Gabriel Krouk

Nitrate-Regulated Auxin Transport by NRT1.1 Defines a Mechanism for Nutrient Sensing in Plants

Nitrate is both a nitrogen source for higher plants and a signal molecule regulating their development. In Arabidopsis, the NRT1.1 nitrate transporter is crucial for nitrate signaling governing root growth, and has been proposed to act as a nitrate sensor. However, the sensing mechanism is unknown. Herein we show that NRT1.1 not only transports nitrate but also facilitates uptake of the phytohormone auxin. Moreover, nitrate inhibits NRT1.1-dependent auxin uptake, suggesting that transduction of nitrate signal by NRT1.1 is associated with a modification of auxin transport. Among other effects, auxin stimulates lateral root development. Mutation of NRT1.1 enhances both auxin accumulation in lateral roots and growth of these roots at low, but not high, nitrate concentration. Thus, we propose that NRT1.1 represses lateral root growth at low nitrate availability by promoting basipetal auxin transport out of these roots. This defines a mechanism connecting nutrient and hormone signaling during organ development.

Nitrate signaling: adaptation to fluctuating environments

Nitrate (NO(3)(-)) is a key nutrient as well as a signaling molecule that impacts both metabolism and development of plants. Understanding the complexity of the regulatory networks that control nitrate uptake, metabolism, and associated responses has the potential to provide solutions that address the major issues of nitrate pollution and toxicity that threaten agricultural and ecological sustainability and human health. Recently, major advances have been made in cataloguing the nitrate transcriptome and in identifying key components that mediate nitrate signaling. In this perspective, we describe the genes involved in nitrate regulation and how they influence nitrate transport and assimilation, and we discuss the role of systems biology approaches in elucidating the gene networks involved in NO(3)(-) signaling adaptation to fluctuating environments.

Nitrogen economics of root foraging: Transitive closure of the nitrate–cytokinin relay and distinct systemic signaling for N supply vs. demand

As sessile organisms, root plasticity enables plants to forage for and acquire nutrients in a fluctuating underground environment. Here, we use genetic and genomic approaches in a “split-root” framework—in which physically isolated root systems of the same plant are challenged with different nitrogen (N) environments—to investigate how systemic signaling affects genome-wide reprogramming and root development. The integration of transcriptome and root phenotypes enables us to identify distinct mechanisms underlying “N economy” (i.e., N supply and demand) of plants as a system. Under nitrate-limited conditions, plant roots adopt an “active-foraging strategy”, characterized by lateral root outgrowth and a shared pattern of transcriptome reprogramming, in response to either local or distal nitrate deprivation. By contrast, in nitrate-replete conditions, plant roots adopt a “dormant strategy”, characterized by a repression of lateral root outgrowth and a shared pattern of transcriptome reprogramming, in response to either local or distal nitrate supply. Sentinel genes responding to systemic N signaling identified by genome-wide comparisons of heterogeneous vs. homogeneous split-root N treatments were used to probe systemic N responses in Arabidopsis mutants impaired in nitrate reduction and hormone synthesis and also in decapitated plants. This combined analysis identified genetically distinct systemic signaling underlying plant N economy: (i) N supply, corresponding to a long-distance systemic signaling triggered by nitrate sensing; and (ii) N demand, experimental support for the transitive closure of a previously inferred nitrate–cytokinin shoot–root relay system that reports the nitrate demand of the whole plant, promoting a compensatory root growth in nitrate-rich patches of heterogeneous soil.

A framework integrating plant growth with hormones and nutrients

It is well known that nutrient availability controls plant development. Moreover, plant development is finely tuned by a myriad of hormonal signals. Thus, it is not surprising to see increasing evidence of coordination between nutritional and hormonal signaling. In this opinion article, we discuss how nitrogen signals control the hormonal status of plants and how hormonal signals interplay with nitrogen nutrition. We further expand the discussion to include other nutrient-hormone pairs. We propose that nutrition and growth are linked by a multi-level, feed-forward cycle that regulates plant growth, development and metabolism via dedicated signaling pathways that mediate nutrient and hormonal regulation. We believe this model will provide a useful concept for past and future research in this field.

