Abstract
ENOD40 is induced most early in response to interaction of legume plants with Rhizobium bacteria. Lotus japonicus has two ENOD40 genes, LjENOD40-1 and -2 closely related each other. We generated transgenic lines of L. japonicus with a construct that expresses hairpin double-stranded RNA for LjENOD40-1 to induce sequence-specific RNA silencing. In the transgenic plants, expression of both LjENOD40-1 and -2 was significantly reduced, and no accumulation of ENOD40 transcripts was detected upon M. loti inoculation. The transgenic plants exhibited very poor nodulation (only 0-2 nodules/plant). Microscopic observation of the infection process using lacZ-labelled M. loti, together with expression analysis of infection-related nodulin genes, demonstrated that ENOD40 knockdown did not inhibit bacterial infection process. In contrast, nodule primordium initiation and subsequent nodule development were significantly suppressed in the transgenic plants. These results clearly indicate that ENOD40 is required for nodule initiation and subsequent organogenesis, but not involved in early infection events.
