基于单细胞水平的孕二烯酮诱导雌性西部食蚊鱼雄性化的研究  

Exploration of GES-induced masculinization of Gambusia affi nis Based on Single Cell Level

作　　者：冯婷茵 邓希楷 邢珂 FENG Tingyin;DENG Xikai;XING Ke(College of Life Sciences,Guangzhou University,Guangzhou 510006,China)

机构地区：[1]广州大学生命科学学院,广东广州510006

出　　处：《广东农业科学》2023年第6期126-136,共11页Guangdong Agricultural Sciences

基　　金：广东省重点领域研发计划项目(2018B030335001)。

摘　　要：【目的】从单细胞转录组水平研究雌性西部食蚊鱼(Gambusia affinis)性别逆转过程中涉及到的性腺细胞分化发育和基因表达变化,探讨孕二烯酮(Gestodene,GES)作用下食蚊鱼性别反转的分子机制。【方法】将培养驯化2个月后的性成熟雌性食蚊鱼暴露于浓度为500 ng/L的孕二烯酮溶液中进行培养,于第4、8周取样观察其形态变化,并分别取正常雌性对照、雄性对照、暴露4周和8周样本的性腺制成单细胞悬液,进行单细胞转录组测序并分析。【结果】经孕二烯酮处理后的雌性食蚊鱼形态上出现臀鳍类生殖足现象。单细胞转录组测序结果发现,雄性食蚊鱼和雌性食蚊鱼的性腺具有完全不同的细胞构成。雄性食蚊鱼性腺主要由成纤维细胞、雄性生殖细胞、上皮细胞和原始生殖细胞构成,而雌性性腺主要由内皮细胞、成纤维细胞、卵母细胞和原始生殖细胞构成。暴露4、8周后雌性食蚊鱼性腺中出现少量的雄性生殖细胞,且原始生殖细胞数量明显上升。进一步分析发现,原始生殖细胞中性别决定关键基因出现差异化表达情况。细胞轨迹分析表明,部分原始生殖细胞出现向雄性分化的特征,其中有2477个基因在两个不同的分化方向表现出不同的表达模式。KEGG通路富集分析结果表明,Toll-like receptor、TGF-beta、Notch等信号通路参与原始生殖细胞向雄性化逆转的过程,甘氨酸,丝氨酸、苏氨酸、半胱氨酸、甲硫氨酸代谢、氨基糖和核苷酸糖代谢等途径也与该过程相关。【结论】雌性食蚊鱼经孕二烯酮诱导后,在形态学上出现臀鳍雄性化特征,性腺中有部分原始生殖细胞向雄性生殖细胞转化。原始生殖细胞中性别决定关键基因amh、fabp3、dmrt1、wt1a、wt1b、cyp19a1a和fgf16出现差异化表达情况。Toll-like receptor、TGF-beta、Notch和一些氨基酸代谢信号通路参与了原始生殖细胞向雄性化逆转的过程。【Objective】To investigate the molecular mechanism of sex reversal in female Western mosquitofish(Gambusia affinis)under the effect of gestodene(GES)by studying the development of gonadal cell differentiation and gene expression changes involved in the process of sex reversal in female mosquitofish at the single-cell transcriptome level.【Method】Sexually mature female mosquitofish were exposed to 500 ng/L GES after culture and domestication for 2 months,and samples were taken at the 4th and 8th week to observe morphological changes,and single-cell suspensions were made from the gonads of normal female controls,male controls,and samples exposed for 4 and 8 weeks,respectively,for single-cell transcriptome sequencing and analysis.【Result】The morphology of GES-treated female mosquitofish showed gonopodium,and single-cell transcriptome sequencing revealed completely different cellular composition of gonads between male and female mosquitofish.The gonads of male mosquitofish consist mainly of fibroblasts,male germ cells,epithelial cells and primordial germ cells,whereas the gonads of female mosquitofish consist mainly of endothelial cells,fibroblasts,oocytes and primordial germ cells.After 4 and 8 weeks of exposure,a small amount of male germ cells appeared in the gonads of female mosquitofish,and the number of primordial germ cells increased significantly.Further analysis revealed that several crucial genes related to the sex-differentiation were differentially expressed in primordial germ cells.Cell trajectory analysis showed that some primordial germ cells showed characteristics of differentiation towards males,in which 2477 genes showed different expression patterns in two different different directions of differentiation.The results of KEGG pathway enrichment analysis revealed that signaling pathways,such as Toll-like receptor,TGF-beta,Notch,etc.,were involved in the process of reversal of primordial germ cells towards maleness.Glycine,serine,threonine,cysteine,methionine metabolism,amino sugar and nucle

关 键 词：孕二烯酮(GES) 西部食蚊鱼 硬骨鱼 性别逆转 单细胞转录组 

分 类 号：S917.4[农业科学—水产科学]