Sinorhizobium Fredii HH103 Research Articles

Exopolysaccharide is detrimental for the symbiotic performance of Sinorhizobium fredii HH103 mutants with a truncated lipopolysaccharide core.

The nitrogen-fixing rhizobia-legume symbiosis relies on a complex interchange of molecular signals between the two partners during the whole interaction. On the bacterial side, different surface polysaccharides, such as lipopolysaccharide (LPS) and exopolysaccharide (EPS), might play important roles for the success of the interaction. In a previous work we studied two Sinorhizobium fredii HH103 mutants affected in the rkpK and lpsL genes, which are responsible for the production of glucuronic acid and galacturonic acid, respectively. Both mutants produced an altered LPS, and the rkpK mutant, in addition, lacked EPS. These mutants were differently affected in symbiosis with Glycine max and Vigna unguiculata, with the lpsL mutant showing a stronger impairment than the rkpK mutant. In the present work we have further investigated the LPS structure and the symbiotic abilities of the HH103 lpsL and rkpK mutants. We demonstrate that both strains produce the same LPS, with a truncated core oligosaccharide devoid of uronic acids. We show that the symbiotic performance of the lpsL mutant with Macroptilium atropurpureum and Glycyrrhiza uralensis is worse than that of the rkpK mutant. Introduction of an exoA mutation (which avoids EPS production) in HH103 lpsL improved its symbiotic performance with G. max, M. atropurpureum, and G. uralensis to the level exhibited by HH103 rkpK, suggesting that the presence of EPS might hide the truncated LPS produced by the former mutant.

Insights into the Early Steps of the Symbiotic Interaction between Soybean (Glycine max) and Sinorhizobium fredii Symbiosis Using Transcriptome, Small RNA, and Degradome Sequencing.

Symbiotic nitrogen fixation carried out by the soybean-rhizobia symbiosis increases soybean yield and reduces the amount of nitrogen fertilizer that has been applied. MicroRNAs (miRNAs) are crucial in plant growth and development, prompting an investigation into their role in the symbiotic interaction of soybean with partner rhizobia. Through integrated small RNA, transcriptome, and degradome sequencing analysis, 1215 known miRNAs, 314 of them conserved, and 187 novel miRNAs were identified, with 44 differentially expressed miRNAs in soybean roots inoculated with Sinorhizobium fredii HH103 and a ttsI mutant. The study unveiled that the known miRNA gma-MIR398a-p5 was downregulated in the presence of the ttsI mutation, while the target gene of gma-MIR398a-p5, Glyma.06G007500, associated with nitrogen metabolism, was upregulated. The results of this study offer insights for breeding high-efficiency nitrogen-fixing soybean varieties, enhancing crop yield and quality.

The type III effector NopL interacts with GmREM1a and GmNFR5 to promote symbiosis in soybean

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.

Sinorhizobium fredii HH103 flgJ is a flagellar gene induced by genistein in a NodD1- and TtsI- dependent manner

Sinorhizobium fredii HH103 flgJ is a flagellar gene induced by genistein in a NodD1- and TtsI- dependent manner

A complex regulatory network governs the expression of symbiotic genes in Sinorhizobium fredii HH103.

The establishment of the rhizobium-legume nitrogen-fixing symbiosis relies on the interchange of molecular signals between the two symbionts. We have previously studied by RNA-seq the effect of the symbiotic regulators NodD1, SyrM, and TtsI on the expression of the symbiotic genes (the nod regulon) of Sinorhizobium fredii HH103 upon treatment with the isoflavone genistein. In this work we have further investigated this regulatory network by incorporating new RNA-seq data of HH103 mutants in two other regulatory genes, nodD2 and nolR. Both genes code for global regulators with a predominant repressor effect on the nod regulon, although NodD2 acts as an activator of a small number of HH103 symbiotic genes. By combining RNA-seq data, qPCR experiments, and b-galactosidase assays of HH103 mutants harbouring a lacZ gene inserted into a regulatory gene, we have analysed the regulatory relations between the nodD1, nodD2, nolR, syrM, and ttsI genes, confirming previous data and discovering previously unknown relations. Previously we showed that HH103 mutants in the nodD2, nolR, syrM, or ttsI genes gain effective nodulation with Lotus japonicus, a model legume, although with different symbiotic performances. Here we show that the combinations of mutations in these genes led, in most cases, to a decrease in symbiotic effectiveness, although all of them retained the ability to induce the formation of nitrogen-fixing nodules. In fact, the nodD2, nolR, and syrM single and double mutants share a set of Nod factors, either overproduced by them or not generated by the wild-type strain, that might be responsible for gaining effective nodulation with L. japonicus.