Nitrate transceptor(s) in plants

The availability of mineral nutrients in the soil dramatically fluctuates in both time and space. In order to optimize their nutrition, plants need efficient sensing systems that rapidly signal the local external concentrations of the individual nutrients. Until recently, the most upstream actors of the nutrient signalling pathways, i.e. the sensors/receptors that perceive the extracellular nutrients, were unknown. In Arabidopsis, increasing evidence suggests that, for nitrate, the main nitrogen source for most plant species, a major sensor is the NRT1.1 nitrate transporter, also contributing to nitrate uptake by the roots. Membrane proteins that fulfil a dual nutrient transport/signalling function have been described in yeast and animals, and are called transceptors. This review aims to illustrate the nutrient transceptor concept in plants by presenting the current evidence indicating that NRT1.1 is a representative of this class of protein. The various facets, as well as the mechanisms of nitrate sensing by NRT1.1 are considered, and the possible occurrence of other nitrate transceptors is discussed.

ABA transport and transporters

Abscisic acid (ABA) metabolism, perception, and transport form a triptych allowing higher plants to use ABA as a signaling molecule. The molecular bases of ABA metabolism are now well described and, over the past few years, several ABA receptors have been discovered. Although ABA transport has long been demonstrated in planta, the first breakthroughs in identifying plasma membrane-localized ABA transporters came in 2010, with the identification of two ATP-binding cassette (ABC) proteins. More recently, two ABA transporters in the nitrate transporter 1/peptide transporter (NRT1/PTR) family have been identified. In this review, we discuss the role of these different ABA transporters and examine the scientific impact of their identification. Given that the NRT1/PTR family is involved in the transport of nitrogen (N) compounds, further work should determine whether an interaction between ABA and N signaling or nutrition occurs.

Predictive network modeling of the high-resolution dynamic plant transcriptome in response to nitrate

BackgroundNitrate, acting as both a nitrogen source and a signaling molecule, controls many aspects of plant development. However, gene networks involved in plant adaptation to fluctuating nitrate environments have not yet been identified.ResultsHere we use time-series transcriptome data to decipher gene relationships and consequently to build core regulatory networks involved in Arabidopsis root adaptation to nitrate provision. The experimental approach has been to monitor genome-wide responses to nitrate at 3, 6, 9, 12, 15 and 20 minutes using Affymetrix ATH1 gene chips. This high-resolution time course analysis demonstrated that the previously known primary nitrate response is actually preceded by a very fast gene expression modulation, involving genes and functions needed to prepare plants to use or reduce nitrate. A state-space model inferred from this microarray time-series data successfully predicts gene behavior in unlearnt conditions.ConclusionsThe experiments and methods allow us to propose a temporal working model for nitrate-driven gene networks. This network model is tested both in silico and experimentally. For example, the over-expression of a predicted gene hub encoding a transcription factor induced early in the cascade indeed leads to the modification of the kinetic nitrate response of sentinel genes such as NIR, NIA2, and NRT1.1, and several other transcription factors. The potential nitrate/hormone connections implicated by this time-series data are also evaluated.

Multiple mechanisms of nitrate sensing by Arabidopsis nitrate transceptor NRT1.1

In Arabidopsis the plasma membrane nitrate transceptor (transporter/receptor) NRT1.1 governs many physiological and developmental responses to nitrate. Alongside facilitating nitrate uptake, NRT1.1 regulates the expression levels of many nitrate assimilation pathway genes, modulates root system architecture, relieves seed dormancy and protects plants from ammonium toxicity. Here, we assess the functional and phenotypic consequences of point mutations in two key residues of NRT1.1 (P492 and T101). We show that the point mutations differentially affect several of the NRT1.1-dependent responses to nitrate, namely the repression of lateral root development at low nitrate concentrations, and the short-term upregulation of the nitrate-uptake gene NRT2.1, and its longer-term downregulation, at high nitrate concentrations. We also show that these mutations have differential effects on genome-wide gene expression. Our findings indicate that NRT1.1 activates four separate signalling mechanisms, which have independent structural bases in the protein. In particular, we present evidence to suggest that the phosphorylated and non-phosphorylated forms of NRT1.1 at T101 have distinct signalling functions, and that the nitrate-dependent regulation of root development depends on the phosphorylated form. Our findings add to the evidence that NRT1.1 is able to trigger independent signalling pathways in Arabidopsis in response to different environmental conditions.

