Bruno Touraine

N Demand and the Regulation of Nitrate Uptake

Uptake of nitrate by root cells followed by reduction and assimilation in plant tissues is the main route by which mineral N is converted into organic N by living organisms. Like photosynthesis, these are life-dependent processes that members of the animal kingdom are unable to perform for themselves. Nitrate and other mineral nutrients required for optimal plant growth and development frequently exist at relatively low concentrations in soil. To thrive on these dilute nutrients, plants have developed high-performance uptake systems in their root cells. To cope with wide variations in mineral concentrations in soil, plants have evolved mechanisms to regulate the activity of uptake systems so that net intake of a nutrient depends on the plants need for this element rather than its concentration in the rooting medium. Indeed, uptake rates of most ions are seemingly controlled by specific demand-driven regulatory mechanisms. Such processes set the uptake rate of a given element to match the plants current growth rate and developmental stage. Nitrate uptake is of special interest because nitrate is absorbed at a relatively high rate and because compounds that function as uptake sensors may have been identified. This paper focuses on whole-plant signaling processes involved in the regulation of nitrate uptake by N demand.

Plant growth-promoting rhizobacteria and root system functioning

The rhizosphere supports the development and activity of a huge and diversified microbial community, including microorganisms capable to promote plant growth. Among the latter, plant growth-promoting rhizobacteria (PGPR) colonize roots of monocots and dicots, and enhance plant growth by direct and indirect mechanisms. Modification of root system architecture by PGPR implicates the production of phytohormones and other signals that lead, mostly, to enhanced lateral root branching and development of root hairs. PGPR also modify root functioning, improve plant nutrition and influence the physiology of the whole plant. Recent results provided first clues as to how PGPR signals could trigger these plant responses. Whether local and/or systemic, the plant molecular pathways involved remain often unknown. From an ecological point of view, it emerged that PGPR form coherent functional groups, whose rhizosphere ecology is influenced by a myriad of abiotic and biotic factors in natural and agricultural soils, and these factors can in turn modulate PGPR effects on roots. In this paper, we address novel knowledge and gaps on PGPR modes of action and signals, and highlight recent progress on the links between plant morphological and physiological effects induced by PGPR. We also show the importance of taking into account the size, diversity, and gene expression patterns of PGPR assemblages in the rhizosphere to better understand their impact on plant growth and functioning. Integrating mechanistic and ecological knowledge on PGPR populations in soil will be a prerequisite to develop novel management strategies for sustainable agriculture.

Regulation of the nitrate transporter gene AtNRT2.1 in Arabidopsis thaliana: responses to nitrate, amino acids and developmental stage

AbstractThe NRT2.1 gene codes for a high-affinity nitrate transporter in Arabidopsis thaliana. To examine the regulation of NRT2.1 gene expression, we used a promoter-β-glucuronidase (GUS) fusion and found that the NRT2.1 promoter directs expression to the epidermal, cortical and endodermal cell layers of mature root parts. The gene appeared to be expressed essentially in roots, but was also present in the leaf hydathodes. Investigation of NRT2.1 expression pattern during the plant developmental cycle showed that it increased rapidly during early vegetative growth, peaked prior to floral stem emergence, and decreased to very low levels in flowering and silique-bearing plants. Experiments with various nitrogen supply regimes demonstrated the induction of NRT2.1 expression by nitrate and repression by amino acids. Amino acid analysis showed that this repression was specifically related to increased internal glutamine, suggesting a role for this particular amino acid in nitrogen signalling responsible for nitrate uptake regulation. Taken together, our results support the hypothesis that the NRT2.1 gene codes for a major component of the inducible high-affinity transport system for nitrate, which is spatially and developmentally controlled at the transcriptional level. Surprisingly, NRT2.1 was not expressed in younger root parts, although a similar rate of nitrate influx was observed in both young and old root samples. This lack of correlation between nitrate influx and NRT2.1 expression suggests that another high-affinity nitrate transporter operates in root tips. Abbreviation: GUS, β-glucuronidase

Cloning and characterization of a root specific high-affinity sulfate transporter from Arabidopsis thaliana

