Establishment of developmental gene silencing by ordered polycomb complex recruitment in early zebrafish embryos

Vertebrate embryos achieve developmental competency during zygotic genome activation (ZGA) by establishing chromatin states that silence yet poise developmental genes for subsequent lineage-specific activation. Here, we reveal the order of chromatin states in establishing developmental gene poising in preZGA zebrafish embryos. Poising is established at promoters and enhancers that initially contain open/permissive chromatin with 'Placeholder' nucleosomes (bearing H2A.Z, H3K4me1, and H3K27ac), and DNA hypomethylation. Silencing is initiated by the recruitment of Polycomb Repressive Complex 1 (PRC1), and H2Aub1 deposition by catalytic Rnf2 during preZGA and ZGA stages. During postZGA, H2Aub1 enables Aebp2-containing PRC2 recruitment and H3K27me3 deposition. Notably, preventing H2Aub1 (via Rnf2 inhibition) eliminates recruitment of Aebp2-PRC2 and H3K27me3, and elicits transcriptional upregulation of certain developmental genes during ZGA. However, upregulation is independent of H3K27me3 - establishing H2Aub1 as the critical silencing modification at ZGA. Taken together, we reveal the logic and mechanism for establishing poised/silent developmental genes in early vertebrate embryos.

Signalling dynamics in embryonic development

In multicellular organisms, cellular behaviour is tightly regulated to allow proper embryonic development and maintenance of adult tissue. A critical component in this control is the communication between cells via signalling pathways, as errors in intercellular communication can induce developmental defects or diseases such as cancer. It has become clear over the last years that signalling is not static but varies in activity over time. Feedback mechanisms present in every signalling pathway lead to diverse dynamic phenotypes, such as transient activation, signal ramping or oscillations, occurring in a cell type- and stage-dependent manner. In cells, such dynamics can exert various functions that allow organisms to develop in a robust and reproducible way. Here, we focus on Erk, Wnt and Notch signalling pathways, which are dynamic in several tissue types and organisms, including the periodic segmentation of vertebrate embryos, and are often dysregulated in cancer. We will discuss how biochemical processes influence their dynamics and how these impact on cellular behaviour within multicellular systems.

Maternally-inherited anti-sense piRNAs antagonize transposon expression in teleost embryos

Transposable elements threaten genome stability, and the Piwi-piRNA system has evolved to silence transposons in the germline. However, it remains largely unknown what mechanisms are utilized in early vertebrate embryos prior to germline establishment and ping-pong piRNA production. To address this, we first characterized small RNAs in early zebrafish embryos and detected abundant maternally-deposited, Ziwi-associated, antisense piRNAs that map largely to evolutionarily young long terminal repeat (LTR) retrotransposons. Notably, the focal establishment of the repressive modification H3K9me2/3 coincides with these young LTR elements, is deposited independent of transcription, and is required for LTR silencing. We find piRNAs highly enriched and maintained in primordial germ cells (PGCs), which display lower LTR expression than somatic cells. To examine the consequences of piRNA loss, we used reciprocal zebrafish-medaka hybrids, which display selective activation of LTRs that lack maternally-contributed targeting piRNAs. Thus, the Piwi-piRNA system actively antagonizes transposons in the soma and PGCs during early vertebrate embryogenesis.

Developing Inside a Layer of Germs—A Potential Role for Multiciliated Surface Cells in Vertebrate Embryos

This paper reviews current research on the microbial life that surrounds vertebrate embryos. Several clades are believed to develop inside sterile—or near-sterile—embryonic microhabitats, while others thrive within a veritable zoo of microbial life. The occurrence of embryo-associated microbes in some groups, but not others, is an under-appreciated transition (possibly transitions) in vertebrate evolution. A lack of comparable studies makes it currently impossible to correlate embryo-associated microbiomes with other aspects of vertebrate evolution. However, there are embryonic features that should instruct a more targeted survey. This paper concludes with a hypothesis for the role of multiciliated surface cells in amphibian and some fish embryos, which may contribute to managing embryo-associated microbial consortia. These cells are known to exist in some species that harbor in ovo microbes or have relatively porous egg capsules, although most have not been assayed for embryo-associated microbiota. Whether the currents generated within these extraembryonic microhabitats contribute to culturing consistent microbial communities remains to be seen.

Segregation of neural crest specific lineage trajectories from a heterogeneous neural plate border territory only emerges at neurulation

The epiblast of vertebrate embryos is comprised of neural and non-neural ectoderm, with the border territory at their intersection harbouring neural crest and cranial placode progenitors. Here we profile avian epiblast cells as a function of time using single-cell RNA-seq to define transcriptional changes in the emerging ‘neural plate border’. The results reveal gradual establishment of heterogeneous neural plate border signatures, including novel genes that we validate by fluorescent in situ hybridisation. Developmental trajectory analysis shows that segregation of neural plate border lineages only commences at early neurulation, rather than at gastrulation as previously predicted. We find that cells expressing the prospective neural crest marker Pax7 contribute to multiple lineages, and a subset of premigratory neural crest cells shares a transcriptional signature with their border precursors. Together, our results suggest that cells at the neural plate border remain heterogeneous until early neurulation, at which time progenitors become progressively allocated toward defined lineages.

