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Nature Biotechnology Sep 2023Integrated in vitro models of human organogenesis are needed to elucidate the multi-systemic events underlying development and disease. Here we report the generation of...
Integrated in vitro models of human organogenesis are needed to elucidate the multi-systemic events underlying development and disease. Here we report the generation of human trunk-like structures that model the co-morphogenesis, patterning and differentiation of the human spine and spinal cord. We identified differentiation conditions for human pluripotent stem cells favoring the formation of an embryo-like extending antero-posterior (AP) axis. Single-cell and spatial transcriptomics show that somitic and spinal cord differentiation trajectories organize along this axis and can self-assemble into a neural tube surrounded by somites upon extracellular matrix addition. Morphogenesis is coupled with AP patterning mechanisms, which results, at later stages of organogenesis, in in vivo-like arrays of neural subtypes along a neural tube surrounded by spine and muscle progenitors contacted by neuronal projections. This integrated system of trunk development indicates that in vivo-like multi-tissue co-morphogenesis and topographic organization of terminal cell types can be achieved in human organoids, opening windows for the development of more complex models of organogenesis.
PubMed: 37709912
DOI: 10.1038/s41587-023-01956-9 -
Nature Reviews. Molecular Cell Biology Jul 2024Segmentation is a fundamental feature of the vertebrate body plan. This metameric organization is first implemented by somitogenesis in the early embryo, when paired... (Review)
Review
Segmentation is a fundamental feature of the vertebrate body plan. This metameric organization is first implemented by somitogenesis in the early embryo, when paired epithelial blocks called somites are rhythmically formed to flank the neural tube. Recent advances in in vitro models have offered new opportunities to elucidate the mechanisms that underlie somitogenesis. Notably, models derived from human pluripotent stem cells introduced an efficient proxy for studying this process during human development. In this Review, we summarize the current understanding of somitogenesis gained from both in vivo studies and in vitro studies. We deconstruct the spatiotemporal dynamics of somitogenesis into four distinct modules: dynamic events in the presomitic mesoderm, segmental determination, somite anteroposterior polarity patterning, and epithelial morphogenesis. We first focus on the segmentation clock, as well as signalling and metabolic gradients along the tissue, before discussing the clock and wavefront and other models that account for segmental determination. We then detail the molecular and cellular mechanisms of anteroposterior polarity patterning and somite epithelialization.
Topics: Somites; Animals; Humans; Body Patterning; Vertebrates; Gene Expression Regulation, Developmental; Embryonic Development; Mesoderm; Signal Transduction; Morphogenesis
PubMed: 38418851
DOI: 10.1038/s41580-024-00709-z -
Nature Communications Jan 2024Embryonic cells exhibit diverse metabolic states. Recent studies have demonstrated that metabolic reprogramming drives changes in cell identity by affecting gene...
Embryonic cells exhibit diverse metabolic states. Recent studies have demonstrated that metabolic reprogramming drives changes in cell identity by affecting gene expression. However, the connection between cellular metabolism and gene expression remains poorly understood. Here we report that glycolysis-regulated histone lactylation couples the metabolic state of embryonic cells with chromatin organization and gene regulatory network (GRN) activation. We found that lactylation marks genomic regions of glycolytic embryonic tissues, like the neural crest (NC) and pre-somitic mesoderm. Histone lactylation occurs in the loci of NC genes as these cells upregulate glycolysis. This process promotes the accessibility of active enhancers and the deployment of the NC GRN. Reducing the deposition of the mark by targeting LDHA/B leads to the downregulation of NC genes and the impairment of cell migration. The deposition of lactyl-CoA on histones at NC enhancers is supported by a mechanism that involves transcription factors SOX9 and YAP/TEAD. These findings define an epigenetic mechanism that integrates cellular metabolism with the GRNs that orchestrate embryonic development.
