Formation Of Apoplastic Barriers Research Articles

Early differentiation of Casparian strip mediated by nitric oxide is required for efficient K transport under low K conditions in Arabidopsis.

The Casparian strip (CS) is a cell wall modification made of lignin that functions as an apoplastic barrier in the root endodermis to restrict nutrient and water transport between the soil and stele. CS formation is affected by nutritional conditions and its physiological roles have been discussed. In this study, we found that low K condition affects CS permeability, lignin deposition and MYB36 mRNA accumulation. To understand the mechanism underling these findings, we focused on nitric oxide (NO). NO is known to act as a signaling molecule and participates in cell wall synthesis, especially for lignin composition. However, the mechanism by which NO affects lignin deposition and corrects CS formation in the plant roots remains unclear. Through the combining fluorescent observation with histological stains, we demonstrated that the root endodermal cell lignification response to low-potassium (K) conditions is mediated by NO through the MYB36-associated lignin-polymerizing pathway. Furthermore, we discovered the noteworthy ability of NO to maintain nutrient homeostasis for adaptation to low-K conditions by affecting the correct apoplastic barrier formation of CS. Collectively, our results suggest that NO is required for the lignification and apoplastic barrier formation in the root endodermis during adaptation to low-K conditions, which revealing the novel physiological roles of CS under low nutrient conditions and making a significant contribution to CS biology.

Apoplastic barriers of Populus × canescens roots in reaction to different cultivation conditions and abiotic stress treatments

Populus is an important tree genus frequently cultivated for economical purposes. However, the high sensitivity of poplars towards water deficit, drought, and salt accumulation significantly affects plant productivity and limits biomass yield. Various cultivation and abiotic stress conditions have been described to significantly induce the formation of apoplastic barriers (Casparian bands and suberin lamellae) in roots of different monocotyledonous crop species. Thus, this study aimed to investigate to which degree the roots of the dicotyledonous gray poplar (Populus × canescens) react to a set of selected cultivation conditions (hydroponics, aeroponics, or soil) and abiotic stress treatments (abscisic acid, oxygen deficiency) because a differing stress response could potentially help in explaining the observed higher stress susceptibility. The apoplastic barriers of poplar roots cultivated in different environments were analyzed by means of histochemistry and gas chromatography and compared to the available literature on monocotyledonous crop species. Overall, dicotyledonous poplar roots showed only a remarkably low induction or enhancement of apoplastic barriers in response to the different cultivation conditions and abiotic stress treatments. The genetic optimization (e.g., overexpression of biosynthesis key genes) of the apoplastic barrier development in poplar roots might result in more stress-tolerant cultivars in the future.

Effect of ipt Gene Induction in Transgenic Tobacco Plants on Hydraulic Conductance, Formation of Apoplastic Barriers and Aquaporin Activity under Heat Shock.

Cytokinins are known to keep stomata open, which supports gas exchange and correlates with increased photosynthesis. However, keeping the stomata open can be detrimental if the increased transpiration is not compensated for by water supply to the shoots. In this study, we traced the effect of ipt (isopentenyl transferase) gene induction, which increases the concentration of cytokinins in transgenic tobacco plants, on transpiration and hydraulic conductivity. Since water flow depends on the conductivity of the apoplast, the deposition of lignin and suberin in the apoplast was studied by staining with berberine. The effect of an increased concentration of cytokinins on the flow of water through aquaporins (AQPs) was revealed by inhibition of AQPs with HgCl2. It was shown that an elevated concentration of cytokinins in ipt-transgenic plants increases hydraulic conductivity by enhancing the activity of aquaporins and reducing the formation of apoplastic barriers. The simultaneous effect of cytokinins on both stomatal and hydraulic conductivity makes it possible to coordinate the evaporation of water from leaves and its flow from roots to leaves, thereby maintaining the water balance and leaf hydration.

Influence of Plant Growth-Promoting Rhizobacteria on the Formation of Apoplastic Barriers and Uptake of Water and Potassium by Wheat Plants.

