Organic-inorganic hybrid material for hole transport in inverted perovskite solar cells.

Hole mobility is critical to the power conversion efficiencies of perovskite solar cells (PSCs). Organic small-molecule hole-transporting materials (HTMs) have attracted considerable interest in PSCs due to their structural flexibility and operational durability, but they suffer from modest hole mobility. On the other hand, inorganic HTMs with good hole mobility are inflexible in structural variation and exhibit unsatisfactory cell efficiency. In this study, a ligand BT28 and its zinc-based coordination complex BTZ30 were synthesized, characterized, and investigated as HTMs for PSC applications. The mixed-halide perovskites can be grown uniformly with large crystalline grains on both HTMs, which exhibit similar optical and electrochemical properties. However, it was discovered that the BTZ30-based solar cell exhibited an open-circuit voltage of 1.0817 V and a high short-circuit current density of 23.1392 mA cm-2 with a champion power conversion efficiency of close to 20 %. The performance difference between the two HTMs can be attributed to the difference in their hole mobilities, which is 63.31 % higher for BTZ30 than BT28. The comparison of non-metal and metal HTMs revealed the importance of considering hybrid structures to overcome some shortcomings associated with organic and inorganic HTMs and achieve high-performance PSCs.

The adaptation of root cell wall pectin to copper toxicity in two citrus species differing in copper tolerance: remodeling and responding.

Citrus species are prone to suffer from copper (Cu) toxicity because of improper application of Cu-based agrochemicals. Copper immobilization mediated by pectin methylesterase (PME) in the root cell wall (CW) is effective for Cu detoxification. However, the underlying mechanisms of the structural modification and stress responses of citrus root CW pectin to Cu toxicity have been less discussed. In the present study, seedlings of 'Shatian pummelo' (Citrus grandis L. Osbeck) and 'Xuegan' (Citrus sinensis L. Osbeck), which differ in Cu tolerance, were irrigated with nutrient solution containing 0.5 (as control), 100, 300 or 500 µM Cu for 18 weeks in sandy culture or 24 h in hydroponics. At the end of treatments in the 18-week sandy culture, Cu toxicity on CW pectin content, Cu distribution, degree of pectin methylesterification (DPM) and the PME enzyme activity were discussed. At the genome-wide level, PME gene family was identified from the two citrus species, and qRT-PCR array of citrus PMEs under control and 300 µM Cu stress for 18 weeks were performed to screen the Cu-responsive PME genes. Moreover, the candidate genes that responded to Cu toxicity were further examined within 24 h. The results showed that Cu toxicity increased the root CW pectin content. The root CW pectin under Cu toxicity was remodeled by upregulation of the expression of the Cu-responsive PME genes followed by increasing PME activity, which mainly promoted low methylesterased pectin level and the Cu content on root CW pectin. Compared with C. sinensis, C. grandis root CW had a lower DPM and higher Cu content on the Cu-stressed root CW pectin, contributing to its higher Cu tolerance. Our present study provided theoretical evidence for root CW pectin remodeling in response to Cu toxicity of citrus species.

Elevated pH-mediated mitigation of aluminum-toxicity in sweet orange (Citrus sinensis) roots involved the regulation of energy-rich compounds and phytohormones.

For the first time, we used targeted metabolome to investigate the effects of pH-aluminum (Al) interactions on energy-rich compounds and their metabolites (ECMs) and phytohormones in sweet orange (Citrus sinensis) roots. The concentration of total ECMs (TECMs) was reduced by Al-toxicity in 4.0-treated roots, but unaffected significantly in pH 3.0-treated roots. However, the concentrations of most ECMs and TECMs were not lower in pH 4.0 + 1.0 mM Al-treated roots (P4AR) than in pH 3.0 + 1.0 mM Al-treated roots (P3AR). Increased pH improved the adaptability of ECMs to Al-toxicity in roots. For example, increased pH improved the utilization efficiency of ECMs and the conversion of organic phosphorus (P) from P-containing ECMs into available phosphate in Al-treated roots. We identified upregulated cytokinins (CKs), downregulated jasmonic acid (JA), methyl jasmonate (MEJA) and jasmonates (JAs), and unaltered indole-3-acetic acid (IAA) and salicylic acid (SA) in P3AR vs pH 3.0 + 0 mM Al-treated roots (P3R); upregulated JA, JAs and IAA, downregulated total CKs, and unaltered MEJA and SA in P4AR vs pH 4.0 + 0 mM Al-treated roots (P4R); and upregulated CKs, downregulated JA, MEJA, JAs and SA, and unaltered IAA in P3AR vs P4AR. Generally viewed, raised pH-mediated increments of JA, MEJA, total JAs, SA and IAA concentrations and reduction of CKs concentration in Al-treated roots might help to maintain nutrient homeostasis, increase Al-toxicity-induced exudation of organic acid anions and the compartmentation of Al in vacuole, and reduce oxidative stress and Al uptake, thereby conferring root Al-tolerance. In short, elevated pH-mediated mitigation of root Al-stress involved the regulation of ECMs and phytohormones.