Nitrate Transport, Sensing, and Responses in Plants

Nitrogen (N) is an essential macronutrient that affects plant growth and development. N is an important component of chlorophyll, amino acids, nucleic acids, and secondary metabolites. Nitrate is one of the most abundant N sources in the soil. Because nitrate and other N nutrients are often limiting, plants have developed sophisticated mechanisms to ensure adequate supply ofxa0nutrients in a variable environment. Nitrate is absorbed in the root and mobilized to other organs by nitrate transporters. Nitrate sensing activates signaling pathways that impinge upon molecular, metabolic, physiological, and developmental responses locally and at the whole plant level. With the advent of genomics technologies and genetic tools, important advances in our understanding of nitrate and other N nutrient responses have been achieved in the past decade. Furthermore, techniques that take advantage of natural polymorphisms present in divergent individuals from a single species have been essential in uncovering new components. However, there are still gaps in our understanding of how nitrate signaling affects biological processes in plants. Moreover, we still lack an integrated view of how all the regulatory factors identified interact or crosstalk to orchestrate the myriad N responses plants typically exhibit. In this review, we provide an updated overview of mechanisms by which nitrate is sensed and transported throughout the plant. We discuss signaling components and how nitrate sensing crosstalks with hormonal pathways for developmental responsesxa0locally and globally in the plant. Understanding how nitrate impacts on plant metabolism, physiology, and growth and development in plants is key to improving crops for sustainable agriculture.

Hit-and-run transcriptional control by bZIP1 mediates rapid nutrient signaling in Arabidopsis

Significance Cellular signals evoke rapid and broad changes in gene regulatory networks. To uncover these network dynamics, we developed an approach able to monitor primary targets of a transcription factor (TF) based solely on gene regulation, in the absence of detectable binding. This enabled us to follow the transient propagation of a nitrogen (N) nutrient signal as a direct impact of the master TF Basic Leucine Zipper 1 (bZIP1). Unexpectedly, the largest class of primary targets that exhibit transient associations with bZIP1 is uniquely relevant to the rapid and dynamic propagation of the N signal. Our ability to uncover this transient network architecture has revealed the “dark matter” of dynamic N nutrient signaling in plants that has previously eluded detection. The dynamic nature of gene regulatory networks allows cells to rapidly respond to environmental change. However, the underlying temporal connections are missed, even in kinetic studies, as transcription factor (TF) binding within at least one time point is required to identify primary targets. The TF-regulated but unbound genes are dismissed as secondary targets. Instead, we report that these genes comprise transient TF–target interactions most relevant to rapid signal transduction. We temporally perturbed a master TF (Basic Leucine Zipper 1, bZIP1) and the nitrogen (N) signal it transduces and integrated TF regulation and binding data from the same cell samples. Our enabling approach could identify primary TF targets based solely on gene regulation, in the absence of TF binding. We uncovered three classes of primary TF targets: (i) poised (TF-bound but not TF-regulated), (ii) stable (TF-bound and TF-regulated), and (iii) transient (TF-regulated but not TF-bound), the largest class. Unexpectedly, the transient bZIP1 targets are uniquely relevant to rapid N signaling in planta, enriched in dynamic N-responsive genes, and regulated by TF and N signal interactions. These transient targets include early N responders nitrate transporter 2.1 and NIN-like protein 3, bound by bZIP1 at 1–5 min, but not at later time points following TF perturbation. Moreover, promoters of these transient targets are uniquely enriched with cis-regulatory motifs coinherited with bZIP1 binding sites, suggesting a recruitment role for bZIP1. This transient mode of TF action supports a classic, but forgotten, “hit-and-run” transcription model, which enables a “catalyst TF” to activate a large set of targets within minutes of signal perturbation.