The formation of apoplastic barriers is important for controlling the uptake of water and ions by plants, thereby influencing plant growth. However, the effects of plant growth-promoting bacteria on the formation of apoplastic barriers, and the relationship between these effects and the ability of bacteria to influence the content of hormones in plants, have not been sufficiently studied. The content of cytokinins, auxins and potassium, characteristics of water relations, deposition of lignin and suberin and the formation of Casparian bands in the root endodermis of durum wheat (Triticum durum Desf.) plants were evaluated after the introduction of the cytokinin-producing bacterium Bacillus subtilis IB-22 or the auxin-producing bacterium Pseudomonas mandelii IB-Ki14 into their rhizosphere. The experiments were carried out in laboratory conditions in pots with agrochernozem at an optimal level of illumination and watering. Both strains increased shoot biomass, leaf area and chlorophyll content in leaves. Bacteria enhanced the formation of apoplastic barriers, which were most pronounced when plants were treated with P. mandelii IB-Ki14. At the same time, P. mandelii IB-Ki14 caused no decrease in the hydraulic conductivity, while inoculation with B. subtilis IB-22, increased hydraulic conductivity. Cell wall lignification reduced the potassium content in the roots, but did not affect its content in the shoots of plants inoculated with P. mandelii IB-Ki14. Inoculation with B. subtilis IB-22 did not change the potassium content in the roots, but increased it in the shoots.

ФОРМИРОВАНИЕ ПОЯСКОВ КАСПАРИ В КОРНЯХ ПОЧТИ ИЗОГЕННЫХ ЛИНИЙ ЯЧМЕНЯ, ОТЛИЧАЮЩИХСЯ НАЛИЧИЕМ МЕЛАНИНОВОЙ ОКРАСКИ КОЛОСА

The influence of the BLP locus genes, which control the formation of black coloration of the spike glumes and grain pericarp on the formation of Casparian bands in Hordeum vulgare L. plants, was studied. The experiment was carried out with plants of the Bowman variety and three almost isogenic lines obtained on its basis: the BLP line, carrying one recombinant section from the donor of the trait of black coloration of the ear; a line with a dark gray mBLP grain and a line with a light gray gBLP grain, carrying recombinant regions in chromosomes 1H, 2H, 3H. Lignin and suberin deposition were assessed on transverse sections of root tissue. Lignin and suberin on histological sections were stained with an aqueous solution of berberine hemisulfate. On the 7th day after the start of germination, on sections made in the basal part of the root, luminescence with berberine revealed no differences between plants of different genotypes in the level of accumulation of lignin and suberin. On the 10th day, on sections of the basal part of the roots of the gBLP line, staining of the radial walls of the endoderm is noticeable, which indicates the deposition of lignin in the endoderm area, characteristic of the formation of Casparian bands, as well as in the area of metaxylem vessels. Thus, it was revealed that the gBLP genotype is characterized by accelerated formation of apoplastic barriers. The Bowman and gBLP genotypes we selected, which contrast in the rate of formation of Casparian bands, are a convenient object for further identifying the relationship between the rate of formation of Casparian bands and salt tolerance of plants.

Localization and synthesis of lipid-transfer proteins in pea plants after treatment by strain of growth-stimulating bacteria and their role in the formation of Casparian bands and the regulation of hydraulic conductance

The presence of PGP bacteria in the rhizosphere has a positive effect on plant growth and increases their productivity both under favorable and stressful conditions. Despite the fact that much attention is paid to the various mechanisms of action of growth-promoting bacteria on plants, the aspect of the effect of bacteria on changes in the formation of apoplastic barriers and their effect on water transport remains almost unexplored. Previously, it was shown that Pseudomonas mandelii IB-Ki14 accelerated the formation of Casparian bands in wheat plants, although the involvement of LTPs (Lipid Transfer Proteins) in this process has not been shown. The aim of this study was to elucidate the effect of bacterial inoculation on LTP synthesis in the roots of the pea (Pisum sativum) and its relation to the formation of apoplastic barriers and hydraulic conductance. We aimed to test the hypothesis that the decrease in hydraulic conductance due to increased formation of apoplastic barriers can be compensated by increased activity of aquaporin water channels. Activation of genes encoding PsLTP and an increase in the amount of lipid-transporting proteins in pea root phloem induced by P. mandelii IB-Ki14 treatment were found. Inoculation was accompanied by an increase in suberin deposition in the Casparian bands, while hydraulic conductivity did not decrease. The use of Fenton's reagent, which is an inhibitor of aquaporins, showed that the hydraulic conductivity of plants treated with bacteria decreased to a greater extent compared to the control group, which indicates an increase in the activity of aquaporins under the influence of bacteria. We conclude that P. mandelii IB-Ki14 stimulates the deposition of suberin, the biosynthesis of which involves lipid-transporting proteins, and increases the activity of aquaporins, which, in turn, prevents a possible decrease in hydraulic conductivity due to the formation of apoplastic barriers in pea roots.

