The neural crest is an evolutionary novelty that fostered the emergence of vertebrate anatomical ... more The neural crest is an evolutionary novelty that fostered the emergence of vertebrate anatomical innovations such as the cranium and jaws. During embryonic development, multipotent neural crest cells are specified at the lateral borders of the neural plate before delaminating, migrating and differentiating into various cell types. In invertebrate chordates (cephalochordates and tunicates), neural plate border cells express conserved factors such as Msx, Snail and Pax3/7 and generate melanin-containing pigment cells, a derivative of the neural crest in vertebrates. However, invertebrate neural plate border cells have not been shown to generate homologues of other neural crest derivatives. Thus, proposed models of neural crest evolution postulate vertebrate-specific elaborations on an ancestral neural plate border program, through acquisition of migratory capabilities and the potential to generate several cell types. Here we show that a particular neuronal cell type in the tadpole larva of the tunicate Ciona intestinalis, the bipolar tail neuron, shares a set of features with neural-crest-derived spinal ganglia neurons in vertebrates. Bipolar tail neuron precursors derive from caudal neural plate border cells, delaminate and migrate along the paraxial mesoderm on either side of the neural tube, eventually differentiating into afferent neurons that form synaptic contacts with both epidermal sensory cells and motor neurons. We propose that the neural plate borders of the chordate ancestor already produced migratory peripheral neurons and pigment cells, and that the neural crest evolved through the acquisition of a multipotent progenitor regulatory state upstream of multiple, pre-existing neural plate border cell differentiation programs.
The anterior-most ectoderm of ascidian larvae contains the adhesive papillae, or palps, which pla... more The anterior-most ectoderm of ascidian larvae contains the adhesive papillae, or palps, which play an important role in triggering the metamorphosis of swimming tadpoles. In Ciona intestinalis, the palps consist of three conical protrusions within a field of thickened epithelium that form late in embryogenesis, as tailbuds mature into larvae. The palp protrusions express the LIM-homeodomain transcription factor Islet. Protrusion occurs through differential cell elongation, probably mediated by Islet, as we find that ectopic expression of Islet is sufficient to promote cell lengthening. FGF signaling is required for both Islet expression and palp morphogenesis. Importantly, we show that Islet expression can rescue the palp-deficient phenotype that results from inhibition of FGF signaling. We conclude that Islet is a key regulatory factor governing morphogenesis of the palps. It is conceivable that Islet is also essential for the cellular morphogenesis of placode-derived sensory neuro...
The Journal of neuroscience : the official journal of the Society for Neuroscience, Jan 4, 2012
Astrocytes, one of the most common cell types in the brain, are essential for processes ranging f... more Astrocytes, one of the most common cell types in the brain, are essential for processes ranging from neural development through potassium homeostasis to synaptic plasticity. Surprisingly, the developmental origins of astrocytes in the neocortex are still controversial. To investigate the patterns of astrocyte development in the neocortex we examined cortical development in a transgenic mouse in which a random, sparse subset of neural progenitors undergoes CRE/lox recombination, permanently labeling their progeny. We demonstrate that neural progenitors in neocortex generate discrete columnar structures that contain both projection neurons and protoplasmic astrocytes. Ninety-five percent of developmental cortical columns labeled in our system contained both astrocytes and neurons. The astrocyte to neuron ratio of labeled cells in a developmental column was 1:7.4, similar to the overall ratio of 1:8.4 across the entire gray matter of the neocortex, indicating that column-associated ast...
Gene regulatory networks control the progressive specification of cell types and govern morphogen... more Gene regulatory networks control the progressive specification of cell types and govern morphogenetic events during development. However, how morphogenetic events reciprocally affect gene expression remains poorly understood. Here, we analyzed the function of BMP signaling and expression of downstream target genes during cell migration of the precardiac mesoderm (trunk ventral cells, TVCs) in ascidian embryos. Our results indicate that migrating TVCs experience increasing BMP signaling as they migrate towards the ventral trunk epidermis, which expresses sustained levels of Bmp2/4. This increasing signaling intensity allows the successive activation of GATAa, Tolloid, Bmp2/4 and NK4. Initial activation of GATAa, Tolloid and Bmp2/4 contribute to a positive feedback loop involving cell migration, chordin inhibition and BMP ligand production. Sustained levels of BMP signaling become sufficient to activate NK4 expression, which in turn contributes to a negative feedback loop inhibiting Bmp2/4 and Tolloid expression. In addition, NK4 appears to inhibit cell migration thus providing a "transcriptional brake" to stop TVC migration. Our observations led us to propose a model for the coordination of cell migration and gene expression based on the temporal unfolding of a gene regulatory sub-network in a relevant developmental context.
