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The continuous global escalation of soil salinization areas presents severe challenges to the stability and growth of agricultural development across the world. In-depth research on sugar beet (Beta vulgaris L.), an important economic and sugar crop with salt tolerance characteristics, is crucial for to determine its salt-tolerance mechanisms, which has important practical implications for production. This review summarizes the multifaceted effects of salt stress on sugar beet, ranging from individual plant responses to cellular and molecular adaptations. Sugar beet exhibits robust salt-tolerance mechanisms, including osmotic regulation, ion balance management, and the compartmentalization of toxic ions. Omics technologies, including genomics, transcriptomics, proteomics, post-translational modification omics and metabolomics, have played crucial roles in elucidating these mechanisms. Key genes and pathways involved in salt tolerance in sugar beet have been identified, paving the way for targeted breeding strategies and biotechnological advancements. Understanding these mechanisms not only enhances our knowledge of sugar beet's adaptation strategies but also provides insights for improving salt tolerance in other crops. Future studies should focus on analyzing gene expression changes in sugar beet under salt stress to gain insight into the molecular aspects of its salt-tolerance mechanisms. Meanwhile, the effects of different environmental conditions on sugar beet adaptation strategies should also be investigated to improve their growth potential in salinized soils. [ABSTRACT FROM AUTHOR]

Azadirachta indica or neem is recognized for its unique properties, both against insects and in improving human health. However, proteomics studies of this plant's response to abiotic stresses have yet to be reported. Application of this technique highly depends on efficient and optimized sample preparation, in deriving highly reproducible proteomics data. The present study highlights the comparison of protein extraction method which effectively recovers proteins from recalcitrant phenolic-rich tissues of neem callus. Four protocols based on TCA-acetone or/and phenol extraction methods were compared. The TCA-acetone complemented with PVPP method was the most effective approach and generated the greatest total protein yields, spot number, good resolution, and spot intensities. Subsequently, the differential proteomics performed revealed that proteins associated with oxidoreduction, energy, transcriptional, stress response, respiration, and cell division were differentially regulated in salinity stress treatment. Meanwhile, the expression of proteins classified in the group of energy, photosynthesis, protein synthesis, oxidoreduction, and post-transcriptional regulation was altered in the JA-stressed callus. The accumulation of AZA was also noticeably reduced in salinity- and JA-treated callus, suggesting these tested stress parameters were not effective in improving AZA production in the neem callus. Overall, this analysis provides valuable information on the application of plant tissue culture as an alternative for sustainable production of major bioactive constituents in A. indica. Together, the proteomics results brought forward a possible fundamental representation of the mechanism associated with salinity and JA tolerance in the neem callus. [ABSTRACT FROM AUTHOR]

The Nobel Prize in Chemistry for 2022 was awarded to the pioneers of click chemistry. In Lego-like 'click chemistry', molecular building blocks 'snap' together quickly and specifically. The field of click chemistry has received comparatively little attention within the plant science community, but it holds great potential. Classical click reactions (CuAAC, SPAAC, and iEDDA) have been exploited to study different biomolecules, physiological processes, and biofunctions of plants. Bioorthogonal non-classical amino acid tagging (BONCAT) has provided new pathways and insights to identify plant proteomics and interrogate plant–microbe interactions. Bioorthogonal click chemistry coupled with activity-based probes and metabolic labeling can overcome the limitations of current tagging strategies and have served as effective platforms for the visualization of plant biological processes. The Nobel Prize in Chemistry for 2022 was awarded to the pioneers of Lego-like 'click chemistry': combinatorial chemistry with remarkable modularity and diversity. It has been applied to a wide variety of biological systems, from microorganisms to plants and animals, including humans. Although click chemistry is a powerful chemical biology tool, comparatively few studies have examined its potential in plant science. Here, we review click chemistry reactions and their applications in plant systems, highlighting the activity-based probes and metabolic labeling strategies combined with bioorthogonal click chemistry to visualize plant biological processes. These applications offer new opportunities to explore and understand the underlying molecular mechanisms regulating plant composition, growth, metabolism, defense, and immune responses. [ABSTRACT FROM AUTHOR]

We examined the proteomic profiles of three registered opium poppy cultivars (Papaver somniferum L.) with varying alkaloid contents. The study was conducted on both the stem and capsule organs. A high number of differentially expressed proteins (DEPs) were identified between the cultivars and the organs. We analyzed DEPs for their contribution in GO terms and KEGG pathways. The upregulated DEPs were significantly enriched in photosynthesis and translation for morphine-rich and noscapine-rich cultivars, respectively. The data indicated that photosynthesis is crucial for benzylisoquinoline alkaloid (BIA) biosynthesis, but different processes are also effective in morphine and noscapine biosynthesis, which occur at different branches in the biosynthetic pathway. The proteomics profiles revealed that energy demand is more effective in morphine biosynthesis, while translational control plays a leading role in noscapine biosynthesis. This study represents the first report demonstrating organ-based and cultivar-based protein expression differences in mature poppy plants. [ABSTRACT FROM AUTHOR]

In recent years, ultrafast liquid chromatography/mass spectrometry methods have been extensively developed for the use in proteome profiling in biochemical studies. These methods are intended for express monitoring of cell response to biotic stimuli and elucidation of correlation of molecular changes with biological processes and phenotypical changes. New technologies, including the use of nanomaterials, are actively introduced to increase agricultural production. However, this requires complex approbation of new fertilizers and investigation of mechanisms underlying the biotic effects on the germination, growth, and development of plants. The aim of this work was to adapt the method of ultrafast chromatography/mass spectrometry for rapid quantitative profiling of molecular changes in 7-day-old wheat seedlings in response to pre-sowing seed treatment with iron compounds. The used method allows to analyze up to 200 samples per day; its practical value lies in the possibility of express proteomic diagnostics of the biotic action of new treatments, including those intended for agricultural needs. Changes in the regulation of photosynthesis, biosynthesis of chlorophyll and porphyrin- and tetrapyrrole-containing compounds, glycolysis (in shoot tissues), and polysaccharide metabolism (in root tissues) were shown after seed treatment with suspensions containing film-forming polymers (PEG 400, Na-CMC, Na2-EDTA), iron (II, III) nanoparticles, or iron (II) sulfate. Observations at the protein levels were consistent with the results of morphometry, superoxide dismutase activity assay, and microelement analysis of 3-day-old germinated seeds and shoots and roots of 7-day-old seedlings. A characteristic molecular signature involving proteins participating in the regulation of photosynthesis and glycolytic process was suggested as a potential marker of the biotic effects of seed treatment with iron compounds, which will be confirmed in further studies. [ABSTRACT FROM AUTHOR]