Effects of Phytohormone-Producing Rhizobacteria on Casparian Band Formation, Ion Homeostasis and Salt Tolerance of Durum Wheat

Inoculation with plant growth-promoting rhizobacteria can increase plant salt resistance. We aimed to reveal bacterial effects on the formation of apoplastic barriers and hormone concentration in relation to maintaining ion homeostasis and growth of salt-stressed plants. The rhizosphere of a durum wheat variety was inoculated with cytokinin-producing Bacillus subtilis and auxin-producing Pseudomonas mandelii strains. Plant growth, deposition of lignin and suberin and concentrations of sodium, potassium, phosphorus and hormones were studied in the plants exposed to salinity. Accumulation of sodium inhibited plant growth accompanied by a decline in potassium in roots and phosphorus in shoots of the salt-stressed plants. Inoculation with both bacterial strains resulted in faster appearance of Casparian bands in root endodermis and an increased growth of salt-stressed plants. B. subtilis prevented the decline in both potassium and phosphorus concentrations and increased concentration of cytokinins in salt-stressed plants. P. mandelii decreased the level of sodium accumulation and increased the concentration of auxin. Growth promotion was greater in plants inoculated with B. subtilis. Increased ion homeostasis may be related to the capacity of bacteria to accelerate the formation of Casparian bands preventing uncontrolled diffusion of solutes through the apoplast. We discuss the relative impacts of the decline in Na accumulation and maintenance of K and P content for growth improvement of salt-stressed plants and their possible relation to the changes in hormone concentration in plants.

ЛОКАЛИЗАЦИЯ АКВАПОРИНОВ В ПРОЦЕССЕ ФОРМИРОВАНИЯ ПОЯСКОВ КАСПАРИ В КОРНЯХ РАСТЕНИЙ ЯЧМЕНЯ

There are two main pathways for water transport in plants. One of them is water transport along the cell walls and xylem vessels, called apoplastic pathway, and the other is from cell to cell through the membrane water channels aquaporins. Formation of Casparian bands by suberin and lignin deposition reduces conductivity of the apoplastic pathway. So the role of membrane water transport should increase, as mentioned in many reviews. The aim of this research was to localize barley aquaporins of the HvPIP2 family in different root zones depending on the differentiation of cell walls and deposition of Casparian bands. It was shown that the intensity of berberine fluorescence was insignificant in the zone closer to the root tip (at a distance of 3–4 cm), however fluorescence increased in the endodermis and the central cylinder of the root basal zone, indicating high lignification of cell walls and deposition of lignin and suberin in developing Casparian bands. Localization of HvPIP2;1 and HvPIP2;5 aquaporins showed no differences in their level between different root zones. The use of antibodies against aquaporins HvPIP2;2 and HvPIP2;3/4 revealed a local increase in fluorescence in the endodermal region of the basal zone of the roots characterized by high level of lignification and suberinization which apparently should compensate for the expected decrease in hydraulic conductivity as a result of the formation of apoplastic barriers. It is possible that the role of different aquaporins in water transport varies depending on the growing conditions of plants.

ВЛИЯНИЕ ГИПОКСИИ НА ФОРМИРОВАНИЕ АЭРЕНХИМЫ И РОСТ РАСТЕНИЙ ПШЕНИЦЫ

Hypoxia as a stress condition occurs due to a decrease in the availability of oxygen in the environment as a result of natural waterlogging of soils or suboptimal watering. In laboratory conditions, when growing plants in hydroponics, hypoxia was modeled by stopping the aeration of the nutrient solution, which led to a rapid decrease of oxygen concentration in it. Under these conditions was observed, the formation of aerenchyma, through which oxygen can penetrate the roots from the shoot. In addition, the luminescence of berberine complexes with lignin and suberin in the cell membranes of the outer layers of the cortex increased, which indicates the beginning of the formation of apoplastic barriers in this area, which in turn can reduce the diffusion of oxygen into the solution on the way of its transport to the apical part of the root along aerenchyma. In the morphological features the most striking was the decrease in the length of the root under hypoxic conditions, which reduses the path of oxygen transport from the shoot to the root tips. However, the decrease in the rate of shoot growth and photosynthesis under the influence of hypoxia was insignificant, which confirms the adaptive significance of the aerenchyma and apoplast barriers formation that provide oxygen supply to the roots.