NopC/T/L Signal Crosstalk Gene GmPHT1-4.

Symbiotic nodulation between leguminous plants and rhizobia is a critical biological interaction. The type III secretion system (T3SS) employed by rhizobia manipulates the host's nodulation signaling, analogous to mechanisms used by certain bacterial pathogens for effector protein delivery into host cells. This investigation explores the interactive signaling among type III effectors HH103ΩNopC, HH103ΩNopT, and HH103ΩNopL from SinoRhizobium fredii HH103. Experimental results revealed that these effectors positively regulate nodule formation. Transcriptomic analysis pinpointed GmPHT1-4 as the key gene facilitating this effector-mediated signaling. Overexpression of GmPHT1-4 enhances nodulation, indicating a dual function in nodulation and phosphorus homeostasis. This research elucidates the intricate regulatory network governing Rhizobium-soybean (Glycine max (L.) Merr) interactions and the complex interplay between type III effectors.

QTL Mapping and Functional Identification of Candidate Genes Regulated by Sinorhizobium fredii HH103 and Associated with Nodulation Traits in Soybean

Large amounts of nitrogen fertilizer are annually applied to improve soybean yield. However, the overuse of nitrogen fertilizers has resulted in significant environmental pollution. Biological nitrogen fixation is an ecological and environmentally friendly method to increase soybean yield. However, the signaling pathway and function of genes in the plant host regulated by rhizobia under the symbiotic relationship remain unknown. In this study, the chromosome region in soybean responsive to Sinorhizobium fredii HH103 was identified using chromosome-segment-substituted lines produced from a cross between cultivated soybean SN14 and wild soybean Zyd00006. In addition, candidate genes associated with nodulation traits and regulated by S. fredii HH103 were identified. In total, three quantitative trait loci (QTLs) containing 68 genes were identified on chromosomes 02, 05, and 20. The differentially expressed genes among the QTL regions were determined using RNA-seq and qRT-PCR. Glyma. 05G240500, a potential gene responding to S. fredii HH103 and positively regulating soybean nodulation, was identified. To explore the relationships between haplotypes and soybean nodulation phenotypes, SNPs found in the regulatory areas of Glyma.05G240500 haplotype were investigated. Our study revealed the role of Glyma.05G240500 in symbiosis and provided a reference for facilitating symbiotic nitrogen fixation in the field and for marker-assisted selection.

GmTNRP1, associated with rhizobial type‐III effector NopT, regulates nitrogenase activity in the nodules of soybean (Glycine max)

AbstractSoybean is a rich source of protein and oil for humans. It forms a symbiotic association with rhizobia to fix atmospheric nitrogen for its growth and development. Rhizobial type‐III effectors act as important signaling molecules in symbiosis. However, they are poorly studied. In this study, mutation of TtsI in Sinorhizobium fredii HH103 reduced nodule numbers and dry weights in SN14 (cultivated soybean) and ZYD00006 (wild soybean). Moreover, it reduced nitrogenase activity in SN14 but not in ZYD00006. Based on the differences in nitrogenase activity between SN14 and ZYD00006 inoculated with HH103 and the TtsI mutant, two conditional quantitative trait loci associated with nitrogenase activity were identified in chromosome segment substitution lines derived from SN14 and ZYD00006. Using transcriptome analysis, single‐nucleotide polymorphism (SNP) analysis, and qRT‐PCR, Glyma.13G150000 (GmTNRP1) was identified as a candidate gene in soybean that was associated with nitrogenase activity, along with NopT (a type‐III effector). In addition, GmTNRP1 (an LRR‐RK family protein located on the cell membrane) was observed to be induced by NopT to negatively regulate nitrogenase activity. In transgenic hairy roots with GmTNRP1 silencing, no significant difference was observed in terms of nitrogenase activity between nodules inoculated with the NopT mutant and those inoculated with HH103 (wild type). However, the nitrogenase activity in the nodules decreased after overexpression of GmTNRP1. SNP located in the GmTNRP1 promoter affected its expression in response to NopT. Haplotype analysis revealed that SN14 belonged to Hap1 and ZYD00006 belonged to Hap2, which explained the different results in the two varieties. To the best of our knowledge, this is the first study to identify a soybean gene associated with a rhizobial type‐III effector in the nodules. GmTNRP1 can negatively regulate nitrogenase activity in the nodules. This study enhanced the understanding of the mechanism underlying symbiotic association between soybean and rhizobia.

