Exploring the puzzle of Reactive Oxygen Species (ROS) acting on root hair cells.

Reactive oxygen species (ROS) are essential signaling molecules that enable cells to respond rapidly to a range of stimuli. The capacity of plants to recognize various stressors, incorporate a variety of environmental inputs, and initiate stress-response networks depends on ROS. Plants develop resilience and defensive systems as a result of these processes. Root hairs (RHs) are central components of the root biology since they increase the surface area of the root, anchor it in the soil, increase its ability to absorb water and nutrients, and foster interactions between microorganisms. In this review, we specifically focused on RHs cells and we highlighted the identification of ROS receptors, important new regulatory hubs that connect ROS production, transport, and signaling in the context of two hormonal pathways (auxin and ethylene) and under low temperature environmental input related to nutrients. As ROS plays a crucial role in regulating cell elongation rates, RHs are rapidly gaining traction as a very valuable single plant cell model for investigating ROS homeostasis and signaling. These promising findings might soon aid in the development of plants and roots that are more resilient to environmental stressors.

The exception to the rule? TORC1 triggers growth under low nutrient environments.

Eukaryotic cells' proliferation and growth are controlled by the target of rapamycin kinase (TOR). TOR usually activates in favorable energy and nutritional circumstances. This is challenged by recent research, suggesting that plant cells optimized for nutrient absorption in low nutritional conditions may activate the TOR pathway in a polarized manner.

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.

Transcription factor NAC1 activates expression of peptidase-encoding AtCEPs in roots to limit root hair growth.

Plant genomes encode a unique group of papain-type Cysteine EndoPeptidases (CysEPs) containing a KDEL endoplasmic reticulum (ER) retention signal (KDEL-CysEPs or CEPs). CEPs process the cell-wall scaffolding EXTENSIN (EXT) proteins that regulate de novo cell-wall formation and cell expansion. Since CEPs cleave EXTs and EXT-related proteins, acting as cell-wall-weakening agents, they may play a role in cell elongation. The Arabidopsis (Arabidopsis thaliana) genome encodes 3 CEPs (AtCPE1-AtCEP3). Here, we report that the genes encoding these 3 Arabidopsis CEPs are highly expressed in root-hair (RH) cell files. Single mutants have no evident abnormal RH phenotype, but atcep1-3 atcep3-2 and atcep1-3 atcep2-2 double mutants have longer RHs than wild-type (Wt) plants, suggesting that expression of AtCEPs in root trichoblasts restrains polar elongation of the RH. We provide evidence that the transcription factor NAC1 (petunia NAM and Arabidopsis ATAF1, ATAF2, and CUC2) activates AtCEPs expression in roots to limit RH growth. Chromatin immunoprecipitation indicates that NAC1 binds to the promoter of AtCEP1, AtCEP2, and, to a lower extent, AtCEP3 and may directly regulate their expression. Inducible NAC1 overexpression increases AtCEP1 and AtCEP2 transcript levels in roots and leads to reduced RH growth while the loss of function nac1-2 mutation reduces AtCEP1-AtCEP3 gene expression and enhances RH growth. Likewise, expression of a dominant chimeric NAC1-SRDX repressor construct leads to increased RH length. Finally, we show that RH cell walls in the atcep1-3 atcep3-2 double mutant have reduced levels of EXT deposition, suggesting that the defects in RH elongation are linked to alterations in EXT processing and accumulation. Our results support the involvement of AtCEPs in controlling RH polar growth through EXT processing and insolubilization at the cell wall.

A spatially concerted epidermal auxin signaling framework steers the root hair foraging response under low nitrogen.

