Abstract
Symbiosis between legumes and rhizobia is of major importance in terrestrial ecosystems due to its ability to fix atmospheric nitrogen. The molecular dialogue between the two partners, which ultimately leads to development of nodules hosting the N2 fixing bacteria, can be initiated by the binding of Nod factors (NF) secreted by the rhizobial partner on NF receptors at the legume root hair plasma membrane (PM). This triggers a Ca2+ influx through the PM, followed by a cascade of ionic signaling events, involving changes in H+, K+, and Cl- fluxes at the PM. My objective was to characterize molecular mechanisms underlying these early ionic signaling events in the legume model Medicago truncatula. By using the patch-clamp technique on protoplasts obtained either by cell wall enzymatic digestion or laser-assisted ablation from growing root hairs, I have contributed to characterize several ion conductances from this cell type. I especially focused on a cationic conductance activated by membrane hyperpolarization (HACC), uniquely found to be most permeable to Ca2+. This conductance was quickly activated (within less than 1 minute) following NF addition at physiological concentration. Its activation was dependent on the presence of functional NFP (“Nod Factor Perception”) receptors, which suggested that this conductance mediates the early Ca2+ influx triggered by NF perception. In addition, cationic transport systems expressed in M. truncatula root hairs and belonging to the HKT and Glutamate receptor-like (GLR) families were investigated as potential contributors to the early ionic signaling events. Loss-of-function mutant analysis for 3 highly expressed GLRs suggested that these genes did not play major roles in the expression/activity of the HACC conductance, and were not indispensable for nodulation. On the other hand, the HKT transporters, which were found to be Na+-selective, were expressed in nodules, which suggested a role in symbiosis.