Leaf Proteome Analysis Research Articles

Inhibition of sulfur assimilation by S-benzyl-L-cysteine: Impacts on growth, photosynthesis, and leaf proteome of maize plants

Sulfur is an essential nutrient for various physiological processes, including protein synthesis and enzyme activation. We aimed to evaluate how S-benzyl-L-cysteine (SBC), an inhibitor of the sulfur assimilation pathway, affects maize plants' growth, photosynthesis, and leaf proteomic profile. Thus, maize plants were grown for 14 days in vermiculite supplemented with SBC. Photosynthesis was assessed using light and CO2 response curves and chlorophyll a fluorescence. Leaf proteome analysis was conducted to evaluate photosynthetic protein biosynthesis, and ROS content was quantified to assess oxidative stress. Applying SBC resulted in a significant decrease in the growth of maize plants. The gas exchange analysis revealed that maize plants exhibited a diminished rate of CO2 assimilation attributable to both stomatal and non-stomatal limitations. Furthermore, SBC suppressed the activity of important elements involved in the photosynthetic electron transport chain (including photosystems I and II, cytochrome b6f, and ATP synthase) and enzymes responsible for the Calvin cycle, some of which have sulfur-containing prosthetic groups. Consequently, the diminished electron flow rate resulted in a substantial increase in the levels of ROS within the leaves. Our research highlights the crucial role of SBC in disrupting maize photosynthesis by limiting L-cysteine and assimilated sulfur availability, which are essential for the synthesis of protein and prosthetic groups and photosynthetic processes, emphasizing the potential of OAS-TL as a new herbicide site of action.

Leaf Proteomic Analysis in Seedlings of Two Maize Landraces with Different Tolerance to Boron Toxicity.

Boron (B) toxicity is an important stressor that negatively affects maize yield and the quality of the produce. The excessive B content in agricultural lands is a growing problem due to the increase in arid and semi-arid areas because of climate change. Recently, two Peruvian maize landraces, Sama and Pachía, were physiologically characterized based on their tolerance to B toxicity, the former being more tolerant to B excess than Pachía. However, many aspects regarding the molecular mechanisms of these two maize landraces against B toxicity are still unknown. In this study, a leaf proteomic analysis of Sama and Pachía was performed. Out of a total of 2793 proteins identified, only 303 proteins were differentially accumulated. Functional analysis indicated that many of these proteins are involved in transcription and translation processes, amino acid metabolism, photosynthesis, carbohydrate metabolism, protein degradation, and protein stabilization and folding. Compared to Sama, Pachía had a higher number of differentially expressed proteins related to protein degradation, and transcription and translation processes under B toxicity conditions, which might reflect the greater protein damage caused by B toxicity in Pachía. Our results suggest that the higher tolerance to B toxicity of Sama can be attributed to more stable photosynthesis, which can prevent damage caused by stromal over-reduction under this stress condition.

Comparative protein analysis of two maize genotypes with contrasting tolerance to low temperature

BackgroundLow temperature (LT) stress is one of the major environmental stress factors affecting the growth and yield of maize (Zea mays L.). Hence, it is important to unravel the molecular mechanisms behind LT stress tolerance to improve molecular breeding in LT tolerant genotypes. In the present study, two maize genotypes viz. Gurez local from Kashmir Himalaya and tropical grown GM6, were dissected for their LT stress response in terms of accumulation of differentially regulated proteins (DRPs). Leaf proteome analysis at three-leaf stage of maize seedlings subjected to LT stress of 6 °C for a total of 12 h duration was performed using two dimensional gel electrophoresis (2D-PAGE) followed by subsequent identification of the proteins involved.ResultsAfter MALDI-TOF (Matrix-assisted laser desorption/ionization-time of flight) and bioinformatics analysis, 19 proteins were successfully identified in Gurez local, while as 10 proteins were found to get successful identification in GM6. The interesting observations from the present investigation is the identification of three novel proteins viz. threonine dehydratase biosynthetic chloroplastic, thylakoidal processing peptidase 1 chloroplastic, and nodulin-like protein, whose role in abiotic stress tolerance, in general, and LT stress, in particular, has not been reported so far. It is important to highlight here that most of LT responsive proteins including the three novel proteins were identified from Gurez local only, owing to its exceptional LT tolerance. From the protein profiles, obtained in both genotypes immediately after LT stress perception, it was inferred that stress responsive protein accumulation and their expression fashion help the Gurez local in seedling establishment and withstand unfavorable conditions as compared to GM6. This was inferred from the findings of pathway enrichment analysis like regulation of seed growth, timing of floral transition, lipid glycosylation, and aspartate family amino acid catabolic processes, besides other key stress defense mechanisms. However, in GM6, metabolic pathways enriched were found to be involved in more general processes including cell cycle DNA replication and regulation of phenylpropanoid metabolism. Furthermore, majority of the qRT-PCR results of the selected proteins demonstrated positive correlation between protein levels and transcript abundance, thereby strengthening our findings.ConclusionsIn conclusion, our findings reported majority of the identified proteins in Gurez local exhibiting up-regulated pattern under LT stress as compared to GM6. Furthermore, three novel proteins induced by LT stress were found in Gurez local, requiring further functional validation. Therefore, our results offer more insights for elucidating the molecular networks mediating LT stress tolerance in maize.

