RESUMO
Comparative studies of great apes provide a window into our evolutionary past, but the extent and identity of cellular differences that emerged during hominin evolution remain largely unexplored. We established a comparative loss-of-function approach to evaluate whether human cells exhibit distinct genetic dependencies. By performing genome-wide CRISPR interference screens in human and chimpanzee pluripotent stem cells, we identified 75 genes with species-specific effects on cellular proliferation. These genes comprised coherent processes, including cell-cycle progression and lysosomal signaling, which we determined to be human-derived by comparison with orangutan cells. Human-specific robustness to CDK2 and CCNE1 depletion persisted in neural progenitor cells and cerebral organoids, supporting the G1-phase length hypothesis as a potential evolutionary mechanism in human brain expansion. Our findings demonstrate that evolutionary changes in human cells reshaped the landscape of essential genes and establish a platform for systematically uncovering latent cellular and molecular differences between species.
Assuntos
Hominidae , Células-Tronco Neurais , Células-Tronco Pluripotentes , Células-Tronco , Animais , Humanos , Pan troglodytes/genéticaRESUMO
The human brain undergoes rapid development at mid-gestation from a pool of neural stem and progenitor cells (NSPCs) that give rise to the neurons, oligodendrocytes, and astrocytes of the mature brain. Functional study of these cell types has been hampered by a lack of precise purification methods. We describe a method for prospectively isolating ten distinct NSPC types from the developing human brain using cell-surface markers. CD24-THY1-/lo cells were enriched for radial glia, which robustly engrafted and differentiated into all three neural lineages in the mouse brain. THY1hi cells marked unipotent oligodendrocyte precursors committed to an oligodendroglial fate, and CD24+THY1-/lo cells marked committed excitatory and inhibitory neuronal lineages. Notably, we identify and functionally characterize a transcriptomically distinct THY1hiEGFRhiPDGFRA- bipotent glial progenitor cell (GPC), which is lineage-restricted to astrocytes and oligodendrocytes, but not to neurons. Our study provides a framework for the functional study of distinct cell types in human neurodevelopment.
Assuntos
Células-Tronco Neurais , Camundongos , Animais , Humanos , Células-Tronco Neurais/metabolismo , Neurônios , Diferenciação Celular/fisiologia , Neuroglia/metabolismo , Encéfalo , AstrócitosRESUMO
Neural stem cells (NSCs) persist in the adult mammalian brain and are able to give rise to new neurons and glia throughout life. The largest stem cell niche in the adult mouse brain is the ventricular-subventricular zone (V-SVZ) lining the lateral ventricles. Adult NSCs in the V-SVZ coexist in quiescent and actively proliferating states, and they exhibit a regionalized molecular identity. The importance of such spatial diversity is just emerging, as depending on their position within the niche, adult NSCs give rise to distinct subtypes of olfactory bulb interneurons and different types of glia. However, the functional relevance of stem cell heterogeneity in the V-SVZ is still poorly understood. Here, we put into perspective findings highlighting the importance of adult NSC diversity for brain plasticity, and how the body signals to brain stem cells in different physiological states to regulate their behavior.
Assuntos
Células-Tronco Adultas , Células-Tronco Neurais , Nicho de Células-Tronco , Animais , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Adultas/citologia , Células-Tronco Adultas/metabolismo , Humanos , Ventrículos Laterais/citologia , Neurogênese , Bulbo Olfatório/citologia , Bulbo Olfatório/metabolismo , Camundongos , Encéfalo/citologia , Diferenciação CelularRESUMO
Neural stem cells (NSCs) in the adult brain transit from the quiescent state to proliferation to produce new neurons. The mechanisms regulating this transition in freely behaving animals are, however, poorly understood. We customized in vivo imaging protocols to follow NSCs for several days up to months, observing their activation kinetics in freely behaving mice. Strikingly, NSC division is more frequent during daylight and is inhibited by darkness-induced melatonin signaling. The inhibition of melatonin receptors affected intracellular Ca2+ dynamics and promoted NSC activation. We further discovered a Ca2+ signature of quiescent versus activated NSCs and showed that several microenvironmental signals converge on intracellular Ca2+ pathways to regulate NSC quiescence and activation. In vivo NSC-specific optogenetic modulation of Ca2+ fluxes to mimic quiescent-state-like Ca2+ dynamics in freely behaving mice blocked NSC activation and maintained their quiescence, pointing to the regulatory mechanisms mediating NSC activation in freely behaving animals.
