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Nature Sep 2016Maternal-to-zygotic transition (MZT) is essential for the formation of a new individual, but is still poorly understood despite recent progress in analysis of gene...
Maternal-to-zygotic transition (MZT) is essential for the formation of a new individual, but is still poorly understood despite recent progress in analysis of gene expression and DNA methylation in early embryogenesis. Dynamic histone modifications may have important roles in MZT, but direct measurements of chromatin states have been hindered by technical difficulties in profiling histone modifications from small quantities of cells. Recent improvements allow for 500 cell-equivalents of chromatin per reaction, but require 10,000 cells for initial steps or require a highly specialized microfluidics device that is not readily available. We developed a micro-scale chromatin immunoprecipitation and sequencing (μChIP-seq) method, which we used to profile genome-wide histone H3 lysine methylation (H3K4me3) and acetylation (H3K27ac) in mouse immature and metaphase II oocytes and in 2-cell and 8-cell embryos. Notably, we show that ~22% of the oocyte genome is associated with broad H3K4me3 domains that are anti-correlated with DNA methylation. The H3K4me3 signal becomes confined to transcriptional-start-site regions in 2-cell embryos, concomitant with the onset of major zygotic genome activation. Active removal of broad H3K4me3 domains by the lysine demethylases KDM5A and KDM5B is required for normal zygotic genome activation and is essential for early embryo development. Our results provide insight into the onset of the developmental program in mouse embryos and demonstrate a role for broad H3K4me3 domains in MZT.
Topics: Acetylation; Animals; Cell Line, Tumor; Chromatin; Chromatin Immunoprecipitation; DNA Methylation; Embryonic Development; Female; Gene Expression Regulation, Developmental; Genome; Histones; Humans; Lysine; Male; Methylation; Mice; Oocytes; Sequence Analysis, DNA; Transcription Initiation Site; Zygote
PubMed: 27626377
DOI: 10.1038/nature19360 -
Nature Jun 2023Translation regulation is critical for early mammalian embryonic development. However, previous studies had been restricted to bulk measurements, precluding precise...
Translation regulation is critical for early mammalian embryonic development. However, previous studies had been restricted to bulk measurements, precluding precise determination of translation regulation including allele-specific analyses. Here, to address this challenge, we developed a novel microfluidic isotachophoresis (ITP) approach, named RIBOsome profiling via ITP (Ribo-ITP), and characterized translation in single oocytes and embryos during early mouse development. We identified differential translation efficiency as a key mechanism regulating genes involved in centrosome organization and N-methyladenosine modification of RNAs. Our high-coverage measurements enabled, to our knowledge, the first analysis of allele-specific ribosome engagement in early development. These led to the discovery of stage-specific differential engagement of zygotic RNAs with ribosomes and reduced translation efficiency of transcripts exhibiting allele-biased expression. By integrating our measurements with proteomics data, we discovered that ribosome occupancy in germinal vesicle-stage oocytes is the predominant determinant of protein abundance in the zygote. The Ribo-ITP approach will enable numerous applications by providing high-coverage and high-resolution ribosome occupancy measurements from ultra-low input samples including single cells.
Topics: Animals; Mice; Protein Biosynthesis; Proteomics; Ribosomes; RNA, Messenger; Single-Cell Analysis; Embryonic Development; Alleles; Microfluidic Analytical Techniques; Oocytes; Isotachophoresis; Ribosome Profiling; Centrosome; Zygote
PubMed: 37344592
DOI: 10.1038/s41586-023-06228-9 -
Cell Systems Jul 2020Zygotic genome activation (ZGA) is an essential transcriptional event in embryonic development that coincides with extensive epigenetic reprogramming. Complex...