Topics: Histones; Gene Regulatory Networks; Transcription Factors; Embryonic Development; Mesoderm
PubMed: 38167340
DOI: 10.1038/s41467-023-44121-1 -
Development (Cambridge, England) May 2024Proper embryonic development depends on the timely progression of a genetic program. One of the key mechanisms for achieving precise control of developmental timing is... (Review)
Review
Proper embryonic development depends on the timely progression of a genetic program. One of the key mechanisms for achieving precise control of developmental timing is to use gene expression oscillations. In this Review, we examine how gene expression oscillations encode temporal information during vertebrate embryonic development by discussing the gene expression oscillations occurring during somitogenesis, neurogenesis, myogenesis and pancreas development. These oscillations play important but varied physiological functions in different contexts. Oscillations control the period of somite formation during somitogenesis, whereas they regulate the proliferation-to-differentiation switch of stem cells and progenitor cells during neurogenesis, myogenesis and pancreas development. We describe the similarities and differences of the expression pattern in space (i.e. whether oscillations are synchronous or asynchronous across neighboring cells) and in time (i.e. different time scales) of mammalian Hes/zebrafish Her genes and their targets in different tissues. We further summarize experimental evidence for the functional role of their oscillations. Finally, we discuss the outstanding questions for future research.
Topics: Animals; Embryonic Development; Gene Expression Regulation, Developmental; Humans; Somites; Muscle Development; Neurogenesis; Pancreas; Cell Differentiation
PubMed: 38727565
DOI: 10.1242/dev.202191 -
Nature Feb 2024The house mouse (Mus musculus) is an exceptional model system, combining genetic tractability with close evolutionary affinity to humans. Mouse gestation lasts only 3...
The house mouse (Mus musculus) is an exceptional model system, combining genetic tractability with close evolutionary affinity to humans. Mouse gestation lasts only 3 weeks, during which the genome orchestrates the astonishing transformation of a single-cell zygote into a free-living pup composed of more than 500 million cells. Here, to establish a global framework for exploring mammalian development, we applied optimized single-cell combinatorial indexing to profile the transcriptional states of 12.4 million nuclei from 83 embryos, precisely staged at 2- to 6-hour intervals spanning late gastrulation (embryonic day 8) to birth (postnatal day 0). From these data, we annotate hundreds of cell types and explore the ontogenesis of the posterior embryo during somitogenesis and of kidney, mesenchyme, retina and early neurons. We leverage the temporal resolution and sampling depth of these whole-embryo snapshots, together with published data from earlier timepoints, to construct a rooted tree of cell-type relationships that spans the entirety of prenatal development, from zygote to birth. Throughout this tree, we systematically nominate genes encoding transcription factors and other proteins as candidate drivers of the in vivo differentiation of hundreds of cell types. Remarkably, the most marked temporal shifts in cell states are observed within one hour of birth and presumably underlie the massive physiological adaptations that must accompany the successful transition of a mammalian fetus to life outside the womb.
Topics: Animals; Female; Mice; Pregnancy; Animals, Newborn; Cell Differentiation; Embryo, Mammalian; Embryonic Development; Gastrula; Gastrulation; Kidney; Mesoderm; Neurons; Retina; Single-Cell Analysis; Somites; Time Factors; Time-Lapse Imaging; Transcription Factors; Transcription, Genetic; Organ Specificity
PubMed: 38355799
DOI: 10.1038/s41586-024-07069-w -
Frontiers in Cell and Developmental... 2023Morphological phenotyping of the mouse embryo is described at neurulation stages, primarily as a guide to evaluating the outcome of whole embryo cultures between... (Review)
Review
Morphological phenotyping of the mouse embryo is described at neurulation stages, primarily as a guide to evaluating the outcome of whole embryo cultures between embryonic days 8.5 and 9.5. During this period, neural tube closure is initiated and progresses to completion in the cranial region. Spinal closure is still underway at the end of the culture period. The focus of this article is particularly on phenotyping that can be performed at the bench, using a stereomicroscope. This involves assessment of embryonic health, through observation and scoring of yolk sac blood circulation, measurement of developmental stage by somite counting, and determination of crown-rump length as a measure of growth. Axial rotation ("turning") can also be assessed using a simple scoring system. Neural tube closure assessment includes: 1) determining whether closure has been initiated at the Closure 1 site; 2) evaluating the complex steps of cranial neurulation including initiation at Closure sites 2 and 3, and completion of closure at the anterior and hindbrain neuropores; 3) assessment of spinal closure by measurement of posterior neuropore length. Interpretation of defects in neural tube closure requires an appreciation of, first, the stages that particular events are expected to be completed and, second, the correspondence between embryonic landmarks, for example, somite position, and the resulting adult axial levels. Detailed embryonic phenotyping, as described in this article, when combined with the versatile method of whole embryo culture, can form the basis for a wide range of experimental studies in early mouse neural development.
