Gurmeen Rakhra

Expression analysis of BsAPase14 acid phosphatase, a stress responsive boiling-stable protein from Triticum aestivum

Developmental regulation and effect of abiotic stresses (osmotic, salt, heat, and drought) on induction of boiling-stable acid phosphatases (BsAPases) was studied in the growing wheat seedlings. SDS-PAGE revealed the induction of several boiling-stable polypeptides in a spatial and temporal manner in the growing seedlings. Western blot analysis, using polyclonal acid phosphatase antiserum, revealed the induction of a cross-reacting one 14 kDa boiling-stable acid phosphatase band (14 kDa, BsAPase14) in endosperm at 7 and 14 days of germination (DOG). Various abiotic stresses (heat, salt, and osmotic) also elicit accumulation of BsAPase14 in a tissue-dependent manner. Imposition of drought stress also resulted in an induction of boiling-stable protein (BsAPase-14) which disappeared after removing the stress. Based upon these observations, the possible role of BsAPase14 in water-stress tolerance is discussed.

Anti-oxidative potential of boiling soluble antioxidant enzymes inAmelioration of drought-induced oxidative stress in tolerant and sensitive cultivars of Triticum aestivum

Drought is one of the most important abiotic stress factors that limit plant growth and productivity but studies documenting how drought stress regulates boiling soluble antioxidants in cultivars differing in drought tolerance has not been critically evaluated. In this study, we analysed the indices of oxidative stress H2O2, O2-, Membrane Injury Index (MII), Membrane Stability Index (MSI) and lipid peroxidation (MDA) and activities of ROS-scavenging boiling soluble antioxidant enzymes in caryopses of sensitive and tolerant cultivars of wheat at different stages of development. Water Content (WC) recorded a decline in both the sensitive cvs. PBW 621 and 343 while in tolerant cvs. PBW 175 and 527, it remained unresponsive to drought conditions at 35 DPA. Oxidative stress indicators increased in a cultivar genotype and developmental stage dependent manner under stress. MII increased in all the cultivars under stress conditions in a stage specific manner. Basal activities of various boiling soluble antioxidants like BsMDAR, BsCAT, BsAPX, and BsGR were significantly higher in caryopses of tolerant cvs. PBW 527 and 175 as compared to sensitive cvs.PBW 343 and 621. Activities of BsMDAR, BsGST, BsPDI, BsGR, BsSOD, BsGPX, and BsAPX were induced or increased in the tolerant cvs. PBW 527 and 175 under drought stress while nil or decreased activities were detected in the sensitive cvs. PBW 343 and 621 at different developmental stages. Based upon our findings, it can be inferred that tolerant cultivars may have more biochemical capacity to perform biological antioxidative reactions to combat drought-induced oxidative stress.

Time-Course Studies on Accumulation of Hydrophilic Antioxidants to Different Stress Regimes Followed by Recovery in Contrasting Cultivars of Wheat

Under unpredictable climatic scenarios, drought is one of the major environmental constraints limiting plant growth and productivity in arid and semi-arid regions. Rapid recovery from drought is of paramount importance for the persistence and survival of different crops growing worldwide. The boiling soluble proteins, BSPs (proteins remaining soluble upon boiling in aqueous solution) forms an instrumental part of the response to water deficit conditions and might be of key importance for the survival of plants under unfavourable environmental conditions. These BSPs are typified by two unique properties: high hydrophilicity and high thermal stability. The main objective of the study was to determine drought-induced changes in the markers of oxidative stress along with modulation in the activity of the boiling soluble antioxidants in response to different stress regimes followed by re-watering in Triticum aestivum L. In this study, we determined the indices of oxidative stress (membrane injury index (MII) and lipid peroxidation in terms of malondialdehyde (MDA) content) and activities of boiling soluble antioxidant enzymes in seeds of sensitive and tolerant cultivars of wheat at different duration of stress (3, 6 and 10 days) followed by recovery (post stress harvest). Water content recorded a decline in the sensitive (PBW 343 and PBW 621) as well as tolerant (PBW 527 and PBW 175) cultivars in stress duration and cultivar dependent manner and this was reversed following re-watering in all the cultivars. Oxidative stress indicators also increased in all the cultivars at different stress intensities but this was reversed following re-watering in the tolerant cvs. PBW 175 and PBW 527. At 3 and 6 days, boiling soluble monodehydroascorbate reductase (BsMDAR), boiling soluble protein disulphide isomerase (BsPDI) activity increased in both the tolerant cvs. PBW 175 and PBW 527 whereas boiling soluble guaiacol peroxidase (BsGPX) increased in the sensitive cv. PBW 343. However, as the stress intensity increased to 10 days, BsMDAR, boiling soluble glutathione-S-transferase (BsGST) and BsGPX increased only in the tolerant cvs. PBW 175 and PBW 527, thus accentuating their cardinal roles in stress tolerance under harsh drought conditions. Upon re-watering the stress plants after 10 days, BsMDAR increased only in the tolerant cv. PBW 175. On the other hand, boiling soluble protein disulphide isomerase (BsPDI) increased in both the tolerant cv. PBW 175 and susceptible cv. PBW 343, but with a greater enhancement in the cv. PBW 175. Based upon our results, biochemical significance of the boiling soluble antioxidants in the cultivars of wheat differing in drought resistance during different stress intensities and recovery is discussed.

