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Proneural genes are conserved drivers of neurogenesis across the animal kingdom. How their functions have adapted to guide humanspecific neurodevelopmental features is poorly understood. Here, we mined transcriptomic data from human fetal cortices and generated from human embryonic stem cell-derived cortical organoids (COs) to show that NEUROG1 and NEUROG2 are most highly expressed in basal neural progenitor cells, with pseudotime trajectory analyses indicating that NEUROG1-derived lineages predominate early and NEUROG2 lineages later. Using ChIP-qPCR, gene silencing and overexpression studies in COs, we show that NEUROG2 is necessary and sufficient to directly transactivate known target genes (NEUROD1, EOMES, RND2). To identify new targets, we engineered NEUROG2-mCherry knock-in human embryonic stem cells for CO generation. The mCherry-high CO cell transcriptome is enriched in extracellular matrix-associated genes, and two genes associated with human-accelerated regions: PPP1R17 and FZD8.We show that NEUROG2 binds COL1A1, COL3A1 and PPP1R17 regulatory elements, and induces their ectopic expression in COs, although NEUROG2 is not required for this expression. Neurog2 similarly induces Col3a1 and Ppp1r17 in murine P19 cells. These data are consistent with a conservation of NEUROG2 function across mammalian species. [ABSTRACT FROM AUTHOR]

Transit-amplifying (TA) cells are progenitors that undergo an amplification phase followed by transition into an extinction phase. A long postulated epidermal TA progenitor with biphasic behavior has not yet been experimentally observed in vivo. Here, we identify such a TA population using clonal analysis of Aspm-CreER genetic cell-marking in mice, which uncovers contribution to both homeostasis and injury repair of adult skin. This TA population is more frequently dividing than a Dlx1-CreER-marked long-term self-renewing (e.g. stem cell) population. Newly developed generalized birth-death modeling of long-term lineage tracing data shows that both TA progenitors and stem cells display neutral competition, but only the stem cells display neutral drift. The quantitative evolution of a nascent TA cell and its direct descendants shows that TA progenitors indeed amplify the basal layer before transition and that the homeostatic TA population is mostly in extinction phase. This model will be broadly useful for analyzing progenitors whose behavior changes with their clone age. This work identifies a long-missing class of non-self-renewing biphasic epidermal TA progenitors and has broad implications for understanding tissue renewal mechanisms. [ABSTRACT FROM AUTHOR]

Tissue-resident lymphoid cells (TLCs) span the spectrum of innateto-adaptive immune function. Unlike traditional, circulating lymphocytes that are continuously generated from hematopoietic stem cells (HSCs), many TLCs are of fetal origin and poorly generated from adult HSCs. Here, we sought to further understand murine TLC development and the roles of Flk2 and IL7Ra, two cytokine receptors with known function in traditional lymphopoiesis. Using Flk2- and Il7r-Cre lineage tracing, we found that peritoneal B1a cells, splenic marginal zone B (MZB) cells, lung ILC2s and regulatory T cells (Tregs) were highly labeled. Despite high labeling, loss of Flk2 minimally affected the generation of these cells. In contrast, loss of IL7Ra, or combined deletion of Flk2 and IL7Ra, dramatically reduced the number of B1a cells, MZBs, ILC2s and Tregs, both in situ and upon transplantation, indicating an intrinsic and essential role for IL7Ra. Surprisingly, reciprocal transplants of wild-type HSCs showed that an IL7Ra-/-environment selectively impaired reconstitution of TLCs when compared with TLC numbers in situ. Taken together, our data defined Flk2- and IL7Ra-positive TLC differentiation paths, and revealed functional roles of Flk2 and IL7Ra in TLC establishment. [ABSTRACT FROM AUTHOR]

In zebrafish, transgenic labeling approaches, robust regenerative responses and excellent in vivo imaging conditions enable precise characterization of immune cell behavior in response to injury. Here, we monitored osteoblast-immune cell interactions in bone, a tissuewhich is particularly difficult to in vivo image in tetrapod species. Ablation of individual osteoblasts leads to recruitment of neutrophils and macrophages in varying numbers, depending on the extent of the initial insult, and initiates generation of cathepsin K+ osteoclasts from macrophages. Osteoblast ablation triggers the production of pro-inflammatory cytokines and reactive oxygen species, which are needed for successful macrophage recruitment. Excess glucocorticoid signaling as it occurs during the stress response inhibits macrophage recruitment, maximum speed and changes the macrophage phenotype. Although osteoblast loss is compensated for within a day by contribution of committed osteoblasts, macrophages continue to populate the region. Their presence is required for osteoblasts to fill the lesion site. Our model enables visualization of bone repair after microlesions at single-cell resolution and demonstrates a proosteogenic function of tissue-resident macrophages in nonmammalian vertebrates. [ABSTRACT FROM AUTHOR]

