Ethylene signaling increases reactive oxygen species accumulation to drive root hair initiation in Arabidopsis.

Root hair initiation is a highly regulated aspect of root development. The plant hormone ethylene and its precursor, 1-amino-cyclopropane-1-carboxylic acid, induce formation and elongation of root hairs. Using confocal microscopy paired with redox biosensors and dyes, we demonstrated that treatments that elevate ethylene levels lead to increased hydrogen peroxide accumulation in hair cells prior to root hair formation. In the ethylene-insensitive receptor mutant, etr1-3, and the signaling double mutant, ein3eil1, the increase in root hair number or reactive oxygen species (ROS) accumulation after ACC and ethylene treatment was lost. Conversely, etr1-7, a constitutive ethylene signaling receptor mutant, has increased root hair formation and ROS accumulation, similar to ethylene-treated Col-0 seedlings. The caprice and werewolf transcription factor mutants have decreased and elevated ROS levels, respectively, which are correlated with levels of root hair initiation. The rhd2-6 mutant, with a defect in the gene encoding the ROS-synthesizing RESPIRATORY BURST OXIDASE HOMOLOG C (RBOHC), and the prx44-2 mutant, which is defective in a class III peroxidase, showed impaired ethylene-dependent ROS synthesis and root hair formation via EIN3EIL1-dependent transcriptional regulation. Together, these results indicate that ethylene increases ROS accumulation through RBOHC and PRX44 to drive root hair formation.

Filling the gaps on root hair development under salt stress and phosphate starvation using current evidence coupled with a meta-analysis approach.

Population expansion is a global issue, especially for food production. Meanwhile, global climate change is damaging our soils, making it difficult for crops to thrive and lowering both production and quality. Poor nutrition and salinity stress affect plant growth and development. Although the impact of individual plant stresses has been studied for decades, the real stress scenario is more complex due to the exposure to multiple stresses at the same time. Here we investigate using existing evidence and a meta-analysis approach to determine molecular linkages between two contemporaneous abiotic stimuli, phosphate (Pi) deficiency and salinity, on a single plant cell model, the root hairs (RHs), which is the first plant cell exposed to them. Understanding how these two stresses work molecularly in RHs may help us build super-adaptable crops and sustainable agriculture in the face of global climate change.

Amino acids biosynthesis in root hair development: a mini-review.

Metabolic factors are essential for developmental biology of an organism. In plants, roots fulfill important functions, in part due to the development of specific epidermal cells, called hair cells that form root hairs (RHs) responsible for water and mineral uptake. RH development consists in (a) patterning processes involved in formation of hair and non-hair cells developed from trichoblasts and atrichoblasts; (b) RH initiation; and (c) apical (tip) growth of the RH. Here we review how these processes depend on pools of different amino acids and what is known about RH phenotypes of mutants disrupted in amino acid biosynthesis. This analysis shows that some amino acids, particularly aromatic ones, are required for RH apical (tip) growth, and that not much is known about the role of amino acids at earlier stages of RH formation. We also address the role of amino acids in rhizosphere, inhibitory and stimulating effects of amino acids on RH growth, amino acids as N source in plant nutrition, and amino acid transporters and their expression in the RHs. Amino acids form conjugates with auxin, a hormone essential for RH growth, and respective genes are overviewed. Finally, we outline missing links and envision some perspectives in the field.

Keep in touch: the soil-root hydraulic continuum and its role in drought resistance in crops.

Drought is a major threat to food security worldwide. Recently, the root-soil interface has emerged as a major site of hydraulic resistance during water stress. Here, we review the impact of soil drying on whole-plant hydraulics and discuss mechanisms by which plants can adapt by modifying the properties of the rhizosphere either directly or through interactions with the soil microbiome.

Nodulating another way: what can we learn from lateral root base nodulation in legumes?

