RESUMEN
Dorsal interneurons (dIs) in the spinal cord encode the perception of touch, pain, heat, itchiness and proprioception. Previous studies using genetic strategies in animal models have revealed important insights into dI development, but the molecular details of how dIs arise as distinct populations of neurons remain incomplete. We have developed a resource to investigate dI fate specification by combining a single-cell RNA-Seq atlas of mouse embryonic stem cell-derived dIs with pseudotime analyses. To validate this in silico resource as a useful tool, we used it to first identify genes that are candidates for directing the transition states that lead to distinct dI lineage trajectories, and then validated them using in situ hybridization analyses in the developing mouse spinal cord in vivo. We have also identified an endpoint of the dI5 lineage trajectory and found that dIs become more transcriptionally homogeneous during terminal differentiation. This study introduces a valuable tool for further discovery about the timing of gene expression during dI differentiation and demonstrates its utility in clarifying dI lineage relationships.
Asunto(s)
Diferenciación Celular , Linaje de la Célula , Regulación del Desarrollo de la Expresión Génica , Interneuronas , Médula Espinal , Animales , Ratones , Médula Espinal/metabolismo , Médula Espinal/embriología , Linaje de la Célula/genética , Interneuronas/metabolismo , Interneuronas/citología , Diferenciación Celular/genética , Análisis de la Célula Individual , Células Madre Embrionarias de Ratones/metabolismo , Células Madre Embrionarias de Ratones/citología , RNA-SeqRESUMEN
Germline mutations in the RAS/ERK signaling pathway underlie several related developmental disorders collectively termed neuro-cardio-facial-cutaneous (NCFC) syndromes. NCFC patients manifest varying degrees of cognitive impairment, but the developmental basis of their brain abnormalities remains largely unknown. Neurofibromatosis type 1 (NF1), an NCFC syndrome, is caused by loss-of-function heterozygous mutations in the NF1 gene, which encodes neurofibromin, a RAS GTPase-activating protein. Here, we show that biallelic Nf1 inactivation promotes Erk-dependent, ectopic Olig2 expression specifically in transit-amplifying progenitors, leading to increased gliogenesis at the expense of neurogenesis in neonatal and adult subventricular zone (SVZ). Nf1-deficient brains exhibit enlarged corpus callosum, a structural defect linked to severe learning deficits in NF1 patients. Strikingly, these NF1-associated developmental defects are rescued by transient treatment with an MEK/ERK inhibitor during neonatal stages. This study reveals a critical role for Nf1 in maintaining postnatal SVZ-derived neurogenesis and identifies a potential therapeutic window for treating NF1-associated brain abnormalities.
Asunto(s)
Encéfalo/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células-Madre Neurales/patología , Neurofibromatosis 1/patología , Neurofibromina 1/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Cuerpo Calloso/patología , Humanos , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/metabolismo , Neurofibromatosis 1/embriología , Neurofibromatosis 1/metabolismo , Neurofibromina 1/genética , Neuroglía/patología , Factor de Transcripción 2 de los OligodendrocitosRESUMEN
Zika virus (ZIKV) has become a public health threat due to its global transmission and link to severe congenital disorders. The host immune responses to ZIKV infection have not been fully elucidated, and effective therapeutics are not currently available. Herein, we demonstrated that cholesterol-25-hydroxylase (CH25H) was induced in response to ZIKV infection and that its enzymatic product, 25-hydroxycholesterol (25HC), was a critical mediator of host protection against ZIKV. Synthetic 25HC addition inhibited ZIKV infection in vitro by blocking viral entry, and treatment with 25HC reduced viremia and conferred protection against ZIKV in mice and rhesus macaques. 25HC suppressed ZIKV infection and reduced tissue damage in human cortical organoids and the embryonic brain of the ZIKV-induced mouse microcephaly model. Our findings highlight the protective role of CH25H during ZIKV infection and the potential use of 25HC as a natural antiviral agent to combat ZIKV infection and prevent ZIKV-associated outcomes, such as microcephaly.