An interview with James Wells

ABSTRACT James (Jim) Wells is a Professor in the Division of Developmental Biology at Cincinnati Children's Hospital Medical Center, and Chief Scientific Officer of the Center for Stem Cell & Organoid Medicine (CuSTOM) at Cincinnati Children's. Using both vertebrate embryos and human organoids as model systems, Jim's research focuses on the mechanisms by which gastrointestinal and endocrine organs form. Earlier this year, Jim joined the Development team as an Academic Editor. We caught up with Jim to find out more about his research career, the stem cell and organoid fields, and why he decided to get involved with the journal.

Sustained experimental activation of FGF8/ERK in the developing chicken spinal cord reproducibly models early events in ERK-mediated tumorigenesis

Most human cancers demonstrate activated MAPK/ERK pathway signaling as a key tumor initiation step, but the immediate steps of further oncogenic progression are poorly understood due to a lack of appropriate models. Spinal cord differentiation follows caudal elongation in vertebrate embryos; both processes are regulated by a FGF8 gradient highest in neuromesodermal progenitors (NMP), where kinase effectors ERK1/2 maintain an undifferentiated state. FGF8/ERK signal attenuation is necessary for NMPs to progress to differentiation. We show that sustained ERK1/2 activity, using a constitutively active form of the kinase MEK1 (MEK1ca) in the chicken embryo, reproducibly provokes neopasia in the developing spinal cord. Transcriptomic data show that neoplasia not only relies on the maintenance of NMP gene expression, and the inhibition of genes expressed in the differentiating spinal cord, but also on a profound change in the transcriptional signature of the spinal cord cells leading to a complete loss of cell-type identity. MEK1ca expression in the developing spinal cord of the chicken embryo is therefore a tractable in vivo model to identify the critical factors fostering malignancy in ERK-induced tumorigenesis.

Maternal and zygotic factors sequentially shape the tissue regionalization of chromatin landscapes in early vertebrate embryos

One of the first steps in cellular differentiation of vertebrate embryos is the formation of the three germ layers. Maternal pioneer transcription factors (TFs) bind to the regulatory regions of the embryonic genome prior to zygotic genome activation and initiate germ layer specification. While the involvement of maternal TFs in establishing epigenetic marks in whole embryos was addressed previously, how early pluripotent cells acquire spatially restricted epigenetic identity in embryos remain unknown. Here, we report that the H3K4me1 enhancer mark in each germ layer becomes distinct in germ layer specific regulatory regions, forming super-enhancers (SEs), by early gastrula stage. Distinct SEs are established in these germ layers near robustly regulated germ layer identity genes, suggesting that SEs are important for the canalization of development. Establishment of these enhancers requires a sequential function of maternal and zygotic TFs. By knocking down the expression of a critical set of maternal endodermal TFs, an overwhelming majority of the endodermal H3K4me1 marks are lost. Interestingly, this disappearance of endodermal marking coincides with the appearance of ectodermal and mesodermal H3K4me1 marks in the endoderm, suggesting a transformation in the chromatin state of these nuclei towards a more ecto-mesodermal state. De novo motif analysis to identify TFs responsible for the transformation recovers a profile for endodermal maternal TFs as well as their downstream target TFs. We demonstrate the importance of coordinated activities of maternal and zygotic TFs in defining a spatially resolved dynamic process of chromatin state establishment.

Establishment of Developmental Gene Silencing by Ordered Polycomb Complex Recruitment in Early Zebrafish Embryos

Vertebrate embryos achieve developmental competency during zygotic genome activation (ZGA) by establishing chromatin states that silence yet poise developmental genes for subsequent lineage-specific activation. Here, we reveal how developmental gene poising is established de novo in preZGA zebrafish embryos. Poising is established at promoters and enhancers that initially contain open/permissive chromatin with 'Placeholder' nucleosomes (bearing H2A.Z, H3K4me1, and H3K27ac), and DNA hypomethylation. Silencing is initiated by the recruitment of Polycomb Repressive Complex 1 (PRC1), and H2Aub1 deposition by catalytic Rnf2 during preZGA and ZGA stages. During postZGA, H2Aub1 enables Aebp2-containing PRC2 recruitment and H3K27me3 deposition. Notably, preventing H2Aub1 (via Rnf2 inhibition) eliminates recruitment of Aebp2-PRC2 and H3K27me3, and elicits transcriptional upregulation of certain developmental genes during ZGA. However, upregulation is independent of H3K27me3 - establishing H2Aub1 as the critical silencing modification at ZGA. Taken together, we reveal the logic and mechanism for establishing poised/silent developmental genes in early vertebrate embryos.