拟南芥转运蛋白AtTSPO的渗透调节在种子萌发中的作用机制

Osmotic regulation plays important roles in plant development and growth while ABA, a stress phytohormone, is involved in many stress signaling pathways. Arabidopsis translocator protein AtTSPO is induced by ABA and osmotic stress supporting that AtTSPO is related to stress responses. A preliminary observation indicates that AtTSPO may be involved in seed germination. Yet, it remains unclear whether and how AtTSPO regulates seed germination. In this proposal, we plan to perform following experiments to address these scientific questions. (1)Elucidate the roles of AtTSPO in osmotic and ABA regulation of seed maturation and germination via penotypic analyses of AtTSPO related genetic plant materials including overexpression and mutation plants; Analyze the expression patterns of AtTSPO gene in seed development and germination, and its regulation by ABA and osmotic pressure, by using GUS reporter and quantitative RT-PCR approaches, to uncover the gene expression basis; Study dynamics of protein levels, and cellular localization in seed development and in seed germination subject to ABA and osmotic stress, by using Western blot and GFP reporter assays, to explore the protein dynamics basis for AtTSPO functioning; Decipher the possible relation of AtTSPO to ABA signaling pathways, by using multiple approaches including genetic analyses, and assays of gene expression, protein dynamics, and ABA content, and screening for AtTSPO-interacting proteins.

渗透调节在植物生长发育过程中起重要的作用,同时ABA作为一种植物逆境激素，参与很多逆境信号过程。转运蛋白AtTSPO与逆境响应有关，其表达受ABA和渗透胁迫的诱导，初步的观察提示它可能影响种子萌发，但其调节种子萌发的分子机制尚不清楚。我们通过遗传材料的表型分析研究AtTSPO在种子成熟和萌发渗透调节和ABA响应中的作用；用GUS染色和定量RT-PCR研究AtTSPO基因在种子发育和萌发中的表达模式、以及渗透压和ABA对它的表达调节，揭示AtTSPO作用的基因表达基础；用Western blot和GFP融合蛋白检测AtTSPO蛋白在种子发育成熟和萌发中的蛋白动态和细胞定位，揭示其作用的蛋白动态和细胞定位基础；通过遗传分析、基因表达和蛋白动态分析、ABA含量分析、以及AtTSPO互作蛋白分析，研究AtTSPO与ABA信号途径的可能关系。

ABA对植物生长发育和逆境响应具有重要作用，初步的研究提示AtTSPO可能与ABA和逆境响应相关。通过突变体和过表达植物材料的表型分析，证明了AtTSPO在种子萌发和休眠中的调节中发挥重要作用。用qRT-PCR和GUS染色技术研究了种子发育（包括成熟阶段）、后熟和萌发过程中的AtTSPO基因表达模式，用Western blot技术研究了蛋白量和定位的动态变化规律，揭示了AtTSPO作用的基因表达和蛋白动态基础。qRT-PCR和Western blot技术证明了AtTSPO作为敏感的渗透压感受器介导渗透压对种子成熟和萌发的调节。通过基因表达和遗传分析确定了AtTSPO与ABA信号途径（主要蛋白）的关系。筛选到了AtTSPO的互作蛋白，用多种技术验证了它们之间的互作，并对VAP27的功能进行了初步研究，正在通过双突变体进行它们之间遗传关系的分析，以及它们定位关系的细胞学分析。我们运用多种研究技术和手段，初步揭示AtTSPO在植物种子萌发响应ABA、低渗和高渗环境的分子机制。这些结果对阐明种子萌发和休眠的分子机制具有重要参考价值。