Asunto(s)
Antivirales/farmacología , Hidroxicolesteroles/farmacología , Microcefalia/virología , Infección por el Virus Zika/complicaciones , Animales , Encéfalo/efectos de los fármacos , Modelos Animales de Enfermedad , Técnica del Anticuerpo Fluorescente , Humanos , Macaca mulatta , Ratones , Microscopía Confocal , Internalización del Virus/efectos de los fármacos , Virus Zika/efectos de los fármacos , Virus Zika/fisiologíaRESUMEN
Neural progenitor cells within the cerebral cortex undergo a characteristic switch between symmetric self-renewing cell divisions early in development and asymmetric neurogenic divisions later. Yet, the mechanisms controlling this transition remain unclear. Previous work has shown that early but not late neural progenitor cells (NPCs) endogenously express the autism-linked transcription factor Foxp1, and both loss and gain of Foxp1 function can alter NPC activity and fate choices. Here, we show that premature loss of Foxp1 upregulates transcriptional programs regulating angiogenesis, glycolysis, and cellular responses to hypoxia. These changes coincide with a premature destabilization of HIF-1α, an elevation in HIF-1α target genes, including Vegfa in NPCs, and precocious vascular network development. In vitro experiments demonstrate that stabilization of HIF-1α in Foxp1-deficient NPCs rescues the premature differentiation phenotype and restores NPC maintenance. Our data indicate that the endogenous decline in Foxp1 expression activates the HIF-1α transcriptional program leading to changes in the tissue environment adjacent to NPCs, which, in turn, might alter their self-renewal and neurogenic capacities.
Asunto(s)
Corteza Cerebral , Factores de Transcripción Forkhead , Subunidad alfa del Factor 1 Inducible por Hipoxia , Células-Madre Neurales , Proteínas Represoras , Transducción de Señal , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Factores de Transcripción Forkhead/metabolismo , Factores de Transcripción Forkhead/genética , Células-Madre Neurales/metabolismo , Células-Madre Neurales/citología , Animales , Ratones , Corteza Cerebral/metabolismo , Corteza Cerebral/citología , Proteínas Represoras/metabolismo , Proteínas Represoras/genética , Neovascularización Fisiológica/genética , Diferenciación Celular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismo , Factor A de Crecimiento Endotelial Vascular/genética , Neurogénesis/genética , Glucólisis , AngiogénesisRESUMEN
Chromobox homolog 3 (Cbx3/heterochromatin protein 1γ [HP1γ]) stimulates cell differentiation, but its mechanism is unknown. We found that Cbx3 binds to gene promoters upon differentiation of murine embryonic stem cells (ESCs) to neural progenitor cells (NPCs) and recruits the Mediator subunit Med26. RNAi knockdown of either Cbx3 or Med26 inhibits neural differentiation while up-regulating genes involved in mesodermal lineage decisions. Thus, Cbx3 and Med26 together ensure the fidelity of lineage specification by enhancing the expression of neural genes and down-regulating genes specific to alternative fates.
Asunto(s)
Diferenciación Celular , Linaje de la Célula , Proteínas Cromosómicas no Histona/metabolismo , Células Madre Embrionarias/citología , Regulación de la Expresión Génica , Complejo Mediador/metabolismo , Células-Madre Neurales/citología , Animales , Células Cultivadas , Proteínas Cromosómicas no Histona/antagonistas & inhibidores , Proteínas Cromosómicas no Histona/genética , Quinasa 8 Dependiente de Ciclina/genética , Quinasa 8 Dependiente de Ciclina/metabolismo , Células Madre Embrionarias/metabolismo , Complejo Mediador/genética , Mesodermo/citología , Mesodermo/metabolismo , Ratones , Células-Madre Neurales/metabolismo , Regiones Promotoras Genéticas , ARN Interferente Pequeño/genéticaRESUMEN
In the developing nervous system, neural progenitors exit the cell cycle and differentiate on a precise schedule, yet the mechanisms driving this process remain poorly defined. Yan et al. (2009) now identify a thiol-redox reaction mediated by the membrane protein GDE2 and the peroxiredoxin protein Prdx1 that promotes neurogenesis.