Auditory and vestibular mechanosensory hair cells do not regenerate following injury or aging in the adult mammalian inner ear, inducing irreversible hearing loss and balance disorders for millions of people. Research on model systems showing replacement of mechanosensory cells can provide mechanistic insights into developing new regenerative therapies. Here, we developed lineage tracing systems to reveal the generation of mechanosensory neurons in the Johnston's organ (JO) of intact adult Drosophila, which are the functional counterparts to hair cells in vertebrates. New JO neurons develop cilia and target central brain circuitry. Unexpectedly, mitotic recombination clones point to JOneuron self-replication as a likely source of neuronal plasticity. This mechanism is further enhanced upon treatment with experimental and ototoxic compounds. Our findings introduce a new platform to expedite research on mechanisms and compounds mediating mechanosensory cell regeneration, with nascent implications for hearing and balance restoration. [ABSTRACT FROM AUTHOR]

Adult tissues contain label-retaining cells (LRCs), which are relatively slow-cycling and considered to represent a property of tissue stem cells (SCs). In the ocular surface epithelium, LRCs are present in the limbus and conjunctival fornix; however, the character of these LRCs remains unclear, owing to lack of appropriate molecular markers. Using three CreER transgenic mouse lines, we demonstrate that the ocular surface epithelium accommodates spatially distinct populations with different cell division dynamics. In the limbus, long-lived Slc1a3CreER-labeled SCs either migrate centripetally toward the central cornea or slowly expand their clones laterally within the limbal region. In the central cornea, non-LRCs labeled with Dlx1CreER and K14CreER behave as short-lived progenitor cells. The conjunctival epithelium in the bulbar, fornix and palpebral compartment is regenerated by regionally unique SC populations. Severe damage to the cornea leads to the cancellation of SC compartments and conjunctivalization, whereas milder limbal injury induces a rapid increase of laterally expanding clones in the limbus. Taken together, our work defines compartmentalized multiple SC/ progenitor populations of the mouse eye in homeostasis and their behavioral changes in response to injury. [ABSTRACT FROM AUTHOR]

Salivary glands exert exocrine secretory function to provide saliva for lubrication and protection of the oral cavity. Its epithelium consists of several differentiated cell types, including acinar, ductal and myoepithelial cells, that are maintained in a lineage-restricted manner during homeostasis or after mild injuries. Glandular regeneration following a near complete loss of secretory cells, however, may involve cellular plasticity, although the mechanism and extent of such plasticity remain unclear. Here, by combining lineage-tracing experiments with a model of severe glandular injury in the mouse submandibular gland, we show that de novo formation of acini involves induction of cellular plasticity in multiple non-acinar cell populations. Fate-mapping analysis revealed that, although ductal stem cells marked by cytokeratin K14 and Axin2 undergo a multipotency switch, they do not make a significant contribution to acinar regeneration. Intriguingly, more than 80% of regenerated acini derive from differentiated cells, including myoepithelial and ductal cells, that appear to dedifferentiate to a progenitor-like state before re-differentiation into acinar cells. The potential of diverse cell populations serving as a reserve source for acini widens the therapeutic options for hyposalivation. [ABSTRACT FROM AUTHOR]

New neurons are generated in the postnatal rodent hypothalamus, with a subset of tanycytes in the third ventricular (3V) wall serving as neural stem/progenitor cells. However, the precise stem cell niche organization, the intermediate steps and the endogenous regulators of postnatal hypothalamic neurogenesis remain elusive. Quantitative lineage-tracing in vivo revealed that conditional deletion of fibroblast growth factor 10 (Fgf10) from Fgf10-expressing ß-tanycytes at postnatal days (P)4-5 results in the generation of significantly more parenchymal cells by P28, composed mostly of ventromedial and dorsomedial neurons and some glial cells, which persist into adulthood. A closer scrutiny in vivo and ex vivo revealed that the 3V wall is not static and is amenable to cell movements. Furthermore, normally ß-tanycytes give rise to parenchymal cells via an intermediate population of a-tanycytes with transient amplifying cell characteristics. Loss of Fgf10 temporarily attenuates the amplification of ß-tanycytes but also appears to delay the exit of their a-tanycyte descendants from the germinal 3V wall. Our findings suggest that transience of cells through the a-tanycyte domain is a key feature, and Fgf10 is a negative regulator of postnatal hypothalamic neurogenesis. [ABSTRACT FROM AUTHOR]

