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Just as in clay moulding or glass blowing, physically sculpting biological structures requires the constituent material to locally flow like a fluid while maintaining overall mechanical integrity like a solid. Disordered soft materials, such as foams, emulsions and colloidal suspensions, switch from fluid-like to solid-like behaviours at a jamming transition 1 – 4 . Similarly, cell collectives have been shown to display glassy dynamics in 2D and 3D 5 , 6 and jamming in cultured epithelial monolayers 7 , 8 , behaviours recently predicted theoretically 9 – 11 and proposed to influence asthma pathobiology 8 and tumour progression 12 . However, little is known about whether these seemingly universal behaviours occur in vivo 13 and, specifically, whether they play any functional part during embryonic morphogenesis. Here, by combining direct in vivo measurements of tissue mechanics with analysis of cellular dynamics, we show that during vertebrate body axis elongation, posterior tissues undergo a jamming transition from a fluid-like behaviour at the extending end, the mesodermal progenitor zone, to a solid-like behaviour in the presomitic mesoderm. We uncover an anteroposterior, N-cadherin-dependent gradient in yield stress that provides increasing mechanical integrity to the presomitic mesoderm, consistent with the tissue transiting from a wetter to a dryer foam-like architecture. Our results show that cell-scale stresses fluctuate rapidly (within about 1 min), enabling cell rearrangements and effectively ‘melting’ the tissue at the growing end. Persistent (more than 0.5 h) stresses at supracellular scales, rather than cell-scale stresses, guide morphogenetic flows in fluid-like tissue regions. Unidirectional axis extension is sustained by the reported rigidification of the presomitic mesoderm, which mechanically supports posterior, fluid-like tissues during remodelling before their maturation. The spatiotemporal control of fluid-like and solid-like tissue states may represent a generic physical mechanism of embryonic morphogenesis. Cell collectives in embryonic tissues undergo a fluid-to-solid jamming transition, similar to those that occur in soft materials such as foams, emulsions and colloidal suspensions, to physically sculpt the vertebrate body axis.

N 6 -methyladenosine modification of mRNA determines cell fate decisions during the specification of the first haematopoietic stem cells in zebrafish. Stem cell specification by m 6 A Adding modified N 6 -methyladenosine (m 6 A) on messenger RNA (mRNA) has recently been reported to modulate mRNA translation and degradation. However, the effects of the modification on vertebrate development are so far unclear. Feng Liu and colleagues show that addition of m 6 A determines cell fate during the specification of the earliest haematopoietic stem cell in zebrafish. Blocking addition of this modification on genes that regulate Notch signalling maintains the endothelial fate of progenitors that should be specified as the earliest haematopoietic stem cell progenitors. N 6 -methyladenosine (m 6 A) has been identified as the most abundant modification on eukaryote messenger RNA (mRNA) 1 . Although the rapid development of high-throughput sequencing technologies has enabled insight into the biological functions of m 6 A modification 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , the function of m 6 A during vertebrate embryogenesis remains poorly understood. Here we show that m 6 A determines cell fate during the endothelial-to-haematopoietic transition (EHT) to specify the earliest haematopoietic stem/progenitor cells (HSPCs) during zebrafish embryogenesis. m 6 A-specific methylated RNA immunoprecipitation combined with high-throughput sequencing (MeRIP–seq) and m 6 A individual-nucleotide-resolution cross-linking and immunoprecipitation with sequencing (miCLIP–seq) analyses reveal conserved features on zebrafish m 6 A methylome and preferential distribution of m 6 A peaks near the stop codon with a consensus RRACH motif. In mettl3 -deficient embryos, levels of m 6 A are significantly decreased and emergence of HSPCs is blocked. Mechanistically, we identify that the delayed YTHDF2-mediated mRNA decay of the arterial endothelial genes notch1a and rhoca contributes to this deleterious effect. The continuous activation of Notch signalling in arterial endothelial cells of mettl3 -deficient embryos blocks EHT, thereby repressing the generation of the earliest HSPCs. Furthermore, knockdown of Mettl3 in mice confers a similar phenotype. Collectively, our findings demonstrate the critical function of m 6 A modification in the fate determination of HSPCs during vertebrate embryogenesis.

