Biological nitrogen fixation by rhizobia is an important process in the cultivation of leguminous crops such as soybean. Soybean undergoes a symbiosis with Bradyrhizobium species, where bacteria enter specific root organs (“nodules") and differentiate into nitrogen fixing organelles, the symbiosomes. Whilst the molecular cross-talk that governs the initial steps of the symbiosis is increasingly understood, the molecular landscape at the symbiosome interface is poorly characterised. This project will use different electron microscopy techniques together with biophysics to decipher nutrient exchange and communication processes at this plant-bacterial interface. First, a novel in situ structural biology pipeline will be established, where symbiosomes and protoplasts from infected nodule cells are vitrified and subjected to thinning and cryo-electron tomography analysis. Next, this pipeline will be used to characterise structural features of the symbiosome interface, sampling soybean nodules over time and under nitrate stress. Finally, specific protein complexes active at the symbiosome interface will be fluorescently tagged and structurally analysed using biophysical methods, mass spectrometry analysis and single particle cryo-EM. Together, these techniques will offer unprecedented detail of molecular processes and protein composition at the symbiosome membrane and will provide a novel in situ structural biology tool for the (inter)national molecular plant biology communities.