Legumes can acquire nitrogen (N) from NO 3 2 , NH 4 1 , and N 2 (through symbiosis with Rhizobium bacteria); however, the mechanisms by which uptake and assimilation of these N forms are coordinately regulated to match the N demand of the plant are currently unknown. Here, we find by use of the split-root approach in Medicago truncatula plants that NO 3 2 uptake, NH 4 1 uptake, and N 2 fixation are under general control by systemic signaling of plant N status. Indeed, irrespective of the nature of the N source, N acquisition by one side of the root system is repressed by high N supply to the other side. Transcriptome analysis facilitated the identification of over 3,000 genes that were regulated by systemic signaling of the plant N status. However, detailed scrutiny of the data revealed that the observation of differential gene expression was highly dependent on the N source. Localized N starvation results, in the unstarved roots of the same plant, in a strong compensatory up-regulation of NO 3 2 uptake but not of either NH 4 1 uptake or N 2 fixation. This indicates that the three N acquisition pathways do not always respond similarly to a change in plant N status. When taken together, these data indicate that although systemic signals of N status control root N acquisition, the regulatory gene networks targeted by these signals, as well as the functional response of the N acquisition systems, are predominantly determined by the nature of the N source.

Summary• Adaptation of Medicago truncatula to local nitrogen (N) limitation was investigated to provide new insights into local and systemic N signaling.• The split-root technique allowed a characterization of the local and systemic responses of NO 3 ) or N 2 -fed plants to localized N limitation. 15 N and 13 C labeling were used to monitor plant nutrition. Plants expressing pMtENOD11-GUS and the sunn-2 hypernodulating mutant were used to unravel mechanisms involved in these responses.• Unlike NO 3 ) -fed plants, N 2 -fixing plants lacked the ability to compensate rapidly for a localized N limitation by up-regulating the N 2 -fixation activity of roots supplied elsewhere with N. However they displayed a long-term response via a growth stimulation of pre-existing nodules, and the generation of new nodules, likely through a decreased abortion rate of early nodulation events. Both these responses involve systemic signaling. The latter response is abolished in the sunn mutant, but the mutation does not prevent the first response.• Local but also systemic regulatory mechanisms related to plant N status regulate de novo nodule development in Mt, and SUNN is required for this systemic regulation. By contrast, the stimulation of nodule growth triggered by systemic N signaling does not involve SUNN, indicating SUNN-independent signaling.

The fluxes of (1) exogenous nitrogen (N) assimilation and (2) remobilization of endogenous N from vegetative plant compartments were measured by 15 N labeling during the seed-filling period in pea (Pisum sativum L. cv Caméor), to better understand the mechanism of N remobilization. While the majority (86%) of exogenous N was allocated to the vegetative organs before the beginning of seed filling, this fraction decreased to 45% at the onset of seed filling, the remainder being directed to seeds. Nitrogen remobilization from vegetative parts contributed to 71% of the total N in mature seeds borne on the first two nodes (first stratum). The contribution of remobilized N to total seed N varied, with the highest proportion at the beginning of filling; it was independent of the developmental stage of each stratum of seeds, suggesting that remobilized N forms a unique pool, managed at the whole-plant level and supplied to all filling seeds whatever their position on the plant. Once seed filling starts, N is remobilized from all vegetative organs: 30% of the total N accumulated in seeds was remobilized from leaves, 20% from pod walls, 11% from roots, and 10% from stems. The rate of N remobilization was maximal when seeds of all the different strata were filling, consistent with regulation according to the N demand of seeds. At later stages of seed filling, the rate of remobilization decreases and may become controlled by the amount of residual N in vegetative tissues.Pea (Pisum sativum) is an important agricultural crop grown primarily for its high seed protein content. However, the protein yield of the pea crop remains too low and variable between cropping area and years (http://apps.fao/faostat/, http://www.prolea.com/ unip/) to sustain the needs in plant protein of European countries. To extend the pea crop in Europe and to increase use of pea seed in the feed industry, breeders have to develop varieties with better harvest and nitrogen (N) indices. Toward this aim, a better understanding of the processes involved in the elaboration of seed protein content is needed. The final protein yield of seeds depends both upon the genotype and on environmental factors during seed filling (Lhuillier-Soundélé et al., 1999a). Nitrogen accumulation by seeds during filling depends upon the external N supply: soil mineral N assimilation and/or symbiotic fixation of atmospheric N 2 . However, exogenous N cannot generally sustain the high N demand of filling seeds, so endogenous N previously accumulated in vegetative parts is largely remobilized to fulfill this demand (Sinclair and de Wit, 1976;Salon et al., 2001). Seed N concentration is correlated to N availability within plants, and the extent of the contribution of N remobilization to seed N yield varies among legumes: 70% in field-grown pea (Atta et al., 2004), 80% to 90% in soybean (Glycine max; Warembourg and Fernandez, 1985;Grandgirard, 2002), 43% to 94% in rain-fed grown lentil (Lens culinaris; Kurdali et al., 1997), 84% in bean plants (Phaseolus vulgaris;Westermann et al., 1...

BackgroundIn order to maintain high yields while saving water and preserving non-renewable resources and thus limiting the use of chemical fertilizer, it is crucial to select plants with more efficient root systems. This could be achieved through an optimization of both root architecture and root uptake ability and/or through the improvement of positive plant interactions with microorganisms in the rhizosphere. The development of devices suitable for high-throughput phenotyping of root structures remains a major bottleneck.ResultsRhizotrons suitable for plant growth in controlled conditions and non-invasive image acquisition of plant shoot and root systems (RhizoTubes) are described. These RhizoTubes allow growing one to six plants simultaneously, having a maximum height of 1.1 m, up to 8 weeks, depending on plant species. Both shoot and root compartment can be imaged automatically and non-destructively throughout the experiment thanks to an imaging cabin (RhizoCab). RhizoCab contains robots and imaging equipment for obtaining high-resolution pictures of plant roots. Using this versatile experimental setup, we illustrate how some morphometric root traits can be determined for various species including model (Medicago truncatula), crops (Pisum sativum, Brassica napus, Vitis vinifera, Triticum aestivum) and weed (Vulpia myuros) species grown under non-limiting conditions or submitted to various abiotic and biotic constraints. The measurement of the root phenotypic traits using this system was compared to that obtained using “classic” growth conditions in pots.ConclusionsThis integrated system, to include 1200 Rhizotubes, will allow high-throughput phenotyping of plant shoots and roots under various abiotic and biotic environmental conditions. Our system allows an easy visualization or extraction of roots and measurement of root traits for high-throughput or kinetic analyses. The utility of this system for studying root system architecture will greatly facilitate the identification of genetic and environmental determinants of key root traits involved in crop responses to stresses, including interactions with soil microorganisms.