Drought-induced changes in the accumulation of boiling-soluble proteins (p40, GST, HSP90) in the grains of drought-tolerant and drought-sensitive cultivars of Triticum aestivum

Abstract. Approximately 70% of crop yield losses are caused by abiotic stresses, with drought being the most serious threat to crop production in many areas of the world. Plants have developed physiological and biochemical responses at multiple levels to allow them to grow and survive under drought stress. Among these, hydrophilins (BSPs, proteins soluble after boiling), representing 0.2% of the total genome, play an important role in the stress adaptation in plants. In this study, we examined the effect of drought on BSPs at different developmental stages of leaves and seeds in drought-tolerant (cv. PBW 175) and drought-susceptible (cv. PBW 621) cultivars of Triticum aestivum. The BSP profiles of seeds were outlined via SDS-PAGE followed by immunoblot analysis using anti-HSP (heat shock protein-90), anti-GST (glutathione S-transferases) and anti-p40 (protein 40). In SDS-PAGE profile, BSPs were detected in a genotype- and treatment-dependent manner. Notably, no BSPs were detected in shoots at any stage, whereas in seeds, many BSPs were detected, indicating organ-specific regulation of BSPs. In western blotting, the induced accumulation of protein bands Bsp40-51 and 59 and presence of differential band of BsHSP44 under drought conditions was observed only in tolerant cv. PBW 175, not in sensitive cv. PBW 621, indicating the roles of such proteins in drought-stress adaptation. BSPs were accumulated at different developmental stages in a cultivar- and stage-dependent manner. The induced expression of different BSPs under drought conditions in tolerant cv. PBW 175 implies the relevance of these BSPs under drought conditions. Notably, the different BSPs were also expressed under normal growth and developmental stages at 57 and 76 days post-anthesis, implying their key role in earlier stages and maturity of grain development.

2-D and In Silico Analysis of Some Putative Drought-Induced Differential Boiling Soluble Proteins (hydrophilins) of Triticum aestivum

The analysis of stress response in plants is an important route to the discovery of genetic elements conferring stress tolerance. Cereal seeds are model system to study desiccation tolerance due to their ability to survive extreme water loss during late embryogenesis. In an attempt to determine the adaptation strategy to drought stress, comparative proteome analysis of boiling-soluble proteins were conducted in wheat cultivar (cv.) having tolerance to drought stress. Evaluation of 2-dimensional electrophoresis gels revealed differential expression of many proteins in drought-tolerant cv. Five differentially expressed hydrophilic protein bands were further analyzed by matrix-assisted laser desorption/ionization-time of flight mass spectrophotometer. By bioinformatics approach, five differential drought-induced hydrophilic proteins were functionally annotated and identified. Based on results, it was argued that elucidating the function of hydrophilic proteins expressed by genes in stress-tolerant plants will not only advance our understanding of plant adaptation but also tolerance to environmental stresses. Based on hydrophilic proteins, which have been identified by proteomics and bioinformatics approach, the possible functions have been discussed.