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.
The establishment of the nitrogen‐fixing symbiosis between soybean and
- Award ID(s):
- 1734145
- NSF-PAR ID:
- 10456896
- Publisher / Repository:
- Wiley-Blackwell
- Date Published:
- Journal Name:
- The Plant Journal
- Volume:
- 103
- Issue:
- 5
- ISSN:
- 0960-7412
- Page Range / eLocation ID:
- p. 1937-1958
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
more » « less
Abstract -
more » « less
Summary Despite of important functions of strigolactones (SLs) and karrikins (KARs) in plant development, plant–parasite and plant–fungi interactions, their roles in soybean–rhizobia interaction remain elusive. SL/KAR signaling genes
GmMAX2a, GmD14s, andGmKAIs are activated by rhizobia infection. GmMAX2a restoredatmax2 root hair defects and soybean root hairs were changed inGmMAX2a overexpression (GmMAX2a ‐OE ) or knockdown (GmMAX2a ‐KD ) mutants.GmMAX2a ‐KD gave fewer, whereasGmMAX2a ‐OE produced more nodules than GUS hairy roots. Mutation ofGmMAX2a in itsKD orOE transgenic hairy roots affected the rhizobia infection‐induced increases in early nodulation gene expression. Both mutant hairy roots also displayed the altered auxin, jasmonate and abscisic acid levels, as further verified by transcriptomic analyses of their synthetic genes. Overexpression of an auxin synthetic geneGmYUC2a also affected SL and KAR signaling genes. GmMAX2a physically interacted with SL/KAR receptors GmD14s, GmKAIs, and GmD14Ls with different binding affinities, depending on variations in the critical amino acids, forming active D14/KAI‐SCFMAX2complexes. The knockdown mutant roots of the nodule‐specifically expressingGmKAI s andGmD14L s gave fewer nodules, with altered expression of several early nodulation genes. The expression levels ofGmKAI s, andGmD14L s were markedly changed inGmMAX2a mutant roots, so did their target repressor genesGmD53 s andGmSMAX1 s. Thus, SL and KAR signaling were involved in soybean–rhizobia interaction and nodulation partly through interactions with hormones, and this may explain the different effects of MXA2 orthologs on legume determinate and indeterminate nodulation. The study provides fresh insights into the roles of GmMAX2‐mediated SL/KAR signaling in soybean root hair and nodule formation. -
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
-
more » « less
Premise Polyploidy is a major genetic driver of ecological and evolutionary processes in plants, yet its effects on plant interactions with mutualistic microbes remain unresolved. The legume–rhizobium symbiosis regulates global nutrient cycles and plays a role in the diversification of legume species. In this mutualism, rhizobia bacteria fix nitrogen in exchange for carbon provided by legume hosts. This exchange occurs inside root nodules, which house bacterial cells and represent the interface of legume–rhizobium interactions. Although polyploidy may directly impact the legume–rhizobium mutualism, no studies have explored how it alters the internal structure of nodules.
Methods We created synthetic autotetraploids using
Medicago sativa subsp.caerulea . Neotetraploid plants and their diploid progenitors were singly inoculated with two strains of rhizobia,Sinorhizobium meliloti andS. medicae . Confocal microscopy was used to quantify internal traits of nodules produced by diploid and neotetraploid plants.Results Autotetraploid plants produced larger nodules with larger nitrogen fixation zones than diploids for both strains of rhizobia, although the significance of these differences was limited by power. Neotetraploid
M. sativa subsp.caerulea plants also produced symbiosomes that were significantly larger, nearly twice the size, than those present in diploids.Conclusions This study sheds light on how polyploidy directly affects a plant–bacterium mutualism and uncovers novel mechanisms. Changes in plant–microbe interactions that directly result from polyploidy likely contribute to the increased ability of polyploid legumes to establish in diverse environments.
-
more » « less
Abstract In agriculture, plant growth promoting bacteria (PGPB) are increasingly used for reducing environmental stress‐related crop losses through mutualistic actions of these microorganisms, activating physiological and biochemical responses, building tolerances within their hosts. Here we report the use of radioactive carbon‐11 (
t ½20.4 min) to examine the metabolic and physiological responses ofZea mays toAzospirillum brasilense (HM053) inoculation while plants were subjected to salinity and low nitrogen stresses. Host metabolism of “new” carbon resources (as11C) and physiology including [11C]‐photosynthate translocation were measured in response to imposed growth conditions. Salinity stress caused shortened, dense root growth with a 6‐fold increase in foliar [11C]‐raffinose, a potent osmolyte. ICP‐MS analyses revealed increased foliar Na+levels at the expense of K+. HM053 inoculation relieved these effects, reinstating normal root growth, lowering [11C]‐raffinose levels while increasing [11C]‐sucrose and its translocation to the roots. Na+levels remained elevated with inoculation, but K+levels were boosted slightly. Low nitrogen stress yielded longer roots possessing high levels of anthocyanins. Metabolic analysis revealed significant shifts in “new” carbon partitioning into the amino acid pool under low nitrogen stress, with significant increases in foliar [11C]‐glutamate, [11C]‐aspartate, and [11C]‐asparagine, a noted osmoprotectant.11CO2fixation and [11C]‐photosynthate translocation also decreased, limiting carbon supply to roots. However, starch levels in roots were reduced under nitrogen limitation, suggesting that carbon repartitioning could be a compensatory action to support root growth. Finally, inoculation with HM053 re‐instated normal root growth, reduced anthocyanin, boosted root starch, and returned11C‐allocation levels back to those of unstressed plants.
