Comparative proteomics of sugarcane smut fungus - Sporisorium scitamineum unravels dynamic proteomic alterations during the dimorphic transition.

Life cycle of the dimorphic sugarcane smut fungi, Sporisorium scitamineum, involves recognition and mating of compatible saprophytic yeast-like haploid sporidia (MAT-1 and MAT-2) that upon fusion, develop into infective dikaryotic mycelia. Although the dimorphic transition is intrinsically linked with the pathogenicity and virulence of S. scitamineum, it has never been studied using a proteomic approach. In the present study, an iTRAQ-based comparative proteomic analysis of three distinct stages was carried out. The stages were: the dimorphic transition period - haploid sporidial stage (MAT-1 and MAT-2); the transition phase (24 h post co-culturing (hpc)) and the dikaryotic mycelial stage (48 hpc). Functional categorization of differentially abundant proteins showed that the most altered biological processes were energy production, primary metabolism, especially, carbohydrate, amino acid, fatty acid, followed by translation, post-translation and protein turnover. Several differentially abundant proteins (DAPs), especially in the dikaryotic mycelial stage were predicted as effectors. Taken together, key molecular mechanisms underpinning the dimorphic transition in S. scitamineum at the proteome level were highlighted. The catalogue of stage-specific and dimorphic transition-associated-proteins and potential effectors identified herein represents a list of potential candidates for defective mutant screening to elucidate their functional role in the dimorphic transition and pathogenicity in S. scitamineum. BIOLOGICAL SIGNIFICANCE: Being the first comparative proteomics analysis of S. scitamineum, this study comprehensively examined three pivotal life cycle stages of the pathogen: the non-pathogenic haploid phase, the transition phase, and the pathogenic dikaryotic mycelial stage. While previous studies have reported the sugarcane and S. scitamineum interactions, this study endeavored to specifically identify the proteins responsible for pathogenicity. By analyzing the proteomic alterations between the haploid and dikaryotic mycelial phases, the study revealed significant changes in metabolic pathway-associated proteins linked to energy production, notably oxidative phosphorylation, and the citrate cycle. Furthermore, this study successfully identified key metabolic pathways that undergo reprogramming during the transition from the non-pathogenic to the pathogenic stage. The study also deciphered the underlying mechanisms driving the morphological and physiological alterations crucial for the S. scitamineum virulence. By studying its life cycle stages, identifying the key metabolic pathways and stage-specific proteins, it provides unprecedented insights into the pathogenicity and potential avenues for intervention. As proteomics continues to advance, such studies pave the way for a deeper understanding of plant-pathogen interactions and the development of innovative strategies to mitigate the impact of devastating pathogens like S. scitamineum.

Uptake and distribution of perfluoroalkyl substances by grafted tomato plants cultivated in a contaminated site in northern Italy.

Poly- and perfluoroalkyl substances (PFAS) are highly persistent and mobile pollutants raising alarming concerns due to their capability to accumulate in living organisms and exert toxic effects on human health. We studied the accumulation of different PFAS in the leaves and fruits of tomato plants grown on a PFAS-polluted soil in North-East Italy. Tomato plants were grafted with different rootstocks characterized by different vigor, and irrigated with PFAS-polluted groundwater. Leaves and fruits of the first and sixth truss were analyzed at full plant maturity. All tomato varieties accumulated PFAS in leaves and fruits, with the highest concentrations detected in the most vigorous rootstock and reflecting the PFAS concentration profile of the irrigation water. PFAS with a chain length from 4 to 8 C atoms and with carboxylic and sulfonic functional groups were detected in plant leaves, whereas only carboxylic C4, C5, and C6 PFAS were detected in tomato fruits. A general trend of decreasing PFAS concentrations in fruits upon increasing height of the plant trusses was observed. Calculation of the target hazard quotient (THQ) showed increasing values depending on the plant vigor. The hazard index (HI) values showed values slightly higher than 1 for the most vigorous plants, indicating potential risks to human health associated with the consumption of contaminated tomato fruits.

