Dissection of grasspea (Lathyrus sativus L.) root exoproteome reveals critical insights and novel proteins.

The plant exoproteome is crucial because its constituents greatly influence plant phenotype by regulating physiological characteristics to adapt to environmental stresses. The root exudates constitute a dynamic aspect of plant exoproteome, as its molecular composition ensures a beneficial rhizosphere in a species-specific manner. We investigated the root exoproteome of grasspea, a stress-resilient pulse and identified 2861 non-redundant proteins, belonging to a myriad of functional classes, including root development, rhizosphere augmentation as well as defense functions against soil-borne pathogens. Significantly, we identified 1986 novel exoproteome constituents of grasspea, potentially involved in cell-to-cell communication and root meristem maintenance, among other critical roles. Sequence-based comparison revealed that grasspea shares less than 30 % of its exoproteome with the reports so far from model plants as well as crop species. Further, the exoproteome revealed 65 % proteins to be extracellular in nature and of these, 37 % constituents were predicted to follow unconventional protein secretion (UPS) mode. We validated the UPS for four stress-responsive proteins, which were otherwise predicted to follow classical protein secretion (CPS). Conclusively, we recognized not only the highest number of root exudate proteins, but also pinpointed novel signatures of dicot root exoproteome.

Proteomic dissection of rice cytoskeleton reveals the dominance of microtubule and microfilament proteins, and novel components in the cytoskeleton-bound polysome.

The plant cytoskeleton persistently undergoes remodeling to achieve its roles in supporting cell division, differentiation, cell expansion and organelle transport. However, the links between cell metabolism and cytoskeletal networks, particularly how the proteinaceous components execute such processes remain poorly understood. We investigated the cytoskeletal proteome landscape of rice to gain better understanding of such events. Proteins were extracted from highly enriched cytoskeletal fraction of four-week-old rice seedlings, and the purity of the fraction was stringently monitored. A total of 2577 non-redundant proteins were identified using both gel-based and gel-free approaches, which constitutes the most comprehensive dataset, thus far, for plant cytoskeleton. The data set includes both microtubule and microfilament-associated proteins and their binding proteins comprising hypothetical as well as novel cytoskeletal proteins. Further, various in-silico analyses were performed, and the proteins were functionally classified on the basis of their gene ontology. The catalogued proteins were validated through their sequence analysis. Extensive comparative analysis of our dataset with the non-redundant set of cytoskeletal proteins across plant species affirms unique as well as overlapping candidates. Together, these findings unveil new insights of how cytoskeletons undergo dynamic remodeling in rice to drive seedling development processes in rapidly changing in planta environment.

Physiological plasticity to high temperature stress in chickpea: Adaptive responses and variable tolerance.

High temperature stress (HTS) is one of the most crucial factors that limits plant growth and development, and reduces crop yields worldwide. Cool-season crops, particularly the legumes, are severely affected by increasing ambient temperature associated with global climate change. We characterized the HTS-induced modulations of morpho-physicochemical traits and gene expression of several chickpea genotypes and the metabolic profile of the tolerant cultivar. Higher water use efficiency and photosynthetic capacity, minimal membrane lipid peroxidation in conjunction with increased abundance of osmolytes and secondary metabolites depicted thermotolerance of ICC 1205. The adaptive responses were accompanied by high transcript abundance of heat shock proteins and antioxidant enzymes. To integrate stress-responsive signalling and metabolic networks, the HTS-induced physicochemical analysis was further extended to metabolite profiling of the thermotolerant cultivar. The screening of the metabolome landscape led to the identification of 49 HTS-responsive metabolites that include polycarboxylic acid, sugar acids, sugar alcohols and amino acids which might confer thermotolerance in chickpea. The present study, to our knowledge, is the most comprehensive of its kind in dissecting cultivar-specific differential adaptive responses to HTS in chickpea, which might potentiate the identification of genetic traits extendible to improvement of thermotolerance of crops.

Metabolite signatures of grasspea suspension-cultured cells illustrate the complexity of dehydration response.

