仿刺参FoxO1介导夏眠抗氧化防御调控的分子基础研究

The sea cucumber Apostichopus japonicus is an important aquaculture species in China. During the period of aestivation, it tends to get damaged from the shock of reactive oxygen species, which increases the risk of aestivating aquaculture. Antioxidant defenses is the primary way to resist oxidative stress, and also an important guarantee for sea cucumber to go through aestivation safely. However, there is very limited information in terms of the molecular basis of antioxidant defenses regulation during aestivation in sea cucumber. In preliminary work of aestivation in sea cucumber, a unique regulating mode was observed in antioxidant defenses. Meanwhile, FoxO1 was found to play an important role in the regulation. In this research, the functional and expression mode will be studied by qRT-PCR and Western blot; the regulatory factors and target gene will be studied by Co-IP and CHIP-Seq; the regulation mode will be verified by RNAi and protein injection. The research achievements will elucidate the molecular basis of antioxidant defenses regulation mediated by FoxO1 during aestivation in sea cucumber, and promote the processes of selective breeding of resistant variety.

仿刺参是我国重要的海水养殖品种，其在夏眠过程中易遭受活性氧冲击而发生损伤，持续的氧化压力加大了夏眠过程的养殖风险。抗氧化防御是仿刺参夏眠期间抵御氧化冲击的主要手段，是其安全度过夏眠的重要保障。然而，仿刺参夏眠期间抗氧化防御调控的分子基础研究还十分有限。项目组前期研究发现，仿刺参在夏眠过程中可能具有独特的抗氧化防御调控模式；同时发现，FoxO1（叉头盒因子O1）可能在介导调控过程中发挥了重要作用。基于前期研究和项目组已掌握的仿刺参基因组数据，本研究由FoxO1入手，拟通过重组蛋白注射、RNA干扰、Co-IP与质谱联用、以及ChIP-Seq等技术，探明FoxO1的功能和表达模式，鉴定与FoxO1结合的调节因子和靶基因，并对其调控模式进行验证和分析。研究结果将阐明仿刺参夏眠过程中FoxO1介导抗氧化防御调控的分子基础，为提高仿刺参夏眠防御能力、推动仿刺参抗逆优良品种的选育工作提供重要依据。

仿刺参在夏眠期间，机体会处于一种高度缺氧状态，易造成氧化抑制或氧化应激，从而对机体造成损伤。抗氧化防御作用是仿刺参夏眠期间应对氧化冲击的主要防御手段，也是其安全度过夏眠的重要保障。在国家自然科学基金的资助下，本研究分析了FoxO1及其它抗氧化防御调控网络功能基因在仿刺参夏眠中的功能，以期进一步理解仿刺参夏眠抗氧化防御调控的分子机制。项目研究结果主要包括以下内容。项目分析了FoxO1基因的结构与进化特征，从仿刺参夏眠不同阶段、不同组织、以及成参与幼参对比等多个角度解析了FoxO1在仿刺参夏眠不同阶段的水平与表达变化以及对夏眠抗氧化防御特征的影响，并通过多样性关联分析鉴定了SNP位点-310 A-C与仿刺参夏眠抗氧化防御能力显著关联；对仿刺参夏眠抗氧化防御过程的mRNA转录组学和miRNA变化进行了解析，鉴定了10个在仿刺参夏眠抗氧化防御过程中调控FoxO1表达的miRNA，同时鉴定了FoxO1调控相关基因等仿刺参夏眠抗氧化防御调控网络功能基因并分析了调控关系；对Keap1、Nrf1、Prx和HSP等夏眠抗氧化防御调控网络功能基因结构、进化与表达特征进行了全面解析，研究结果有助于全面理解仿刺参夏眠抗氧化防御调控网络和调控分子机制，为仿刺参分子标记辅助育种提供技术支撑。