植物源烟水对丹参次生代谢产物积累的影响及“smoke signals”机制研究

Plant-derived smoke water is produced by combusting plant materials and bubbling smoke through water. The ecological effects of smoke water caused the attention of ecologists. Research abroad is very active while less attention in China. It was found that great changes had taken place in the content and composition of secondary metabolites in Salvia miltiorrhiza which were planted in after "burning" field. Furthermore, it has been confirmed by controlled experiments that smoke water contains chemical signal molecules which could affect the accumulation of secondary metabolites in S. miltiorrhiza. A scientific hypothesis was proposed in this project that plant-derived smoke water could regulate the accumulation of secondary metabolites in S. miltiorrhiza based on “smoke signals” mechanism. Based on the preliminary verification of signal molecule (KAR1) in the smoke water, signal molecules will be separated from smoke water on the basis of “precision discovery”strategy. The effects of smoke water on the secondary metabolites of S. miltiorrhiza will be studied using metabolomics technologies. Then, the signal transduction of secondary metabolism in S. miltiorrhiza regulated by smoke water will be discussed using means of transcriptomics and proteomics. It is hopeful that this project will be the pioneering study of smoke water in resource ecology of traditional Chinese medicine and endow the ancient tradition of "burning" with new ecological connotation.

植物源烟水是指植物燃烧产生的烟溶入水中所得的溶液，烟水的生态学效应引起了生态学界的极大关注，相关研究在国外非常活跃，国内关注较少。申请人发现野外“烧荒”后的过火痕地上种植的丹参植物体内次生代谢产物的含量与组成发生了很大变化，进一步实验证明了烟水中存在调控丹参次生代谢的化学信号分子，由此提出“基于‘smoke signals’机制的植物源烟水调控丹参次生代谢产物积累”的科学假说。本项目将在初步明确烟水中信号分子（丁烯羟酸内酯，KAR1）的基础上，采用“精准”发现策略分离烟水中的信号分子；采用代谢组学的手段揭示信号分子对丹参次生代谢整体轮廓的影响；采用转录组学和蛋白组学的手段揭示信号分子调控丹参次生代谢的信号转导机制。本项目将开拓烟水在我国中药资源生态学领域的研究，赋予“烧荒”这一古老传统以新的生态学内涵。

植物源烟水是指植物焖烧产烟入水所得的溶液，烟水对植物的生理生态功能引起了生态学界的极大关注，相关研究在国外非常活跃，国内关注较少。本项目采用代谢组学手段揭示烟水中信号分子KAR1对丹参次生代谢整体轮廓的影响；采用转录组学和蛋白组学的手段揭示信号分子调控丹参次生代谢的信号转导机制。结果表明：①自主研发了烟水制取设备，获得高效稳定的烟水。基于高速逆流色谱技术建立了KAR1的分离方法。以糠醇为原料对KAR1进行了化学全合成。②KAR1依赖内源性信号分子NO和JA调控丹参中丹参酮I的积累，且NO位于JA的上游。内源性钙离子（Ca2+）-钙调素（CaM）、油菜素内酯（BL）和双氧水（H2O2）作为信号分子参与KAR1诱导丹酚酸B的合成，且Ca2+-CaM和BL位于H2O2的上游，Ca2+-CaM与BL无明显上下游单向调节关系。③SW和KAR1对丹参整体代谢轮廓表现出相似的影响，均能显著促进黄酮类（葛根素、车前子苷、三叶豆紫檀甙等）和萜类化合物（特女贞苷、獐芽菜苷、栀子苷、橄榄苦苷等）的积累。④SW和KAR1启动丹参体内糖酵解途径、糖基转移酶、萜类和丹酚酸类代谢途径，诱导丹参酮类合成中AACT、HMGS、HMGR、DXS、DXR、CMK以及丹酚酸类合成中PAL、C4H、RAS、TAT、HPPR、4CL等基因的表达。⑤SW和KAR1能显著上调丹参中与丹酚酸生物合成相关的蛋白（4-香豆酸酯-CoA连接酶和莽草酸O-羟基肉桂酸-转移酶）以及与木质素生物合成相关的酶（肉桂醇脱氢酶和过氧化物酶）。上述结果为阐明烟水调控丹参次生代谢产物积累的分子机制奠定基础。本项目开拓了烟水在我国中药资源生态学领域的研究，赋予“烧荒”这一古老传统以新的生态学内涵。发表相关SCI论文9篇，获得授权专利5项，培养硕士研究生3名。超额完成任务书中的各项指标。