Assuntos
Células-Tronco Adultas/metabolismo , Cálcio/metabolismo , Ritmo Circadiano , Espaço Intracelular/metabolismo , Células-Tronco Neurais/metabolismo , Células-Tronco Adultas/citologia , Células-Tronco Adultas/efeitos dos fármacos , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Comportamento Animal/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ritmo Circadiano/efeitos dos fármacos , Citosol/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Melatonina/metabolismo , Camundongos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/efeitos dos fármacos , Optogenética , Transdução de Sinais/efeitos dos fármacos , Triptaminas/farmacologiaRESUMO
Although oncogenic mutations predispose tissue stem cells to tumor initiation, the rate-limiting processes for stem cell immortalization remain unknown. In this issue of Cell, Bonnay et al. identify enhanced electron transport chain activity as a critical determinant of this process, establishing metabolic reprogramming as limiting for tumor initiation.
Assuntos
Transformação Celular Neoplásica , Células-Tronco Neurais , Carcinogênese , Humanos , Estresse OxidativoRESUMO
Metabolic reprogramming is a key feature of many cancers, but how and when it contributes to tumorigenesis remains unclear. Here we demonstrate that metabolic reprogramming induced by mitochondrial fusion can be rate-limiting for immortalization of tumor-initiating cells (TICs) and trigger their irreversible dedication to tumorigenesis. Using single-cell transcriptomics, we find that Drosophila brain tumors contain a rapidly dividing stem cell population defined by upregulation of oxidative phosphorylation (OxPhos). We combine targeted metabolomics and in vivo genetic screening to demonstrate that OxPhos is required for tumor cell immortalization but dispensable in neural stem cells (NSCs) giving rise to tumors. Employing an in vivo NADH/NAD+ sensor, we show that NSCs precisely increase OxPhos during immortalization. Blocking OxPhos or mitochondrial fusion stalls TICs in quiescence and prevents tumorigenesis through impaired NAD+ regeneration. Our work establishes a unique connection between cellular metabolism and immortalization of tumor-initiating cells.
Assuntos
Neoplasias Encefálicas/metabolismo , Carcinogênese/metabolismo , Transformação Celular Neoplásica/metabolismo , Dinâmica Mitocondrial , NAD/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neurais/metabolismo , Fosforilação Oxidativa , Animais , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Carcinogênese/genética , Carcinogênese/patologia , Transformação Celular Neoplásica/patologia , Ciclo do Ácido Cítrico/genética , Biologia Computacional , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Drosophila , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Glicólise/genética , Espectrometria de Massas , Metabolômica , Microscopia Eletrônica de Transmissão , Família Multigênica , Células-Tronco Neurais/patologia , Consumo de Oxigênio/genética , Interferência de RNA , Espécies Reativas de Oxigênio/metabolismo , Análise de Célula Única , Transcriptoma/genéticaRESUMO
Histone H3.3 glycine 34 to arginine/valine (G34R/V) mutations drive deadly gliomas and show exquisite regional and temporal specificity, suggesting a developmental context permissive to their effects. Here we show that 50% of G34R/V tumors (n = 95) bear activating PDGFRA mutations that display strong selection pressure at recurrence. Although considered gliomas, G34R/V tumors actually arise in GSX2/DLX-expressing interneuron progenitors, where G34R/V mutations impair neuronal differentiation. The lineage of origin may facilitate PDGFRA co-option through a chromatin loop connecting PDGFRA to GSX2 regulatory elements, promoting PDGFRA overexpression and mutation. At the single-cell level, G34R/V tumors harbor dual neuronal/astroglial identity and lack oligodendroglial programs, actively repressed by GSX2/DLX-mediated cell fate specification. G34R/V may become dispensable for tumor maintenance, whereas mutant-PDGFRA is potently oncogenic. Collectively, our results open novel research avenues in deadly tumors. G34R/V gliomas are neuronal malignancies where interneuron progenitors are stalled in differentiation by G34R/V mutations and malignant gliogenesis is promoted by co-option of a potentially targetable pathway, PDGFRA signaling.