Autophagy is a catabolic pathway capable of degrading cellular components ranging from individual molecules to organelles. Autophagy helps cells cope with stress by removing superfluous or hazardous material. In a previous work, we demonstrated that transcriptional upregulation of two autophagy-related genes, ATG5 and ATG7, in Arabidopsis thaliana positively affected agronomically important traits: biomass, seed yield, tolerance to pathogens and oxidative stress. Although the occurrence of these traits correlated with enhanced autophagic activity, it is possible that autophagy-independent roles of ATG5 and ATG7 also contributed to the phenotypes. In this study, we employed affinity purification and LC-MS/MS to identify the interactome of wild-type ATG5 and its autophagy-inactive substitution mutant, ATG5K128R Here we present the first interactome of plant ATG5, encompassing not only known autophagy regulators but also stress-response factors, components of the ubiquitin-proteasome system, proteins involved in endomembrane trafficking, and potential partners of the nuclear fraction of ATG5. Furthermore, we discovered post-translational modifications, such as phosphorylation and acetylation present on ATG5 complex components that are likely to play regulatory functions. These results strongly indicate that plant ATG5 complex proteins have roles beyond autophagy itself, opening avenues for further investigations on the complex roles of autophagy in plant growth and stress responses. [ABSTRACT FROM AUTHOR]

Bottle gourd, a common vegetable in the human diet, has been valued for its medicinal and energetic properties. In this experiment, the time-resolved analysis of the changes in the proteins’ electrophoretic patterning of the seed development at different crossing periods was studied in bottle gourd using label-free quantitative proteomics. Hybrid HBGH-35 had the highest observed protein levels at the 4th week of the crossing period (F4) compared to the parental lines, viz. G-2 (M) and Pusa Naveen (F). The crossing period is significantly correlated with grain filling and reserve accumulation. The observed protein expression profile after storage was related to seed maturation and grain filling in bottle gourds. A total of 2517 proteins were identified in differentially treated bottle gourd fruits, and 372 proteins were differentially expressed between different crossing periods. Proteins related to carbohydrate and energy metabolism, anthocyanin biosynthesis, cell stress response, and fruit firmness were characterized and quantified. Some proteins were involved in the development, while others were engaged in desiccation and the early grain-filling stage. F4 was distinguished by an increase in the accumulation of low molecular weight proteins and enzymes such as amylase, a serine protease, and trypsin inhibitors. The seed vigor also followed similar patterns of differential expression of seed storage proteins. Our findings defined a new window during seed production, which showed that at F4, maximum photosynthetic assimilates accumulated, resulting in an enhanced source–sink relationship and improved seed production. Our study attempts to observe the protein expression profiling pattern under different crossing periods using label-free quantitative proteomics in bottle gourd. It will facilitate future detailed investigation of the protein associated with quality traits and the agronomic importance of bottle gourd through selective breeding programs.

Information regarding the physiological and molecular plant responses to the treatment with new biofertilizers is limited. In this study, a fast-composting soil amendment obtained from solid waste by means of a Fenton reaction was assessed to evaluate the effects on the growth of Lactuca sativa L. var. longifolia seedlings. Growth rate, root biomass, chlorophyll concentration, and total soluble proteins of seedlings treated with the 2% fast-composting soil amendment showed significant increases in comparison with the control seedlings. Proteomic analysis revealed that the soil amendment induced the up-regulation of proteins belonging to photosynthesis machinery, carbohydrate metabolism, and promoted energy metabolism. Root proteomics indicated that the fast-composting soil amendment strongly induced the organs morphogenesis and development; root cap development, lateral root formation, and post-embryonic root morphogenesis were the main biological processes enriched by the treatment. Overall, our data suggest that the addition of the fast-composting soil amendment formulation to the base soils might ameliorate plant growth by inducing carbohydrate primary metabolism and the differentiation of a robust root system. [ABSTRACT FROM AUTHOR]

The use of sealing lids with microporous membranes (MM) is relevant to understanding the role of gas exchange on changes in ethylene and polyamine (PA) contents and proteome profiles during seedling growth. This study aimed to evaluate the influence of sealing lids and culture media, Murashige and Skoog (MS) and woody plant medium (WPM), on the in vitro seed germination and seedling development of Dalbergia nigra and on the content of ethylene, CO2, PAs, photosynthetic index, and protein profile. Sealing lids with MM allowed higher germination of seeds. Higher CO2 and leaf abscission were observed in seedlings grown in all culture media without an MM. The contents of total free PAs and free putrescine were significantly affected by the different culture media but not by the MM. A total of 545 proteins were identified, of which 101 were differentially accumulated (DAP) in seedlings grown in MS culture medium with MM compared to those grown without MM. Among DAPs, the presence of aspartate aminotransferase protein, related to ethylene biosynthesis, in seedlings grown in culture tubes containing MS culture medium without an MM was associated with greater ethylene accumulation and, consequently, greater leaf abscission. Photosynthesis- and carbon metabolism-related proteins accumulated in seedlings grown in MS culture medium sealed with MM were affected by gas exchange and associated with a higher photosynthetic index. This is the first study showing the effects of gas exchange promotion on in vitro seed germination and seedling development in D. nigra and its interaction with ethylene and protein accumulation and is relevant for optimizing in vitro micropropagation studies in this endangered woody species from the Brazilian rainforest. Key message: The type of sealing lids and culture media affects in vitro seedling growth of Dalbergia nigra by accumulation of ethylene and changes in the protein profile. [ABSTRACT FROM AUTHOR]

Castor bean (Ricinus communis L.), is one of the top 10 oilseed crops in the world and, therefore, of high economic value. Hybridization is one of the most effective ways to breed new varieties with high yield, high oil content, and better stress resistance. Therefore, prediction of desired traits in castor hybrid offspring is particularly important. In this study, proteomic analysis was performed to identify differentially expressed proteins (DEPs) in seeds between castor hybrid offspring and their female (Lm female line aLmAB2) and male parents (CSR·181). Among the DEPs upregulated in the seeds of hybrid offspring, the majority were related to seed yield and stress tolerance, while some were related to oil synthesis and fatty acid synthesis and metabolism in seeds. In other words, the hybrid offspring showed heterosis for seed yield, stress tolerance, oil synthesis, and fatty acid synthesis and metabolism when compared with their parents. Further, real-time quantitative polymerase chain reaction assays were performed on 12 genes encoding DEPs involved in oil synthesis, pollen abortion, yield, and stress tolerance of seeds. The results showed that the expression levels of the 12 genes were consistent with those of the DEPs. [ABSTRACT FROM AUTHOR]

This review is a compilation of proteomic studies on forest tree species published in the last decade (2012-2022), mostly focused on the most investigated species, including Eucalyptus, Pinus, and Quercus. Improvements in equipment, platforms, and methods in addition to the increasing availability of genomic data have favored the biological knowledge of these species at the molecular, organismal, and community levels. Integration of proteomics with physiological, biochemical and other large-scale omics in the direction of the Systems Biology, will provide a comprehensive understanding of different biological processes, from growth and development to responses to biotic and abiotic stresses. As main issue we envisage that proteomics in long-living plants will thrive light on the plant responses and resilience to global climate change, contributing to climate mitigation strategies and molecular breeding programs. Proteomics not only will provide a molecular knowledge of the mechanisms of resilience to either biotic or abiotic stresses, but also will allow the identification on key gene products and its interaction. Proteomics research has also a translational character being applied to the characterization of the variability and biodiversity, as well as to wood and non-wood derived products, traceability, allergen and bioactive peptides identification, among others. Even thought, the full potential of proteomics is far from being fully exploited in forest tree research, with PTMs and interactomics being reserved to plant model systems. The most outstanding achievements in forest tree proteomics in the last decade as well as prospects are discussed. [ABSTRACT FROM AUTHOR]