Asunto(s)
Neuronas/citología , Médula Espinal/citología , Animales , Diferenciación Celular , Neurogénesis , Peroxirredoxinas/metabolismo , Hidrolasas Diéster Fosfóricas/metabolismo , Médula Espinal/embriologíaRESUMEN
The oligodendrocyte (OL) lineage transcription factor Olig2 is expressed throughout oligodendroglial development and is essential for oligodendroglial progenitor specification and differentiation. It was previously reported that deletion of Olig2 enhanced the maturation and myelination of immature OLs and accelerated the remyelination process. However, by analyzing multiple Olig2 conditional KO mouse lines (male and female), we conclude that Olig2 has the opposite effect and is required for OL maturation and remyelination. We found that deletion of Olig2 in immature OLs driven by an immature OL-expressing Plp1 promoter resulted in defects in OL maturation and myelination, and did not enhance remyelination after demyelination. Similarly, Olig2 deletion during premyelinating stages in immature OLs using Mobp or Mog promoter-driven Cre lines also did not enhance OL maturation in the CNS. Further, we found that Olig2 was not required for myelin maintenance in mature OLs but was critical for remyelination after lysolecithin-induced demyelinating injury. Analysis of genomic occupancy in immature and mature OLs revealed that Olig2 targets the enhancers of key myelination-related genes for OL maturation from immature OLs. Together, by leveraging multiple immature OL-expressing Cre lines, these studies indicate that Olig2 is essential for differentiation and myelination of immature OLs and myelin repair. Our findings raise fundamental questions about the previously proposed role of Olig2 in opposing OL myelination and highlight the importance of using Cre-dependent reporter(s) for lineage tracing in studying cell state progression.SIGNIFICANCE STATEMENT Identification of the regulators that promote oligodendrocyte (OL) myelination and remyelination is important for promoting myelin repair in devastating demyelinating diseases. Olig2 is expressed throughout OL lineage development. Ablation of Olig2 was reported to induce maturation, myelination, and remyelination from immature OLs. However, lineage-mapping analysis of Olig2-ablated cells was not conducted. Here, by leveraging multiple immature OL-expressing Cre lines, we observed no evidence that Olig2 ablation promotes maturation or remyelination of immature OLs. Instead, we find that Olig2 is required for immature OL maturation, myelination, and myelin repair. These data raise fundamental questions about the proposed inhibitory role of Olig2 against OL maturation and remyelination. Our findings highlight the importance of validating genetic manipulation with cell lineage tracing in studying myelination.
