Guozhang Kang

Proteomics Reveals the Effects of Salicylic Acid on Growth and Tolerance to Subsequent Drought Stress in Wheat

Pretreatment with 0.5 mM salicylic acid (SA) for 3 days significantly enhanced the growth and tolerance to subsequent drought stress (PEG-6000, 15%) in wheat seedlings, manifesting as increased shoot and root dry weights, and decreased lipid peroxidation. Total proteins from wheat leaves exposed to (i) 0.5 mM SA pretreatment, (ii) drought stress, and (iii) 0.5 mM SA treatment plus drought-stress treatments were analyzed using a proteomics method. Eighty-two stress-responsive protein spots showed significant changes, of which 76 were successfully identified by MALDI-TOF-TOF. Analysis of protein expression patterns revealed that proteins associated with signal transduction, stress defense, photosynthesis, carbohydrate metabolism, protein metabolism, and energy production could by involved in SA-induced growth and drought tolerance in wheat seedlings. Furthermore, the SA-responsive protein interaction network revealed 35 key proteins, suggesting that these proteins are critical for SA-induced tolerance.

Exogenous salicylic acid enhances wheat drought tolerance by influence on the expression of genes related to ascorbate-glutathione cycle

Treatment with 0.5 mM salicylic acid (SA) significantly alleviated growth inhibition induced by drought in wheat seedlings, manifested by less decreassed fresh mass, dry mass, plant height, root length, and less increased lipid peroxidation. Under drought stress, SA significantly increased the content of ascorbate (ASA) and glutathione (GSH). We determined the full-length cDNA sequences of genes encoding the glutathione-S-transferase 1 (GST1) and 2 (GST2) and we also measured the transcription of eight genes related to ASA-GSH cycle. The results indicated that exogenous SA significantly enhanced the transcription of GST1, GST2, glutathione reductase (GR), and monodehydroascorbate reductase (MDHAR) genes during almost the entire drought period, but only increased those of dehydroascorbate reductase (DHAR) at 12 h, glutathione peroxidase (GPX1) at 48 h, phospholipid hydroperoxide glutathione peroxidase (GPX2) at 12 and 24 h, and glutathione synthetase (GSHS) at 12, 24, and 48 h. This implies that SA alleviates the detrimental effects of drought stress on wheat seedling growth by influencing the ASA-GSH cycle.

Salicylic acid increases the contents of glutathione and ascorbate and temporally regulates the related gene expression in salt-stressed wheat seedlings

Exogenous salicylic acid (SA) significantly improved abiotic tolerance in higher plants, and ascorbate (ASA) and glutathione (GSH) play important roles in abiotic tolerance. In this study, SA (0.5mM) markedly increased the contents of ASA and GSH in SA-treated plants during salt stress (250mM NaCl). The transcript levels of the genes encoding ASA and GSH cycle enzymes were measured using quantitative real-time PCR. The results indicated that, during salt stress, exogenous SA significantly enhanced the transcripts of glutathione peroxidase (GPX1), phospholipid hydroperoxide glutathione peroxidase (GPX2) and dehydroascorbate reductase (DHAR) genes at 12h, glutathione reductase (GR) at 24h, 48h and 72h, glutathione-S-transferase 1 (GST1), 2 (GST2), monodehydroascorbate reductase (MDHAR) and glutathione synthetase (GS) at the 48h and 72h after salt stress, respectively. The results implied that SA temporally regulated the transcript levels of the genes encoding ASA-GSH cycle enzymes, resulting in the increased contents of GSH and ASA and enhanced salt tolerance.

Proteomic analysis of leaves and roots of common wheat (Triticum aestivum L.) under copper-stress conditions.