Hst1At (accession number AB018695) was identified from the Arabidopsis thaliana sequencing project on BAC T3F12, and the corresponding cDNA was isolated by reverse transcription‐PCR. Southern blot analysis reveals a single copy of this gene. The cDNA encodes a root specific sulfate transporter of 649 amino acids. Heterologous expression of hst1At in a sulfate transport deficient yeast mutant shows that this gene encodes a high‐affinity transport system (∼2 μM). The transcript relative abundance increases in roots in response to sulfate deprivation, which correlated with increased root SO4 2− influx capacity. These patterns were reversed upon sulfate addition to the medium and were accompanied by an increased glutathione level in roots. Feeding plants with cysteine or glutathione led to similar responses. Using buthionine sulfoximine, an inhibitor of glutathione synthesis, we show that glutathione rather than cysteine controls hst1At expression.

Simultaneous Interaction of Arabidopsis thaliana with Bradyrhizobium Sp. Strain ORS278 and Pseudomonas syringae pv. tomato DC3000 Leads to Complex Transcriptome Changes

Induced systemic resistance (ISR) is a process elicited by telluric microbes, referred to as plant growth-promoting rhizobacteria (PGPR), that protect the host plant against pathogen attacks. ISR has been defined from studies using Pseudomonas strains as the biocontrol agent. Here, we show for the first time that a photosynthetic Bradyrhizobium sp. strain, ORS278, also exhibits the ability to promote ISR in Arabidopsis thaliana, indicating that the ISR effect may be a widespread ability. To investigate the molecular bases of this response, we performed a transcriptome analysis designed to reveal the changes in gene expression induced by the PGPR, the pathogen alone, or by both. The results confirm the priming pattern of ISR described previously, meaning that a set of genes, of which the majority was predicted to be influenced by jasmonic acid or ethylene, was induced upon pathogen attack when plants were previously colonized by PGPR. The analysis and interpretation of transcriptome data revealed that 12-oxo-phytodienoic acid, an intermediate of the jasmonic acid biosynthesis pathway, is likely to be an actor in the signaling cascade involved in ISR. In addition, we show that the PGPR counterbalanced the pathogen-induced changes in expression of a series of genes.

Nitrate Uptake and Its Regulation

NO3 - uptake by the roots of higher plants is the main pathway for entry of N into the global food chain. In quantitative terms, N is the most important element after C, H and O, i.e. the most important of the mineral elements that have to be acquired from the soil. It is thus not surprising that it commonly limits plant growth, especially in agricultural systems, because a large proportion of the N accumulated in crops is removed from the plant-soil system at harvest. Since the mid-19th century, the use of NO3 - — based fertilisers has continually increased to sustain high crop yields.

The auxin-signaling pathway is required for the lateral root response of Arabidopsis to the rhizobacterium Phyllobacterium brassicacearum

Plant root development is highly responsive both to changes in nitrate availability and beneficial microorganisms in the rhizosphere. We previously showed that Phyllobacterium brassicacearum STM196, a plant growth-promoting rhizobacteria strain isolated from rapeseed roots, alleviates the inhibition exerted by high nitrate supply on lateral root growth. Since soil-borne bacteria can produce IAA and since this plant hormone may be implicated in the high nitrate-dependent control of lateral root development, we investigated its role in the root development response of Arabidopsis thaliana to STM196. Inoculation with STM196 resulted in a 50% increase of lateral root growth in Arabidopsis wild-type seedlings. This effect was completely abolished in aux1 and axr1 mutants, altered in IAA transport and signaling, respectively, indicating that these pathways are required. The STM196 strain, however, appeared to be a very low IAA producer when compared with the high-IAA-producing Azospirillum brasilense sp245 strain and its low-IAA-producing ipdc mutant. Consistent with the hypothesis that STM196 does not release significant amounts of IAA to the host roots, inoculation with this strain failed to increase root IAA content. Inoculation with STM196 led to increased expression levels of several IAA biosynthesis genes in shoots, increased Trp concentration in shoots, and increased auxin-dependent GUS staining in the root apices of DR5::GUS transgenic plants. All together, our results suggest that STM196 inoculation triggers changes in IAA distribution and homeostasis independently from IAA release by the bacteria.