Assuntos
Neoplasias Encefálicas/genética , Carcinogênese/genética , Glioma/genética , Histonas/genética , Interneurônios/metabolismo , Mutação/genética , Células-Tronco Neurais/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Neoplasias Encefálicas/patologia , Carcinogênese/patologia , Linhagem da Célula , Reprogramação Celular/genética , Cromatina/metabolismo , Embrião de Mamíferos/metabolismo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Glioma/patologia , Histonas/metabolismo , Lisina/metabolismo , Camundongos Endogâmicos C57BL , Modelos Biológicos , Gradação de Tumores , Oligodendroglia/metabolismo , Regiões Promotoras Genéticas/genética , Prosencéfalo/embriologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transcrição Gênica , Transcriptoma/genéticaRESUMO
The function of somatic stem cells declines with age. Understanding the molecular underpinnings of this decline is key to counteract age-related disease. Here, we report a dramatic drop in the neural stem cells (NSCs) number in the aging murine brain. We find that this smaller stem cell reservoir is protected from full depletion by an increase in quiescence that makes old NSCs more resistant to regenerate the injured brain. Once activated, however, young and old NSCs show similar proliferation and differentiation capacity. Single-cell transcriptomics of NSCs indicate that aging changes NSCs minimally. In the aging brain, niche-derived inflammatory signals and the Wnt antagonist sFRP5 induce quiescence. Indeed, intervention to neutralize them increases activation of old NSCs during homeostasis and following injury. Our study identifies quiescence as a key feature of old NSCs imposed by the niche and uncovers ways to activate NSCs to repair the aging brain.
Assuntos
Encéfalo/fisiologia , Fatores Etários , Animais , Encéfalo/citologia , Diferenciação Celular/fisiologia , Divisão Celular/fisiologia , Proliferação de Células/fisiologia , Senescência Celular/fisiologia , Homeostase , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Regeneração Nervosa , Células-Tronco Neurais/citologia , Células-Tronco Neurais/fisiologia , Neurogênese , Nicho de Células-TroncoRESUMO
New neurons arise from quiescent adult neural progenitors throughout life in specific regions of the mammalian brain. Little is known about the embryonic origin and establishment of adult neural progenitors. Here, we show that Hopx+ precursors in the mouse dentate neuroepithelium at embryonic day 11.5 give rise to proliferative Hopx+ neural progenitors in the primitive dentate region, and they, in turn, generate granule neurons, but not other neurons, throughout development and then transition into Hopx+ quiescent radial glial-like neural progenitors during an early postnatal period. RNA-seq and ATAC-seq analyses of Hopx+ embryonic, early postnatal, and adult dentate neural progenitors further reveal common molecular and epigenetic signatures and developmental dynamics. Together, our findings support a "continuous" model wherein a common neural progenitor population exclusively contributes to dentate neurogenesis throughout development and adulthood. Adult dentate neurogenesis may therefore represent a lifelong extension of development that maintains heightened plasticity in the mammalian hippocampus.
