Epithelial-mesenchymal transition is an indispensable mechanism during morphogenesis, as without mesenchymal cells, tissues and organs will never be formed. However, epithelial-cell plasticity, coupled to the transient or permanent formation of mesenchyme, goes far beyond the problem of cell-lineage segregation. Understanding how mesenchymal cells arise from an epithelial default status will also have a strong impact in unravelling the mechanisms that control fibrosis and cancer progression.

Complex networks orchestrate epithelial–mesenchymal transitions

Next-generation sequencing technology is a powerful tool for transcriptome analysis. However, under certain conditions, only a small amount of material is available, which requires more sensitive techniques that can preferably be used at the single-cell level. Here we describe a single-cell digital gene expression profiling assay. Using our mRNA-Seq assay with only a single mouse blastomere, we detected the expression of 75% (5,270) more genes than microarray techniques and identified 1,753 previously unknown splice junctions called by at least 5 reads. Moreover, 8-19% of the genes with multiple known transcript isoforms expressed at least two isoforms in the same blastomere or oocyte, which unambiguously demonstrated the complexity of the transcript variants at whole-genome scale in individual cells. Finally, for Dicer1(-/-) and Ago2(-/-) (Eif2c2(-/-)) oocytes, we found that 1,696 and 1,553 genes, respectively, were abnormally upregulated compared to wild-type controls, with 619 genes in common.

/pdf/mrna-seq-whole-transcriptome-analysis-of-a-single-cell-lljqtkg28j.pdf

mRNA-Seq whole-transcriptome analysis of a single cell.

DNA methylation is among the best studied epigenetic modifications and is essential to mammalian development. Although the methylation status of most CpG dinucleotides in the genome is stably propagated through mitosis, improvements to methods for measuring methylation have identified numerous regions in which it is dynamically regulated. In this Review, we discuss key concepts in the function of DNA methylation in mammals, stemming from more than two decades of research, including many recent studies that have elucidated when and where DNA methylation has a regulatory role in the genome. We include insights from early development, embryonic stem cells and adult lineages, particularly haematopoiesis, to highlight the general features of this modification as it participates in both global and localized epigenetic regulation.

DNA methylation: roles in mammalian development

The term non-coding RNA (ncRNA) is commonly employed for RNA that does not encode a protein, but this does not mean that such RNAs do not contain information nor have function. Although it has been generally assumed that most genetic information is transacted by proteins, recent evidence suggests that the majority of the genomes of mammals and other complex organisms is in fact transcribed into ncRNAs, many of which are alternatively spliced and/or processed into smaller products. These ncRNAs include microRNAs and snoRNAs (many if not most of which remain to be identified), as well as likely other classes of yet-to-be-discovered small regulatory RNAs, and tens of thousands of longer transcripts (including complex patterns of interlacing and overlapping sense and antisense transcripts), most of whose functions are unknown. These RNAs (including those derived from introns) appear to comprise a hidden layer of internal signals that control various levels of gene expression in physiology and development, including chromatin architecture/epigenetic memory, transcription, RNA splicing, editing, translation and turnover. RNA regulatory networks may determine most of our complex characteristics, play a significant role in disease and constitute an unexplored world of genetic variation both within and between species.

Non-coding RNA

Overlapping epigenetic mechanisms have evolved in eukaryotic cells to silence the expression and mobility of transposable elements (TEs). Owing to their ability to recruit the silencing machinery, TEs have served as building blocks for epigenetic phenomena, both at the level of single genes and across larger chromosomal regions. Important progress has been made recently in understanding these silencing mechanisms. In addition, new insights have been gained into how this silencing has been co-opted to serve essential functions in 'host' cells, highlighting the importance of TEs in the epigenetic regulation of the genome.

http://depa.fquim.unam.mx/amyd//archivero/Tarea3_23133.pdf

Transposable elements and the epigenetic regulation of the genome.

Retrotransposons regulate host genes in mouse oocytes and preimplantation embryos.