Asunto(s)
Enfermedades Desmielinizantes , Remielinización , Animales , Femenino , Masculino , Ratones , Diferenciación Celular , Enfermedades Desmielinizantes/metabolismo , Vaina de Mielina/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos/genética , Factor de Transcripción 2 de los Oligodendrocitos/metabolismo , Oligodendroglía/metabolismo , Ratones NoqueadosRESUMEN
During tissue development, multipotent progenitors differentiate into specific cell types in characteristic spatial and temporal patterns. We addressed the mechanism linking progenitor identity and differentiation rate in the neural tube, where motor neuron (MN) progenitors differentiate more rapidly than other progenitors. Using single cell transcriptomics, we defined the transcriptional changes associated with the transition of neural progenitors into MNs. Reconstruction of gene expression dynamics from these data indicate a pivotal role for the MN determinant Olig2 just prior to MN differentiation. Olig2 represses expression of the Notch signaling pathway effectors Hes1 and Hes5. Olig2 repression of Hes5 appears to be direct, via a conserved regulatory element within the Hes5 locus that restricts expression from MN progenitors. These findings reveal a tight coupling between the regulatory networks that control patterning and neuronal differentiation and demonstrate how Olig2 acts as the developmental pacemaker coordinating the spatial and temporal pattern of MN generation.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/fisiología , Ciclo Celular/genética , Neuronas Motoras/citología , Neurogénesis/genética , Factor de Transcripción 2 de los Oligodendrocitos/fisiología , Proteínas Represoras/fisiología , Análisis de la Célula Individual , Factor de Transcripción HES-1/fisiología , Transcriptoma , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Colorantes Fluorescentes/metabolismo , Regulación de la Expresión Génica/fisiología , Genes Reporteros , Interneuronas/citología , Ratones Transgénicos , Factor de Transcripción 2 de los Oligodendrocitos/genética , Receptores Notch/metabolismo , Secuencias Reguladoras de Ácidos Nucleicos , Proteínas Represoras/genética , Transducción de Señal , Factor de Transcripción HES-1/genéticaRESUMEN
Fibrils and oligomers are the aggregated protein agents of neuronal dysfunction in ALS diseases. Whereas we now know much about fibril architecture, atomic structures of disease-related oligomers have eluded determination. Here, we determine the corkscrew-like structure of a cytotoxic segment of superoxide dismutase 1 (SOD1) in its oligomeric state. Mutations that prevent formation of this structure eliminate cytotoxicity of the segment in isolation as well as cytotoxicity of the ALS-linked mutants of SOD1 in primary motor neurons and in a Danio rerio (zebrafish) model of ALS. Cytotoxicity assays suggest that toxicity is a property of soluble oligomers, and not large insoluble aggregates. Our work adds to evidence that the toxic oligomeric entities in protein aggregation diseases contain antiparallel, out-of-register ß-sheet structures and identifies a target for structure-based therapeutics in ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral/metabolismo , Superóxido Dismutasa-1/metabolismo , Esclerosis Amiotrófica Lateral/genética , Animales , Cristalografía por Rayos X/métodos , Ratones , Neuronas Motoras/metabolismo , Mutación/genética , Conformación Proteica en Lámina beta , Superóxido Dismutasa-1/genéticaRESUMEN
Orofacial muscles are critical for life-sustaining behaviors, such as feeding and breathing. Centuries of work by neuroanatomists and surgeons resulted in the mapping of bulbar motor neurons in the brainstem and the course of the cranial nerves that carry their axons. Despite the sophisticated understanding of the anatomy of the region, the molecular mechanisms that dictate the development and maturation of facial motor neurons remain poorly understood. This fundamental problem has been recently revisited by physiologists with novel techniques of studying the rhythmic contraction of orofacial muscles in relationship to breathing. The molecular understanding of facial motor neuron development will not only lead to the comprehension of the neural basis of facial expression but may also unlock new avenues to generate stem cell-derived replacements. This review summarizes the current understanding of molecular programs involved in facial motor neuron generation, migration, and maturation, including neural circuit assembly.
Asunto(s)
Nervio Facial/metabolismo , Neuronas Motoras/metabolismo , Neurogénesis/genética , Animales , Tipificación del Cuerpo/genética , Movimiento Celular/genética , Nervio Facial/embriología , Vertebrados/embriología , Vertebrados/metabolismoRESUMEN
Distinct classes of neurons and glial cells in the developing spinal cord arise at specific times and in specific quantities from spatially discrete neural progenitor domains. Thus, adjacent domains can exhibit marked differences in their proliferative potential and timing of differentiation. However, remarkably little is known about the mechanisms that account for this regional control. Here, we show that the transcription factor Promyelocytic Leukemia Zinc Finger (PLZF) plays a critical role shaping patterns of neuronal differentiation by gating the expression of Fibroblast Growth Factor (FGF) Receptor 3 and responsiveness of progenitors to FGFs. PLZF elevation increases FGFR3 expression and STAT3 pathway activity, suppresses neurogenesis, and biases progenitors towards glial cell production. In contrast, PLZF loss reduces FGFR3 levels, leading to premature neuronal differentiation. Together, these findings reveal a novel transcriptional strategy for spatially tuning the responsiveness of distinct neural progenitor groups to broadly distributed mitogenic signals in the embryonic environment.
