Heterologous Expression of Serine Hydroxymethyltransferase-3 From Rice Confers Tolerance to Salinity Stress in E. coli and Arabidopsis.

Among abiotic stresses, salt stress adversely affects growth and development in rice. Contrasting salt tolerant (CSR27), and salt sensitive (MI48) rice varieties provided information on an array of genes that may contribute for salt tolerance of rice. Earlier studies on transcriptome and proteome profiling led to the identification of salt stress-induced serine hydroxymethyltransferase-3 (SHMT3) gene. In the present study, the SHMT3 gene was isolated from salt-tolerant (CSR27) rice. OsSHMT3 exhibited salinity-stress induced accentuated and differential expression levels in different tissues of rice. OsSHMT3 was overexpressed in Escherichia coli and assayed for enzymatic activity and modeling protein structure. Further, Arabidopsis transgenic plants overexpressing OsSHMT3 exhibited tolerance toward salt stress. Comparative analyses of OsSHMT3 vis a vis wild type by ionomic, transcriptomic, and metabolic profiling, protein expression and analysis of various traits revealed a pivotal role of OsSHMT3 in conferring tolerance toward salt stress. The gene can further be used in developing gene-based markers for salt stress to be employed in marker assisted breeding programs. HIGHLIGHTS: - The study provides information on mechanistic details of serine hydroxymethyl transferase gene for its salt tolerance in rice.

Identification of cis-regulatory elements associated with salinity and drought stress tolerance in rice from co-expressed gene interaction networks.

Rice, a staple food crop, is often subjected to drought and salinity stresses thereby limiting its yield potential. Since there is a cross talk between these abiotic stresses, identification of common and/or overlapping regulatory elements is pivotal for generating rice cultivars that showed tolerance towards them. Analysis of the gene interaction network (GIN) facilitates identifying the role of individual genes and their interactions with others that constitute important molecular determinants in sensing and signaling cascade governing drought and/or salinity stresses. Identification of the various cis-regulatory elements of the genes constituting GIN is equally important. Here, in this study graphical Gaussian model (GGM) was used for generating GIN for an array of genes that were differentially regulated during salinity and/or drought stresses to contrasting rice cultivars (salt-tolerant [CSR11], salt-sensitive [VSR156], drought-tolerant [Vandana], drought-sensitive [IR64]). Whole genome transcriptom profiling by using microarray were employed in this study. Markov Chain completed co-expression analyses of differentially expressed genes using Dynamic Bayesian Network, Probabilistic Boolean Network and Steady State Analysis. A compact GIN was identified for commonly co-expressed genes during salinity and drought stresses with three major hubs constituted by Myb2 transcription factor (TF), phosphoglycerate kinase and heat shock protein (Hsp). The analysis suggested a pivotal role of these genes in salinity and/or drought stress responses. Further, analysis of cis-regulatory elements (CREs) of commonly differentially expressed genes during salinity and drought stresses revealed the presence of 20 different motifs.

Comparative Proteomic and Nutritional Composition Analysis of Independent Transgenic Pigeon Pea Seeds Harboring cry1AcF and cry2Aa Genes and Their Nontransgenic Counterparts.

Safety assessment of genetically modified plants is an important aspect prior to deregulation. Demonstration of substantial equivalence of the transgenics compared to their nontransgenic counterparts can be performed using different techniques at various molecular levels. The present study is a first-ever comprehensive evaluation of pigeon pea transgenics harboring two independent cry genes, cry2Aa and cry1AcF. The absence of unintended effects in the transgenic seed components was demonstrated by proteome and nutritional composition profiling. Analysis revealed that no significant differences were found in the various nutritional compositional analyses performed. Additionally, 2-DGE-based proteome analysis of the transgenic and nontransgenic seed protein revealed that there were no major changes in the protein profile, although a minor fold change in the expression of a few proteins was observed. Furthermore, the study also demonstrated that neither the integration of T-DNA nor the expression of the cry genes resulted in the production of unintended effects in the form of new toxins or allergens.