Cells respond to physical stimuli, such as stiffness 1 , fluid shear stress 2 and hydraulic pressure 3 , 4 . Extracellular fluid viscosity is a key physical cue that varies under physiological and pathological conditions, such as cancer 5 . However, its influence on cancer biology and the mechanism by which cells sense and respond to changes in viscosity are unknown. Here we demonstrate that elevated viscosity counterintuitively increases the motility of various cell types on two-dimensional surfaces and in confinement, and increases cell dissemination from three-dimensional tumour spheroids. Increased mechanical loading imposed by elevated viscosity induces an actin-related protein 2/3 (ARP2/3)-complex-dependent dense actin network, which enhances Na + /H + exchanger 1 (NHE1) polarization through its actin-binding partner ezrin. NHE1 promotes cell swelling and increased membrane tension, which, in turn, activates transient receptor potential cation vanilloid 4 (TRPV4) and mediates calcium influx, leading to increased RHOA-dependent cell contractility. The coordinated action of actin remodelling/dynamics, NHE1-mediated swelling and RHOA-based contractility facilitates enhanced motility at elevated viscosities. Breast cancer cells pre-exposed to elevated viscosity acquire TRPV4-dependent mechanical memory through transcriptional control of the Hippo pathway, leading to increased migration in zebrafish, extravasation in chick embryos and lung colonization in mice. Cumulatively, extracellular viscosity is a physical cue that regulates both short- and long-term cellular processes with pathophysiological relevance to cancer biology. Elevated viscosity counterintuitively increases the motility of various cell types in vitro and imprints mechanical memory to tumour cells, which enables them to disseminate more efficiently in vivo.

Gomez Perdiguero and Geissmann discuss the origin of tissue macrophages as a layered system composed of resident macrophages originating mostly from yolk-sac progenitor cells and transitory myeloid cells that originate and renew from bone marrow hematopoietic stem cells. The molecular and cellular mechanisms that underlie the many roles of macrophages in health and disease states in vivo remain poorly understood. The purpose of this Review is to present and discuss current knowledge on the developmental biology of macrophages, as it underlies the concept of a layered myeloid system composed of 'resident' macrophages that originate mainly from progenitor cells generated in the yolk sac and of 'passenger' or 'transitory' myeloid cells that originate and renew from bone marrow hematopoietic stem cells, and to provide a framework for investigating the functions of macrophages in vivo .

Lithium-ion batteries (LiBs) are used globally as a key component of clean and sustainable energy infrastructure, and emerging LiB technologies have incorporated a class of per- and polyfluoroalkyl substances (PFAS) known as bis-perfluoroalkyl sulfonimides (bis-FASIs). PFAS are recognized internationally as recalcitrant contaminants, a subset of which are known to be mobile and toxic, but little is known about environmental impacts of bis-FASIs released during LiB manufacture, use, and disposal. Here we demonstrate that environmental concentrations proximal to manufacturers, ecotoxicity, and treatability of bis-FASIs are comparable to PFAS such as perfluorooctanoic acid that are now prohibited and highly regulated worldwide, and we confirm the clean energy sector as an unrecognized and potentially growing source of international PFAS release. Results underscore that environmental impacts of clean energy infrastructure merit scrutiny to ensure that reduced CO 2 emissions are not achieved at the expense of increasing global releases of persistent organic pollutants. A new class of PFAS (bis-perfluoroalkyl sulfonamides) used in lithium-ion batteries have been released to the environment internationally. This places lithium-ion batteries at the nexus of CO2 reduction and release of recalcitrant aquatic contaminants.