Proteomic studies were performed to identify the protein species involved in copper (Cu) stress responses in common wheat. Two-week-old wheat seedlings were exposed to 100 μM CuSO4 treatment for 3 days. Growth of shoots and roots was markedly inhibited and lipid peroxidation was greatly increased. Cu was readily absorbed by wheat seedlings, with greater Cu contents in roots than in leaves. Using 2-DE method, 98 protein spots showed significantly enhanced or reduced abundance, of which 93 were successfully identified. Of these identified protein species, 49 and 44 were found in roots and leaves, respectively. Abundance of most of identified protein species, which function in signal transduction, stress defense, and energy production, was significantly enhanced, while that of many protein species involved in carbohydrate metabolism, protein metabolism, and photosynthesis was severely reduced. The Cu-responsive protein interaction network revealed 36 key proteins, most of which may be regulated by abscisic acid (ABA), ethylene, jasmonic acid (JA), and so on. Exogenous JA application showed a protective effect against Cu stress and significantly increased transcripts of the glutathione S-transferase (GST) gene. This study provides insight into the molecular mechanisms of Cu responses in higher plants.

Molecular mechanism of salicylic acid-induced abiotic stress tolerance in higher plants

Salicylic acid (SA), a key signaling molecule in higher plants, has been found to play a role in the response to a diverse range of phytopathogens and is essential for the establishment of both local and systemic-acquired resistance. Recent studies have indicated that SA also plays an important role in abiotic stress-induced signaling, and studies on SA-modulated abiotic tolerance have mainly focused on the antioxidant capacity of plants by altering the activity of anti-oxidative enzymes. However, little information is available about the molecular mechanisms of SA-induced abiotic stress tolerance. Here, we review recent progress toward characterizing the SA-regulated genes and proteins, the SA signaling pathway, the connections and differences between SA-induced tolerances to biotic and abiotic stresses, and the interaction of SA with other plant hormones under conditions of abiotic stress. The future prospects related to molecular tolerance of SA in response to abiotic stresses are also further summarized.

Proteomic analysis of spring freeze-stress responsive proteins in leaves of bread wheat (Triticum aestivum L.)

Following three-day exposure to -5 °C simulated spring freeze stress, wheat plants at the anther connective tissue formation phase of spike development displayed the drooping and wilting of leaves and markedly increased rates of relative electrolyte leakage. We analysed freeze-stress responsive proteins in wheat leaves at one and three days following freeze-stress exposure, using two-dimensional electrophoresis and matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. Our results indicate that out of 75 protein spots successfully identified under freeze-stress conditions 52 spots were upregulated and 18 were downregulated. These spring freeze-stress responsive proteins were involved in signal transduction, stress/defence/detoxification, protein metabolism (i.e. translation, processing, and degradation), photosynthesis, amino acid metabolism, carbohydrate metabolism, and energy pathways, and may therefore be functionally relevant for many biological processes. The enhanced accumulation of signal transduction proteins such as a C2H2 zinc finger protein, stress/defence/detoxification proteins including LEA-related COR protein, disease resistance protein, Cu/Zn superoxide dismutase, and two ascorbate peroxidases may play crucial roles in the mechanisms of response to spring freeze stress in wheat plants.

Increasing the starch content and grain weight of common wheat by overexpression of the cytosolic AGPase large subunit gene

ADP-glucose pyrophosphorylase (AGPase) catalyzes the first committed step of starch synthesis. AGPase is a heterotetramer composed of two large subunits and two small subunits, has cytosolic and plastidial isoforms, and is detected mainly in the cytosol of endosperm in cereal crops. To investigate the effects of AGPase cytosolic large subunit gene (LSU I) on starch biosynthesis in higher plant, in this study, a TaLSU I gene from wheat was overexpressed under the control of an endosperm-specific promoter in a wheat cultivar (Yumai 34). PCR, Southern blot, and real-time RT-PCR analyses indicated that the transgene was integrated into the genome of transgenic plants and was overexpressed in their progeny. The overexpression of the TaLSU I gene remarkably enhanced AGPase activity, endosperm starch weight, grain number per spike, and single grain weight. Therefore, we conclude that overexpression of the TaLSU I gene enhances the starch biosynthesis in endosperm of wheat grains, having potential applications in wheat breeding to develop a high-yield wheat cultivar with high starch weight and kernel weight.

Proteomic analysis on salicylic acid-induced salt tolerance in common wheat seedlings (Triticum aestivum L.)