The past decade has seen a renewed appreciation of the central importance of cellular lineages to many questions in biology (especially organogenesis, stem cells and tumor biology). This has been driven in part by a renaissance in genetic clonal-labeling techniques. Recent approaches are based on accelerated mutation of DNA sequences, which can then be sequenced from individual cells to re-create a 'phylogenetic' tree of cell lineage. However, current approaches depend on making transgenic alterations to the genome in question, which limit their application. Here, we introduce a new method that completely avoids the need for prior genetic engineering, by identifying endogenous CRISPR/Cas9 target arrays suitable for lineage analysis. In both mouse and zebrafish, we identify the highest quality compact arrays as judged by equal base composition, 5'G sequence, minimal likelihood of residing in the functional genome, minimal off targets and ease of amplification. We validate multiple high-quality endogenous CRISPR/Cas9 arrays, demonstrating their utility for lineage tracing. Our pragmatically scalable technique thus can produce deep and broad lineages in vivo, while removing the dependence on genetic engineering. [ABSTRACT FROM AUTHOR]

Neural remodeling is essential for the development of a functional nervous system and has been extensively studied in the metamorphosis of Drosophila. Despite the crucial roles of glial cells in brain functions, including learning and behavior, little is known of how adult glial cells develop in the context of neural remodeling. Here, we show that the architecture of neuropil-glia in the adult Drosophila brain, composed of astrocyte-like glia (ALG) and ensheathing glia (EG), robustly develops from two different populations in the larva: the larval EG and glial cell missing-positive (gcm+) cells. While larval EG dedifferentiate, proliferate, and redifferentiate into adult ALG and EG, gcm+ cells also generate the same glial subtypes. Each glial lineage occupies a certain brain area complementary to the other, and together they form the adult neuropil-glia architecture. Both lineages require the FGF receptor Heartless to proliferate, and the homeoprotein Prospero to differentiate into ALG. Lineage-specific inhibition of gliogenesis revealed that each lineage compensates for deficiency in the proliferation of the other. Together, the lineages ensure the robust development of adult neuropil-glia, thereby ensuring a functional brain. [ABSTRACT FROM AUTHOR]

Alveologenesis is an essential developmental process that increases the surface area of the lung through the formation of septal ridges. In the mouse, septation occurs postnatally and is thought to require the alveolar myofibroblast (AMF). Though abundant during alveologenesis, markers for AMFs are minimally detected in the adult. After septation, the alveolar walls thin to allow efficient gas exchange. Both loss of AMFs or retention and differentiation into another cell type during septal thinning have been proposed. Using a novel Fgf18:CreERT2 allele to lineage trace AMFs, we demonstrate that most AMFs are developmentally cleared during alveologenesis. Lung mesenchyme also contains other poorly described cell types, including alveolar lipofibroblasts (ALF). We show that Gli1:CreERT2 marks both AMFs as well as ALFs, and lineage tracing shows that ALFs are retained in adult alveoli while AMFs are lost. We further show that multiple immune cell populations contain lineage-labeled particles, suggesting a phagocytic role in the clearance of AMFs. The demonstration that the AMF lineage is depleted during septal thinning through a phagocytic process provides a mechanism for the clearance of a transient developmental cell population. [ABSTRACT FROM AUTHOR]