Assuntos
Células-Tronco Embrionárias/metabolismo , Neurogênese , Animais , Diferenciação Celular , Giro Denteado/metabolismo , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias/citologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Hipocampo/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismoRESUMO
In situ transgenesis methods such as viruses and electroporation can rapidly create somatic transgenic mice but lack control over copy number, zygosity, and locus specificity. Here we establish mosaic analysis by dual recombinase-mediated cassette exchange (MADR), which permits stable labeling of mutant cells expressing transgenic elements from precisely defined chromosomal loci. We provide a toolkit of MADR elements for combination labeling, inducible and reversible transgene manipulation, VCre recombinase expression, and transgenesis of human cells. Further, we demonstrate the versatility of MADR by creating glioma models with mixed reporter-identified zygosity or with "personalized" driver mutations from pediatric glioma. MADR is extensible to thousands of existing mouse lines, providing a flexible platform to democratize the generation of somatic mosaic mice. VIDEO ABSTRACT.
Assuntos
Neoplasias Encefálicas/genética , Modelos Animais de Doenças , Marcação de Genes/métodos , Loci Gênicos/genética , Glioma/genética , Mutagênese Insercional/métodos , Transgenes/genética , Animais , Linhagem Celular Tumoral , Feminino , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células-Tronco Neurais/metabolismo , Recombinases/metabolismo , TransfecçãoRESUMO
Neural induction in vertebrates generates a CNS that extends the rostral-caudal length of the body. The prevailing view is that neural cells are initially induced with anterior (forebrain) identity; caudalizing signals then convert a proportion to posterior fates (spinal cord). To test this model, we used chromatin accessibility to define how cells adopt region-specific neural fates. Together with genetic and biochemical perturbations, this identified a developmental time window in which genome-wide chromatin-remodeling events preconfigure epiblast cells for neural induction. Contrary to the established model, this revealed that cells commit to a regional identity before acquiring neural identity. This "primary regionalization" allocates cells to anterior or posterior regions of the nervous system, explaining how cranial and spinal neurons are generated at appropriate axial positions. These findings prompt a revision to models of neural induction and support the proposed dual evolutionary origin of the vertebrate CNS.
Assuntos
Montagem e Desmontagem da Cromatina , Indução Embrionária , Neurogênese , Animais , Linhagem Celular , Células Cultivadas , Embrião de Galinha , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Medula Espinal/citologia , Medula Espinal/crescimento & desenvolvimento , Medula Espinal/metabolismoRESUMO
The development of interventions to prevent congenital Zika syndrome (CZS) has been limited by the lack of an established nonhuman primate model. Here we show that infection of female rhesus monkeys early in pregnancy with Zika virus (ZIKV) recapitulates many features of CZS in humans. We infected 9 pregnant monkeys with ZIKV, 6 early in pregnancy (weeks 6-7 of gestation) and 3 later in pregnancy (weeks 12-14 of gestation), and compared findings with uninfected controls. 100% (6 of 6) of monkeys infected early in pregnancy exhibited prolonged maternal viremia and fetal neuropathology, including fetal loss, smaller brain size, and histopathologic brain lesions, including microcalcifications, hemorrhage, necrosis, vasculitis, gliosis, and apoptosis of neuroprogenitor cells. High-resolution MRI demonstrated concordant lesions indicative of deep gray matter injury. We also observed spinal, ocular, and neuromuscular pathology. Our data show that vascular compromise and neuroprogenitor cell dysfunction are hallmarks of CZS pathogenesis, suggesting novel strategies to prevent and to treat this disease.
Assuntos
Feto/virologia , Neurônios/patologia , Infecção por Zika virus/patologia , Zika virus/patogenicidade , Animais , Animais Recém-Nascidos , Apoptose , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Calcinose/patologia , Calcinose/veterinária , Feminino , Idade Gestacional , Macaca mulatta , Imageamento por Ressonância Magnética , Necrose , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/virologia , Neurônios/virologia , Gravidez , Índice de Gravidade de Doença , Vasculite/patologia , Vasculite/veterinária , Infecção por Zika virus/veterinária , Infecção por Zika virus/virologiaRESUMO
Type I interferon (IFN) is produced when host sensors detect foreign nucleic acids, but how sensors differentiate self from nonself nucleic acids, such as double-stranded RNA (dsRNA), is incompletely understood. Mutations in ADAR1, an adenosine-to-inosine editing enzyme of dsRNA, cause Aicardi-Goutières syndrome, an autoinflammatory disorder associated with spontaneous interferon production and neurologic sequelae. We generated ADAR1 knockout human cells to explore ADAR1 substrates and function. ADAR1 primarily edited Alu elements in RNA polymerase II (pol II)-transcribed mRNAs, but not putative pol III-transcribed Alus. During the IFN response, ADAR1 blocked translational shutdown by inhibiting hyperactivation of PKR, a dsRNA sensor. ADAR1 dsRNA binding and catalytic activities were required to fully prevent endogenous RNA from activating PKR. Remarkably, ADAR1 knockout neuronal progenitor cells exhibited MDA5 (dsRNA sensor)-dependent spontaneous interferon production, PKR activation, and cell death. Thus, human ADAR1 regulates sensing of self versus nonself RNA, allowing pathogen detection while avoiding autoinflammation.