Molecular and Physiological Responses of Citrus sinensis Leaves to Long-Term Low pH Revealed by RNA-Seq Integrated with Targeted Metabolomics.

Low pH-induced alterations in gene expression profiles and organic acids (OA) and free amino acid (FAA) abundances were investigated in sweet orange [Citrus sinensis (L.) Osbeck cv. Xuegan] leaves. We identified 503 downregulated and 349 upregulated genes in low pH-treated leaves. Further analysis indicated that low pH impaired light reaction and carbon fixation in photosynthetic organisms, thereby lowering photosynthesis in leaves. Low pH reduced carbon and carbohydrate metabolisms, OA biosynthesis and ATP production in leaves. Low pH downregulated the biosynthesis of nitrogen compounds, proteins, and FAAs in leaves, which might be conducive to maintaining energy homeostasis during ATP deprivation. Low pH-treated leaves displayed some adaptive responses to phosphate starvation, including phosphate recycling, lipid remodeling, and phosphate transport, thus enhancing leaf acid-tolerance. Low pH upregulated the expression of some reactive oxygen species (ROS) and aldehyde detoxifying enzyme (peroxidase and superoxidase) genes and the concentrations of some antioxidants (L-tryptophan, L-proline, nicotinic acid, pantothenic acid, and pyroglutamic acid), but it impaired the pentose phosphate pathway and VE and secondary metabolite biosynthesis and downregulated the expression of some ROS and aldehyde detoxifying enzyme (ascorbate peroxidase, aldo-keto reductase, and 2-alkenal reductase) genes and the concentrations of some antioxidants (pyridoxine and γ-aminobutyric acid), thus disturbing the balance between production and detoxification of ROS and aldehydes and causing oxidative damage to leaves.

The aluminum distribution and translocation in two citrus species differing in aluminum tolerance.

BACKGROUND: Many citrus orchards of south China suffer from soil acidification, which induces aluminum (Al) toxicity. The Al-immobilization in vivo is crucial for Al detoxification. However, the distribution and translocation of excess Al in citrus species are not well understood. RESULTS: The seedlings of 'Xuegan' [Citrus sinensis (L.) Osbeck] and 'Shatianyou' [Citrus grandis (L.) Osbeck], that differ in Al tolerance, were hydroponically treated with a nutrient solution (Control) or supplemented by 1.0 mM Al3+ (Al toxicity) for 21 days after three months of pre-culture. The Al distribution at the tissue level of citrus species followed the order: lateral roots > primary roots > leaves > stems. The concentration of Al extracted from the cell wall (CW) of lateral roots was found to be about 8 to 10 times higher than in the lateral roots under Al toxicity, suggesting that the CW was the primary Al-binding site at the subcellular level. Furthermore, the Al distribution in CW components of the lateral roots showed that pectin had the highest affinity for binding Al. The relative expression level of genes directly relevant to Al transport indicated a dominant role of Cs6g03670.1 and Cg1g021320.1 in the Al distribution of two citrus species. Compared to C. grandis, C. sinensis had a significantly higher Al concentration on the CW of lateral roots, whereas remarkably lower Al levels in the leaves and stems. Furthermore, Al translocation revealed by the absorption kinetics of the CW demonstrated that C. sinensis had a higher Al retention and stronger Al affinity on the root CW than C. grandis. According to the FTIR (Fourier transform infrared spectroscopy) analysis, the Al distribution and translocation might be affected by a modification in the structure and components of the citrus lateral root CW. CONCLUSIONS: A higher Al-retention, mainly attributable to pectin of the root CW, and a lower Al translocation efficiency from roots to shoots contributed to a higher Al tolerance of C. sinensis than C. grandis. The aluminum distribution and translocation of two citrus species differing in aluminum tolerance were associated with the transcriptional regulation of genes related to Al transport and the structural modification of root CW.