Plant proteins that are secreted without a classical signal peptide leader sequence are termed leaderless secretory proteins (LSPs) and are implicated in both plant development and (a)biotic stress responses. In plant proteomics experimental workflows, identification of LSPs is hindered by the possibility of contamination from other subcellar compartments upon purification of the secretome. Applying machine learning algorithms to predict LSPs in plants is also challenging due to the rarity of experimentally validated examples for training purposes. This work attempts to address this issue by establishing criteria for identifying potential plant LSPs based on experimental observations and training random forest classifiers on the putative datasets. The resultant plant protein database LSPDB and bioinformatic prediction tools LSPpred and SPLpred are available at lsppred.lspdb.org. The LSPpred and SPLpred modules are internally validated on the training dataset, with false positives controlled at 5%, and are also able to classify the limited number of established plant LSPs (SPLpred (3/4, LSPpred 4/4). Until such time as a larger set of bona fide (independently experimentally validated) LSPs is established using imaging technologies (light/fluorescence/electron microscopy) to confirm sub-cellular location, these tools represent a bridging method for predicting and identifying plant putative LSPs for subsequent experimental validation. [ABSTRACT FROM AUTHOR]

Plants adjust their complex molecular, biochemical, and metabolic processes to overcome salt stress. Here, we investigated the proteomic and epigenetic alterations involved in the morphophysiological responses of Pfaffia glomerata, a medicinal plant, to salt stress and the demethylating agent 5-azacytidine (5-azaC). Moreover, we investigated how these changes affected the biosynthesis of 20-hydroxyecdysone (20-E), a pharmacologically important specialized metabolite. Plants were cultivated in vitro for 40 days in Murashige and Skoog medium supplemented with NaCl (50 mM), 5-azaC (25 μM), and NaCl + 5-azaC. Compared with the control (medium only), the treatments reduced growth, photosynthetic rates, and photosynthetic pigment content, with increase in sucrose, total amino acids, and proline contents, but a reduction in starch and protein. Comparative proteomic analysis revealed 282 common differentially accumulated proteins involved in 87 metabolic pathways, most of them related to amino acid and carbohydrate metabolism, and specialized metabolism. 5-azaC and NaCl + 5-azaC lowered global DNA methylation levels and 20-E content, suggesting that 20-E biosynthesis may be regulated by epigenetic mechanisms. Moreover, downregulation of a key protein in jasmonate biosynthesis indicates the fundamental role of this hormone in the 20-E biosynthesis. Taken together, our results highlight possible regulatory proteins and epigenetic changes related to salt stress tolerance and 20-E biosynthesis in P. glomerata, paving the way for future studies of the mechanisms involved in this regulation. [ABSTRACT FROM AUTHOR]

Plants are an indispensable cornerstone of sustainable global food supply. While immense progress has been made in decoding the genomes of crops in recent decades, the composition of their proteomes, the entirety of all expressed proteins of a species, is virtually unknown. In contrast to the model plant Arabidopsis thaliana, proteomic analyses of crop plants have often been hindered by the presence of extreme concentrations of secondary metabolites such as pigments, phenolic compounds, lipids, carbohydrates or terpenes. As a consequence, crop proteomic experiments have, thus far, required individually optimized protein extraction protocols to obtain samples of acceptable quality for downstream analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS). In this article, we present a universal protein extraction protocol originally developed for gel-based experiments and combined it with an automated single-pot solid-phase-enhanced sample preparation (SP3) protocol on a liquid handling robot to prepare high-quality samples for proteomic analysis of crop plants. We also report an automated offline peptide separation protocol and optimized micro-LC-MS/MS conditions that enables the identification and quantification of ~10,000 proteins from plant tissue within 6 h of instrument time. We illustrate the utility of the workflow by analyzing the proteomes of mature tomato fruits to an unprecedented depth. The data demonstrate the robustness of the approach which we propose for use in upcoming large-scale projects that aim to map crop tissue proteomes. [ABSTRACT FROM AUTHOR]

This study evaluates the effects of alkaline and micellisation extraction methods, alongside freeze-drying and spray-drying, on the protein subunits, amino acid profiles, and proteome data of hempseed protein isolate (HPI). Findings revealed that the extraction methods affect protein profiles more than the drying methods. Micellisation-extracted HPI showed higher albumin, oleosin, and sulphur-containing protein levels than alkaline-extracted HPI. The alkali-extracted undried sample (AU) gave more potentially allergenic proteins, including Hsp70 and triosephosphate isomerase, than its micellization-extracted counterpart (MU). Unique potential allergens were identified, including malate dehydrogenase and enolase in AU, and RuBisCo in MU samples. Both drying processes impacted the HPI proteome and reduced RuBisCo in the micellisation-extracted HPI. These insights highlight the crucial role of method selection in HPI processing for optimising production in the food industry. [Display omitted] • Extraction methods impact HPI patterns more than drying methods. • Micellisation yields more albumin, oleosin, sulphur amino acids than alkaline extraction. • Freeze-drying is better for amino acid retention in micellisation-extracted HPI. • Alkaline-extracted HPI has a higher abundance of potentially allergenic proteins. • Drying reduces potentially allergenic proteins in micellisation-extracted HPI. [ABSTRACT FROM AUTHOR]

Proteomics is one of the applied sciences in biology, this century. Using the mass spectrometer device and the development of databases and bioinformatics tools, a fundamental evolution has taken place in molecular biology and new perspectives have been found in agriculture, pharmaceuticals, molecular biology, etc. Two-dimensional gel electrophoresis methods are a popular technique for protein separation because they allow researchers to quantify protein changes on a large scale. Proteomics is a large-scale evaluation of proteins. The term proteomics was produced in 1997 after the introduction of genomics. Although the use of databases and bioinformatics to interpret the results of proteomic findings is being seriously developed. In the study of huge proteomic data, it is very desirable to use the capacity of multivariate statistical methods, due to having many and extensive variables, because these multivariate statistical methods provide the possibility of statistical analysis of several variables, therefore, the use of this statistical technique recently, it has attracted the attention of proteomics scientists. In addition to understanding the tools of proteomics, we are looking for the practical use of proteomics, especially in agriculture, because today the pressure is on plant breeders to provide "smart plant varieties" that are more suitable genotypes with the ability to tolerate biotic and abiotic stress a wider range of climate changes for dealing with the food insecurity of future generations. Therefore, modern plant breeders need precise genetic modification with a gene tracking system for the modified trait. The only caveat in the application of proteomics in biotechnology applications is that genetic modification must be expressed at the protein level. The purpose of this research is to examine the concepts of some main topics in the field of proteomics in simple language, and of course, case studies of proteomics in plants, especially in industrial oilseed plants. [ABSTRACT FROM AUTHOR]