Assuntos
Adenosina Desaminase/metabolismo , Elementos Alu , Doenças Autoimunes do Sistema Nervoso/metabolismo , Malformações do Sistema Nervoso/metabolismo , Células-Tronco Neurais/metabolismo , Biossíntese de Proteínas , RNA de Cadeia Dupla/metabolismo , Proteínas de Ligação a RNA/metabolismo , Adenosina Desaminase/genética , Adenosina Desaminase/imunologia , Doenças Autoimunes do Sistema Nervoso/genética , Doenças Autoimunes do Sistema Nervoso/imunologia , Morte Celular/genética , Morte Celular/imunologia , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Inflamação/genética , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Helicase IFIH1 Induzida por Interferon/genética , Helicase IFIH1 Induzida por Interferon/imunologia , Helicase IFIH1 Induzida por Interferon/metabolismo , Malformações do Sistema Nervoso/genética , Malformações do Sistema Nervoso/imunologia , Células-Tronco Neurais/citologia , Células-Tronco Neurais/imunologia , Células-Tronco Neurais/patologia , RNA Polimerase II/genética , RNA Polimerase II/imunologia , RNA Polimerase II/metabolismo , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/imunologia , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/imunologia , eIF-2 Quinase/genética , eIF-2 Quinase/imunologia , eIF-2 Quinase/metabolismoRESUMO
Many patients with advanced cancers achieve dramatic responses to a panoply of therapeutics yet retain minimal residual disease (MRD), which ultimately results in relapse. To gain insights into the biology of MRD, we applied single-cell RNA sequencing to malignant cells isolated from BRAF mutant patient-derived xenograft melanoma cohorts exposed to concurrent RAF/MEK-inhibition. We identified distinct drug-tolerant transcriptional states, varying combinations of which co-occurred within MRDs from PDXs and biopsies of patients on treatment. One of these exhibited a neural crest stem cell (NCSC) transcriptional program largely driven by the nuclear receptor RXRG. An RXR antagonist mitigated accumulation of NCSCs in MRD and delayed the development of resistance. These data identify NCSCs as key drivers of resistance and illustrate the therapeutic potential of MRD-directed therapy. They also highlight how gene regulatory network architecture reprogramming may be therapeutically exploited to limit cellular heterogeneity, a key driver of disease progression and therapy resistance.
Assuntos
Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Melanoma/tratamento farmacológico , Neoplasia Residual/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neurais/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Receptor X Retinoide gama/antagonistas & inibidores , Animais , Biomarcadores Tumorais , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Humanos , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 1/genética , Masculino , Melanoma/metabolismo , Melanoma/patologia , Camundongos SCID , Mutação , Neoplasia Residual/metabolismo , Neoplasia Residual/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/patologia , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The cerebral cortex underwent rapid expansion and increased complexity during recent hominid evolution. Gene duplications constitute a major evolutionary force, but their impact on human brain development remains unclear. Using tailored RNA sequencing (RNA-seq), we profiled the spatial and temporal expression of hominid-specific duplicated (HS) genes in the human fetal cortex and identified a repertoire of 35 HS genes displaying robust and dynamic patterns during cortical neurogenesis. Among them NOTCH2NL, human-specific paralogs of the NOTCH2 receptor, stood out for their ability to promote cortical progenitor maintenance. NOTCH2NL promote the clonal expansion of human cortical progenitors, ultimately leading to higher neuronal output. At the molecular level, NOTCH2NL function by activating the Notch pathway through inhibition of cis Delta/Notch interactions. Our study uncovers a large repertoire of recently evolved genes active during human corticogenesis and reveals how human-specific NOTCH paralogs may have contributed to the expansion of the human cortex.
