The landscape of accessible chromatin in bovine oocytes and early embryos.
TL;DR: It is found that chromatin accessibility is low in oocytes and 2-/4-cell embryos, followed by a significant increase in embryos during major embryonic genome activation (EGA), and peaked in elongating day 14 embryos.
Abstract: Chromatin reorganization governs the regulation of gene expression during preimplantation development. However, the landscape of chromatin dynamics in this period has not been explored in bovine. In this study, we constructed a genome-wide map of accessible chromatin in bovine oocytes and early embryos using an improved assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) which revealed unique features of the accessible chromatin during bovine early embryo development. We found that chromatin accessibility is low in oocytes and 2-/4-cell embryos, followed by a significant increase in embryos during major embryonic genome activation (EGA), and peaked in elongating day 14 embryos. Genome-wide characteristics of open chromatin showed that ATAC-seq signals in both transcription start sites (TSS) and transcription end sites (TES) were strong. Additionally, the distal ATAC-seq peaks were enriched in repeat elements in a type-specific and stage-specific manner. We further unveiled a series of transcription factor (TF) motifs with distinct variation of enrichment from distal ATAC-seq peaks. By integrated analysis of chromatin accessibility with transcriptomes and DNA methylomes in bovine early embryos, we showed that promoter accessibility was positively correlated with gene expression, especially during major EGA, and was strongly correlated to DNA methylation and CpG density. Finally, we identified the critical chromatin signatures and TFs that differ between in vivo and in vitro derived blastocysts, which provides insights to the potential mechanisms leading to low quality of embryos produced in vitro. Together, this comprehensive analysis revealed critical features of chromatin landscape and epigenetic reprogramming during bovine preimplantation embryo development.
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TL;DR: In this article, the effects of parental environmental exposures on the epigenetics of gametes and the early embryo, and evidence for transgenerational inheritance in livestock species was discussed.
Abstract: The epigenome is dynamic and forged by epigenetic mechanisms, such as DNA methylation, histone modifications, chromatin remodeling, and non-coding RNA species. Increasing lines of evidence support the concept that certain acquired traits are derived from environmental exposure during early embryonic and fetal development, i.e., fetal programming, and can even be "memorized" in the germline as epigenetic information and transmitted to future generations. Advances in technology are now driving the global profiling and precise editing of germline and embryonic epigenomes, thereby improving our understanding of epigenetic regulation and inheritance. These achievements open new avenues for the development of technologies or potential management interventions to counteract adverse conditions or improve performance in livestock species. In this article, we review the epigenetic analyses (DNA methylation, histone modification, chromatin remodeling, and non-coding RNAs) of germ cells and embryos in mammalian livestock species (cattle, sheep, goats, and pigs) and the epigenetic determinants of gamete and embryo viability. We also discuss the effects of parental environmental exposures on the epigenetics of gametes and the early embryo, and evidence for transgenerational inheritance in livestock.
17 citations
TL;DR: The Ruminant Genome Database (RGD) v2.0 as mentioned in this paper provides visualization and analysis tools for ruminant comparative genomics and functional annotations, including the LiftOver tool, which allows coordinate conversion between different RGD assemblies.
Abstract: Ruminant Genome Database (RGD; http://animal.nwsuaf.edu.cn/RGD) provides visualization and analysis tools for ruminant comparative genomics and functional annotations. As more high-quality ruminant genome assemblies have become available, we have redesigned the user interface, integrated and expanded multi-omics data, and developed novel features to improve the database. The new version, RGD v2.0, houses 78 ruminant genomes; 110-species synteny alignments for major livestock (including cattle, sheep, goat) and wild ungulates; 21 012 orthologous gene clusters with Gene Ontology and pathway annotation; ∼8 600 000 conserved elements; and ∼1 000 000 cis-regulatory elements by utilizing 1053 epigenomic data sets. The transcriptome data in RGD v2.0 has nearly doubled, currently with 1936 RNA-seq data sets, and 155 174 phenotypic data sets have been newly added. New and updated features include: (i) The UCSC Genome Browser, BLAT, BLAST and Table Browser tools were updated for six available ruminant livestock species. (ii) The LiftOver tool was newly introduced into our browser to allow coordinate conversion between different ruminant assemblies. And (iii) tissue specificity index, tau, was calculated to facilitate batch screening of specifically expressed genes. The enhanced genome annotations and improved functionality in RGD v2.0 will be useful for study of genome evolution, environmental adaption, livestock breeding and biomedicine.
