MicroRNA 4407 modulates nodulation in soybean by repressing a root-specific ISOPENTENYLTRANSFERASE (GmIPT3).

MicroRNAs (miRNAs) are important regulators of plant biological processes, including soybean nodulation. One miRNA, miR4407, was identified in soybean roots and nodules. However, the function of miR4407 in soybean is still unknown. MiR4407, unique to soybean, positively regulates lateral root emergence and root structures and represses a root-specific ISOPENTENYLTRANSFERASE (GmIPT3). By altering the expression of miR4407 and GmIPT3, we investigated the role of miR4407 in lateral root and nodule development. Both miR4407 and GmIPT3 are expressed in the inner root cortex and nodule primordia. Upon rhizobial inoculation, miR4407 was downregulated while GmIPT3 was upregulated. Overexpressing miR4407 reduced the number of nodules in transgenic soybean hairy roots while overexpressing the wild-type GmIPT3 or a miR4407-resistant GmIPT3 mutant (mGmIPT3) significantly increased the nodule number. The mechanism of miR4407 and GmIPT3 functions was also linked to autoregulation of nodulation (AON), where miR4407 overexpression repressed miR172c and activated its target, GmNNC1, turning on AON. Exogenous CK mimicked the effects of GmIPT3 overexpression on miR172c, supporting the notion that GmIPT3 regulates nodulation by enhancing root-derived CK. Overall, our data revealed a new miRNA-mediated regulatory mechanism of nodulation in soybean. MiR4407 showed a dual role in lateral root and nodule development.

Differential RNA Editing and Intron Splicing in Soybean Mitochondria during Nodulation.

Nitrogen fixation in soybean consumes a tremendous amount of energy, leading to substantial differences in energy metabolism and mitochondrial activities between nodules and uninoculated roots. While C-to-U RNA editing and intron splicing of mitochondrial transcripts are common in plant species, their roles in relation to nodule functions are still elusive. In this study, we performed RNA-seq to compare transcript profiles and RNA editing of mitochondrial genes in soybean nodules and roots. A total of 631 RNA editing sites were identified on mitochondrial transcripts, with 12% or 74 sites differentially edited among the transcripts isolated from nodules, stripped roots, and uninoculated roots. Eight out of these 74 differentially edited sites are located on the matR transcript, of which the degrees of RNA editing were the highest in the nodule sample. The degree of mitochondrial intron splicing was also examined. The splicing efficiencies of several introns in nodules and stripped roots were higher than in uninoculated roots. These include nad1 introns 2/3/4, nad4 intron 3, nad5 introns 2/3, cox2 intron 1, and ccmFc intron 1. A greater splicing efficiency of nad4 intron 1, a higher NAD4 protein abundance, and a reduction in supercomplex I + III2 were also observed in nodules, although the causal relationship between these observations requires further investigation.

Using genomic information to improve soybean adaptability to climate change.

Climate change has brought severe challenges to agriculture. It is anticipated that there will be a drop in crop yield - including that of soybean - due to climatic stress factors that include drastic fluctuations in temperature, drought, flooding and high salinity. Genomic information on soybean has been accumulating rapidly since initial publication of its reference genome, providing a valuable tool for the improvement of cultivated soybean. Not only are many molecular markers that are associated with important quantitative trait loci now identified, but we also have a more detailed picture of the genomic variations among soybean germplasms, enabling us to utilize these as tools to assist crop breeding. In this review, we will summarize and discuss the currently available soybean genomic approaches, including whole-genome sequencing, sequencing-based genotyping, functional genomics, proteomics, and epigenomics. The information uncovered through these techniques will help further pinpoint important gene candidates and genomic loci associated with adaptive traits, as well as achieving a better understanding of how soybeans cope with the changing climate.

Roles of non-coding RNAs in the hormonal and nutritional regulation in nodulation and nitrogen fixation.

Symbiotic nitrogen fixation is an important component in the nitrogen cycle and is a potential solution for sustainable agriculture. It is the result of the interactions between the plant host, mostly restricted to legume species, and the rhizobial symbiont. From the first encounter between the host and the symbiont to eventual successful nitrogen fixation, there are delicate processes involved, such as nodule organogenesis, rhizobial infection thread progression, differentiation of the bacteroid, deregulation of the host defense systems, and reallocation of resources. All these processes are tightly regulated at different levels. Recent evidence revealed that non-coding RNAs (ncRNAs), including microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), participate in these processes by controlling the transcription and translation of effector genes. In general, ncRNAs are functional transcripts without translation potential and are important gene regulators. MiRNAs, negative gene regulators, bind to the target mRNAs and repress protein production by causing the cleavage of mRNA and translational silencing. LncRNAs affect the formation of chromosomal loops, DNA methylation, histone modification, and alternative splicing to modulate gene expression. Both lncRNAs and circRNAs could serve as target mimics of miRNA to inhibit miRNA functions. In this review, we summarized and discussed the current understanding of the roles of ncRNAs in legume nodulation and nitrogen fixation in the root nodule, mainly focusing on their regulation of hormone signal transduction, the autoregulation of nodulation (AON) pathway and nutrient homeostasis in nodules. Unraveling the mediation of legume nodulation by ncRNAs will give us new insights into designing higher-performance leguminous crops for sustainable agriculture.

The Identification of MATE Antisense Transcripts in Soybean Using Strand-Specific RNA-Seq Datasets.

