An ABC Transporter Is Involved in the Silicon-Induced Formation of Casparian Bands in the Exodermis of Rice.
ABSTRACT: Silicon (Si) promotes the formation of Casparian bands (CB) in rice and reduces radial oxygen loss (ROL). Further transcriptomic approaches revealed several candidate genes involved in the Si-induced formation of CB such as ATP binding cassette (ABC) transporter, Class III peroxidases, ligases and transferases. Investigation of these genes by means of overexpression (OE) and knockout (KO) mutants revealed the contribution of the ABC transporter (OsABCG25) to CB formation in the exodermis, which was also reflected in the expression of other OsABCG25 in the Si-promoted formation of CB genes related to the phenylpropanoid pathway, such as phenylalanine-ammonia-lyase, diacylglycerol O-acyltransferase and 4-coumarate-CoA ligase. Differential CB development in mutants and Si supply also affected the barrier function of the exodermis. OE of the ABC transporter and Si supply reduced the ROL from roots and Fe uptake. No effect on ROL and Fe uptake could be observed for the KO mutant. The presented research confirms the impact of the OsABCG25 in the Si-promoted formation of CB and its barrier functions.
Project description:We studied the effect of Silicon (Si) on Casparian band (CB) development, chemical composition of the exodermal CB and Si deposition across the root in the Si accumulators rice and maize and the Si non-accumulator onion. Plants were cultivated in nutrient solution with and without Si supply. The CB development was determined in stained root cross-sections. The outer part of the roots containing the exodermis was isolated after enzymatic treatment. The exodermal suberin was transesterified with MeOH/BF3 and the chemical composition was measured using gas chromatography-mass spectroscopy (GC-MS) and flame ionization detector (GC-FID). Laser ablation-inductively coupled plasma-mass spectroscopy (LA-ICP-MS) was used to determine the Si deposition across root cross sections. Si promoted CB formation in the roots of Si-accumulator and Si non-accumulator species. The exodermal suberin was decreased in rice and maize due to decreased amounts of aromatic suberin fractions. Si did not affect the concentration of lignin and lignin-like polymers in the outer part of rice, maize and onion roots. The highest Si depositions were found in the tissues containing CB. These data along with literature were used to suggest a mechanism how Si promotes the CB development by forming complexes with phenols.
Project description:Si has a wide range of effects in plant growth. One of these effects is the Si mediated formation of Casparian bands (CB), and thereby a reduced radial oxygen loss (ROL). The GeneChip provided evidence for involved genes in plant Si reaction. The given GeneChip-data revealed and influence of Si in the uptake of Fe. Additionally several genes involved in CB formation were found regulated through Si.
Project description:Gramineous plants take up silicon (Si) that enhances the formation of exodermal Casparian bands (CBs) in the roots of rice (Oryza sativa L.). Furthermore, it is known that Si supply reduces the concentration of Fe in rice shoots. We hypothesized that the Si-enhanced CB formation in the exodermis reduces in the flux of Fe in the apoplast and the uptake of Fe loaded deoxymugineic acid. Thus, the effect of silicic acid supply at varied Fe concentrations and Fe forms was investigated in nutrient solution. The Fe concentrations in the shoot and apoplastic Fe concentrations in the root were determined and an Affymetrix GeneChip experiment was carried out together with qRT-PCR measurements for observation of transcriptomic reactions. Additionally, the Fe uptake of an overexpression mutant of OsABCG25 with an enhanced exodermal CB formation was investigated. The application of silicic acid reduced the Fe concentrations in shoot DM independently of the supplied Fe concentration and Fe form. As a reaction to the Fe shortage, the full cascade of Fe-homeostasis-related genes in the roots was upregulated. Silicic acid supply also decreased the apoplastic Fe concentrations in roots. In addition, an overexpression mutant of OsABCG25 with an enhanced CB formation showed a reduced uptake of Fe in excess Fe conditions. The results suggest that the Si-induced CB formation in the exodermis hampers the flux of Fe into the apoplast of the cortex and, thus, Fe uptake of rice grown in nutrient solution which is reflected in the upregulation of Fe homeostasis-related genes.
Project description:The beneficial element silicon (Si) may affect radial oxygen loss (ROL) of rice roots depending on suberization of the exodermis and lignification of sclerenchyma. Thus, the effect of Si nutrition on the oxidation power of rice roots, suberization and lignification was examined. In addition, Si-induced alterations of the transcript levels of 265 genes related to suberin and lignin synthesis were studied by custom-made microarray and quantitative Real Time-PCR. Without Si supply, the oxidation zone of 12 cm long adventitious roots extended along the entire root length but with Si supply the oxidation zone was restricted to 5 cm behind the root tip. This pattern coincided with enhanced suberization of the exodermis and lignification of sclerenchyma by Si supply. Suberization of the exodermis started, with and without Si supply, at 4-5 cm and 8-9 cm distance from the root tip (drt), respectively. Si significantly increased transcript abundance of 12 genes, while two genes had a reduced transcript level. A gene coding for a leucine-rich repeat protein exhibited a 25-fold higher transcript level with Si nutrition. Physiological, histochemical, and molecular-biological data showing that Si has an active impact on rice root anatomy and gene transcription is presented here.