Asunto(s)
Factores de Crecimiento de Fibroblastos/farmacología , Factores de Transcripción de Tipo Kruppel/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Animales , Astrocitos/citología , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Diferenciación Celular/efectos de los fármacos , Pollos , Epistasis Genética/efectos de los fármacos , Humanos , Interneuronas/citología , Interneuronas/efectos de los fármacos , Interneuronas/metabolismo , Ratones , Células-Madre Neurales/efectos de los fármacos , Neurogénesis/efectos de los fármacos , Neurogénesis/genética , Neuroglía/citología , Neuroglía/metabolismo , Proteína de la Leucemia Promielocítica con Dedos de Zinc , Receptor Tipo 3 de Factor de Crecimiento de Fibroblastos/metabolismo , Proteínas Represoras/metabolismo , Factor de Transcripción STAT3/metabolismo , Médula Espinal/efectos de los fármacos , Médula Espinal/metabolismo , Transcripción Genética/efectos de los fármacosRESUMEN
To date, only the five most posterior groups of Hox genes, Hox9-Hox13, have demonstrated loss-of-function roles in limb patterning. Individual paralog groups control proximodistal patterning of the limb skeletal elements. Hox9 genes also initiate the onset of Hand2 expression in the posterior forelimb compartment, and collectively, the posterior HoxA/D genes maintain posterior Sonic Hedgehog (Shh) expression. Here we show that an anterior Hox paralog group, Hox5, is required for forelimb anterior patterning. Deletion of all three Hox5 genes (Hoxa5, Hoxb5, and Hoxc5) leads to anterior forelimb defects resulting from derepression of Shh expression. The phenotype requires the loss of all three Hox5 genes, demonstrating the high level of redundancy in this Hox paralogous group. Further analyses reveal that Hox5 interacts with promyelocytic leukemia zinc finger biochemically and genetically to restrict Shh expression. These findings, along with previous reports showing that point mutations in the Shh limb enhancer lead to similar anterior limb defects, highlight the importance of Shh repression for proper patterning of the vertebrate limb.
Asunto(s)
Miembro Anterior/embriología , Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas Hedgehog/metabolismo , Factores de Transcripción de Tipo Kruppel/metabolismo , Organogénesis/fisiología , Factores de Transcripción/metabolismo , Animales , Miembro Anterior/metabolismo , Células HEK293 , Humanos , Hibridación in Situ , Ratones , Proteína de la Leucemia Promielocítica con Dedos de Zinc , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
During development, spinal motoneurons (MNs) diversify into a variety of subtypes that are specifically dedicated to the motor control of particular sets of skeletal muscles or visceral organs. MN diversification depends on the coordinated action of several transcriptional regulators including the LIM-HD factor Isl1, which is crucial for MN survival and fate determination. However, how these regulators cooperate to establish each MN subtype remains poorly understood. Here, using phenotypic analyses of single or compound mutant mouse embryos combined with gain-of-function experiments in chick embryonic spinal cord, we demonstrate that the transcriptional activators of the Onecut family critically regulate MN subtype diversification during spinal cord development. We provide evidence that Onecut factors directly stimulate Isl1 expression in specific MN subtypes and are therefore required to maintain Isl1 production at the time of MN diversification. In the absence of Onecut factors, we observed major alterations in MN fate decision characterized by the conversion of somatic to visceral MNs at the thoracic levels of the spinal cord and of medial to lateral MNs in the motor columns that innervate the limbs. Furthermore, we identify Sip1 (Zeb2) as a novel developmental regulator of visceral MN differentiation. Taken together, these data elucidate a comprehensive model wherein Onecut factors control multiple aspects of MN subtype diversification. They also shed light on the late roles of Isl1 in MN fate decision.
