芳香化酶抑制剂诱导成年斑马鱼雌到雄性逆转的分子机制研究

Fish sex is determined in extremely complex ways and exhibits plasticity with the combination of genetic and environmental influences. However, the underlying mechanism of its sexual plasticity is still unclear even in zebrafish, which is the well-established teleost research model. Our previous study in Drosophila melanogaster, whose sex is controlled by sex chromosome, demonstrates large differences in transcriptomes between wildtype females and female-to-male transformed mutants, suggesting the feasibility of studying the underlying mechanism of sexual plasticity with transcriptomic changes in sex reversal. Here, to find out the molecular mechanisms underlying the environmental effects on sex reversal, we 1) construct female-to-male transformed zebrafish with Aromatase inhibitor treatment, as well as female and male zebrafish with and without the treatment; 2) measure the expression profiles of individuals from each group by RNA-seq; and 3) identify the differentially expressed genes among groups to determine the candidate genes and pathways related to the sex reversal caused by aromatase inhibitor treatment. The implementation of the project will provide a solid foundation for revealing environmental influences on sexual plasticity, and is important for studying the evolution of sex-determining mechanisms and evaluating the effects of chemical pollution.

鱼类性别的决定方式多样，且受到基因和环境的共同影响，具有显著的可塑性。但是即使对于目前科学研究中重要的鱼类模式生物斑马鱼来说，其性别发育受到环境影响的机制也尚未阐明。本项目申请人以往在果蝇中（其性别受性染色体控制，而受环境的影响小）的研究发现，野生型雌性果蝇和雌转雄性突变体果蝇在转录组水平具有较大差异，表明性别可塑性的分子机制可以通过性反转过程中的转录组变化来研究。为了找到环境对性别影响的分子基础，本项目以斑马鱼为研究材料，构建用芳香化酶抑制剂处理后雌转雄性和未变性的斑马鱼，以及对照组的正常雌雄性斑马鱼。通过RNA-seq测定各类斑马鱼的表达谱，找出各类斑马鱼之间的差异性表达基因，从而确定与芳香化酶抑制剂对斑马鱼性别发育影响相关的候选基因及其通路。本研究的顺利实施将为揭示环境对性别发育的影响提供理论基础，对性别决定机制演化的研究和化学污染效果的评估具有重要意义。

斑马鱼在幼年时难以区分性别，极大制约了人们对其早期性别分化机制的研究。而芳香化酶抑制剂的处理可以定向诱导幼鱼分化为雄性，甚至使成年雌鱼发生性逆转，从而为解决这一难题提供了办法。为了确定幼鱼性别分化和成鱼早期性逆转中的转录组变化，我们对芳香化酶抑制剂处理32天（此时幼鱼的性别分化完成）的幼鱼和成鱼进行了转录组测序。在成年雌鱼中，芳香化酶抑制剂的处理使雌激素水平下降，并导致gsdf, macf1a和paqr5a等与雌激素代谢、雌配子生成和卵子发生过程相关的基因上调，以及vtg6, vtg2, vtg4和 vtg7等卵黄原蛋白基因下调。而在幼鱼中，芳香化酶抑制剂的处理使bcl2l16和anax1c等与细胞死亡和凋亡相关的基因上调，而zar1和zpcx等与正调控生殖过程和卵细胞分化相关的基因下调。这些结果表明，芳香化酶抑制剂的处理使成年雌鱼而非幼鱼发生抗雄性化，这可能是导致成鱼比幼鱼性别可塑性弱的原因。