Defense mechanisms promoting tolerance to aggressive Phytophthora species in hybrid poplar

Poplars are among the fastest-growing trees and significant resources in agriculture and forestry. However, rapid growth requires a large water consumption, and irrigation water provides a natural means for pathogen spread. That includes members of Phytophthora spp. that have proven to be a global enemy to forests. With the known adaptability to new hosts, it is only a matter of time for more aggressive Phytophthora species to become a threat to poplar forests and plantations. Here, the effects of artificial inoculation with two different representatives of aggressive species (P. cactorum and P. plurivora) were analyzed in the proteome of the Phytophthora-tolerant hybrid poplar clone T-14 [Populus tremula L. 70 × (Populus × canescens (Ait.) Sm. 23)]. Wood microcore samples were collected at the active necrosis borders to provide insight into the molecular processes underlying the observed tolerance to Phytophthora. The analysis revealed the impact of Phytophthora on poplar primary and secondary metabolism, including carbohydrate-active enzymes, amino acid biosynthesis, phenolic metabolism, and lipid metabolism, all of which were confirmed by consecutive metabolome and lipidome profiling. Modulations of enzymes indicating systemic response were confirmed by the analysis of leaf proteome, and sampling of wood microcores in distal locations revealed proteins with abundance correlating with proximity to the infection, including germin-like proteins, components of proteosynthesis, glutamate carboxypeptidase, and an enzyme that likely promotes anthocyanin stability. Finally, the identified Phytophthora-responsive proteins were compared to those previously found in trees with compromised defense against Phytophthora, namely, Quercus spp. and Castanea sativa. That provided a subset of candidate markers of Phytophthora tolerance, including certain ribosomal proteins, auxin metabolism enzymes, dioxygenases, polyphenol oxidases, trehalose-phosphate synthase, mannose-1-phosphate guanylyltransferase, and rhamnose biosynthetic enzymes. In summary, this analysis provided the first insight into the molecular mechanisms of hybrid poplar defense against Phytophthora and identified prospective targets for improving Phytophthora tolerance in trees.

Metabolomic and proteomic signature of Gloriosa superba leaves treated with mercuric chloride and phenylalanine, a precursor of colchicine alkaloid

Gloriosa superba is a tropical, medicinally important plant used in the treatment of gout, rheumatism, and other ailments. It produces pharmaceutically important alkaloids like colchicine, gloriosine, thiocolchicoside and others. In the present study, gas chromatography (GC)-mass spectrometry (MS) method has been deployed for the identification of low abundance phytochemicals in mercuric chloride (elicitor)-treated leaves. The analysis revealed nearly 500 molecules including 15 key secondary metabolites like estragole, N-methylloline (alkaloid), aphidocolin, 3-hydroxykynurenine, octyl salicylate, butibufen, anonaine (aporhine alkaloid), bolasterone, austricin, bolandione, octahydrocoumarin, jacaranone, bonducellin, quinacridone, and β-carotene that may have medicinal importance. Liquid chromatography-mass spectrometry (LC-MS) analysis of leaf proteome in the control and phenylalanine (a precursor of colchicine)-treated tissues revealed a total of 982 and 937 proteins respectively. In precursor-treated tissues, 364 differentially expressed proteins were noticed besides others. Key proteins involved in shikimate/chorismate pathway such as phenylalanine ammonia-lyase, chalcone-flavone isomerase (associated with flavonoid biosynthesis), chalcone synthase (involved in the synthesis of bioactive compounds in plants), chorismate synthetase, chorismate mutase, tryptophan synthase, and medium chain triglyceride protein were detected. Importantly, detection of nearly 154 proteins with unknown functions may hold key and play a role in colchicine biosynthetic pathway. These studies suggest that while metabolomic studies help to detect new secondary plant products, proteomic studies assist us in identifying key enzymes implicated in the biosynthetic pathway of alkaloids in G. superba.

