Genome duplication has played a pivotal role in the evolution of many eukaryotic lineages, including the vertebrates. A relatively recent vertebrate genome duplication is that in Xenopus laevis, ...which resulted from the hybridization of two closely related species about 17 million years ago. However, little is known about the consequences of this duplication at the level of the genome, the epigenome, and gene expression.
The X. laevis genome consists of two subgenomes, referred to as L (long chromosomes) and S (short chromosomes), that originated from distinct diploid progenitors. Of the parental subgenomes, S chromosomes have degraded faster than L chromosomes from the point of genome duplication until the present day. Deletions appear to have the largest effect on pseudogene formation and loss of regulatory regions. Deleted regions are enriched for long DNA repeats and the flanking regions have high alignment scores, suggesting that non-allelic homologous recombination has played a significant role in the loss of DNA. To assess innovations in the X. laevis subgenomes we examined p300-bound enhancer peaks that are unique to one subgenome and absent from X. tropicalis. A large majority of new enhancers comprise transposable elements. Finally, to dissect early and late events following interspecific hybridization, we examined the epigenome and the enhancer landscape in X. tropicalis × X. laevis hybrid embryos. Strikingly, young X. tropicalis DNA transposons are derepressed and recruit p300 in hybrid embryos.
The results show that erosion of X. laevis genes and functional regulatory elements is associated with repeats and non-allelic homologous recombination and furthermore that young repeats have also contributed to the p300-bound regulatory landscape following hybridization and whole-genome duplication.
In this study we analyzed transcript abundance and promoters of genes encoding mitochondrial proteins to identify signaling pathways that regulate stress-induced gene expression. We used Arabidopsis ...(Arabidopsis thaliana) alternative oxidase AOX1a, external NADP H-dehydrogenase NDB2, and two additional highly stress-responsive genes, At2g21640 and BCS1. As a starting point, the promoter region of AOX1a was analyzed and functional analysis identified 10 cis-acting regulatory elements (CAREs), which played a role in response to treatment with H₂O₂, rotenone, or both. Six of these elements were also functional in the NDB2 promoter. The promoter region of At2g21640, previously defined as a hallmark of oxidative stress, shared two functional CAREs with AOX1a and was responsive to treatment with H₂O₂ but not rotenone. Microarray analysis further supported that signaling pathways induced by H₂O₂ and rotenone are not identical. The promoter of BCS1 was not responsive to H₂O₂ or rotenone, but highly responsive to salicylic acid (SA), whereas the promoters of AOX1a and NDB2 were unresponsive to SA. Analysis of transcript abundance of these genes in a variety of defense signaling mutants confirmed that BCS1 expression is regulated in a different manner compared to AOX1a, NDB2, and At2g21640. These mutants also revealed a pathway associated with programmed cell death that regulated AOX1a in a manner distinct from the other genes. Thus, at least three distinctive pathways regulate mitochondrial stress response at a transcriptional level, an SA-dependent pathway represented by BCS1, a second pathway that represents a convergence point for signals generated by H₂O₂ and rotenone on multiple CAREs, some of which are shared between responsive genes, and a third pathway that acts via EDS1 and PAD4 regulating only AOX1a. Furthermore, posttranscriptional regulation accounts for changes in transcript abundance by SA treatment for some genes.
Our current understanding of induced pluripotent stem cell (iPSC) generation has almost entirely been shaped by studies performed on reprogramming fibroblasts. However, whether the resulting model ...universally applies to the reprogramming process of other cell types is still largely unknown. By characterizing and profiling the reprogramming pathways of fibroblasts, neutrophils, and keratinocytes, we unveil that key events of the process, including loss of original cell identity, mesenchymal to epithelial transition, the extent of developmental reversion, and reactivation of the pluripotency network, are to a large degree cell-type specific. Thus, we reveal limitations for the use of fibroblasts as a universal model for the study of the reprogramming process and provide crucial insights about iPSC generation from alternative cell sources.
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•Cell-type-specific molecular reprogramming trajectories converge at the iPSC stage•Two transcriptional waves are common to all reprogramming pathways•Transient changes during reprogramming are mostly cell-type specific•Egr1 downregulation is part of a conserved transcriptional program
Nefzger et al. find that the molecular reprogramming trajectories of fibroblasts, neutrophils, and keratinocytes have a cell-type-specific component that only fully converges in induced pluripotent stem cells. The authors also identify universal changes shared by all three cell types, including two transcriptional waves and a conserved transcriptional program involving Egr1 downregulation.
