Activation of Ca2+ oscillations in rice (Oryza sativa) root hairs expressing the legume-specific Nod factor receptor kinases MtNFP and MtLYK3

Abstract Nitrogen is a critical nutrient for rice production. Chemical fertilizers have been the main source of nitrogen for crop production, but their indiscriminate use has adversely affected human and environmental health. Legumes form symbiosis with rhizobia as well as arbuscular mycorrhizal fungi (AMF) to fix atmospheric nitrogen and uptake of phosphorus, respectively. Formation of symbiotic associations in legumes with rhizobia and AMF utilize a shared genetic network known as a common symbiotic signalling pathway (CSSP). Similar to legumes rice forms a symbiotic association only with AMF and possesses a similar CSSP that promotes the formation of arbuscular mycorrhizal symbiosis (AMS). Some vital genes for Nod factor (NF) perception are not conserved in rice, while a part of the genetic network for rhizobial symbiosis in legumes is conserved in rice. Hence, to make a symbiotic association with Rhizobium, we developed transgenic rice lines expressing legume nodulation-specific receptor kinase genes (MtLYK3 and MtNFP) and evaluated their ability to perceive rhizobial NF. Besides, we crossed these transgenic plants with others expressing the Cameleon calcium (Ca²⁺) sensor and assessed the NF-induced Ca²⁺ fluxes and spiking around nuclei in root hairs. Confocal microscopy of the root hairs in the hybrid transgenic plants revealed that NF can promote Ca²⁺ fluxes at root hair tips as well as Ca²⁺ spiking around nuclei. These results showed that the rice plants expressing MtLYK3 and MtNFP gained the ability to perceive NF and transmit nod signals appropriately to trigger calcium fluxes and spiking akin to that in legume roots.