Zygotic genome activation (ZGA) is an essential transcriptional event in embryonic development that coincides with extensive epigenetic reprogramming. Complex manipulation techniques and maternal stores of proteins preclude large-scale functional screens for ZGA regulators within early embryos. Here, we combined pooled CRISPR activation (CRISPRa) with single-cell transcriptomics to identify regulators of ZGA-like transcription in mouse embryonic stem cells, which serve as a tractable, in vitro proxy of early mouse embryos. Using multi-omics factor analysis (MOFA+) applied to ∼200,000 single-cell transcriptomes comprising 230 CRISPRa perturbations, we characterized molecular signatures of ZGA and uncovered 24 factors that promote a ZGA-like response. Follow-up assays validated top screen hits, including the DNA-binding protein Dppa2, the chromatin remodeler Smarca5, and the transcription factor Patz1, and functional experiments revealed that Smarca5's regulation of ZGA-like transcription is dependent on Dppa2. Together, our single-cell transcriptomic profiling of CRISPRa-perturbed cells provides both system-level and molecular insights into the mechanisms that orchestrate ZGA.
Topics: Clustered Regularly Interspaced Short Palindromic Repeats; Epigenesis, Genetic; Genome; Humans; Transcriptome; Zygote
PubMed: 32634384
DOI: 10.1016/j.cels.2020.06.004 -
Nature Jan 2024DNA replication enables genetic inheritance across the kingdoms of life. Replication occurs with a defined temporal order known as the replication timing (RT) programme,...
DNA replication enables genetic inheritance across the kingdoms of life. Replication occurs with a defined temporal order known as the replication timing (RT) programme, leading to organization of the genome into early- or late-replicating regions. RT is cell-type specific, is tightly linked to the three-dimensional nuclear organization of the genome and is considered an epigenetic fingerprint. In spite of its importance in maintaining the epigenome, the developmental regulation of RT in mammals in vivo has not been explored. Here, using single-cell Repli-seq, we generated genome-wide RT maps of mouse embryos from the zygote to the blastocyst stage. Our data show that RT is initially not well defined but becomes defined progressively from the 4-cell stage, coinciding with strengthening of the A and B compartments. We show that transcription contributes to the precision of the RT programme and that the difference in RT between the A and B compartments depends on RNA polymerase II at zygotic genome activation. Our data indicate that the establishment of nuclear organization precedes the acquisition of defined RT features and primes the partitioning of the genome into early- and late-replicating domains. Our work sheds light on the establishment of the epigenome at the beginning of mammalian development and reveals the organizing principles of genome organization.
Topics: Animals; Mice; Blastocyst; Chromatin; DNA Replication Timing; Epigenome; Genome; RNA Polymerase II; Zygote; Embryo, Mammalian
PubMed: 38123678
DOI: 10.1038/s41586-023-06872-1 -
Seminars in Cell & Developmental Biology Jan 2023Brown algae are a group of multicellular, heterokont algae that have convergently evolved developmental complexity that rivals that of embryophytes, animals or fungi.... (Review)
Review
Brown algae are a group of multicellular, heterokont algae that have convergently evolved developmental complexity that rivals that of embryophytes, animals or fungi. Early in development, brown algal zygotes establish a basal and an apical pole, which will become respectively the basal system (holdfast) and the apical system (thallus) of the adult alga. Brown algae are interesting models for understanding the establishment of cell polarity in a broad evolutionary context, because they exhibit a large diversity of life cycles, reproductive strategies and, importantly, their zygotes are produced in large quantities free of parental tissue, with symmetry breaking and asymmetric division taking place in a highly synchronous manner. This review describes the current knowledge about the establishment of the apical-basal axis in the model brown seaweeds Ectocarpus, Dictyota, Fucus and Saccharina, highlighting the advantages and specific interests of each system. Ectocarpus is a genetic model system that allows access to the molecular basis of early development and life-cycle control over apical-basal polarity. The oogamous brown alga Fucus, together with emerging comparative models Dictyota and Saccharina, emphasize the diversity of strategies of symmetry breaking in determining a cell polarity vector in brown algae. A comparison with symmetry-breaking mechanisms in land plants, animals and fungi, reveals that the one-step zygote polarisation of Fucus compares well to Saccharomyces budding and Arabidopsis stomata development, while the two-phased symmetry breaking in the Dictyota zygote compares to Schizosaccharomyces fission, the Caenorhabditis anterior-posterior zygote polarisation and Arabidopsis prolate pollen polarisation. The apical-basal patterning in Saccharina zygotes on the other hand, may be seen as analogous to that of land plants. Overall, brown algae have the potential to bring exciting new information on how a single cell gives rise to an entire complex body plan.
