Haruko Imaizumi-Anraku

The nuclear pore complex controls the movement of proteins into and out of the nucleus, allowing cells to regulate protein localization and abundance. This process influences how organisms respond to environmental stimuli. Components of the nuclear pore complex, including the NUP107-160 sub-complex, NUP133, NUP85, and NENA, are required for root nodulation and arbuscular mycorrhization in Lotus japonicus. However, the specific role of these nucleoporins in symbiotic signaling was poorly understood. Through reverse genetics, we discovered that NUP133 is also required for symbiosis in Medicago truncatula, although the mutant phenotypes were less pronounced than in Lotus. Overexpression of the symbiotic ion channels Medicago DMI1 and Lotus Castor and Pollux in the Lotus Ljnup133, Ljnup85, and Ljnena mutants partially alleviated the nodulation defects. Notably, in NUP107-160 sub-complex mutants of Lotus and Medicago, the accumulation of GFP-labeled Pollux and DMI1 on the inner nuclear membrane was reduced, indicating the NUP107-160 sub-complex plays a key role in regulating the distribution of DMI1 and Pollux on the nuclear envelope. This highlights the extreme sensitivity of nodulation in Lotus to changes in the abundance of Pollux on the inner nuclear membrane. In contrast, Medicago appears to exhibit greater tolerance to alterations in the distribution of DMI1 on the nuclear envelope.

Soybeans fix atmospheric N₂ through symbiosis with rhizobia. The relationship between rhizobia and soybeans, particularly those with high nitrous oxide (N₂O)-reducing (N₂OR) activities, can be leveraged to reduce N₂O emissions from agricultural soils. However, inoculating soybeans with these rhizobia under field conditions often fails because of the competition from indigenous rhizobia that possess low or no N₂OR activity. In this work, we utilize natural incompatibility systems between soybean and rhizobia to address this challenge. Specifically, Rj2 and GmNNL1 inhibit certain rhizobial infections in response to NopP, an effector protein. By combining a soybean line with a hybrid accumulation of the Rj2 and GmNNL1 genes and bradyrhizobia lacking the nopP gene, we develop a soybean-bradyrhizobial symbiosis system in which strains with high N₂OR activity predominantly infect. Our optimize symbiotic system substantially reduces N₂O emissions in field and laboratory tests, presenting a promising approach for sustainable agricultural practices.

Soybeans fix atmospheric N₂ through symbiosis with rhizobia, N₂-fixing bacteria. The relationship between rhizobia and soybeans, particularly those with high nitrous oxide (N₂O)-reducing activities (Nos⁺⁺), can be used for reducing N₂O emissions from agricultural soils. However, inoculating soybeans with Nos⁺⁺ rhizobia under field conditions often fails because of the competition from indigenous Nos⁻ (no N₂O-reducing activities) and Nos⁺ (normal N₂O-reducing activities) rhizobia. Here, we utilized natural incompatibility systems between soybean and rhizobia to address this challenge. Specifically, Rj2 and GmNNL1 inhibit certain rhizobial infections in response to NopP, an effector protein. By combining a soybean line with a hybrid accumulation of the Rj2 and GmNNL1 genes and Nos⁺⁺ bradyrhizobia lacking the nopP effector gene, we developed a soybean-bradyrhizobial symbiosis system that Nos⁺⁺ rhizobial inoculants predominantly infect. This optimized symbiotic system substantially reduced N₂O emissions in field and laboratory tests, presenting a promising approach for sustainable agricultural practices.

Co-inoculation of soybeans with Bradyrhizobium and plant growth-promoting bacteria has displayed promise for enhancing plant growth, but concrete evidence of its impact on soybean yields is limited. Therefore, this study assessed the comparative efficacy of two 1-aminocyclopropane-1-carboxylate deaminase-producing Pseudomonas species (OFT2 and OFT5) co-inoculated with Bradyrhizobium ottawaense (SG09) on the growth, physiology, nodulation efficiency, and grain yield of three major Japanese soybean cultivars: Enrei, Fukuyutaka, and Satonohohoemi. The experiments were conducted in a warehouse under natural light conditions. The treatments included the inoculation of SG09, SG09 + OFT2, and SG09 + OFT5. Compared with Bradyrhizobium inoculation alone, co-inoculation led to significant improvements in nodulation efficiency, growth, and physiological performance in the Enrei and Fukuyutaka cultivars, but not in the Satonohohoemi cultivar. Furthermore, co-inoculation significantly boosted the total nitrogen content and ion uptake in the shoots, ultimately leading to a remarkable improvement in the grain yield in the Enrei and Fukuyutaka cultivars. These findings contribute to clarifying the interplay among Bradyrhizobium, Pseudomonas, and the plant host cultivar. Notably, Bradyrhizobium–Pseudomonas co-inoculation represents a potentially effective biofertilization strategy for soybean production, highlighting promising avenues for sustainable agricultural practices.

