Symbiosome membrane-localized cationic amino acid transporters support symbiotic nitrogen fixation in Medicago truncatula

Legumes engage in nitrogen-fixing symbiosis with rhizobia, in which host legumes supply dicarboxylates as a carbon source to rhizobia, while rhizobia reciprocate by providing ammonium to the host plants. Beyond this classical model, accumulating evidence suggests that amino acid exchange is also essential for legume-rhizobium symbiosis. However, it remains unclear whether amino acid transporters are present on the symbiosome membrane (SM) to mediate amino acid exchange during symbiotic nitrogen fixation (SNF). In this study, we identified three amino acid transporters in Medicago truncatula-MtCAT1a, MtCAT1b, and MtCAT1c-which belong to a clade of the plant Cationic Amino acid Transporter (CAT) family known to transport a wide range of amino acids. Notably, MtCAT1b and MtCAT1c are predominantly expressed in infected nodule cells and localize to the SM. Genetic analyses further demonstrate that both MtCAT1b and MtCAT1c are required for amino acid exchange at the SM, with additional evidence indicating that bacteroid metabolism is disturbed in the mutants. Transport assays show that both MtCAT1b and MtCAT1c exhibit broad substrate specificity. Collectively, these findings identify MtCAT1b and MtCAT1c as key mediators of cross-kingdom amino acid exchange, which is essential for maintaining efficient SNF in root nodules.