Alveologenesis is an essential developmental process that increases the surface area of the lung through the formation of septal ridges. In the mouse, septation occurs postnatally and is thought to require the alveolar myofibroblast (AMF). Though abundant during alveologenesis, markers for AMFs are minimally detected in the adult. After septation, the alveolar walls thin to allow efficient gas exchange. Both loss of AMFs or retention and differentiation into another cell type during septal thinning have been proposed. Using a novel Fgf18:CreERT2 allele to lineage trace AMFs, we demonstrate that most AMFs are developmentally cleared during alveologenesis. Lung mesenchyme also contains other poorly described cell types including alveolar lipofibroblasts (ALF). We show that Gli1:CreERT2 marks both AMFs as well as ALFs and lineage tracing shows that ALFs are retained in adult alveoli while AMFs are lost. We further show that multiple immune cell populations contain lineage labeled particles suggesting a phagocytic role in the clearance of AMFs. The demonstration that the AMF lineage is depleted during septal thinning through a phagocytic process provides a mechanism for the clearance of a transient developmental cell population. [ABSTRACT FROM AUTHOR]

The discovery of a fetal origin for tissue-resident macrophages (trMacs) has inspired an intense search for the mechanisms underlying their development. Here, we performed in vivo lineage tracing of cells with an expression history of IL7Rα, a marker exclusively associated with the lymphoid lineage in adult hematopoiesis. Surprisingly, we found that Il7r-Cre labeled fetalderived, adult trMacs. Labeling was almost complete in some tissues and partial in others. The putative progenitors of trMacs, yolk sac (YS) erythromyeloid progenitors, did not express IL7R, and YS hematopoiesis was unperturbed in IL7R-deficient mice. In contrast, tracking of IL7Rα message levels, surface expression, and Il7r-Cremediated labeling across fetal development revealed dynamic regulation of Il7r mRNA expression and rapid upregulation of IL7Rα surface protein upon transition from monocyte to macrophage within fetal tissues. Fetal monocyte differentiation in vitro produced IL7R+ macrophages, supporting a direct progenitor-progeny relationship. Additionally, blockade of IL7R function during late gestation specifically impaired the establishment of fetal-derived trMacs in vivo. These data provide evidence for a distinct function of IL7Rα in fetal myelopoiesis and identify IL7R as a novel regulator of trMac development. [ABSTRACT FROM AUTHOR]

The embryonic origin of distinct fat depots and the role for ontogeny in specifying the functional differences among adipocyte lineages between and within depots is unclear. Using a Cre/Lox-based strategy to track the fate of major mesodermal subcompartments in mice we present evidence that <50% of interscapular brown adipocytes are derived from progenitors of the central dermomyotome. Furthermore, we demonstrate that depot-specific adipocyte lineages spatially diverge as early as gastrulation, and that perigonadal adipocytes arise from separate mesodermal subcompartments in males and females. Last, we show adipocyte precursors (APs) of distinct lineages within the same depot exhibit indistinguishable responses to a high fat diet, indicating that ontogenetic differences between APs do not necessarily correspond to functional differences in this context. Altogether, these findings shed light on adipose tissue patterning and suggest that the behavior of adipocyte lineage cells is not strictly determined by developmental history. [ABSTRACT FROM AUTHOR]

During pregnancy the trophoblast cells of the placenta are the only fetal cells in direct contact with maternal blood and decidua. Their functions include transport of nutrients and oxygen, secretion of pregnancy hormones, remodelling of the uterine arteries, and communicating with maternal cells. Despite the importance of trophoblast cells in placental development and successful pregnancy, little is known about the identity, location and differentiation of human trophoblast progenitors. We identify a proliferative trophoblast niche at the base of the cytotrophoblast cell columns in first trimester placentas that is characterised by integrin α2 (ITGA2) expression. Pulse-chase experiments with 5-iodo-2...-deoxyuridine indicate that these cells might contribute to both villous (VCT) and extravillous (EVT) lineages. These proliferating trophoblast cells can be isolated by flow cytometry using ITGA2 as a marker and express genes from both VCT and EVT. Microarray expression analysis shows that ITAG2+ cells display a unique transcriptional signature, including genes involved in NOTCH signalling, and exhibit a combination of epithelial and mesenchymal characteristics. ITGA2 thus marks a niche allowing the study of pure populations of trophoblast progenitor cells. [ABSTRACT FROM AUTHOR]