Subclinical Mastitis from Streptococcus agalactiae and Prototheca spp. Induces Changes in Milk Peptidome in Holstein Cattle.

Early detection of bovine subclinical mastitis may improve treatment strategies and reduce the use of antibiotics. Herein, individual milk samples from Holstein cows affected by subclinical mastitis induced by S. agalactiae and Prototheca spp. were analyzed by untargeted and targeted mass spectrometry approaches to assess changes in their peptidome profiles and identify new potential biomarkers of the pathological condition. Results showed a higher amount of peptides in milk positive on the bacteriological examination when compared with the negative control. However, the different pathogens seemed not to trigger specific effects on the milk peptidome. The peptides that best distinguish positive from negative samples are mainly derived from the most abundant milk proteins, especially from ß- and αs1-casein, but also include the antimicrobial peptide casecidin 17. These results provide new insights into the physiopathology of mastitis. Upon further validation, the panel of potential discriminant peptides could help the development of new diagnostic and therapeutic tools.

Perfluoroalkyl substances exposure alters stomatal opening and xylem hydraulics in willow plants.

Climate change and pollution are increasingly important stress factors for life on Earth. Dispersal of poly- and perfluoroalkyl substances (PFAS) are causing worldwide contamination of soils and water tables. PFAS are partially hydrophobic and can easily bioaccumulate in living organisms, causing metabolic alterations. Different plant species can uptake large amounts of PFAS, but little is known about its consequences for the plant water relation and other physiological processes, especially in woody plants. In this study, we investigated the fractionation of PFAS bioaccumulation from roots to leaves and its effects on the conductive elements of willow plants. Additionally, we focused on the stomal opening and the phytohormonal content. For this purpose, willow cuttings were exposed to a mixture of 11 PFAS compounds and the uptake was evaluated by LC-MS/MS. Stomatal conductance was measured and the xylem vulnerability to air embolism was tested and further, the abscisic acid and salicylic acid contents were quantified using LC-MS/MS. PFAS accumulated from roots to leaves based on their chemical structure. PFAS-exposed plants showed reduced stomatal conductance, while no differences were observed in abscisic acid and salicylic acid contents. Interestingly, PFAS exposure caused a higher vulnerability to drought-induced xylem embolism in treated plants. Our study provides novel information about the PFAS effects on the xylem hydraulics, suggesting that the plant water balance may be affected by PFAS exposure. In this perspective, drought events may be more stressful for PFAS-exposed plants, thus reducing their potential for phytoremediation.

The impact of milk storage temperatures on cheese quality and microbial communities at dairy processing plant scale.

Cheese production is an applied biotechnology whose proper outcome relies strictly on the complex interactive dynamics which unfold within defined microbial groups. These may start being active from the collection of milk and continue up to its final stages of maturation. One of the critical parameters playing a major role is the milk refrigeration temperature before pasteurization as it can affect the proportion of psychrotrophic taxa abundance in the total milk bacterial population. While a standard temperature of 4 °C is the common choice, due to its general growth control effect, it does have a potential drawback. This is due to the fact that some cold-tolerant genera present a proteolytic activity with uncompleted proliferation, which could negatively affect curd clotting and regular cheese maturation. Moreover, accidental thermal variations of milk before cheese-making, in a plus or minus direction, can occur both at farm collection sites and during transfer to dairy plant. This present research, directly commissioned by a major fresh cheese-producing company, includes an in-factory trial. In this trial, a gradient of temperatures from 4 °C to 13 °C, which were subsequently reversed, was purposely adopted to: (a) verify sensory alterations in the resulting product at different maturation stages, and, (b) analyze, in parallel, using DNA extraction and 16S-metabarcoding sequencing from the same samples, the presence, abundance and corresponding taxonomical identity of all the bacteria featured in communities found in milk and cheese samples. Overall, 1,714 different variants were detected and sorted into 394 identified taxa. Significant bacterial community shifts in cheese were observed in response to milk refrigeration temperature and subsequently associated with samples having altered scores in sensory panel tests. In particular, proteolytic psychrotrophes were outcompeted by Enterobacteriales and by other taxa at the peak temperature of 13 °C, but aggressively increased in the descent phases, upon the cooling down of milk to values of 7 °C. Relevant clues have been collected for better anticipation of thermal abuse effects or parameter variations allowing for improved handling of technical processing conditions by the cheese manufacturing industry.