Due to its outstanding throughput and analytical resolution, gel-free LC-based shotgun proteomics represents the gold standard of proteome analysis. Thereby, the efficiency of sample preparation dramatically affects the correctness and reliability of protein quantification. Thus, the steps of protein isolation, solubilization, and proteolysis represent the principal bottleneck of shotgun proteomics. The desired performance of the sample preparation protocols can be achieved by the application of detergents. However, these compounds ultimately compromise reverse-phase chromatographic separation and disrupt electrospray ionization. Filter-aided sample preparation (FASP) represents an elegant approach to overcome these limitations. Although this method is comprehensively validated for cell proteomics, its applicability to plants and compatibility with plant-specific protein isolation protocols remain to be confirmed. Thereby, the most important gap is the absence of the data on the linearity of underlying protein quantification methods for plant matrices. To fill this gap, we address here the potential of FASP in combination with two protein isolation protocols for quantitative analysis of pea (Pisum sativum) seed and Arabidopsis thaliana leaf proteomes by the shotgun approach. For this aim, in comprehensive spiking experiments with bovine serum albumin (BSA), we evaluated the linear dynamic range (LDR) of protein quantification in the presence of plant matrices. Furthermore, we addressed the interference of two different plant matrices in quantitative experiments, accomplished with two alternative sample preparation workflows in comparison to conventional FASP-based digestion of cell lysates, considered here as a reference. The spiking experiments revealed high sensitivities (LODs of up to 4 fmol) for spiked BSA and LDRs of at least 0.6 × 10². Thereby, phenol extraction yielded slightly better recoveries, whereas the detergent-based method showed better linearity. Thus, our results indicate the very good applicability of FASP to quantitative plant proteomics with only limited impact of the protein isolation technique on the method's overall performance. [ABSTRACT FROM AUTHOR]

The natural environment of plants comprises a complex set of biotic and abiotic stresses, and plant responses to these stresses are complex as well. Plant proteomics approaches have significantly revealed dynamic changes in plant proteome responses to stress and developmental processes. Thus, we reviewed the recent advances in cotton proteomics research under changing environmental conditions, considering the progress and challenging factors. Finally, we highlight how single-cell proteomics is revolutionizing plant research at the proteomics level. We envision that future cotton proteomics research at the single-cell level will provide a more complete understanding of cotton's response to stresses. [ABSTRACT FROM AUTHOR]

Dalbergia nigra is an endangered species from the Brazilian Atlantic Rainforest, and in vitro propagation can be applied for its conservation. The aim of this study was to establish in vitro propagation and ex vitro rooting in D. nigra and evaluate the alterations in polyamines (PAs) and protein profiles during shoot development. The effects of MS and WPM culture media on in vitro germination were tested. For shoot induction, explants of apical and cotyledonary nodal segments from 45 day-old seedlings were inoculated in WPM culture medium supplemented with benzyladenine (BA; 0, 2.5 and 5 µM). Shoots obtained in vitro without and with 2.5 µM BA were rooted ex vitro with different concentrations (0, 100 and 500 µM) of indole-3-butyric acid (IBA). The seedlings grew best in WPM culture medium. Treatment with 2.5 µM BA significantly increased the shoot length by increasing free putrescine contents and the accumulation of proteins associated with shoot elongation, such as aspartate aminotransferase, elongation factor, calreticulin-3, and cell division cycle protein 48. Ex vitro rooting was obtained in all treatments, and the use of auxin was not necessary. The BA used for shoot multiplication significantly affected rooting, reducing the induction and number of roots but increasing their length. This study showed the relevance of cytokinin, PAs and proteomic profiles to in vitro shoot development, as well as the auxin and cytokinin balance to ex vitro rooting in D. nigra. Key message: BA significantly affected shoot multiplication and rooting and induced changes in PAs and proteomic profiles during micropropagation of Dalbergia nigra. [ABSTRACT FROM AUTHOR]

Due to its outstanding throughput and analytical resolution, gel-free LC-based shotgun proteomics represents the gold standard of proteome analysis. Thereby, the efficiency of sample preparation dramatically affects the correctness and reliability of protein quantification. Thus, the steps of protein isolation, solubilization, and proteolysis represent the principal bottleneck of shotgun proteomics. The desired performance of the sample preparation protocols can be achieved by the application of detergents. However, these compounds ultimately compromise reverse-phase chromatographic separation and disrupt electrospray ionization. Filter-aided sample preparation (FASP) represents an elegant approach to overcome these limitations. Although this method is comprehensively validated for cell proteomics, its applicability to plants and compatibility with plant-specific protein isolation protocols remain to be confirmed. Thereby, the most important gap is the absence of the data on the linearity of underlying protein quantification methods for plant matrices. To fill this gap, we address here the potential of FASP in combination with two protein isolation protocols for quantitative analysis of pea (Pisum sativum) seed and Arabidopsis thaliana leaf proteomes by the shotgun approach. For this aim, in comprehensive spiking experiments with bovine serum albumin (BSA), we evaluated the linear dynamic range (LDR) of protein quantification in the presence of plant matrices. Furthermore, we addressed the interference of two different plant matrices in quantitative experiments, accomplished with two alternative sample preparation workflows in comparison to conventional FASP-based digestion of cell lysates, considered here as a reference. The spiking experiments revealed high sensitivities (LODs of up to 4 fmol) for spiked BSA and LDRs of at least 0.6 × 102. Thereby, phenol extraction yielded slightly better recoveries, whereas the detergent-based method showed better linearity. Thus, our results indicate the very good applicability of FASP to quantitative plant proteomics with only limited impact of the protein isolation technique on the method’s overall performance.

The expression of heat shock proteins is considered a central adaptive mechanism to heat stress. This study investigated the expression of heat shock proteins (HSPs) and other stress-protective proteins against heat stress in cowpea genotypes under field (IT-96D-610 and IT-16) and controlled (IT-96D-610) conditions. Heat stress response analysis of proteins at 72 h in the controlled environment showed 270 differentially regulated proteins identified using label-free quantitative proteomics in IT-96D-610 plants. These plants expressed HSPs and chaperones [BAG family molecular chaperone 6 (BAG6), Multiprotein bridging factor1c (MBF1C) and cold shock domain protein 1 (CSDP1) in the controlled environment]. However, IT-96D-610 plants expressed a wider variety of small HSPs and more HSPs in the field. IT-96D-610 plants also responded to heat stress by exclusively expressing chaperones [DnaJ chaperones, universal stress protein and heat shock binding protein (HSBP)] and non-HSP proteins (Deg1, EGY3, ROS protective proteins, temperature-induced lipocalin and succinic dehydrogenase). Photosynthesis recovery and induction of proteins related to photosynthesis were better in IT-96D-610 because of the concurrent induction of heat stress response proteins for chaperone functions, protein degradation for repair and ROS scavenging proteins and PSII operating efficiency (Fq'/Fm') than IT-16. This study contributes to identification of thermotolerance mechanisms in cowpea that can be useful in knowledge-based crop improvement. [ABSTRACT FROM AUTHOR]