Light Regulates the Cytokinin-Dependent Cold Stress Responses in Arabidopsis.

To elucidate the effect of light intensity on the cold response (5°C; 7 days) in Arabidopsis thaliana, we compared the following parameters under standard light (150 μmol m–2 s–1), low light (20 μmol m–2 s–1), and dark conditions: membrane damage, photosynthetic parameters, cytokinin oxidase/dehydrogenase (CKX) activity, phytohormone levels, and transcription of selected stress- and hormone-related genes and proteome. The impact of cytokinins (CKs), hormones directly interacting with the light signaling pathway, on cold responses was evaluated using transformants overexpressing CK biosynthetic gene isopentenyl transferase (DEX:IPT) or CK degradation gene HvCKX2 (DEX:CKX) under a dexamethasone-inducible promoter. In wild-type plants, cold treatment under light conditions caused down-regulation of CKs (in shoots) and auxin, while abscisic acid (ABA), jasmonates, and salicylic acid (SA) were up-regulated, especially under low light. Cold treatment in the dark strongly suppressed all phytohormones, except ABA. DEX:IPT plants showed enhanced stress tolerance associated with elevated CK and SA levels in shoots and auxin in apices. Contrarily, DEX:CKX plants had weaker stress tolerance accompanied by lowered levels of CKs and auxins. Nevertheless, cold substantially diminished the impact from the inserted genes. Cold stress in dark minimized differences among the genotypes. Cold treatments in light strongly up-regulated stress marker genes RD29A, especially in roots, and CBF1-3 in shoots. Under control conditions, their levels were higher in DEX:CKX plants, but after 7-day stress, DEX:IPT plants exhibited the highest transcription. Transcription of genes related to CK metabolism and signaling showed a tendency to re-establish, at least partially, CK homeostasis in both transformants. Up-regulation of strigolactone-related genes in apices and leaves indicated their role in suppressing shoot growth. The analysis of leaf proteome revealed over 20,000 peptides, representing 3,800 proteins and 2,212 protein families (data available via ProteomeXchange, identifier PXD020480). Cold stress induced proteins involved in ABA and jasmonate metabolism, antioxidant enzymes, and enzymes of flavonoid and glucosinolate biosynthesis. DEX:IPT plants up-regulated phospholipase D and MAP-kinase 4. Cold stress response at the proteome level was similar in all genotypes under optimal light intensity, differing significantly under low light. The data characterized the decisive effect of light–CK cross-talk in the regulation of cold stress responses.

Analysis of Proteomic Profile of Contrasting Phosphorus Responsive Rice Cultivars Grown under Phosphorus Deficiency

Phosphorus (P) deficiency is one of the major limiting factors for crop productivity. The yield of rice (Oryza sativa L.) is severely limited by phosphorus deficiency. An attempt has been made in this study to identify P deficiency responsive differentially expressed proteins of rice through analysis of leaf proteome of contrasting P-responsive rice cultivars under P deficiency conditions because genetic variability has been found in the rice cultivars for adaptive response to P deficiency and a controlled regulatory system is involved in the P deficiency adaptation response. Phosphorus-efficient (cv. Panvel) and P-inefficient (cv. Nagina 22) rice cultivars were hydroponically grown in the nutrient medium under control environmental conditions at low-P level (2.0 µM) and optimum-P level (320 µM) treatments. Expression patterns of the proteins of the leaves of both the cultivars were analyzed in 30-day-old plants. The identification of these proteins through mass spectrometry and MASCOT software (Matrix Science Inc., Boston, USA) revealed that these differentially expressed proteins were homologous to known functional proteins involved in energy metabolism, biosynthesis, photosynthesis, signaling, protein synthesis, protein folding, phospholipid metabolism, oxidative stress, transcription factors, and phosphorus metabolism. It has been observed that rice cultivars responded differently to low-P treatment through modification in protein expressions pattern to maintain the growth of the plants. Therefore, the expression patterns of proteins were different in both of the cultivars under low-P treatment. Higher potential of protein stability, stress tolerance, osmo-protection, and regulation of phosphorus uptake was observed in cv. Panvel than cv. Nagina 22. This study could help to unravel the complex regulatory process that is involved in adaptation to P deficiency in rice.