Buckwheat OMICS: present status and future prospects.

Buckwheat (Fagopyrum spp.) is an underutilized resilient crop of North Western Himalayas belonging to the family Polygonaceae and is a source of essential nutrients and therapeutics. Common Buckwheat and Tatary Buckwheat are the two main cultivated species used as food. It is the only grain crop possessing rutin, an important metabolite with high nutraceutical potential. Due to its inherent tolerance to various biotic and abiotic stresses and a short life cycle, Buckwheat has been proposed as a model crop plant. Nutritional security is one of the major concerns, breeding for a nutrient-dense crop such as Buckwheat will provide a sustainable solution. Efforts toward improving Buckwheat for nutrition and yield are limited due to the lack of available: genetic resources, genomics, transcriptomics and metabolomics. In order to harness the agricultural importance of Buckwheat, an integrated breeding and OMICS platforms needs to be established that can pave the way for a better understanding of crop biology and developing commercial varieties. This, coupled with the availability of the genome sequences of both Buckwheat species in the public domain, should facilitate the identification of alleles/QTLs and candidate genes. There is a need to further our understanding of the molecular basis of the genetic regulation that controls various economically important traits. The present review focuses on: the food and nutritional importance of Buckwheat, its various omics resources, utilization of omics approaches in understanding Buckwheat biology and, finally, how an integrated platform of breeding and omics will help in developing commercially high yielding nutrient rich cultivars in Buckwheat.

Metabolic-GWAS provides insights into genetic architecture of seed metabolome in buckwheat.

BACKGROUND: Buckwheat (Fagopyrum spp.), belonging to the Polygonaceae family, is an ancient pseudo-cereal with high nutritional and nutraceutical properties. Buckwheat proteins are gluten-free and show balanced amino acid and micronutrient profiles, with higher content of health-promoting bioactive flavonoids that make it a golden crop of the future. Plant metabolome is increasingly gaining importance as a crucial component to understand the connection between plant physiology and environment and as a potential link between the genome and phenome. However, the genetic architecture governing the metabolome and thus, the phenome is not well understood. Here, we aim to obtain a deeper insight into the genetic architecture of seed metabolome in buckwheat by integrating high throughput metabolomics and genotyping-by-sequencing applying an array of bioinformatics tools for data analysis. RESULTS: High throughput metabolomic analysis identified 24 metabolites in seed endosperm of 130 diverse buckwheat genotypes. The genotyping-by-sequencing (GBS) of these genotypes revealed 3,728,028 SNPs. The Genome Association and Prediction Integrated Tool (GAPIT) assisted in the identification of 27 SNPs/QTLs linked to 18 metabolites. Candidate genes were identified near 100 Kb of QTLs, providing insights into several metabolic and biosynthetic pathways. CONCLUSIONS: We established the metabolome inventory of 130 germplasm lines of buckwheat, identified QTLs through marker trait association and positions of potential candidate genes. This will pave the way for future dissection of complex economic traits in buckwheat.

Investigation of sulfur-containing compounds in spears of green and white Asparagus officinalis through LC-MS and HS-GC-MS.