Root characteristics critical for cadmium tolerance and reduced accumulation in wheat (Triticum aestivum L.)

Root radial transport is important for cadmium (Cd) absorption and root-shoot translocation. However, the relationship between root structural characteristics and radial transport of Cd in wheat is still unclear. Six wheat cultivars with different Cd tolerance and accumulation characteristics were used to investigate the roles of root phenotype, microstructure, and apoplastic and symplastic pathways in Cd uptake and root-shoot transport in pot culture. Longer root length, smaller root diameter, and more numerous root tips were more conducive to Cd absorption, while thicker roots were able to retain more Cd, thus reducing root-shoot transport and improving Cd tolerance of shoots. Cd stress can induce the deposition of apoplastic barriers in wheat roots, and the deposition of the apoplastic barrier increases under greater stress. The formation of apoplastic barriers can reduce Cd absorption and transfer to the shoot, and the presence of passage cells can weaken this effect. The cell wall thickening induced by Cd stress enhanced Cd adsorption capacity in wheat roots, but there was no significant correlation between Cd content and polysaccharide content in the cell wall. The up-regulated expression of TaHMA3 and TaVP1, which encode proteins related to Cd compartmentalization, was associated with increased Cd tolerance in wheat and decreased Cd translocation to aboveground parts. The morphology and anatomy of roots appear to play critical roles in Cd tolerance, uptake, and translocation in wheat. The present study provides useful information for the selection of wheat cultivars with low Cd accumulation.

The barrier to radial oxygen loss impedes the apoplastic entry of iron into the roots of Urochloa humidicola.

Lack of O2 and high concentrations of iron (Fe) and manganese (Mn) commonly occur in waterlogged soils. The development of a barrier to impede radial O2 loss (ROL) is a key trait improving internal O2 transport and waterlogging tolerance in plants. We evaluated the ability of the barrier to ROL to impede the entry of excess Fe into the roots of the waterlogging-tolerant grass Urochloa humidicola. Plants were grown in aerated or stagnant deoxygenated nutrient solution with 5 µM or 900 µM Fe. Quantitative X-ray microanalysis was used to determine cell-specific Fe concentrations at two positions behind the root apex in relation to ROL and the formation of apoplastic barriers. At a mature zone of the root, Fe was 'excluded' at the exodermis where a suberized lamella was evident, a feature also associated with a strong barrier to ROL. In contrast, the potassium (K) concentration was similar in all root cells, indicating that K uptake was not affected by apoplastic barriers. The hypothesis that the formation of a tight barrier to ROL impedes the apoplastic entry of toxic concentrations of Fe into the mature zones of roots was supported by the significantly higher accumulation of Fe on the outer side of the exodermis.

ИЗМЕНЕНИЕ ВКЛАДА АКВАПОРИНОВ В ГИДРАВЛИЧЕСКУЮ ПРОВОДИМОСТЬ РАСТЕНИЙ НА ФОНЕ ФОРМИРОВАНИЯ АПОПЛАСТНЫХ БАРЬЕРОВ ПОД ВЛИЯНИЕМ ЗАСОЛЕНИЯ

The study of plant adaptation mechanisms during the salt stress is required to provide an increase in plant productivity under such conditions. Along with a decrease in the availability of water for plants, the NaCL-induced inhibition of plant growth is associated with the toxic effect of sodium ions. The formation of apoplastic barriers due to the deposition of suberin and lignin restricts passive ion diffusion. However, the formation of such barriers reduces the capacity of the apoplastic pathway for water movement. In these conditions the role of transmembrane water transport is increased. This process is provided by aquaporin water channels. Thus the purpose of this work was to determine the contribution of aquaporins to hydraulic conductivity of peas plants under salinity-induced apoplastic barrier formation. An only slight decrease in plants transpiration caused by mercury chloride in the absence of salinization was in accordance with the ideas the apoplast is the dominant pathway when the Casparian bands is not formed yet. Salt stress in our experiments accelerated the development of the Casparian bands formation which could be visualized as an appearance of suberin strips in root endodermis which in turn was accompanied by a decrease in hydraulic conductivity. The decrease in hydraulic conductivity in 2 times during the mercury chloride treatment under salinity confirmed that contribution of aquaporins to the total hydraulic conductivity was increased under conditions when Casparian bands have had formed.