Topics: Animals; Zygote; Arabidopsis; Phaeophyceae; Cell Polarity; Cell Division; Plants
PubMed: 35317961
DOI: 10.1016/j.semcdb.2022.03.008 -
Current Topics in Developmental Biology 2020Soon after fertilization the zebrafish embryo generates the pool of cells that will give rise to the germline and the three somatic germ layers of the embryo (ectoderm,... (Review)
Review
Soon after fertilization the zebrafish embryo generates the pool of cells that will give rise to the germline and the three somatic germ layers of the embryo (ectoderm, mesoderm and endoderm). As the basic body plan of the vertebrate embryo emerges, evolutionarily conserved developmental signaling pathways, including Bmp, Nodal, Wnt, and Fgf, direct the nearly totipotent cells of the early embryo to adopt gene expression profiles and patterns of cell behavior specific to their eventual fates. Several decades of molecular genetics research in zebrafish has yielded significant insight into the maternal and zygotic contributions and mechanisms that pattern this vertebrate embryo. This new understanding is the product of advances in genetic manipulations and imaging technologies that have allowed the field to probe the cellular, molecular and biophysical aspects underlying early patterning. The current state of the field indicates that patterning is governed by the integration of key signaling pathways and physical interactions between cells, rather than a patterning system in which distinct pathways are deployed to specify a particular cell fate. This chapter focuses on recent advances in our understanding of the genetic and molecular control of the events that impart cell identity and initiate the patterning of tissues that are prerequisites for or concurrent with movements of gastrulation.
Topics: Animals; Body Patterning; Embryo, Nonmammalian; Gastrula; Gastrulation; Gene Expression Regulation, Developmental; Signal Transduction; Zebrafish; Zebrafish Proteins; Zygote
PubMed: 31959294
DOI: 10.1016/bs.ctdb.2019.08.002 -
Seminars in Cell & Developmental Biology Dec 2018Zygotic genome activation (ZGA) denotes the initiation of gene expression after fertilization. It is part of the complex oocyte-to-embryo transition (OET) in which a... (Review)
Review
Zygotic genome activation (ZGA) denotes the initiation of gene expression after fertilization. It is part of the complex oocyte-to-embryo transition (OET) in which a highly specialized cell - the oocyte - is fertilized and transformed into a zygote that gives rise to an embryo that will develop into a newborn. From the perspective of gene expression, the OET reflects reprogramming of germ cell gene expression into the new developmental program of the zygote. This reprogramming occurs at transcriptional and post-transcriptional levels. This review will discuss selected aspects of mammalian ZGA, highlighting shared features and evolved differences observed in commonly investigated mammals and non-mammalian model animals.
Topics: Animals; Embryo, Mammalian; Fertilization; Gene Expression Regulation, Developmental; Humans; Mammals; Oocytes; Zygote
PubMed: 29233752
DOI: 10.1016/j.semcdb.2017.12.006 -
Current Biology : CB Jun 2024Rapid cleavage divisions and the transition from maternal to zygotic control of gene expression are the hallmarks of early embryonic development in most species. Early... (Review)
Review
Rapid cleavage divisions and the transition from maternal to zygotic control of gene expression are the hallmarks of early embryonic development in most species. Early development in insects, fish and amphibians is characterized by several short cell cycles with no gap phases, necessary for the rapid production of cells prior to patterning and morphogenesis. Maternal mRNAs and proteins loaded into the egg during oogenesis are essential to drive these rapid early divisions. Once the function of these maternal inputs is complete, the maternal-to-zygotic transition (MZT) marks the handover of developmental control to the gene products synthesized from the zygotic genome. The MZT requires three major events: the removal of a subset of maternal mRNAs, the initiation of zygotic transcription, and the remodeling of the cell cycle. In each species, the MZT occurs at a highly reproducible time during development due to a series of feedback mechanisms that tightly couple these three processes. Dissecting these feedback mechanisms and their spatiotemporal control will be essential to understanding the control of the MZT. In this primer, we outline the mechanisms that govern the major events of the MZT across species and highlight the role of feedback mechanisms that ensure the MZT is precisely timed and orchestrated.