Non-Ionic Osmotic Stress Induces the Biosynthesis of Nodulation Factors and Affects Other Symbiotic Traits in Sinorhizobium fredii HH103.

(1) Background: Some rhizobia, such as Rhizobium tropici CIAT 899, activate nodulation genes when grown under osmotic stress. This work aims to determine whether this phenomenon also takes place in Sinorhizobium fredii HH103. (2) Methods: HH103 was grown with and without 400 mM mannitol. β-galactosidase assays, nodulation factor extraction, purification and identification by mass spectrometry, transcriptomics by RNA sequencing, motility assays, analysis of acyl-homoserine lactones, and indole acetic acid quantification were performed. (3) Results: Non-ionic osmotic stress induced the production of nodulation factors. Forty-two different factors were detected, compared to 14 found in the absence of mannitol. Transcriptomics indicated that hundreds of genes were either activated or repressed upon non-ionic osmotic stress. The presence of 400 mM mannitol induced the production of indole acetic acid and acyl homoserine lactones, abolished swimming, and promoted surface motility. (4) Conclusions: In this work, we show that non-ionic stress in S. fredii HH103, caused by growth in the presence of 400 mM mannitol, provokes notable changes not only in gene expression but also in various bacterial traits, including the production of nodulation factors and other symbiotic signals.

Genetic variation in GmCRP contributes to nodulation in soybean (Glycine max Merr.)

Symbiosis between soybean and rhizobia contributes to soybean yield and quality. Although secreted rhizobial type III effectors are known to regulate infection and promote nitrogen fixation, much about them remains unknown. Mutation of NopC, a type III effector from Sinorhizobium fredii HH103, reduced nodule numbers and dry weights in 310 soybean accessions, and expression of NopC in soybean hairy roots promoted symbiosis. Based on observed differences in nodule traits between Suinong 14 and Zyd 00,006 inoculated with HH103 and the NopC mutant, 11 QTL associated with rhizobia were identified in chromosome segment substitution lines (CSSLs) derived from Suinong 14 and Zyd 00006. Using chromosome fragment insertion, whole-genome sequencing of Suinong 14 and Zyd 00006, and qRT-PCR, Glyma.19G176300 (GmCRP) was identified as a candidate gene associated with NopC, and GmCRP was found to be induced by NopC to positively regulate nodulation. SNPs located in the regulatory regions of GmCRP influenced its expression response to NopC, with SNPs contributing to nodulation having been selected during domestication. Our findings reveal the function of a soybean gene encoding a rhizobial type III effector that contributes to symbiosis, and will facilitate the practical application of symbiotic nitrogen fixation in molecular breeding.

The Rhizobium tropici CIAT 899 NodD2 protein promotes symbiosis and extends rhizobial nodulation range by constitutive nodulation factor synthesis.

In the symbiotic associations between rhizobia and legumes, the NodD regulators orchestrate the transcription of the specific nodulation genes. This set of genes is involved in the synthesis of nodulation factors, which are responsible for initiating the nodulation process. Rhizobium tropici CIAT 899 is the most successful symbiont of Phaseolus vulgaris and can nodulate a variety of legumes. Among the five NodD regulators present in this rhizobium, only NodD1 and NodD2 seem to have a role in the symbiotic process. However, the individual role of each NodD in the absence of the other proteins has remained elusive. In this work, we show that the CIAT 899 NodD2 does not require activation by inducers to promote the synthesis of nodulation factors. A CIAT 899 strain overexpressing nodD2, but lacking all additional nodD genes, can nodulate three different legumes as efficiently as the wild type. Interestingly, CIAT 899 NodD2-mediated gain of nodulation can be extended to another rhizobial species, since its overproduction in Sinorhizobium fredii HH103 not only increases the number of nitrogen-fixing nodules in two host legumes but also results in nodule development in incompatible legumes. These findings potentially open exciting opportunities to develop rhizobial inoculants and increase legume crop production.

Surface Motility Regulation of Sinorhizobium fredii HH103 by Plant Flavonoids and the NodD1, TtsI, NolR, and MucR1 Symbiotic Bacterial Regulators.