A redundant hydraulic function of root hairs in barley plants grown in hydroponics.

The root hair-less brb of Hordeum vulgare L. (bald root barley) mutant was used to assess the significance that root hairs have for the hydraulic properties of roots and response to a limited supply of mineral nutrients in plants grown on hydroponics. The barley brb mutant and its parent wild-type (H. vulgare cv. Pallas) were grown under nutrient sufficient control conditions, and under conditions of low supply of P and N. Plants were analysed when they were 14-18 days old. Root hydraulic conductivity (Lp) was determined for excised root systems and intact transpiring plants, and cell Lp was determined through cell pressure probe measurements. The formation of Casparian bands and suberin lamellae was followed through staining of cross-sections. The presence or absence of root hairs had no effect on the overall hydraulic response of plants to nutritional treatments. Root and cell Lp did not differ between the two genotypes. The most apparent difference between brb and wild-type plants was the consistently reduced formation of apoplastic barriers in brb plants. Any hydraulic function of root hairs can be redundant in barley, at least under the hydroponic conditions tested.

Salt stress reduces root water uptake in barley (Hordeum vulgare L.) through modification of the transcellular transport path.

The aim of the study was to understand the hydraulic response to salt stress of the root system of the comparatively salt-tolerant crop barley (Hordeum vulgare L.). We focused on the transcellular path of water movement across the root cylinder that involves the crossing of membranes. This path allows for selective water uptake, while excluding salt ions. Hydroponically grown plants were exposed to 100 mM NaCl for 5-7 days and analysed when 15-17 days old. A range of complementary and novel approaches was used to determine hydraulic conductivity (Lp). This included analyses at cell, root and plant level and modelling of water flow. Apoplastic barrier formation and gene expression level of aquaporins (AQPs) was analysed. Salt stress reduced the Lp of root system through reducing water flow along the transcellular path. This involved changes in the activity and gene expression level of AQPs. Modelling of root-Lp showed that the reduction in root-Lp did not require added hydraulic resistances through apoplastic barriers at the endodermis. The bulk of data points to a near-perfect semi-permeability of roots of control plants (solute reflection coefficient σ ~1.0). Roots of salt-stressed plants are almost as semi-permeable (σ > 0.8).

Regulation of a Cytochrome P450 Gene CYP94B1 by WRKY33 Transcription Factor Controls Apoplastic Barrier Formation in Roots to Confer Salt Tolerance.

Salinity is an environmental stress that causes decline in crop yield. Avicennia officinalis and other mangroves have adaptations such as ultrafiltration at the roots aided by apoplastic cell wall barriers to thrive in saline conditions. We studied a cytochrome P450 gene from A. officinalis, AoCYP94B1, and its putative ortholog in Arabidopsis (Arabidopsis thaliana), AtCYP94B1, which are involved in apoplastic barrier formation. Both genes were induced by 30 min of salt treatment in the roots. Heterologous expression of AoCYP94B1 in the atcyp94b1 Arabidopsis mutant and wild-type rice (Oryza sativa) conferred increased NaCl tolerance to seedlings by enhancing root suberin deposition. Histochemical staining and gas chromatography-tandem mass spectrometry quantification of suberin precursors confirmed the role of CYP94B1 in suberin biosynthesis. Using chromatin immunoprecipitation and yeast one-hybrid and luciferase assays, we identified AtWRKY33 as the upstream regulator of AtCYP94B1 in Arabidopsis. In addition, atwrky33 mutants exhibited reduced suberin and salt-sensitive phenotypes, which were rescued by expressing 35S::AtCYP94B1 in the atwrky33 background. This further confirmed that AtWRKY33-mediated regulation of AtCYP94B1 is part of the salt tolerance mechanism. Our findings may help efforts aimed at generating salt-tolerant crops.

Photosynthetically active radiation impacts significantly on root and cell hydraulics in barley (Hordeum vulgare L.).

