Here, we report a detailed fate map of the zebrafish pancreas at the early gastrula stage of development 6 hours postfertilization. Fate maps were developed as a way of tracing a particular region as it develops from an early embryo into a differentiated body plan. Lineage tracing and fate map studies in zebrafish, chick. Background pancreas development in zebrafish shares many features with mammals, including the participation of epithelial progenitor cells expressing pancreas transcription factor 1a ptf1a. Three research groups have used single cell rna sequencing to analyze the transcriptional changes accompanying development of vertebrate embryos see the perspective by harland. We also discuss using zebrafish for live cell imaging experiments. However, to date it has remained unclear whether, as in mammals, ptf1aexpressing zebrafish pancreatic progenitors are able to contribute to multiple exocrine and endocrine lineages.
Fatemapping experiments of the zebrafish blastula demonstrate that blood and endothelial cells as well as posterior somites derive from the ventral mesoderm. Fate map of the deep cell layer del at gastrula onset, at the. Further, fate mapping studies and analysis of cells in vitro should confirm these earlier predictions. Cell proliferation in the developing lateral line system. Incredibly detailed embryo maps chart each cells developmental fate. Photoactivated gene expression for cell fate mapping and cell. Thus, a fate mapping analysis with high temporalspatial resolution is required to clarify the precise source of microglia.
Single cell fate mapping in zebrafish protocol jove. Conklin, a biologist and zoologist, conducted the first cell lineage experiments, which involved following the progenitor cells of the embryo of the tunicate styela partita. Live fatemapping of jointassociated fibroblasts visualizes expansion of cell contributions during zebrafish fin regeneration. Michael parsons department of developmental and cell biology. The number of hsc clones born has been estimated through transplantation, but experimental approaches to assess the absolute number of forming hscs in a native setting have remained challenging. Zebrafish has natural advantages on hematopoietic fate mapping owing to the. Klein also teamed up with megason and others to map gene expression in 92,000 cells collected from zebrafish danio rerio embryos over the. We show that, at this stage, both pancreas and liver progenitors are symmetrically localized in two broad domains relative to the dorsal organizer. To this end, we have developed a technique that allows for spatial an. For instance, determining how hematopoietic, lymphatic, and blood vessel lineages arise in developing embryos requires fate mapping and lineage tracing of undifferentiated precursor cells. Labeling single cell for invivo study of cell fate. Here, in this study we show that mirna1423p participates in the haematopoiesis regulation and vascular development of the early stages of erythropoiesis during zebra fish early developmental stages. We develop an infrared laserevoked gene operator heatshock microscope system to achieve single cell labeling in zebrafish.
It requires labeling individual cells and tracing the development of their progeny. Using lightinduced temporalspatial resolution fate mapping, xu and zhu et al. Our findings indicate that natural epicardial fates are limited to nonmyocardial cell types in zebrafish. Mar 18, 2005 fate map studies indicate that the cells that will generate the different placodes are already roughly aligned at 6haf kozlowski et al. Eos 1, 2, pamcherry 3, kaede 47, pkindling 8, and kikgr 9, 10 have received wide interest as cell. Recent advances have addressed the relationship between early cleavage planes and the future dorsal axis, the pattern of cell mixing during blastula and gastrula. Genetic inducible fate mapping in larval zebrafish reveals. In vivo singlecell lineage tracing in zebrafish using highresolution. We have identified in the zebrafish brain two distinct microglial populations.
Cell fates are pictorially depicted with migration denoted by wavy lines. All clones except one also included endothelial cells. Singlecell mapping of gene expression landscapes and lineage. Current fate mapping and imaging platforms are limited in their ability to capture dynamic behaviors of epithelial cells. Mechanisms for shaping and folding sheets of cells during development are poorly understood. An example is the complex reorganisation of the forebrain neural plate during neurulation, which must fold a sheet into a tube while evaginating two eyes from a single contiguous domain within the neural plate. In the spinal cord, shh is directed by both the notochord and floor plate regions which ultimately drives the. With a fluorescent thermometry technique, we measure the. Single cell fate mapping in zebrafish europe pmc article. This video will provide an overview of fate mapping, explain a protocol used to generate a fate map in zebrafish, and show some ways in which this technique is currently being applied in labs. Aug 15, 2017 live fate mapping of jointassociated fibroblasts visualizes expansion of cell contributions during zebrafish fin regeneration. Their fate map of the 32 cell stage embryo divided the cells into four tiers each containing eight cells, labeled ad a and b corresponding to the animal pole, c to the marginal zone, and d to the vegetal pole. High resolution fate map of the zebrafish diencephalon.