Bacteria can spread on surfaces to colonize new environments and access more resources. Rhizobia, a group of α- and β-Proteobacteria, establish nitrogen-fixing symbioses with legumes that rely on a complex signal interchange between the partners. Flavonoids exuded by plant roots and the bacterial transcriptional activator NodD control the transcription of different rhizobial genes (the so-called nod regulon) and, together with additional bacterial regulatory proteins (such as TtsI, MucR or NolR), influence the production of different rhizobial molecular signals. In Sinorhizobium fredii HH103, flavonoids and NodD have a negative effect on exopolysaccharide production and biofilm production. Since biofilm formation and motility are often inversely regulated, we have analysed whether flavonoids may influence the translocation of S. fredii HH103 on surfaces. We show that the presence of nod gene-inducing flavonoids does not affect swimming but promotes a mode of surface translocation, which involves both flagella-dependent and -independent mechanisms. This surface motility is regulated in a flavonoid-NodD1-TtsI-dependent manner, relies on the assembly of the symbiotic type 3 secretion system (T3SS), and involves the participation of additional modulators of the nod regulon (NolR and MucR1). To our knowledge, this is the first evidence indicating the participation of T3SS in surface motility in a plant-interacting bacterium. Interestingly, flavonoids acting as nod-gene inducers also participate in the inverse regulation of surface motility and biofilm formation, which could contribute to a more efficient plant colonisation.

GmARP is Related to the Type III Effector NopAA to Promote Nodulation in Soybean (Glycine max).

Type III effectors secreted by rhizobia regulate nodulation in the host plant and are important modulators of symbiosis between rhizobia and soybean (Glycine max), although the underlying mechanisms are poorly understood. Here, we studied the type III effector NopAA in Sinorhizobium fredii HH103, confirming its secretion into the extracellular environment under the action of genistein. The enzyme activity of NopAA was investigated in vitro, using xyloglucan and β-glucan as substrates. NopAA functions were investigated by the generation of a NopAA mutant and the effects of NopAA deficiency on symbiosis were analyzed. Soybean genes associated with NopAA were identified in a recombinant inbred line (RIL) population and their functions were verified. NopAA was confirmed to be a type III effector with glycosyl hydrolase activity, and its mutant did not promote nodulation. Quantitative trait locus (QTL) analysis identified 10 QTLs with one, Glyma.19g074200 (GmARP), found to be associated with NopAA and to positively regulate the establishment of symbiosis. All these results support the hypothesis that type III effectors interact with host proteins to regulate the establishment of symbiosis and suggest the possibility of manipulating the symbiotic soybean–rhizobia interaction to promote efficient nitrogen fixation.

Mapping of Quantitative Trait Loci Underlying Nodule Traits in Soybean (Glycine max (L.) Merr.) and Identification of Genes Whose Expression Is Affected by the Sinorhizobium fredii HH103 Effector Proteins NopL and NopT

Symbiotic nitrogen fixation provides most of the nitrogen required for soybean growth. Rhizobial nodulation outer proteins (Nops) have been reported to influence host specificity during symbiosis establishment. However, the host proteins that interact with Nops remain unknown. In this study, we generated Sinorhizobium fredii HH103 mutants (HH103ΩNopL, HH103ΩNopT, and HH103ΩNopLΩNopT) and analysed the nodule number (NN) and nodule dry weight (NDW) of 12 soybean germplasms after inoculation with wild-type S. fredii HH103 or the mutant strains. The analysis of chromosome segment substitution lines revealed quantitative trait loci (QTLs) associated with NopL and NopT interactions. A total of 22 QTLs for the 2 nodule traits were detected and mapped to 12 different chromosomes in the soybean genome. Eight and fifteen QTLs were found to be associated with NN and NDW, respectively. Furthermore, 17 candidate genes were selected for further analyses. Considering the results of reverse-transcription quantitative PCR, we propose that the protein products of these 17 candidate genes interact with NopL and NopT.

Analysis of Outer Membrane Vesicles Indicates That Glycerophospholipid Metabolism Contributes to Early Symbiosis Between Sinorhizobium fredii HH103 and Soybean.