The expression of heat shock proteins is considered a central adaptive mechanism to heat stress. This study investigated the expression of heat shock proteins (HSPs) and other stress-protective proteins against heat stress in cowpea genotypes under field (IT-96D-610 and IT-16) and controlled (IT-96D-610) conditions. Heat stress response analysis of proteins at 72 h in the controlled environment showed 270 differentially regulated proteins identified using label-free quantitative proteomics in IT-96D-610 plants. These plants expressed HSPs and chaperones [BAG family molecular chaperone 6 (BAG6), Multiprotein bridging factor1c (MBF1C) and cold shock domain protein 1 (CSDP1) in the controlled environment]. However, IT-96D-610 plants expressed a wider variety of small HSPs and more HSPs in the field. IT-96D-610 plants also responded to heat stress by exclusively expressing chaperones [DnaJ chaperones, universal stress protein and heat shock binding protein (HSBP)] and non-HSP proteins (Deg1, EGY3, ROS protective proteins, temperature-induced lipocalin and succinic dehydrogenase). Photosynthesis recovery and induction of proteins related to photosynthesis were better in IT-96D-610 because of the concurrent induction of heat stress response proteins for chaperone functions, protein degradation for repair and ROS scavenging proteins and PSII operating efficiency (Fq’/Fm′) than IT-16. This study contributes to identification of thermotolerance mechanisms in cowpea that can be useful in knowledge-based crop improvement.

Information regarding the physiological and molecular plant responses to the treatment with new biofertilizers is limited. In this study, a fast-composting soil amendment obtained from solid waste by means of a Fenton reaction was assessed to evaluate the effects on the growth of Lactuca sativa L. var. longifolia seedlings. Growth rate, root biomass, chlorophyll concentration, and total soluble proteins of seedlings treated with the 2% fast-composting soil amendment showed significant increases in comparison with the control seedlings. Proteomic analysis revealed that the soil amendment induced the up-regulation of proteins belonging to photosynthesis machinery, carbohydrate metabolism, and promoted energy metabolism. Root proteomics indicated that the fast-composting soil amendment strongly induced the organs morphogenesis and development; root cap development, lateral root formation, and post-embryonic root morphogenesis were the main biological processes enriched by the treatment. Overall, our data suggest that the addition of the fast-composting soil amendment formulation to the base soils might ameliorate plant growth by inducing carbohydrate primary metabolism and the differentiation of a robust root system.

The influence of protein and starch profiles of chickpea, lentil, red lentil, white bean, quinoa, amaranth and oat flours on their techno-functional properties was studied in detail. Proteome of flours was approached through an affordable proteomic pipeline supported by liquid chromatography ion trap mass spectrometry (LC-MS) research coupled to quantitative polyacrylamide gel image analysis. Vicilins characterized pulse flours with a minimum of 45% of their total proteome and conferred their remarkable emulsifying, foaming and gelling capacities. Poor-vicilin quinoa (20% of total proteome) and vicilin-free amaranth and oat flours exhibited a good oil retention capacity that was exclusively provided by their high legumin content that comprised a minimum of 48% of their total proteome. Large starch values found in non-pulse flours (above 53% w/w versus less than 47% in pulse samples) mainly contributed to their noteworthy water holding capacity, freeze-thaw stability and high viscosity of pastes. This study can promote the creation of new insights on the use of alternative vegetable flours by industry to obtain innovative and sustainable high-protein food products. [Display omitted] • There was characterized proteome of seven vegetable flours. • Protein profiles of flours were correlated to their techno-functional properties. • Pulse flour vicilins conferred strong emulsion, foam and gelling capacities. • Legumins provided oil retention capacity in quinoa, amaranth and oat. [ABSTRACT FROM AUTHOR]

Food safety and quality are rising global issues while foodomics is rapidly becoming an important field of study, which promises significant outcomes related to food science due to the urgent need to address many of the most important questions facing humanity today. Food and plant proteomics as a tool of foodomics are relatively new entries in the field of food safety and quality as well as food bioactivity and health, which would yield new integrative and quantitative information, and consequently new functions of food and improved biomarkers for food safety and quality as well as bioactivity. The use of powerful analytical instruments such as nano-liquid chromatography (nano-LC) is needed in order to achieve its great analytical potential. Nano-LC coupled with mass spectrometry (Nano-LC/MS) has become the primary tool for proteomics as an indispensable technology for gaining understanding of complex food molecular processes. In recent decade, significant advances in relation to food and plant proteomics analysis by means of nano-LC/MS have been achieved. Among other advances, food nanoproteomics analyzing biological assemblies in food such as exosome, probiotics and microbial ingredients can reveal critical information in modern food science. This review article will focus on the role of nano-LC in food and plant proteomics analysis towards advanced foodomics. Special attention is given to recent food and plant proteomics applications and overview of recent studies (mostly the years between 2018-early 2024) by means of nano-LC/MS. In addition, it will give an overview of the current state of various food and plant proteomics strategies with discussion on future directions and perspectives of this technique as well as recent achievements in view of new findings regarding foodomics. • New advances in nano-LC in view of modern food science. • New approaches towards advanced foodomics. • Advanced food safety and quality applications. • Food nanoproteomics applications by nano-LC/MS. • Applications of new food biomaterials (e.g. exosomes). [ABSTRACT FROM AUTHOR]

Potentially toxic elements (PTEs) such as cadmium (Cd), lead (Pb), chromium (Cr), and arsenic (As), polluting the environment, pose a significant risk and cause a wide array of adverse changes in plant physiology. Above threshold accumulation of PTEs is alarming which makes them prone to ascend along the food chain, making their environmental prevention a critical intervention. On a global scale, current initiatives to remove the PTEs are costly and might lead to more pollution. An emerging technology that may help in the removal of PTEs is phytoremediation. Compared to traditional methods, phytoremediation is eco-friendly and less expensive. While many studies have reported several plants with high PTEs tolerance, uptake, and then storage capacity in their roots, stem, and leaves. However, the wide application of such a promising strategy still needs to be achieved, partly due to a poor understanding of the molecular mechanism at the proteome level controlling the phytoremediation process to optimize the plant's performance. The present study aims to discuss the detailed mechanism and proteomic response, which play pivotal roles in the uptake of PTEs from the environment into the plant's body, then scavenge/detoxify, and finally bioaccumulate the PTEs in different plant organs. In this review, the following aspects are highlighted as: (i) PTE's stress and phytoremediation strategies adopted by plants and (ii) PTEs induced expressional changes in the plant proteome more specifically with arsenic, cadmium, copper, chromium, mercury, and lead with models describing the metal uptake and plant proteome response. Recently, interest in the comparative proteomics study of plants exposed to PTEs toxicity results in appreciable progress in this area. This article overviews the proteomics approach to elucidate the mechanisms underlying plant's PTEs tolerance and bioaccumulation for optimized phytoremediation of polluted environments. [ABSTRACT FROM AUTHOR]