Certain legumes provide a special pathway for rhizobia to invade the root and develop nitrogen-fixing nodules, a process known as lateral root base (LRB) nodulation. This pathway involves intercellular infection at the junction of the lateral roots with the taproot, leading to nodule formation in the lateral root cortex. Remarkably, this LRB pathway serves as a backbone for various adaptative symbiotic processes. Here, we describe different aspects of LRB nodulation and highlight directions for future research to elucidate the mechanisms of this as yet little known but original pathway that will help in broadening our knowledge on the rhizobium-legume symbiosis.

Endoplasmic reticulum membrane receptors of the GET pathway are conserved throughout eukaryotes.

Type II tail-anchored (TA) membrane proteins are involved in diverse cellular processes, including protein translocation, vesicle trafficking, and apoptosis. They are characterized by a single C-terminal transmembrane domain that mediates posttranslational targeting and insertion into the endoplasmic reticulum (ER) via the Guided-Entry of TA proteins (GET) pathway. The GET system was originally described in mammals and yeast but was recently shown to be partially conserved in other eukaryotes, such as higher plants. A newly synthesized TA protein is shielded from the cytosol by a pretargeting complex and an ATPase that delivers the protein to the ER, where membrane receptors (Get1/WRB and Get2/CAML) facilitate insertion. In the model plant Arabidopsis thaliana, most components of the pathway were identified through in silico sequence comparison, however, a functional homolog of the coreceptor Get2/CAML remained elusive. We performed immunoprecipitation-mass spectrometry analysis to detect in vivo interactors of AtGET1 and identified a membrane protein of unknown function with low sequence homology but high structural homology to both yeast Get2 and mammalian CAML. The protein localizes to the ER membrane, coexpresses with AtGET1, and binds to Arabidopsis GET pathway components. While loss-of-function lines phenocopy the stunted root hair phenotype of other Atget lines, its heterologous expression together with the coreceptor AtGET1 rescues growth defects of Δget1get2 yeast. Ectopic expression of the cytosolic, positively charged N terminus is sufficient to block TA protein insertion in vitro. Our results collectively confirm that we have identified a plant-specific GET2 in Arabidopsis, and its sequence allows the analysis of cross-kingdom pathway conservation.

SUPPRESSOR of MAX2 1 (SMAX1) and SMAX1-LIKE2 (SMXL2) Negatively Regulate Drought Resistance in Arabidopsis thaliana.

Recent investigations in Arabidopsis thaliana suggest that SUPPRESSOR of MORE AXILLARY GROWTH 2 1 (SMAX1) and SMAX1-LIKE2 (SMXL2) are negative regulators of karrikin (KAR) and strigolactone (SL) signaling during plant growth and development, but their functions in drought resistance and related mechanisms of action remain unclear. To understand the roles and mechanisms of SMAX1 and SMXL2 in drought resistance, we investigated the drought-resistance phenotypes and transcriptome profiles of smax1 smxl2 (s1,2) double-mutant plants in response to drought stress. The s1,2 mutant plants showed enhanced drought-resistance and lower leaf water loss when compared with wild-type (WT) plants. Transcriptome comparison of rosette leaves from the s1,2 mutant and the WT under normal and dehydration conditions suggested that the mechanism related to cuticle formation was involved in drought resistance. This possibility was supported by enhanced cuticle formation in the rosette leaves of the s1,2 mutant. We also found that the s1,2 mutant plants were more sensitive to abscisic acid in assays of stomatal closure, cotyledon opening, chlorophyll degradation and growth inhibition, and they showed a higher reactive oxygen species detoxification capacity than WT plants. In addition, the s1,2 mutant plants had longer root hairs and a higher root-to-shoot ratio than the WT plants, suggesting that the mutant had a greater capacity for water absorption than the WT. Taken together, our results indicate that SMAX1 and SMXL2 negatively regulate drought resistance, and disruption of these KAR- and SL-signaling-related genes may therefore provide a novel means for improving crop drought resistance.

Genome wide association analysis of root hair traits in rice reveals novel genomic regions controlling epidermal cell differentiation.

