Thesis

The role of podJ in attachment during Sinorhizobium meliloti-medicago truncatula symbiosis

Nitrogen is an essential nutrient that is inaccessible to many organisms in its abundant atmospheric form. Rhizobia, a group of bacteria capable of fixing molecular dinitrogen into more useful forms, symbiotically colonize the root nodules of leguminous plants and provide these hosts with nitrogenous compounds. The exploitation of nitrogen-fixing bacteria has many applications in agriculture, since synthetic fertilizers have adverse effects on the natural environment. Sinorhizobium meliloti is an alpha-proteobacterium that engages in symbiosis with the model legume, Medicago truncatula. The S. meliloti polar organelle development protein (PodJ) is an important factor for symbiosis. Originally identified in Caulobacter crescentus, PodJ is required for the production of holdfast, which is an adhesive organelle synthesized at the bacterial cell pole that enhances attachment to surfaces. A nonsense mutation was identified in the podJ gene in the domesticated S. meliloti Rm1021 lab strain. When the mutation was corrected, the new Rm 1021 -derived podJ+ strain effectively outcompeted Rm 1021 for root nodule invasion in M. truncatula, suggesting that PodJ enhances attachment of S. meliloti to the host plant. To investigate this, competition experiments between Rm1021 and podJ+ strains were conducted. Confocal microscopy analysis showed that the p o d f strain attaches more robustly to the root compared to the Rm1021 strain. Quantitative polymerase chain reaction experiments were used to assess the relative abundance of the Rm1021 and podJ+ strain on root hair. Collectively, the data indicate that PodJ is a conserved polarity factor that contributes to effective symbiosis. Identification of additional molecular factors that may make these types of symbioses more efficient will ideally aid in the development of sustainable substitutes for synthetic fertilizer.

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