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Integrating physiological and proteomics signatures for understanding drought, salinity and alkalinity stress tolerance mechansisms in lentil

ABSTRACT: Abiotic stresses—such as drought, salinity, and alkalinity severely limit global lentil production by impairing physiological processes including osmolytes accumulation, chlorophyll synthesis, H2O2 production and antioxidant activities. However, no proteomic signature studies have dissected the molecular mechanisms underlying tolerance to drought, salinity and alkalinity in lentil. In this study, proteomic profiling revealed differentially abundant proteins (DAPs) distinguishing tolerant from sensitive lines under multiple abiotic stress conditions including drought, salinity and alkalinity. Furthermore, the aforementioned physiological traits were phenotyped to compare the responses of tolerant versus sensitive genotypes toward these stresses. By integrating proteomics and physiological analyses, this study provides new insights into the physiological and molecular regulatory mechanisms of lentil in response to multiple abiotic stresses.Physiological analysis revealed genotypic differences in osmolyte regulation, chlorophyll content, H2O2 production and antioxidant activities. Ultra performance liquid chromatography with tandem mass spectrometery (UPLC-MS/MS) analysis identified 707 differentially abundant proteins (DAPs) between tolerant v/s sensitive genotypes across stresses, with 283, 213 and 211 DAPs under drought, salinity and alkalinity, respectively. Ninety-five DAPs involved in protein biogenesis, cellular transport, photosynthesis, RNA regulation, defense and detoxification were common across stresses. Proteomic findings were validated by quantitative real-time polymerase chain reaction (qRT-PCR) of five DAPs at 1, 2 and 3 days post-stress. Functional annotation revealed enrichment in photosynthesis/energy metabolism, transcription/translational, protein folding/degradation, and antioxidative defense. Integrative transcriptomic and proteomic analysis highlighted superoxide dismutase and monodehydroascorbate reductase as central components of antioxidative defense. This represents the first proteomics study of lentil seedling response toward drought, salinity and alkalinity stresses.The differential abundance of proteins involved in varied regulatory pathways suggests a wide, yet complex network of proteins mediating lentil’s metabolic adaptation to drought, salinity and alkalinity stresses. Proteins identified in this study will provide new insight to lentil’s interconnected regulatory mechanisms against abiotic stresses.

INSTRUMENT(S):

ORGANISM(S): Lens Culinaris

TISSUE(S): Plant Cell, Leaf

DISEASE(S): Disease Free

SUBMITTER: Dharmendra Singh

LAB HEAD: SINGH DHARMENDRA

PROVIDER: PXD078023 | Pride | 2026-06-15

REPOSITORIES: Pride

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Project description:Purpose: To identify high temperature, sailinity and drought-responsive miRNAs at genome wide level in B. juncea var varuna. Results: In this study four small RNA libraries viz. B. juncea control (BJC), B. juncea high temperature stressed (BJH), B. juncea salinity stressed (BJS) and B. juncea drought stressed (BJD) were prepared and sequenced. With the help of UEA sRNA workbench software package 51 conserved miRNAs belonging to 30 miRNA families were identified. As there was limited genomic information available for B. juncea, we generated and assembled its genome sequence at a very low coverage. Using the generated sequence and other publically available Brassica genomic/transcriptomic resources as mapping reference, 126 novel (not reported so far in any plant species) were discovered for the first time in B. juncea. Further analysis also revealed existence of 32 and 37 star sequences for conserved and novel miRNAs, respectively. The expression of a few selected conserved and novel miRNAs under conditions of different abiotic stresses was revalidated through universal TaqMan based real time PCR. Putative targets of identified conserved and novel miRNAs were predicted in B. rapa to gain insights into functional roles manifested by B. juncea miRNAs. Furthermore, SPL2-like, ARF17-like and a NAC domain containing protein were experimentally validated as targets of miR156, miR160 and miR164 respectively. Investigation of gene ontologies linked with targets of known and novel miRNAs forecasted their involvement in various biological functions. Conclusions: We have generated in this study, the first comprehensive abiotic stress influenced small RNA dataset in B. juncea. The combinatorial approach of NGS and computational methods led to the discovery of 51 conserved and 126 novel miRNAs. The present study provides a holistic view of B. juncea miRNAome under conditions of high temperature, salinity and drought. The catalogue of miRNA sequences, their expression and putative targets, generated in this study can be utilized to design crop improvement strategies in B. juncea and related species. Total four small RNA libraries were prepared and sequenced independently [B. juncea control (BJC), B. juncea high temperature stressed (BJH), B. juncea salinity stressed (BJS) and B. juncea drought stressed (BJD)] on Illumina GAIIx

Project description:In low rainfall regions soils are naturally conditioned with frequent co-occurrence of salinity and alkalinity. Plant salinity responses both at physiological and molecular level have been extensively researched. However, effects of the combined treatment of alkaline salinity that could greatly reduce plant growth and the mechanisms responsible for tolerance remain indeterminate. In Brassica juncea, large reductions in biomass and increased leaf Na+ concentration under alkaline salinity indicates that the combined treatment had greater negative effect than salinity on both growth and the physiological responses of the plant. To determine molecular mechanisms potentially controlling adaptive tolerance responses to salinity and alkaline salinity, the moderately tolerant genotype NDR 8501 was further investigated using microarray analysis. The transcripts of treated leaf tissues verses those of the untreated control sample were analysed after prolonged stress of four weeks. In total, 528 salinity responsive and 1245 alkaline salinity responsive genes were indentified and only 101 genes were expressed jointly in either of the two treatments. Transcription of 37% more genes involved in response to alkaline salinity than salinity alone, which suggests the increased impact and severity of the combined stress on the plant, indicating the transcription of a far greater number of genes likely involved in mitigation and damage control. Transcription of KUP2 and KUP7 genes involved in potassium homeostasis under salinity alone and NHX1 and ENH1 genes for ion (K+ and Na+) homeostasis under alkaline salinity, clearly demonstrated that different genes and genetic pathways are involved in response to each stress. They further provide supporting evidence for the physiological responses that occur in the plant, with massive reprogramming of the transcriptome leading to partial ion exclusion, shuttling and compartmentation. Salinity and alkaline salinity induced gene expression in Brassica juncea leaf was measured at 4 weeks of prolonged treatment of 50 mM NaCl alone and combined with 2.5 mM HCO3- versus non-stressed control. A single experiment was conducted using Brassica juncea genotype NDR 8501 at a single time point (fours weeks after treatment) with three replications per treatment.

			Action	DRS
	A1C-A1T.xlsx	Xlsx
	A1C_A1T.fasta	Fasta
	A1T-A2T.xlsx	Xlsx
	A1T_A2T.fasta	Fasta
	A2C-A2T.xlsx	Xlsx

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Integrating physiological and proteomics signatures for understanding drought, salinity and alkalinity stress tolerance mechansisms in lentil

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