1. In addition to nitrogen (N), phosphorus (P) is a limiting factor for plant growth in many terrestrial ecosystems. To cope with the shortage of P, plants have evolved to adapt in various ways, in...

We investigated the potential dual application of two Bradyrhizobium strains (B. diazoefficiens USDA110 and B. ottawaense SG09) and plant growth-promoting bacteria, PGPB (Pseudomonas spp.: OFT2 and OFT5), to improve nodulation and N₂-fixation in soybean plants. The growth-promoting effects of dual inoculation were observed on plant growth, physiology, and nodulation of soybean under normal conditions compared with plants individually inoculated with either USDA110 or SG09. Both OFT2 and OFT5 promoted N₂-fixation by 11% and 56%, respectively, when dual inoculation with USDA110 and by 76% and 81%, respectively, when dual inoculation with SG09. Salinity stress significantly reduces soybean growth, physiology, nutrient uptake, nodulation, and N₂-fixation. However, these adverse effects were attenuated by the dual inoculation of PGPB and rhizobia depending on the combination of inoculants. In particular, dual inoculation of PGPB with SG09 was more effective in enhancing the salt tolerance of soybean by reducing salt-induced ethylene production and improving nutrient uptake. However, no such effect was observed with the combined inoculation of USDA110 and OFT5. An effective symbiotic association between SG09 and two Pseudomonas bacteria can be considered a beneficial approach to improving the symbiotic efficiency of nodulation and mitigating salinity stress in soybeans.

Most plants interact with various soil microorganisms as they grow through the soil. Root nodule symbiosis by legumes and rhizobia is a well-known phenomenon of plant–microbe interactions in the soil. Although microscopic observations are useful for understanding the infection processes of rhizobia, nondestructive observation methods have not been established for monitoring interactions between rhizobia and soil-grown roots. In this study, we constructed Bradyrhizobium diazoefficiens strains that constitutively express different fluorescent proteins, which allows identification of tagged rhizobia by the type of fluorophores. In addition, we constructed a plant cultivation device, Rhizosphere Frame (RhizoFrame), which is a soil-filled container made of transparent acrylic plates that allows observation of roots growing along the acrylic plates. Combining fluorescent rhizobia with RhizoFrame, we established a live imaging system, RhizoFrame system, that enabled us to track the nodulation processes with fluorescence stereomicroscope while retaining spatial information about roots, rhizobia, and soil. Mixed inoculation with different fluorescent rhizobia using RhizoFrame enabled the visualization of mixed infection of a single nodule with two strains. In addition, observation of transgenic Lotus japonicus expressing auxin-responsive reporter genes indicated that RhizoFrame system could be used for a real-time and nondestructive reporter assay. Thus, the use of RhizoFrame system is expected to enhance the study of the spatiotemporal dynamics of plant–microbe interactions in the soil.

The original article has been updated.

Spatiotemporal features of anthocyanin accumulation in a model legume Lotus japonicus (Regel) K.Larsen were elucidated to develop criteria for the genetic analysis of flavonoid biosynthesis. Artificial mutants and wild accessions, with lower anthocyanin accumulation in the stem than the standard wild type (B-129 ‘Gifu’), were obtained by ethyl methanesulfonate (EMS) mutagenesis and from a collection of wild-grown variants, respectively. The loci responsible for the green stem of the mutants were named as VIRIDICAULIS (VIC). Genetic and chemical analysis identified two loci, namely, VIC1 and VIC2, required for the production of both anthocyanins and proanthocyanidins (condensed tannins), and two loci, namely, VIC3 and VIC4, required for the steps specific to anthocyanin biosynthesis. A mutation in VIC5 significantly reduced the anthocyanin accumulation. These mutants will serve as a useful system for examining the effects of anthocyanins and proanthocyanidins on the interactions with herbivorous pests, pathogenic microorganisms and nitrogen-fixing symbiotic bacteria, Mesorhizobium loti.

Achlorophylous and early developmental stages of chorolophylous orchids are highly dependent on carbon and other nutrients provided by mycorrhizal fungi, in a nutritional mode termed mycoheterotrophy. Previous findings have implied that some common properties at least partially underlie the mycorrhizal symbioses of mycoheterotrophic orchids and that of autotrophic arbuscular mycorrhizal (AM) plants; however, information about the molecular mechanisms of the relationship between orchids and their mycorrhizal fungi is limited. In this study, we characterized the molecular basis of an orchid-mycorrhizal (OM) symbiosis by analyzing the transcriptome of Bletilla striata at an early developmental stage associated with the mycorrhizal fungus Tulasnella sp. The essential components required for the establishment of mutual symbioses with AM fungi or rhizobia in most terrestrial plants were identified from the B. striata gene set. A cross-species gene complementation analysis showed one of the component genes, calcium and calmodulin-dependent protein kinase gene CCaMK in B. striata, retains functional characteristics of that in AM plants. The expression analysis revealed the activation of homologs of AM-related genes during the OM symbiosis. Our results suggest that orchids possess, at least partly, the molecular mechanisms common to AM plants.