Assuntos
Córtex Cerebral/metabolismo , Regulação da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Membrana/metabolismo , Neurogênese , Neurônios/metabolismo , Receptor Notch2/genética , Sequência de Aminoácidos , Proteínas de Ligação ao Cálcio , Diferenciação Celular/genética , Análise por Conglomerados , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Hibridização In Situ , Células-Tronco Neurais/metabolismo , Transdução de SinaisRESUMO
Ependymal cells are multi-ciliated cells that form the brain's ventricular epithelium and a niche for neural stem cells (NSCs) in the ventricular-subventricular zone (V-SVZ). In addition, ependymal cells are suggested to be latent NSCs with a capacity to acquire neurogenic function. This remains highly controversial due to a lack of prospective in vivo labeling techniques that can effectively distinguish ependymal cells from neighboring V-SVZ NSCs. We describe a transgenic system that allows for targeted labeling of ependymal cells within the V-SVZ. Single-cell RNA-seq revealed that ependymal cells are enriched for cilia-related genes and share several stem-cell-associated genes with neural stem or progenitors. Under in vivo and in vitro neural-stem- or progenitor-stimulating environments, ependymal cells failed to demonstrate any suggestion of latent neural-stem-cell function. These findings suggest remarkable stability of ependymal cell function and provide fundamental insights into the molecular signature of the V-SVZ niche.
Assuntos
Epêndima/metabolismo , Genômica , Actinas/genética , Actinas/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Epêndima/citologia , Epêndima/efeitos dos fármacos , Feminino , Fator 2 de Crescimento de Fibroblastos/farmacologia , Ventrículos Laterais/citologia , Ventrículos Laterais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Análise de Célula Única , Nicho de Células-Tronco , Transcriptoma , Fator A de Crescimento do Endotélio Vascular/farmacologia , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
During corticogenesis, ventricular zone progenitors sequentially generate distinct subtypes of neurons, accounting for the diversity of neocortical cells and the circuits they form. While activity-dependent processes are critical for the differentiation and circuit assembly of postmitotic neurons, how bioelectrical processes affect nonexcitable cells, such as progenitors, remains largely unknown. Here, we reveal that, in the developing mouse neocortex, ventricular zone progenitors become more hyperpolarized as they generate successive subtypes of neurons. Experimental in vivo hyperpolarization shifted the transcriptional programs and division modes of these progenitors to a later developmental status, with precocious generation of intermediate progenitors and a forward shift in the laminar, molecular, morphological, and circuit features of their neuronal progeny. These effects occurred through inhibition of the Wnt-beta-catenin signaling pathway by hyperpolarization. Thus, during corticogenesis, bioelectric membrane properties are permissive for specific molecular pathways to coordinate the temporal progression of progenitor developmental programs and thus neocortical neuron diversity.
