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Abstract Background Root nodule symbiosis (RNS) is a fascinating evolutionary event. Given that limited genes conferring the evolution of RNS in Leguminosae have been functionally validated, the genetic basis of the evolution of RNS remains largely unknown. Identifying the genes involved in the evolution of RNS will help to reveal the mystery. Results Here, we investigate the gene loss event during the evolution of RNS in Leguminosae through phylogenomic and synteny analyses in 48 species including 16 Leguminosae species. We reveal that loss of the Lateral suppressor gene, a member of the GRAS-domain protein family, is associated with the evolution of RNS in Leguminosae. Ectopic expression of the Lateral suppressor (Ls) gene from tomato and its homolog MONOCULM 1 (MOC1) and Os7 from rice in soybean and Medicago truncatula result in almost completely lost nodulation capability. Further investigation shows that Lateral suppressor protein, Ls, MOC1, and Os7 might function through an interaction with NODULATION SIGNALING PATHWAY 2 (NSP2) and CYCLOPS to repress the transcription of NODULE INCEPTION (NIN) to inhibit the nodulation in Leguminosae. Additionally, we find that the cathepsin H (CTSH), a conserved protein, could interact with Lateral suppressor protein, Ls, MOC1, and Os7 and affect the nodulation. Conclusions This study sheds light on uncovering the genetic basis of the evolution of RNS in Leguminosae and suggests that gene loss plays an essential role.

Abstract Seed size/weight plays an important role in determining crop yield, yet only few genes controlling seed size have been characterized in soybean. Here, we perform a genome-wide association study and identify a major quantitative trait locus (QTL), named GmSW17 (Seed Width 17), on chromosome 17 that determine soybean seed width/weight in natural population. GmSW17 encodes a ubiquitin-specific protease, an ortholog to UBP22, belonging to the ubiquitin-specific protease (USPs/UBPs) family. Further functional investigations reveal that GmSW17 interacts with GmSGF11 and GmENY2 to form a deubiquitinase (DUB) module, which influences H2Bub levels and negatively regulates the expression of GmDP-E2F-1, thereby inhibiting the G1-to-S transition. Population analysis demonstrates that GmSW17 undergo artificial selection during soybean domestication but has not been fixed in modern breeding. In summary, our study identifies a predominant gene related to soybean seed weight, providing potential advantages for high-yield breeding in soybean.

Abstract The establishment of symbiotic interactions between leguminous plants and rhizobia requires complex cellular programming activated by Rhizobium Nod factors (NFs) as well as type III effector (T3E)-mediated symbiotic signaling. However, the mechanisms by which different signals jointly affect symbiosis are still unclear. Here we describe the mechanisms mediating the cross-talk between the broad host range rhizobia Sinorhizobium fredii HH103 T3E Nodulation Outer Protein L (NopL) effector and NF signaling in soybean. NopL physically interacts with the Glycine max Remorin 1a (GmREM1a) and the NFs receptor NFR5 (GmNFR5) and promotes GmNFR5 recruitment by GmREM1a. Furthermore, NopL and NF influence the expression of GmRINRK1, a receptor-like kinase (LRR-RLK) ortholog of the Lotus RINRK1, that mediates NF signaling. Taken together, our work indicates that S. fredii NopL can interact with the NF signaling cascade components to promote the symbiotic interaction in soybean.

The automated collection of plant phenotypic information has become a trend in breeding and smart agriculture. Four YOLOv8-based models were used to segment mature soybean plants placed in a simple background in a laboratory environment, identify pods, distinguish the number of soybeans in each pod, and obtain soybean phenotypes. The YOLOv8-Repvit model yielded the most optimal recognition results, with an R2 coefficient value of 0.96 for both pods and beans, and the RMSE values were 2.89 and 6.90, respectively. Moreover, a novel algorithm was devised to efficiently differentiate between the main stem and branches of soybean plants, called the midpoint coordinate algorithm (MCA). This was accomplished by linking the white pixels representing the stems in each column of the binary image to draw curves that represent the plant structure. The proposed method reduces computational time and spatial complexity in comparison to the A* algorithm, thereby providing an efficient and accurate approach for measuring the phenotypic characteristics of soybean plants. This research lays a technical foundation for obtaining the phenotypic data of densely overlapped and partitioned mature soybean plants under field conditions at harvest.