Gram-negative bacteria can produce outer membrane vesicles (OMVs), and most functional studies of OMVs have been focused on mammalian-bacterial interactions. However, research on the OMVs of rhizobia is still limited. In this work, we isolated and purified OMVs from Sinorhizobium fredii HH103 under free-living conditions that were set as control (C-OMVs) and symbiosis-mimicking conditions that were induced by genistein (G-OMVs). The soybean roots treated with G-OMVs displayed significant deformation of root hairs. G-OMVs significantly induced the expression of nodulation genes related to early symbiosis, while they inhibited that of the defense genes of soybean. Proteomics analysis identified a total of 93 differential proteins between C-OMVs and G-OMVs, which are mainly associated with ribosome synthesis, flagellar assembly, two-component system, ABC transporters, oxidative phosphorylation, nitrogen metabolism, quorum sensing, glycerophospholipid metabolism, and peptidoglycan biosynthesis. A total of 45 differential lipids were identified through lipidomics analysis. Correlation analysis of OMV proteome and lipidome data revealed that glycerophospholipid metabolism is the enriched Kyoto Encyclopedia of Genes and Genomes metabolic pathway, and the expression of phosphatidylserine decarboxylase was significantly up-regulated in G-OMVs. The changes in three lipids related to symbiosis in the glycerophospholipid metabolism pathway were verified by enzyme-linked immunosorbent assay. Our results indicate that glycerophospholipid metabolism contributes to rhizobia-soybean symbiosis via OMVs.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The nodD1 Gene of Sinorhizobium fredii HH103 Restores Nodulation Capacity on Bean in a Rhizobium tropici CIAT 899 nodD1/nodD2 Mutant, but the Secondary Symbiotic Regulators nolR, nodD2 or syrM Prevent HH103 to Nodulate with This Legume.

Rhizobial NodD proteins and appropriate flavonoids induce rhizobial nodulation gene expression. In this study, we show that the nodD1 gene of Sinorhizobium fredii HH103, but not the nodD2 gene, can restore the nodulation capacity of a double nodD1/nodD2 mutant of Rhizobium tropici CIAT 899 in bean plants (Phaseolus vulgaris). S. fredii HH103 only induces pseudonodules in beans. We have also studied whether the mutation of different symbiotic regulatory genes may affect the symbiotic interaction of HH103 with beans: ttsI (the positive regulator of the symbiotic type 3 protein secretion system), and nodD2, nolR and syrM (all of them controlling the level of Nod factor production). Inactivation of either nodD2, nolR or syrM, but not that of ttsI, affected positively the symbiotic behavior of HH103 with beans, leading to the formation of colonized nodules. Acetylene reduction assays showed certain levels of nitrogenase activity that were higher in the case of the nodD2 and nolR mutants. Similar results have been previously obtained by our group with the model legume Lotus japonicus. Hence, the results obtained in the present work confirm that repression of Nod factor production, provided by either NodD2, NolR or SyrM, prevents HH103 to effectively nodulate several putative host plants.

Structure of the unusual Sinorhizobium fredii HH103 lipopolysaccharide and its role in symbiosis

Rhizobia are soil bacteria that form important symbiotic associations with legumes, and rhizobial surface polysaccharides, such as K-antigen polysaccharide (KPS) and lipopolysaccharide (LPS), might be important for symbiosis. Previously, we obtained a mutant of Sinorhizobium fredii HH103, rkpA, that does not produce KPS, a homopolysaccharide of a pseudaminic acid derivative, but whose LPS electrophoretic profile was indistinguishable from that of the WT strain. We also previously demonstrated that the HH103 rkpLMNOPQ operon is responsible for 5-acetamido-3,5,7,9-tetradeoxy-7-(3-hydroxybutyramido)-l-glycero-l-manno-nonulosonic acid [Pse5NAc7(3OHBu)] production and is involved in HH103 KPS and LPS biosynthesis and that an HH103 rkpM mutant cannot produce KPS and displays an altered LPS structure. Here, we analyzed the LPS structure of HH103 rkpA, focusing on the carbohydrate portion, and found that it contains a highly heterogeneous lipid A and a peculiar core oligosaccharide composed of an unusually high number of hexuronic acids containing β-configured Pse5NAc7(3OHBu). This pseudaminic acid derivative, in its α-configuration, was the only structural component of the S. fredii HH103 KPS and, to the best of our knowledge, has never been reported from any other rhizobial LPS. We also show that Pse5NAc7(3OHBu) is the complete or partial epitope for a mAb, NB6-228.22, that can recognize the HH103 LPS, but not those of most of the S. fredii strains tested here. We also show that the LPS from HH103 rkpM is identical to that of HH103 rkpA but devoid of any Pse5NAc7(3OHBu) residues. Notably, this rkpM mutant was severely impaired in symbiosis with its host, Macroptilium atropurpureum.