The influence of salicylic acid (SA) on the salt tolerance mechanism in seedlings of common wheat (Triticum aestivum L.) was investigated using physiological measurements combined with global expression profiling (proteomics). In the present study, 0.5mM SA significantly reduced NaCl-induced growth inhibition in wheat seedlings, manifesting as increased fresh weights, dry weights, and photosynthetic pigments, but decreased lipid peroxidation. Two-week-old wheat seedlings treated with 0.5mM SA, 250 mM NaCl and 250 mM NaCl+0.5mM SA for 3 days were used for the proteomic analyses. In total, 39 proteins differentially regulated by both salt and SA were revealed by 2D PAGE, and 38 proteins were identified by MALDI-TOF/TOF MS. The identified proteins were involved in various cellular responses and metabolic processes including signal transduction, stress defense, energy, metabolism, photosynthesis, and others of unknown function. All protein spots involved in signal transduction and the defense response were significantly upregulated by SA under salt stress, suggesting that these proteins could play a role in the SA-induced salt resistance in wheat seedlings.

Abscisic acid enhances tolerance of wheat seedlings to drought and regulates transcript levels of genes encoding ascorbate-glutathione biosynthesis

Glutathione (GSH) and ascorbate (ASA) are associated with the abscisic acid (ABA)-induced abiotic tolerance in higher plant, however, its molecular mechanism remains obscure. In this study, exogenous application (10 μM) of ABA significantly increased the tolerance of seedlings of common wheat (Triticum aestivum L.) suffering from 5 days of 15% polyethylene glycol (PEG)-stimulated drought stress, as demonstrated by increased shoot lengths and shoot and root dry weights, while showing decreased content of hydrogen peroxide (H2O2) and malondialdehyde (MDA). Under drought stress conditions, ABA markedly increased content of GSH and ASA in both leaves and roots of ABA-treated plants. Temporal and spatial expression patterns of eight genes encoding ASA and GSH synthesis-related enzymes were measured using quantitative real-time reverse transcription polymerase chain reaction (qPCR). The results showed that ABA temporally regulated the transcript levels of genes encoding ASA-GSH cycle enzymes. Moreover, these genes exhibited differential expression patterns between the root and leaf organs of ABA-treated wheat seedlings during drought stress. These results implied that exogenous ABA increased the levels of GSH and ASA in drought-stressed wheat seedlings in time- and organ-specific manners. Moreover, the transcriptional profiles of ASA-GSH synthesis-related enzyme genes in the leaf tissue were compared between ABA- and salicylic acid (SA)-treated wheat seedlings under PEG-stimulated drought stress, suggesting that they increased the content of ASA and GSH by differentially regulating expression levels of ASA-GSH synthesis enzyme genes. Our results increase our understanding of the molecular mechanism of ABA-induced drought tolerance in higher plants.

Hg-Responsive Proteins Identified in Wheat Seedlings Using iTRAQ Analysis and the Role of ABA in Hg Stress

Wheat seedlings exposed to 100 μM HgCl2 for 3 days exhibited high-level mercury (Hg) accumulation, which led to inhibited growth, increased lipid peroxidation, and disrupted cellular ultrastructures. And root growth and ultrastructural changes of wheat seedlings were inhibited more severely than those of leaves. To identify the wheat protein response to Hg stress, the iTRAQ method was used to determine the proteome profiles of the roots and leaves of wheat seedlings exposed to high-Hg conditions. 249 proteins were identified with significantly altered abundance. 117 were found in roots and 132 in leaves. These proteins were classified into signal transduction, stress defense, carbohydrate metabolism, protein metabolism, energy production, and transport functional groups. The majority of proteins identified in Hg-stressed roots and leaves displayed differently altered abundance, revealing organ-specific differences in adaption to Hg stress. Pathway Studio software was used to identify the Hg-responsive protein interaction network that included 49 putative key proteins, and they were potentially regulated by abscisic acid (ABA). Exogenous ABA application conferred protection against Hg stress and increased activities of peroxidase enzyme, suggesting that it may be an important factor in the Hg signaling pathway. These findings can provide useful insights into the molecular mechanisms of Hg responses in higher plants.