Asunto(s)
Diferenciación Celular/fisiología , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas con Homeodominio LIM/metabolismo , Neuronas Motoras/fisiología , Factores de Transcripción Onecut/metabolismo , Médula Espinal/citología , Factores de Transcripción/metabolismo , Animales , Embrión de Pollo , Inmunoprecipitación de Cromatina , Cartilla de ADN/genética , Electroporación , Técnica del Anticuerpo Fluorescente , Regulación del Desarrollo de la Expresión Génica/fisiología , Hibridación in Situ , RatonesRESUMEN
The neural circuits that control motor activities depend on the spatially and temporally ordered generation of distinct classes of spinal interneurons. Despite the importance of these interneurons, the mechanisms underlying their genesis are poorly understood. Here, we demonstrate that the Olig-related transcription factor Bhlhb5 (recently renamed Bhlhe22) plays two central roles in this process. Our findings suggest that Bhlhb5 repressor activity acts downstream of retinoid signaling and homeodomain proteins to promote the formation of dI6, V1 and V2 interneuron progenitors and their differentiated progeny. In addition, Bhlhb5 is required to organize the spatially restricted expression of the Notch ligands and Fringe proteins that both elicit the formation of the interneuron populations that arise adjacent to Bhlhb5(+) cells and influence the global pattern of neuronal differentiation. Through these actions, Bhlhb5 helps transform the spatial information established by morphogen signaling into local cell-cell interactions associated with Notch signaling that control the progression of neurogenesis and extend neuronal diversity within the developing spinal cord.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Interneuronas/fisiología , Receptores Notch/metabolismo , Transducción de Señal/fisiología , Médula Espinal/citología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Diferenciación Celular/fisiología , Embrión de Pollo , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Interneuronas/clasificación , Interneuronas/citología , Neuronas/citología , Neuronas/fisiología , Receptores Notch/genética , Médula Espinal/embriología , Células Madre/citología , Células Madre/fisiologíaRESUMEN
The initiation, execution, and completion of complex locomotor behaviors are depending on precisely integrated neural circuitries consisting of motor pathways that activate muscles in the extremities and sensory afferents that deliver feedback to motoneurons. These projections form in tight temporal and spatial vicinities during development, yet the molecular mechanisms and cues coordinating these processes are not well understood. Using cell-type specific ablation of the axon guidance receptor Neuropilin-1 (Npn-1) in spinal motoneurons or in sensory neurons in the dorsal root ganglia (DRG), we have explored the contribution of this signaling pathway to correct innervation of the limb. We show that Npn-1 controls the fasciculation of both projections and mediates inter-axonal communication. Removal of Npn-1 from sensory neurons results in defasciculation of sensory axons and, surprisingly, also of motor axons. In addition, the tight coupling between these two heterotypic axonal populations is lifted with sensory fibers now leading the spinal nerve projection. These findings are corroborated by partial genetic elimination of sensory neurons, which causes defasciculation of motor projections to the limb. Deletion of Npn-1 from motoneurons leads to severe defasciculation of motor axons in the distal limb and dorsal-ventral pathfinding errors, while outgrowth and fasciculation of sensory trajectories into the limb remain unaffected. Genetic elimination of motoneurons, however, revealed that sensory axons need only minimal scaffolding by motor axons to establish their projections in the distal limb. Thus, motor and sensory axons are mutually dependent on each other for the generation of their trajectories and interact in part through Npn-1-mediated fasciculation before and within the plexus region of the limbs.