Protein hydrolysates modulate leaf proteome and metabolome in water-stressed grapevines

Biostimulants are innovative tools for the modern vineyard management, especially in relation to the new challenges posed by climate change. These products can modulate vine physiology and improve biotic/abiotic stress tolerance, but their mechanisms of action are still unclear. The present work aims at investigating the effects of two vegetal-derived protein hydrolysates on water-stressed vines, clarifying the involved mechanisms of action.Potted vines of Vitis vinifera L. cv. Montepulciano were subjected to a progressive water-stress. Vines sprayed pre-veraison with ‘Trainer’ (PH1) and ‘Stimtide’ (PH2) were compared with untreated vines. Leaves were sampled to perform non-target analysis of leaf proteome and metabolome. Gas exchanges performances were measured after re-watering and, at harvest, vine productivity and fruit composition were evaluated.PH1 significantly up-regulated metabolites involved in plant growth regulations and promoted the delay of senescence. At leaf Ψ= −1.4 MPa, both biostimulants up-regulated 6 and 7 photosynthesis-related proteins, respectively. However, 5 days past re-watering, no effects were found on assimilation rates, but leaf transpiration was improved by both treatments, as compared to control. The two formulates did not change vine yield and bunch morphology, but significantly reduced grapes total soluble solids (−3.5°Brix) and pH (−6%). PH2 significantly increased grapes TA (+1.87 g/L). The two biostimulants did not significantly affect the grapes total anthocyanins or their phenolic profile.In conclusion, PH1 and PH2 produced deep changes in leaf proteome and metabolome that delayed physiological maturity and maintained higher acidity, with no negative effects on anthocyanins and phenolics. Considering the negative effects of warming trends, today affecting most of traditional and modern wine regions, protein hydrolysates can be considered valuable tools to improve fruit quality and vineyard sustainability.

Leaf proteome pattern of two bread wheat varieties under water deficit stress conditions

This investigation was carried out to study the effect of water deficit stress on leaf proteome pattern and agronomic and physiological characteristics of two spring bread wheat varieties (Arg and Moghan3) with contrasting responses to drought stress. For this purpose, an experiment was conducted as a 2 × 2 factorial arranged as the randomized complete block design with five replications in a greenhouse of University of Tabriz, Iran. The factors consisted of two Arg and Moghan3 varieties and irrigation conditions (normal condition and drought stress, where water was withheld at the pollination stage). Based on the analysis of variance, significant differences were observed between two wheat varieties with respect to all studied characters except leaf chlorophyll content. Drought stress increased leaf temperature and reduced other characters. Leaf proteome analysis was performed by two-dimensional electrophoresis for both wheat varieties at two irrigation conditions. After quantification of spots with PDQuest software, 274 and 202 repeatable protein spots were identified in Arg and Moghan3 varieties, respectively. Among them, based on the induction factor and t-test, 73 and 64 protein spots showed significant changes in abundance in Arg and Moghan3 varieties, respectively. Out of 73 spots identified in Arg, 46 showed an increased expression and 27 showed a decreased expression. In Moghan3, out of 64 protein spots, 18 and 46 protein spots were up- and down-regulated, respectively. Moreover, 15 common protein spots in Arg and Moghan3 with significant changes in expression between normal and drought stress conditions were identified by applying MALDI-TOF/TOF mass spectrometry. Identified proteins were assigned to seven functional categories: photosynthesis, stress, defense and detoxification, glycolysis, transport, protein synthesis and transcription regulation. In general, proteins related to photosynthesis, stress, and defense and detoxification showed major changes which may be considered as important proteins contributing to drought tolerance of Arg in this study.

ITRAQ-based comparative proteomic analysis of two coconut varieties reveals aromatic coconut cold-sensitive in response to low temperature