Summary: Stop codon readthrough (SCR) has important biological implications but remains largely uncharacterized. Here, we identify 1,009 SCR events in plants using a proteogenomic strategy. Plant SCR candidates tend to have shorter transcript lengths and fewer exons and splice variants than non-SCR transcripts. Mass spectrometry evidence shows that stop codons involved in SCR events can be recoded as 20 standard amino acids, some of which are also supported by suppressor tRNA analysis. We also observe multiple functional signals in 34 maize extended proteins and characterize the structural and subcellular localization changes in the extended protein of basic transcription factor 3. Furthermore, the SCR events exhibit non-conserved signature, and the extensions likely undergo protein-coding selection. Overall, our study not only characterizes that SCR events are commonly present in plants but also identifies the recoding plasticity of stop codons, which provides important insights into the flexibility of genetic decoding.

Abstract Background High-order chromatin structure plays important roles in gene regulation. However, the diversity of the three-dimensional (3D) genome across plant accessions are seldom reported. Results Here, we perform the pan-3D genome analysis using Hi-C sequencing data from 27 soybean accessions and comprehensively investigate the relationships between 3D genomic variations and structural variations (SVs) as well as gene expression. We find that intersection regions between A/B compartments largely contribute to compartment divergence. Topologically associating domain (TAD) boundaries in A compartments exhibit significantly higher density compared to those in B compartments. Pan-3D genome analysis shows that core TAD boundaries have the highest transcription start site (TSS) density and lowest GC content and repeat percentage. Further investigation shows that non-long terminal repeat (non-LTR) retrotransposons play important roles in maintaining TAD boundaries, while Gypsy elements and satellite repeats are associated with private TAD boundaries. Moreover, presence and absence variation (PAV) is found to be the major contributor to 3D genome variations. Nevertheless, approximately 55% of 3D genome variations are not associated with obvious genetic variations, and half of them affect the flanking gene expression. In addition, we find that the 3D genome may also undergo selection during soybean domestication. Conclusion Our study sheds light on the role of 3D genomes in plant genetic diversity and provides a valuable resource for studying gene regulation and genome evolution.

Shoot branching is critical in determining soybean yield. Here, the authors report natural variation of Dt2 in controlling soybean branching number and the interaction of Dt2 with GmAgl22 and GmSoc1a to activate transcription of GmAp1a and GmAp1d.

Tong et al. describe a super pangenome assembled from long-read sequences of 545 wild and domesticated silkworms. Naturally selected (diapause, aposemantic coloration) or artificially selected (silk yield and fineness) sets of genes are delineated.

MicroRNAs (miRNAs) are endogenous small RNAs, that are vital for gene expression regulation in eukaryotes. Whenever a pri-miRNA precursor includes another miRNA precursor, and both of these precursors may generate independent, non-overlapping mature miRNAs, we named them nested miRNAs. However, the extent of nested miR159 structural evolutionary conservation and its promoter characterization remains unknown. In this study, the sequence alignment and phylogenetic analysis reveal that the MIR159 family is ancient, and its nested miR159 structures are evolutionary conserved in different plant species. The overexpression of ath-MIR159a, including the 1.2 kb downstream region, has no effect on rescuing the mir159ab phenotype. The promoter truncation results revealed that the 1.0 kb promoter of ath-MIR159a is sufficient for rescuing the mir159ab phenotype. The cis-regulatory elements in the ath-miR159a promoters indicated functions related to different phytohormones, abiotic stresses, and transcriptional activation. While the MybSt1 motif-containing region is not responsible for activating the regulation of the miR159a promoter. The qRT-PCR results showed that overexpression of ath-MIR159a led to high expression levels of miR159a.1–5 and miR159a.1–3 and complemented the growth defect of mir159ab via downregulation of MYB33 and MYB65. Furthermore, continuously higher expression of the miR159a.2 duplex in transgenic lines with the curly leaf phenotype indicates that miR159a.2 is functional in Arabidopsis and suggests that it is possible for a miRNA precursor to encode several regulatory small RNAs in plants. Taken together, our study demonstrates that the nested miR159 structure is evolutionary conserved and miRNA-mediated gene regulation is more complex than previously thought.

