Abstract
Symbiotic nitrogen fixation (SNF) between rhizobia and legumes requires metabolic coordination within specialized root organs called nodules. Nodules formed in the symbiosis between S. medicae and barrel medic (M. truncatula) are indeterminate, cylindrical, and contain spatially distinct developmental zones. Bacteria in the infection zone II (ZII), interzone II-III (IZ), and nitrogen fixation zone III (ZIII) represent different stages in the metabolic progression from free-living bacteria into nitrogen fixing bacteroids. To better understand the coordination of plant and bacterial metabolism within the nodule, we used liquid and gas chromatography coupled to tandem mass spectrometry (MS) to observe protein and metabolite profiles representative of ZII, IZ, ZIII, whole-nodule, and primary root. Our MS-based approach confidently identified 361 S. medicae proteins and 888 M. truncatula proteins, as well as 160 metabolites from each tissue. The data are consistent with several organ- and zone-specific protein and metabolite localization patterns characterized previously. We used our comprehensive dataset to demonstrate how multiple branches of primary metabolism are coordinated between symbionts and zones, including central carbon, fatty acid, and amino acid metabolism. For example, M. truncatula glycolysis enzymes accumulate from zone I to zone III within the nodule, while equivalent S. medicae enzymes decrease in abundance. We also show the localization of S. medicae's transition to dicarboxylic acid-dependent carbon metabolism within the IZ. The spatial abundance patterns of S. medicae fatty acid (FA) biosynthesis enzymes indicate an increased demand for FA production in the IZ and ZIII as compared to ZI. These observations provide a resource for those seeking to understand coordinated physiological changes during the development of SNF.
Highlights
The largest contribution of nitrogen into the biosphere results from symbiotic interactions between legumes and soil bacteria of the family Rhizobiaceae [1,2,3]
We report here the use of nano-ultra performance liquid chromatography (UPLC) coupled to label-free mass spectrometry (MS) to catalogue the protein profiles of whole nodules, roots, and three developmental fractions of nodules formed in the M. truncatula-S. medicae plantbacterial system
Highly distinct protein profiles of five plant and bacterial symbiotic nitrogen fixation (SNF) tissues To evaluate the physiology of tissues associated with SNF, we developed a method for generating comprehensive protein profiles using label free nano-ultra performance liquid chromatography coupled to mass spectrometry (MS)
Summary
The largest contribution of nitrogen into the biosphere results from symbiotic interactions between legumes and soil bacteria of the family Rhizobiaceae [1,2,3]. The developmental transition leading to SNF is initiated by nitrogen deficient legumes, which release flavonoids and other diffusible metabolites into the rhizosphere [5]. Rhizobia perceive these signals and respond by excreting Nod factors, lipochitooligosaccharides (LCOs) that interact with plant lysin-motif (LysM) receptors to stimulate the common symbiotic pathway (CSP) [6,7]. Within the infected plant cells, rhizobia begin a metabolic transition from bacteria to bacteroids, a differentiated state that is capable of fixing nitrogen [10]
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