Coconut (Cocos nucifera L.) is an important economic fruit and oil crop largely cultivated in humid and sub-humid tropical coastal zones worldwide. To date proteomic profile analysis of coconut under cold stress yet not been conducted. In order to understand the cold stress tolerance in coconut, the iTRAQ approach was employed to dissect proteomic response of two coconut varieties Hainan Tall, BenDi (BD) and Aromatic coconut, XiangShui (XS) under cold stress. Under cold treatment at (8 °C) for 2 days, 193 up and 134 down-regulated in BD (Cn-DB-0_VS_Cn-DB-2) and 140 up and 155 down-regulated DEPs in XS (Cn-XS-0_VS_Cn-XS-2) were identified. The 5 days post cold treatment also identified increased abundance of up-regulated proteins in BD compared to XS. The 5 days post treatment (dpt) depicted 172-up/127-down and 108-up/134-down accumulated proteins for BD (Cn-DB-0_VS_Cn-DB-5) and XS (Cn-XS-0_VS_Cn-XS-5) respectively. A total of 22, 12 and 14 DEP categories were enriched in biological process, cellular component and molecular function respectively in Gene Ontology (GO) analysis of two coconut varieties. Metabolic and biosynthesis of secondary metabolites pathways were highly enriched in KEGG pathway analysis of DEPs between two varieties. Twenty-two different functional classes revealed differentially expressed proteins in two varieties. Among those, four major categories involved in metabolism, stress response, photosynthesis and respiration related DEPs increased abundance in two varieties. However, general function perdition only (GFPO) and stress-responsive proteins were greatly up-regulated in BD than XS. Increased abundance of stress response related proteins up-regulation under cold stress suggested that BD is cold-tolerant variety. Collectively, iTRAQ-based coconut leaf proteomic analysis showed that XS (aromatic) coconut variety is cold-sensitive compared to BD (Hainan Tall) variety. This study provided a basis for further functional analyses to understand the molecular mechanisms of tropical crops adapting to cold stress. SignificanceLeaf proteomic approach determines the role of differentially expressed proteins (DEPs) under cold stress in crops. However, cold stress could damage the coconut fruit lead to decrease in crop yield during winter in China. Here, we report the first ever iTRAQ-based proteomic analysis of two coconut varieties in response to cold stress. The study identified the proteins involved in biosynthesis of secondary metabolites, photosynthesis, respiration, biotic and abiotic stresses under cold stress in two coconut varieties. Moreover, the increased abundance of stress-responsive and general function proteins in BD under cold stress suggested that Hainan Tall is cold-tolerant compared to aromatic coconut variety. Inhibition abundance of photosynthesis related proteins may reduce photodamage owing to the over energized state of thylakoid membrane lead to ROS generation during oxidative stress. This could be the reason for adaption of BD to low temperature stress. Nonetheless, further research may insight the mechanism involved in cold tolerance/sensitive in coconut in response to low temperature.

Elevated CO2 and temperature influence key proteins and metabolites associated with photosynthesis, antioxidant and carbon metabolism in Picrorhiza kurroa

Global warming is currently one of the most serious issues in ecology. Rising CO2 level and temperature have begun to impact life cycles, distribution and yield of various plants yet, how medicinal plants will respond to changing environment is largely unknown. Picrorhiza kurroa Royle ex Benth. (Plantaginaceae) is a medicinal plant species that has been used for treatment of various diseases, particularly hepatic disease. Here, we have performed leaf and rhizome specific proteomic and metabolomic analysis to investigate the effect of elevated CO2 and temperature on adaptive responses of P. kurroa. We observed differentially abundant proteins related to photosynthesis and carbon metabolism under free air carbon dioxide enhancement, whereas cytoskeleton proteins in free air temperature increase besides signaling, antioxidant, stress-responsive and chromatin remodeling proteins in both conditions. We also found an increased accumulation of metabolites, particularly picroside-I and picroside-II, sugars and sugar alcohol in rhizomes, whereas, decrease in picroside-I and increase in picroside-II content in leaves at FACE condition. Biochemical indices like total protein, phenolics, flavonoids and antioxidant activity were altered in a tissue-specific manner to elevated CO2 and temperature. The results would provide new insights into possible adaptive mechanism, particularly in P. kurroa and medicinal plants in general. SignificancePicrorhiza kurroa is an endangered medicinal plant of Himalayan region having immense medicinal values due to the presence of iridoid glycosides. This endangered plant species is particularly, more vulnerable to climate change. P. kurroa provides an opportunity to investigate the interaction between high altitude plant and their environment. Therefore, it will be essential to elucidate the adaptation mechanism at molecular level under e[CO2] and e[t] conditions. The e[CO2] and e[t] will likely alter the proteome and metabolite composition of plant and thereby, enhance plant adaptation. Proteomic and metabolomic studies would facilitate to explore the adaptive mechanism of P. kurroa which is poorly understood. Collectively, the findings will be helpful for better understanding of plant response to future CO2 and temperature enriched environment and are of key importance to agriculture and ecosystem.