MicroRNAs (miRNAs) are 20- to 24-nucleotide small RNAs, and whenever a pri-miRNA precursor includes another miRNA precursor, and both of these precursors may generate independent non overlapping mature miRNAs, we called them nested miRNAs. However, the functional and regulatory roles of nested miRNA structures in plants are still unknown. In this study, the Arabidopsis nested miR159a structure, which consists of two nested miRNAs, miR159a.1, and miR159a.2, was used as a model to determine miRNA-mediated gene silencing in plants. Complementation analysis of nested miR159a structures revealed that the miR159a structure can differentially complement the mir159ab phenotype, and a duplex nested structure in the tail end region of the pre-miR159a fold back may have a possible dominant function, indicating the importance of the flanking sequence of the stem in the cleavage of the mature miRNA. Furthermore, continuously higher expression of the miR159a.2 duplex in the severe leaf curl phenotype indicates that miR159a.2 is functional in Arabidopsis and suggests that in plants, a miRNA precursor may encode multiple regulatory small RNAs. Taken together, our study demonstrates that the nested miR159a structure regulated by duplex mutations of miR159a has a unique pattern and provides novel insight into silencing efficacy of Arabidopsis miR159a.

The rhizosphere microbiota plays a critical and crucial role in plant health and growth, assisting plants in resisting adverse stresses, including soil salinity. Plastic film mulching is an important method to adjust soil properties and improve crop yield, especially in saline–alkali soil. However, it remains unclear whether and to what extent the association between these improvements and rhizosphere microbiota exists. Here, from a field survey and a greenhouse mesocosm experiment, we found that mulching plastic films on saline–alkali soil can promote the growth of soybeans in the field. Results of the greenhouse experiment showed that soybeans grew better in unsterilized saline–alkali soil than in sterilized saline–alkali soil under plastic film mulching. By detecting the variations in soil properties and analyzing the high-throughput sequencing data, we found that with the effect of film mulching, soil moisture content was effectively maintained, soil salinity was obviously reduced, and rhizosphere bacterial and fungal communities were significantly changed. Ulteriorly, correlation analysis methods were applied. The optimization of soil properties ameliorated the survival conditions of soil microbes and promoted the increase in relative abundance of potential beneficial microorganisms, contributing to the growth of soybeans. Furthermore, the classification of potential key rhizosphere microbial OTUs were identified. In summary, our study suggests the important influence of soil properties as drivers on the alteration of rhizosphere microbial communities and indicates the important role of rhizosphere microbiota in promoting plant performance in saline–alkali soil under plastic film mulching.

Abstract Background The genetic mechanisms underlying the domestication of animals and plants have been of great interest to biologists since Darwin. To date, little is known about the global pattern of gene expression changes during domestication. Results We generated and collected transcriptome data for seven pairs of domestic animals and plants including dog, silkworm, chicken, rice, cotton, soybean and maize and their wild progenitors and compared the expression profiles between the domestic and wild species. Intriguingly, although the number of expressed genes varied little, the domestic species generally exhibited lower gene expression diversity than did the wild species, and this lower diversity was observed for both domestic plants and different kinds of domestic animals including insect, bird and mammal in the whole-genome gene set (WGGS), candidate selected gene set (CSGS) and non-CSGS, with CSGS exhibiting a higher degree of decreased expression diversity. Moreover, different from previous reports which found 2 to 4% of genes were selected by human, we identified 6892 candidate selected genes accounting for 7.57% of the whole-genome genes in rice and revealed that fewer than 8% of the whole-genome genes had been affected by domestication. Conclusions Our results showed that domestication affected the pattern of variation in gene expression throughout the genome and generally decreased the expression diversity across species, and this decrease may have been associated with decreased genetic diversity. This pattern might have profound effects on the phenotypic and physiological changes of domestic animals and plants and provide insights into the genetic mechanisms at the transcriptome level other than decreased genetic diversity and increased linkage disequilibrium underpinning artificial selection.

