Legumes can enter into symbiotic relationships with both nitrogen-fixing bacteria (rhizobia) and mycorrhizal fungi. Nodulation by rhizobia results from a signal transduction pathway induced in legume roots by rhizobial Nod factors. DMI3 ,a Medicago truncatula gene that acts immediately downstream of calcium spiking in this signaling pathway and is required for both nodulation and mycorrhizal infection, has high sequence similarity to genes encoding calcium and calmodulin-dependent protein kinases (CCaMKs). This indicates that calcium spiking is likely an essential component of the signaling cascade leading to nodule development and mycorrhizal infection, and sheds light on the biological role of plant CCaMKs.

Dependent Protein Kinase Required for Bacterial and Fungal Symbioses

Humans are currently confronted by many global challenges. These include achieving food security for a rapidly expanding population, lowering the risk of climate change by reducing the net release of greenhouse gases into the atmosphere due to human activity, and meeting the increasing demand for energy in the face of dwindling reserves of fossil energy and uncertainties about future reliability of supply. Legumes deliver several important services to societies. They provide important sources of oil, fiber, and protein-rich food and feed while supplying nitrogen (N) to agro-ecosystems via their unique ability to fix atmospheric N2 in symbiosis with the soil bacteria rhizobia, increasing soil carbon content, and stimulating the productivity of the crops that follow. However, the role of legumes has rarely been considered in the context of their potential to contribute to the mitigation of climate change by reducing fossil fuel use or by providing feedstock for the emerging biobased economies where fossil sources of energy and industrial raw materials are replaced in part by sustainable and renewable biomass resources. The aim of this review was to collate the current knowledge regarding the capacity of legumes to (1) lower the emissions of the key greenhouse gases carbon dioxide (CO2) and nitrous oxide (N2O) compared to N-fertilized systems, (2) reduce the fossil energy used in the production of food and forage, (3) contribute to the sequestration of carbon (C) in soils, and (4) provide a viable source of biomass for the generation of biofuels and other materials in future biorefinery concepts. We estimated that globally between 350 and 500 Tg CO2 could be emitted as a result of the 33 to 46 Tg N that is biologically fixed by agricultural legumes each year. This compares to around 300 Tg CO2 released annually from the manufacture of 100 Tg fertilizer N. The main difference is that the CO2 respired from the nodulated roots of N2-fixing legumes originated from photosynthesis and will not represent a net contribution to atmospheric concentrations of CO2, whereas the CO2 generated during the synthesis of N fertilizer was derived from fossil fuels. Experimental measures of total N2O fluxes from legumes and N-fertilized systems were found to vary enormously (0.03–7.09 and 0.09–18.16 kg N2O–N ha−1, respectively). This reflected the data being collated from a diverse range of studies using different rates of N inputs, as well as the large number of climatic, soil, and management variables known to influence denitrification and the portion of the total N lost as N2O. Averages across 71 site-years of data, soils under legumes emitted a total of 1.29 kg N2O–N ha−1 during a growing season. This compared to a mean of 3.22 kg N2O–N ha−1 from 67 site-years of N-fertilized crops and pastures, and 1.20 kg N2O–N ha−1 from 33 site-years of data collected from unplanted soils or unfertilized non-legumes. It was concluded that there was little evidence that biological N2 fixation substantially contributed to total N2O emissions, and that losses of N2O from legume soil were generally lower than N-fertilized systems, especially when commercial rates of N fertilizer were applied. Elevated rates of N2O losses can occur following the termination of legume-based pastures, or where legumes had been green- or brown-manured and there was a rapid build-up of high concentrations of nitrate in soil. Legume crops and legume-based pastures use 35% to 60% less fossil energy than N-fertilized cereals or grasslands, and the inclusion of legumes in cropping sequences reduced the average annual energy usage over a rotation by 12% to 34%. The reduced energy use was primarily due to the removal of the need to apply N fertilizer and the subsequently lower N fertilizer requirements for crops grown following legumes. Life cycle energy balances of legume-based rotations were also assisted by a lower use of agrichemicals for crop protection as diversification of cropping sequences reduce the incidence of cereal pathogens and pests and assisted weed control, although it was noted that differences in fossil energy use between legumes and N-fertilized systems were greatly diminished if energy use was expressed per unit of biomass or grain produced. For a change in land use to result in a net increase C sequestration in soil, the inputs of C remaining in plant residues need to exceed the CO2 respired by soil microbes during the decomposition of plant residues or soil organic C, and the C lost through wind or water erosion. The net N-balance of the system was a key driver of changes in soil C stocks in many environments, and data collected from pasture, cropping, and agroforestry systems all indicated that legumes played a pivotal role in providing the additional organic N required to encourage the accumulation of soil C at rates greater than can be achieved by cereals or grasses even when they were supplied with N fertilizer. Legumes contain a range of compounds, which could be refined to produce raw industrial materials currently manufactured from petroleum-based sources, pharmaceuticals, surfactants, or food additives as valuable by-products if legume biomass was to be used to generate biodiesel, bioethanol, biojet A1 fuel, or biogas. The attraction of using leguminous material feedstock is that they do not need the inputs of N fertilizer that would otherwise be necessary to support the production of high grain yields or large amounts of plant biomass since it is the high fossil energy use in the synthesis, transport, and application of N fertilizers that often negates much of the net C benefits of many other bioenergy sources. The use of legume biomass for biorefineries needs careful thought as there will be significant trade-offs with the current role of legumes in contributing to the organic fertility of soils. Agricultural systems will require novel management and plant breeding solutions to provide the range of options that will be required to mitigate climate change. Given their array of ecosystem services and their ability to reduce greenhouse gas emissions, lower the use of fossil energy, accelerate rates of C sequestration in soil, and provide a valuable source of feedstock for biorefineries, legumes should be considered as important components in the development of future agroecosystems.

