基于单细胞RNA测序解析黄鳝性逆转过程中生殖细胞谱系的发生和分化

Swamp eel (Monopterus albus), a hermaphrodite fish with female premature, possesses a characteristic of natural sex-reversal from female-to-male transition through an intersex stage. It is an excellent specie to study vertebrate sexual development. However, the existence of germ cells in ovary, ovotestis and testis, and their differentiation and development patterns remain elusive. We have completed the work of whole genome sequencing and assembly of the Monopterus, and analyzed the gene expression profile during gonadal reversal using RNA-seq data, identifying a large number of genes related to sex-reversal. Moreover, a potential ovary specific germline stem cell group were identified by immunohistochemistry method. Based on the previous studies, we will investigate the cell types in the three gonads, reconstruct the differentiation processes of each cell lineage, uncover the origin of male germline stem cells, and establish the model of germ cell transition during sex-reversal by the single-cell transcriptome sequencing technology. This project will lay a foundation for revealing the molecular mechanism of germ cell generation and differentiation during sex-reversal of the Monopterus.

黄鳝为雌性先熟型雌雄同体鱼，幼年雌性黄鳝第一次性成熟产卵后发生性逆转转变为雄性，是研究脊椎动物性别发育的重要物种。然而黄鳝雌、间、雄性腺生殖细胞的类型及分化发育模式仍然未知。我们前期已完成了黄鳝的全基因组测序和三种性腺转录组测序工作，鉴定出大量与性逆转相关的基因，进一步通过免疫荧光实验发现一种潜在的雌性性腺特异的生殖干细胞类型。本研究在此基础上，拟通过单细胞转录组测序技术鉴定不同发育时期三种性腺中的细胞类群，构建相关的细胞谱系发生过程，明确雄性生殖干细胞的起源，建立性逆转过程中生殖细胞谱系转变模式，为最终揭示黄鳝性逆转过程中生殖细胞谱系发生和分化的分子机制奠定基础。

黄鳝为雌性先熟型雌雄同体鱼，幼年雌性黄鳝第一次性成熟产卵后发生性逆转转变为雄性，是研究脊椎动物性别发育的重要物种，鉴定黄鳝不同类型性腺中的生殖干细胞类群，并分析其在性逆转过程中的动态变化，有望揭示黄鳝性逆转的本质。本项目对5例包含有雌、间、雄三种类型的黄鳝性腺进行了单细胞转录组测序，并且进行了深入的生物信息分析与数据挖掘，鉴定出黄鳝不同类型性腺中的生殖细胞类群、性腺基质细胞类群、性腺免疫细胞类群以及生殖干细胞类群，分别构建了雌性生殖细胞和雄性生殖细胞的成熟分化轨迹。通过对生殖干细胞类群的深入分析，我们在雌性性腺中鉴定出一种特异高表达wdr17的生殖干细胞类群，这群细胞特异存在于雌性性腺中，并且表现出原始生殖干细胞的特征。进化分析表明这群细胞可以通过性逆转过程分化为雄性生殖干细胞。通过分析wdr17highGSC向雄性分化过程中变化的基因和信号通路，鉴定出Wnt信号通路可能在性逆转过程中参与生殖干细胞的转变。

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