The PGPR strain Phyllobacterium brassicacearum STM196 induces a reproductive delay and physiological changes that result in improved drought tolerance in Arabidopsis

Understanding how biotic interactions can improve plant tolerance to drought is a challenging prospect for agronomy and ecology. Plant growth-promoting rhizobacteria (PGPR) are promising candidates but the phenotypic changes induced by PGPR under drought remain to be elucidated. We investigated the effects of Phyllobacterium brassicacearum STM196 strain, a PGPR isolated from the rhizosphere of oilseed rape, on two accessions of Arabidopsis thaliana with contrasting flowering time. We measured multiple morphophysiological traits related to plant growth and development in order to quantify the added value of the bacteria to drought-response strategies of Arabidopsis in soil conditions. A delay in reproductive development induced by the bacteria resulted in a gain of biomass that was independent of the accession and the watering regime. Coordinated changes in transpiration, ABA content, photosynthesis and development resulted in higher water-use efficiency and a better tolerance to drought of inoculated plants. Our findings give new insights into the ecophysiological bases by which PGPR can confer stress tolerance to plants. Rhizobacteria-induced delay in flowering time could represent a valuable strategy for increasing biomass yield, whereas rhizobacteria-induced improvement of water use is of particular interest in multiple scenarios of water availability.

PGPR-Arabidopsis interactions is a useful system to study signaling pathways involved in plant developmental control

Using their 1 amino cyclopropane-1-carboxylic acid (ACC) deaminase activity, many rhizobacteria can divert ACC from the ethylene biosynthesis pathway in plant roots. To investigate the role of this microbial activity in plant responses to plant growth-promoting rhizobacteria (PGPR), we analyzed the effects of acdS knock-out and wild-type PGPR strains on two phenotypic responses to inoculation –root hair elongation and root system architecture– in Arabidopsis thaliana. Our work shows that rhizobacterial AcdS activity has a negative effect on root hair elongation, as expected from the reduction of ethylene production rate in root cells, while it has no impact on root system architecture. This suggests that PGPR triggered root hair elongation is independent of ethylene biosynthesis or signaling pathway. In addition, it does indicate that AcdS activity alters local regulatory processes, but not systemic regulations such as those that control root architecture. Our work also indicates that root hair elongation induced by PGPR inoculation is probably an auxin-independent mechanism. These findings were unexpected since genetic screens for abnormal root hair development mutants led to the isolation of ethylene and auxin mutants. Our work hence shows that studying the interaction between a PGPR and the model plant Arabidopsis is a useful system to uncover new pathways involved in plant plasticity.

The NRT2.5 and NRT2.6 genes are involved in growth promotion of Arabidopsis by the plant growth‐promoting rhizobacterium (PGPR) strain Phyllobacterium brassicacearum STM196

The Phyllobacterium brassicacearum STM196 strain stimulates Arabidopsis thaliana growth and antagonizes high nitrate inhibition of lateral root development. A previous study identified two STM196-responsive genes, NRT2.5 and NRT2.6 (Mantelin et al., 2006, Planta 223: 591-603). We investigated the role of NRT2.5 and NRT2.6 in the plant response to STM196 using single and double Arabidopsis mutants. The single mutants were also crossed with an nrt2.1 mutant, lacking the major nitrate root transporter, to distinguish the effects of NRT2.5 and NRT2.6 from potential indirect effects of nitrate pools. The nrt2.5 and nrt2.6 mutations abolished the plant growth and root system architecture responses to STM196. The determination of nitrate content revealed that NRT2.5 and NRT2.6 do not play an important role in nitrate distribution between plant organs. Conversely, NRT2.5 and NRT2.6 appeared to play a role in the plant response independent of nitrate uptake. Using a nitrate reductase mutant, it was confirmed that the NRT2.5/NRT2.6-dependent plant signalling pathway is independent of nitrate-dependent regulation of root development. Our findings demonstrate that NRT2.5 and NRT2.6, which are preferentially expressed in leaves, play an essential role in plant growth promotion by the rhizospheric bacterium STM196.