The soybean NADPH oxidase gene GmRbohK is crucial for nodule formation in symbiosis

Plants regulate various biological activities in cells by producing reactive oxygen species (ROS) as signal molecules. Plant NADPH oxidases, respiratory burst oxidase homologues (Rbohs), are the main source of ROS and participate in a variety of plant physiological processes. However, there are few studies regarding the function of Rbohs in rhizobium–legume symbiotic nodulation. A Rbohs subfamily gene GmRbohK was cloned and the gene expression and its subcellular localization were studied. The GmRbohK gene was highly expressed in soybean roots and nodules and greatly induced by rhizobia. GmRbohK was located at the cell membrane of Arabidopsis protoplasts. Moreover, RNA interference (RNAi) of GmRbohK resulted in a decrease in the number of nodules and nitrogenase activity. Meanwhile, the expression levels of nodulation marker genes decreased in association with the decrease in GmRbohK expression level. RNAi of GmRbohK inhibited the production of ROS and blocked infection events at the initial stage of nodule formation. Further analysis of nodule tissue sections revealed that RNAi of GmRbohK significantly reduced the number of bacteroids in the nodule infection area. In conclusion, GmRbohK is heavily involved in the regulation of soybean nodule formation.