MAIN CONCLUSION: This represents the first report deciphering the dehydration response of suspension-cultured cells of a crop species, highlighting unique and shared pathways, and adaptive mechanisms via profiling of 330 metabolites. Grasspea, being a hardy legume, is an ideal model system to study stress tolerance mechanisms in plants. In this study, we investigated the dehydration-responsive metabolome in grasspea suspension-cultured cells (SCCs) to identify the unique and shared metabolites crucial in imparting dehydration tolerance. To reveal the dehydration-induced metabolite signatures, SCCs of grasspea were exposed to 10% PEG, followed by metabolomic profiling. Chromatographic separation by HPLC coupled with MRM-MS led to the identification of 330 metabolites, designated dehydration-responsive metabolites (DRMs), which belonged to 28 varied functional classes. The metabolome was found to be constituted by carboxylic acids (17%), amino acids (13.5%), flavonoids (10.9%) and plant growth regulators (10%), among others. Pathway enrichment analysis revealed predominance of metabolites involved in phytohormone biosynthesis, secondary metabolism and osmotic adjustment. Exogenous application of DRMs, arbutin and acetylcholine, displayed improved physiological status in stress-resilient grasspea as well as hypersensitive pea, while administration of lauric acid imparted detrimental effects. This represents the first report on stress-induced metabolomic landscape of a crop species via a suspension culture system, which would provide new insights into the molecular mechanism of stress responses and adaptation in crop species.

Transcriptome profiling illustrates expression signatures of dehydration tolerance in developing grasspea seedlings.

MAIN CONCLUSION: This study highlights dehydration-mediated temporal changes in physicochemical, transcriptome and metabolome profiles indicating altered gene expression and metabolic shifts, underlying endurance and adaptation to stress tolerance in the marginalized crop, grasspea. Grasspea, often regarded as an orphan legume, is recognized to be fairly tolerant to water-deficit stress. In the present study, 3-week-old grasspea seedlings were subjected to dehydration by withholding water over a period of 144 h. While there were no detectable phenotypic changes in the seedlings till 48 h, the symptoms appeared during 72 h and aggravated upon prolonged dehydration. The physiological responses to water-deficit stress during 72-96 h displayed a decrease in pigments, disruption in membrane integrity and osmotic imbalance. We evaluated the temporal effects of dehydration at the transcriptome and metabolome levels. In total, 5201 genes of various functional classes including transcription factors, cytoplasmic enzymes and structural cell wall proteins, among others, were found to be dehydration-responsive. Further, metabolome profiling revealed 59 dehydration-responsive metabolites including sugar alcohols and amino acids. Despite the lack of genome information of grasspea, the time course of physicochemical and molecular responses suggest a synchronized dehydration response. The cross-species comparison of the transcriptomes and metabolomes with other legumes provides evidence for marked molecular diversity. We propose a hypothetical model that highlights novel biomarkers and explain their relevance in dehydration-response, which would facilitate targeted breeding and aid in commencing crop improvement efforts.

Variety-specific nutrient acquisition and dehydration-induced proteomic landscape of grasspea (Lathyrus sativus L.).

Grasspea, a stress-resilient pulse crop, has largely remained outside the realm of phytochemical and functional genomics analyses despite its high nutritional significance. To unravel the intervarietal variability in nutrient acquisition of grasspea, we conducted a series of physicochemical experiments using two cultivated varieties, LP-24 and Prateek. The analyses revealed high percentage of starch, cellulose, peroxides, carotenoids, phytic acid and minerals in cv. LP-24, whereas large amounts of protein, soluble carbohydrates and antioxidants in Prateek. To dissect the mechanism of stress tolerance, 3-week-old seedlings of cv. LP-24 and Prateek were afflicted with dehydration for a period of 144 h. The physicochemical indices indicated better adaptation in cv. LP-24, with high abundance of proline, phenolics and flavonoids. Dehydration-responsive proteome landscape of cv. LP-24 revealed 152 proteins with variance at a statistically 94% significance level. The comparative proteomics analysis led to the identification of 120 dehydration-responsive proteins (DRPs), most of which were associated with carbohydrate metabolism, amino acid synthesis, antioxidant reactions and cell defense. We report, for the first time, the dehydration-induced proteome landscape of grasspea, whose genome is yet to be sequenced. The results provide unique insights into variety-specific nutrient acquisition attributes and dehydration-tolerance of grasspea. BIOLOGICAL SIGNIFICANCE: Grasspea is a great source of protein and antioxidants with nitrogen fixing ability, besides its tolerance to multivariate environmental stress as compared to major legume species. This represents the first report on nutrient profile and health-promoting attributes of grasspea. The cultivars under study are nutritionally enriched that possess high protein, amino acids and health-promoting factors and may therefore be projected as a vital part of a healthy diet. Grasspea is known for its hardy nature, water-use efficiency and efficacy as a stress-tolerant pulse. Further, this study portrays the dehydration-responsive proteomic landscape of grasspea. The proteomics analyses provide crucial insights into the dehydration response, presumably orchestrated by proteins belonging to an array of functional classes including photosynthesis, protein and RNA metabolism, protein folding, antioxidant enzymes and defense. The interplay of the differentially regulated proteins might aid in reinforcing the mechanisms of dehydration avoidance and/or tolerance.