Comparative proteomics analysis of two maize hybrids revealed drought-stress tolerance mechanisms

Drought is a major abiotic stress factor that affects the yield of maize (Zea mays L.) during the grain filling period. Therefore, it is critical to reveal the molecular mechanisms underlying maize drought stress tolerance. To analyze the characteristics and functions of differentially abundant proteins (DAPs) in response to drought stress during the maize kernel-filling stage, here, two maize hybrid varieties with contrasting drought tolerance (tolerant ND476 and sensitive ZX978) were subjected to water sufficient (control) and water deficit (drought) treatment conditions for 12 days at the grain filling stage. Physiological assays and leaf proteome analysis were performed. We conducted iTRAQ analysis on 12 samples from 4 groups and obtained 1,655 DAPs. Amongst these, five essential sets of drought responsive DAPs were identified by bioinformatics techniques. Four significantly enriched metabolic pathways were identified in ND476, which were shown to be associated with the ribosome, metabolic, basal transcription factors and photosynthesis pathways. Further, the phenotypic characterization and qRT-PCR analysis confirmed our iTRAQ sequencing data. Remarkably, significantly higher drought tolerance of ND476 may be attributed to the following responses: (a) oxidoreductase, peroxidase and hydrolytic enzyme activities to promote cell redox homeostasis maintenance; (b) elevated expression of stress defense proteins; and (c) reduced synthesis of redundant proteins in order to help plants preserve energy to fight drought stress. In conclusion, these results offer more insights into the molecular mechanisms underlying maize drought tolerance at the grain filling stage and can be a foundational basis for molecular breeding of drought tolerant maize varieties.

Regulation of the leaf proteome by inoculation of Populus \xd7 canescens with two Paxillus involutus isolates differing in root colonization rates

During ectomycorrhizal symbioses, up to 30% of the carbon produced in leaves may be translocated to the fungal partner. Given that the leaf response to root colonization is largely unknown, we performed a leaf proteome analysis of Populus × canescens inoculated in vitro with two isolates of Paxillus involutus significantly differing in root colonization rates (65 ± 7% vs 14 ± 7%), together with plant growth and leaf biochemistry analyses to determine the response of plant leaves to ectomycorrhizal root colonization. The isolate that more efficiently colonized roots (isolate H) affected 9.1% of the leaf proteome compared with control plants. Simultaneously, ectomycorrhiza in isolate H-inoculated plants led to improved plant growth and an increased abundance of leaf proteins involved in protein turnover, stress response, carbohydrate metabolism, and photosynthesis. The protein increment was also correlated with increases in chlorophyll, foliar carbon, and carbohydrate contents. Although inoculation of P. × canescens roots with the other P. involutus isolate (isolate L, characterized by a low root colonization ratio) affected 6.8% of the leaf proteome compared with control plants, most proteins were downregulated. The proteomic signals of increased carbohydrate biosynthesis were not detected, and carbohydrate, carbon, and leaf pigment levels and plant biomass did not differ from the noninoculated plants. Our results revealed that the upregulation of the photosynthetic protein abundance and levels of leaf carbohydrate are positively related to rates of root colonization. Upregulation of photosynthetic proteins, chlorophyll, and leaf carbohydrate levels in ectomycorrhizal plants was positively related to root colonization rates and resulted in increased carbon translocation and sequestration underground.

Leaf proteome analysis signified that photosynthesis and antioxidants are key indicators of salinity tolerance in canola (Brassica napus L.)

Leaf proteome analysis signified that photosynthesis and antioxidants are key indicators of salinity tolerance in canola (Brassica napus L.)

Comparative physiological and leaf proteome analysis between drought-tolerant chickpea Cicer reticulatum and drought-sensitive chickpea C. arietinum

Comparative physiological and proteomic analysis were performed to understand the stress responses of two chickpea species (C. reticulatum and C. arietinum) against drought. Our study revealed that drought stress reduced root length, leaf water content, and enhanced free proline content in both species. Effect of drought stress appeared to be greater in C. arietinum compared to C. reticulatum. A total of 24 differently expressed proteins were identified by using MALDI-TOF/ TOF-MS/MS in response to drought. The proteins involved in photosynthesis and energy mechanisms were up-regulated in C. reticulatum and down-regulated in C. arietinum under drought. Our results suggest that the photosynthesis capacity of C. reticulatum is greater than that of C. arietinum under drought stress. Abundance of proline and sucrose biosynthesis related proteins, glutamine synthetase and cyctosolic fructose-bisphosphate aldolase, respectively, also increased in C. reticulatum under drought stress. The findings of this proteome analysis will help in understanding the mechanism of drought resistance in chickpea and may be also helpful in developing drought-resistant transgenic plants.