As a major determinant of the nutrient-acquiring root surface, root hairs (RHs) provide a low-input strategy to enhance nutrient uptake. Although primary and lateral roots exhibit elongation responses under mild nitrogen (N) deficiency, the foraging response of RHs and underlying regulatory mechanisms remain elusive. Employing transcriptomics and functional studies revealed a framework of molecular components composing a cascade of auxin synthesis, transport, and signaling that triggers RH elongation for N acquisition. Through upregulation of Tryptophan Aminotransferase of Arabidopsis 1 (TAA1) and YUCCA8, low N increases auxin accumulation in the root apex. Auxin is then directed to the RH differentiation zone via the auxin transport machinery, AUXIN TRANSPORTER PROTEIN 1 (AUX1) and PIN-FORMED 2 (PIN2). Upon arrival to the RH zone, auxin activates the transcription factors AUXIN RESPONSE FACTOR 6 and 8 (ARF6/8) to promote the epidermal and auxin-inducible transcriptional module ROOT HAIR DEFECTIVE 6 (RHD6)-LOTUS JAPONICA ROOT HAIRLESS-LIKE 3 (LRL3) to steer RH elongation in response to low N. Our study uncovers a spatially defined regulatory signaling cascade for N foraging by RHs, expanding the mechanistic framework of hormone-regulated nutrient sensing in plant roots.

Understanding signaling pathways governing the polar development of root hairs in low-temperature, nutrient-deficient environments.

Plants exposed to freezing and above-freezing low temperatures must employ a variety of strategies to minimize fitness loss. There is a considerable knowledge gap regarding how mild low temperatures (around 10 °C) affect plant growth and developmental processes, even though the majority of the molecular mechanisms that plants use to adapt to extremely low temperatures are well understood. Root hairs (RH) have become a useful model system for studying how plants regulate their growth in response to both cell-intrinsic cues and environmental inputs. Here, we'll focus on recent advances in the molecular mechanisms underpinning Arabidopsis thaliana RH growth at mild low temperatures and how these discoveries may influence our understanding of nutrient sensing mechanisms by the roots. This highlights how intricately linked mechanisms are necessary for plant development to take place under specific circumstances and to produce a coherent response, even at the level of a single RH cell.

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.

Morphometric and molecular characterization of Kudoa encrasicoli n. sp. (Myxozoa: Myxosporea) from the European anchovy, Engraulis encrasicolus (L.) (Clupeiformes: Engraulidae).

The European anchovy represents the main fisheries for countries in the Mediterranean and Black Sea basins. The skeletal muscle of 13 of 48 (27.1%) Engraulis encrasicolus (L.) specimens from North East Atlantic waters (FAO 27.8.c) was found infected with interfibrillar elongated plasmodia (130-980 µm in length) containing mature myxospores belonging to the genus Kudoa Meglitsch, 1947. No flesh softening was found associated with infection. Fresh myxospores were 10.8 ± 0.7 (9.1-12.3) µm in width 1, 11.3 ± 0.9 (9.5-13.4) µm in width 2, 6.7 ± 0.4 (5.8-7.4) µm in thickness, and 6.9 ± 0.5 (5.8-7.5) µm in length. They were almost stellate in apical view having three pointed-edged shell valves bearing three small polar capsules equal in size 5.0 ± 0.3 (4.4-5.4) µm long and 2.4 ± 0.2 (2.0-3.0) µm wide, and one rounded- to rarely bluntly pointed-edged shell valve bearing a large and particularly wide polar capsule 6.8 ± 0.4 (5.9-7.6) µm long and 4.1 ± 0.2 (3.6-4.4) µm wide. Morphological and morphometrical comparisons between these myxospores and those of Kudoa thyrsites (Gilchrist, 1923) from the clupeid Sardina pilchardus (Walbaum) (North East Atlantic waters, FAO 27.9.a), with which exhibited a similarity of 98.9% and 96.2% using SSU and LSU rDNA sequences, respectively, support the creation of Kudoa encrasicoli n. sp. Morphometrical analysis of the polar capsules of flattened myxospores is suggested as a useful approach to differentiate phylogenetically related kudoids with stellate or almost stellate myxospores bearing four polar capsules.

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.

Class III Peroxidases PRX01, PRX44, and PRX73 Control Root Hair Growth in Arabidopsis thaliana.

