Singlet oxygen induces cell wall thickening and stomatal density reducing by transcriptome reprogramming.

Singlet oxygen (1O2) has a very short half-life of 10-5 s; however, it is a strong oxidant that causes growth arrest and necrotic lesions on plants. Its signaling pathway remains largely unknown. The Arabidopsis flu (fluorescent) mutant accumulates a high level of 1O2 and shows drastic changes in nuclear gene expression. Only two plastid proteins, EX1 (executer 1) and EX2 (executer 2), have been identified in the singlet oxygen signaling. Here, we found that the transcription factor abscisic acid insensitive 4 (ABI4) binds the promoters of genes responsive to 1O2-signals. Inactivation of the ABI4 protein in the flu/abi4 double mutant was sufficient to compromise the changes of almost all 1O2-responsive-genes and rescued the lethal phenotype of flu grown under light/dark cycles, similar to the flu/ex1/ex2 triple mutant. In addition to cell death, we reported for the first time that 1O2 also induces cell wall thickening and stomatal development defect. Contrastingly, no apparent growth arrest was observed for the flu mutant under normal light/dim light cycles, but the cell wall thickening (doubled) and stomatal density reduction (by two-thirds) still occurred. These results offer a new idea for breeding stress tolerant plants.

Nitrate reductase is a key enzyme responsible for nitrogen-regulated auxin accumulation in Arabidopsis roots.

Nitrate reductase (NR) is one of the key enzymes for plant nitrogen assimilation and root architecture remodeling. However, crosstalk between NR-mediated signaling and auxin-mediated root development in nitrogen-status responses has not been investigated in details before. In this study, root phenotype and auxin distribution in nia1/nia2 (nitrate reductase) double mutant and chl1-5 (nitrate transporter NRT1.1) mutant under different nitrogen availabilities were compared. The nia1/nia2 mutant showed very low expression levels of auxin biosynthetic/signaling genes and was insensitive to nitrogen changes. While the chl1-5 mutant showed a high NR activity with a high level of auxin in the meristematic zone and a weaker response to nitrogen changes, when compared with the wild-type plants. We firstly found that NR activity was roughly positive-correlated with the root auxin level, and there is a crosstalk between nitrate signaling and auxin signaling. The putative signaling pathways downstream of NR have been discussed.

Vitamin E Is Superior to Vitamin C in Delaying Seedling Senescence and Improving Resistance in Arabidopsis Deficient in Macro-Elements.

Nitrogen (N), phosphorus (P), and potassium (K) are three essential macro-elements for plant growth and development. Used to improve yield in agricultural production, the excessive use of chemical fertilizers often leads to increased production costs and ecological environmental pollution. Vitamins C and E are antioxidants that play an important role in alleviating abiotic stress. However, there are few studies on alleviating oxidative stress caused by macro-element deficiency. Here, we used Arabidopsis vitamin E synthesis-deficient mutant vte4 and vitamin C synthesis-deficient mutant vtc1 on which exogenous vitamin E and vitamin C, respectively, were applied at the bolting stage. In the deficiency of macro-elements, the Arabidopsis chlorophyll content decreased, malondialdehyde (MDA) content and relative electric conductivity increased, and reactive oxygen species (ROS) accumulated. The mutants vtc1 and vte4 are more severely stressed than the wild-type plants. Adding exogenous vitamin E was found to better alleviate stress than adding vitamin C. Vitamin C barely affected and vitamin E significantly inhibited the synthesis of ethylene (ETH) and jasmonic acid (JA) genes, thereby reducing the accumulation of ETH and JA that alleviated the senescence caused by macro-element deficiency at the later stage of bolting in Arabidopsis. A deficiency of macro-elements also reduced the yield and germination rate of the seeds, which were more apparent in vtc1 and vte4, and adding exogenous vitamin C and vitamin E, respectively, could restore them. This study reported, for the first time, that vitamin E is better than vitamin C in delaying seedling senescence caused by macro-element deficiency in Arabidopsis.

Two-factor ANOVA of SSH and RNA-seq analysis reveal development-associated Pi-starvation genes in oilseed rape.