BACKGROUND: Genome wide association (GWA) studies demonstrate linkages between genetic variants and traits of interest. Here, we tested associations between single nucleotide polymorphisms (SNPs) in rice (Oryza sativa) and two root hair traits, root hair length (RHL) and root hair density (RHD). Root hairs are outgrowths of single cells on the root epidermis that aid in nutrient and water acquisition and have also served as a model system to study cell differentiation and tip growth. Using lines from the Rice Diversity Panel-1, we explored the diversity of root hair length and density across four subpopulations of rice (aus, indica, temperate japonica, and tropical japonica). GWA analysis was completed using the high-density rice array (HDRA) and the rice reference panel (RICE-RP) SNP sets. RESULTS: We identified 18 genomic regions related to root hair traits, 14 of which related to RHD and four to RHL. No genomic regions were significantly associated with both traits. Two regions overlapped with previously identified quantitative trait loci (QTL) associated with root hair density in rice. We identified candidate genes in these regions and present those with previously published expression data relevant to root hair development. We re-phenotyped a subset of lines with extreme RHD phenotypes and found that the variation in RHD was due to differences in cell differentiation, not cell size, indicating genes in an associated genomic region may influence root hair cell fate. The candidate genes that we identified showed little overlap with previously characterized genes in rice and Arabidopsis. CONCLUSIONS: Root hair length and density are quantitative traits with complex and independent genetic control in rice. The genomic regions described here could be used as the basis for QTL development and further analysis of the genetic control of root hair length and density. We present a list of candidate genes involved in root hair formation and growth in rice, many of which have not been previously identified as having a relation to root hair growth. Since little is known about root hair growth in grasses, these provide a guide for further research and crop improvement.

Cell surface receptor kinase FERONIA linked to nutrient sensor TORC signaling controls root hair growth at low temperature linked to low nitrate in Arabidopsis thaliana.

Root hairs (RH) are excellent model systems for studying cell size and polarity since they elongate several hundred-fold their original size. Their tip growth is determined both by intrinsic and environmental signals. Although nutrient availability and temperature are key factors for a sustained plant growth, the molecular mechanisms underlying their sensing and downstream signaling pathways remain unclear. We use genetics to address the roles of the cell surface receptor kinase FERONIA (FER) and the nutrient sensing TOR Complex 1 (TORC) in RH growth. We identified that low temperature (10°C) triggers a strong RH elongation response in Arabidopsis thaliana involving FER and TORC. We found that FER is required to perceive limited nutrient availability caused by low temperature. FERONIA interacts with and activates TORC-downstream components to trigger RH growth. In addition, the small GTPase Rho of plants 2 (ROP2) is also involved in this RH growth response linking FER and TOR. We also found that limited nitrogen nutrient availability can mimic the RH growth response at 10°C in a NRT1.1-dependent manner. These results uncover a molecular mechanism by which a central hub composed by FER-ROP2-TORC is involved in the control of RH elongation under low temperature and nitrogen deficiency.

The spatial distribution of rhizosphere microbial activities under drought: water availability is more important than root-hair-controlled exudation.

Root hairs and soil water content are crucial in controlling the release and diffusion of root exudates and shaping profiles of biochemical properties in the rhizosphere. But whether root hairs can offset the negative impacts of drought on microbial activity remains unknown. Soil zymography, 14 C imaging and neutron radiography were combined to identify how root hairs and soil moisture affect rhizosphere biochemical properties. To achieve this, we cultivated two maize genotypes (wild-type and root-hair-defective rth3 mutant) under ambient and drought conditions. Root hairs and optimal soil moisture increased hotspot area, rhizosphere extent and kinetic parameters (Vmax and Km ) of ß-glucosidase activities. Drought enlarged the rhizosphere extent of root exudates and water content. Colocalization analysis showed that enzymatic hotspots were more colocalized with root exudate hotspots under optimal moisture, whereas they showed higher dependency on water hotspots when soil water and carbon were scarce. We conclude that root hairs are essential in adapting rhizosphere properties under drought to maintain plant nutrition when a continuous mass flow of water transporting nutrients to the root is interrupted. In the rhizosphere, soil water was more important than root exudates for hydrolytic enzyme activities under water and carbon colimitation.