In the field of heart regeneration, the proliferative potential of cardiomyocytes in postnatal mice is under intense investigation. However, solely relying on immunostaining of proliferation markers, the long-term proliferation dynamics and potential of the cardiomyocytes cannot be readily addressed. Previously, we found that a p53 promoter-driving reporter predominantly marked the proliferating lineages in mice. Here, we established a p53-based genetic tracing system to investigate postnatal cardiomyocyte proliferation and heart regeneration. By selectively tracing proliferative cardiomyocytes, a differential pattern of clonal expansion in p53+ cardiac myocytes was revealed in neonatal, adolescent and adult stages. In addition, the percentage of p53+ lineage cardiomyocytes increased continuously in the first month. Furthermore, these cells rapidly responded to heart injury and greatly contributed to the replenished myocardium. Therefore, this study reveals complex proliferating dynamics in postnatal cardiomyocytes and heart repair, and provides a novel genetic tracing strategy for studying postnatal cardiac turnover and regeneration. [ABSTRACT FROM AUTHOR]

Genome-editing technology has revolutionized the field of biology. Here, we report a novel de novo gene-targeting method mediated by in utero electroporation into the developing mammalian brain. Electroporation of donor DNA with the CRISPR/Cas9 system vectors successfully leads to knock-in of the donor sequence, such as EGFP, to the target site via the homology-directed repair mechanism. We developed a targeting vector system optimized to prevent anomalous leaky expression of the donor gene from the plasmid, which otherwise often occurs depending on the donor sequence. The knock-in efficiency of the electroporated progenitors reached up to 40% in the early stage and 20% in the late stage of the developing mouse brain. Furthermore, we inserted different fluorescent markers into the target gene in each homologous chromosome, successfully distinguishing homozygous knock-in cells by color. We also applied this de novo gene targeting to the ferret model for the study of complex mammalian brains. Our results demonstrate that this technique is widely applicable for monitoring gene expression, visualizing protein localization, lineage analysis and gene knockout, all at the single-cell level, in developmental tissues. [ABSTRACT FROM AUTHOR]

Regenerative responses in the vertebrate CNS depend on quiescent radial glia stem cells, which re-enter the cell cycle and eventually differentiate into neurons. The entry into the cell cycle and the differentiation into neurons are events of opposite nature, and therefore efforts to force quiescent radial glia into neurons require different factors. Here, we use fish to show that a single neurogenic factor, Atoh7, directs retinal radial glia (Mü ller glia, MG) into proliferation. The resulting neurogenic clusters differentiate in vivo into various retinal neurons. We use signaling reporters to demonstrate that the Atoh7-induced regeneration-like response of MG cells is mimicked by Notch, resembling the behavior of early progenitors during retinogenesis. Activation of Notch signaling in MG cells is sufficient to trigger proliferation and differentiation. Our results uncover a new role for Atoh7 as a universal neurogenic factor, and illustrate how signaling modules are re-employed in diverse contexts to trigger different biological responses. [ABSTRACT FROM AUTHOR]

Macrophages are specialized phagocytic cells, present in all tissues, which engulf and digest pathogens, infected and dying cells, and debris, and can recruit and regulate other immune cells and the inflammatory response and aid in tissue repair. Macrophage subpopulations play distinct roles in these processes and in disease, and are typically recognized by differences in marker expression, immune function, or tissue of residency. Although macrophage subpopulations in the brain have been found to have distinct developmental origins, the extent to which development contributes to macrophage diversity between tissues and within tissues is not well understood. Here, we investigate the development and maintenance of mouse lung macrophages by marker expression patterns, genetic lineage tracing and parabiosis. We show that macrophages populate the lung in three developmental waves, each giving rise to a distinct lineage. These lineages express different markers, reside in different locations, renew in different ways, and show little or no interconversion. Thus, development contributes significantly to lung macrophage diversity and targets each lineage to a different anatomical domain. [ABSTRACT FROM AUTHOR]

Recent lineage-tracing studies based on inducible genetic labelling have emphasized a crucial role for stochasticity in the maintenance and regeneration of cycling adult tissues. These studies have revealed that stem cells are frequently lost through differentiation and that this is compensated for by the duplication of neighbours, leading to the consolidation of clonal diversity. Through the combination of long-term lineage-tracing assays with short-term in vivo live imaging, the cellular basis of this stochastic stem cell loss and replacement has begun to be resolved. With a focus on mammalian spermatogenesis, intestinal maintenance and the hair cycle, we review the role of dynamic heterogeneity in the regulation of adult stem cell populations. [ABSTRACT FROM AUTHOR]