MAIN CONCLUSION: The 5-leaf-stage rape seedlings were more insensitive to Pi starvation than that of the 3-leaf-stage plants, which may be attributed to the higher expression levels of ethylene signaling and sugar-metabolism genes in more mature seedlings. Traditional suppression subtractive hybridization (SSH) and RNA-Seq usually screen out thousands of differentially expressed genes. However, identification of the most important regulators has not been performed to date. Here, we employed two methods, namely, a two-round SSH and two-factor transcriptome analysis derived from the two-factor ANOVA that is commonly used in the statistics, to identify development-associated inorganic phosphate (Pi) starvation-induced genes in Brassica napus. Several of these genes are related to ethylene signaling (such as EIN3, ACO3, ACS8, ERF1A, and ERF2) or sugar metabolism (such as ACC2, GH3, LHCB1.4, XTH4, and SUS2). Although sucrose and ethylene may counteract each other at the biosynthetic level, they may also work synergistically on Pi-starvation-induced gene expression (such as PT1, PT2, RNS1, ACP5, AT4, and IPS1) and root acid phosphatase activation. Furthermore, three new transcription factors that are responsive to Pi starvation were identified: the zinc-finger MYND domain-containing protein 15 (MYND), a Magonashi family protein (MAGO), and a B-box zinc-finger family salt-tolerance protein. This study indicates that the two methods are highly efficient for functional gene screening in non-model organisms.

The interplay of singlet oxygen and ABI4 in plant growth regulation.

Abscisic acid (ABA) and the AP2/ERF (APETALA 2/ETHYLENE-RESPONSIVE FACTOR)-type transcription factor ABA INSENSITIVE 4 (ABI4) control plant growth and development. We review how singlet oxygen, which is produced in chloroplasts of the fluorescent mutant of Arabidopsis thaliana (arabidopsis), and ABI4 may cooperate in transcriptional and translational reprogramming to cause plants to halt growth or demise. Key elements of singlet oxygen- and ABI4-dependent chloroplast-to-nucleus retrograde signaling involve the chloroplast EXECUTER (EX) 1 and EX2 proteins as well as nuclear WRKY transcription factors. Mutants designed to study singlet oxygen signaling, that lack either ABI4 or the EX1 and EX2 proteins, do not show most of the growth effects of singlet oxygen. We propose a model that positions ABI4 downstream of WRKY transcription factors and EX1 and EX2.

Salicylate and glutamate mediate different Cd accumulation and tolerance between Brassica napus and B. juncea.

Most hyperaccumulator plants have little economic values, and therefore have not been widely used in Cd-contaminated soils. Rape species are Cd hyperaccumulators with high economic values. Black mustard seed (Brassica juncea) has a higher accumulation ability and a higher tolerance for Cd than oilseed rape (Brassica napus), but its biomass is relatively low and its geographical distribution is limited. However, it is unknown why B. juncea (Bj) is more tolerant to and accumulates more Cd than B. napus (Bn). Here, we found that the differences in Cd accumulation and tolerance between the two species is mainly because Bj plants have higher levels of salicylic acid and glutamic acid than Bn plants. Exogenous salicylate and glutamate treatments enhanced Cd accumulation (salicylate + glutamate co-treatment doubled Cd accumulation level in Bn seedlings) but reduced oxidative stresses by increasing glutathione biosynthesis and activating phytochelatin-based sequestration of Cd into vacuoles. Our results provide a new idea to simultaneously improve Cd accumulation and Cd tolerance in B. napus.

Shade Avoidance 3 Mediates Crosstalk Between Shade and Nitrogen in Arabidopsis Leaf Development.

After nitrogen treatments, plant leaves become narrower and thicker, and the chlorophyll content increases. However, the molecular mechanisms behind these regulations remain unknown. Here, we found that the changes in leaf width and thickness were largely compromised in the shade avoidance 3 (sav3) mutant. The SAV3 gene encodes an amino-transferase in the auxin biosynthesis pathway. Thus, the crosstalk between shade and nitrogen in Arabidopsis leaf development was investigated. Both hypocotyl elongation and leaf expansion promoted by the shade treatment were reduced by the high-N treatment; high-N-induced leaf narrowing and thickening were reduced by the shade treatment; and all of these developmental changes were largely compromised in the sav3 mutant. Shade treatment promoted SAV3 expression, while high-N treatment repressed SAV3 expression, which then increased or decreased auxin accumulation in cotyledons/leaves, respectively. SAV3 also regulates chlorophyll accumulation and nitrogen assimilation and thus may function as a master switch responsive to multiple environmental stimuli.

Nitrogen and nitric oxide regulate Arabidopsis flowering differently.

