Symbiotic nitrogen (N) fixation entails successful interaction between legume hosts and rhizobia that occur in specialized organs called nodules. N-fixing legumes have a higher demand for phosphorus (P) than legumes grown on mineral N. Medicago truncatula is an important model plant for characterization of effects of P deficiency at the molecular level. Hence, a study was carried out to address the alteration in metabolite levels of M. truncatula grown aeroponically and subjected to 4 weeks of P stress. First, GC-MS-based untargeted metabolomics initially revealed changes in the metabolic profile of nodules, with increased levels of amino acids and sugars and a decline in amounts of organic acids. Subsequently, LC-MS/MS was used to quantify these compounds including phosphorylated metabolites in the whole plant. Our results showed a drastic reduction in levels of organic acids and phosphorylated compounds in –P leaves, with a moderate reduction in –P roots and nodules. Additionally, sugars and amino acids were elevated in the whole plant under P deprivation. These findings provide evidence that N fixation in M. truncatula is mediated through a N feedback mechanism that in parallel is related to carbon and P metabolism.
Phosphorus deprivation affects composition and spatial distribution of membrane lipids in legume nodules.
In legumes, symbiotic nitrogen (N) fixation (SNF) occurs in specialized organs called nodules after successful interactions
between legume hosts and rhizobia. In a nodule, N-fixing rhizobia are surrounded by symbiosome membranes, through
which the exchange of nutrients and ammonium occurs between bacteria and the host legume. Phosphorus (P) is an
essential macronutrient, and N2-fixing legumes have a higher requirement for P than legumes grown on mineral N. As in
the previous studies, in P deficiency, barrel medic (Medicago truncatula) plants had impaired SNF activity, reduced growth,
and accumulated less phosphate in leaves, roots, and nodules compared with the plants grown in P sufficient conditions.
Membrane lipids in M. truncatula tissues were assessed using electrospray ionization–mass spectrometry. Galactolipids
were found to increase in P deficiency, with declines in phospholipids (PL), especially in leaves. Lower PL losses were found
in roots and nodules. Subsequently, matrix-assisted laser desorption/ionization–mass spectrometry imaging was used to
spatially map the distribution of the positively charged phosphatidylcholine (PC) species in nodules in both P-replete and
P-deficient conditions. Our results reveal heterogeneous distribution of several PC species in nodules, with homogeneous
distribution of other PC classes. In P poor conditions, some PC species distributions were observed to change. The results
suggest that specific PC species may be differentially important in diverse nodule zones and cell types, and that membrane
lipid remodeling during P stress is not uniform across the nodule.
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- Award ID(s):
- 2139351
- NSF-PAR ID:
- 10292568
- Date Published:
- Journal Name:
- Plant physiology
- ISSN:
- 1532-2548
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract -
Stabb, Eric V. (Ed.)ABSTRACT Some soil bacteria, called rhizobia, can interact symbiotically with legumes, in which they form nodules on the plant roots, where they can reduce atmospheric dinitrogen to ammonia, a form of nitrogen that can be used by growing plants. Rhizobium-plant combinations can differ in how successful this symbiosis is: for example, Sinorhizobium meliloti Rm1021 forms a relatively ineffective symbiosis with Medicago truncatula Jemalong A17, but Sinorhizobium medicae WSM419 is able to support more vigorous plant growth. Using proteomic data from free-living and symbiotic S. medicae WSM419, we previously identified a subset of proteins that were not closely related to any S. meliloti Rm1021 proteins and speculated that adding one or more of these proteins to S. meliloti Rm1021 would increase its effectiveness on M. truncatula A17. Three genes, Smed_3503, Smed_5985, and Smed_6456, were cloned into S. meliloti Rm1021 downstream of the E. coli lacZ promoter. Strains with these genes increased nodulation and improved plant growth, individually and in combination with one another. Smed_3503, renamed iseA ( i ncreased s ymbiotic e ffectiveness), had the largest impact, increasing M. truncatula biomass by 61%. iseA homologs were present in all currently sequenced S. medicae strains but were infrequent in other Sinorhizobium isolates. Rhizobium leguminosarum bv. viciae 3841 containing iseA led to more nodules on pea and lentil. Split-root experiments with M. truncatula A17 indicated that S. meliloti Rm1021 carrying the S. medicae iseA is less sensitive to plant-induced resistance to rhizobial infection, suggesting an interaction with the plant’s regulation of nodule formation. IMPORTANCE Legume symbiosis with rhizobia is highly specific. Rhizobia that can nodulate and fix nitrogen on one legume species are often unable to associate with a different species. The interaction can be more subtle. Symbiotically enhanced growth of the host plant can differ substantially when nodules are formed by different rhizobial isolates of a species, much like disease severity can differ when conspecific isolates of pathogenic bacteria infect different cultivars. Much is known about bacterial genes essential for a productive symbiosis, but less is understood about genes that marginally improve performance. We used a proteomic strategy to identify Sinorhizobium genes that contribute to plant growth differences that are seen when two different strains nodulate M. truncatula A17. These genes could also alter the symbiosis between R. leguminosarum bv. viciae 3841 and pea or lentil, suggesting that this approach identifies new genes that may more generally contribute to symbiotic productivity.more » « less
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Abstract Legume roots engage in a symbiotic relationship with rhizobia, leading to the development of nitrogen-fixing nodules. Nodule development is a sophisticated process and is under the tight regulation of the plant. The symbiosis initiates with a signal exchange between the two partners, followed by the development of a new organ colonized by rhizobia. Over two decades of study have shed light on the transcriptional regulation of rhizobium–legume symbiosis. A large number of transcription factors (TFs) have been implicated in one or more stages of this symbiosis. Legumes must monitor nodule development amidst a dynamic physical environment. Some environmental factors are conducive to nodulation, whereas others are stressful. The modulation of rhizobium–legume symbiosis by the abiotic environment adds another layer of complexity and is also transcriptionally regulated. Several symbiotic TFs act as integrators between symbiosis and the response to the abiotic environment. In this review, we trace the role of various TFs involved in rhizobium–legume symbiosis along its developmental route and highlight the ones that also act as communicators between this symbiosis and the response to the abiotic environment. Finally, we discuss contemporary approaches to study TF-target interactions in plants and probe their potential utility in the field of rhizobium–legume symbiosis.