To test models of cardiac chamber specification, we generated a highresolution fate map of cardiac chamber progenitors in the zebrafish embryo at 40% epiboly, a stage prior to the initiation of gastrulation. Temporalspatial resolution fate mapping reveals distinct origins for embryonic and adult microglia in zebrafish dev cell. We have characterized the differentiation of the upper rhombic lip derived granule cell population in stable gfptransgenic zebrafish in the context of zebrafish cerebellar morphogenesis. Microglia are tissueresident macrophages that reside in the brain, spinal cord, and retina. Organization of cardiac chamber progenitors in the. Origin and organization of the zebrafish fate map development. Sep 15, 2014 a research article fate mapping of ptf1aexpressing cells during pancreatic organogenesis and regeneration in zebra. The first fate maps date back to the 1880s and in 1905 the first comprehensive collection of ascidian sea squirt fate maps was published by edwin conklin. Uncaging of caged fluoresceindextran in single cells is achieved using twophoton absorption from a mai tai femtosecond laser coupled to the zeiss lsm 510 meta confocal microscope. The information vogt gathered from his tracing experiments of distinct cells and populations of cells in xenopus was then pooled to construct a fate map. Clonal fate mapping quantifies the number of haematopoietic stem cells that arise during development nat cell biol. The horizontal line through each symbol shows the anteroposterior spread of each clone. The development and growth of vertebrate axial muscle have been studied for decades at both the descriptive and molecular level. Labeling single cell for invivo study of cell fate mapping.
Determination of fate maps and cell lineage tracing have previously been carried out in the zebrafish embryo by following the progeny of individual cells injected with fluorescent dyes. Fatemapping experiments in the chick, xenopus, and zebra fish show that presumptive placodal cells arise from a horseshoeshaped domain around the anterior neural plate. We, for the first time, track these cell rearrangements to show that forebrain morphogenesis. Pdf specification of cell fates at the dorsal margin of. Cell fate mapping and lineage tracing are significant ways to understanding the developmental origins of biological tissues.
A fate map emerges as the del cell lineages become restricted in the late blastula. Key developmental regulators suggest multiple origins of. Our results confirm the presence of multipotent ptf1aexpressing progenitor cells in developing zebrafish pancreas, with reduced ptf1a dosage promoting greater contributions towards nonacinar lineages. The ability to differentially label single cells has important implications in developmental biology. Singlecell mapping of gene expression landscapes and.
Fate mapping studies in teleost fish, mouse and chick have shown that a significant anteriorposterior rearrangement of tissue takes place to relocate the hypothalamic anlagen ventral. What makes the zebrafish particularly compelling for research into. For example, the development of the complete cell lineage of c. Here, we summarise recent findings in zebrafish embryonic myogenesis with a focus. Select 25 cell interactions and morphogenetic motions pattern the zebrafish nervous system. The first transplantable leukemia modeled in zebrafish was t cell acute. Fate maps for frogs and zebrafish are also quite detailed, but these organisms are too large to keep track of every single cell. Commitment of cell fate in the early zebrafish embryo. Work with lineage tracing and fate mapping has shown that the marginal cells of the blastoderm. Furthermore, fatemapping experiments show a degree of intermingling between cells for the different placodes, suggesting that the ppr is competent to generate different placodal derivatives.
