Guohui Zhu

ABA Controls H2O2 Accumulation Through the Induction of OsCATB in Rice Leaves Under Water Stress

The production of both ABA and H₂O₂ is induced by drought and can act as signals under stress conditions. We investigated the relationships between ABA, H₂O₂ and catalase (CAT) in rice leaves when rice seedlings were treated with polyethylene glycol as water stress treatment. As a key gene in ABA biosynthesis, OsNCED3 was significantly induced in rice by water stress treatment and such induction preceded the rapid increase in ABA. Water stress inhibited the expression of CATA and CATC but substantially enhanced the expression of CATB. Exogenously applied ABA promoted the expression of CATB also and inhibited the expression of CATC in a concentration-dependent manner. When ABA production was inhibited by using ABA biosynthesis inhibitors nordihydroguaiaretic acid and tungstate, expression of CATB was also subdued while CATC was enhanced under the water stress. Accumulation of H₂O₂ was also reduced when endogenous ABA production was inhibited and showed a correlation with the total activity of catalases. Our results suggest that water stress-induced ABA prevents the excessive accumulation of H₂O₂, through the induction of the expression of CATB gene during water stress.

Regulation of expression of starch synthesis genes by ethylene and ABA in relation to the development of rice inferior and superior spikelets

Later-flowering spikelets in a rice panicle, referred to as the inferior spikelets, are usually poorly filled and often limit the yield potential of some rice cultivars. The physiological and molecular mechanism for such poor grain filling remains unclear. In this study the differentially expressed genes in starch synthesis and hormone signalling between inferior and superior spikelets were comprehensively analysed and their relationships with grain filling was investigated. DNA microarray and real-time PCR analysis revealed that a group of starch metabolism-related genes showed enhanced expression profiles and had higher transcript levels in superior spikelets than in inferior ones at the early and middle grain-filling stages. Expression of the abscisic acid (ABA) synthesis genes, 9-cis-epoxycarotenoid dioxygenase 1 (NCED1) and NCED5, and the ethylene synthesis genes, 1-aminocyclopropane-1-carboxylate oxidase 1 (ACO1) and ACO3, declined with development of the caryopses. Meanwhile, if compared with inferior spikelets, expression of these genes in superior spikelets decreased faster and had lower transcript profiles, especially for ethylene. ABA concentration and ethylene evolution rate showed similar trends to their gene expression. Exogenous supply of ABA reduced the sucrose synthase (SUS) mRNA level and its enzyme activity in detached rice grains, while exogenously supplied ethephon (an ethylene-releasing reagent) suppressed the expression of most starch synthesis genes; that is, SUS, ADP-glucose pyrophosphorylase (AGPase), and soluble starch synthase (SSS), and down-regulated their enzyme activities. In summary, it is concluded that the relatively high concentrations of ethylene and ABA in inferior spikelets suppress the expression of starch synthesis genes and their enzyme activities and consequently lead to a low grain-filling rate.

Ascorbic acid and reactive oxygen species are involved in the inhibition of seed germination by abscisic acid in rice seeds

The antagonism between abscisic acid (ABA) and gibberellin (GA) plays a key role in controlling seed germination, but the mechanism of antagonism during this process is not known. The possible links among ABA, reactive oxygen species (ROS), ascorbic acid (ASC), and GA during rice seed germination were investigated. Unlike in non-seed tissues where ROS production is increased by ABA, ABA reduced ROS production in imbibed rice seeds, especially in the embryo region. Such reduced ROS also led to an inhibition of ASC production. GA accumulation was also suppressed by a reduced ROS and ASC level, which was indicated by the inhibited expression of GA biosynthesis genes, amylase genes, and enzyme activity. Application of exogenous ASC can partially rescue seed germination from ABA treatment. Production of ASC, which acts as a substrate in GA biosynthesis, was significantly inhibited by lycorine which thus suppressed the accumulation of GA. Consequently, expression of GA biosynthesis genes was suppressed by the low levels of ROS and ASC in ABA-treated seeds. It can be concluded that ABA regulates seed germination in multiple dimensions. ROS and ASC are involved in its inhibition of GA biosynthesis.