Both nitrogen (N) and nitric oxide (NO) postpone plant flowering. However, we still don't know whether N and NO trigger the same signaling pathways leading to flowering delay. Our previous study found that ferredoxin NADP+ oxidoreductase (FNR1) and the blue-light receptor cryptochrome 1 (CRY1) are involved in nitrogen-regulated flowering-time control. However, NO-induced late-flowering does not require FNR1 or CRY1. Sucrose supply counteracts the flowering delay induced by NO. However high-N-induced late-flowering could not be reversed by 5% sucrose supplementation. The high nitrogen condition decreased the amplitudes of all transcripts of the circadian clock. While NO increased the amplitudes of circadian transcripts of CRY1, LHY (LATE ELONGATED HYPOCOTYL), CCA1 (CIRCADIAN CLOCK ASSOCIATED 1) and TOC1 (TIMING OF CAB EXPRESSION 1), but decreased the amplitudes of circadian transcripts of CO (CONSTANS) and GI (GIGANTEA). 5% sucrose supplementation reversed the declines in amplitudes of circadian transcripts of CO and GI after the NO treatment. NO induced S-nitrosation modification on oscillators CO and GI, but not on the other oscillators of the circadian clock. Sucrose supply interestingly reduced S-nitrosation levels of GI and CO proteins. Thus N and NO rely on overlapping but distinct signaling pathways on plant flowering.

Mg-Protoporphyrin IX Signals Enhance Plant's Tolerance to Cold Stress.

The relationship between Mg-protoporphyrin IX (Mg-Proto IX) signals and plant's tolerance to cold stress is investigated. Arabidopsis seedlings grown for 3 weeks were pretreated with 2 mM glutamate (Glu) and 2 mM MgCl2 for 48 h at room temperature to induce Mg-Proto IX accumulation. Then cold stress was performed at 4°C for additional 72 h. Glu + MgCl2 pre-treatments alleviated the subsequent cold stress significantly by rising the leaf temperature through inducing Mg-Proto IX signals. The protective role of Glu + MgCl2 treatment was greatly compromised in the mutants of Mg-Proto IX synthesis, Mg-Proto IX signaling, and cyanide-resistant respiration. And the enhancement of cold-responsive gene expression was greatly compromised in the mutants of Mg-Proto IX synthesis, Mg-Proto IX signaling and ABA signaling, but not in the mutant of cyanide-resistant respiration. Cold stress promoted cyanide-resistant respiration and leaf total respiration exponentially, which could be further induced by the Glu + MgCl2 treatment. Mg-Proto IX signals also activate antioxidant enzymes and increase non-enzymatic antioxidants [glutathione but not ascorbic acid (AsA)] to maintain redox equilibrium during the cold stress.

[Root morphological characteristics of barley genotype with high phosphorus efficiency under phosphorus stress].

A pot experiment was carried out to test the effects of phosphorus (P) supply levels (25, 50, and 75 mg P2O5 . kg-1) with two P genotype (efficient DH110+ and DH147, inefficient DH49) barleys on root morphology and the relationships between root morphology and P uptake. The results showed that barley biomass and P uptake were significantly reduced by low P stress. Efficient genotype barley biomass and P uptake were 1.24-1.70 and 1.18-1.83 times as much as those of inefficient genotype barley respectively. The total root length, total root surface area, average root diameter, adventitious root length and root surface area, lateral root length and root surface area of P efficient genotype barley were significantly reduced with decreasing the P supply level in soil. The total root length, total root surface area, specific root length, lateral root length and surface area of P efficient genotype barley were 1.46-2.06, 1.12-1.51, 1.35-1.72, 1.69-2.42; and 1.40-1.78 times as much as that of those of P inefficient genotype barley, respectively, while the average root diameter was 70.6% - 90.2% of P inefficient genotype barley. Principal component analysis showed that the average root diameter, specific root surface area and specific root length could be used to distinguish two P genotype barleys. Partial least squares regression analysis showed that the total root length, total root surface area made great contributions to P uptake of barley in soil. The contribution of the adventitious root length and surface area on P uptake of barley decreased significantly and the average root diameter, specific root length, lateral root length and root surface area increased with the decreasing P supply level in soil. P efficient genotype barley adapted to low P stress through maintaining the lateral root growth, increasing the specific root length and root fineness.

[Effects of phosphorus sources on phosphorus fractions in rhizosphere soil of wild barley genotypes with high phosphorus utilization efficiency].

