类外泌体miR-1085介导微拟球藻MAPK信号通路以应答环境的分子机制研究

外泌体miRNA能介导细胞通讯以应答环境。研究表明，MAPK通路与环境应答密切相关。然而，外泌体miRNA介导微藻MPAK通路以应答环境尚无报道。申请人的前期研究显示，微拟球藻通过膜融合凹陷胞吐的方式分泌脂滴，方式与动植物外泌体一致，因此将分泌的脂滴命名为类外泌体。借助小RNA高通量组学发现，类外泌体miR-1085可能调控MAPK信号通路相关的CaM4，但调控机制不明确。本研究拟对miR-1085的调控机制开展研究。拟解决两个关键问题：借助分子生物学等手段阐明miR-1085的前体加工成熟过程、与CaM4结合位点互作介导细胞通讯的分子机制；借助蛋白质组学、蛋白质修饰组学、细胞生物学、生物化学等方法明晰miR-1085对MAPK通路多级传导过程影响及对微藻应答能力的影响，以阐明类外泌体miR-1085调控微藻MAPK通路以应答环境变化的机制，为微藻类外泌体介导细胞通讯奠定基础。

Exosomal miRNA can mediate cell-cell communication in response to the environmental conditions. Previous studies have shown the regulatory role of MAPK pathway in response to environmental conditions. However, there is no study available about the role of exosomal miRNA-mediated MAPK pathway regulation in microalgae in response to environmental conditions. Our previous research data demonstrated the secretion of overproduced lipid droplets in oleaginous heterokont Nannochloropsis oceanica across exoplasmosis with invaginated cellular membrane, which is consistent with the exosomes of vascular plants and animal cells and thus, these secreted components were designated as exosome-like particles. With small RNA high-throughput omics, we found that exosomal miR-1085 could possibly regulate CaM4 related to MAPK signaling pathway, however, the molecular mechanism underpinning this phenomenon is unclear. The present research proposal aims to dissect the mechanistic role of miR-1085. It is proposed to solve the following two key issues. To characterize the miR-1085 precursor processing and maturation and it's molecular mechanism of interaction with CaM4 binding sites to mediate cell communication using molecular biology tools; Then, Proteomics, proteomics modification, cell biology, biochemistry and other methods were used to investigate the influence of miR-1085 on the multi-stage transduction process of MAPK pathway and the response ability of microalgae. To elucidate the mechanism by which the exosomal miR-1085 regulates the MAPK pathway of microalgae in response to environmental changes, together these results could pave the way to advance our understanding on exosomes-mediated cell-cell communication in microalgae.