Hundred-seed weight (HSW) is an important measure of yield and a useful indicator to monitor the inheritance of quantitative traits affected by genotype and environmental conditions. To identify quantitative trait nucleotides (QTNs) and mine genes useful for breeding high-yielding and high-quality soybean (Glycine max) cultivars, we conducted a multilocus genome-wide association study (GWAS) on HSW of soybean based on phenotypic data from 20 different environments and genotypic data for 109,676 single-nucleotide polymorphisms (SNPs) in 144 four-way recombinant inbred lines. Using five multilocus GWAS methods, we identified 118 QTNs controlling HSW. Among these, 31 common QTNs were detected by various methods or across multiple environments. Pathway analysis identified three potential candidate genes associated with HSW in soybean. We used allele information to study the common QTNs in 20 large-seed and 20 small-seed lines and identified a higher percentage of superior alleles in the large-seed lines than in small-seed lines. These observations will contribute to construct the gene networks controlling HSW in soybean, which can improve the genetic understanding of HSW, and provide assistance for molecular breeding of soybean large-seed varieties.

Soybean varieties suitable for high planting density allow greater yields. However, the seed protein and oil contents, which determine the value of this crop, can be influenced by planting density. Thus, it is important to understand the genetic basis of the responses of different soybean genotypes to planting density. In this study, we quantified the protein and oil contents in a four-way recombinant inbred line (FW-RIL) soybean population under two planting densities and the response to density. We performed quantitative trait locus (QTL) mapping using a single nucleotide polymorphism (SNP) linkage map generated by inclusive composite interval mapping. We identified 14 QTLs for protein content and 17 for oil content at a planting density of 2.15 × 105 plant/ha (D1) and 14 QTLs for protein content and 20 for oil content at a planting density 3.0 × 105 plant/ha (D2). Among the QTLs detected, two oil-content QTLs was detected at both plant densities. In addition, we identified 38 QTLs for the responses of protein and oil contents to planting density. Of the QTLs detected, 70 were identified in previous studies, while 33 were newly identified. Fourty-five QTLs accounted for over 10% of the phenotypic variation of the corresponding trait, based on 23 QTLs at a marker interval distance of ~600 kb detected under different densities and with the responses to density difference. Pathway analysis revealed four candidate genes involved in protein and oil biosynthesis/metabolism. These results improve our understanding of the genetic underpinnings of protein and oil biosynthesis in soybean, laying the foundation for enhancing protein and oil contents and increasing yields in soybean.

Plant height is an important target for soybean breeding. It is a typical quantitative trait controlled by multiple genes and is susceptible to environmental influences. Here, we carried out phenotypic analysis of 156 recombinant inbred lines derived from “Dongnong L13” and “Henong 60” in nine environments at four locations over 6 years using interval mapping and inclusive composite interval mapping methods. We performed quantitative trait locus (QTL) analysis by applying pre-built simple-sequence repeat maps. We detected 48 QTLs, including nine significant QTLs detected by multiple methods and in multiple environments. Meanwhile, genotyping of all lines using the SoySNP660k BeadChip produced 54,836 non-redundant single-nucleotide polymorphism (SNP) genotypes. We used five multi-locus genome-wide association analysis methods to locate 10 quantitative trait nucleotides (QTNs), four of which overlap with previously located QTLs. Five candidate genes related to plant height are predicted to lie within 200 kb of these four QTNs. We identified 19 homologous genes in Arabidopsis, two of which may be associated with plant height. These findings further our understanding of the multi-gene regulatory network and genetic determinants of soybean plant height, which will be important for breeding high-yielding soybean.

