alterations tether and untether enhancers to their cognate promoters or refashion insulated neighborhoods, thus transforming the landscape of enhancer-promoter interactions. Details of the 3D interactome remodeling process support loop extrusion initiating at random sites as a mechanism for the establishment of CTCF/cohesin loops.
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http://dx.doi.org/10.1016/j.molcel.2018.07.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6214669PMC
September 2018

Analysis of Chromatin Interactions Mediated by Specific Architectural Proteins in Drosophila Cells.

Methods Mol Biol 2018 ;1766:239-256

Department of Biology, Emory University, Atlanta, GA, USA.

Chromosome conformation capture assays have been established, modified, and enhanced for over a decade with the purpose of studying nuclear organization. A recently published method uses in situ Hi-C followed by chromatin immunoprecipitation (HiChIP) to enrich the overall yield of significant genome-wide interactions mediated by a specific protein. Here we applied a modified version of the HiChIP protocol to retrieve the significant contacts mediated by architectural protein CP190 in D. melanogaster cells.
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http://dx.doi.org/10.1007/978-1-4939-7768-0_14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6334841PMC
May 2019

Genome-Wide Mapping of Protein-DNA Interactions on Nascent Chromatin.

Methods Mol Biol 2018 ;1766:231-238

Department of Biology, Emory University, Atlanta, GA, USA.

Chromatin immunoprecipitation (ChIP) is the most widely used method to analyze protein-DNA interactions in vivo. Coupled with next generation sequencing, ChIP-seq experiments map protein-DNA interactions in a genome-wide fashion. Here we describe a novel method called nasChIP-seq for mapping genome-wide occupancy of posttranslationally modified histones or transcription factors on newly replicated DNA.
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http://dx.doi.org/10.1007/978-1-4939-7768-0_13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6311136PMC
May 2019

Nascent DNA methylome mapping reveals inheritance of hemimethylation at CTCF/cohesin sites.

Science 2018 03;359(6380):1166-1170

Department of Biology, Emory University, 1510 Clifton Road NE, Atlanta, GA 30322, USA.

The faithful inheritance of the epigenome is critical for cells to maintain gene expression programs and cellular identity across cell divisions. We mapped strand-specific DNA methylation after replication forks and show maintenance of the vast majority of the DNA methylome within 20 minutes of replication and inheritance of some hemimethylated CpG dinucleotides (hemiCpGs). Mapping the nascent DNA methylome targeted by each of the three DNA methyltransferases (DNMTs) reveals interactions between DNMTs and substrate daughter cytosines en route to maintenance methylation or hemimethylation. Finally, we show the inheritance of hemiCpGs at short regions flanking CCCTC-binding factor (CTCF)/cohesin binding sites in pluripotent cells. Elimination of hemimethylation causes reduced frequency of chromatin interactions emanating from these sites, suggesting a role for hemimethylation as a stable epigenetic mark regulating CTCF-mediated chromatin interactions.
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http://dx.doi.org/10.1126/science.aan5480DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6359960PMC
March 2018

Developing in 3D: the role of CTCF in cell differentiation.

Development 2018 03 22;145(6). Epub 2018 Mar 22.

Department of Biology, Emory University, Atlanta, GA 30322, USA

CTCF is a highly conserved zinc-finger DNA-binding protein that mediates interactions between distant sequences in the genome. As a consequence, CTCF regulates enhancer-promoter interactions and contributes to the three-dimensional organization of the genome. Recent studies indicate that CTCF is developmentally regulated, suggesting that it plays a role in cell type-specific genome organization. Here, we review these studies and discuss how CTCF functions during the development of various cell and tissue types, ranging from embryonic stem cells and gametes, to neural, muscle and cardiac cells. We propose that the lineage-specific control of CTCF levels, and its partnership with lineage-specific transcription factors, allows for the control of cell type-specific gene expression via chromatin looping.
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http://dx.doi.org/10.1242/dev.137729DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5897592PMC
March 2018

Not just heads and tails: The complexity of the sperm epigenome.

J Biol Chem 2018 09 5;293(36):13815-13820. Epub 2018 Mar 5.

From the Department of Biology, Emory University, Atlanta, Georgia 30322

Transgenerational inheritance requires mechanisms by which epigenetic information is transferred via gametes. Canonical thought holds that mammalian sperm chromatin would be incapable of carrying epigenetic information as post-translational modifications of histones because of their replacement with protamine proteins. Furthermore, compaction of the sperm genome would hinder DNA accessibility of proteins involved in transcriptional regulation and genome architecture. In this Minireview, we delineate the paternal chromatin remodeling events during spermatogenesis and fertilization. Sperm chromatin is epigenetically modified at various time points throughout its development. This allows for the addition of environment-specific modifications that can be passed from parents to offspring.
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http://dx.doi.org/10.1074/jbc.R117.001561DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6130957PMC
September 2018

Evolutionarily Conserved Principles Predict 3D Chromatin Organization.

Mol Cell 2017 Sep 17;67(5):837-852.e7. Epub 2017 Aug 17.

Department of Biology, Emory University, 1510 Clifton Road Northeast, Atlanta, GA 30322, USA. Electronic address:

Topologically associating domains (TADs), CTCF loop domains, and A/B compartments have been identified as important structural and functional components of 3D chromatin organization, yet the relationship between these features is not well understood. Using high-resolution Hi-C and HiChIP, we show that Drosophila chromatin is organized into domains we term compartmental domains that correspond precisely with A/B compartments at high resolution. We find that transcriptional state is a major predictor of Hi-C contact maps in several eukaryotes tested, including C. elegans and A. thaliana. Architectural proteins insulate compartmental domains by reducing interaction frequencies between neighboring regions in Drosophila, but CTCF loops do not play a distinct role in this organism. In mammals, compartmental domains exist alongside CTCF loop domains to form topological domains. The results suggest that compartmental domains are responsible for domain structure in all eukaryotes, with CTCF playing an important role in domain formation in mammals.
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http://dx.doi.org/10.1016/j.molcel.2017.07.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5591081PMC
September 2017
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