Abstract Background Aphid attack induces defense responses in plants activating several signaling cascades that led to the production of toxic, repellent or antinutritive compounds and the consequent reorganization of the plant primary metabolism. Pepper (Capsicum annuum L.) leaf proteomic response against Myzus persicae (Sulzer) has been investigated and analyzed by LC-MS/MS coupled with bioinformatics tools. Results Infestation with an initially low density (20 aphids/plant) of aphids restricted to a single leaf taking advantage of clip cages resulted in 6 differentially expressed proteins relative to control leaves (3 proteins at 2 days post-infestation and 3 proteins at 4 days post-infestation). Conversely, when plants were infested with a high density of infestation (200 aphids/plant) 140 proteins resulted differentially expressed relative to control leaves (97 proteins at 2 days post-infestation, 112 proteins at 4 days post-infestation and 105 proteins at 7 days post-infestation). The majority of proteins altered by aphid attack were involved in photosynthesis and photorespiration, oxidative stress, translation, protein folding and degradation and amino acid metabolism. Other proteins identified were involved in lipid, carbohydrate and hormone metabolism, transcription, transport, energy production and cell organization. However proteins directly involved in defense were scarce and were mostly downregulated in response to aphids. Conclusions The unexpectedly very low number of regulated proteins found in the experiment with a low aphid density suggests an active mitigation of plant defensive response by aphids or alternatively an aphid strategy to remain undetected by the plant. Under a high density of aphids, pepper leaf proteome however changed significantly revealing nearly all routes of plant primary metabolism being altered. Photosynthesis was so far the process with the highest number of proteins being regulated by the presence of aphids. In general, at short times of infestation (2 days) most of the altered proteins were upregulated. However, at longer times of infestation (7 days) the protein downregulation prevailed. Proteins involved in plant defense and in hormone signaling were scarce and mostly downregulated.

Proteomics offers one of the best approaches for the functional analysis of the genome, generating detailed information that can be integrated with that obtained by other classic and omics approaches. These results showed that maize seedlings rapidly respond to cold shock, providing a comprehensive landscape for the cold-responsive proteins and phosphoproteins in maize seedlings that can be a significant resource to understand how C SB 4 sb plants respond to a sudden temperature drop. Furthermore, post-translational modifications, subcellular localization, and protein-protein interactions provide deep insight into protein molecular functions. [Extracted from the article]

As one of the serious environmental problems worldwide, acid rain (AR) has always caused continuous damage to the forestry ecosystem. Studies have shown that AR can leach calcium ions from plants and soil. Calcium (Ca) is also a crucial regulator of the plant stress response, whereas there are few reports on how Ca regulates the response of AR-resistant woody plants to AR stress. In this study, by setting different exogenous Ca levels, we study the physiological and molecular mechanism of Ca in regulating the Taxus wallichiana var. mairei response to AR stress. Our results showed that low Ca level leads to photosynthesis, and antioxidant defense system decreases in T. wallichiana var. mairei leaves; however, these negative effects could be reversed at high Ca level. In addition, proteomic analyses identified 44 differentially expressed proteins in different Ca level treatments of T. wallichiana var. mairei under AR stress. These proteins were classified into seven groups, which include metabolic process, photosynthesis and energy pathway, cell rescue and defense, transcription and translation, protein modification and degradation, signal transduction, etc. Furthermore, the study found that low Ca level leads to an obvious increase of Ca-related gene expression under AR stress in T. wallichiana var. mairei using qRT-PCR analyses and however can be reversed at high Ca level. These findings would enrich and extend the Ca signaling pathways of AR stress in AR-resistant woody plants and are expected to have important theoretical and practical significance in revealing the mechanism of woody plants tolerating AR stress and protecting forestry ecosystem in soil environment under different Ca levels. [ABSTRACT FROM AUTHOR]

The protein interactome mediates crucial functions in transcription, chromatin remodeling, and higher-order structural organization. Elucidating the proteins that interact with chromatin-associated RNA or proteins is key to understanding fundamental epigenetic regulatory pathways. We discuss the opportunities and challenges of mass spectrometry (MS)-based proteomics for characterizing the plant chromatin interactome. [ABSTRACT FROM AUTHOR]

Proteins are intimately involved in executing and controlling virtually all cellular processes. To understand the molecular mechanisms that underlie plant phenotypes, it is essential to investigate protein expression, interactions, and modifications, to name a few. The proteome is highly dynamic in time and space, and a plethora of protein modifications, protein interactions, and network constellations are at play under specific conditions and developmental stages. Analysis of proteomes aims to characterize the entire protein complement of a particular cell type, tissue, or organism—a challenging task, given the dynamic nature of the proteome. Modern mass spectrometry–based proteomics technology can be used to address this complexity at a system-wide scale by the global identification and quantification of thousands of proteins. In this review, we present current methods and technologies employed in mass spectrometry–based proteomics and provide examples of dynamic changes in the plant proteome elucidated by proteomic approaches. [ABSTRACT FROM AUTHOR]

Long-term subculture plays an essential role in the large-scale multiplication and production of somatic plantlets. We investigated the effects of long-term subculture on in vitro shoot development and ex vitro rooting associated with changes in the hormones and protein profiles in Cedrela fissilis. The number of subcultures of shoots induced a decrease in the ex vitro rooting response. The reduction in adventitious root (AR) formation was associated with decreases in the contents of indole-3-acetic acid (IAA), abscisic acid (ABA), 12-oxo-phytodienoic acid (OPDA), putrescine (Put), and spermine and increases in jasmonic acid (JA), jasmonoyl-isoleucine, trans-cinnamic acid, and salicylic acid contents in shoots at the fourth subculture compared to the first. The ornithine decarboxylase enzyme preferentially functions in the Put biosynthesis pathway, and it was related to the highest AR formation in shoots at the first subculture. Down-accumulation of the auxin-binding protein ABP19a in shoots from the fourth subculture compared to the first subculture was related to a decrease in both the IAA content and AR formation. In addition, down-accumulation of glucose-6-phosphate isomerase; glutamine synthetase leaf isozyme, chloroplastic; 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase; l-ascorbate peroxidase, cytosolic; monodehydroascorbate reductase; and 2-Cys peroxiredoxin BAS1-like, chloroplastic and up-accumulation of caffeoyl-CoA O-methyltransferase 1 and isoforms of peroxidase 4 proteins in shoots from the fourth relative to the first subculture were associated with a reduction in AR formation. These results showed that the understanding of hormonal and molecular mechanisms related to the potential of AR formation in shoots under successive subcultures is relevant to improving large-scale plantlet production in C. fissilis. Key message: Long-term subculture affects the competence to ex vitro rooting of C. fissilis shoots by changes in endogenous hormones and protein profiles. [ABSTRACT FROM AUTHOR]

As one of the serious environmental problems worldwide, acid rain (AR) has always caused continuous damage to the forestry ecosystem. Studies have shown that AR can leach calcium ions from plants and soil. Calcium (Ca) is also a crucial regulator of the plant stress response, whereas there are few reports on how Ca regulates the response of AR-resistant woody plants to AR stress. In this study, by setting different exogenous Ca levels, we study the physiological and molecular mechanism of Ca in regulating the Taxus wallichiana var. mairei response to AR stress. Our results showed that low Ca level leads to photosynthesis, and antioxidant defense system decreases in T. wallichiana var. mairei leaves; however, these negative effects could be reversed at high Ca level. In addition, proteomic analyses identified 44 differentially expressed proteins in different Ca level treatments of T. wallichiana var. mairei under AR stress. These proteins were classified into seven groups, which include metabolic process, photosynthesis and energy pathway, cell rescue and defense, transcription and translation, protein modification and degradation, signal transduction, etc. Furthermore, the study found that low Ca level leads to an obvious increase of Ca-related gene expression under AR stress in T. wallichiana var. mairei using qRT-PCR analyses and however can be reversed at high Ca level. These findings would enrich and extend the Ca signaling pathways of AR stress in AR-resistant woody plants and are expected to have important theoretical and practical significance in revealing the mechanism of woody plants tolerating AR stress and protecting forestry ecosystem in soil environment under different Ca levels.

