GH3-mediated auxin inactivation attenuates multiple stages of lateral root development.

Lateral root (LR) positioning and development rely on the dynamic interplay between auxin production, transport but also inactivation. Nonetheless, how the latter affects LR organogenesis remains largely uninvestigated. Here, we systematically analyze the impact of the major auxin inactivation pathway defined by GRETCHEN HAGEN3-type (GH3) auxin conjugating enzymes and DIOXYGENASE FOR AUXIN OXIDATION1 (DAO1) in all stages of LR development using reporters, genetics and inhibitors in Arabidopsis thaliana. Our data demonstrate that the gh3.1/2/3/4/5/6 hextuple (gh3hex) mutants display a higher LR density due to increased LR initiation and faster LR developmental progression, acting epistatically over dao1-1. Grafting and local inhibitor applications reveal that root and shoot GH3 activities control LR formation. The faster LR development in gh3hex is associated with GH3 expression domains in and around developing LRs. The increase in LR initiation is associated with accelerated auxin response oscillations coinciding with increases in apical meristem size and LR cap cell death rates. Our research reveals how GH3-mediated auxin inactivation attenuates LR development. Local GH3 expression in LR primordia attenuates development and emergence, whereas GH3 effects on pre-initiation stages are indirect, by modulating meristem activities that in turn coordinate root growth with LR spacing.

Plastid-localized amino acid metabolism coordinates rice ammonium tolerance and nitrogen use efficiency.

Ammonium toxicity affecting plant metabolism and development is a worldwide problem impeding crop production. Remarkably, rice (Oryza sativa L.) favours ammonium as its major nitrogen source in paddy fields. We set up a forward-genetic screen to decipher the molecular mechanisms conferring rice ammonium tolerance and identified rohan showing root hypersensitivity to ammonium due to a missense mutation in an argininosuccinate lyase (ASL)-encoding gene. ASL localizes to plastids and its expression is induced by ammonium. ASL alleviates ammonium-inhibited root elongation by converting the excessive glutamine to arginine. Consequently, arginine leads to auxin accumulation in the root meristem, thereby stimulating root elongation under high ammonium. Furthermore, we identified natural variation in the ASL allele between japonica and indica subspecies explaining their different root sensitivity towards ammonium. Finally, we show that ASL expression positively correlates with root ammonium tolerance and that nitrogen use efficiency and yield can be improved through a gain-of-function approach.

Hydraulic flux-responsive hormone redistribution determines root branching.

Plant roots exhibit plasticity in their branching patterns to forage efficiently for heterogeneously distributed resources, such as soil water. The xerobranching response represses lateral root formation when roots lose contact with water. Here, we show that xerobranching is regulated by radial movement of the phloem-derived hormone abscisic acid, which disrupts intercellular communication between inner and outer cell layers through plasmodesmata. Closure of these intercellular pores disrupts the inward movement of the hormone signal auxin, blocking lateral root branching. Once root tips regain contact with moisture, the abscisic acid response rapidly attenuates. Our study reveals how roots adapt their branching pattern to heterogeneous soil water conditions by linking changes in hydraulic flux with dynamic hormone redistribution.

Root system size and root hair length are key phenes for nitrate acquisition and biomass production across natural variation in Arabidopsis.

The role of root phenes in nitrogen (N) acquisition and biomass production was evaluated in 10 contrasting natural accessions of Arabidopsis thaliana L. Seedlings were grown on vertical agar plates with two different nitrate supplies. The low N treatment increased the root to shoot biomass ratio and promoted the proliferation of lateral roots and root hairs. The cost of a larger root system did not impact shoot biomass. Greater biomass production could be achieved through increased root length or through specific root hair characteristics. A greater number of root hairs may provide a low-resistance pathway under elevated N conditions, while root hair length may enhance root zone exploration under low N conditions. The variability of N uptake and the expression levels of genes encoding nitrate transporters were measured. A positive correlation was found between root system size and high-affinity nitrate uptake, emphasizing the benefits of an exploratory root organ in N acquisition. The expression levels of NRT1.2/NPF4.6, NRT2.2, and NRT1.5/NPF7.3 negatively correlated with some root morphological traits. Such basic knowledge in Arabidopsis demonstrates the importance of root phenes to improve N acquisition and paves the way to design eudicot ideotypes.

A reflux-and-growth mechanism explains oscillatory patterning of lateral root branching sites.

Modular, repetitive structures are a key component of complex multicellular body plans across the tree of life. Typically, these structures are prepatterned by temporal oscillations in gene expression or signaling. Although a clock-and-wavefront mechanism was identified and plant leaf phyllotaxis arises from a Turing-type patterning for vertebrate somitogenesis and arthropod segmentation, the mechanism underlying lateral root patterning has remained elusive. To resolve this enigma, we combined computational modeling with in planta experiments. Intriguingly, auxin oscillations automatically emerge in our model from the interplay between a reflux-loop-generated auxin loading zone and stem-cell-driven growth dynamics generating periodic cell-size variations. In contrast to the clock-and-wavefront mechanism and Turing patterning, the uncovered mechanism predicts both frequency and spacing of lateral-root-forming sites to positively correlate with root meristem growth. We validate this prediction experimentally. Combined, our model and experimental results support that a reflux-and-growth patterning mechanism underlies lateral root priming.