., 1998). Thesematernally derived transcripts are responsible for thecompletion of meiosis, initiation of mitosis, activation ofthe embryonic genome, and completion of the transfor-mation of the highly differentiated oocyte into a totipo-tent embryonic cell. To study the effect of speciﬁc ma-ternal transcripts on these processes, we have sought toestablish a Cre-

https://onlinelibrary.wiley.com/doi/pdf/10.1002/%28SICI%291526-968X%28200002%2926%3A2%3C110%3A%3AAID-GENE2%3E3.0.CO%3B2-8

Expression of Cre recombinase in mouse oocytes: A means to study maternal effect genes

Phenotypic variability in genetic disease is usually attributed to genetic background variation or environmental influence. Here, we show that deletion of a single gene, Trim28 (Kap1 or Tif1β), from the maternal germ line alone, on an otherwise identical genetic background, results in severe phenotypic and epigenetic variability that leads to embryonic lethality. We identify early and minute epigenetic variations in blastomeres of the preimplantation embryo of these animals, suggesting that the embryonic lethality may result from the misregulation of genomic imprinting in mice lacking maternal Trim28. Our results reveal the long-range effects of a maternal gene deletion on epigenetic memory and illustrate the delicate equilibrium of maternal and zygotic factors during nuclear reprogramming.

http://science.sciencemag.org/content/sci/335/6075/1499.full.pdf

Trim28 Is Required for Epigenetic Stability During Mouse Oocyte to Embryo Transition

The oocyte to embryo transition in metazoans depends on maternal proteins and transcripts to ensure the successful initiation of development, and the correct and timely activation of the embryonic genome We conditionally eliminated the maternal gene encoding the cell adhesion molecule E-cadherin and partially eliminated the beta-catenin gene from the mouse oocyte Oocytes lacking E-cadherin, or expressing a truncated allele of beta-catenin without the N-terminal part of the protein, give rise to embryos whose blastomeres do not adhere Blastomere adhesion is restored after translation of protein from the wild-type paternal alleles: at the morula stage in embryos lacking maternal E-cadherin, and at the late four-cell stage in embryos expressing truncated beta-catenin This suggests that adhesion per se is not essential in the early cleavage stage embryos, that embryos develop normally if compaction does not occur until the morula stage, and that the zona pellucida suffices to maintain blastomere proximity Although maternal E-cadherin is not essential for the completion of the oocyte-to-embryo transition, absence of wild-type beta-catenin in oocytes does statistically compromise developmental success rates This developmental deficit is alleviated by the simultaneous absence of maternal E-cadherin, suggesting that E-cadherin regulates nuclear beta-catenin availability during embryonic genome activation

/pdf/maternal-b-catenin-and-e-cadherin-in-mouse-development-2erwchhp54.pdf

Maternal β-catenin and E-cadherin in mouse development

Many components of the Wnt/β-catenin signaling pathway are expressed
during mouse pre-implantation embryo development, suggesting that this pathway
may control cell proliferation and differentiation at this time. We find no
evidence for a functional activity of this pathway in cleavage-stage embryos
using the Wnt-reporter line, BAT-gal. To further probe the activity of this
pathway, we activated β-catenin signaling by mating a zona pellucida3-cre
(Zp3-cre) transgenic mouse line with a mouse line containing an exon3-floxedβ
-catenin allele. The result is expression of a stabilized form ofβ
-catenin, resistant to degradation by the GSK3β-mediated proteasome
pathway, expressed in the developing oocyte and in each cell of the resulting
embryos. Nuclear localization and signaling function of β-catenin were
not observed in cleavage-stage embryos derived from these oocytes. These
results indicate that in pre-implantation embryos, molecular mechanisms
independent of the GSK3β-mediated ubiquitination and proteasome
degradation pathway inhibit the nuclear function of β-catenin. Although
the mutant blastocysts initially developed normally, they then exhibited a
specific phenotype in the embryonic ectoderm layer of early post-implantation
embryos. We show a nuclear function of β-catenin in the mutant epiblast
that leads to activation of Wnt/β-catenin target genes. As a consequence,
cells of the embryonic ectoderm change their fate, resulting in a premature
epithelial-mesenchymal transition.

Wilhelmine N. de Vries

Papers

Retrotransposons regulate host genes in mouse oocytes and preimplantation embryos.

Expression of Cre recombinase in mouse oocytes: A means to study maternal effect genes

Trim28 Is Required for Epigenetic Stability During Mouse Oocyte to Embryo Transition

Maternal β-catenin and E-cadherin in mouse development

Stabilization of β-catenin in the mouse zygote leads to premature epithelial-mesenchymal transition in the epiblast