Asunto(s)
Axones/metabolismo , Extremidades/inervación , Neuronas Motoras/metabolismo , Neuropilina-1/metabolismo , Células Receptoras Sensoriales/metabolismo , Animales , Fasciculación/metabolismo , Fasciculación/patología , Eliminación de Gen , Integrasas/metabolismo , Ratones , Neuronas Motoras/patología , Células Receptoras Sensoriales/patología , Factores de TiempoRESUMEN
Dorsal interneurons (dIs) in the spinal cord encode the perception of touch, pain, heat, itch, and proprioception. While previous studies using genetic strategies in animal models have revealed important insights into dI development, the molecular details by which dIs arise as distinct populations of neurons remain incomplete. We have developed a resource to investigate dI fate specification by combining a single-cell RNA-Seq atlas of mouse ESC-derived dIs with pseudotime analyses. To validate this in silico resource as a useful tool, we used it to first identify novel genes that are candidates for directing the transition states that lead to distinct dI lineage trajectories, and then validated them using in situ hybridization analyses in the developing mouse spinal cord in vivo . We have also identified a novel endpoint of the dI5 lineage trajectory and found that dIs become more transcriptionally homogenous during terminal differentiation. Together, this study introduces a valuable tool for further discovery about the timing of gene expression during dI differentiation and demonstrates its utility clarifying dI lineage relationships. Summary statement: Pseudotime analyses of embryonic stem cell-derived dorsal spinal interneurons reveals both novel regulators and lineage relationships between different interneuron populations.
RESUMEN
Topographic neuronal maps arise as a consequence of axon trajectory choice correlated with the localisation of neuronal soma, but the identity of the pathways coordinating these processes is unknown. We addressed this question in the context of the myotopic map formed by limb muscles innervated by spinal lateral motor column (LMC) motor axons where the Eph receptor signals specifying growth cone trajectory are restricted by Foxp1 and Lhx1 transcription factors. We show that the localisation of LMC neuron cell bodies can be dissociated from axon trajectory choice by either the loss or gain of function of the Reelin signalling pathway. The response of LMC motor neurons to Reelin is gated by Foxp1- and Lhx1-mediated regulation of expression of the critical Reelin signalling intermediate Dab1. Together, these observations point to identical transcription factors that control motor axon guidance and soma migration and reveal the molecular hierarchy of myotopic organisation.
Asunto(s)
Axones/fisiología , Moléculas de Adhesión Celular Neuronal/metabolismo , Movimiento Celular/fisiología , Proteínas de la Matriz Extracelular/metabolismo , Factores de Transcripción Forkhead/metabolismo , Proteínas de Homeodominio/metabolismo , Neuronas Motoras/fisiología , Proteínas del Tejido Nervioso/metabolismo , Proteínas Represoras/metabolismo , Serina Endopeptidasas/metabolismo , Transducción de Señal , Axones/metabolismo , Moléculas de Adhesión Celular Neuronal/genética , Movimiento Celular/efectos de los fármacos , Proteínas de la Matriz Extracelular/genética , Extremidades/inervación , Factores de Transcripción Forkhead/genética , Regulación del Desarrollo de la Expresión Génica , Conos de Crecimiento/metabolismo , Proteínas de Homeodominio/genética , Proteínas con Homeodominio LIM , Neuronas Motoras/metabolismo , Proteínas del Tejido Nervioso/genética , Proteína Reelina , Proteínas Represoras/genética , Serina Endopeptidasas/genética , Médula Espinal/metabolismo , Factores de TranscripciónRESUMEN
Morphogens are secreted signalling molecules that act in a graded manner to control the pattern of cellular differentiation in developing tissues. An example is Sonic hedgehog (Shh), which acts in several developing vertebrate tissues, including the central nervous system, to provide positional information during embryonic patterning. Here we address how Shh signalling assigns the positional identities of distinct neuronal subtype progenitors throughout the ventral neural tube. Assays of intracellular signal transduction and gene expression indicate that the duration as well as level of signalling is critical for morphogen interpretation. Progenitors of the ventral neuronal subtypes are established sequentially, with progressively more ventral identities requiring correspondingly higher levels and longer periods of Shh signalling. Moreover, cells remain sensitive to changes in Shh signalling for an extended time, reverting to antecedent identities if signalling levels fall below a threshold. Thus, the duration of signalling is important not only for the assignment but also for the refinement and maintenance of positional identity. Together the data suggest a dynamic model for ventral neural tube patterning in which positional information corresponds to the time integral of Shh signalling. This suggests an alternative to conventional models of morphogen action that rely solely on the level of signalling.