16 citations
TL;DR: In this paper, the authors review parental non-genetic effects retained in gametes on early embryo development of dairy cows, with emphasis on parental age (around puberty), the metabolism of the mother at the time of conception and in vitro culture (IVC) conditions.
Abstract: The bovine represents an important agriculture species and dairy breeds have experienced intense genetic selection over the last decades. The selection of breeders focused initially on milk production, but now includes feed efficiency, health, and fertility, although these traits show lower heritability. The non-genetic paternal and maternal effects on the next generation represent a new research topic that is part of epigenetics. The evidence for embryo programming from both parents is increasing. Both oocytes and spermatozoa carry methylation marks, histones modifications, small RNAs, and chromatin state variations. These epigenetic modifications may remain active in the early zygote and influence the embryonic period and beyond. In this paper, we review parental non-genetic effects retained in gametes on early embryo development of dairy cows, with emphasis on parental age (around puberty), the metabolism of the mother at the time of conception and in vitro culture (IVC) conditions. In our recent findings, transcriptomic signatures and DNA methylation patterns of blastocysts and gametes originating from various parental and IVC conditions revealed surprisingly similar results. Embryos from all these experiments displayed a metabolic signature that could be described as an "economy" mode where protein synthesis is reduced, mitochondria are considered less functional. In the absence of any significant phenotype, these results indicated a possible similar adaptation of the embryo to younger parental age, post-partum metabolic status and IVC conditions mediated by epigenetic factors.
14 citations
TL;DR: This work indicates that chromatin accessibility plays an important regulatory role in the embryonic muscle development of pigs and regulates the temporal and spatial expression patterns of key genes in muscle development by influencing the binding of transcription factors.
Abstract: The development of skeletal muscle in pigs during the embryonic stage is precisely regulated by transcriptional mechanisms, which depend on chromatin accessibility However, how chromatin accessibility plays a regulatory role during embryonic skeletal muscle development in pigs has not been reported To gain insight into the landscape of chromatin accessibility and the associated genome-wide transcriptome during embryonic muscle development, we performed ATAC-seq and RNA-seq analyses of skeletal muscle from pig embryos at 45, 70 and 100 days post coitus (dpc) In total, 21,638, 35,447 and 60,181 unique regions (or peaks) were found across the embryos at 45 dpc (LW45), 70 dpc (LW70) and 100 dpc (LW100), respectively More than 91% of the peaks were annotated within − 1 kb to 100 bp of transcription start sites (TSSs) First, widespread increases in specific accessible chromatin regions (ACRs) from embryos at 45 to 100 dpc suggested that the regulatory mechanisms became increasingly complicated during embryonic development Second, the findings from integrated ATAC-seq and RNA-seq analyses showed that not only the numbers but also the intensities of ACRs could control the expression of associated genes Moreover, the motif screening of stage-specific ACRs revealed some transcription factors that regulate muscle development-related genes, such as MyoG, Mef2c, and Mef2d Several potential transcriptional repressors, including E2F6, OTX2 and CTCF, were identified among the genes that exhibited different regulation trends between the ATAC-seq and RNA-seq data This work indicates that chromatin accessibility plays an important regulatory role in the embryonic muscle development of pigs and regulates the temporal and spatial expression patterns of key genes in muscle development by influencing the binding of transcription factors Our results contribute to a better understanding of the regulatory dynamics of genes involved in pig embryonic skeletal muscle development
12 citations
TL;DR: In this paper, gene expression profiling and genome-wide accessible chromatin mapping of mouse cardiac fibroblasts isolated from the uninjured myocardium and the infarct at multiple time points corresponding to different differentiation states were performed by RNA sequencing (RNA-seq) and the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq), respectively.