Topics: Zygote; Animals; Gene Expression Regulation, Developmental; Embryonic Development; Female; RNA, Messenger, Stored
PubMed: 38834020
DOI: 10.1016/j.cub.2024.04.044 -
Zygote (Cambridge, England) Feb 2021In higher plants, fertilization induces many structural and physiological changes in the fertilized egg that reflect the transition from the haploid female gamete to the... (Review)
Review
In higher plants, fertilization induces many structural and physiological changes in the fertilized egg that reflect the transition from the haploid female gamete to the diploid zygote - the first cell of the sporophyte. After fusion of the egg nucleus with the sperm nucleus, many molecular changes occur in the zygote during the process of zygote activation during embryogenesis. The zygote originates from the egg, from which some pre-stored translation initiation factors transfer into the zygote and function during zygote activation. This indicates that the control of zygote activation is pre-set in the egg. After the egg and sperm nuclei fuse, gene expression is activated in the zygote, and paternal and maternal gene expression patterns are displayed. This highlights the diversity of zygotic genome activation in higher plants. In addition to new gene expression in the zygote, some genes show quantitative changes in expression. The asymmetrical division of the zygote produces an apical cell and a basal cell that have different destinies during plant reconstruction; these destinies are determined in the zygote. This review describes significant advances in research on the mechanisms controlling zygote activation in higher plants.
Topics: Diploidy; Haploidy; Oryza; Seeds; Zygote
PubMed: 33054867
DOI: 10.1017/S0967199420000568 -
Current Topics in Developmental Biology 2020Gastrulation is a critical early morphogenetic process of animal development, during which the three germ layers; mesoderm, endoderm and ectoderm, are rearranged by... (Review)
Review
Gastrulation is a critical early morphogenetic process of animal development, during which the three germ layers; mesoderm, endoderm and ectoderm, are rearranged by internalization movements. Concurrent epiboly movements spread and thin the germ layers while convergence and extension movements shape them into an anteroposteriorly elongated body with head, trunk, tail and organ rudiments. In zebrafish, gastrulation follows the proliferative and inductive events that establish the embryonic and extraembryonic tissues and the embryonic axis. Specification of these tissues and embryonic axes are controlled by the maternal gene products deposited in the egg. These early maternally controlled processes need to generate sufficient cell numbers and establish the embryonic polarity to ensure normal gastrulation. Subsequently, after activation of the zygotic genome, the zygotic gene products govern mesoderm and endoderm induction and germ layer patterning. Gastrulation is initiated during the maternal-to-zygotic transition, a process that entails both activation of the zygotic genome and downregulation of the maternal transcripts. Genomic studies indicate that gastrulation is largely controlled by the zygotic genome. Nonetheless, genetic studies that investigate the relative contributions of maternal and zygotic gene function by comparing zygotic, maternal and maternal zygotic mutant phenotypes, reveal significant contribution of maternal gene products, transcripts and/or proteins, that persist through gastrulation, to the control of gastrulation movements. Therefore, in zebrafish, the maternally expressed gene products not only set the stage for, but they also actively participate in gastrulation morphogenesis.
Topics: Animals; Blastoderm; Blastula; Embryo, Nonmammalian; Gastrulation; Gene Expression Regulation, Developmental; Maternal Inheritance; Morphogenesis; Zebrafish; Zygote
PubMed: 32591082
DOI: 10.1016/bs.ctdb.2020.05.001