| Nitrogen availability is a limiting factor for cereal crop production leading to an excessive use of fertilizers. Taking advantage of plant-microbe symbioses like legume-rhizobia symbiosis is one option for exploring a natural nitrogen source for cereal plants, such as rice. Legumes can develop a symbiotic association with soil bacteria, rhizobia, that leads to the formation of specialized root structures, nodules, inside which nitrogen fixation can occur. Unfortunately, this symbiosis is species-specific and is restricted only to legumes. Therefore, cereals are still heavily dependent on fertilizers for their nitrogen needs. Several studies have shown that plant hormones, like auxin, play important roles during legume-rhizobia symbiosis. For instance, application of auxin transport inhibitors to legume roots induces the formation of nodule-like structures (NLS) in the absence of rhizobia. Furthermore, these NLS can be colonized by rhizobia and successful nitrogen fixation can occur inside them. Interestingly, auxin also induces similar NLS in cereal roots. While several genetic studies have identified plant genes controlling NLS formation in legumes, unfortunately, no studies have investigated the genes involved in NLS formation in cereals. In this study, we were successful in inducing NLS in rice and Medicago roots, using auxin, 2,4-D, consistently at a high frequency (>90%) under controlled sterile conditions. These structures were characterized by a broad base, a diffuse meristem, and increased cell differentiation in the vasculature. Interestingly, they were found to be structurally similar in rice and Medicago, suggesting a similar developmental program across cereals and legumes. Genetic studies in rice and Medicago using available symbiotic mutants indicated that formation of NLS did not depend on the 'Common Symbiotic Pathway', which is required for both legume-rhizobia and arbuscular mycorrhizal symbioses. These results further suggested that a similar genetic program might control NLS formation across cereals and legumes. Next, we performed a comprehensive RNA-Sequencing experiment to identify the differentially expressed genes during NLS formation in rice. We identified genes at early and later stages of NLS formation. Interestingly, we identified several orthologs of genes, which have been shown to play a role in legume-rhizobia symbiosis. Finally, we validated the expression pattern of thirteen candidate genes belonging to five gene classes (hormone related, growth and development, transcription factors, receptor kinases, and transport related) using RT-PCR. Future efforts can focus on identifying and validating the expression pattern of additional candidate genes and genetically characterizing them. These studies will make important advances towards improving nitrogen fixation in cereals and will have a positive impact on sustainable agriculture worldwide. |
