Collaborative Research: Elucidating the Role of Rhizobial tRNA-Derived Fragments in Soybean Nodule Development and Symbiotic Nitrogen Fixation

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Since the Green Revolution of the 1960s, nitrogen fertilizers have been widely deployed to boost crop yields to meet the demands of the growing world population. However, excessive reliance on fertilizers leads to water, soil and air pollution; therefore, environmentally friendly sources of nitrogen are needed for sustainable agriculture. Legume crops such as soybean, common bean, and peanuts have the capability to harness atmospheric nitrogen for their growth by establishing symbiotic relationships with a group of nitrogen-fixing soil bacteria known as rhizobia. The symbiotic interactions begin with a process called nodulation that forms root nodules, within which rhizobia convert atmospheric nitrogen into ammonia that can be utilized by host plants – a process called symbiotic nitrogen fixation. Approximately 50-60% of the nitrogen needed for soybean production is provided by symbiotic nitrogen fixation, thus positioning soybean as an important component of the biennial rotation with corn in the US agricultural system. Both nodulation and nitrogen fixation require coordinated exchanges of molecular signals between the symbiotic partners, and transfer RNA-derived small RNA fragments produced in rhizobia have recently been found to be one type of molecule mediating rhizobia-host plant communications. This project will address how and to what extent such RNA fragments mediate the nodulation and nitrogen fixation processes. Such knowledge will facilitate the development of new biotechnological tools for optimizing the legume-rhizobia symbioses for sustainable crop production. This project will provide hands-on experience to underrepresented minority undergraduate and high school students through an annual “Bioinformatics in Agricultural Science” summer workshop.Plant-microbe interactions require cross-kingdom small RNA (sRNA) interference. While the roles of eukaryotic sRNAs in plant-fungus interactions have been well characterized, sRNA-mediated prokaryote-eukaryote communications have just begun to garner attention. Transfer RNA-derived sRNA fragments (tRFs) are often viewed as degradation products and thus have been largely ignored in previous studies. Recently, the Ma laboratory demonstrated that three rhizobial tRFs were able to regulate the expression of five soybean genes affecting nodulation via soybean Argonaute 1-guided mRNA cleavage. This project will integrate genomic, molecular, and biochemical tools to elucidate the extent to which rhizobial tRFs hijack the host cellular machineries to regulate nodulation and symbiotic nitrogen fixation (SNF) and to further dissect mechanisms by which rhizobial tRFs regulate these processes using soybean-Bradyrhizobium diazoefficiens symbiotic partners as an experimental system. In Aim 1, the dynamics of rhizobial tRFs abundance and putative target gene expression in developing, mature, and senescing nodules will be investigated; in Aim 2, the mechanisms of rhizobial tRF-host gene interactions and relative magnitudes will be determined; in Aim 3, the functions of a subset of tRFs and their target genes underlying nodule development and SNF will be characterized; in Aim 4, experimental soybean lines with quantitative variation of nodule traits will be developed through gene-editing and evaluated for nodulation and SNF efficiencies and the plant productivity. This project may shed light on the potential roles of tRFs in diverse host-microbe interactions, towards translation of the discoveries into practice for control of bacterial pathogens or to promote beneficial interactions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.