Photosynthetically active radiation (PAR) affects transpirational water loss, yet we do not know through which mechanisms root water uptake is adjusted in parallel. Here, we exposed hydroponically grown barley plants to three levels of PAR [Normal (control), Low, High] and focused on the role which aquaporins (AQPs), apoplastic barriers (Casparian bands, suberin lamellae) and root morphology play in the adjustment of root hydraulic conductivity (Lp). Plants were analyzed when they were 14-18 days (d) old. Root and cell Lp, which involves AQP activity, was determined through exudation and cell pressure probe measurements, respectively. Gene expression of AQPs was analyzed through qPCR. The formation of apoplastic barriers was studied through staining of cross-sections. The rate of transpirational water loss per plant and unit leaf area increased in response to high-PAR and decreased in response to low-PAR treatments, both during day and night. Hydraulic conductivity in roots decreased significantly at organ and cell level in response to Low-PAR, and increased (organ) or did not change (cell level) in response to High-PAR. The formation of apoplastic barriers was little affected by PAR. Gene expression of AQPs tended to be highest in the Low-PAR treatment. Lateral roots, showing few apoplastic barriers, contributed the least in Low- and the most to root surface area in High-PAR plants. It is concluded that barley plants which experience changes in shoot transpirational water loss in response to PAR adjust root water uptake through changes in root Lp, and that these changes are mediated through altered AQP activity and root morphology.

Ethylene-mediated apoplastic barriers development involved in cadmium accumulation in root of hyperaccumulator Sedum alfredii

Ethylene is an important phytohormone for plant adaptation to heavy metal stress. However, the effects of ethylene on radial apoplastic transport of Cd remain elusive. This study investigated the role of ethylene on apoplastic barriers development and consequences for Cd uptake in Sedum alfredii. In response to Cd, endogenous ethylene production in hyperaccumulating ecotype (HE) roots was decreased due to the down-regulated expressions of ethylene biosynthesis genes, while the opposite result was observed in non-hyperaccumulating ecotype (NHE). Interestingly, the ethylene emission in HE was always higher than that in NHE, regardless of Cd concentrations. Results of exogenous application of ethylene biosynthesis precursor/inhibitor indicate that ethylene with high level would delay the formation of apoplastic barriers in HE through restraining phenylalanine ammonia lyase activity and gene expressions related to lignin/suberin biosynthesis. Simultaneously, correlation analyses suggest that Cd-induced apoplastic barriers formation may be also regulated by ethylene signaling. By using an apoplastic bypass tracer and scanning ion-selected electrode, we observed that the delayed deposition of apoplastic barriers significantly promoted Cd influx in roots. Taken together, high endogenous ethylene in HE postponed the formation of apoplastic barriers and thus promoted the Cd accumulation in the apoplast of roots.

The developmental dynamics of the sweet sorghum root transcriptome elucidate the differentiation of apoplastic barriers

ABSTRACTApoplastic barriers in the endodermis, such as Casparian strips and suberin lamellae, control the passage of water and minerals into the stele. Apoplastic barriers are thus thought to contribute to salt exclusion in salt-excluding plants such as sweet sorghum (Sorghum bicolor). However, little is known about the genes involved in the development of the apoplastic barrier. Here, we identified candidate genes involved in Casparian strip and suberin lamella development in the roots of a sweet sorghum line (M-81E). Three distinct developmental regions (no differentiation, developing, and mature) were identified based on Casparian strip and suberin lamella staining in root cross sections. Sequencing of RNA extracted from these distinct sections identified key genes participating in the differentiation of the apoplastic barrier. The different sections were structurally distinct, presumably due to differences in gene expression. Genes controlling the phenylpropanoid pathway, fatty acid elongation, and fatty acid ω-hydroxylation appeared to be directly responsible for the formation of the apoplastic barrier. Our dataset elucidates the molecular processes underpinning apoplastic barrier development and provides a basis for future research on molecular mechanisms of apoplastic barrier formation and salt exclusion.Abbreviations: SHR, SHORTROOT; MYB, MYB DOMAIN PROTEIN; CIFs, Casparian strip integrity factors; CASP, Casparian strip domain proteins; PER, peroxidase; ESB1, ENHANCED SUBERIN1; CS, Casparian strip; RPKM, reads per kilobase per million reads; DEGs, differentially expressed genes; FDR, false discovery rate; GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; RNA-seq, RNA sequencing; PAL, phenylalanine ammonia-lyase; CYP, cytochrome P450 monooxygenases; 4CL, 4-coumarate-CoA ligase; AAE5, ACYL-ACTIVATING ENZYME5; CCR, cinnamoyl CoA reductase; TKPR, TETRAKETIDE ALPHA-PYRONE REDUCTASE1; CAD, cinnamyl alcohol dehydrogenase; HST, shikimate O-hydroxycinnamoyltransferase; PMAT2, PHENOLIC GLUCOSIDE MALONYLTRANSFERASE2; CCOAOMT, caffeoyl-CoA O-methyltransferase; KCS, β-ketoacyl-CoA synthase; CUT1, CUTICULAR PROTEIN1; DET2, 5-alpha-reductase; TAX, 3ʹ-N-debenzoyl-2ʹ-deoxytaxol N-benzoyltransferase; CER1, ECERIFERUM1; FAR, fatty acyl reductase; AF-CoA, alcohol-forming fatty acyl-CoA reductase; ABCG, ATP-binding cassette, subfamily G; ERF, ethylene-responsive transcription factor; HSF, heat stress transcription factor; NTF, NUCLEAR TRANSCRIPTION FACTOR Y SUBUNIT B-5; GPAT, glycerol 3-phosphate acyltransferase.

