The Lotus japonicus alpha Expansin EXPA1 is recruited during intracellular and intercellular rhizobial colonization

Most legumes establish a mutualistic association with rhizobia, a group of nitrogen-fixing bacteria. In Lotus japonicus, the symbiotic colonization occurs intracellularly, via root-hair infection threads by Mesorhizobium loti, or intercellularly, with Agrobacterium pusense IRBG74. In both mechanisms, cell wall remodeling is presumably an essential process. In plants, alpha-Expansins (EXPA) promote cell wall loosening by non-enzymatically triggering a pH-dependent relaxation. In this study, we show that LjEXPA1, is critical for the intracellular and intercellular symbiotic program in L. japonicus. Promoter activity and subcellular localization analyses revealed that EXPA1 is recruited at essential compartments and structures of epidermal and cortical cells in both mechanisms of rhizobial infection, such as the infection chambers, infection pockets and transcellular infection threads. Additionally, EXPA1-YFP abundantly accumulated in dividing cortical cells during nodule formation. The expression profile of EXPA1 correlates with the symbiotic phenotype observed in homozygous mutants disrupted in the EXPA1 gene (expA1-1 and expA1-2). Infection thread formation and intercellular colonization was drastically reduced in expA1-1 and expA1-2 mutants, respect to wild type plants. Similarly, nodule formation was significantly reduced in these mutants after M. loti or IRBG74 inoculation. Our results indicate that non-enzymatic cell wall remodeling by the alpha Expansin EXPA1 is crucial for the successful establishment of Lotus-rhizobia symbiosis, regardless of the infection mechanism.