Genetic inducible fate mapping in larval zebrafish reveals origins of adult insulinproducing. Skn1 is repressed in p2 cells and is unable to specify a ems fate in these cells. Nov 26, 1998 a previous fate map of zebrafish cranial neural crest showed that lineagerestricted clones of pigment cells arise from medial cells near the neural keel, and that clones of neurons arise from. A method is described to photoactivate single cells containing a caged fluorescent protein using twophoton absorption from a. Skn1 is a cytoplasmic determinant that is localized in the p1 cell lineage and determines ems cell fate. A fate mapping study has confirmed that the neuroectoderm and embryonic ectoderm arise within the epiblast, and histological studies suggest that the epiblast gives rise to all cell lineages of the embryo and extraembryonic mesoderm. We discuss the classical fate mapping of the endoderm and the more recent progress in characterizing.
Clonal fate mapping quantifies the number of haematopoietic. Hematopoietic generation, lineage tracing, and differentiation. Chicken embryos are more difficult to work with because the eggs. Pdf specification of cell fates at the dorsal margin of the. We define a domain in the early gastrula that is located just. Fate mapping is a method for tracing cell lineages and a fundamental tool of developmental biology and embryology. October 5th, 2011 vikram kohli 1, kira rehn 1, saulius sumanas 1,2. Another objective is to perturb the behavior of selected cells and follow the ensuing effects. Timelapse analysis followed by individual granule cell tracing demonstrates that the zebrafish upper rhombic lip is spatially patterned along its. Uncaging of caged fluoresceindextran in single cells is achieved using twophoton absorption from a mai tai femtosecond laser coupled to the.
Two phenotypically and functionally distinct microglial populations in. The fate of each cell or group of cells is mapped onto the embryo, showing which parts of the embryo will develop into which tissue. Analysis of cell movements in zebrafish embryos springerlink. Cell proliferation in the developing lateral line system of. Upon tamoxifen induction, the hematopoietic cells in these compartments might be labeled undesirably, potentially confounding the isolation of bona fide sources for microglia.
We now report on using crebased fate mapping to indelibly label pancreatic notchresponsive cells pncs at larval stages and follow their fate in the adult pancreas. This paper describes a versatile method for single cell fate mapping based on the photoactivation of caged fluoresceindextran. Since then, advances in transgenesis, microscopy and computing has allowed the analysis of cell fates and movements in real time, in live specimens. May 07, 2009 microinjection in zebrafish has been widely used for generating transgenic animals, and mapping cell fate in early blastula embryos 5,6,7,8,9. Photoactivated gene expression for cell fate mapping and. The cellular and molecular mechanisms responsible for the separation of ventricular and atrial lineages are not well understood. Fate mapping and cell lineage are similar but distinct topics, although there is often overlap. Sep 28, 2015 in this study, we used a high temporalspatial resolution fate mapping approach to investigate the origin of microglia in zebrafish.
We show an example of such a fate map in the figure above, which has been computed using the data of panc19. Pie1 is localized in the p2 cell lineage and is a general repressor of transcription. Fate mapping is a method used in developmental biology to study the embryonic origin of various adult tissues and structures. A longstanding goal of developmental biologists is to create developmental fate maps by tracking individual cells through development. Headwaters of the zebrafish washington university genetics. Furthermore, fatemapping experiments show a degree of intermingling between cells for the different placodes, suggesting that the ppr is competent to generate. A dynamic fate map of the forebrain shows how vertebrate eyes. How fate mapping is used to track cell development video. In contrast to mammals, stem cell niches are abundant in zebrafish and also found. Definitive hematopoietic stemprogenitor cells manifest. Origin of the zebrafish endocrine and exocrine pancreas. Using fate mapping techniques, we have analyzed development of cells of the dorsal marginal region in wildtype and mutant zebrafish. Regional cell movement and tissue patterning in the zebrafish.
Cellrank probabilistic fate mapping using rna velocity. Haematopoietic stem cells hscs arise in the developing aorta during embryogenesis. Mar 18, 2008 we completed a single cell fate map of the olfactory placode domains at 45 somitestage zebrafish embryo 1112 hours postfertilization at 28. Cell fate mapping and analysis of zebrafish runx1w84x and vlad tepes vltm651 mutants revealed that hspcs in the pbi gave rise to both erythroid and myeloid lineages. Contribution of early cells to the fate map of the zebrafish. Fate mapping is also used to study cell migration and differentiation during development, as well as regeneration and repair during adulthood. Jul, 2011 cell behavior can be analyzed in detail by cell transplantation and by fate mapping. The fate map simulation of gastrula stage zebrafish embryo was generated using the pioneering work of charles kimmel group kimmel et al, 1990. Single cells in embryos injected with caged fluoresceindextran are photoactivated with nearinfrared. The ability to differentially label single cells has important implications in.
