The role of glycolipids at the interface of plant-microbe interactions during nodulation and myorrhiza formation in Lotus japonicus

糖脂是植物和微生物交界处的膜的组成成分。我们正在研究半乳甘油脂、甾醇糖苷和葡萄糖神经酰胺在莲花植物在结瘤期间与中生根瘤菌或在菌根形成期间与真菌球囊菌共生相互作用中的作用。由于每一类糖脂都由许多分子物种组成，因此建立了一种基于四极飞行时间质谱仪(Q-TOF MS)的糖甘油脂类、甾醇类脂类和鞘磷脂类化合物的测定方法。这包括首次开发一种测量植物中所有类固醇脂类(游离类固醇、类固醇葡萄糖苷、类固醇酯类、酰化类固醇类葡萄糖苷)的方法。携带糖脂合成相关基因(DGD1、DGD2、SGT1、SGT2、GCS)的RNAi构建的转基因莲花植株是在之前的资助期间培育出来的。Q-TOF MS测定表明，在SGT2和GCS RNAi系中，甾醇糖苷和葡萄糖神经酰胺分别被减少。我们获得了莲花基因DGD1、DGD2、SGT2和GCS的分蘖突变体。转基因RNAi品系和突变品系的结瘤和菌丝化实验正在进行中。此外，我们开发了一种非靶向的Q-TOF MS方法，我们将使用该方法来寻找对莲花结瘤或菌丝化反应的脂质。

Glycolipids are constituents of membranes at the boundary between plants and microorganisms. We are studying the role of galactoglycerolipids, sterol glucosides and glucosylceramides in the symbiotic interaction of Lotus japonicus plants with Mesorhizobium during nodulation or with the fungus Glomus during mycorrhiza formation. Because each glycolipid class consists of numerous molecular species, a quadrupole-time of flight mass spectrometry (Q-TOF MS) based method was developed for the measurement of glycoglycerolipids, sterol lipids and sphingolipids. This includes the first development of a method for the measurement of all sterol lipid classes in plants (free sterols, sterol glucosides, sterol esters, acylated sterol glucosides). Transgenic Lotus plants carrying RNAi constructs for the genes involved in the synthesis of glycolipids (DGD1, DGD2, SGT1, SGT2, GCS) were generated during the previous funding period. Lipid measurements by Q-TOF MS revealed that sterol glucosides and glucosylceramides are reduced in SGT2 and GCS RNAi lines, respectively. We obtained TILLING mutants for the Lotus genes DGD1, DGD2, SGT2 and GCS. Nodulation and mycorrhization experiments of the transgenic RNAi lines and of the mutant lines are underway. Furthermore, we developed a non-targeted Q-TOF MS approach that we will employ to search for lipids responding to nodulation or mycorrhization in Lotus.