Salinity-induced inhibition of growth in the aquatic pteridophyte Azolla microphylla primarily involves inhibition of photosynthetic components and signaling molecules as revealed by proteome analysis.

Salinity stress causes adverse physiological and biochemical changes in the growth and productivity of a plant. Azolla, a symbiotic pteridophyte and potent candidate for biofertilizer due to its nitrogen fixation ability, shows reduced growth and nitrogen fixation during saline stress. To better understand regulatory components involved in salinity-induced physiological changes, in the present study, Azolla microphylla plants were exposed to NaCl (6.74 and 8.61 ds/m) and growth, photochemical reactions of photosynthesis, ion accumulation, and changes in cellular proteome were studied. Maximum dry weight was accumulated in control and untreated plant while a substantial decrease in dry weight was observed in the plants exposed to salinity. Exposure of the organism to different concentrations of salt in hydroponic conditions resulted in differential level of Na+ and K+ ion accumulation. Comparative analysis of salinity-induced proteome changes in A. microphylla revealed 58 salt responsive proteins which were differentially expressed during the salt exposure. Moreover, 42 % spots among differentially expressed proteins were involved in different signaling events. The identified proteins are involved in photosynthesis, energy metabolism, amino acid biosynthesis, protein synthesis, and defense. Downregulation of these key metabolic proteins appears to inhibit the growth of A. microphylla in response to salinity. Altogether, the study revealed that in Azolla, increased salinity primarily affected signaling and photosynthesis that in turn leads to reduced biomass.

Elucidation of salt-tolerance metabolic pathways in contrasting rice genotypes and their segregating progenies.

KEY MESSAGE: Differentially expressed antioxidant enzymes, amino acids and proteins in contrasting rice genotypes, and co-location of their genes in the QTLs mapped using bi-parental population, indicated their role in salt tolerance. Soil salinity is a major environmental constraint limiting rice productivity. Salt-tolerant 'CSR27', salt-sensitive 'MI48'and their extreme tolerant and sensitive recombinant inbred line (RIL) progenies were used for the elucidation of salt stress tolerance metabolic pathways. Salt stress-mediated biochemical and molecular changes were analyzed in the two parents along with bulked-tolerant (BT) and bulked-sensitive (BS) extreme RILs. The tolerant parent and BT RILs suffered much lower reduction in the chlorophyll as compared to their sensitive counterparts. Activities of antioxidant enzymes superoxide dismutase (SOD) and peroxidase (POD) and non-enzymatic antioxidant ascorbic acid were much higher in salt-stressed CSR27 and BT RILs than MI48 and BS RILs. Further, the tolerant lines showed significant enhancement in the levels of amino acids methionine and proline in response to salt stress in comparison to the sensitive lines. Similarly, the tolerant genotypes showed minimal reduction in cysteine content whereas sensitive genotypes showed a sharp reduction. Real time PCR analysis confirmed the induction of methionine biosynthetic pathway (MBP) enzymes cystathionine-ß synthase (CbS), S-adenosyl methionine synthase (SAMS), S-adenosyl methionine decarboxylase (SAMDC) and serine hydroxymethyl transferase (SHMT) genes in tolerant lines, suggesting potential role of the MBP in conferring salt tolerance in rice variety CSR27. Proteome profiling also confirmed higher expression of SOD, POD and plastidic CbS and other proteins in the tolerant lines, whose genes were co-located in the QTL intervals for salt tolerance mapped in the RIL population. The study signifies integrated biochemical-molecular approach for identifying salt tolerance genes for genetic improvement for stress tolerant rice varieties.

Proteomic changes in rice leaves grown under open field high temperature stress conditions.