It is similar in organization to that of an amphibian embryo. Tornini va 1 2, thompson jd 1 2, allen rl 1, poss kd 3 2. Photoactivatable caged fluorescein as a cell tracer for. However, we surprisingly found that hspcs in the vda were not quiescent but were uniquely adapted to generate myeloid but not erythroid lineage cells. Wang y, rovira m, yusuff s, huang w and parsons mj. Fate mapping of ptf1aexpressing cells during pancreatic. Numerous transgenic zebrafish with fluorescently marked blood cells enable live cell. Aug 01, 1995 fate maps of the zebrafish embryo woo, shih and fraser 441 to perform a direct assay of the distribution of progenitors in the region of the forming embryonic shield, we have performed singlecell fate mapping experiments using dye injections 15. Commitment of cell fate in the early zebrafish embryo science. Temporalspatial resolution fate mapping reveals distinct. Fate maps of the zebrafish embryo katherine woo, john shih and scott e fraser california institute of technology, pasadena, usa in the past few years, we have seen a surge of interest in the zebrafish as a model system for the study of embryonic induction and patterning.
Because a large number of mutations have been identified in recent years, these methods can be applied in a variety of contexts to provide a deep understanding of gene function that is often more difficult to achieve in other vertebrate model systems. The organization of the zebrafish fate map, therefore, is similar to the one of xenopus. In addition, the application of this technique serves as a key step in generating germline chimeras by cell transplantation methods 10. Using a combination of live imaging, retrospective and indicative fate mapping, and genetic studies, we show that mp and ssf precursors segregate at the beginning of segmentation and that they arise from distinct regions along the anteriorposterior ap and dorsalventral dv axes of the adaxial cell. When do single cells in the early zebrafish embryo become irreversibly committed to a specific fate. Classical fate mapping approaches in any embryo typically involve labeling of single cells with nondiffusible dyes by pressure or iontophoretic microinjection followed by tracing the fate s of those cells and their descendants throughout development. Order and coherence in the fate map of the zebrafish. Fate mapping techniques the embryo project encyclopedia. Feb 06, 2014 however, the epigenetic regulation of haematopoiesis regulation via micrornas mirnas remains partially understood.
A dynamic fate map of the forebrain shows how vertebrate. Further reduction of ptf1a gene dosage converts pancreatic progenitor cells to gall bladder and other nonpancreatic cell fates. Control of neural crest cell fate by the wnt signalling. However, to date it has remained unclear whether, as in mammals, ptf1aexpressing zebrafish pancreatic progenitors are able to contribute to multiple. To deconstruct regenerating adult epithelial tissue at single cell resolution, we created a multicolor system, skinbow, that barcodes the superficial epithelial cell sec population of zebrafish skin with dozens of. The fate map was developed by staining each individual cell and tracing each through gastrulation.
Ascl1b and neurod1, instead of neurog3, control pancreatic. Live fatemapping of jointassociated fibroblasts visualizes. We have developed new methods to address this class of problem and in particular the early morphogenesis of the eye, using the transparent zebrafish as a model. Live fate mapping of jointassociated fibroblasts visualizes expansion of cell contributions during zebrafish fin regeneration development. Mir1423p regulates cardiovascular system during zebrafish. Embryological manipulations in zebrafish springerlink. In the central nervous system cns, overall cell fate mapping is typically directed by the sonic hedgehog shh morphogen signaling pathway. Work with lineage tracing and fate mapping has shown that the marginal cells of the. The zebrafish has provided an attractive model system for investigating both muscle patterning and growth due to its simple axial musculature with spatially separated fibre types, which contrasts to complex muscle groups often deployed in amniotes.
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