The identification of subpharyngeal cardiac precursors has had a strong influence on the way we t... more The identification of subpharyngeal cardiac precursors has had a strong influence on the way we think about early cardiac development. From this discovery was born the concept of multiple heart fields. Early support for the concept came from gene expression, genetic retrospective fate mapping, and gene targeting studies, which collectively suggested the existence of a second heart field (SHF) on the basis of specific Islet-1 (Isl-1) expression, presence of two cardiac ancestral lineages, and compatible cardiac knockout phenotypes, respectively. A decade after the original studies, support for the SHF concept is dwindling. This is because in all bilaterian models studied, Isl expression in heart progenitors is not SHF-specific, because lineage data are best explained by alternative models including an older, truly ancestral, lineage of cardiac pioneers with unrestricted contribution to all cardiac segments and, finally, because the inflow-to-outflow segmental nature of the early vertebrate peristaltic heart has been reaffirmed with novel, less invasive, methodologies. Altogether, the paradigms derived from the discovery of subpharyngeal cardiac progenitors helped us shift from relatively simple models, which rely predominantly either on patterning, gene expression patterns or lineages, to a much more sophisticated body of knowledge in which all these parameters must be accounted. Thus, it is well possible that due consideration of the key elements contained in the inflow/outflow, pioneer/scaffold, ballooning, and SHF hypotheses may provide us with a unified framework of the early stages of cardiac development. Here, we advance into this direction by suggesting an intuitive model of early heart development based on the concept of an inflow/outflow scaffold erected by cardiac pioneers, one that is required to assemble all the subsequent cell contribution that emigrates from cardiac progenitor areas.
Knowing the normal patterns of embryonic cell proliferation, migration, and differentiation is a ... more Knowing the normal patterns of embryonic cell proliferation, migration, and differentiation is a cornerstone for understanding development. Yet for most species, the precision with which embryonic cell lineages can be determined is limited by technical considerations (the large numbers of cells, extended developmental times, opacity of the embryos), and these are exacerbated by the inherent variability of the lineages themselves. Here, we present an improved method of cell lineage tracing in the leech Helobdella, driving the expression of a nuclearly localized histone H2B:GFP (green fluorescent protein) fusion protein in selected lineages by microinjection of a plasmid vector. This construct generates a long lasting and minimally mosaic signal with single cell resolution, and does not disrupt the development of most lineages tested. We have validated this technique by elucidating details of cell lineages contributing to segmental and prostomial tissues that could not be observed with standard dextran lineage tracers.
Gene regulatory networks control the progressive specification of cell types and govern morphogen... more Gene regulatory networks control the progressive specification of cell types and govern morphogenetic events during development. However, how morphogenetic events reciprocally affect gene expression remains poorly understood. Here, we analyzed the function of BMP signaling and expression of downstream target genes during cell migration of the precardiac mesoderm (trunk ventral cells, TVCs) in ascidian embryos. Our results indicate that migrating TVCs experience increasing BMP signaling as they migrate towards the ventral trunk epidermis, which expresses sustained levels of Bmp2/4. This increasing signaling intensity allows the successive activation of GATAa, Tolloid, Bmp2/4 and NK4. Initial activation of GATAa, Tolloid and Bmp2/4 contribute to a positive feedback loop involving cell migration, chordin inhibition and BMP ligand production. Sustained levels of BMP signaling become sufficient to activate NK4 expression, which in turn contributes to a negative feedback loop inhibiting Bmp2/4 and Tolloid expression. In addition, NK4 appears to inhibit cell migration thus providing a "transcriptional brake" to stop TVC migration. Our observations led us to propose a model for the coordination of cell migration and gene expression based on the temporal unfolding of a gene regulatory sub-network in a relevant developmental context.
The motor ganglion (MG) controls the rhythmic swimming behavior of the Ciona intestinalis tadpole... more The motor ganglion (MG) controls the rhythmic swimming behavior of the Ciona intestinalis tadpole. Despite its cellular simplicity (five pairs of neurons), the MG exhibits conservation of transcription factor expression with the spinal cord of vertebrates. Evidence is presented that the developing MG is patterned by sequential Ephrin/FGF/MAPK and Delta/Notch signaling events. FGF/MAPK attenuation by a localized EphrinAb signal specifies posterior neuronal subtypes, which in turn relay a Delta2/Notch signal that specifies anterior fates. This short-range relay is distinct from the patterning of the vertebrate spinal cord, which is a result of opposing BMP and Shh morphogen gradients. Nonetheless, both mechanisms lead to localized expression of related homeodomain codes for the specification of distinct neuronal subtypes. This MG regulatory network provides a foundation for elucidating the genetic and cellular basis of a model chordate central pattern generator.