The Sinorhizobium fredii HH103 type III secretion system effector NopC blocks nodulation with Lotus japonicus Gifu.

The broad-host-range bacterium Sinorhizobium fredii HH103 cannot nodulate the model legume Lotus japonicus Gifu. This bacterium possesses a type III secretion system (T3SS), a specialized secretion apparatus used to deliver effector proteins (T3Es) into the host cell cytosol to alter host signaling and/or suppress host defence responses to promote infection. However, some of these T3Es are recognized by specific plant receptors and hence trigger a strong defence response to block infection. In rhizobia, T3Es are involved in nodulation efficiency and host-range determination, and in some cases directly activate host symbiosis signalling in a Nod factor-independent manner. In this work, we show that HH103 RifR T3SS mutants, unable to secrete T3Es, gain nodulation with L. japonicus Gifu through infection threads, suggesting that plant recognition of a T3E could block the infection process. To identify the T3E involved, we performed nodulation assays with a collection of mutants that affect secretion of each T3E identified in HH103 RifR so far. The nopC mutant could infect L. japonicus Gifu by infection thread invasion and switch the infection mechanism in Lotus burttii from intercellular infection to infection thread formation. Lotus japonicus gene expression analysis indicated that the infection-blocking event occurs at early stages of the symbiosis.

QTL Mapping and Data Mining to Identify Genes Associated With the Sinorhizobium fredii HH103 T3SS Effector NopD in Soybean.

In some legume–rhizobium symbioses, host specificity is influenced by rhizobial type III effectors-nodulation outer proteins (Nops). However, the genes encoding host proteins that interact with Nops remain unknown. In this study, we aimed to identify candidate soybean genes associated with NopD, one of the type III effectors of Sinorhizobium fredii HH103. The results showed that the expression pattern of NopD was analyzed in rhizobia induced by genistein. We also found NopD can be induced by TtsI, and NopD as a toxic effector can induce tobacco leaf death. In 10 soybean germplasms, NopD played a positively effect on nodule number (NN) and nodule dry weight (NDW) in nine germplasms, but not in Kenjian28. Significant phenotype of NN and NDW were identified between Dongnong594 and Charleston, Suinong14 and ZYD00006, respectively. To map the quantitative trait locus (QTL) associated with NopD, a recombinant inbred line (RIL) population derived from the cross between Dongnong594 and Charleston, and chromosome segment substitution lines (CSSLs) derived from Suinong14 and ZYD00006 were used. Two overlapping conditional QTL associated with NopD on chromosome 19 were identified. Two candidate genes were identified in the confident region of QTL, we found that NopD could influence the expression of Glyma.19g068600 (FBD/LRR) and expression of Glyma.19g069200 (PP2C) after HH103 infection. Haplotype analysis showed that different types of Glyma.19g069200 haplotypes could cause significant nodule phenotypic differences, but Glyma.19g068600 (FBD/LRR) was not. These results suggest that NopD promotes S. fredii HH103 infection via directly or indirectly regulating Glyma.19g068600 and Glyma.19g069200 expression during the establishment of symbiosis between rhizobia and soybean plants.

Sinorhizobium fredii HH103 syrM inactivation affects the expression of a large number of genes, impairs nodulation with soybean and extends the host-range to Lotus japonicus.

Sinorhizobium fredii HH103 RifR is a broad host-range rhizobial strain able to nodulate with soybean and Lotus burttii, but it is ineffective with L. japonicus. Here, we study the role of the HH103 RifR SyrM protein in the regulation of gene expression and its relevance in symbiosis with those three legumes. RNAseq analyses show that HH103 SyrM is an important transcriptional regulator not only in the presence of inducer flavonoids but also in its absence. Lack of SyrM increases Nod factors production and decreases genistein-mediated repression of exopolysaccharide production in HH103. In symbiosis, mutation of syrM partially impaired interaction with soybean but improves effectiveness with L. burttii and extends the host-rage to L. japonicus Gifu. In addition, HH103 syrM mutants enter in both Lotus species by infection threads, whereas HH103 uses the more primitive intercellular infection to enter into L. burttii roots These symbiotic phenotypes were previously observed in two other HH103 mutants affected in symbiotic regulators, nodD2 and nolR, revealing that in S. fredii HH103 numerous transcriptional regulators finely modulate symbiotic gene expression.