Molecular mechanisms for magnesium-deficiency-induced leaf vein lignification, enlargement and cracking in Citrus sinensis revealed by RNA-Seq.

Citrus sinensis (L.) Osbeck seedlings were fertigated with nutrient solution containing 2 [magnesium (Mg)-sufficiency] or 0 mM (Mg-deficiency) Mg(NO3)2 for 16 weeks. Thereafter, RNA-Seq was used to investigate Mg-deficiency-responsive genes in the veins of upper and lower leaves in order to understand the molecular mechanisms for Mg-deficiency-induced vein lignification, enlargement and cracking, which appeared only in the lower leaves. In this study, 3065 upregulated and 1220 downregulated, and 1390 upregulated and 375 downregulated genes were identified in Mg-deficiency veins of lower leaves (MDVLL) vs Mg-sufficiency veins of lower leaves (MSVLL) and Mg-deficiency veins of upper leaves (MDVUL) vs Mg-sufficiency veins of upper leaves (MSVUL), respectively. There were 1473 common differentially expressed genes (DEGs) between MDVLL vs MSVLL and MDVUL vs MSVUL, 1463 of which displayed the same expression trend. Magnesium-deficiency-induced lignification, enlargement and cracking in veins of lower leaves might be related to the following factors: (i) numerous transciption factors and genes involved in lignin biosynthesis pathways, regulation of cell cycle and cell wall metabolism were upregulated; and (ii) reactive oxygen species, phytohormone and cell wall integrity signalings were activated. Conjoint analysis of proteome and transcriptome indicated that there were 287 and 56 common elements between DEGs and differentially abundant proteins (DAPs) identified in MDVLL vs MSVLL and MDVUL vs MSVUL, respectively, and that among these common elements, the abundances of 198 and 55 DAPs matched well with the transcript levels of the corresponding DEGs in MDVLL vs MSVLL and MDVUL vs MSVUL, respectively, indicating the existence of concordances between protein and transcript levels.

Raised pH conferred the ability to maintain a balance between production and detoxification of reactive oxygen species and methylglyoxal in aluminum-toxic Citrus sinensis leaves and roots.

Little is known about interactive effects of pH-aluminum (Al) on reactive oxygen species (ROS) and methylglyoxal (MG) metabolisms in plants. Citrus sinensis seedlings were fertilized with nutrient solution at an Al concentration of 1 or 0 mM and a pH of 4.0, 3.5, 3.0 or 2.5 for 18 weeks. Thereafter, gas exchange and chlorophylls in leaves, H2O2 generation, electrolyte leakage, total soluble proteins, MG, malondialdehyde (MDA), antioxidants, sulfur-containing compounds, enzymes [viz., antioxidant enzymes, sulfur metabolism-related enzymes, ascorbate oxidase, phosphomannose isomerase, glyoxalase I and glyoxalase II] involved in ROS and MG detoxification in leaves and roots were measured. Effects of low pH and Al-toxicity on these parameters displayed obvious synergism. Without Al-toxicity, low pH increased H2O2 production, electrolyte leakage, MDA and MG concentrations by 45.7%-90.3% (52.4%-73.6%), 24.3%-74.5% (26.7%-86.2%), 18.6%-44.8% (35.6%-53.7%) and 16.3%-47.1% (13.8%-51.7%) in leaves (roots) relative to pH 4, respectively; low pH-induced upregulation of enzymes involved in ROS and MG detoxification and sulfur-containing compounds in leaves and/or roots could not protect them against oxidative damage. At pH 2.5-3.0, Al-toxicity increased H2O2 production, electrolyte leakage, MDA and MG concentrations by 34.2%-35.5% (23.9%-72.7%), 10.2%-29.5% (23.7%-56.8%), 15.6%-35.7% (27.5%-33.9%) and 21.5%-26.8% (21.0%-49.2%) in leaves (roots), respectively, and decreased total soluble protein concentration by 46.2%-47.4% (18.8%-20.8%) in leaves (roots); at pH 3.5-4.0, Al-toxicity did not affect significantly the five parameters in leaves and roots except for Al-induced increases in root MDA concentration at pH 3.5-4.0 and root electrolyte leakage at pH 3.5, and Al-induced decrease in root total soluble protein concentration at pH 4.0. Raised pH conferred the ability to maintain a balance between production and detoxification of ROS and MG in leaves and roots, thus protecting them against oxidative damage, and hence alleviating Al-induced increase in electrolyte leakage and decrease in total soluble protein level.