The neural crest is an evolutionary novelty that fostered the emergence of vertebrate anatomical ... more The neural crest is an evolutionary novelty that fostered the emergence of vertebrate anatomical innovations such as the cranium and jaws. During embryonic development, multipotent neural crest cells are specified at the lateral borders of the neural plate before delaminating, migrating and differentiating into various cell types. In invertebrate chordates (cephalochordates and tunicates), neural plate border cells express conserved factors such as Msx, Snail and Pax3/7 and generate melanin-containing pigment cells, a derivative of the neural crest in vertebrates. However, invertebrate neural plate border cells have not been shown to generate homologues of other neural crest derivatives. Thus, proposed models of neural crest evolution postulate vertebrate-specific elaborations on an ancestral neural plate border program, through acquisition of migratory capabilities and the potential to generate several cell types. Here we show that a particular neuronal cell type in the tadpole larva of the tunicate Ciona intestinalis, the bipolar tail neuron, shares a set of features with neural-crest-derived spinal ganglia neurons in vertebrates. Bipolar tail neuron precursors derive from caudal neural plate border cells, delaminate and migrate along the paraxial mesoderm on either side of the neural tube, eventually differentiating into afferent neurons that form synaptic contacts with both epidermal sensory cells and motor neurons. We propose that the neural plate borders of the chordate ancestor already produced migratory peripheral neurons and pigment cells, and that the neural crest evolved through the acquisition of a multipotent progenitor regulatory state upstream of multiple, pre-existing neural plate border cell differentiation programs.
The anterior-most ectoderm of ascidian larvae contains the adhesive papillae, or palps, which pla... more The anterior-most ectoderm of ascidian larvae contains the adhesive papillae, or palps, which play an important role in triggering the metamorphosis of swimming tadpoles. In Ciona intestinalis, the palps consist of three conical protrusions within a field of thickened epithelium that form late in embryogenesis, as tailbuds mature into larvae. The palp protrusions express the LIM-homeodomain transcription factor Islet. Protrusion occurs through differential cell elongation, probably mediated by Islet, as we find that ectopic expression of Islet is sufficient to promote cell lengthening. FGF signaling is required for both Islet expression and palp morphogenesis. Importantly, we show that Islet expression can rescue the palp-deficient phenotype that results from inhibition of FGF signaling. We conclude that Islet is a key regulatory factor governing morphogenesis of the palps. It is conceivable that Islet is also essential for the cellular morphogenesis of placode-derived sensory neuro...
The Journal of neuroscience : the official journal of the Society for Neuroscience, Jan 4, 2012
Astrocytes, one of the most common cell types in the brain, are essential for processes ranging f... more Astrocytes, one of the most common cell types in the brain, are essential for processes ranging from neural development through potassium homeostasis to synaptic plasticity. Surprisingly, the developmental origins of astrocytes in the neocortex are still controversial. To investigate the patterns of astrocyte development in the neocortex we examined cortical development in a transgenic mouse in which a random, sparse subset of neural progenitors undergoes CRE/lox recombination, permanently labeling their progeny. We demonstrate that neural progenitors in neocortex generate discrete columnar structures that contain both projection neurons and protoplasmic astrocytes. Ninety-five percent of developmental cortical columns labeled in our system contained both astrocytes and neurons. The astrocyte to neuron ratio of labeled cells in a developmental column was 1:7.4, similar to the overall ratio of 1:8.4 across the entire gray matter of the neocortex, indicating that column-associated ast...
Gene regulatory networks control the progressive specification of cell types and govern morphogen... more Gene regulatory networks control the progressive specification of cell types and govern morphogenetic events during development. However, how morphogenetic events reciprocally affect gene expression remains poorly understood. Here, we analyzed the function of BMP signaling and expression of downstream target genes during cell migration of the precardiac mesoderm (trunk ventral cells, TVCs) in ascidian embryos. Our results indicate that migrating TVCs experience increasing BMP signaling as they migrate towards the ventral trunk epidermis, which expresses sustained levels of Bmp2/4. This increasing signaling intensity allows the successive activation of GATAa, Tolloid, Bmp2/4 and NK4. Initial activation of GATAa, Tolloid and Bmp2/4 contribute to a positive feedback loop involving cell migration, chordin inhibition and BMP ligand production. Sustained levels of BMP signaling become sufficient to activate NK4 expression, which in turn contributes to a negative feedback loop inhibiting Bmp2/4 and Tolloid expression. In addition, NK4 appears to inhibit cell migration thus providing a "transcriptional brake" to stop TVC migration. Our observations led us to propose a model for the coordination of cell migration and gene expression based on the temporal unfolding of a gene regulatory sub-network in a relevant developmental context.