An auxin-regulable oscillatory circuit drives the root clock in Arabidopsis.

In Arabidopsis, the root clock regulates the spacing of lateral organs along the primary root through oscillating gene expression. The core molecular mechanism that drives the root clock periodicity and how it is modified by exogenous cues such as auxin and gravity remain unknown. We identified the key elements of the oscillator (AUXIN RESPONSE FACTOR 7, its auxin-sensitive inhibitor IAA18/POTENT, and auxin) that form a negative regulatory loop circuit in the oscillation zone. Through multilevel computer modeling fitted to experimental data, we explain how gene expression oscillations coordinate with cell division and growth to create the periodic pattern of organ spacing. Furthermore, gravistimulation experiments based on the model predictions show that external auxin stimuli can lead to entrainment of the root clock. Our work demonstrates the mechanism underlying a robust biological clock and how it can respond to external stimuli.

GOLVEN peptide signalling through RGI receptors and MPK6 restricts asymmetric cell division during lateral root initiation.

During lateral root initiation, lateral root founder cells undergo asymmetric cell divisions that generate daughter cells with different sizes and fates, a prerequisite for correct primordium organogenesis. An excess of the GLV6/RGF8 peptide disrupts these initial asymmetric cell divisions, resulting in more symmetric divisions and the failure to achieve lateral root organogenesis. Here, we show that loss-of-function GLV6 and its homologue GLV10 increase asymmetric cell divisions during lateral root initiation, and we identified three members of the RGF1 INSENSITIVE/RGF1 receptor subfamily as likely GLV receptors in this process. Through a suppressor screen, we found that MITOGEN-ACTIVATED PROTEIN KINASE6 is a downstream regulator of the GLV pathway. Our data indicate that GLV6 and GLV10 act as inhibitors of asymmetric cell divisions and signal through RGF1 INSENSITIVE receptors and MITOGEN-ACTIVATED PROTEIN KINASE6 to restrict the number of initial asymmetric cell divisions that take place during lateral root initiation.

The dynamic nature and regulation of the root clock.

Plants explore the soil by continuously expanding their root system, a process that depends on the production of lateral roots (LRs). Sites where LRs can be produced are specified in the primary root axis through a pre-patterning mechanism, determined by a biological clock that is coordinated by temporal signals and positional cues. This 'root clock' generates an oscillatory signal that is translated into a developmental cue to specify a set of founder cells for LR formation. In this Review, we summarize recent findings that shed light on the mechanisms underlying the oscillatory signal and discuss how a periodic signal contributes to the conversion of founder cells into LR primordia. We also provide an overview of the phases of the root clock that may be influenced by endogenous factors, such as the plant hormone auxin, and by exogenous environmental cues. Finally, we discuss additional aspects of the root-branching process that act independently of the root clock.

Unraveling a Local Inhibitory Mechanism Safeguarding Regular Lateral Root Spacing.

The mechanisms underlying even spacing of lateral roots remains incompletely understood. In this issue of Developmental Cell, Toyokura et al. make an important step forward by showing how a local inhibitory mechanism involving a novel peptide hormone-receptor cascade acts as a safeguarding mechanism to regulate lateral root spacing.

A Comparative Study of Ethylene Emanation upon Nitrogen Deficiency in Natural Accessions of Arabidopsis thaliana.

An original approach to develop sustainable agriculture with less nitrogen fertilizer inputs is to tackle the cross-talk between nitrogen nutrition and plant growth regulators. In particular the gaseous hormone, ethylene, is a prime target for that purpose. The variation of ethylene production in natural accessions of the model species Arabidopsis thaliana was explored in response to the nitrate supply. Ethylene was measured with a laser-based photoacoustic detector. First, experimental conditions were established with Columbia-0 (Col-0) accession, which was grown in vitro on horizontal plates across a range of five nitrate concentrations (0.5, 1, 2.5, 5, or 10 mM). The concentrations of 1 and 10 mM nitrate were retained for further characterization. Along with a decrease of total dry biomass and higher biomass allocation to the roots, the ethylene production was 50% more important at 1 mM than at 10 mM nitrate. The total transcript levels of 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID SYNTHASES (ACS) in roots and those of ACC OXIDASES (ACO) in shoots increased by 100% between the same treatments. This was mainly due to higher transcript levels of ACS6 and of ACO2 and ACO4 respectively. The assumption was that during nitrogen deficiency, the greater biomass allocation in favor of the roots was controlled by ethylene being released in the shoots after conversion of ACC originating from the roots. Second, biomass and ethylene productions were measured in 20 additional accessions. Across all accessions, the total dry biomass and ethylene production were correlated negatively at 1 mM but positively at 10 mM nitrate. Furthermore, polymorphism was surveyed in ACC and ethylene biosynthesis genes and gene products among accessions. Very few substitutions modifying the amino acids properties in conserved motifs of the enzymes were found in the accessions. Natural variation of ethylene production could be further explored to improve Nitrogen Use Efficiency (NUE), in particular by manipulating features like the biomass production and the timing of senescence upon nitrogen limitation.