The interactive effect of temperature with other climatic and soil factors has profound influences on the growth and development of rice. The responses of rice to high temperatures under field conditions are more important than those under the controlled conditions. To understand the genes associated with high temperature stress response in general and tolerance in particular, the expression of all those genes associated with adaptation and tolerance in rice requires proteomic analysis. High temperature stress-tolerant cv. N22 was subjected to 28/18 °C (control) and 42/32 °C (high temperature stress) at flowering stage. The plants were grown in the field under the free air temperature increment condition. The proteomic changes in rice leaves due to high temperature stress were discussed. The proteomes of leaves had about 3000 protein spots, reproducibly detected on 2-dimensional electrophoretic gels with 573 proteins differentially expressed between the control and the high temperature treatments. Putative physiological functions suggested five categories such as growth (15.4%), heat shock proteins (7.7%), regulatory proteins (26.9%), redox homeostasis proteins (11.5%) and energy and metabolism (38.5%) related proteins. The results of the present study suggest that cv. N22, an agronomically recognized temperature tolerant rice cultivar copes with high temperature stress in a complex manner. Several functional proteins play important roles in its responses. The predicted climate change events necessitate more studies using this cultivar under different simulated ecological conditions to identify proteomic changes and the associated genes to be used as biomarkers and to gain a better understanding on the biochemical pathways involved in tolerance.

Deciphering the dynamics of changing proteins of tolerant and intolerant wheat seedlings subjected to heat stress.

Indulgence of heat defense mechanism is crucial to allay undesirable effects by developing significant heat tolerant plants. Translation of heat stress related genes into proteins is a key tolerance strategy tailored by plants. In order to understand the possible mechanisms of heat tolerance in wheat at proteomic level, two wheat genotypes (WH 730-heat tolerant; Raj 4014-heat intolerant) along with their 10 extreme recombinant inbred lines (RILs) were exposed to heat stress (35 °C for 6 h) to identify important stress related proteins. 2-DE coupled with MALDI TOF/TOF of wheat seedlings revealed 14 differentially regulated protein spots. Compared to Raj 4014, 3 proteins viz. Rubisco activase A, Con A and PEP carboxylase 1 were differentially regulated only in WH 730 implying their practical role in heat tolerance. Above and beyond, increased expression of cytochrome b6f complex and catalase in tolerant RIL population signifies their role in accelerated electron flow during heat stress to cope up with the stress. Our results suggests that, compared to intolerant parent and RILs, tolerant parent and RILs might be actively modulating protein involved in photosynthesis, signal transduction and defense which signifies the activation of adaptation mechanism under heat stress.

Nickel and ultraviolet-B stresses induce differential growth and photosynthetic responses in Pisum sativum L. seedlings.

Enhanced level of UV-B radiation and heavy metals in irrigated soils due to anthropogenic activities are deteriorating the environmental conditions necessary for growth and development of plants. The present study was undertaken to study the individual and interactive effects of heavy metal nickel (NiCl(2)·6H(2)O; 0.01, 0.1, 1.0 mM) and UV-B exposure (0.4 W m(-2); 45 min corresponds to 1.08 KJ m(-2)) on growth performance and photosynthetic activity of pea (Pisum sativum L.) seedlings. Ni treatment at high doses (0.1 and 1.0 mM Ni) and UV-B alone reduced chlorophyll content and photosynthetic activity (oxygen yield, carbon fixation, photorespiration, and PSI, PSII, and whole chain electron transport activities), and declining trends continued with combined doses. In contrast to this, Ni at 0.01 mM appeared to be stimulatory for photosynthetic pigments and photosynthetic activity, thereby enhanced biomass was observed at this concentration. However, combined dose (UV-B + 0.01 mM Ni) caused inhibitory effects. Carotenoids showed different responses to each stress. Nickel at high doses strongly inhibited PSII activity and the inhibition was further intensified when chloroplasts were simultaneously exposed to UV-B radiation. PSI activity appeared to be more resistant to each stress. High doses of Ni (0.1 and 1.0 mM) and UV-B alone interrupted electron flow at the oxygen evolving complex. Similar damaging effects were caused by 0.01 and 0.1 mM Ni together with UV-B, but the damage extended to PSII reaction center in case of 1.0 mM Ni in combination with UV-B. In conclusion, the results demonstrate that low dose of Ni stimulated the growth performance of pea seedlings in contrast to its inhibitory role at high doses. However, UV-B alone and together with low as well as high doses of Ni proved to be toxic for P. sativum L.