Root hair cells are important sensors of soil conditions. They grow towards and absorb water-soluble nutrients. This fast and oscillatory growth is mediated by continuous remodeling of the cell wall. Root hair cell walls contain polysaccharides and hydroxyproline-rich glycoproteins, including extensins (EXTs). Class-III peroxidases (PRXs) are secreted into the apoplastic space and are thought to trigger either cell wall loosening or polymerization of cell wall components, such as Tyr-mediated assembly of EXT networks (EXT-PRXs). The precise role of these EXT-PRXs is unknown. Using genetic, biochemical, and modeling approaches, we identified and characterized three root-hair-specific putative EXT-PRXs, PRX01, PRX44, and PRX73. prx01,44,73 triple mutation and PRX44 and PRX73 overexpression had opposite effects on root hair growth, peroxidase activity, and ROS production, with a clear impact on cell wall thickness. We use an EXT fluorescent reporter with contrasting levels of cell wall insolubilization in prx01,44,73 and PRX44-overexpressing background plants. In this study, we propose that PRX01, PRX44, and PRX73 control EXT-mediated cell wall properties during polar expansion of root hair cells.

Apoplastic class III peroxidases PRX62 and PRX69 promote Arabidopsis root hair growth at low temperature.

Root Hairs (RHs) growth is influenced by endogenous and by external environmental signals that coordinately regulate its final cell size. We have recently determined that RH growth was unexpectedly boosted when Arabidopsis thaliana seedlings are cultivated at low temperatures. It was proposed that RH growth plasticity in response to low temperature was linked to a reduced nutrient availability in the media. Here, we explore the molecular basis of this RH growth response by using a Genome Wide Association Study (GWAS) approach using Arabidopsis thaliana natural accessions. We identify the poorly characterized PEROXIDASE 62 (PRX62) and a related protein PRX69 as key proteins under moderate low temperature stress. Strikingly, a cell wall protein extensin (EXT) reporter reveals the effect of peroxidase activity on EXT cell wall association at 10 °C in the RH apical zone. Collectively, our results indicate that PRX62, and to a lesser extent PRX69, are key apoplastic PRXs that modulate ROS-homeostasis and cell wall EXT-insolubilization linked to RH elongation at low temperature.

Proline-rich extensin-like receptor kinases PERK5 and PERK12 are involved in pollen tube growth.

Proline-rich extensin-like receptor kinases (PERKs) belong to the hydroxyproline-rich glycoprotein (HRGP) superfamily known to be involved in many plant developmental processes. Here, we characterized two pollen-expressed PERKs from Arabidopsis thaliana, PERK5 and PERK12. Pollen tube growth was impaired in single and double perk5-1 perk12-1 loss of function mutants, with an impact on seed production. When the segregation was analysed, a male gametophytic defect was found, indicating that perk5-1 and perk12-1 mutants carry deficient pollen transmission. Furthermore, perk5-1 perk12-1 displayed an excessive accumulation of pectins and cellulose at the cell wall of the pollen tubes. Our results indicate that PERK5 and PERK12 are necessary for proper pollen tube growth, highlighting their role in cell wall assembly and reactive oxygen species homeostasis.

Salt stress on Lotus tenuis triggers cell wall polysaccharide changes affecting their digestibility by ruminants.

Lotus tenuis is a glycophytic forage legume (Fabaceae) used in feeding ruminants that can grow under salinity and waterlogging stresses. Plants obtained in controlled conditions were affected negatively in their growth by the effect of salt. Results from sequential extraction of plant cell wall polysaccharides and chemical characterization were related to those from nutritional parameters used to assess ruminants feedstuffs (Van Soest detergent system). Shoots and leaves were analyzed, and the most important differences were found for shoots. The salt-stressed shoots gave lower values of neutral detergent fiber and acid detergent fiber; they produced higher amounts of reserve α-glucans, and hemicelluloses (xyloglucans and glucuronoxylans from primary and secondary cell walls, respectively) and pectins, leaving less material resistant to extraction. This effect was clearly confirmed by an in vitro gas production assay. In addition, observations by light microcopy (LM) and transmission electron microscopy (TEM), showed in some tissues thicker walls and more opened cell wall structures in regard to control samples, which could allow easier access of degrading enzymes in the rumen. Although the plant biomass of Lotus tenuis produced under salt stress was lower, its quality as forage improved due to production of increased quantities of more digestible polysaccharides.