Molecular and physiological mechanisms underlying magnesium-deficiency-induced enlargement, cracking and lignification of Citrus sinensis leaf veins.

Little is known about the physiological and molecular mechanisms underlying magnesium (Mg)-deficiency-induced enlargement, cracking and lignification of midribs and main lateral veins of Citrus leaves. Citrus sinensis (L.) Osbeck seedlings were irrigated with nutrient solution at a concentration of 0 (Mg-deficiency) or 2 (Mg-sufficiency) mM Mg(NO3)2 for 16 weeks. Enlargement, cracking and lignification of veins occurred only in lower leaves, but not in upper leaves. Total soluble sugars (glucose + fructose + sucrose), starch and cellulose concentrations were less in Mg-deficiency veins of lower leaves (MDVLL) than those in Mg-sufficiency veins of lower leaves (MSVLL), but lignin concentration was higher in MDVLL than that in MSVLL. However, all four parameters were similar between Mg-deficiency veins of upper leaves (MDVUL) and Mg-sufficiency veins of upper leaves (MSVUL). Using label-free, liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, we identified 1229 and 492 differentially abundant proteins (DAPs) in MDVLL vs MSVLL and MDVUL vs MSVUL, respectively. Magnesium-deficiency-induced alterations of Mg, nonstructural carbohydrates, cell wall components, and protein profiles were greater in veins of lower leaves than those in veins of upper leaves. The increased concentration of lignin in MDVLL vs MSVLL might be caused by the following factors: (i) repression of cellulose and starch accumulation promoted lignin biosynthesis; (ii) abundances of proteins involved in phenylpropanoid biosynthesis pathway, hormone biosynthesis and glutathione metabolism were increased; and (iii) the abundances of the other DAPs [viz., copper/zinc-superoxide dismutase, ascorbate oxidase (AO) and ABC transporters] involved in lignin biosynthesis were elevated. Also, the abundances of several proteins involved in cell wall metabolism (viz., expansins, Rho GTPase-activating protein gacA, AO, monocopper oxidase-like protein and xyloglucan endotransglucosylase/hydrolase) were increased in MDVLL vs MSVLL, which might be responsible for the enlargement and cracking of leaf veins.

Low pH effects on reactive oxygen species and methylglyoxal metabolisms in Citrus roots and leaves.

BACKGROUND: Limited data are available on the responses of reactive oxygen species (ROS) and methylglyoxal (MG) metabolisms to low pH in roots and leaves. In China, quite a few of Citrus are cultivated in acidic soils (pH < 5.0). 'Xuegan' (Citrus sinensis) and 'Sour pummelo' (Citrus grandis) (C. sinensis were more tolerant to low pH than C. grandis) seedlings were irrigated daily with nutrient solution at a pH of 2.5, 3 or 5 for nine months. Thereafter, we examined low pH effects on growth, and superoxide anion production rate (SAP), malondialdehyde (MDA), MG, antioxidants, and enzymes related to ROS and MG detoxification in roots and leaves in order to (a) test the hypothesis that low pH affected ROS and MG metabolisms more in roots than those of leaves, and (b) understand the roles of ROS and MG metabolisms in Citrus low pH-tolerance and -toxicity. RESULTS: Compared with control, most of the physiological parameters related to ROS and MG metabolisms were greatly altered at pH 2.5, but almost unaffected at pH 3. In addition to decreased root growth, many fibrous roots became rotten and died at pH 2.5. pH 2.5-induced changes in SAP, the levels of MDA, MG and antioxidants, and the activities of most enzymes related to ROS and MG metabolisms were greater in roots than those of leaves. Impairment of root ascorbate metabolism was the most serious, especially in C. grandis roots. pH 2.5-induced increases in MDA and MG levels in roots and leaves, decreases in the ratios of ascorbate/(ascorbate+dehydroascorbate) in roots and leaves and of reduced glutathione/(reduced+oxidized glutathione) in roots were greater in C. grandis than those in C. sinensis. CONCLUSIONS: Low pH affected MG and ROS metabolisms more in roots than those in leaves. The most seriously impaired ascorbate metabolism in roots was suggested to play a role in low pH-induced root death and growth inhibition. Low pH-treated C. sinensis roots and leaves had higher capacity to maintain a balance between ROS and MG production and their removal via detoxification systems than low pH-treated C. grandis ones, thus contribute to the higher acid-tolerance of C. sinensis.