/pdf/legumes-for-mitigation-of-climate-change-and-the-provision-jfs23ve9qk.pdf

Legumes for mitigation of climate change and the provision of feedstock for biofuels and biorefineries. A review

Despite long-standing observations on diverse cytokinin actions, the discovery path to cytokinin signaling mechanisms was tortuous. Unyielding to conventional genetic screens, experimental innovations were paramount in unraveling the core cytokinin signaling circuitry, which employs a large repertoire of genes with overlapping and specific functions. The canonical two-component transcription circuitry involves His kinases that perceive cytokinin and initiate signaling, as well as His-to-Asp phosphorelay proteins that transfer phosphoryl groups to response regulators, transcriptional activators, or repressors. Recent advances have revealed the complex physiological functions of cytokinins, including interactions with auxin and other signal transduction pathways. This review begins by outlining the historical path to cytokinin discovery and then elucidates the diverse cytokinin functions and key signaling components. Highlights focus on the integration of cytokinin signaling components into regulatory networks in specific contexts, ranging from molecular, cellular, and developmental regulations in the embryo, root apical meristem, shoot apical meristem, stem and root vasculature, and nodule organogenesis to organismal responses underlying immunity, stress tolerance, and senescence.

/pdf/cytokinin-signaling-networks-3428h56930.pdf

Cytokinin Signaling Networks

Legumes are highly important food, feed and biofuel crops. With few exceptions, they can enter into an intricate symbiotic relationship with specific soil bacteria called rhizobia. This interaction results in the formation of a new root organ called the nodule in which the rhizobia convert atmospheric nitrogen gas into forms of nitrogen that are useable by the plant. The plant tightly controls the number of nodules it forms, via a complex root-to-shoot-to-root signaling loop called autoregulation of nodulation (AON). This regulatory process involves peptide hormones, receptor kinases and small metabolites. Using modern genetic and genomic techniques, many of the components required for nodule formation and AON have now been isolated. This review addresses these recent findings, presents detailed models of the nodulation and AON processes, and identifies gaps in our understanding of these process that have yet to be fully explained.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1744-7909.2010.00899.x

Molecular Analysis of Legume Nodule Development and Autoregulation

https://web.nmsu.edu/~plantgen/supplemental_reading_files/j.1744-7909.2010.00899.x.pdf

Invited Expert Review Molecular Analysis of Legume Nodule Development and Autoregulation

This protocol is used to induce transgenic roots on soybean to study the function of genes required in biological processes of the root. Young seedlings with unfolded cotyledons are infected at the cotyledonary node and/or hypocotyl with Agrobacterium rhizogenes carrying the gene construct to be tested and the infection sites are kept in an environment of high humidity. When the emerged hairy roots can support the plants, the main roots are removed and the transgenic roots can be tested. Using this method, almost 100% of the infected plants form hairy roots within 1 month from the start of the experiments.