Asparagus officinalis is largely consumed as food in many parts of the world, and due to its content in secondary metabolites can be considered as a vegetable with health-promoting value. Part of its organoleptic qualities can be ascribed to the presence of sulfur-containing compounds, nevertheless qualitative data about the volatile and non-volatile pools of these compounds in green and white spears of asparagus are poorly investigated. Due to the wide alimentary use of this crop and the potential biological properties of S-containing compounds, research aimed at filling this gap is required. In this paper, a comprehensive characterization of S-metabolites in asparagus performed by LC-MS and GC-MS is reported. Both green and white varieties of asparagus were considered. The fresh vegetal material was subjected to different sample preparation procedures, such as solvent extraction, distillation, and headspace sampling. Furthermore, a derivatization protocol with 4,4'-dithiodipyridine was used for low-molecular weight thiols, and both derivatized and underivatized compounds were analysed by LC-MS. The methods allowed to identify 80 S-containing metabolites in asparagus samples, and to assess the distribution of these compounds in different parts of the spears. Results were discussed comparing the literature, and the identified compounds were considered to explain some peculiar taste and odorous properties of green and white asparagus, although further research is required to confirm our hypotheses. Overall, in this work we report for the first time an exhaustive characterization of S-compounds profile in spears of green and white Asparagus varieties. Furthermore, results indicate that multiple approaches should be used to study the S-containing metabolites of this plant, due to their diverse chemical properties.

Perfluorinated alkyl substances affect the growth, physiology and root proteome of hydroponically grown maize plants.

Poly- and perfluorinated alkyl substances (PFAS) are a group of persistent organic pollutants causing serious global concern. Plants can accumulate PFAS but their effect on plant physiology, especially at the molecular level is not very well understood. Hence, we used hydroponically-grown maize plants treated with a combination of eleven different PFAS (each at 100 µg L-1) to investigate their bioaccumulation and effects on the growth, physiology and their impact on the root proteome. A dose-dependent decrease in root growth parameters was evidenced with a significant reduction in the relative growth rate, fresh weight of leaves and roots and altered photosynthetic parameters in PFAS-treated plants. Higher concentration of shorter PFAS (C < 8) was detected in the leaves, while long-chain PFAS (C ≥ 8) were more retained in roots. From the root proteome analysis, we identified 75 differentially abundant proteins, mostly involved in cellular metabolic and biosynthetic processes, translation and cytoskeletal reorganization. Validating the altered protein abundance using quantitative real-time PCR, the results were further substantiated using amino acid and fatty acid profiling, thus, providing first insight into the altered metabolic state of plants exposed to PFAS from a proteomics perspective.

Low Temperature Stress Tolerance: An Insight Into the Omics Approaches for Legume Crops.

The change in climatic conditions is the major cause for decline in crop production worldwide. Decreasing crop productivity will further lead to increase in global hunger rate. Climate change results in environmental stress which has negative impact on plant-like deficiencies in growth, crop yield, permanent damage, or death if the plant remains in the stress conditions for prolonged period. Cold stress is one of the main abiotic stresses which have already affected the global crop production. Cold stress adversely affects the plants leading to necrosis, chlorosis, and growth retardation. Various physiological, biochemical, and molecular responses under cold stress have revealed that the cold resistance is more complex than perceived which involves multiple pathways. Like other crops, legumes are also affected by cold stress and therefore, an effective technique to mitigate cold-mediated damage is critical for long-term legume production. Earlier, crop improvement for any stress was challenging for scientific community as conventional breeding approaches like inter-specific or inter-generic hybridization had limited success in crop improvement. The availability of genome sequence, transcriptome, and proteome data provides in-depth sight into different complex mechanisms under cold stress. Identification of QTLs, genes, and proteins responsible for cold stress tolerance will help in improving or developing stress-tolerant legume crop. Cold stress can alter gene expression which further leads to increases in stress protecting metabolites to cope up the plant against the temperature fluctuations. Moreover, genetic engineering can help in development of new cold stress-tolerant varieties of legume crop. This paper provides a general insight into the "omics" approaches for cold stress in legume crops.

Desiccation Tolerance in Ramonda serbica Panc.: An Integrative Transcriptomic, Proteomic, Metabolite and Photosynthetic Study.

