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Neural stem cell heterogeneity and adult neuronal diversity
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RESUMO: Uma vasta diversidade celular pode ser observada no sistema nervoso animal. Na mosca da fruta
são os neuroblastos (NB), células estaminais multipotentes, que geram todos os neurónios do
indivíduo adulto. Estes dividem-se assimetricamente gerando células filhas que de seguida se
dividem originando neurónios ou células da glia. Há mais de 30 anos que as linhagens de NBs são
estudadas como modelo para entender os mecanismos moleculares básicos por detrás do
desenvolvimento neuronal. Estudos mostram que diferentes genes ligados à especificação celular
podem ser diferencialmente expressos entre as variadas linhagens. Como resultado, diferentes
subtipos neuronais são então gerados a partir de cada NB. No entanto, ainda não temos um
entendimento global de todas as instruções genéticas necessárias em cada linhagem para a
construção do sistema nervoso central do adulto. De modo a contribuirmos para o seu estudo,
recentemente o nosso laboratório revisitou a especificação da diversidade neuronal utilizando o
método de sequenciação de célula-única, um método que permite a identificação de
heterogeneidade celular. Neste trabalho, dados de sequenciação de célula-única de linhagens de
NBs foram analisados com recurso a métodos bioinformáticos. Isto levou à identificação de todos
os tipos celulares característicos das linhagens de NBs. Através da comparação dos perfis de
expressão das várias categorias celulares, foi possível identificar múltiplos genes que se
encontravam diferencialmente expressos ao longo da diferenciação das linhagens de NBs. Incluídos
estavam vários genes conhecidos como estando envolvidos na especificação das linhagens de NBs,
mas também alguns genes que ainda não foram estudados neste contexto. Além disto, também
observámos uma notável heterogeneidade entre NBs e neurónios. Curiosamente, vários genes que
foram identificados como estando diferencialmente expressos entre os NBs da larva, também
apresentam este padrão no embrião. Isto apoia a ideia de que a identidade e os programas
transcricionais das várias linhagens são conservados desde a fase de neurogénese do embrião até
à larva. Por último, a expressão de alguns genes marcadores que tinham sido identificados como
estando heterogeneamente expressos foi também validada in vivo. No geral, este trabalho poderá
providenciar estudos futuros um mapa dos genes potencialmente envolvidos no desenvolvimento
de linhagens de NBs e na criação da diversidade neuronal.
ABSTRACT: Vast cellular diversity can be observed in the animal nervous system. In the fruit fly, multipotent neural stem cells, neuroblasts (NB), represent the fundamental unit for the production of all neurons of the adult. These divide asymmetrically to give rise to daughter cells which then produce neurons or glia. For more than 30 years, Drosophila NB lineages have been a powerful model for neurodevelopmental studies, which has allowed the identification of conserved basic molecular mechanisms. Previous work has shown that various fate determinant genes are differentially expressed in between lineages, which results in the production of diverse sets of neurons by each NB in a stereotyped manner. However, we still lack a global understanding of all genetic instructions necessary in each lineage for the construction of the adult central nervous system. To try to contribute to this ultimate goal, our lab has revisited the specification of neuronal diversity through single-cell technologies, a powerful method to identify cellular heterogeneity. Specifically, in this work, we have curated a dataset of single-cell sequencing of larval NB lineages through bioinformatic methods. This allowed the identification of populations of cells that seem to match all the NB lineage cell types. By comparing the transcriptomic profiles of these cell types, we have identified multiple genes that become differentially expressed along NB lineage development. This included known NB lineage fate regulators, but also unstudied genes within this context. Furthermore, we have observed notable expression heterogeneity within NBs and neurons. Interestingly, we found that multiple genes differentially expressed between larval NBs show the same expression pattern in embryonic NBs. This supports the idea that NB lineage identity programs are conserved from embryo to larval neurogenesis. Finally, we have also validated in vivo the expression of a few marker genes which were found to be distinctively expressed among NBs. Overall, this work should provide future studies with a useful molecular blueprint of players potentially driving NB lineage development and the generation of neuronal diversity.
ABSTRACT: Vast cellular diversity can be observed in the animal nervous system. In the fruit fly, multipotent neural stem cells, neuroblasts (NB), represent the fundamental unit for the production of all neurons of the adult. These divide asymmetrically to give rise to daughter cells which then produce neurons or glia. For more than 30 years, Drosophila NB lineages have been a powerful model for neurodevelopmental studies, which has allowed the identification of conserved basic molecular mechanisms. Previous work has shown that various fate determinant genes are differentially expressed in between lineages, which results in the production of diverse sets of neurons by each NB in a stereotyped manner. However, we still lack a global understanding of all genetic instructions necessary in each lineage for the construction of the adult central nervous system. To try to contribute to this ultimate goal, our lab has revisited the specification of neuronal diversity through single-cell technologies, a powerful method to identify cellular heterogeneity. Specifically, in this work, we have curated a dataset of single-cell sequencing of larval NB lineages through bioinformatic methods. This allowed the identification of populations of cells that seem to match all the NB lineage cell types. By comparing the transcriptomic profiles of these cell types, we have identified multiple genes that become differentially expressed along NB lineage development. This included known NB lineage fate regulators, but also unstudied genes within this context. Furthermore, we have observed notable expression heterogeneity within NBs and neurons. Interestingly, we found that multiple genes differentially expressed between larval NBs show the same expression pattern in embryonic NBs. This supports the idea that NB lineage identity programs are conserved from embryo to larval neurogenesis. Finally, we have also validated in vivo the expression of a few marker genes which were found to be distinctively expressed among NBs. Overall, this work should provide future studies with a useful molecular blueprint of players potentially driving NB lineage development and the generation of neuronal diversity.
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Palavras-chave
Drosophila Diversidade neuronal Linhagens de neuroblastos Sequenciação de célula-única Drosophila Neuronal diversity Nuroblast lineages Single-cell sequencing