ITRAQ-based analysis of leaf proteome identifies important proteins in secondary metabolite biosynthesis and defence pathways crucial to cross-protection against TMV

Cross-protection is a phenomenon in which infection with a mild virus strain protects host plants against subsequent infection with a closely related severe virus strain. This study showed that a mild strain mutant virus, Tobacco mosaic virus (TMV)-43A could cross protect Nicotiana benthamiana plants against wild-type TMV. Furthermore, we investigated the host responses at the proteome level to identify important host proteins involved in cross-protection. We used the isobaric tags for relative and absolute quantification (iTRAQ) technique to analyze the proteome profiles of TMV, TMV-43A and cross-protected plants at different time-points. Our results showed that TMV-43A can cross-protect N. benthamiana plants from TMV. In cross-protected plants, photosynthetic activities were augmented, as supported by the increased accumulation of 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) and geranylgeranyl diphosphate synthase (GGPS) enzymes, which are crucial for chlorophyll biosynthesis. The increased abundance of ROS scavenging enzymes like thioredoxins and L-ascorbate peroxidase would prevent oxidative damage in cross-protected plants. Interestingly, the abundance of defence-related proteins (14–3-3 and NbSGT1) decreased, along with a reduction in virus accumulation during cross-protection. In conclusion, we have identified several important host proteins that are crucial in cross-protection to counter TMV infection in N. benthamiana plants. Biological significanceTMV is the most studied model for host-virus interaction in plants. It can infect wide varieties of plant species, causing significant economic losses. Cross protection is one of the methods to combat virus infection. A few cross-protection mechanisms have been proposed, including replicase/coat protein-mediated resistance, RNA silencing, and exclusion/spatial separation between virus strains. However, knowledge on host responses at the proteome level during cross protection is limited. To address this knowledge gap, we have leveraged on a global proteomics analysis approach to study cross protection. We discovered that TMV-43A (protector) protects N. benthamiana plants from TMV (challenger) infection through multiple host pathways: secondary metabolite biosynthesis, photosynthesis, defence, carbon metabolism, protein translation and processing and amino acid biosynthesis. In the secondary metabolite biosynthesis pathway, enzymes 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) and geranylgeranyl diphosphate synthase (GGPS) play crucial roles in chlorophyll biosynthesis during cross protection. In addition, accumulation of ROS scavenging enzymes was also found in cross-protected plants, providing rescues from excessive oxidative damage. Reduced abundance of plant defence proteins is correlated to reduced virus accumulation in host plants. These findings have increased our knowledge in host responses during cross-protection.

Soybean (Glycine max L. Merr.) seedlings response to shading: leaf structure, photosynthesis and proteomic analysis

BackgroundIntercropping and close planting are important cultivation methods that increase soybean yield in agricultural production. However, plant shading is a major abiotic stress factor that influences soybean growth and development. Although shade affects leaf morphological parameters and decreases leaf photosynthesis capacity, information on the responses of soybean leaf photosynthesis to shading at proteomic level is still lacking.ResultsCompared with leaves under normal light (CK) treatment, leaves under shading treatment exhibited decreased palisade and spongy tissue thicknesses but significantly increased cell gap. Although shade increased the number of the chloroplast, the thickness of the grana lamella and the photosynthetic pigments per unit mass, but the size of the chloroplast and starch grains and the rate of net photosynthesis decreased compared with those of under CK treatment. A total of 248 differentially expressed proteins, among which 138 were upregulated, and 110 were downregulated, in soybean leaves under shading and CK treatments were detected via isobaric tags for relative and absolute quantification labeling in the three biological repeats. Differentially expressed proteins were classified into 3 large and 20 small groups. Most proteins involved in porphyrin and chlorophyll metabolism, photosynthesis-antenna proteins and carbon fixation in photosynthetic organisms were upregulated. By contrast, proteins involved in photosynthesis were downregulated. The gene family members corresponding to differentially expressed proteins, including protochlorophyllide reductase (Glyma06g247100), geranylgeranyl hydrogenase (Ggh), LHCB1 (Lhcb1) and ferredoxin (N/A) involved in the porphyrin and chlorophyll metabolism, photosynthesis-antenna proteins and photosynthesis pathway were verified with real-time qPCR. The results showed that the expression patterns of the genes were consistent with the expression patterns of the corresponding proteins.ConclusionsThis study combined the variation of the soybean leaf structure and differentially expressed proteins of soybean leaves under shading. These results demonstrated that shade condition increased the light capture efficiency of photosystem II (PSII) in soybean leaves but decreased the capacity from PSII transmitted to photosystem II (PSI). This maybe the major reason that the photosynthetic capacity was decreased in shading.

Response of Lablab purpureus L. to high temperature stress and role of exogenous protectants in mitigating high temperature induced oxidative damages.