The resurrection plant Ramonda serbica Panc. survives long desiccation periods and fully recovers metabolic functions within one day upon watering. This study aimed to identify key candidates and pathways involved in desiccation tolerance in R. serbica. We combined differential transcriptomics and proteomics, phenolic and sugar analysis, FTIR analysis of the cell wall polymers, and detailed analysis of the photosynthetic electron transport (PET) chain. The proteomic analysis allowed the relative quantification of 1192 different protein groups, of which 408 were differentially abundant between hydrated (HL) and desiccated leaves (DL). Almost all differentially abundant proteins related to photosynthetic processes were less abundant, while chlorophyll fluorescence measurements implied shifting from linear PET to cyclic electron transport (CET). The levels of H2O2 scavenging enzymes, ascorbate-glutathione cycle components, catalases, peroxiredoxins, Fe-, and Mn superoxide dismutase (SOD) were reduced in DL. However, six germin-like proteins (GLPs), four Cu/ZnSOD isoforms, three polyphenol oxidases, and 22 late embryogenesis abundant proteins (LEAPs; mainly LEA4 and dehydrins), were desiccation-inducible. Desiccation provoked cell wall remodeling related to GLP-derived H2O2/HO● activity and pectin demethylesterification. This comprehensive study contributes to understanding the role and regulation of the main metabolic pathways during desiccation aiming at crop drought tolerance improvement.

Novel echocardiographic score to predict duct-dependency after percutaneous relief of critical pulmonary valve stenosis/atresia.

OBJECTIVES: This study aimed to identify clinical, hemodynamic, or echocardiographic predictive features of persistent duct-dependency of pulmonary circulation (PDDPC) after effective percutaneous relief of pulmonary atresia with the intact ventricular septum (PA-IVS) or critical pulmonary stenosis (CPS). METHODS: From 2010 to 2021, 55 neonates with PA-IVS or CPS underwent percutaneous right ventricle (RV) decompression at our Institution. After successfully relief of critical obstruction, 27 patients (group I) showed PDDPC, whereas RV was able to support the pulmonary circulation in the remaining 28 patients (group II). Clinical, hemodynamic, and echocardiographic features of these two groups were compared. RESULTS: No significant difference in clinical and hemodynamic data was found between the groups, although the group I had a lower oxygen saturation at hospital admission. However, tricuspid valve (TV) diameter <8.8 mm, TV z-score ←2.12, tricuspid/mitral valve annular ratio <.78, pulmonary valve diameter <6.7 mm, pulmonary valve z-score ←1.17, end-diastolic RV area <1.35 cm2 , end-systolic right atrium area >2.45 cm2 , percentage amount of interatrial right-to-left shunt >69.5%, moderate/severe tricuspid regurgitation, RV systolic pressure >42.5 mmHg, tricuspid E/E' ratio >6.6 showed each significant predictive value of PDDPC. These parameters were used to build a composite echocardiographic score (PDDPC-score), assigning one point each above the respective cut-off value. A score ≥4.00 showed high sensitivity (100%) and specificity (86%) in predicting PDDPC. CONCLUSION: Clinical and hemodynamic features fail to predict the short-term fate of the pulmonary circulation after successful treatment of PA-IVS/CPS. However, a simple, composite echocardiographic score is useful to predict PDDPC and could be crucial in the management of this frail subset of patients.

Fe and Zn stress induced gene expression analysis unraveled mechanisms of mineral homeostasis in common bean (Phaseolus vulgaris L.).

Iron (Fe) and zinc (Zn) stress significantly affects fundamental metabolic and physiological processes in plants that results in reduction of plant growth and development. In the present study, common bean variety; Shalimar French Bean-1 (SFB-1) was used as an experimental material. Four different MGRL media i.e. normal MGRL medium (Control), media without Fe (0-Fe), media without Zn (0-Zn) and media with excess Zn (300-Zn) were used for growing seeds of SFB-1 under in vitro condition for three weeks under optimum conditions. Three week old shoot and root tissues were harvested from the plants grown in these four different in vitro conditions and were, subjected to Fe and Zn estimation. Further, extraction of total RNA for differential gene expression of ten candidate genes selected based on our in silico investigation and their classification, phylogeny and expression pattern was unraveled. Expression analysis of three candidate genes (OPT3, NRAMP2 and NRAMP3) in roots revealed possible cross talk among Fe/Zn stress that was further confirmed by observing less accumulation of Fe in roots under both these conditions. However, we observed, higher accumulation of Fe in shoots under 0-Fe condition compared to control that suggests precise sensing for priority based compartmentalization and partitioning leading to higher accumulation of Fe in shoots. Furthermore, the expression analysis of IRT1, FRO1 and Ferritin 1 genes under Fe/Zn stress suggested their role in uptake/transport and signaling of Fe and Zn, whereas the expression of ZIP2, NRAMP1, HA2 and GLP1 genes were highly responsive to Zn in Phaseolus vulgaris. The identified genes highly responsive to Fe and Zn stress condition can be potential candidates for overcoming mineral stress in dicot crop plants.

