Abstract

Bacterial extracellular vesicles (bEVs) are emerging as key players in interkingdom communication, yet their role in delivering functional proteins to host cells during symbiosis remains unexplored. This study shows that Sinorhizobium fredii HH103 packages a PIN-like auxin transporter, AuxT, into bEVs that traffic within the peribacteroid space of soybean nodules. AuxT is chromosomally encoded and constitutively expressed, genetically uncoupled from the flavonoid-inducible auxin biosynthesis machinery located on the symbiotic plasmid. Structural prediction reveals that AuxT adopts an eight-transmembrane-helix architecture with striking homology to plant PIN auxin exporters, despite negligible sequence identity. Molecular docking demonstrates that AuxT binds indole-3-acetic acid within a central cavity, with dimerization inducing ligand-specific conformational changes consistent with transport activity. The auxT mutant exhibits significant symbiotic defects including reduced shoot biomass, nodule number, and nodule mass that are fully restored by complementation. Critically, AuxT-enriched bEVs contain elevated auxin levels, and nodules colonized by the complemented strain accumulate more auxin specifically within the bEVs peribacteroid space, while bacteroids themselves show no auxin retention. We postulate that bEV-associated AuxT mediates localized auxin export into the symbiosome, modulating the host hormonal environment to optimize symbiotic development. This work reveals a previously unrecognized mechanism of interdomain hormonal modulation, where a bacterium uses a structurally convergent transporter and vesicular delivery to actively shape host physiology and to improve the symbiotic performance.