Project description:A barrier to radial oxygen loss (ROL), which reduces the loss of oxygen transported via the aerenchyma to the root tips, enables the roots of wetland plants to grow into anoxic/hypoxic waterlogged soil. However, little is known about its genetic regulation. Quantitative trait loci (QTLs) mapping can help to understand the factors that regulate barrier formation. Rice (Oryza sativa) inducibly forms an ROL barrier under stagnant conditions, while a few wetland plants constitutively form one under aerated conditions. Here, we evaluated the formation of a constitutive ROL barrier in a total of four accessions from two wild rice species. Three of the accessions were wetland accessions of O. glumaepatula, and the fourth was a non-wetland species of O. rufipogon. These species have an AA type genome, which allows them to be crossed with cultivated rice. The three O. glumaepatula accessions (W2165, W2149, and W1183) formed an ROL barrier under aerated conditions. The O. rufipogon accession (W1962) did not form a constitutive ROL barrier, but it formed an inducible ROL barrier under stagnant conditions. The three O. glumaepatula accessions should be useful for QTL mapping to understand how a constitutive ROL barrier forms. The constitutive barrier of W2165 was closely associated with suberization and resistance to penetration by an apoplastic tracer (periodic acid) at the exodermis but did not include lignin at the sclerenchyma.
Project description:<h4>Background and aims</h4>Many wetland species form aerenchyma and a barrier to radial O(2) loss (ROL) in roots. These features enhance internal O(2) diffusion to the root apex. Barrier formation in rice is induced by growth in stagnant solution, but knowledge of the dynamics of barrier induction and early anatomical changes was lacking.<h4>Methods</h4>ROL barrier induction in short and long roots of rice (Oryza sativa L. 'Nipponbare') was assessed using cylindrical root-sleeving O(2) electrodes and methylene blue indicator dye for O(2) leakage. Aerenchyma formation was also monitored in root cross-sections. Microstructure of hypodermal/exodermal layers was observed by transmission electron microscopy (TEM).<h4>Key results</h4>In stagnant medium, barrier to ROL formation commenced in long adventitious roots within a few hours and the barrier was well formed within 24 h. By contrast, barrier formation took longer than 48 h in short roots. The timing of enhancement of aerenchyma formation was the same in short and long roots. Comparison of ROL data and subsequent methylene blue staining determined the apparent ROL threshold for the dye method, and the dye method confirmed that barrier induction was faster for long roots than for short roots. Barrier formation might be related to deposition of new electron-dense materials in the cell walls at the peripheral side of the exodermis. Histochemical staining indicated suberin depositions were enhanced prior to increases in lignin.<h4>Conclusions</h4>As root length affected formation of the barrier to ROL, but not aerenchyma, these two acclimations are differentially regulated in roots of rice. Moreover, ROL barrier induction occurred before histochemically detectable changes in putative suberin and lignin deposits could be seen, whereas TEM showed deposition of new electron-dense materials in exodermal cell walls, so structural changes required for barrier functioning appear to be more subtle than previously described.
Project description:Internal aeration is crucial for root growth under waterlogged conditions. Some wetland plants have a structural barrier that impedes oxygen leakage from the basal part of roots called a radial oxygen loss (ROL) barrier. The ROL barrier reduces loss of oxygen transported via the aerenchyma to the root tips, enabling root growth into anoxic soil. The roots of some plants develop an ROL barrier under waterlogged conditions, while they remain leaky to oxygen under well-drained or aerated conditions. The main components of the inducible ROL barrier are thought to be suberin and lignin deposited at the outer cellular space (apoplast) in the outer part of roots. On the other hand, a few wetland plants including a species of Echinochloa form a constitutive ROL barrier, i.e., it is formed even in the absence of waterlogging. However, little is known about the components of constitutive ROL barriers. An ROL barrier is considered to be a characteristic of wetland species because it has not been found in any non-wetland species so far. Here, we examined whether Echinochloa species from non-waterlogged fields also form an inducible or constitutive ROL barrier. We found that three species of Echinochloa from non-waterlogged fields constitutively developed an ROL barrier under aerated conditions. Over 85% of their root exodermis cells were covered with suberin lamellae and had well-developed Casparian strips. These substances inhibited the infiltration of an apoplastic tracer (periodic acid), suggesting that the ROL barrier can also prevent the entry of phytotoxic compounds from the soil. Unlike the other Echinochloa species, E. oryzicola, which mainly inhabits rice paddies, was found to lack a constitutive ROL barrier under aerated conditions. Although close to 90% of its sclerenchyma was well lignified, it leaked oxygen from the basal part of roots. A high percentage (55%) of the root exodermis cells were not fortified with suberin lamellae. These results suggest that suberin is an important component of constitutive ROL barriers.