PubMed: 37601098
DOI: 10.3389/fcell.2023.1223849 -
Current Topics in Developmental Biology 2024The Segmentation Clock is a tissue-level patterning system that enables the segmentation of the vertebral column precursors into transient multicellular blocks called... (Review)
Review
The Segmentation Clock is a tissue-level patterning system that enables the segmentation of the vertebral column precursors into transient multicellular blocks called somites. This patterning system comprises a set of elements that are essential for correct segmentation. Under the so-called "Clock and Wavefront" model, the system consists of two elements, a genetic oscillator that manifests itself as traveling waves of gene expression, and a regressing wavefront that transforms the temporally periodic signal encoded in the oscillations into a permanent spatially periodic pattern of somite boundaries. Over the last twenty years, every new discovery about the Segmentation Clock has been tightly linked to the nomenclature of the "Clock and Wavefront" model. This constrained allocation of discoveries into these two elements has generated long-standing debates in the field as what defines molecularly the wavefront and how and where the interaction between the two elements establishes the future somite boundaries. In this review, we propose an expansion of the "Clock and Wavefront" model into three elements, "Clock", "Wavefront" and signaling gradients. We first provide a detailed description of the components and regulatory mechanisms of each element, and we then examine how the spatiotemporal integration of the three elements leads to the establishment of the presumptive somite boundaries. To be as exhaustive as possible, we focus on the Segmentation Clock in zebrafish. Furthermore, we show how this three-element expansion of the model provides a better understanding of the somite formation process and we emphasize where our current understanding of this patterning system remains obscure.
Topics: Animals; Body Patterning; Gene Expression Regulation, Developmental; Somites; Mesoderm; Zebrafish; Signal Transduction; Biological Clocks
PubMed: 38729682
DOI: 10.1016/bs.ctdb.2023.11.001 -
DNA and Cell Biology Oct 2023Fibroblast growth factor (FGF) signaling is conserved from cnidaria to mammals (Ornitz and Itoh, 2022) and it regulates several critical processes such as... (Review)
Review
Fibroblast growth factor (FGF) signaling is conserved from cnidaria to mammals (Ornitz and Itoh, 2022) and it regulates several critical processes such as differentiation, proliferation, apoptosis, cell migration, and embryonic development. One pivotal process FGF signaling controls is the division of vertebrate paraxial mesoderm into repeated segmented units called somites (i.e., somitogenesis). Somite segmentation occurs periodically and sequentially in a head-to-tail manner, and lays down the plan for compartmentalized development of the vertebrate body axis (Gomez , 2008). These somites later give rise to vertebrae, tendons, and skeletal muscle. Somite segments form sequentially from the anterior end of the presomitic mesoderm (PSM). The periodicity of somite segmentation is conferred by the segmentation clock, comprising oscillatory expression of Hairy and enhancer-of-split (Her/Hes) genes in the PSM. The positional information for somite boundaries is instructed by the double phosphorylated extracellular signal-regulated kinase (ppERK) gradient, which is the relevant readout of FGF signaling during somitogenesis (Sawada , 2001; Delfini , 2005; Simsek and Ozbudak, 2018; Simsek , 2023). In this review, we summarize the crosstalk between the segmentation clock and FGF/ppERK gradient and discuss how that leads to periodic somite boundary formation. We also draw attention to outstanding questions regarding the interconnected roles of the segmentation clock and ppERK gradient, and close with suggested future directions of study.
