Extracellular Vesicle-Driven Crosstalk between Legume Plants and Rhizobia: The Peribacteroid Space of Symbiosomes as a Protein Trafficking Interface

Prokaryotes and eukaryotes secrete extracellular vesicles (EVs) into the surrounding milieu to preserve and transport elevated concentrations of biomolecules across long distances. EVs encapsulate metabolites, DNA, RNA, and proteins, whose abundance and composition fluctuate depending on environmental cues. EVs are involved in eukaryote-to-prokaryote communication owing to their ability to navigate different ecological niches and exchange molecular cargo between the two domains. Among the different bacterium-host relationships, rhizobium-legume symbiosis is one of the closest known to nature. A crucial developmental stage of symbiosis is the formation of N₂-fixing root nodules by the plant. These nodules contain endocytosed rhizobia─called bacteroids─confined by plant-derived peribacteroid membranes. The unrestricted interface between the bacterial external membrane and the peribacteroid membrane is the peribacteroid space. Many molecular aspects of symbiosis have been studied, but the interbacterial and interdomain molecule trafficking by EVs in the peribacteroid space has not been questioned yet. Here, we unveil intensive EV trafficking within the symbiosome interface of several rhizobium-legume dual systems by developing a robust EV isolation procedure. We analyze the EV-encased proteomes from the peribacteroid space of each bacterium-host partnership, uncovering both conserved and differential traits of every symbiotic system. This study opens the gates for designing EV-based biotechnological tools for sustainable agriculture.