Background: Covalent modifications of core histones govern downstream DNA-templated processes such as transcription by altering chromatin structure and function. Previously, we reported that the plant homeodomain protein ALFIN-LIKE 6 (AL6), a bona fide histone reader that preferentially binds trimethylated lysin 4 on histone 3 (H3K4me3), is critical for recalibration of cellular phosphate (Pi) homeostasis and root hair elongation under Pi-deficient conditions. Results: Here, we demonstrate that AL6 is also involved in the response of Arabidopsis seedlings to jasmonic acid (JA) during skotomorphogenesis, possibly by modulating chromatin dynamics that affect the transcriptional regulation of JA-responsive genes. Dark-grown al6 seedlings showed a compromised reduction in hypocotyl elongation upon exogenously supplied JA, a response that was calibrated by the availability of Pi in the growth medium. A comparison of protein profiles between wild-type and al6 mutant seedlings using a quantitative Chromatin Enrichment for Proteomics (ChEP) approach, that we modified for plant tissue and designated ChEP-P (ChEP in Plants), yielded a comprehensive suite of chromatin-associated proteins and candidates that may be causative for the mutant phenotype. Conclusions: Altered abundance of proteins involved in chromatin organization in al6 seedlings suggests a role of AL6 in coordinating the deposition of histone variants upon perception of internal or environmental stimuli. Our study shows that ChEP-P is well suited to gain holistic insights into chromatin-related processes in plants. Data are available via ProteomeXchange with identifier PXD026541. [ABSTRACT FROM AUTHOR]

The cost-effective production of biofuels from lignocellulosic material will likely require manipulation of plant biomass, specifically cell walls. The North American native prairie grass Panicum virgatum (switchgrass) is seen as a potential biofuel crop with an array of genetic resources currently being developed. We have characterized the endomembrane proteome of switchgrass coleoptiles to provide additional information to the switchgrass community. In total, we identified 1750 unique proteins from two biological replicates. These data have been deposited in the ProteomeXchange with the identifier PXD001351 (http://proteomecentral.proteomexchange.org/dataset/PXD001351).

Metalloids are a subset of particular concern to risk assessors and toxicologists because of their well-documented potential hazards to plant system. Most of the metalloids are major environmental contaminants which affect crop productivity when present in high concentrations in soil. Metalloids are coupled with carrier proteins of the plasma membrane and translocated to various organs causing changes in key metabolic processes, damages cell biomolecules, and finally inhibit its growth. Phytoremediation-based approaches help in understanding the molecular and biochemical mechanisms for prerequisite recombinant genetic approaches. Recent advancements in proteomics and plant genomics help in understanding the role of transcription factors, metabolites, and genes in plant system which confers metal tolerance. The present review summarizes our current status of knowledge in this direction related to various physiological responses, detoxification mechanisms, and remediation strategies of metalloids in crop plants in relation to plant-metalloid tolerance. Further, the role of various transcription factors and miRNAs in conferring metal tolerance is also briefed. Hence, the present review mainly focused on the alterations in the physiological activities of plants due to metalloid toxicity and the various mechanisms which get activated inside the plants to mitigate their toxic effects. [ABSTRACT FROM AUTHOR]

The fruit and leaf of God's crown (Phaleria macrocarpa) have been traditionally used to treat a wide variety of diseases. However, the proteins of this tropical plant are still heavily understudied. Three protein extraction methods; phenol (Phe), trichloroacetic acid (TCA)–acetone–phenol (TCA–A–Phe), and ultrasonic (Ult) were compared on the fruit and leaf of P. macrocarpa. The Phe extraction method showed the highest percentage of recovered protein after the resolubilization process for both leaf (12.24%) and fruit (30.41%) based on protein yields of the leaf (6.15 mg/g) and fruit (36.98 mg/g). Phe and TCA–A–Phe extraction methods gave well‐resolved bands over a wide range of molecular weights through sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. Following liquid chromatography‐tandem mass spectrometry analysis, proteins identified through the Phe extraction method were 30%–35% enzymatic proteins, including oxidoreductases, transferases, hydrolases, lyases, isomerases, and ligases that possess various biological functions. Practical applications: Every part of God's crown plant is traditionally consumed to treat various illnesses. While plant's benefits are well known and have led to a plethora of health products, the proteome remains mostly unknown. This study compares three protein extraction methods for the leaf and fruit of P. macrocarpa and identifies their proteins thru LC‐MS/MS coupled with PEAKS. These method comparisons can be a guide for works on other plants as well. In addition, the proteomics data from this study may shed light on the functional properties of these plant parts and their products. [ABSTRACT FROM AUTHOR]

Exogenous application of double-stranded RNA (dsRNA) in the tobacco–Tobacco mosaic virus (TMV) pathosystem was shown previously to induce resistance against TMV providing an alternative approach to transgenesis. In the present study, we employed proteomics technology to elucidate the effect of TMV on tobacco as well as the effect of exogenous application of TMV p126 dsRNA molecules (dsRNAp126) at an early stage of the tobacco–TMV interaction. The proteome of tobacco leaf at 15 min post inoculation (mpi) in the presence or absence of dsRNAp126 molecules was studied. Thirty-six tobacco proteins were differentially accumulated in TMV-infected vs. healthy tobacco leaf tissue. The identified main differential TMV-responsive proteins were found to be involved in photosynthesis, energy metabolism, stress, and defense responses. Most of the virus-induced changes in the tobacco leaf proteome were not observed in the leaves treated with dsRNAp126 + TMV. The results indicated that the protein changes induced by TMV infection were counteracted by the exogenous application of dsRNAp126 molecules. Moreover, using small RNA sequencing, we showed that the exogenously applied dsRNAp126 was efficiently processed in tobacco as early as 15 min post application (mpa) to produce small interfering RNAs (siRNAs); the dicing pattern was not affected by the presence of TMV. The presence of dsRNAp126 reduced TMV p126 RNA abundance suggesting virus titer reduction via a sequence-specific mechanism, since a non-homologous dsRNA did not protect from TMV infection nor affect TMV accumulation.