Assuntos
Potenciais da Membrana , Neocórtex/embriologia , Neurônios/metabolismo , Células-Tronco/citologia , Animais , Encéfalo/citologia , Encéfalo/embriologia , Diferenciação Celular , Progressão da Doença , Eletroporação , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos , Neocórtex/citologia , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/citologia , Neurogênese , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Análise de Sequência de RNA , Transdução de Sinais , Fatores de Tempo , Proteínas Wnt/metabolismo , beta Catenina/metabolismoRESUMO
Genetic changes causing brain size expansion in human evolution have remained elusive. Notch signaling is essential for radial glia stem cell proliferation and is a determinant of neuronal number in the mammalian cortex. We find that three paralogs of human-specific NOTCH2NL are highly expressed in radial glia. Functional analysis reveals that different alleles of NOTCH2NL have varying potencies to enhance Notch signaling by interacting directly with NOTCH receptors. Consistent with a role in Notch signaling, NOTCH2NL ectopic expression delays differentiation of neuronal progenitors, while deletion accelerates differentiation into cortical neurons. Furthermore, NOTCH2NL genes provide the breakpoints in 1q21.1 distal deletion/duplication syndrome, where duplications are associated with macrocephaly and autism and deletions with microcephaly and schizophrenia. Thus, the emergence of human-specific NOTCH2NL genes may have contributed to the rapid evolution of the larger human neocortex, accompanied by loss of genomic stability at the 1q21.1 locus and resulting recurrent neurodevelopmental disorders.
Assuntos
Encéfalo/embriologia , Córtex Cerebral/fisiologia , Neurogênese/fisiologia , Receptor Notch2/metabolismo , Transdução de Sinais , Animais , Diferenciação Celular , Células-Tronco Embrionárias/metabolismo , Feminino , Deleção de Genes , Genes Reporter , Gorilla gorilla , Células HEK293 , Humanos , Neocórtex/citologia , Células-Tronco Neurais/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Pan troglodytes , Receptor Notch2/genética , Análise de Sequência de RNARESUMO
Cerebral cortex size differs dramatically between reptiles, birds, and mammals, owing to developmental differences in neuron production. In mammals, signaling pathways regulating neurogenesis have been identified, but genetic differences behind their evolution across amniotes remain unknown. We show that direct neurogenesis from radial glia cells, with limited neuron production, dominates the avian, reptilian, and mammalian paleocortex, whereas in the evolutionarily recent mammalian neocortex, most neurogenesis is indirect via basal progenitors. Gain- and loss-of-function experiments in mouse, chick, and snake embryos and in human cerebral organoids demonstrate that high Slit/Robo and low Dll1 signaling, via Jag1 and Jag2, are necessary and sufficient to drive direct neurogenesis. Attenuating Robo signaling and enhancing Dll1 in snakes and birds recapitulates the formation of basal progenitors and promotes indirect neurogenesis. Our study identifies modulation in activity levels of conserved signaling pathways as a primary mechanism driving the expansion and increased complexity of the mammalian neocortex during amniote evolution.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurogênese/genética , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo , Animais , Proteínas de Ligação ao Cálcio , Córtex Cerebral/metabolismo , Embrião de Galinha , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Homeodomínio , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteína Jagged-1 , Proteína Jagged-2 , Mamíferos/embriologia , Camundongos , Camundongos Endogâmicos C57BL , Neocórtex/fisiologia , Células-Tronco Neurais , Neurogênese/fisiologia , Neuroglia/fisiologia , Neurônios , Fator de Transcrição PAX6/metabolismo , Proteínas Repressoras , Transdução de Sinais , Serpentes/embriologia , Proteínas RoundaboutRESUMO
X-linked Dystonia-Parkinsonism (XDP) is a Mendelian neurodegenerative disease that is endemic to the Philippines and is associated with a founder haplotype. We integrated multiple genome and transcriptome assembly technologies to narrow the causal mutation to the TAF1 locus, which included a SINE-VNTR-Alu (SVA) retrotransposition into intron 32 of the gene. Transcriptome analyses identified decreased expression of the canonical cTAF1 transcript among XDP probands, and de novo assembly across multiple pluripotent stem-cell-derived neuronal lineages discovered aberrant TAF1 transcription that involved alternative splicing and intron retention (IR) in proximity to the SVA that was anti-correlated with overall TAF1 expression. CRISPR/Cas9 excision of the SVA rescued this XDP-specific transcriptional signature and normalized TAF1 expression in probands. These data suggest an SVA-mediated aberrant transcriptional mechanism associated with XDP and may provide a roadmap for layered technologies and integrated assembly-based analyses for other unsolved Mendelian disorders.