Tendon attaches to bone across a specialized tissue called the enthesis. This tissue modulates the transfer of muscle forces between two materials, i.e. tendon and bone, with vastly different mechanical properties. The enthesis for many tendons consists of a mineralized graded fibrocartilage that develops postnatally, concurrent with epiphyseal mineralization. Although it is well described that the mineralization and development of functional maturity requires muscle loading, the biological factors that modulate enthesis development are poorly understood. By genetically demarcating cells expressingGli1 in response to Hedgehog(Hh) signaling, we discovered a unique population of Hh-responsive cells in the developing murine enthesis that were distinct from tendon fibroblasts and epiphyseal chondrocytes. Lineage-tracing experiments revealed that the Gli1 lineage cells that originate in utero eventually populate the entiremature enthesis. Muscle paralysis increased the number of Hh-responsive cells in the enthesis, demonstrating that responsiveness to Hh is modulated in part by muscle loading. Ablation of the Hh-responsive cells during the first week of postnatal development resulted in a loss of mineralized fibrocartilage, with very little tissue remodeling 5 weeks after cell ablation. Conditional deletion of smoothened, a molecule necessary for responsiveness to Ihh, from the developing tendon and enthesis altered the differentiation of enthesis progenitor cells, resulting in significantly reduced fibrocartilage mineralization and decreased biomechanical function. Taken together, these results demonstrate that Hh signaling within developing enthesis fibrocartilage cells is required for enthesis formation. [ABSTRACT FROM AUTHOR]

The Lef/Tcf-family transcription factor Tcf3 has important roles in development, stem cell function and malignancy. Previous gain- and loss-of-function studies have suggested that Tcf3 is a mediator of selfrenewal and an undifferentiated state in stem and progenitor cells in skin, but little is known of its role in other postnatal tissues. Here, we explore the distribution and behavior of Tcf3-expressing cells in several adult tissues using a novel Tcf3-CreER knock-in mouse model. By lineage tracing in dorsal skin, we verify that Tcf3-expressing cells in the hair follicle bulge are self-renewing stem cells with multilineage potential. We then demonstrate, for the first time, the presence of Tcf3-expressing cells in the basal layer of several other stratified epithelia, including the paw skin, tongue and esophagus. By lineage tracing, we demonstrate that the Tcf3-expressing population in these tissues includes persistent stem cells, transient progenitors and cells undergoing active differentiation. Our observations here suggest that the role of Tcf3 in cell-fate decision is more complex than previously appreciated and is highly dependent on cellular context. [ABSTRACT FROM AUTHOR]

Observation of how cells divide, grow, migrate and form different parts of a developing organism is crucial for understanding developmental programs. Here, we describe a multicolor imaging tool named Raeppli (after the colorful confetti used at the carnival in Basel). Raeppli allows whole-tissue labeling such that the descendants of the majority of cells in a single organ are labeled and can be followed simultaneously relative to one another. We tested the use of Raeppli in the Drosophila melanogaster wing imaginal disc. Induction of Raeppli during larval stages irreversibly labels >90% of the cells with one of four spectrally separable, bright fluorescent proteins with low bias of selection. To understand the global growth characteristics of imaginal discs better, we induced Raeppli at various stages of development, imaged multiple fixed discs at the end of their larval development and estimated the size of their pouch primordium at those developmental stages. We also imaged the same wing disc through the larval cuticle at different stages of its development; the clones marked by Raeppli provide landmarks that can be correlated between multiple time points. Finally, we used Raeppli for continuous live imaging of prepupal eversion of the wing disc. [ABSTRACT FROM AUTHOR]

The lung mesenchyme consists of a widely heterogeneous population of cells that play crucial roles during development and homeostasis after birth. These cells belong to myogenic, adipogenic, chondrogenic, neuronal and other lineages. Yet, no clear hierarchy for these lineages has been established. We have previously generated a novel Fgf10iCre knock-in mouse line that allows lineage tracing of Fgf10-positive cells during development and postnatally. Using these mice, we hereby demonstrate the presence of two waves of Fgf10 expression during embryonic lung development: the first wave, comprising Fgf10-positive cells residing in the submesothelial mesenchyme at early pseudoglandular stage (as well as their descendants); and the second wave, comprising Fgf10- positive cells from late pseudoglandular stage (as well as their descendants). Our lineage-tracing data reveal that the first wave contributes to the formation of parabronchial and vascular smooth muscle cells as well as lipofibroblasts at later developmental stages, whereas the second wave does not give rise to smooth muscle cells but to lipofibroblasts as well as an Nkx2.1- E-Cad- Epcam+ Pro-Spc+ lineage that requires further in-depth analysis. During alveologenesis, Fgf10-positive cells give rise to lipofibroblasts rather than alveolar myofibroblasts, and during adult life, a subpopulation of Fgf10- expressing cells represents a pool of resident mesenchymal stromal (stem) cells (MSCs) (Cd45- Cd31- Sca-1+). Taken together, we show for the first time that Fgf10-expressing cells represent a pool of mesenchymal progenitors in the embryonic and postnatal lung. Our findings suggest that Fgf10-positive cells could be useful for developing stem cell-based therapies for treating interstitial lung diseases. [ABSTRACT FROM AUTHOR]