Exogenous application of double-stranded RNA (dsRNA) in the tobacco– Tobacco mosaic virus (TMV) pathosystem was shown previously to induce resistance against TMV providing an alternative approach to transgenesis. In the present study, we employed proteomics technology to elucidate the effect of TMV on tobacco as well as the effect of exogenous application of TMV p126 dsRNA molecules (dsRNAp126) at an early stage of the tobacco–TMV interaction. The proteome of tobacco leaf at 15 min post inoculation (mpi) in the presence or absence of dsRNAp126 molecules was studied. Thirty-six tobacco proteins were differentially accumulated in TMV-infected vs. healthy tobacco leaf tissue. The identified main differential TMV-responsive proteins were found to be involved in photosynthesis, energy metabolism, stress, and defense responses. Most of the virus-induced changes in the tobacco leaf proteome were not observed in the leaves treated with dsRNAp126 + TMV. The results indicated that the protein changes induced by TMV infection were counteracted by the exogenous application of dsRNAp126 molecules. Moreover, using small RNA sequencing, we showed that the exogenously applied dsRNAp126 was efficiently processed in tobacco as early as 15 min post application (mpa) to produce small interfering RNAs (siRNAs); the dicing pattern was not affected by the presence of TMV. The presence of dsRNAp126 reduced TMV p126 RNA abundance suggesting virus titer reduction via a sequence-specific mechanism, since a non-homologous dsRNA did not protect from TMV infection nor affect TMV accumulation. [ABSTRACT FROM AUTHOR]

This review presents the view of the author, that is opinionable and even speculative, on the field of proteomics, its application to plant biology knowledge, and translation to biotechnology. Written in a more academic than scientific style, it is based on past original and review articles by the author´s group, and those published by leading scientists in the last two years. Starting with a general definition and references to historical milestones, it covers sections devoted to the different platforms employed, the plant biology discourse in the protein language, challenges and future prospects, ending with the author opinion. In 25 years, five proteomics platform generations have appeared. We are now moving from proteomics to Systems Biology. While feasible with model organisms, proteomics of orphan species remains challenging. Proteomics, even in its simplest approach, sheds light on plant biological processes, central dogma, and molecular bases of phenotypes of interest, and it can be translated to areas such as food traceability and allergen detection. Proteomics should be validated and optimized to each experimental system, objectives, and hypothesis. It has limitations, artifacts, and biases. We should not blindly accept proteomics data and just create a list of proteins, networks, and avoid speculative biological interpretations. From the hundred to thousand proteins identified and quantified, it is important to obtain a focus and validate some of them, otherwise it is merely. We are starting to have the protein pieces, so let, from now, build the proteomics and biological puzzle. [ABSTRACT FROM AUTHOR]

Background: Aphid attack induces defense responses in plants activating several signaling cascades that led to the production of toxic, repellent or antinutritive compounds and the consequent reorganization of the plant primary metabolism. Pepper (Capsicum annuum L.) leaf proteomic response against Myzus persicae (Sulzer) has been investigated and analyzed by LC-MS/MS coupled with bioinformatics tools. Results: Infestation with an initially low density (20 aphids/plant) of aphids restricted to a single leaf taking advantage of clip cages resulted in 6 differentially expressed proteins relative to control leaves (3 proteins at 2 days post-infestation and 3 proteins at 4 days post-infestation). Conversely, when plants were infested with a high density of infestation (200 aphids/plant) 140 proteins resulted differentially expressed relative to control leaves (97 proteins at 2 days post-infestation, 112 proteins at 4 days post-infestation and 105 proteins at 7 days post-infestation). The majority of proteins altered by aphid attack were involved in photosynthesis and photorespiration, oxidative stress, translation, protein folding and degradation and amino acid metabolism. Other proteins identified were involved in lipid, carbohydrate and hormone metabolism, transcription, transport, energy production and cell organization. However proteins directly involved in defense were scarce and were mostly downregulated in response to aphids. Conclusions: The unexpectedly very low number of regulated proteins found in the experiment with a low aphid density suggests an active mitigation of plant defensive response by aphids or alternatively an aphid strategy to remain undetected by the plant. Under a high density of aphids, pepper leaf proteome however changed significantly revealing nearly all routes of plant primary metabolism being altered. Photosynthesis was so far the process with the highest number of proteins being regulated by the presence of aphids. In general, at short times of infestation (2 days) most of the altered proteins were upregulated. However, at longer times of infestation (7 days) the protein downregulation prevailed. Proteins involved in plant defense and in hormone signaling were scarce and mostly downregulated. [ABSTRACT FROM AUTHOR]

Low-abundance proteins (LAPs) play a very important role in interaction, regulation, and metabolism of plant biological processes. A combinatorial peptide ligand library (CPLL) can solve the problem of high-abundance proteins (HAPs) masking LAPs and enlarging the dynamic range of protein concentrations perfectly and be considered as one of the most advanced approaches for plant proteomics research. In this paper, a proper CPLL method to rice leaf proteins was established for the first time and 1056 proteins were identified in rice leaf extracts, and 624 (59.1%) LAPs were newly detected after CPLL. Based on this technology, we detected the response of rice to Cd stress and analyzed the differential LAPs and the biological significance of misexpressed proteins before and after Cd stress by bioinformatics analysis. An important contribution has also been made to a better understanding of the complex mechanisms by which rice adapts to Cd stress. [ABSTRACT FROM AUTHOR]

Summary: Mitochondria host vital cellular functions, including oxidative phosphorylation and co‐factor biosynthesis, which are reflected in their proteome. At the cellular level plant mitochondria are organized into hundreds of discrete functional entities, which undergo dynamic fission and fusion. It is the individual organelle that operates in the living cell, yet biochemical and physiological assessments have exclusively focused on the characteristics of large populations of mitochondria. Here, we explore the protein composition of an individual average plant mitochondrion to deduce principles of functional and structural organisation. We perform proteomics on purified mitochondria from cultured heterotrophic Arabidopsis cells with intensity‐based absolute quantification and scale the dataset to the single organelle based on criteria that are justified by experimental evidence and theoretical considerations. We estimate that a total of 1.4 million protein molecules make up a single Arabidopsis mitochondrion on average. Copy numbers of the individual proteins span five orders of magnitude, ranging from >40 000 for Voltage‐Dependent Anion Channel 1 to sub‐stoichiometric copy numbers, i.e. less than a single copy per single mitochondrion, for several pentatricopeptide repeat proteins that modify mitochondrial transcripts. For our analysis, we consider the physical and chemical constraints of the single organelle and discuss prominent features of mitochondrial architecture, protein biogenesis, oxidative phosphorylation, metabolism, antioxidant defence, genome maintenance, gene expression, and dynamics. While assessing the limitations of our considerations, we exemplify how our understanding of biochemical function and structural organization of plant mitochondria can be connected in order to obtain global and specific insights into how organelles work. Significance Statement: We explore the structure and function of a single plant mitochondrion by scaling down a quantitative proteomic dataset of Arabidopsis cell culture mitochondria to calculate the copy numbers of individual protein species for a single organelle. [ABSTRACT FROM AUTHOR]

Proteomics approach was used mainly in the samples for microorganisms and animal tissues until early 2000s, the period during which the plant proteomics studies were scarcely available. Tough nature of cell membranes, existence of complex and diverse range of secondary metabolites, high abundance of pigments, proteases, polyphenols, polysaccharides and lipids caused a number of problems for preparation and resolution of the protein samples. However, proteomics approach in plant area had become widespread with the help of continuous efforts in order to overcome these difficulties. This review is prepared comprehensively from the preparation of the samples to the identification of the proteins and it aims to serve as a detailed source for the researchers interested in plant proteomics, especially for young investigators. [ABSTRACT FROM AUTHOR]