Agrobacterium rhizogenes -mediated transformation of soybean to study root biology

Two allelic non-nodulating mutants, nod49 and rj1, were characterized using map-based cloning and candidate gene approaches, and genetic complementation. From our results we propose two highly related lipo-oligochitin LysM-type receptor kinase genes (GmNFR1α and GmNFR1β) as putative Nod factor receptor components in soybean. Both mutants contained frameshift mutations in GmNFR1α that would yield protein truncations. Both mutants contained a seemingly functional GmNFR1β homeologue, characterized by a 374-bp deletion in intron 6 and 20-100 times lower transcript levels than GmNFR1α, yet both mutants were unable to form nodules. Mutations in GmNFR1β within other genotypes had no defects in nodulation, showing that GmNFR1β was redundant. Transgenic overexpression of GmNFR1α, but not of GmNFR1β, increased nodule number per plant, plant nitrogen content and the ability to form nodules with restrictive, ultra-low Bradyrhizobium japonicum titres in transgenic roots of both nod49 and rj1. GmNFR1α overexpressing roots also formed nodules in nodulation-restrictive acid soil (pH 4.7). Our results show that: (i) NFR1α expression controls nodule number in soybean, and (ii) acid soil tolerance for nodulation and suppression of nodulation deficiency at low titre can be achieved by overexpression of GmNFR1α.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1365-313X.2010.04398.x

Nodulation factor receptor kinase 1α controls nodule organ number in soybean (Glycine max L. Merr)

Chemically induced non-nodulating nod139 and nn5 mutants of soybean (Glycine max) show no visible symptoms in response to rhizobial inoculation. Both exhibit recessive Mendelian inheritance suggesting loss of function. By allele determination and genetic complementation in nod139 and nn5, two highly related lipo-oligochitin LysM-type receptor kinase genes in Glycine max were cloned; they are presumed to be the critical nodulation-inducing (Nod) factor receptor similar to those of Lotus japonicus, pea and Medicago truncatula. These duplicated receptor genes were called GmNFR5alpha and GmNFR5beta. Nonsense mutations in GmNFR5alpha and GmNFR5beta were genetically complemented by both wild-type GmNFR5alpha and GmNFR5beta in transgenic roots, indicating that both genes are functional. Both genes lack introns. In cultivar Williams82 GmNFR5alpha is located in chromosome 11 and in tandem with GmLYK7 (a related LysM receptor kinase gene), while GmNFR5beta is in tandem with GmLYK4 in homologous chromosome 1, suggesting ancient synteny and regional segmental duplication. Both genes are wild type in G. soja CPI100070 and Harosoy63; however, a non-functional NFR5beta allele (NFR5beta*) was discovered in parental lines Bragg and Williams, which harbored an identical 1,407 bp retroelement-type insertion. This retroelement (GmRE-1) and related sequences are located in several soybean genome positions. Paradoxically, putatively unrelated soybean cultivars shared the same insertion, suggesting a smaller than anticipated genetic base in this crop. GmNFR5alpha but not GmNFR5beta* was expressed in inoculated and uninoculated tap and lateral root portions at about 10-25% of GmATS1 (ATP synthase subunit 1), but not in trifoliate leaves and shoot tips.

Inactivation of duplicated nod factor receptor 5 (NFR5) genes in recessive loss-of-function non-nodulation mutants of allotetraploid soybean (Glycine max L. Merr.).

Arief Indrasumunar

Papers

Molecular Analysis of Legume Nodule Development and Autoregulation

Invited Expert Review Molecular Analysis of Legume Nodule Development and Autoregulation

Agrobacterium rhizogenes -mediated transformation of soybean to study root biology

Nodulation factor receptor kinase 1α controls nodule organ number in soybean (Glycine max L. Merr)

Inactivation of duplicated nod factor receptor 5 (NFR5) genes in recessive loss-of-function non-nodulation mutants of allotetraploid soybean (Glycine max L. Merr.).