The lncRNA APOLO and the transcription factor WRKY42 target common cell wall EXTENSIN encoding genes to trigger root hair cell elongation.

Plant long noncoding RNAs (lncRNAs) are key chromatin dynamics regulators, directing the transcriptional programs driving a wide variety of developmental outputs. Recently, we uncovered how the lncRNA AUXIN REGULATED PROMOTER LOOP (APOLO) directly recognizes the locus encoding the root hair (RH) master regulator ROOT HAIR DEFECTIVE 6 (RHD6) modulating its transcriptional activation and leading to low temperature-induced RH elongation. We further demonstrated that APOLO interacts with the transcription factor WRKY42 in a novel ribonucleoprotein complex shaping RHD6 epigenetic environment and integrating signals governing RH growth and development. In this work, we expand this model showing that APOLO is able to bind and positively control the expression of several cell wall EXTENSIN (EXT) encoding genes, including EXT3, a key regulator for RH growth. Interestingly, EXT3 emerged as a novel common target of APOLO and WRKY42. Furthermore, we showed that the ROS homeostasis-related gene NADPH OXIDASE C (NOXC) is deregulated upon APOLO overexpression, likely through the RHD6-RSL4 pathway, and that NOXC is required for low temperature-dependent enhancement of RH growth. Collectively, our results uncover an intricate regulatory network involving the APOLO/WRKY42 hub in the control of master and effector genes during RH development.

The lncRNA APOLO interacts with the transcription factor WRKY42 to trigger root hair cell expansion in response to cold.

Plant long noncoding RNAs (lncRNAs) have emerged as important regulators of chromatin dynamics, impacting on transcriptional programs leading to different developmental outputs. The lncRNA AUXIN-REGULATED PROMOTER LOOP (APOLO) directly recognizes multiple independent loci across the Arabidopsis genome and modulates their three-dimensional chromatin conformation, leading to transcriptional shifts. Here, we show that APOLO recognizes the locus encoding the root hair (RH) master regulator ROOT HAIR DEFECTIVE 6 (RHD6) and controls RHD6 transcriptional activity, leading to cold-enhanced RH elongation through the consequent activation of the transcription factor gene RHD6-like RSL4. Furthermore, we demonstrate that APOLO interacts with the transcription factor WRKY42 and modulates its binding to the RHD6 promoter. WRKY42 is required for the activation of RHD6 by low temperatures and WRKY42 deregulation impairs cold-induced RH expansion. Collectively, our results indicate that a novel ribonucleoprotein complex with APOLO and WRKY42 forms a regulatory hub to activate RHD6 by shaping its epigenetic environment and integrate signals governing RH growth and development.

Cracking the "Sugar Code": A Snapshot of N- and O-Glycosylation Pathways and Functions in Plants Cells.

Glycosylation is a fundamental co-translational and/or post-translational modification process where an attachment of sugars onto either proteins or lipids can alter their biological function, subcellular location and modulate the development and physiology of an organism. Glycosylation is not a template driven process and as such produces a vastly larger array of glycan structures through combinatorial use of enzymes and of repeated common scaffolds and as a consequence it provides a huge expansion of both the proteome and lipidome. While the essential role of N- and O-glycan modifications on mammalian glycoproteins is already well documented, we are just starting to decode their biological functions in plants. Although significant advances have been made in plant glycobiology in the last decades, there are still key challenges impeding progress in the field and, as such, holistic modern high throughput approaches may help to address these conceptual gaps. In this snapshot, we present an update of the most common O- and N-glycan structures present on plant glycoproteins as well as (1) the plant glycosyltransferases (GTs) and glycosyl hydrolases (GHs) responsible for their biosynthesis; (2) a summary of microorganism-derived GHs characterized to cleave specific glycosidic linkages; (3) a summary of the available tools ranging from monoclonal antibodies (mAbs), lectins to chemical probes for the detection of specific sugar moieties within these complex macromolecules; (4) selected examples of N- and O-glycoproteins as well as in their related GTs to illustrate the complexity on their mode of action in plant cell growth and stress responses processes, and finally (5) we present the carbohydrate microarray approach that could revolutionize the way in which unknown plant GTs and GHs are identified and their specificities characterized.