Abstract: After myocardial infarction, the massive death of cardiomyocytes leads to cardiac fibroblast proliferation and myofibroblast differentiation, which contributes to the extracellular matrix remodeling of the infarcted myocardium. We recently found that myofibroblasts further differentiate into matrifibrocytes, a newly identified cardiac fibroblast differentiation state. Cardiac fibroblasts of different states have distinct gene expression profiles closely related to their functions. However, the mechanism responsible for the gene expression changes during these activation and differentiation events is still not clear. In this study, the gene expression profiling and genome-wide accessible chromatin mapping of mouse cardiac fibroblasts isolated from the uninjured myocardium and the infarct at multiple time points corresponding to different differentiation states were performed by RNA sequencing (RNA-seq) and the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq), respectively. ATAC-seq peaks were highly enriched in the promoter area and the distal area where enhancers are located. A positive correlation was identified between the expression and promoter accessibility for many dynamically expressed genes, even though evidence showed that mechanisms independent of chromatin accessibility may also contribute to the gene expression changes in cardiac fibroblasts after MI. Moreover, motif enrichment analysis and gene regulatory network construction identified transcription factors that possibly contributed to the differential gene expression between cardiac fibroblasts of different states.
11 citations
References
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TL;DR: This work presents Model-based Analysis of ChIP-Seq data, MACS, which analyzes data generated by short read sequencers such as Solexa's Genome Analyzer, and uses a dynamic Poisson distribution to effectively capture local biases in the genome, allowing for more robust predictions.
Abstract: We present Model-based Analysis of ChIP-Seq data, MACS, which analyzes data generated by short read sequencers such as Solexa's Genome Analyzer. MACS empirically models the shift size of ChIP-Seq tags, and uses it to improve the spatial resolution of predicted binding sites. MACS also uses a dynamic Poisson distribution to effectively capture local biases in the genome, allowing for more robust predictions. MACS compares favorably to existing ChIP-Seq peak-finding algorithms, and is freely available.
13,008 citations
TL;DR: It is shown here that P-clouds predicts >840 Mbp of additional repetitive sequences in the human genome, thus suggesting that 66%–69% of the human chromosome is repetitive or repeat-derived, and that the human genomes consists of substantially more repetitive sequence than previously believed.
Abstract: Transposable elements (TEs) are conventionally identified in eukaryotic genomes by alignment to consensus element sequences. Using this approach, about half of the human genome has been previously identified as TEs and low-complexity repeats. We recently developed a highly sensitive alternative de novo strategy, P-clouds, that instead searches for clusters of high-abundance oligonucleotides that are related in sequence space (oligo “clouds”). We show here that P-clouds predicts >840 Mbp of additional repetitive sequences in the human genome, thus suggesting that 66%–69% of the human genome is repetitive or repeat-derived. To investigate this remarkable difference, we conducted detailed analyses of the ability of both P-clouds and a commonly used conventional approach, RepeatMasker (RM), to detect different sized fragments of the highly abundant human Alu and MIR SINEs. RM can have surprisingly low sensitivity for even moderately long fragments, in contrast to P-clouds, which has good sensitivity down to small fragment sizes (∼25 bp). Although short fragments have a high intrinsic probability of being false positives, we performed a probabilistic annotation that reflects this fact. We further developed “element-specific” P-clouds (ESPs) to identify novel Alu and MIR SINE elements, and using it we identified ∼100 Mb of previously unannotated human elements. ESP estimates of new MIR sequences are in good agreement with RM-based predictions of the amount that RM missed. These results highlight the need for combined, probabilistic genome annotation approaches and suggest that the human genome consists of substantially more repetitive sequence than previously believed.
983 citations
TL;DR: Oct-4 expression in the germline is regulated separately from epiblast expression, and this provides the first marker for the identification of totipotent cells in the embryo, and suggests that expression of Oct-4 in the Totipotent cycle is dependent on a set of factors unique to the germ line.
Abstract: The totipotent stem cells of the pregastrulation mouse embryo which give rise to all embryonic somatic tissues and germ cells express Oct-4. The expression is downregulated during gastrulation and is thereafter only maintained in the germline lineage. Oct-4/lacZ transgenes were used to determine how this pattern of expression was achieved, and resulted in the identification of two separate regulatory elements. The distal element drives Oct-4 expression in preimplantation embryos, in migratory and postmigratory primordial germ cells but is inactive in cells of the epiblast. In cell lines this element is specifically active in embryonic stem and embryonic germ cells. The proximal element directs the epiblast-specific expression pattern, including downregulation during gastrulation; in cell lines its activity is restricted to epiblast-derived cells. Thus, Oct-4 expression in the germline is regulated separately from epiblast expression. This provides the first marker for the identification of totipotent cells in the embryo, and suggests that expression of Oct-4 in the totipotent cycle is dependent on a set of factors unique to the germline.