Asunto(s)
Proteínas Hedgehog/fisiología , Tubo Neural/embriología , Vertebrados/embriología , Animales , Proteínas Hedgehog/metabolismo , Transducción de SeñalRESUMEN
Morphogens act in developing tissues to control the spatial arrangement of cellular differentiation. The activity of a morphogen has generally been viewed as a concentration-dependent response to a diffusible signal, but the duration of morphogen signalling can also affect cellular responses. One such example is the morphogen sonic hedgehog (SHH). In the vertebrate central nervous system and limbs, the pattern of cellular differentiation is controlled by both the amount and the time of SHH exposure. How these two parameters are interpreted at a cellular level has been unclear. Here we provide evidence that changing the concentration or duration of SHH has an equivalent effect on intracellular signalling. Chick neural cells convert different concentrations of SHH into time-limited periods of signal transduction, such that signal duration is proportional to SHH concentration. This depends on the gradual desensitization of cells to ongoing SHH exposure, mediated by the SHH-dependent upregulation of patched 1 (PTC1), a ligand-binding inhibitor of SHH signalling. Thus, in addition to its role in shaping the SHH gradient, PTC1 participates cell autonomously in gradient sensing. Together, the data reveal a novel strategy for morphogen interpretation, in which the temporal adaptation of cells to a morphogen integrates the concentration and duration of a signal to control differential gene expression.
Asunto(s)
Proteínas Hedgehog/metabolismo , Transducción de Señal , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Embrión de Pollo , Regulación de la Expresión Génica/efectos de los fármacos , Proteínas Hedgehog/farmacología , Proteína Homeobox Nkx-2.2 , Proteínas de Homeodominio/metabolismo , Ratones , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Tubo Neural/citología , Tubo Neural/efectos de los fármacos , Tubo Neural/embriología , Tubo Neural/metabolismo , Factor de Transcripción 2 de los Oligodendrocitos , Proteínas Oncogénicas/metabolismo , Factor de Transcripción PAX7/metabolismo , Receptores Patched , Receptor Patched-1 , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Transducción de Señal/efectos de los fármacos , Factores de Tiempo , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Proteínas de Pez Cebra , Proteína con Dedos de Zinc GLI1RESUMEN
Embryonic neural stem cells (NSCs, i.e., radial glia) in the ventricular-subventricular zone (V-SVZ) generate the majority of neurons and glia in the forebrain. Postnatally, embryonic radial glia disappear and a subpopulation of radial glia transition into adult NSCs. As this transition occurs, widespread neurogenesis in brain regions such as the cerebral cortex ends. The mechanisms that regulate the postnatal disappearance of radial glia and the ending of embryonic neurogenesis remain poorly understood. Here, we show that PR domain-containing 16 (Prdm16) promotes the disappearance of radial glia and the ending of neurogenesis in the cerebral cortex. Genetic deletion of Prdm16 from NSCs leads to the persistence of radial glia in the adult V-SVZ and prolonged postnatal cortical neurogenesis. Mechanistically, Prdm16 induces the postnatal reduction in Vascular Cell Adhesion Molecule 1 (Vcam1). The postnatal disappearance of radial glia and the ending of cortical neurogenesis occur normally in Prdm16-Vcam1 double conditional knockout mice. These observations reveal novel molecular regulators of the postnatal disappearance of radial glia and the ending of embryonic neurogenesis, filling a key knowledge gap in NSC biology.