High P-efficiency (IS-22-30, IS-22-25) and low P-efficiency (IS-07-07) wild barley cultivars were chosen to evaluate characteristics of phosphorus uptake and utilization, and properties of phosphorus fractions in rhizosphere and non-rhizosphere in a pot experiment with 0 (CK) and 30 mg P · kg(-1) supplied as only Pi (KH2PO4), only Po (phytate) or Pi + Po (KH2PO4+ phytate). The results showed that dry matter and phosphorus accumulation of wild barley in the different treatments was ranked as Pi > Pi + Po > Po > CK. In addition, dry matter yield and phosphorus uptake of wild barley with high P-efficiency exhibited significantly greater than that with low P-efficiency. The concentration of soil available phosphorus was significantly different after application of different phosphorus sources, which was presented as Pi > Pi + Po > Po. The concentration of soil available phosphorus in high P-efficiency wild barley was significantly higher than that of low P-efficiency in the rhizosphere soil. There was a deficit in rhizosphere available phosphorus of high P-efficiency wild barley, especially in Pi and Pi+Po treatments. The inorganic phosphorus fractions increased with the increasing Pi treatment, and the concentrations of inorganic phosphorus fractions in soil were sorted as follows: Ca10-P > O-P > Fe-P > Al-P > Ca2-P > Ca8-P. The contents of Ca2-P and Ca8-P for high P-efficiency wild barley showed deficits in rhizosphere soil under each phosphorus source treatment. In addition, enrichment of Al-P and Fe-P was observed in Pi treatment in rhizosphere soil. The concentrations of organic phosphorus fractions in soil were sorted as follows: moderate labile organic phosphorus > moderate resistant, resistant organic phosphorus > labile organic phosphorus. The labile and moderate labile organic phosphorus enriched in rhizosphere soil and the greatest enrichment appeared in Pi treatment. Furthermore, the concentrations of moderate resistant organic phosphorus and resistant organic phosphorus decreased in rhizosphere soil. The concentrations of labile and moderate labile organic phosphorus in rhizosphere soil of high P-efficiency wild barley were significantly higher than that of low P-efficiency wild barley in each phosphorus source treatment. However, moderate resistant organic phosphorus and resistant organic phosphorus concentrations had no significant difference between the two genotypes. Wild barley with high P-efficiency demonstrated a greater ability of mobilization and uptake Ca2-P, Ca8-P, Al-P and labile organic phosphorus than that with low P-efficiency under Pi deficiency.

[Characteristics of phosphorus uptake and use efficiency of rice with high yield and high phosphorus use efficiency].

A total of twenty seven middle maturing rice varieties as parent materials were divided into four types based on P use efficiency for grain yield in 2011 by field experiment with normal phosphorus (P) application. The rice variety with high yield and high P efficiency was identified by pot experiment with normal and low P applications, and the contribution rates of various P efficiencies to yield were investigated in 2012. There were significant genotype differences in yield and P efficiency of the test materials. GRLu17/AiTTP//Lu17_2 (QR20) was identified as a variety with high yield and high P efficiency, and its yields at the low and normal rates of P application were 1.96 and 1.92 times of that of Yuxiang B, respectively. The contribution rate of P accumulation to yield was greater than that of P grain production efficiency and P harvest index across field and pot experiments. The contribution rates of P accumulation and P grain production efficiency to yield were not significantly different under the normal P condition, whereas obvious differences were observed under the low P condition (66.5% and 26.6%). The minimal contribution to yield was P harvest index (11.8%). Under the normal P condition, the contribution rates of P accumulation to yield and P harvest index were the highest at the jointing-heading stage, which were 93.4% and 85.7%, respectively. In addition, the contribution rate of P accumulation to grain production efficiency was 41.8%. Under the low P condition, the maximal contribution rates of P accumulation to yield and grain production efficiency were observed at the tillering-jointing stage, which were 56.9% and 20.1% respectively. Furthermore, the contribution rate of P accumulation to P harvest index was 16.0%. The yield, P accumulation, and P harvest index of QR20 significantly increased under the normal P condition by 20.6%, 18.1% and 18.2% respectively compared with that in the low P condition. The rank of the contribution rates of P efficiencies to the yield was in order of P uptake efficiency > P utilization efficiency > P transportation efficiency. The greatest contribution rate of P accumulation to the yield was noticed at the jointing-heading stage with the normal P application while it reached the maximal value at the tillering-jointing stage with the low P application. Therefore, these two stages may be the critical periods to coordinate high yield and high P efficiency in rice.