Present study was conducted to explore the role of exogenous salicylic acid (SA), sodium nitroprusside (SNP), abscisic acid (ABA) and proline (PRO) in mitigating high-temperature (HT) induced oxidative stress in different Lablab purpureus L. cultivars. The attempt was made to examine whether these phytohormones, when applied exogenously, were able to regulate plant morpho-physiological behavior by modulating genes and proteins involved in antioxidant defense system. The HT stress induced membrane damage, degraded chlorophyll, generated redox metabolites and significantly reduced growth and biomass in all the cultivars. Among all the four treatments, foliar application of SA and SNP were most effective in the regulation of growth and physiological processes of the cultivars compared to ABA and PRO applications. Thus, signifying the protective role of SA and SNP in mitigation of HT induced oxidative stress and conferring HT stress tolerance in the cultivars. Gene expression and leaf proteome analysis revealed that these phytohormones were also involved in regulation of defense related gene expression, stress inducible proteins and de novo synthesis of specific proteins under HT stress. The experimental findings depict that foliar applications of SA and SNP enhances HT stress tolerance in lablab cultivars by modulating antioxidant defense system and by regulating bio-physical growth more effectively as compared to ABA and PRO application.

Leaf Physiological and Proteomic Analysis to Elucidate Silicon Induced Adaptive Response under Salt Stress in Rosa hybrida 'Rock Fire'.

Beneficial effects of silicon (Si) on growth and development have been witnessed in several plants. Nevertheless, studies on roses are merely reported. Therefore, the present investigation was carried out to illustrate the impact of Si on photosynthesis, antioxidant defense and leaf proteome of rose under salinity stress. In vitro-grown, acclimatized Rosa hybrida ‘Rock Fire’ were hydroponically treated with four treatments, such as control, Si (1.8 mM), NaCl (50 mM), and Si+NaCl. After 15 days, the consequences of salinity stress and the response of Si addition were analyzed. Scorching of leaf edges and stomatal damages occurred due to salt stress was ameliorated under Si supplementation. Similarly, reduction of gas exchange, photosynthetic pigments, higher lipid peroxidation rate, and accumulation of reactive oxygen species under salinity stress were mitigated in Si treatment. Lesser oxidative stress observed was correlated with the enhanced activity and expression of antioxidant enzymes, such as superoxide dismutase, catalase, and ascorbate peroxidase in Si+NaCl treatment. Importantly, sodium transportation was synergistically restricted with the stimulated counter-uptake of potassium in Si+NaCl treatment. Furthermore, two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) results showed that out of 40 identified proteins, on comparison with control 34 proteins were down-accumulated and six proteins were up-accumulated due to salinity stress. Meanwhile, addition of Si with NaCl treatment enhanced the abundance of 30 proteins and downregulated five proteins. Differentially-expressed proteins were functionally classified into six groups, such as photosynthesis (22%), carbohydrate/energy metabolism (20%), transcription/translation (20%), stress/redox homeostasis (12%), ion binding (13%), and ubiquitination (8%). Hence, the findings reported in this work could facilitate a deeper understanding on potential mechanism(s) adapted by rose due to the exogenous Si supplementation during the salinity stress.

The disadvantages of being a hybrid during drought: A combined analysis of plant morphology, physiology and leaf proteome in maize.

A comparative analysis of various parameters that characterize plant morphology, growth, water status, photosynthesis, cell damage, and antioxidative and osmoprotective systems together with an iTRAQ analysis of the leaf proteome was performed in two inbred lines of maize (Zea mays L.) differing in drought susceptibility and their reciprocal F1 hybrids. The aim of this study was to dissect the parent-hybrid relationships to better understand the mechanisms of the heterotic effect and its potential association with the stress response. The results clearly showed that the four examined genotypes have completely different strategies for coping with limited water availability and that the inherent properties of the F1 hybrids, i.e. positive heterosis in morphological parameters (or, more generally, a larger plant body) becomes a distinct disadvantage when the water supply is limited. However, although a greater loss of photosynthetic efficiency was an inherent disadvantage, the precise causes and consequences of the original predisposition towards faster growth and biomass accumulation differed even between reciprocal hybrids. Both maternal and paternal parents could be imitated by their progeny in some aspects of the drought response (e.g., the absence of general protein down-regulation, changes in the levels of some carbon fixation or other photosynthetic proteins). Nevertheless, other features (e.g., dehydrin or light-harvesting protein contents, reduced chloroplast proteosynthesis) were quite unique to a particular hybrid. Our study also confirmed that the strategy for leaving stomata open even when the water supply is limited (coupled to a smaller body size and some other physiological properties), observed in one of our inbred lines, is associated with drought-resistance not only during mild drought (as we showed previously) but also during more severe drought conditions.