Phosphoproteomic analysis of the response of maize leaves to drought, heat and their combination stress

Drought and heat stress, especially their combination, greatly affect crop production. Many studies have described transcriptome, proteome and phosphoproteome changes in response of plants to drought or heat stress. However, the study about the phosphoproteomic changes in response of crops to the combination stress is scare. To understand the mechanism of maize responses to the drought and heat combination stress, phosphoproteomic analysis was performed on maize leaves by using multiplex iTRAQ-based quantitative proteomic and LC-MS/MS methods. Five-leaf-stage maize was subjected to drought, heat or their combination, and the leaves were collected. Globally, heat, drought and the combined stress significantly changed the phosphorylation levels of 172, 149, and 144 phosphopeptides, respectively. These phosphopeptides corresponded to 282 proteins. Among them, 23 only responded to the combined stress and could not be predicted from their responses to single stressors; 30 and 75 only responded to drought and heat, respectively. Notably, 19 proteins were phosphorylated on different sites in response to the single and combination stresses. Of the seven significantly enriched phosphorylation motifs identified, two were common for all stresses, two were common for heat and the combined stress, and one was specific to the combined stress. The signaling pathways in which the phosphoproteins were involved clearly differed among the three stresses. Functional characterization of the phosphoproteins and the pathways identified here could lead to new targets for the enhancement of crop stress tolerance, which will be particularly important in the face of climate change and the increasing prevalence of abiotic stressors.

A Key ABA Catabolic Gene, OsABA8ox3, Is Involved in Drought Stress Resistance in Rice

Expressions of ABA biosynthesis genes and catabolism genes are generally co-regulated in plant development and responses to environmental stress. Up-regulation of OsNCED3 gene, a key gene in ABA biosynthesis, has been suggested as a way to enhance plant drought resistance but little is known for the role of ABA catabolic genes during drought stress. In this study, we found that OsABA8ox3 was the most highly expressed gene of the OsABA8ox family in rice leaves. Expression of OsABA8ox3 was promptly induced by rehydration after PEG-mimic dehydration, a tendency opposite to the changes of ABA level. We therefore constructed rice OsABA8ox3 silencing (RNA interference, RNAi) and overexpression plants. There were no obvious phenotype differences between the transgenic seedlings and wild type under normal condition. However, OsABA8ox3 RNAi lines showed significant improvement in drought stress tolerance while the overexpression seedlings were hypersensitive to drought stress when compared with wild type in terms of plant survival rates after 10 days of unwatering. Enzyme activity analysis indicated that OsABA8ox3 RNAi plants had higher superoxide dismutase (SOD) and catalase (CAT) activities and less malondialdehyde (MDA) content than those of wild type when the plants were exposed to dehydration treatment, indicating a better anti-oxidative stress capability and less membrane damage. DNA microarray and real-time PCR analysis under dehydration treatment revealed that expressions of a group of stress/drought-related genes, i.e. LEA genes, were enhanced with higher transcript levels in OsABA8ox3 RNAi transgenic seedlings. We therefore conclude that that OsABA8ox3 gene plays an important role in controlling ABA level and drought stress resistance in rice.

Copper Suppresses Abscisic Acid Catabolism and Catalase Activity, and Inhibits Seed Germination of Rice

Although copper (Cu) is an essential micronutrient for plants, a slight excess of Cu in soil can be harmful to plants. Unfortunately, Cu contamination is a growing problem all over the world due to human activities, and poses a soil stress to plant development. As one of the most important biological processes, seed germination is sensitive to Cu stress. However, little is known about the mechanism of Cu-induced inhibition of seed germination. In the present study, we investigated the relationship between Cu and ABA which is the predominant regulator of seed germination. Cu at a concentration of 30 µM effectively inhibited germination of rice caryopsis. ABA content in germinating seeds under copper stress was also higher than that under control conditions. Quantitative real-time PCR (qRT-PCR) revealed that Cu treatment reduced the expression of OsABA8ox2, a key gene of ABA catabolism in rice seeds. In addition, both malondialdehyde (MDA) and H2O2 contents were increased by Cu stress in the germinating seeds. Antioxidant enzyme assays revealed that only catalase activity was reduced by excess Cu, which was consistent with the mRNA profile of OsCATa during seed germination under Cu stress. Together, our results demonstrate that suppression of ABA catabolism and catalase (CAT) activity by excess Cu leads to the inhibition of seed germination of rice.