Silencing against the conserved NAC domain of the potato StNAC103 reveals new NAC candidates to repress the suberin associated waxes in phellem

Both suberin and its associated waxes contribute to the formation of apoplastic barriers that protect plants from the environment. Some transcription factors have emerged as regulators of the suberization process. The potato StNAC103 gene was reported as a repressor of suberin polyester and suberin-associated waxes deposition because its RNAi-mediated downregulation (StNAC103-RNAi) over-accumulated suberin and associated waxes in the tuber phellem concomitantly with the induction of representative biosynthetic genes. Here, to explore if other genes of the large NAC gene family participate to this repressive function, we extended the silencing to other NAC members by targeting the conserved NAC domain of StNAC103 (StNAC103-RNAi-c). Transcript profile of the StNAC103-RNAi-c phellem indicated that StNAC101 gene was an additional potential target. In comparison with StNAC103-RNAi, the silencing with StNAC103-RNAi-c construct resulted in a similar effect in suberin but yielded an increased load of associated waxes in tuber phellem, mainly alkanes and feruloyl esters. Globally, the chemical effects in both silenced lines are supported by the transcript accumulation profile of genes involved in the biosynthesis, transport and regulation of apoplastic lipids. In contrast, the genes of polyamine biosynthesis were downregulated. Altogether these results point out to StNAC101 as a candidate to repress the suberin-associated waxes.

Cortex cell hydraulic conductivity, endodermal apoplastic barriers and root hydraulics change in barley (Hordeum vulgare L.) in response to a low supply of N and P.

Mineral nutrient limitation affects the water flow through plants. We wanted to test on barley whether any change in root-to-shoot ratio in response to low supply of nitrogen and phosphate is accompanied by changes in root and cell hydraulic properties and involves changes in aquaporin (AQP) gene expression and root apoplastic barriers (suberin lamellae, Casparian bands). Plants were grown hydroponically on complete nutrient solution or on solution containing only 3.3 % or 2.5 % of the control level of nutrient. Plants were analysed when they were 14-18 d old. Nutrient-limited plants adjusted water flow to an increased root-to-shoot surface area ratio through a reduction in root hydraulic conductivity (Lp) as determined through exudation analyses. Cortex cell Lp (cell pressure probe analyses) decreased in the immature but not the mature region of the main axis of seminal roots and in primary lateral roots. The aquaporin inhibitor HgCl2 reduced root Lp most in nutrient-sufficient control plants. Exchange of low-nutrient for control media caused a rapid (20-80 min) and partial recovery in Lp, though cortex cell Lp did not increase in any of the root regions analysed. The gene expression level (qPCR analyses) of five plasma membrane-localized AQP isoforms did not change in bulk root extracts, while the formation of apoplastic barriers increased considerably along the main axis of root and lateral roots in low-nutrient treatments. Decrease in root and cortex cell Lp enables the adjustment of root water uptake to increased root-to-shoot area ratio in nutrient-limited plants. Aquaporins are the prime candidate to play a key role in this response. Modelling of water flow suggests that some of the reduction in root Lp is due to increased formation of apoplastic barriers.