Protein profile of commercial soybean milks analyzed by label-free quantitative proteomics.

The consumption of soy milk is increasing worldwide for its nutritional value and health benefits, however, its protein composition after commercialization is not well known. Technological and thermal treatments to which soy milk is subjected could affect the protein composition of the commercial products. This study compared the protein profile of 15 different commercial soy milks using a label-free quantitative proteomics approach. Proteins related to nutrient reservoir activity, endopeptidase inhibitor activity, lipid binding, and seed maturation contribute the most in terms of percentage mass. Their associated Gene Ontology terms are also enriched. Samples clustered into three groups based on their protein composition, with glycinins and beta-conglycinins being the most influential for determining the clustering. Amino acid composition estimated from the proteomics data also reflects the clustering of samples. Twenty allergenic proteins varying in abundance were identified, with Gly m 5 and Gly m 6 being the predominantly abundant allergens.

Rare case of Kawasaki disease with cardiac tamponade and giant coronary artery aneurysms.

Kawasaki disease is an acute systemic vascular disease, generally self-limited and typically affecting children <5 years old, which leads to coronary artery aneurysms in about 25% of untreated cases. Cardiovascular involvement is characterised by transient pancarditis, in acute phase, while coronary illness, ranging from mild dilation to giant CAAs occurs late, rarely before the 10th day since fever onset. Here, we describe a peculiar case of KD, which occurred in a 4-month-old infant and presented with exudate cardiac tamponade and early giant aneurism of both the proximal right coronary artery) and the left circumflex coronary artery, in acute phase of the disease.

Metabarcoding analysis of the bacterial succession during vermicomposting of municipal solid waste employing the earthworm Eisenia fetida.

A culture-independent DNA metabarcoding analysis of the bacterial communities was carried out throughout a complete vermicomposting cycle of municipal solid waste material using the earthworm Eisenia fetida. 16S rRNA amplicons from the initial material (0 days), an intermediate (42 days), and a final stage (84 days) were sequenced in an Illumina NGS platform and compared. A steady increase in community diversity was observed corresponding to a 2.5-fold higher taxa richness and correspondingly risen values of the Shannon and Simpson ecological indexes and the evenness parameter. A total of 49,665 operational taxonomic units (OTUs) were counted. From the qualitative standpoint, a clear successional shift was observed with an initial community dominated by putatively plant-associated groups belonging to the Rhizobiales order within the Alphaproteobacteria class, regressively leaving the scores of relative abundance (RA) to the Firmicutes phylum and in particular to the Bacilli. Vermistabilization of municipal solid waste (MSW) increased (p < 0.001) the TKN and total P content in the final vermicompost, while pH, TOC, and C/N ratio declined (p < 0.001) in the process. Likewise, a progressive decrease was noticed in ß-glucosidase, acid phosphatase, and urease activity while protease and dehydrogenase showed a slight increase, followed by a steep fall. A strong positive correlation was observed among the canonical functions of physico-chemical attributes and enzyme activities. The canonical correspondence analysis (CCA) revealed that significant families did not change on the temporal scale; however, their abundance was influenced by the abiotic environmental factors. In comparison to prior studies on vermicomposting that used different earthworm species (Eisenia andrei) and different substrates, results reflect a considerable degree of substrate specificity for the earthworm species used. The results offer clues to optimize the vermistabilization of MSW along with its potential use in agriculture, to foster improved levels of the circular economy.