Abstract Background In addition to genetic variation, epigenetic variation plays an important role in determining various biological processes. The importance of natural genetic variation to crop domestication and improvement has been widely investigated. However, the contribution of epigenetic variation in crop domestication at population level has rarely been explored. Results To understand the impact of epigenetics on crop domestication, we investigate the variation of DNA methylation during soybean domestication and improvement by whole-genome bisulfite sequencing of 45 soybean accessions, including wild soybeans, landraces, and cultivars. Through methylomic analysis, we identify 5412 differentially methylated regions (DMRs). These DMRs exhibit characters distinct from those of genetically selected regions. In particular, they have significantly higher genetic diversity. Association analyses suggest only 22.54% of DMRs can be explained by local genetic variations. Intriguingly, genes in the DMRs that are not associated with any genetic variation are enriched in carbohydrate metabolism pathways. Conclusions This study provides a valuable map of DNA methylation across diverse accessions and dissects the relationship between DNA methylation variation and genetic variation during soybean domestication, thus expanding our understanding of soybean domestication and improvement.

Abstract Background Organisms need to adapt to keep pace with a changing environment. Examining recent range expansion aids our understanding of how organisms evolve to overcome environmental constraints. However, how organisms adapt to climate changes is a crucial biological question that is still largely unanswered. The plant Arabidopsis thaliana is an excellent system to study this fundamental question. Its origin is in the Iberian Peninsula and North Africa, but it has spread to the Far East, including the most south-eastern edge of its native habitats, the Yangtze River basin, where the climate is very different. Results We sequenced 118 A. thaliana strains from the region surrounding the Yangtze River basin. We found that the Yangtze River basin population is a unique population and diverged about 61,409 years ago, with gene flows occurring at two different time points, followed by a population dispersion into the Yangtze River basin in the last few thousands of years. Positive selection analyses revealed that biological regulation processes, such as flowering time, immune and defense response processes could be correlated with the adaptation event. In particular, we found that the flowering time gene SVP has contributed to A. thaliana adaptation to the Yangtze River basin based on genetic mapping. Conclusions A. thaliana adapted to the Yangtze River basin habitat by promoting the onset of flowering, a finding that sheds light on how a species can adapt to locales with very different climates.

Abstract Background Soybean (Glycine max [L.] Merr.) is one of the most important oil and protein crops. Ever-increasing soybean consumption necessitates the improvement of varieties for more efficient production. However, both correlations among different traits and genetic interactions among genes that affect a single trait pose a challenge to soybean breeding. Results To understand the genetic networks underlying phenotypic correlations, we collected 809 soybean accessions worldwide and phenotyped them for two years at three locations for 84 agronomic traits. Genome-wide association studies identified 245 significant genetic loci, among which 95 genetically interacted with other loci. We determined that 14 oil synthesis-related genes are responsible for fatty acid accumulation in soybean and function in line with an additive model. Network analyses demonstrated that 51 traits could be linked through the linkage disequilibrium of 115 associated loci and these links reflect phenotypic correlations. We revealed that 23 loci, including the known Dt1, E2, E1, Ln, Dt2, Fan, and Fap loci, as well as 16 undefined associated loci, have pleiotropic effects on different traits. Conclusions This study provides insights into the genetic correlation among complex traits and will facilitate future soybean functional studies and breeding through molecular design.

Peach is both an economically important crop species and a model for Rosaceae fruit development research. Here, the authors perform genome-wide association analysis in peach and find candidate genes associated with variation in agronomically important fruit phenotypes.

RNA editing plays a key role in organelle gene expression. Little is known about how RNA editing factors influence soybean plant development. Here, we report the isolation and characterization of a soybean yl (yellow leaf) mutant. The yl plants showed decreased chlorophyll accumulation, lower PS II activity, an impaired net photosynthesis rate, and an altered chloroplast ultrastructure. Fine mapping of YL uncovered a point mutation in Glyma.20G187000, which encodes a chloroplast-localized protein homologous to Arabidopsis thaliana (Arabidopsis) ORRM1. YL is mainly expressed in trifoliate leaves, and its deficiency affects the editing of multiple chloroplast RNA sites, leading to inferior photosynthesis in soybean. Taken together, these results demonstrate the importance of the soybean YL protein in chloroplast RNA editing and photosynthesis.