Second heart field (SHF) progenitors perform essential functions during mammalian cardiogenesis. We recently identified a population of cardiac progenitor cells (CPCs) in zebrafish expressing latent TGFβ-binding protein 3 (ltbp3) that exhibits several defining characteristics of the anterior SHF in mammals. However, ltbp3 transcripts are conspicuously absent in anterior lateral plate mesoderm (ALPM), where SHF progenitors are specified in higher vertebrates. Instead, ltbp3 expression initiates at the arterial pole of the developing heart tube. Because the mechanisms of cardiac development are conserved evolutionarily, we hypothesized that zebrafish SHF specification also occurs in the ALPM. To test this hypothesis, we Cre/loxP lineage traced gata4+ and nkx2.5+ ALPM populations predicted to contain SHF progenitors, based on evolutionary conservation of ALPM patterning. Traced cells were identified in SHF-derived distal ventricular myocardium and in three lineages in the outflow tract (OFT). We confirmed the extent of contributions made by ALPM nkx2.5+ cells using Kaede photoconversion. Taken together, these data demonstrate that, as in higher vertebrates, zebrafish SHF progenitors are specified within the ALPM and express nkx2.5. Furthermore, we tested the hypothesis that Nkx2.5 plays a conserved and essential role during zebrafish SHF development. Embryos injected with an nkx2.5 morpholino exhibited SHF phenotypes caused by compromised progenitor cell proliferation. Co-injecting low doses of nkx2.5 and ltbp3 morpholinos revealed a genetic interaction between these factors. Taken together, our data highlight two conserved features of zebrafish SHF development, reveal a novel genetic relationship between nkx2.5 and ltbp3, and underscore the utility of this model organism for deciphering SHF biology. [ABSTRACT FROM AUTHOR]

Pancreatic multipotent progenitor cells (MPCs) produce acinar, endocrine and duct cells during organogenesis, but their existence and location in the mature organ remain contentious. We used inducible lineage-tracing from the MPC-instructive gene Ptf1a to define systematically in mice the switch of Ptf1a+ MPCs to unipotent proacinar competence during the secondary transition, their rapid decline during organogenesis, and absence from the mature organ. Between E11.5 and E15.5, we describe tip epithelium heterogeneity, suggesting that putative Ptf1a+Sox9+Hnf1β+ MPCs are intermingled with Ptf1aHISox9LO proacinar progenitors. In the adult, pancreatic duct ligation (PDL) caused facultative reactivation of multipotency factors (Sox9 and Hnf1β) in Ptf1a+ acini, which undergo rapid reprogramming to duct cells and longer-term reprogramming to endocrine cells, including insulin+ β-cells that are mature by the criteria of producing Pdx1HI, Nkx6.1+ and MafA+. These Ptf1a lineage-derived endocrine/β-cells are likely formed via Ck19+/Hnf1β+/Sox9+ ductal and Ngn3+ endocrine progenitor intermediates. Acinar to endocrine/β-cell transdifferentiation was enhanced by combining PDL with pharmacological elimination of pre-existing β-cells. Thus, we show that acinar cells, without exogenously introduced factors, can regain aspects of embryonic multipotentiality under injury, and convert into mature β-cells. [ABSTRACT FROM AUTHOR]

