Ecological Function Of Amino Acid Exporter MsiAR System In Early Signal Communication Between Mesorhizobium Tianshanense And Glycyrrhiza Uralensis

Mesorhizobium tianshanense is a nitrogen-fixing bacterium which can establish symbiotic associations with Glycyrrhiza uralensis. Complicated Signal communication are required for Establishment of symbiosis, this communication may occur as early as seed germination. During germination, seeds secret a lot of nutrient small molecular, including sugars, amino acid, which can induce chemotaxis of bacteria in soil.Formation of invasion thread, nodulation and nitrogen fixation are studied significantly, many genes are found necessary for these procedures. But there are few study on the early interaction between Rhizobium and their host plant, especially before attachment of bacteria on root hair. So my project was focused on how the compounds secreted during Glycyrrhiza uralensis seed germination affect M. tianshanense gene transcription profile, those genes whose transcription changed may play important roles in the early interaction between Rhizobium and plants.We put exude of Glycyrrhiza uralensis seed into agar plates to imitate the chemical surroundings of seed germination. Five genes(msiA,B,C,D,E) were identified to be induced by seed exude, by using a transposon containing a promoterless kanamycin resistant gene. msiA, which can be induced 50 times by seed exude, belongs to lysE family which encoding amino acid exporter protein.The induction of seed exude on msiA promoter needs the product of msiR, which located upstream of msiA, MsiR belongs to LysR regulation family. MS result showed that the functional compound in seed exude is canavanine, which is an analogue of arginine and can be misintegrated into polypepetide to form unfuntional protein, leading death of bacteria finally. Arginine and lysine, which can induce the expression of lysE in Escherichia coli and Corynebacterium glutamicum separately, cannot induce msiA expression.Canavanine was 104 times more toxic to msiA or msiR mutant than to wild type strain under culture condition where yeast extract was as low as 0.005%(W/V), also the intracellular canavanine concentration of msiA or msiR mutant was higher than wild type strain, suggesting that MsiA/R system may act as canavanine exporter to maintain intracellular canavanine to a nontoxic level. Root hair attachment experiment showed that the amount of msiA mutant attached to root hair is 100 times lower than wild type strain.We also separated stains from rhizosphere of blank soil or soil in which Glycyrrhiza uralensis were planted, tested the resistance of these strains and calculated the ratio(NCR/ NCS) of canavanine resistant strains(NCR) and canavanine sensitive strains(NCR) from different samples. We found that the ratio ratio(NCR/NCS) from plant soil is 8 times higher than from blank soil, suggesting that Glycyrrhiza uralensis may secret canavanine to change bacteria population in rhizosphere.Based on these data, we put forward the model for early interaction between M. tianshanense MsiA/R system and Glycyrrhiza uralensis:during germination, seed can secret canavanine as an antimicrobial to optimize the bacteria population in the rhizosphere, killing or repressing the potential hazard bacteria. But beneficial bacteria such as M. tianshanense can propagate to be the dominant population paving the way for further symbiosis, cause of MsiA/R system.