CRISPR-Cas9 genome editing has potential to cure diseases without current treatments, but therapies must be safe. Here we show that CRISPR-Cas9 editing can introduce unintended mutations in vivo, which are passed on to the next generation. By editing fertilized zebrafish eggs using four guide RNAs selected for off-target activity in vitro, followed by long-read sequencing of DNA from >1100 larvae, juvenile and adult fish across two generations, we find that structural variants (SVs), i.e., insertions and deletions ≥50 bp, represent 6% of editing outcomes in founder larvae. These SVs occur both at on-target and off-target sites. Our results also illustrate that adult founder zebrafish are mosaic in their germ cells, and that 26% of their offspring carries an off-target mutation and 9% an SV. Hence, pre-testing for off-target activity and SVs using patient material is advisable in clinical applications, to reduce the risk of unanticipated effects with potentially large implications. CRISPR-Cas9 can introduce unintended off-target effects. Here authors show that unintended mutations produced by in vivo of zebrafish can be inherited by their off-spring.

During development, oligodendrocytes contact and wrap neuronal axons with myelin. Similarly to neurons and synapses, excess myelin sheaths are produced and selectively eliminated, but how elimination occurs is unknown. Microglia, the resident immune cells of the central nervous system, engulf surplus neurons and synapses. To determine whether microglia also prune myelin sheaths, we used zebrafish to visualize and manipulate interactions between microglia, oligodendrocytes, and neurons during development. We found that microglia closely associate with oligodendrocytes and specifically phagocytose myelin sheaths. By using a combination of optical, genetic, chemogenetic, and behavioral approaches, we reveal that neuronal activity bidirectionally balances microglial association with neuronal cell bodies and myelin phagocytosis in the optic tectum. Furthermore, multiple strategies to deplete microglia resulted in oligodendrocytes maintaining excessive and ectopic myelin. Our work reveals a neuronal activity-regulated role for microglia in modifying developmental myelin targeting by oligodendrocytes.Microglia refine the developing CNS by engulfing excess neurons and synapses. Hughes and Appel here show that microglia also prune myelin sheaths in a neuronal activity-regulated manner to sculpt developmental myelination.

During tumor progression, cancer cells come into contact with various non-tumor cell types, but it is unclear how tumors adapt to these new environments. Here, we integrate spatially resolved transcriptomics, single-cell RNA-seq, and single-nucleus RNA-seq to characterize tumor-microenvironment interactions at the tumor boundary. Using a zebrafish model of melanoma, we identify a distinct “interface” cell state where the tumor contacts neighboring tissues. This interface is composed of specialized tumor and microenvironment cells that upregulate a common set of cilia genes, and cilia proteins are enriched only where the tumor contacts the microenvironment. Cilia gene expression is regulated by ETS-family transcription factors, which normally act to suppress cilia genes outside of the interface. A cilia-enriched interface is conserved in human patient samples, suggesting it is a conserved feature of human melanoma. Our results demonstrate the power of spatially resolved transcriptomics in uncovering mechanisms that allow tumors to adapt to new environments. During tumor progression, cancer cells contact different neighboring cell types, but it is unclear how these interactions affect cancer cell behavior. Here, the authors use spatially resolved transcriptomics and single-cell RNA-seq to study the role of cilia at the tumormicroenvironment interface.

scGESTALT enables large-scale characterization of cell types and lineage relationships during vertebrate brain development. The lineage relationships among the hundreds of cell types generated during development are difficult to reconstruct. A recent method, GESTALT, used CRISPR–Cas9 barcode editing for large-scale lineage tracing, but was restricted to early development and did not identify cell types. Here we present scGESTALT, which combines the lineage recording capabilities of GESTALT with cell-type identification by single-cell RNA sequencing. The method relies on an inducible system that enables barcodes to be edited at multiple time points, capturing lineage information from later stages of development. Sequencing of ∼60,000 transcriptomes from the juvenile zebrafish brain identified >100 cell types and marker genes. Using these data, we generate lineage trees with hundreds of branches that help uncover restrictions at the level of cell types, brain regions, and gene expression cascades during differentiation. scGESTALT can be applied to other multicellular organisms to simultaneously characterize molecular identities and lineage histories of thousands of cells during development and disease.