Topics: Animals; Fibroblast Growth Factors; Somites; Mesoderm; Signal Transduction; Embryonic Development; Gene Expression Regulation, Developmental; Mammals
PubMed: 37462914
DOI: 10.1089/dna.2023.0226 -
Cardiovascular Research Jul 2023Congenital heart disease (CHD) is the most common genetic birth defect, which has considerable morbidity and mortality. We focused on deciphering key regulators that...
AIMS
Congenital heart disease (CHD) is the most common genetic birth defect, which has considerable morbidity and mortality. We focused on deciphering key regulators that govern cardiac progenitors and cardiogenesis. FOXK1 is a forkhead/winged helix transcription factor known to regulate cell cycle kinetics and is restricted to mesodermal progenitors, somites, and heart. In the present study, we define an essential role for FOXK1 during cardiovascular development.
METHODS AND RESULTS
We used the mouse embryoid body system to differentiate control and Foxk1 KO embryonic stem cells into mesodermal, cardiac progenitor cells and mature cardiac cells. Using flow cytometry, immunohistochemistry, cardiac beating, transcriptional and chromatin immunoprecipitation quantitative polymerase chain reaction assays, bulk RNA sequencing (RNAseq) and assay for transposase-accessible chromatin using sequencing (ATACseq) analyses, FOXK1 was observed to be an important regulator of cardiogenesis. Flow cytometry analyses revealed perturbed cardiogenesis in Foxk1 KO embryoid bodies (EBs). Bulk RNAseq analysis at two developmental stages showed a significant reduction of the cardiac molecular program in Foxk1 KO EBs compared to the control EBs. ATACseq analysis during EB differentiation demonstrated that the chromatin landscape nearby known important regulators of cardiogenesis was significantly relaxed in control EBs compared to Foxk1 KO EBs. Furthermore, we demonstrated that in the absence of FOXK1, cardiac differentiation was markedly impaired by assaying for cardiac Troponin T expression and cardiac contractility. We demonstrate that FOXK1 is an important regulator of cardiogenesis by repressing the Wnt/β-catenin signalling pathway and thereby promoting differentiation.
CONCLUSION
These results identify FOXK1 as an essential transcriptional and epigenetic regulator of cardiovascular development. Mechanistically, FOXK1 represses Wnt signalling to promote the development of cardiac progenitor cells.
Topics: Animals; Mice; Cell Differentiation; Embryonic Stem Cells; Heart; Wnt Signaling Pathway
PubMed: 37036809
DOI: 10.1093/cvr/cvad054 -
Developmental Cell Oct 2023The mammalian body plan is shaped by rhythmic segmentation of mesoderm into somites, which are transient embryonic structures that form down each side of the neural...
The mammalian body plan is shaped by rhythmic segmentation of mesoderm into somites, which are transient embryonic structures that form down each side of the neural tube. We have analyzed the genome-wide transcriptional and chromatin dynamics occurring within nascent somites, from early inception of somitogenesis to the latest stages of body plan establishment. We created matched gene expression and open chromatin maps for the three leading pairs of somites at six time points during mouse embryonic development. We show that the rate of somite differentiation accelerates as development progresses. We identified a conserved maturation program followed by all somites, but somites from more developed embryos concomitantly switch on differentiation programs from derivative cell lineages soon after segmentation. Integrated analysis of the somitic transcriptional and chromatin activities identified opposing regulatory modules controlling the onset of differentiation. Our results provide a powerful, high-resolution view of the molecular genetics underlying somitic development in mammals.
Topics: Pregnancy; Female; Mice; Animals; Somites; Embryonic Development; Mesoderm; Cell Differentiation; Chromatin; Mammals
PubMed: 37499658
DOI: 10.1016/j.devcel.2023.07.003