Root cap is an important determinant of rhizosphere microbiome assembly.

Plants impact the development of their rhizosphere microbial communities. It is yet unclear to what extent the root cap and specific root zones contribute to microbial community assembly. To test the roles of root caps and root hairs in the establishment of microbiomes along maize roots (Zea mays), we compared the composition of prokaryote (archaea and bacteria) and protist (Cercozoa and Endomyxa) microbiomes of intact or decapped primary roots of maize inbred line B73 with its isogenic root hairless (rth3) mutant. In addition, we tracked gene expression along the root axis to identify molecular control points for an active microbiome assembly by roots. Absence of root caps had stronger effects on microbiome composition than the absence of root hairs and affected microbial community composition also at older root zones and at higher trophic levels (protists). Specific bacterial and cercozoan taxa correlated with root genes involved in immune response. Our results indicate a central role of root caps in microbiome assembly with ripple-on effects affecting higher trophic levels and microbiome composition on older root zones.

The effect of root hairs on root water uptake is determined by root-soil contact and root hair shrinkage.

The effect of root hairs on water uptake remains controversial. In particular, the key root hair and soil parameters that determine their importance have been elusive. We grew maize plants (Zea mays) in microcosms and scanned them using synchrotron-based X-ray computed microtomography. By means of image-based modelling, we investigated the parameters determining the effectiveness of root hairs in root water uptake. We explicitly accounted for rhizosphere features (e.g. root-soil contact and pore structure) and took root hair shrinkage of dehydrated root hairs into consideration. Our model suggests that > 85% of the variance in root water uptake is explained by the hair-induced increase in root-soil contact. In dry soil conditions, root hair shrinkage reduces the impact of hairs substantially. We conclude that the effectiveness of root hairs on root water uptake is determined by the hair-induced increase in root-soil contact and root hair shrinkage. Although the latter clearly reduces the effect of hairs on water uptake, our model still indicated facilitation of water uptake by root hairs at soil matric potentials from -1 to -0.1 MPa. Our findings provide new avenues towards a mechanistic understanding of the role of root hairs on water uptake.

Advanced microscopy resolves dynamic localization patterns of stress-induced mitogen-activated protein kinase (SIMK) during alfalfa root hair interactions with Ensifer meliloti.

Leguminous plants have established mutualistic endosymbiotic interactions with nitrogen-fixing rhizobia to secure nitrogen sources in root nodules. Before nodule formation, the development of early symbiotic structures is essential for rhizobia docking, internalization, targeted delivery, and intracellular accommodation. We recently reported that overexpression of stress-induced mitogen-activated protein kinase (SIMK) in alfalfa affects root hair, nodule, and shoot formation, raising the question of how SIMK modulates these processes. In particular, detailed subcellular spatial distribution, activation, and developmental relocation of SIMK during early stages of alfalfa nodulation remain unclear. Here, we characterized SIMK distribution in Ensifer meliloti-infected root hairs using live-cell imaging and immunolocalization, employing alfalfa stable transgenic lines with genetically manipulated SIMK abundance and kinase activity. In the SIMKK-RNAi line, showing down-regulation of SIMKK and SIMK, we found considerably decreased accumulation of phosphorylated SIMK around infection pockets and infection threads. However, this was strongly increased in the GFP-SIMK line, constitutively overexpressing green fluorescent protein (GFP)-tagged SIMK. Thus, genetically manipulated SIMK modulates root hair capacity to form infection pockets and infection threads. Advanced light-sheet fluorescence microscopy on intact plants allowed non-invasive imaging of spatiotemporal interactions between root hairs and symbiotic E. meliloti, while immunofluorescence detection confirmed that SIMK was activated in these locations. Our results shed new light on SIMK spatiotemporal participation in early interactions between alfalfa and E. meliloti, and its internalization into root hairs, showing that local accumulation of active SIMK modulates early nodulation in alfalfa.