The identification of subpharyngeal cardiac precursors has had a strong influence on the way we t... more The identification of subpharyngeal cardiac precursors has had a strong influence on the way we think about early cardiac development. From this discovery was born the concept of multiple heart fields. Early support for the concept came from gene expression, genetic retrospective fate mapping, and gene targeting studies, which collectively suggested the existence of a second heart field (SHF) on the basis of specific Islet-1 (Isl-1) expression, presence of two cardiac ancestral lineages, and compatible cardiac knockout phenotypes, respectively. A decade after the original studies, support for the SHF concept is dwindling. This is because in all bilaterian models studied, Isl expression in heart progenitors is not SHF-specific, because lineage data are best explained by alternative models including an older, truly ancestral, lineage of cardiac pioneers with unrestricted contribution to all cardiac segments and, finally, because the inflow-to-outflow segmental nature of the early vertebrate peristaltic heart has been reaffirmed with novel, less invasive, methodologies. Altogether, the paradigms derived from the discovery of subpharyngeal cardiac progenitors helped us shift from relatively simple models, which rely predominantly either on patterning, gene expression patterns or lineages, to a much more sophisticated body of knowledge in which all these parameters must be accounted. Thus, it is well possible that due consideration of the key elements contained in the inflow/outflow, pioneer/scaffold, ballooning, and SHF hypotheses may provide us with a unified framework of the early stages of cardiac development. Here, we advance into this direction by suggesting an intuitive model of early heart development based on the concept of an inflow/outflow scaffold erected by cardiac pioneers, one that is required to assemble all the subsequent cell contribution that emigrates from cardiac progenitor areas.
Knowing the normal patterns of embryonic cell proliferation, migration, and differentiation is a ... more Knowing the normal patterns of embryonic cell proliferation, migration, and differentiation is a cornerstone for understanding development. Yet for most species, the precision with which embryonic cell lineages can be determined is limited by technical considerations (the large numbers of cells, extended developmental times, opacity of the embryos), and these are exacerbated by the inherent variability of the lineages themselves. Here, we present an improved method of cell lineage tracing in the leech Helobdella, driving the expression of a nuclearly localized histone H2B:GFP (green fluorescent protein) fusion protein in selected lineages by microinjection of a plasmid vector. This construct generates a long lasting and minimally mosaic signal with single cell resolution, and does not disrupt the development of most lineages tested. We have validated this technique by elucidating details of cell lineages contributing to segmental and prostomial tissues that could not be observed with standard dextran lineage tracers.
Gene regulatory networks control the progressive specification of cell types and govern morphogen... more Gene regulatory networks control the progressive specification of cell types and govern morphogenetic events during development. However, how morphogenetic events reciprocally affect gene expression remains poorly understood. Here, we analyzed the function of BMP signaling and expression of downstream target genes during cell migration of the precardiac mesoderm (trunk ventral cells, TVCs) in ascidian embryos. Our results indicate that migrating TVCs experience increasing BMP signaling as they migrate towards the ventral trunk epidermis, which expresses sustained levels of Bmp2/4. This increasing signaling intensity allows the successive activation of GATAa, Tolloid, Bmp2/4 and NK4. Initial activation of GATAa, Tolloid and Bmp2/4 contribute to a positive feedback loop involving cell migration, chordin inhibition and BMP ligand production. Sustained levels of BMP signaling become sufficient to activate NK4 expression, which in turn contributes to a negative feedback loop inhibiting Bmp2/4 and Tolloid expression. In addition, NK4 appears to inhibit cell migration thus providing a "transcriptional brake" to stop TVC migration. Our observations led us to propose a model for the coordination of cell migration and gene expression based on the temporal unfolding of a gene regulatory sub-network in a relevant developmental context.
The motor ganglion (MG) controls the rhythmic swimming behavior of the Ciona intestinalis tadpole... more The motor ganglion (MG) controls the rhythmic swimming behavior of the Ciona intestinalis tadpole. Despite its cellular simplicity (five pairs of neurons), the MG exhibits conservation of transcription factor expression with the spinal cord of vertebrates. Evidence is presented that the developing MG is patterned by sequential Ephrin/FGF/MAPK and Delta/Notch signaling events. FGF/MAPK attenuation by a localized EphrinAb signal specifies posterior neuronal subtypes, which in turn relay a Delta2/Notch signal that specifies anterior fates. This short-range relay is distinct from the patterning of the vertebrate spinal cord, which is a result of opposing BMP and Shh morphogen gradients. Nonetheless, both mechanisms lead to localized expression of related homeodomain codes for the specification of distinct neuronal subtypes. This MG regulatory network provides a foundation for elucidating the genetic and cellular basis of a model chordate central pattern generator.
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Papers by Alberto Stolfi