Magnesium-Deficiency Effects on Pigments, Photosynthesis and Photosynthetic Electron Transport of Leaves, and Nutrients of Leaf Blades and Veins in Citrus sinensis Seedlings.

Citrus sinensis seedlings were irrigated with nutrient solution at a concentration of 0 (Mg-deficiency) or 2 (Mg-sufficiency) mM Mg (NO3)2 for 16 weeks. Mg-deficiency-induced interveinal chlorosis, vein enlargement and corkiness, and alterations of gas exchange, pigments, chlorophyll a fluorescence (OJIP) transients and related parameters were observed in middle and lower leaves, especially in the latter, but not in upper leaves. Mg-deficiency might impair the whole photosynthetic electron transport, including structural damage to thylakoids, ungrouping of photosystem II (PSII), inactivation of oxygen-evolving complex (OEC) and reaction centers (RCs), increased reduction of primary quinone electron acceptor (QA) and plastoquinone pool at PSII acceptor side and oxidation of PSI end-electron acceptors, thus lowering energy transfer and absorption efficiency and the transfer of electrons to the dark reactions, hence, the rate of CO2 assimilation in Mg-deficiency middle and lower leaves. Although potassium, Mg, manganese and zinc concentration in blades displayed a significant and positive relationship with the corresponding element concentration in veins, respectively, great differences existed in Mg-deficiency-induced alterations of nutrient concentrations between leaf blades and veins. For example, Mg-deficiency increased boron level in the blades of upper leaves, decreased boron level in the blades of lower leaves, but did not affect boron level in the blades of middle leaves and veins of upper, middle and lower leaves. To conclude, Mg-deficiency-induced interveinal chlorosis, vein enlargement, and corkiness, and alterations to photosynthesis and related parameters increased with increasing leaf age. Mg-deficiency-induced enlargement and corkiness of veins were not caused by Mg-deficiency-induced boron-starvation.

Increasing Nutrient Solution pH Alleviated Aluminum-Induced Inhibition of Growth and Impairment of Photosynthetic Electron Transport Chain in Citrus sinensis Seedlings.

Although the physiological and molecular responses of Citrus to Al-toxicity or low pH have been examined in some details, little information is available on Citrus responses to pH and aluminum (Al) interactions. Citrus sinensis seedlings were irrigated for 18 weeks with nutrient solution at a concentration of 0 or 1 mM AlCl3•6H2O and a pH of 2.5, 3.0, 3.5, or 4.0. Thereafter, biomass, root, stem, and leaf concentrations of Al and nutrients, leaf gas exchange, chlorophyll a fluorescence (OJIP) transients, and related parameters were investigated to understand the physiological mechanisms underlying the elevated pH-induced alleviation of Citrus toxicity. Increasing the nutrient solution pH from 2.5 to 4.0 alleviated the Al-toxic effects on biomass, photosynthesis, OJIP transients and related parameters, and element concentrations, uptake, and distributions. In addition, low pH effects on the above physiological parameters were intensified by Al-toxicity. Evidently, a synergism existed between low pH and Al-toxicity. Increasing pH decreased Al uptake per root dry weight and its concentration in roots, stems, and leaves and increased nitrogen, phosphorus, calcium, magnesium, sulfur, and boron uptake per plant and their concentrations in roots, stems, and leaves. This might be responsible for the elevated pH-induced alleviation of growth inhibition and the impairment of the whole photosynthetic electron transport chain, thus preventing the decrease of CO2 assimilation.

Low pH-responsive proteins revealed by a 2-DE based MS approach and related physiological responses in Citrus leaves.