Hematopoietic cells have been reported to convert into a number of non-hematopoietic cells types after transplantation/injury. Here, we have used a lineage tracing approach to determine whether hematopoietic plasticity is relevant for the normal development of hepatocytes and endothelial cells, both of which develop in close association with blood cells. Two mouse models were analyzed: vav ancestry mice, in which essentially all hematopoietic cells, including stem cells, irreversibly express yellow fluorescent protein (YFP); and lysozyme ancestry mice, in which all macrophages, as well as a small subset of all other non-myeloid hematopoietic cells, are labeled. Both lines were found to contain YFP+ hepatocytes at similar frequencies, indicating that macrophage to hepatocyte contributions occur in unperturbed mice. However, the YFP+ hepatocytes never formed clusters larger than three cells, suggesting a postnatal origin. In addition, the frequency of these cells was very low (∼1 in 75,000) and only increased two- to threefold after acute liver injury. Analysis of the two mouse models revealed no evidence for a hematopoietic origin of endothelial cells, showing that definitive HSCs do not function as hemangioblasts during normal development. Using endothelial cells and hepatocytes as paradigms, our study indicates that hematopoietic cells are tightly restricted in their differentiation potential during mouse embryo development and that hematopoietic plasticity plays at best a minor role in adult organ maintenance and regeneration. [ABSTRACT FROM AUTHOR]

The conducting airways (bronchi and bronchioles) and peripheral gas exchange (alveolar) regions of the mammalian lung are generated by a process of branching morphogenesis. Evidence suggests that during embryonic development, the undifferentiated epithelial progenitors are located at the distal tips of the branching epithelium. To test this hypothesis, we used an Id2-CreERT2 knock-in mouse strain to lineage trace the distal epithelial tip cells during either the pseudoglandular or canalicular phases of development. During the pseudoglandular stage, the tip cells both self-renew and contribute descendents to all epithelial cell lineages, including neuroendocrine cells. In addition, individual Id2+ tip cells can self-renew and contribute descendents to both the bronchiolar and alveolar compartments. By contrast, during the later canalicular stage, the distal epithelial tip cells only contribute descendents to the alveoli. Taken together, this evidence supports a model in which the distal tip of the developing lung contains a multipotent epithelial population, the fate of which changes during development. [ABSTRACT FROM AUTHOR]

The vertebrate neural retina comprises six classes of neurons and one class of glial cells, all derived from a population of multipotent progenitors. There is little information on the molecular mechanisms governing the specification of cell type identity from multipotent progenitors in the developing retina. We report that Ptf1a, a basic-helix-loop-helix (bHLH) transcription factor, is transiently expressed by post-mitotic precursors in the developing mouse retina. Recombination-based lineage tracing analysis in vivo revealed that Ptf1a expression marks retinal precursors with competence to exclusively produce horizontal and amacrine neurons. Inactivation of Ptf1a leads to a fate-switch in these precursors that causes them to adopt a ganglion cell fate. This mis-specification of neurons results in a complete loss of horizontal cells, a profound decrease of amacrine cells and an increase in ganglion cells. Furthermore, we identify Ptf1a as a primary downstream target for Foxn4, a forkhead transcription factor involved in the genesis of horizontal and amacrine neurons. These data, together with the previous findings on Foxn4, provide a model in which the Foxn4-Ptf1a pathway plays a central role in directing the differentiation of retinal progenitors towards horizontal and amacrine cell fates. [ABSTRACT FROM AUTHOR]

Notch receptor-mediated cell-cell signaling is known to negatively regulate neurogenesis in both vertebrate and invertebrate species, while being implicated in promoting the acquisition of glial fates. We studied Notch1 function directly during retinal neurogenesis by selective Cre/loxP-triggered Notch1 gene inactivation in peripheral retinal progenitor cells (RPCs) prior to the onset of cell differentiation. Consistent with its previously established role, Notch1 inactivation led to dramatic alteration in the expression profile of multiple basic helix-loop-helix transcription factors, consequently prompting premature cell-cycle exit and neuronal specification. Surprisingly, however, Notch1 inactivation led to a striking change in retinal cell composition, with cone-photoreceptor precursors expanding at the expense of other early- as well as late-born cell fates. Intriguingly, the Notch1-deficient precursors adhered to the normal chronological sequence of the cone-photoreceptor differentiation program. Together, these findings reveal an unexpected role of Notch signaling in directly controlling neuronal cell-type composition, and suggest a model by which, during normal retinogenesis, Notch1 functions to suppress cone-photoreceptor fate, allowing for the specification of the diversity of retinal cell types [ABSTRACT FROM AUTHOR]