New molecular components that regulates the transcriptional hub in root hairs: coupling environmental signals to endogenous hormones to coordinate growth.

Root hairs (RH) have become an important model system for studying plant growth and how plants modulate their growth in response to cell-intrinsic and environmental stimuli. Here, we will discuss recent advances in our understanding of the molecular mechanisms underlying the growth of Arabidopsis thaliana RH in the interface between responses to environmental cues (e.g. nutrients such as nitrates, phosphate and microorganism) and hormonal stimuli (e.g. auxin). RH growth is under the control of several transcription factors that are also under strong regulation at different levels. In this review we highlight recent new discoveries along these transcriptional pathways that may increase our capacity to enhance nutrient uptake by the roots in the context of abiotic stresses. We used text-mining capacities of the PlantConnectome database to generate the most updated view of RH growth in these complex biological contexts.

The karrikin signaling regulator SMAX1 controls Lotus japonicus root and root hair development by suppressing ethylene biosynthesis.

An evolutionarily ancient plant hormone receptor complex comprising the α/ß-fold hydrolase receptor KARRIKIN INSENSITIVE 2 (KAI2) and the F-box protein MORE AXILLARY GROWTH 2 (MAX2) mediates a range of developmental responses to smoke-derived butenolides called karrikins (KARs) and to yet elusive endogenous KAI2 ligands (KLs). Degradation of SUPPRESSOR OF MAX2 1 (SMAX1) after ligand perception is considered to be a key step in KAR/KL signaling. However, molecular events which regulate plant development downstream of SMAX1 removal have not been identified. Here we show that Lotus japonicus SMAX1 is specifically degraded in the presence of KAI2 and MAX2 and plays an important role in regulating root and root hair development. smax1 mutants display very short primary roots and elongated root hairs. Their root transcriptome reveals elevated ethylene responses and expression of ACC Synthase 7 (ACS7), which encodes a rate-limiting enzyme in ethylene biosynthesis. smax1 mutants release increased amounts of ethylene and their root phenotype is rescued by treatment with ethylene biosynthesis and signaling inhibitors. KAR treatment induces ACS7 expression in a KAI2-dependent manner and root developmental responses to KAR treatment depend on ethylene signaling. Furthermore, in Arabidopsis, KAR-induced root hair elongation depends on ACS7 Thus, we reveal a connection between KAR/KL and ethylene signaling in which the KAR/KL signaling module (KAI2-MAX2-SMAX1) regulates the biosynthesis of ethylene to fine-tune root and root hair development, which are important for seedling establishment at the beginning of the plant life cycle.

A Nonspecific Lipid Transfer Protein with Potential Functions in Infection and Nodulation.

The response of Alnus glutinosa to Frankia alni ACN14a is driven by several sequential physiological events from calcium spiking and root-hair deformation to the development of the nodule. Early stages of actinorhizal symbiosis were monitored at the transcriptional level to observe plant host responses to Frankia alni. Forty-two genes were significantly upregulated in inoculated compared with noninoculated roots. Most of these genes encode proteins involved in biological processes induced during microbial infection, such as oxidative stress or response to stimuli, but a large number of them are not differentially modulated or downregulated later in the process of nodulation. In contrast, several of them remained upregulated in mature nodules, and this included the gene most upregulated, which encodes a nonspecific lipid transfer protein (nsLTP). Classified as an antimicrobial peptide, this nsLTP was immunolocalized on the deformed root-hair surfaces that are points of contact for Frankia spp. during infection. Later in nodules, it binds to the surface of F. alni ACN14a vesicles, which are the specialized cells for nitrogen fixation. This nsLTP, named AgLTP24, was biologically produced in a heterologous host and purified for assay on F. alni ACN14a to identify physiological effects. Thus, the activation of the plant immunity response occurs upon first contact, while the recognition of F. alni ACN14a genes switches off part of the defense system during nodulation. AgLTP24 constitutes a part of the defense system that is maintained all along the symbiosis, with potential functions such as the formation of infection threads or nodule primordia to the control of F. alni proliferation. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Trihelix transcription factors GTL1 and DF1 prevent aberrant root hair formation in an excess nutrient condition.