921 citations
TL;DR: Four different situations have been identified that result in the large offspring syndrome: in vitro embryo culture, asynchronous embryo transfer into an advanced uterine environment, nuclear transfer and maternal exposure to excessively high urea diets.
Abstract: Bovine and ovine embryos exposed to a variety of unusual environments prior to the blastocyst stage have resulted in the development of unusually large offspring which can also exhibit a number of organ defects. In these animals, the increased incidence of difficult parturition and of fetal and neonatal losses has limited the large-scale use of in vitro embryo production technologies commonly used in humans and other species. Four different situations have been identified that result in the syndrome: in vitro embryo culture, asynchronous embryo transfer into an advanced uterine environment, nuclear transfer and maternal exposure to excessively high urea diets. However, programming of the syndrome by all of these situations is unpredictable and not all of the symptoms described have been observed universally. Neither the environmental factors inducing the large offspring syndrome nor the mechanisms of perturbation occurring in the early embryo and manifesting themselves in the fetus have been identified.
912 citations
TL;DR: These chromatin accessibility profiles identify cancer- and tissue-specific DNA regulatory elements that enable classification of tumor subtypes with newly recognized prognostic importance, and identify distinct TF activities in cancer based on differences in the inferred patterns of TF-DNA interaction and gene expression.
Abstract: INTRODUCTION Cancer is one of the leading causes of death worldwide. Although the 2% of the human genome that encodes proteins has been extensively studied, much remains to be learned about the noncoding genome and gene regulation in cancer. Genes are turned on and off in the proper cell types and cell states by transcription factor (TF) proteins acting on DNA regulatory elements that are scattered over the vast noncoding genome and exert long-range influences. The Cancer Genome Atlas (TCGA) is a global consortium that aims to accelerate the understanding of the molecular basis of cancer. TCGA has systematically collected DNA mutation, methylation, RNA expression, and other comprehensive datasets from primary human cancer tissue. TCGA has served as an invaluable resource for the identification of genomic aberrations, altered transcriptional networks, and cancer subtypes. Nonetheless, the gene regulatory landscapes of these tumors have largely been inferred through indirect means. RATIONALE A hallmark of active DNA regulatory elements is chromatin accessibility. Eukaryotic genomes are compacted in chromatin, a complex of DNA and proteins, and only the active regulatory elements are accessible by the cell’s machinery such as TFs. The assay for transposase-accessible chromatin using sequencing (ATAC-seq) quantifies DNA accessibility through the use of transposase enzymes that insert sequencing adapters at these accessible chromatin sites. ATAC-seq enables the genome-wide profiling of TF binding events that orchestrate gene expression programs and give a cell its identity. RESULTS We generated high-quality ATAC-seq data in 410 tumor samples from TCGA, identifying diverse regulatory landscapes across 23 cancer types. These chromatin accessibility profiles identify cancer- and tissue-specific DNA regulatory elements that enable classification of tumor subtypes with newly recognized prognostic importance. We identify distinct TF activities in cancer based on differences in the inferred patterns of TF-DNA interaction and gene expression. Genome-wide correlation of gene expression and chromatin accessibility predicts tens of thousands of putative interactions between distal regulatory elements and gene promoters, including key oncogenes and targets in cancer immunotherapy, such as MYC , SRC , BCL2 , and PDL1 . Moreover, these regulatory interactions inform known genetic risk loci linked to cancer predisposition, nominating biochemical mechanisms and target genes for many cancer-linked genetic variants. Lastly, integration with mutation profiling by whole-genome sequencing identifies cancer-relevant noncoding mutations that are associated with altered gene expression. A single-base mutation located 12 kilobases upstream of the FGD4 gene, a regulator of the actin cytoskeleton, generates a putative de novo binding site for an NKX TF and is associated with an increase in chromatin accessibility and a concomitant increase in FGD4 gene expression. CONCLUSION The accessible genome of primary human cancers provides a wealth of information on the susceptibility, mechanisms, prognosis, and potential therapeutic strategies of diverse cancer types. Prediction of interactions between DNA regulatory elements and gene promoters sets the stage for future integrative gene regulatory network analyses. The discovery of hundreds of noncoding somatic mutations that exhibit allele-specific regulatory effects suggests a pervasive mechanism for cancer cells to manipulate gene expression and increase cellular fitness. These data may serve as a foundational resource for the cancer research community.
774 citations