BACKGROUND: Rare data are available on the molecular responses of higher plants to low pH. Seedlings of 'Sour pummelo' (Citrus grandis) and 'Xuegan' (Citrus sinensis) were treated daily with nutrient solution at a pH of 2.5, 3, or 6 (control) for nine months. Thereafter, we first used 2-dimensional electrophoresis (2-DE) to investigate low pH-responsive proteins in Citrus leaves. Meanwhile, we examined low pH-effects on leaf gas exchange, carbohydrates, ascorbate, dehydroascorbate and malondialdehyde. The objectives were to understand the adaptive mechanisms of Citrus to low pH and to identify the possible candidate proteins for low pH-tolerance. RESULTS: Our results demonstrated that Citrus were tolerant to low pH, with a slightly higher low pH-tolerance in the C. sinensis than in the C. grandis. Using 2-DE, we identified more pH 2.5-responsive proteins than pH 3-responsive proteins in leaves. This paper discussed mainly on the pH 2.5-responsive proteins. pH 2.5 decreased the abundances of proteins involved in ribulose bisphosphate carboxylase/oxygenase activation, Calvin cycle, carbon fixation, chlorophyll biosynthesis and electron transport, hence lowering chlorophyll level, electron transport rate and photosynthesis. The higher oxidative damage in the pH 2.5-treated C. grandis leaves might be due to a combination of factors including higher production of reactive oxygen species, more proteins decreased in abundance involved in antioxidation and detoxification, and lower ascorbate level. Protein and amino acid metabolisms were less affected in the C. sinensis leaves than those in the C. grandis leaves when exposed to pH 2.5. The abundances of proteins related to jasmonic acid biosynthesis and signal transduction were increased and decreased in the pH 2.5-treated C. sinensis and C. grandis leaves, respectively. CONCLUSIONS: This is the first report on low pH-responsive proteins in higher plants. Thus, our results provide some novel information on low pH-toxicity and -tolerance in higher plants.

Proteome profile analysis of boron-induced alleviation of aluminum-toxicity in Citrus grandis roots.

Aluminum (Al)-toxicity and boron (B)-deficiency are two major factors limiting crop production in tropical and subtropical areas. Elevating B supply can alleviate the Al-induced inhibition of growth in Citrus grandis. Seedlings of C. grandis were irrigated for 18 weeks with nutrient solutions containing two B levels (2.5 and 20 µM H3BO3) and two Al levels (0 and 1.2 mM AlCl3·6H2O). By using 2-dimensional electrophoresis (2-DE) based MALDI-TOF/TOF-MS method, this study successfully identified and quantified sixty-one differentially abundant proteins in Citrus roots in response to B-Al interactions. The mechanisms underlying the B-induced alleviation of Al-toxicity unveiled by 2-DE technique could be summarized as follows: a) remodeling of cell wall by reducing the synthesis of lignin (sugar ATP Binding Cassette (ABC) transporter ATPase and cinnamyl alcohol dehydrogenase) and increasing the modification of cell wall (UDP-forming); b) enhancing the abundances of proteasomes and turnover of dysfunctional proteins (proteasome or protease); c) increasing the abundance of stress response proteins, such as alcohol dehydrogenase, S-adenosylmethionine synthetase (SAMS) and glycosyl hydrolase; d) reinforcing cellular biological regulation and signal transduction (calreticulin-1). For the first time, some proteins, such as cell division protein 48 (CDC48), calreticulin and phospholipase, which might be involved in the downstream signaling of Al in Citrus plants, were successfully identified.

Aluminum-responsive genes revealed by RNA-Seq and related physiological responses in leaves of two Citrus species with contrasting aluminum-tolerance.

Little is known about the physiological and molecular responses of leaves to aluminum (Al)-toxicity. Seedlings of Al-intolerant Citrus grandis and Al-tolerant Citrus sinensis were supplied daily with nutrient solution containing 0 mM (control) and 1.0 mM (Al-toxicity) AlCl3·6H2O for 18 weeks. We found that Al-treatment only decreased CO2 assimilation in C. grandis leaves, and that the Al-induced alterations of gene expression profiles were less in C. sinensis leaves than those in C. grandis leaves, indicating that C. sinensis seedlings were more tolerant to Al-toxicity than C. grandis ones. Al concentration was similar between Al-treated C. sinensis and C. grandis roots, but it was higher in Al-treated C. grandis stems and leaves than that in Al-treated C. sinensis stems and leaves. Al-treated C. sinensis seedlings accumulated relatively more Al in roots and transported relatively little Al to shoots. This might be responsible for the higher Al-tolerance of C. sinensis. Further analysis showed that the following several aspects might account for the higher Al-tolerance of C. sinensis, including: (a) Al-treated C. sinensis leaves had higher capacity to maintain the homeostasis of energy and phosphate, the stability of lipid composition and the integrity of cell wall than did Al-treated C. grandis leaves; (b) Al-triggered production of reactive oxygen species (ROS) and the other cytotoxic compounds was less in Al-treated C. sinensis leaves than that in Al-treated C. grandis leaves, because Al-toxicity decreased CO2 assimilation only in C. grandis leaves; accordingly, more upregulated genes involved in the detoxifications of ROS, aldehydes and methylglyoxal were identified in Al-treated C. grandis leaves; in addition, flavonoid concentration was increased only in Al-treated C. grandis leaves; (c) Al-treated C. sinensis leaves could keep a better balance between protein phosphorylation and dephosphorylation than did Al-treated C. grandis leaves; and (d) both the equilibrium of hormones and hormone-mediated signal transduction were greatly disrupted in Al-treated C. grandis leaves, but less altered in Al-treated C. sinensis leaves. Finally, we discussed the differences in Al-responsive genes between Citrus roots and leaves.