Root hair growth is tuned in response to the environment surrounding plants. While most previous studies focused on the enhancement of root hair growth during nutrient starvation, few studies investigated the root hair response in the presence of excess nutrients. We report that the post-embryonic growth of wild-type Arabidopsis plants is strongly suppressed with increasing nutrient availability, particularly in the case of root hair growth. We further used gene expression profiling to analyze how excess nutrient availability affects root hair growth, and found that RHD6 subfamily genes, which are positive regulators of root hair growth, are downregulated in this condition. However, defects in GTL1 and DF1, which are negative regulators of root hair growth, cause frail and swollen root hairs to form when excess nutrients are supplied. Additionally, we observed that the RHD6 subfamily genes are mis-expressed in gtl1-1 df1-1. Furthermore, overexpression of RSL4, an RHD6 subfamily gene, induces swollen root hairs in the face of a nutrient overload, while mutation of RSL4 in gtl1-1 df1-1 restore root hair swelling phenotype. In conclusion, our data suggest that GTL1 and DF1 prevent unnecessary root hair formation by repressing RSL4 under excess nutrient conditions.

Root hydraulic phenotypes impacting water uptake in drying soils.

Soil drying is a limiting factor for crop production worldwide. Yet, it is not clear how soil drying impacts water uptake across different soils, species, and root phenotypes. Here we ask (1) what root phenotypes improve the water use from drying soils? and (2) what root hydraulic properties impact water flow across the soil-plant continuum? The main objective is to propose a hydraulic framework to investigate the interplay between soil and root hydraulic properties on water uptake. We collected highly resolved data on transpiration, leaf and soil water potential across 11 crops and 10 contrasting soil textures. In drying soils, the drop in water potential at the soil-root interface resulted in a rapid decrease in soil hydraulic conductance, especially at higher transpiration rates. The analysis reveals that water uptake was limited by soil within a wide range of soil water potential (-6 to -1000 kPa), depending on both soil textures and root hydraulic phenotypes. We propose that a root phenotype with low root hydraulic conductance, long roots and/or long and dense root hairs postpones soil limitation in drying soils. The consequence of these root phenotypes on crop water use is discussed.

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.

Nuclear movement in growing Arabidopsis root hairs involves both actin filaments and microtubules.

Nuclear migration during growth and development is a conserved phenomenon among many eukaryotic species. In Arabidopsis, movement of the nucleus is important for root hair growth, but the detailed mechanism behind this movement is not well known. Previous studies in different cell types have reported that the myosin XI-I motor protein is responsible for this nuclear movement by attaching to the nuclear transmembrane protein complex WIT1/WIT2. Here, we analyzed nuclear movement in growing root hairs of wild-type, myosin xi-i, and wit1 wit2 Arabidopsis lines in the presence of actin and microtubule-disrupting inhibitors to determine the individual effects of actin filaments and microtubules on nuclear movement. We discovered that forward nuclear movement during root hair growth can occur in the absence of myosin XI-I, suggesting the presence of an alternative actin-based mechanism that mediates rapid nuclear displacements. By quantifying nuclear movements with high temporal resolution during the initial phase of inhibitor treatment, we determined that microtubules work to dampen erratic nuclear movements during root hair growth. We also observed microtubule-dependent backwards nuclear movement when actin filaments were impaired in the absence of myosin XI-I, indicating the presence of complex interactions between the cytoskeletal arrays during nuclear movements in growing root hairs.