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ABSTRACT Symbiotic nitrogen fixation (SNF) in the interaction between the soil bacteria Sinorhizobium meliloti and legume plant Medicago sativa is carried out in specialized root organs called nodules. During nodule development, each symbiont must drastically alter their proteins, transcripts, and metabolites in order to support nitrogen fixation. Moreover, bacteria within the nodules are under stress, including challenges by plant antimicrobial peptides, low pH, limited oxygen availability, and strongly reducing conditions, all of which challenge proteome integrity. S. meliloti stress adaptation, proteome remodeling, and quality control are controlled in part by the large oligomeric protease complexes HslUV and ClpXP1. To improve understanding of the roles of S. meliloti HslUV and ClpXP1 under free-living conditions and in symbiosis with M. sativa , we generated Δ hslU , Δ hslV , Δ hslUV , and Δ clpP1 knockout mutants. The shoot dry weight of M. sativa plants inoculated with each deletion mutant was significantly reduced, suggesting a role in symbiosis. Further, slower free-living growth of the Δ hslUV and Δ clpP1 mutants suggests that HslUV and ClpP1 were involved in adapting to heat stress, the while Δ hslU and Δ clpP1 mutants were sensitive to kanamycin. All deletion mutants produced less exopolysaccharide and succinoglycan, as shown by replicate spot plating and calcofluor binding. We also generated endogenous C-terminal enhanced green fluorescent protein (eGFP) fusions to HslU, HslV, ClpX, and ClpP1 in S. meliloti . Using anti-eGFP antibodies, native coimmunoprecipitation experiments with proteins from free-living and nodule tissues were performed and analyzed by mass spectrometry. The results suggest that HslUV and ClpXP were closely associated with ribosomal and proteome quality control proteins, and they identified several novel putative protein-protein interactions. IMPORTANCE Symbiotic nitrogen fixation (SNF) is the primary means by which biologically available nitrogen enters the biosphere, and it is therefore a critical component of the global nitrogen cycle and modern agriculture. SNF is the result of highly coordinated interactions between legume plants and soil bacteria collectively referred to as rhizobia, e.g., Medicago sativa and S. meliloti , respectively. Accomplishing SNF requires significant proteome changes in both organisms to create a microaerobic environment suitable for high-level bacterial nitrogenase activity. The bacterial protease systems HslUV and ClpXP are important in proteome quality control, in metabolic remodeling, and in adapting to stress. This work shows that S. meliloti HslUV and ClpXP are involved in SNF, in exopolysaccharide production, and in free-living stress adaptation.more » « less
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null (Ed.)Leguminous plants form symbiotic relationships with rhizobia. These nitrogen-fixing bacteria live in specialized root organs called nodules. While rhizobia form the most notable host relationship within root nodules, other bacterial endophytes also inhabit these root nodules and can influence host-rhizobia interactions as well as exert effects of their own, whether beneficial or detrimental. In this study, we investigate differences in nodule communities between genotypes (A17 and R108) of a single plant species, the model legume Medicago truncatula. While diversity of endophytes in nodules was similar across hosts, both nodule endophyte composition and gene functional groups differed. In contrast to the significant direct effect of host genotype, neither the presence nor identity of a host in the previous generation (either A17 or R108) had a significant effect on the nodule endophyte diversity or composition. However, whether or not a host was present altered gene functional groups. We conclude that genetic variation within a legume host species can play an important role in the establishment of nodule microbiomes. Further studies, including GWAS and functional assays, can open the door for engineering and optimizing nodule endophyte communities that promote growth or have other beneficial qualities.more » « less
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/doi.org/10.1093/plphys/kiaa115