AtDsPTP1 acts as a negative regulator in osmotic stress signalling during Arabidopsis seed germination and seedling establishment

Highlight AtDsPTP1 was found to regulate ABA accumulation and act as a negative regulator in osmotic stress signalling during Arabidospsis seed germination and seedling establishment.

SWATH-MS Quantitative Analysis of Proteins in the Rice Inferior and Superior Spikelets during Grain Filling

Modern rice cultivars have large panicle but their yield potential is often not fully achieved due to poor grain-filling of late-flowering inferior spikelets (IS). Our earlier work suggested a broad transcriptional reprogramming during grain filling and showed a difference in gene expression between IS and earlier-flowering superior spikelets (SS). However, the links between the abundances of transcripts and their corresponding proteins are unclear. In this study, a SWATH-MS (sequential window acquisition of all theoretical spectra-mass spectrometry) -based quantitative proteomic analysis has been applied to investigate SS and IS proteomes. A total of 304 proteins of widely differing functionality were observed to be differentially expressed between IS and SS. Detailed gene ontology analysis indicated that several biological processes including photosynthesis, protein metabolism, and energy metabolism are differentially regulated. Further correlation analysis revealed that abundances of most of the differentially expressed proteins are not correlated to the respective transcript levels, indicating that an extra layer of gene regulation which may exist during rice grain filling. Our findings raised an intriguing possibility that these candidate proteins may be crucial in determining the poor grain-filling of IS. Therefore, we hypothesize that the regulation of proteome changes not only occurs at the transcriptional, but also at the post-transcriptional level, during grain filling in rice.

Characterization of a rice metallothionein type 3 gene with different expression profiles under various nitrogen forms

A cDNA sequence OsMT3 was initially isolated from the subtractive cDNA library of ammonium-fed rice (Oryza sativa L.) leaves, which was further confirmed by Northern blot to be highly ammonium-up-regulated as compared to nitrate. Its full-length cDNA was cloned by RT-PCR, and in silico analysis reveals that the cDNA includes an open reading frame of 186 bp and encodes a rice metallothionein type 3 peptide. Northern blotting analysis showed that OsMT3 gene predominantly expressed in rice leaves, weakly in stems, and barely in buds and roots. The gene transcripts in leaves were significantly induced by polyethylene glycol (PEG), low temperature, NaCl and Cu2+, but not by Pb2+. Activities of three anti-oxidative enzymes (superoxide dismutase, catalase and peroxidases) and two non-enzymic antioxidants (reduced ascorbate and reduced glutathione) little differed in ammonium-and nitrate-fed rice leaves, indicating that the induced OsMT3 expression was not mediated by ammonium-elicited oxidative signals.

Impairment of FtsHi5 Function Affects Cellular Redox Balance and Photorespiratory Metabolism in Arabidopsis

Photorespiration is an essential process for plant photosynthesis, development and growth in aerobic conditions. Recent studies have shown that photorespiration is an open system integrated with the plant primary metabolism network and intracellular redox systems, though the mechanisms of regulating photorespiration are far from clear. Through a forward genetic method, we identified a photorespiratory mutant pr1 (photorespiratory related 1), which produced a chlorotic and smaller photorespiratory growth phenotype with decreased chlorophyll content and accumulation of glycine and serine in ambient air. Morphological and physiological defects in pr1 plants can be largely abolished under elevated CO2 conditions. Genetic mapping and complementation confirmed that PR1 encodes an FtsH (Filamentation temperature-sensitive H)-like protein, FtsHi5. Reduced FtsHi5 expression in DEX-induced RNAi transgenic plants produced a similar growth phenotype with pr1 (ftsHi5-1). Transcriptome analysis suggested a changed expression pattern of redox-related genes and an increased expression of senescence-related genes in DEX: RNAi-FtsHi5 seedlings. Together with the observation that decreased accumulation of D1 and D2 proteins of photosystem II (PSII) and over-accumulation of reactive oxygen species (ROS) in ftsHi5 mutants, we hypothesize that FtsHi5 functions in maintaining the cellular redox balance and thus regulates photorespiratory metabolism.