Long-term manganese-toxicity-induced alterations of physiology and leaf protein profiles in two Citrus species differing in manganese-tolerance.

Manganese (Mn)-intolerant 'Sour pummelo' (Citrus grandis) and Mn-tolerant 'Xuegan' (Citrus sinensis) seedlings were irrigated for 17 weeks with 2 (control) or 600µM (Mn-toxicity or -excess) MnSO4. C. sinensis had higher Mn-tolerance than C. grandis, as indicated by the higher photosynthesis rates in Mn-excess C. sinensis leaves. Under Mn-toxicity, Mn levels were similar between C. sinensis and C. grandis roots, but lower in C. sinensis leaves than in C. grandis leaves. This might be responsible for C. sinensis Mn-tolerance. Using two-dimensional electrophoresis, we identified more differentially abundant proteins (DAPs) in Mn-excess C. grandis than in Mn-excess C. sinensis leaves, which agrees with the higher Mn levels in Mn-excess C. grandis leaves. DAPs were mainly related to carbohydrate and energy metabolism, stress response, and protein and amino acid metabolism. DAPs involved in the cytoskeleton and signal transduction were found only in Mn-excess C. grandis leaves. We isolated more photosynthesis-related proteins with decreased abundances in Mn-excess C. grandis leaves than in Mn-excess C. sinensis leaves, which might account for the larger decrease in photosynthesis rates in C. grandis leaves. The abundances of proteins involved in reactive oxygen species (ROS) scavenging and photorespiration were increased in Mn-excess C. grandis leaves, while only proteins involved in ROS detoxification were increased in Mn-excess C. sinensis leaves. This agrees with the increased requirement for dissipating the excess absorbed light energy, which was higher in Mn-excess C. grandis leaves than Mn-excess C. sinensis leaves because Mn-toxicity inhibited photosynthesis to a greater degree in C. grandis leaves.

Magnesium-deficiency-induced alterations of gas exchange, major metabolites and key enzymes differ among roots, and lower and upper leaves of Citrus sinensis seedlings.

Magnesium (Mg)-deficiency is a widespread problem adversely affecting the quality and yield of crops, including citrus. 'Xuegan' [Citrus sinensis (L.) Osbeck] seedlings were irrigated every other day with nutrient solution at an Mg concentration of 0 mM (Mg-deficiency) or 1 mM (Mg-sufficiency) for 16 weeks. Thereafter, biomass, leaf mass per area, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), pigments in the upper and lower leaves, Mg, gas exchange, organic acids, nonstructural carbohydrates, total soluble proteins, amino acids, phenolics and anthocyanins, and key enzymes related to organic acid, amino acid and phenolic metabolisms in the roots, and upper and lower leaves were assayed in order to test the hypothesis that Mg-deficiency-induced alterations of gas exchange, major metabolites and key enzymes may differ among the roots, and upper and lower leaves. Magnesium-deficiency affected the most measured parameters more in the lower than in the upper leaves except for the nonstructural carbohydrates, but the variation trends were similar between the two. Despite increased accumulation of nonstructural carbohydrates, the lower CO2 assimilation in the Mg-deficient leaves was not caused by the feedback inhibition mechanism via sugar accumulation. Both dark respiration and organic acid metabolism were elevated in the Mg-deficient lower leaves to 'consume' the excess carbohydrates, and inhibited in the Mg-deficient roots with less accumulation of nonstructural carbohydrates to keep the balance of net carbon. More total phenolics and fewer anthocyanins were accumulated in the Mg-deficient lower leaves, whereas the accumulation of both total phenolics and anthocyanins was reduced in the Mg-deficient roots. Interestingly, amino acid biosynthesis was repressed in the Mg-deficient roots and lower leaves, thus lowering the level of total free amino acids in these roots and leaves. To conclude, great differences existed in the Mg-deficiency-induced alterations of gas exchange, major metabolites and key enzymes among the roots, and upper and lower leaves.