Natural antisense transcripts (NATs) have been generally reported as negative regulators of their sense counterparts. Multidrug and toxic compound extrusion (MATE) proteins mediate the transport of various substrates. Although MATEs have been identified genome-wide in various plant species, their transcript regulators remain unclear. Here, using the publicly available strand-specific RNA-seq datasets of Glycine soja (wild soybean) which have the data from various tissues including developing pods, developing seeds, embryos, cotyledons and hypocotyls, roots, apical buds, stems, and flowers, we identified 35 antisense transcripts of MATEs from 28 gene loci after transcriptome assembly. Spearman correlation coefficients suggested the positive expression correlations of eight MATE antisense and sense transcript pairs. By aligning the identified transcripts with the reference genome of Glycine max (cultivated soybean), the MATE antisense and sense transcript pairs were identified. Using soybean C08 (Glycine max), in developing pods and seeds, the positive correlations between MATE antisense and sense transcript pairs were shown by RT-qPCR. These findings suggest that soybean antisense transcripts are not necessarily negative transcription regulators of their sense counterparts. This study enhances the existing knowledge on the transcription regulation of MATE transporters by uncovering the previously unknown MATE antisense transcripts and their potential synergetic effects on sense transcripts.

Differentially expressed microRNAs that target functional genes in mature soybean nodules.

MicroRNAs (miRNAs) are important regulators of biological functions in plants. To find out what roles miRNAs play in regulating symbiotic nitrogen fixation (SNF) in soybean [Glycine max (L.) Merr.], we identified high-confidence differentially expressed (DE) miRNAs from uninoculated roots (UR), rhizobium-inoculated roots (IR), and nodules (NODs) of soybean by robust small RNA sequencing (sRNA-seq). Based on their predicted target messenger RNAs (mRNAs), the expression profiles of some of these DE miRNAs could be linked to nodule functions. In particular, several miRNAs associated with nutrient transportation genes were differentially expressed in IRs and mature NODs. MiR399b, specifically, was highly induced in IRs and NODs, as well as by inorganic phosphate (Pi) starvation. In composite soybean plants overexpressing miR399b, PHOSPHATE2 (PHO2), a known target of miR399b that inhibits the activities of high-affinity Pi transporters, was strongly repressed. In addition, the overexpression of miR399b in the roots of transgenic composite plants significantly improved whole-plant Pi and ureide concentrations and the overall growth in terms of leaf node numbers and whole-plant dry weight. Our findings suggest that the induction of miR399b in NODs could enhance nitrogen fixation and soybean growth, possibly via improving Pi uptake to achieve a better Pi-nitrogen balance to promote SNF in nodules.

Fabricating cellulose nanofibril from licorice residues and its cellulose composite incorporated with natural nanoparticles.

It is important to make full utilization of industrial biomass residues. Pulp was prepared from licorice residues by soda-anthraquinone pulping followed by peroxyacetic acid bleaching. Cellulose nanofibril was obtained by enzymatic pretreatment followed by homogenization of the pulp (ETCNF). The effects of enzymatic pretreatment on ETCNF were investigated. Chitosan nanofibril (CHN) and lignin nanoparticles (LNPs) were prepared and used for ETCNF composites, respectively. The results showed that ETCNF exhibited clear nanofibrillar structure and a highly relative colloidal stability, and a much higher crystallinity index and thermal stability compared to TEMPO-medicated oxidized one; the cellulose composite films incorporated with CHN or LNPs exhibited good thermal stability and hydrophobicity. Compared with ETCNF film, ETCNF@LNPs-5.0% film showed higher UV-blocking ability and thermal stability, but reduced light transmittance, while ETCNF@CHN-5.0% film showed improved mechanical properties and similar light transmittance. This study would expend licorice residues as potential materials for CNF and its applications.

A reference-grade wild soybean genome.

Efficient crop improvement depends on the application of accurate genetic information contained in diverse germplasm resources. Here we report a reference-grade genome of wild soybean accession W05, with a final assembled genome size of 1013.2 Mb and a contig N50 of 3.3 Mb. The analytical power of the W05 genome is demonstrated by several examples. First, we identify an inversion at the locus determining seed coat color during domestication. Second, a translocation event between chromosomes 11 and 13 of some genotypes is shown to interfere with the assignment of QTLs. Third, we find a region containing copy number variations of the Kunitz trypsin inhibitor (KTI) genes. Such findings illustrate the power of this assembly in the analysis of large structural variations in soybean germplasm collections. The wild soybean genome assembly has wide applications in comparative genomic and evolutionary studies, as well as in crop breeding and improvement programs.

Interaction and Regulation of Carbon, Nitrogen, and Phosphorus Metabolisms in Root Nodules of Legumes.

Members of the plant family Leguminosae (Fabaceae) are unique in that they have evolved a symbiotic relationship with rhizobia (a group of soil bacteria that can fix atmospheric nitrogen). Rhizobia infect and form root nodules on their specific host plants before differentiating into bacteroids, the symbiotic form of rhizobia. This complex relationship involves the supply of C4-dicarboxylate and phosphate by the host plants to the microsymbionts that utilize them in the energy-intensive process of fixing atmospheric nitrogen into ammonium, which is in turn made available to the host plants as a source of nitrogen, a macronutrient for growth. Although nitrogen-fixing bacteroids are no longer growing, they are metabolically active. The symbiotic process is complex and tightly regulated by both the host plants and the bacteroids. The metabolic pathways of carbon, nitrogen, and phosphate are heavily regulated in the host plants, as they need to strike a fine balance between satisfying their own needs as well as those of the microsymbionts. A network of transporters for the various metabolites are responsible for the trafficking of these essential molecules between the two partners through the symbiosome membrane (plant-derived membrane surrounding the bacteroid), and these are in turn regulated by various transcription factors that control their expressions under different environmental conditions. Understanding this complex process of symbiotic nitrogen fixation is vital in promoting sustainable agriculture and enhancing soil fertility.