Protein content (PC), an important trait in soybean (Glycine max) breeding, is controlled by multiple genes with relatively small effects. To identify the quantitative trait nucleotides (QTNs) controlling PC, we conducted a multi-locus genome-wide association study (GWAS) for PC in 144 four-way recombinant inbred lines (FW-RILs). All the FW-RILs were phenotyped for PC in 20 environments, including four locations over 4 years with different experimental treatments. Meanwhile, all the FW-RILs were genotyped using SoySNP660k BeadChip, producing genotype data for 109,676 non-redundant single-nucleotide polymorphisms. A total of 129 significant QTNs were identified by five multi-locus GWAS methods. Based on the 22 common QTNs detected by multiple GWAS methods or in multiple environments, pathway analysis identified 8 potential candidate genes that are likely to be involved in protein synthesis and metabolism in soybean seeds. Using superior allele information for 22 common QTNs in 22 elite and 7 inferior lines, we found higher superior allele percentages in the elite lines and lower percentages in the inferior lines. These findings will contribute to the discovery of the polygenic networks controlling PC in soybean, increase our understanding of the genetic foundation and regulation of PC, and be useful for molecular breeding of high-protein soybean varieties.

Isoflavones comprise a group of secondary metabolites produced almost exclusively by plants in the legume family, including soybean [Glycine max (L.) Merr.]. They play vital roles in plant defense and have many beneficial effects on human health. Isoflavone content is a complex quantitative trait controlled by multiple genes, and the genetic mechanisms underlying isoflavone biosynthesis remain largely unknown. Via a genome-wide association study (GWAS), we identified 28 single nucleotide polymorphisms (SNPs) that are significantly associated with isoflavone concentrations in soybean. One of these 28 SNPs was located in the 5'-untranslated region (5'-UTR) of an R2R3-type MYB transcription factor, GmMYB29, and this gene was thus selected as a candidate gene for further analyses. A subcellular localization study confirmed that GmMYB29 was located in the nucleus. Transient reporter gene assays demonstrated that GmMYB29 activated the IFS2 (isoflavone synthase 2) and CHS8 (chalcone synthase 8) gene promoters. Overexpression and RNAi-mediated silencing of GmMYB29 in soybean hairy roots resulted in increased and decreased isoflavone content, respectively. Moreover, a candidate-gene association analysis revealed that 11 natural GmMYB29 polymorphisms were significantly associated with isoflavone contents, and regulation of GmMYB29 expression could partially contribute to the observed phenotypic variation. Taken together, these results provide important genetic insights into the molecular mechanisms underlying isoflavone biosynthesis in soybean.

Centromere repositioning refers to a de novo centromere formation at another chromosomal position without sequence rearrangement. This phenomenon was frequently encountered in both mammalian and plant species and has been implicated in genome evolution and speciation. To understand the dynamic of centromeres on soybean genome, we performed the pan-centromere analysis using CENH3-ChIP-seq data from 27 soybean accessions, including 3 wild soybeans, 9 landraces, and 15 cultivars. Building upon the previous discovery of three centromere satellites in soybean, we have identified two additional centromere satellites that specifically associate with chromosome 1. These satellites reveal significant rearrangements in the centromere structures of chromosome 1 across different accessions, consequently impacting the localization of CENH3. By comparative analysis, we reported a high frequency of centromere repositioning on 14 out of 20 chromosomes. Most newly emerging centromeres formed in close proximity to the native centromeres and some newly emerging centromeres were apparently shared in distantly related accessions, suggesting their emergence is independent. Furthermore, we crossed two accessions with mismatched centromeres to investigate how centromere positions would be influenced in hybrid genetic backgrounds. We found that a significant proportion of centromeres in the S9 generation undergo changes in size and position compared to their parental counterparts. Centromeres preferred to locate at satellites to maintain a stable state, highlighting a significant role of centromere satellites in centromere organization. Taken together, these results revealed extensive centromere repositioning in soybean genome and highlighted how important centromere satellites are in constraining centromere positions and supporting centromere function. [ABSTRACT FROM AUTHOR]