Effects of Low pH on Photosynthesis, Related Physiological Parameters, and Nutrient Profiles of Citrus.

Seedlings of "Xuegan" (Citrus sinensis) and "Sour pummelo" (Citrus grandis) were irrigated daily with a nutrient solution at a pH of 2.5, 3, 4, 5, or 6 for 9 months. Thereafter, the following responses were investigated: seedling growth; root, stem, and leaf concentrations of nutrient elements; leaf gas exchange, pigment concentration, ribulose-1,5-bisphosphate carboxylase/oxygenase activity and chlorophyll a fluorescence; relative water content, total soluble protein level, H2O2 production and electrolyte leakage in roots and leaves. This was done (a) to determine how low pH affects photosynthesis, related physiological parameters, and mineral nutrient profiles; and (b) to understand the mechanisms by which low pH may cause a decrease in leaf CO2 assimilation. The pH 2.5 greatly inhibited seedling growth, and many physiological parameters were altered only at pH 2.5; pH 3 slightly inhibited seedling growth; pH 4 had almost no influence on seedling growth; and seedling growth and many physiological parameters reached their maximum at pH 5. No seedlings died at any given pH. These results demonstrate that citrus survival is insensitive to low pH. H+-toxicity may directly damage citrus roots, thus affecting the uptake of mineral nutrients and water. H+-toxicity and a decreased uptake of nutrients (i.e., nitrogen, phosphorus, potassium, calcium, and magnesium) and water were likely responsible for the low pH-induced inhibition of growth. Leaf CO2 assimilation was inhibited only at pH 2.5. The combinations of an impaired photosynthetic electron transport chain, increased production of reactive oxygen species, and decreased uptake of nutrients and water might account for the pH 2.5-induced decrease in CO2 assimilation. Mottled bleached leaves only occurred in the pH 2.5-treated C. grandis seedlings. Furthermore, the pH 2.5-induced alterations of leaf CO2 assimilation, water-use efficiency, chlorophylls, polyphasic chlorophyll a fluorescence (OJIP) transients and many fluorescence parameters, root and leaf total soluble proteins, H2O2 production, and electrolyte leakage were all slightly greater in C. grandis than in C. sinensis seedlings. Hence, C. sinensis was slightly more tolerant to low pH than C. grandis. In conclusion, our findings provide novel insight into the causes of low pH-induced inhibition of seedling growth and leaf CO2 assimilation.

Root Adaptive Responses to Aluminum-Treatment Revealed by RNA-Seq in Two Citrus Species With Different Aluminum-Tolerance.

Seedlings of aluminum (Al)-tolerant Citrus sinensis and Al-intolerant Citrus grandis were fertigated daily with nutrient solution containing 0 and 1.0 mM AlCl3●6H2O for 18 weeks. The Al-induced decreases of biomass and root total soluble proteins only occurred in C. grandis, demonstrating that C. sinensis had higher Al-tolerance than C. grandis. Under Al-treatment, C. sinensis roots secreted more citrate and malate than C. grandis ones; less Al was accumulated in C. sinenis than in C. grandis leaves. The Al-induced reduction of phosphorus was lesser in C. sinensis roots and leaves than in C. grandis ones, whereas the Al-induced increase of sulfur was greater in C. sinensis roots and leaves. Using RNA-seq, we isolated 1905 and 2670 differentially expressed genes (DEGs) from Al-treated C. sinensis than C. grandis roots, respectively. Among these DEGs, only 649 DEGs were shared by the two species. Further analysis suggested that the following several aspects conferred C. sinensis higher Al-tolerance: (a) Al-treated C. sinensis seedlings had a higher external Al detoxification capacity via enhanced Al-induced secretion of organic acid anions, a higher antioxidant capacity and a more efficient chelation system in roots; (b) Al-treated C. sinensis seedlings displayed a higher level of sulfur in roots and leaves possibly due to increased uptake and decreased export of sulfur and a higher capacity to maintain the cellular phosphorus homeostasis by enhancing phosphorus acquisition and utilization; (c) Cell wall and cytoskeleton metabolism, energy and carbohydrate metabolism and signal transduction displayed higher adaptative responses to Al in C. sinensis than in C. grandis roots; (d) More upregulated than downregulated genes related to fatty acid and amino acid metabolisms were isolated from Al-treated C. sinensis roots, but the reverse was the case for Al-treated C. grandis roots. These results provide a platform for further investigating the roles of genes possibly responsible for citrus Al-tolerance.