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Acknowledgements: We thank Anindya Kundu, Chai Hao Chiu, and Victor Hugo Moura De Souza for their valuable discussion and Eli Marable’s feedback on the early draft of this manuscript. We appreciate Jongho Sun for providing a calcium spiking trace for Fig 1., Our current food production systems are unsustainable, driven in part through the application of chemically fixed nitrogen. We need alternatives to empower farmers to maximise their productivity sustainably. Therefore, we explore the potential for transferring the root nodule symbiosis from legumes to other crops. Studies over the last decades have shown that preexisting developmental and signal transduction processes were recruited during the evolution of legume nodulation. This allows us to utilise these preexisting processes to engineer nitrogen fixation in target crops. Here, we highlight our understanding of legume nodulation and future research directions that might help to overcome the barrier of achieving self-fertilising crops.

Engineering signalling between plants and microbes could be exploited to establish host-specificity between plant-growth-promoting bacteria and target crops in the environment. We previously engineered rhizopine-signalling circuitry facilitating exclusive signalling between rhizopine-producing (RhiP) plants and model bacterial strains. Here, we conduct an in-depth analysis of rhizopine-inducible expression in bacteria. We characterize two rhizopine-inducible promoters and explore the bacterial host-range of rhizopine biosensor plasmids. By tuning the expression of rhizopine uptake genes, we also construct a new biosensor plasmid pSIR05 that has minimal impact on host cell growth in vitro and exhibits markedly improved stability of expression in situ on RhiP barley roots compared to the previously described biosensor plasmid pSIR02. We demonstrate that a sub-population of Azorhizobium caulinodans cells carrying pSIR05 can sense rhizopine and activate gene expression when colonizing RhiP barley roots. However, these bacteria were mildly defective for colonization of RhiP barley roots compared to the wild-type parent strain. This work provides advancement towards establishing more robust plant-dependent control of bacterial gene expression and highlights the key challenges remaining to achieve this goal.

Many plants associate with arbuscular mycorrhizal fungi for nutrient acquisition, while legumes also associate with nitrogen-fixing rhizobial bacteria. Both associations rely on symbiosis signaling and here we show that cereals can perceive lipochitooligosaccharides (LCOs) for activation of symbiosis signaling, surprisingly including Nod factors produced by nitrogen-fixing bacteria. However, legumes show stringent perception of specifically decorated LCOs, that is absent in cereals. LCO perception in plants is activated by nutrient starvation, through transcriptional regulation of Nodulation Signaling Pathway (NSP)1 and NSP2. These transcription factors induce expression of an LCO receptor and act through the control of strigolactone biosynthesis and the karrikin-like receptor DWARF14-LIKE. We conclude that LCO production and perception is coordinately regulated by nutrient starvation to promote engagement with mycorrhizal fungi. Our work has implications for the use of both mycorrhizal and rhizobial associations for sustainable productivity in cereals.

Plants associate with beneficial arbuscular mycorrhizal fungi facilitating nutrient acquisition. Arbuscular mycorrhizal fungi produce chitooligosaccharides (COs) and lipo-chitooligosaccharides (LCOs), that promote symbiosis signalling with resultant oscillations in nuclear-associated calcium. The activation of symbiosis signalling must be balanced with activation of immunity signalling, which in fungal interactions is promoted by COs resulting from the chitinaceous fungal cell wall. Here we demonstrate that COs ranging from CO4-CO8 can induce symbiosis signalling in Medicago truncatula. CO perception is a function of the receptor-like kinases MtCERK1 and LYR4, that activate both immunity and symbiosis signalling. A combination of LCOs and COs act synergistically to enhance symbiosis signalling and suppress immunity signalling and receptors involved in both CO and LCO perception are necessary for mycorrhizal establishment. We conclude that LCOs, when present in a mix with COs, drive a symbiotic outcome and this mix of signals is essential for arbuscular mycorrhizal establishment., Polysaccharide molecules chitooligosaccharides (COs) and peptidoglycan not only activate plant immunity but also trigger plant symbiosis signalling. Here the authors show that a combination of COs and lipochitooligosaccharides (LCOs) act synergistically to suppress immunity and promote symbiosis to facilitate beneficial fungal associations.

The root microbiota is critical for agricultural yield, with growth-promoting bacteria able to solubilise phosphate, produce plant growth hormones, antagonise pathogens and fix N2. Plants control the microorganisms in their immediate environment and this is at least in part through direct selection, the immune system, and interactions with other microorganisms. Considering the importance of the root microbiota for crop yields it is attractive to artificially regulate this environment to optimise agricultural productivity. Towards this aim we express a synthetic pathway for the production of the rhizopine scyllo-inosamine in plants. We demonstrate the production of this bacterial derived signal in both Medicago truncatula and barley and show its perception by rhizosphere bacteria, containing bioluminescent and fluorescent biosensors. This study lays the groundwork for synthetic signalling networks between plants and bacteria, allowing the targeted regulation of bacterial gene expression in the rhizosphere for delivery of useful functions to plants., The root microbiota is critical for promoting crop yield. Here, the authors create a synthetic pathway for the production of the rhizopine scyllo-inosamine in Medicago truncatula and barley, and show its perception by rhizosphere bacteria for targeted regulation of bacterial gene expression.

As primary producers, plants rely on a large aboveground surface area to collect carbon dioxide and sunlight and a large underground surface area to collect the water and mineral nutrients needed to support their growth and development. Accessibility of the essential nutrients nitrogen (N) and phosphorus (P) in the soil is affected by many factors that create a variable spatiotemporal landscape of their availability both at the local and global scale. Plants optimize uptake of the N and P available through modifications to their growth and development and engagement with microorganisms that facilitate their capture. The sensing of these nutrients, as well as the perception of overall nutrient status, shapes the plant's response to its nutrient environment, coordinating its development with microbial engagement to optimize N and P capture and regulate overall plant growth.