DNA methylation patterns are established in early embryogenesis and are critical for cellular differentiation. To investigate the role of CG methylation in potential enhancer formation, we assessed ...H3K4me1 modification in murine embryonic fibroblasts (MEFs) derived from the DNA methylation mutant Lsh ⁻/⁻ mice. We report here de novo formation of putative enhancer elements at CG hypomethylated sites that can be dynamically altered. We found a subset of differentially enriched H3K4me1 regions clustered at neuronal lineage genes and overlapping with known cis-regulatory elements present in brain tissue. Reprogramming of Lsh ⁻/⁻ MEFs into induced pluripotent stem (iPS) cells leads to increased neuronal lineage gene expression of premarked genes and enhanced differentiation potential of Lsh ⁻/⁻ iPS cells toward the neuronal lineage pathway compared with WT iPS cells in vitro and in vivo. The state of CG hypomethylation and H3K4me1 enrichment is partially maintained in Lsh ⁻/⁻ iPS cells. The acquisition of H3K27ac and activity of subcloned fragments in an enhancer reporter assay indicate functional activity of several of de novo H3K4me1-marked sequences. Our results suggest a functional link of H3K4me1 enrichment at CG hypomethylated sites, enhancer formation, and cellular plasticity.
Plant mitochondria contain non-phosphorylating bypasses of the respiratory chain, catalysed by the alternative oxidase (AOX) and alternative NADH dehydrogenases (NDH), as well as uncoupling (UCP) ...protein. Each of these components either circumvents or short-circuits proton translocation pathways, and each is encoded by a small gene family in Arabidopsis. Whole genome microarray experiments were performed with suspension cell cultures to examine the effects of various 3 h treatments designed to induce abiotic stress. The expression of over 60 genes encoding components of the classical, phosphorylating respiratory chain and tricarboxylic acid cycle remained largely constant when cells were subjected to a broad range of abiotic stresses, but expression of the alternative components responded differentially to the various treatments. In detailed time-course quantitative PCR analysis, specific members of both AOX and NDH gene families displayed coordinated responses to treatments. In particular, the co-expression of AOX1a and NDB2 observed under a number of treatments suggested co-regulation that may be directed by common sequence elements arranged hierarchically in the upstream promoter regions of these genes. A series of treatment sets were identified, representing the response of specific AOX and NDH genes to mitochondrial inhibition, plastid inhibition and abiotic stresses. These treatment sets emphasise the multiplicity of pathways affecting alternative electron transport components in plants.
Cytosine DNA methylation (5mC) is a widespread base modification in eukaryotic genomes with critical roles in transcriptional regulation. In recent years, our understanding of 5mC has changed because ...of advances in 5mC detection techniques that allow mapping of this mark on the whole genome scale. Profiling DNA methylomes from organisms across the eukaryotic tree of life has reshaped our views on the evolution of 5mC. In this review, we explore the macroevolution of 5mC in major eukaryotic groups, and then focus on recent advances made in animals. Genomic 5mC patterns as well as the mechanisms of 5mC deposition tend to be evolutionary labile across large phylogenetic distances; however, some common patterns are starting to emerge. Within the animal kingdom, 5mC diversity has proven to be much greater than anticipated. For example, a previously held common view that genome hypermethylation is a trait exclusive to vertebrates has recently been challenged. Also, data from genome-wide studies are starting to yield insights into the potential roles of 5mC in invertebrate cis regulation. Here we provide an evolutionary perspective of both the well-known and enigmatic roles of 5mC across the eukaryotic tree of life.
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•Insights from whole genome bisulphite sequencing have revealed unprecedented diversity in methylome patterning in eukaryotes.•The presence, absence, or diverse combinations of DNMTs in a given genome are poor predictors of genomic 5mC patterns.•Genomic hypermethylation is observed more broadly within the animal kingdom than previously anticipated.
We currently lack a comprehensive understanding of the mechanisms underlying neural tube formation and their contributions to neural tube defects (NTDs). Developing a model to study such a complex ...morphogenetic process, especially one that models human-specific aspects, is critical. Three-dimensional, human embryonic stem cell (hESC)-derived neural rosettes (NRs) provide a powerful resource for in vitro modeling of human neural tube formation. Epigenomic maps reveal enhancer elements unique to NRs relative to 2D systems. A master regulatory network illustrates that key NR properties are related to their epigenomic landscapes. We found that folate-associated DNA methylation changes were enriched within NR regulatory elements near genes involved in neural tube formation and metabolism. Our comprehensive regulatory maps offer insights into the mechanisms by which folate may prevent NTDs. Lastly, our distal regulatory maps provide a better understanding of the potential role of neurological-disorder-associated SNPs.