A novel HPLC-MS/MS approach for the identification of biological thiols in vegetables.

Thiols are important natural molecules with diverse functions, ranging from acting as antioxidants that prevent chronic diseases to contributing aromas to foods and beverages. Biological thiols such as glutathione are of particular interest due to their functional roles, which include helping maintain cellular redox homeostasis and detoxifying reactive oxygen species. However, knowledge of thiol metabolism in plants is limited to studying known compounds, whereas other important thiol-containing metabolites could also exist. This work aimed to develop a new analytical approach for screening of thiols in plants, using four vegetal examples and beginning with HPLC-MS/MS in precursor ion scan mode, after extraction and thiol-specific derivatisation with 4,4'-dithiodipyridine (DTDP). Compound identity for prospective thiols was then proposed using HPLC with high resolution MS, and verified with authentic standards. This approach could lead to prospecting studies that identify thiols with potential roles in metabolic pathways, nutritional value of vegetables, or flavouring of foods.

Physiological and Multi-Omics Approaches for Explaining Drought Stress Tolerance and Supporting Sustainable Production of Rice.

Drought differs from other natural disasters in several respects, largely because of the complexity of a crop's response to it and also because we have the least understanding of a crop's inductive mechanism for addressing drought tolerance among all abiotic stressors. Overall, the growth and productivity of crops at a global level is now thought to be an issue that is more severe and arises more frequently due to climatic change-induced drought stress. Among the major crops, rice is a frontline staple cereal crop of the developing world and is critical to sustaining populations on a daily basis. Worldwide, studies have reported a reduction in rice productivity over the years as a consequence of drought. Plants are evolutionarily primed to withstand a substantial number of environmental cues by undergoing a wide range of changes at the molecular level, involving gene, protein and metabolite interactions to protect the growing plant. Currently, an in-depth, precise and systemic understanding of fundamental biological and cellular mechanisms activated by crop plants during stress is accomplished by an umbrella of -omics technologies, such as transcriptomics, metabolomics and proteomics. This combination of multi-omics approaches provides a comprehensive understanding of cellular dynamics during drought or other stress conditions in comparison to a single -omics approach. Thus a greater need to utilize information (big-omics data) from various molecular pathways to develop drought-resilient crop varieties for cultivation in ever-changing climatic conditions. This review article is focused on assembling current peer-reviewed published knowledge on the use of multi-omics approaches toward expediting the development of drought-tolerant rice plants for sustainable rice production and realizing global food security.

Efficient protein extraction for shotgun proteomics from hydrated and desiccated leaves of resurrection Ramonda serbica plants.

Resurrection plant Ramonda serbica is a suitable model to investigate vegetative desiccation tolerance. However, the detailed study of these mechanisms at the protein level is hampered by the severe tissue water loss, high amount of phenolics and polysaccharide, and possible protein modifications and aggregations during the extraction and purification steps. When applied to R. serbica leaves, widely used protein extraction protocols containing polyvinylpolypyrrolidone and ascorbate, as well as the phenol/SDS/buffer-based protocol recommended for recalcitrant plant tissues failed to eliminate persistent contamination and ensure high protein quality. Here we compared three protein extraction approaches aiming to establish the optimal one for both hydrated and desiccated R. serbica leaves. To evaluate the efficacy of these protocols by shotgun proteomics, we also created the first R. serbica annotated transcriptome database, available at http://www.biomed.unipd.it/filearrigoni/Trinity_Sample_RT2.fasta . The detergent-free phenol-based extraction combined with dodecyl-ß-D-maltoside-assisted extraction enabled high-yield and high-purity protein extracts. The phenol-based protocol improved the protein-band resolution, band number, and intensity upon electrophoresis, and increased the protein yield and the number of identified peptides and protein groups by LC-MS/MS. Additionally, dodecyl-ß-D-maltoside enabled solubilisation and identification of more membrane-associated proteins. The presented study paves the way for investigating the desiccation tolerance in R. serbica, and we recommend this protocol for similar recalcitrant plant material.