Project description:Potential components of the barrier to radial oxygen loss (ROL) are suberin and/or lignin, which accumulate at the cell wall in the cells of peripheral cell layers of the root. Chemical composition of the apoplastic barrier in rice roots was characterized and it was suggested that ROL can be restricted by the formation of a suberized exodermis and/or lignified sclerenchyma in the outer part of the root. To characterize reorganization of primary carbon metabolism in rice roots during the ROL barrier formation, we obtained the profiles of polar metabolites and the profiles of fatty acids of different zones of rice roots from plants growing in stagnant (anaerobic) and in well aerated medium. Biochemical data are combined with the results of microarray analysis.
Project description:Internal aeration is crucial for root growth in waterlogged soil. A barrier to radial oxygen loss (ROL) can enhance long- distance oxygen transport via the aerenchyma to the root tip; a higher oxygen concentration at the apex enables root growth into anoxic soil. The ROL barrier is formed within the outer part of roots (OPR). Suberin and/or lignin depos- ited in cell walls are thought to contribute to the barrier, but it is unclear which compound is the main constituent. This study describes gene expression profiles during ROL barrier formation in rice roots to determine the relative responses of suberin and/or lignin biosyntheses for the barrier. OPR tissues were isolated by laser microdissection and their transcripts were analysed by microarray. A total of 128 genes were significantly up- or downregulated in the OPR during the barrier formation. Genes associated with suberin biosynthesis were strongly upregulated, whereas genes associated with lignin biosynthesis were not. By an ab initio analysis of the promoters of the upregulated genes, the putative cis-elements that could be associated with transcription factors, WRKY, AP2/ERF, NAC, bZIP, MYB, CBT/DREB, and MADS, were elucidated. They were particularly associated with the expression of transcrip- tion factor genes containing WRKY, AP2, and MYB domains. A semiquantitative reverse-transcription PCR analysis of genes associated with suberin biosynthesis (WRKY, CYP, and GPAT) confirmed that they were highly expressed during ROL barrier formation. Overall, these results suggest that suberin is a major constituent of the ROL barrier in roots of rice. 23-d-old plants were either continued in aerated solution or transplanted into N2-flushed or stagnant deoxygenated solution for 9 h. After treating the roots of plants in aerated, stagnant, or N2-flushed conditions for 9h, the basal parts (12.5 -22.5mm below the root - shoot junction) of the adventitious roots were collected. Cells in OPR (including exodermis and sclerenchyma) were isolated using laser microdissection. RNA extracted from the isolated OPR was analysed with a 44k rice oligo-DNA microarray. Total RNAs were labeled with a Quick Amp Labeling Kit (Agilent Technologies) according to the manufacturerM-bM-^@M-^Ys instructions. Aliquots of Cy5-labeled and Cy3-labeled cRNA (10 ng each) were used for hybridization in a rice 44K oligo-DNA microarray.
Project description:Potential components of the barrier to radial oxygen loss (ROL) are suberin and/or lignin, which accumulate at the cell wall in the cells of peripheral cell layers of the root. Chemical composition of the apoplastic barrier in rice roots was characterized and it was suggested that ROL can be restricted by the formation of a suberized exodermis and/or lignified sclerenchyma in the outer part of the root. To characterize reorganization of primary carbon metabolism in rice roots during the ROL barrier formation, we obtained the profiles of polar metabolites and the profiles of fatty acids of different zones of rice roots from plants growing in stagnant (anaerobic) and in well aerated medium. Biochemical data are combined with the results of microarray analysis. Nine days after germination, the seedlings were transferred to well aerated nutrient solution or stagnant deoxygenated nutrient solution. Stagnant solution contained 0.1% (w/v) dissolved agar and was deoxygenated (dissolved oxygen, <0.5 mg lM-bM-^@M-^S1) prior to use by flushing with N2 gas. After 14 d (23 d old), adventitious roots, 100-150 mm long, were harvested from rice plants grown either in aerated or stagnant conditions and RNA was extracted from 10 mm segments from the regions 0-10 mm, 10-20 mm and 20-30 mm from the root apex have been cut with sterile razor blade and collected and processed separately. Total RNAs were labeled with a Quick Amp Labeling Kit (Agilent Technologies) according to the manufacturerM-bM-^@M-^Ys instructions. Aliquots of Cy5-labeled and Cy3-labeled cRNA (825 ng each) were used for hybridization in a rice 44K oligo-DNA microarray.