Biochemical Compatibility In Sinorhizobium-Medicago Symbiosis

Purdue University, West Lafayette IN

Investigators

Abstract

Molecular signaling is essential for the establishment of legume-rhizobia symbiosis, and specific structural features of signal molecules may determine symbiotic compatibility. This project focuses on ecotype-strain compatibility in Medicago truncatula-Sinorhizobium meliloti interactions. Wild-type S. meliloti strains NRG185 and NRG34 infect M. truncatula ecotype A17 (compatible), but fail to establish nitrogen-fixing nodules on ecotype A20 at 28 days post infection (incompatible), and the phenotypes are reversed with S. meliloti strains NRG247 and Rm41. Previous research showed that compatible infection is dependent on bacterial extracellular polysaccharide (EPS) production, suggesting that different ecotypes of M. truncatula have different structural requirements for oligosaccharide activity, and a correlation between EPS oligosaccharide structure and the host specificity of the bacterial strain was identified. The primary objective of the project is to determine the structural requirements for EPS oligosaccharide activity in the compatible infection of M. truncatula A17 versus A20 (i.e., to determine the structural basis for compatibility in each host plant), which will be accomplished by extensive purification and analysis of the oligosaccharides produced by strains NRG185 and Rm41. The purified oligosaccharides will then be applied to "rescue" experiments with EPS mutants and incompatible strains, to determine structure/function relationships in compatible interactions. Finally, the EPS (exo) gene cluster from strains NRG185 and Rm41 will be cloned and sequenced for comparison, and strains producing heterologous EPS will be constructed for compatibility analyses. M. truncatula is the subject of a genomic project for the study of plant-microbe interactions, so the identification of specific signal-molecules can be followed by an analysis (at the genetic level) of the plant response to compatible and incompatible infection. The project also includes elements that broaden the impact of the work: First, undergraduates and rural Indiana high school students will participate in the research (the plant assays), and summer workshops will be held to introduce urban high school students to research at Purdue. Second, the three year study will coincide with a program to build an NMR facility in the Purdue University School of Agriculture (NMR is the principal tool for oligosaccharide structural analysis), which will result in educational and research benefits for students and researchers throughout the School. Third, the results of the project will be disseminated in a website, which will have links to relevant biochemical information and structural features of the plants and bacteria. Nitrogen deficiency is a prime limitation to the growth of agricultural plants in the field; however, legumes, such as soybean and alfalfa, form a symbiotic association with specific bacteria that results in the production of utilizable nitrogen compounds. More than 200 million metric tons of "fixed" nitrogen are added to the earth's agricultural fields each year from this process. Plant-microbe symbiosis involves considerable developmental regulation and molecular signals control the progress of the association: First, bacterial compounds induce the development of small appendages (nodules) on the roots of the host plants, and then the nodules are occupied by the bacteria via a complex infection process, which is promoted by distinct bacterial products. This project involves a study of the bacterial signal molecules that are required for the early stages of infection, and the results should provide insight into general mechanisms in bacterial infection.