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•Neural rosettes have transcriptional and epigenomic landscapes distinct from 2D NSCs•Low methylated regions in NRs predict enhancers active later in neurodevelopment•NR regulatory maps provide insights into mechanisms of neural tube defects
Neural rosettes offer a 3D model of neural tube development. Valensisi et al. show that these cells have distinct epigenomic landscapes compared to 2D neural stem cells. cis-regulatory elements identified in NRs harbor folate-associated CpGs, which may be important during neural tube development, and neurodevelopmental-disorder-associated variants.
Abstract
Summary
Due to the scale and sparsity of single-cell RNA-sequencing data, traditional plots can obscure vital information. Our R package schex overcomes this by implementing hexagonal ...binning, which has the additional advantages of improving speed and reducing storage for resulting plots.
Availability and implementation
schex is freely available from Bioconductor via http://bioconductor.org/packages/release/bioc/html/schex.html and its development version can be accessed on GitHub via https://github.com/SaskiaFreytag/schex.
Supplementary information
Supplementary data are available at Bioinformatics online.
Epithelial‐mesenchymal transition (EMT) is a reversible transcriptional program invoked by cancer cells to drive cancer progression. Transcription factor ZEB1 is a master regulator of EMT, driving ...disease recurrence in poor‐outcome triple negative breast cancers (TNBCs). Here, this work silences ZEB1 in TNBC models by CRISPR/dCas9‐mediated epigenetic editing, resulting in highly‐specific and nearly complete suppression of ZEB1 in vivo, accompanied by long‐lasting tumor inhibition. Integrated “omic” changes promoted by dCas9 linked to the KRAB domain (dCas9‐KRAB) enabled the discovery of a ZEB1‐dependent‐signature of 26 genes differentially‐expressed and ‐methylated, including the reactivation and enhanced chromatin accessibility in cell adhesion loci, outlining epigenetic reprogramming toward a more epithelial state. In the ZEB1 locus transcriptional silencing is associated with induction of locally‐spread heterochromatin, significant changes in DNA methylation at specific CpGs, gain of H3K9me3, and a near complete erasure of H3K4me3 in the ZEB1 promoter. Epigenetic shifts induced by ZEB1‐silencing are enriched in a subset of human breast tumors, illuminating a clinically‐relevant hybrid‐like state. Thus, the synthetic epi‐silencing of ZEB1 induces stable “lock‐in” epigenetic reprogramming of mesenchymal tumors associated with a distinct and stable epigenetic landscape. This work outlines epigenome‐engineering approaches for reversing EMT and customizable precision molecular oncology approaches for targeting poor outcome breast cancers.
This study harnesses CRISPR/dCas9 epigenome‐engineering to reprogram mesenchymal‐enriched breast cancers. Key orchestrator of EMT, ZEB1 is epigenetically silenced at mRNA and protein levels. Silencing is maintained in vivo with impaired tumor growth. There is an up‐regulation of epithelial gene programs, alteration of epigenetic landscape, and a signature of 26‐differentially expressed and methylated genes predictive of prognosis in breast cancer.
DNA methylation in neurons is directly linked to neuronal genome regulation and maturation. Unlike other tissues, vertebrate neurons accumulate high levels of atypical DNA methylation in the CH ...sequence context (mCH) during early postnatal brain development. Here, we investigate to what extent neurons derived in vitro from both mouse and human pluripotent stem cells recapitulate in vivo DNA methylation patterns. While human ESC-derived neurons did not accumulate mCH in either 2D culture or 3D organoid models even after prolonged culture, cortical neurons derived from mouse ESCs acquired in vivo levels of mCH over a similar time period in both primary neuron cultures and in vivo development. mESC-derived neuron mCH deposition was coincident with a transient increase in Dnmt3a, preceded by the postmitotic marker Rbfox3 (NeuN), was enriched at the nuclear lamina, and negatively correlated with gene expression. We further found that methylation patterning subtly differed between in vitro mES-derived and in vivo neurons, suggesting the involvement of additional noncell autonomous processes. Our findings show that mouse ESC-derived neurons, in contrast to those of humans, can recapitulate the unique DNA methylation landscape of adult neurons in vitro over experimentally tractable timeframes, which allows their use as a model system to study epigenome maturation over development.
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