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    Isolation of cytoskeleton and cytoskeleton-bound polysome, and identification of cytoskeletal proteins from a grain legume
    (Springer Nature Publishing AG, 2026) Kumar, Sunil; Chakraborty, Sohela; Chakraborty, Subhra; Chakraborty, Niranjan
    The plant cytoskeleton is an essential component of cellular architecture, enabling various critical metabolic processes, including cell division, differentiation, expansion and intracellular transport. It consists primarily of three distinct filamentous structures: microtubules, microfilaments and intermediate filaments. These structures are not static; they undergo continuous remodeling in response to environmental signals and developmental cues, which allow plants to adapt to changing conditions. The microtubules and actin filaments have previously been successfully isolated from various plant tissues, contributing to our understanding of their functions. Among the diverse plant families, legumes (Fabaceae) stand out as the third largest, encompassing approximately 20,000 species. They hold significant agricultural importance, ranking second to cereals in global crop production. To fully grasp the developmental and adaptive processes in legumes, it is essential to identify and understand their regulatory components. This chapter focuses on the isolation of cytoskeletal proteins from chickpea, a prominent grain legume, facilitating biochemical and proteomic analyses that may uncover new insights into the functioning of the cytoskeleton in legumes.
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    Proteo-metabolomic dissection of extracellular matrix reveals alterations in cell wall integrity and calcium signaling governs wall-associated susceptibility during stem rot disease in jute
    (American Chemical Society, 2024) Arafat, Md Yasir; Narula, Kanika; Kumar, Mohit; Chakraborty, Niranjan; Chakraborty, Subhra
    The plant surveillance system confers specificity to disease and immune states by activating distinct molecular pathways linked to cellular functionality. The extracellular matrix (ECM), a preformed passive barrier, is dynamically remodeled at sites of interaction with pathogenic microbes. Stem rot, caused by Macrophomina phaseolina, adversely affects fiber production in jute. However, how wall related susceptibility affects the ECM proteome and metabolome remains undetermined in bast fiber crops. Here, stem rot responsive quantitative temporal ECM proteome and metabolome were developed in jute upon M. phaseolina infection. Morpho-histological examination revealed that leaf shredding was accompanied by reactive oxygen species production in patho-stressed jute. Electron microscopy showed disease progression and ECM architecture remodeling due to necrosis in the later phase of fungal attack. Using isobaric tags for relative and absolute quantitative proteomics and liquid chromatography-tandem mass spectrometry, we identified 415 disease-responsive proteins involved in wall integrity, acidification, proteostasis, hydration, and redox homeostasis. The disease-related correlation network identified functional hubs centered on α-galactosidase, pectinesterase, and thaumatin. Gas chromatography–mass spectrometry analysis pointed toward enrichment of disease-responsive metabolites associated with the glutathione pathway, TCA cycle, and cutin, suberin, and wax metabolism. Data demonstrated that wall-degrading enzymes, structural carbohydrates, and calcium signaling govern rot responsive wall-susceptibility. Proteomics data were deposited in Pride (PXD046937; PXD046939).
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    Extracellular matrix proteome: Isolation of ECM proteins for proteomics studies
    (Springer Nature Publishing AG, 2020) Elagamey, Eman; Narula, Kanika; Chakraborty, Niranjan; Chakraborty, Subhra
    Understanding molecular mechanisms and cellular metabolism in varied plant processes necessitates knowledge of the expressed proteins and their subcellular distribution. Spatial partitioning of organelles generates an enclosed milieu for physiochemical reactions designed and tightly linked to a specific organelle function. Of which, extracellular matrix (ECM)/cell wall (CW) is a dynamic and chemically active compartment. The ECM proteins are organized into complex structural and functional networks involved in several metabolic processes, including carbon and nitrogen metabolism. Organellar proteomics aim for comprehensive identification of resident proteins that rely on the isolation of highly purified organelle free from contamination by other intracellular components. Extraction and isolation of plant ECM proteins features key caveats due to the lack of adjoining membrane, the presence of a polysaccharide–protein network that traps contaminants, and the existence of high phenolic content. Furthermore, due to diverse biochemical forces, including labile, weakly bound and strongly bound protein in the protein–polysaccharide matrix different elution procedures are required to enrich ECM proteins. Here, we describe a method that allows efficient fractionation of plant ECM, extraction of ECM proteins and protein profiling from variety of crop plants, including rice, chickpea and potato. This method can easily be adapted to other plant species for varied experimental conditions.
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    Heat shock proteins and abiotic stress tolerance in plants
    (Springer Nature, 2018) Mishra, Divya; Shekhar, Shubhendu; Singh, Deepika; Chakraborty, Subhra; Chakraborty, Niranjan
    Abiotic stresses restrict plant growth and development, and reduce harvest index of many crop species worldwide. Maintenance of native conformation of proteins and reducing the accumulation of non-native proteins are imperative for survival under stress conditions as such stresses frequently lead to protein aggregation causing metabolic dysfunction. Heat shock proteins (HSP) play a key role in conferring abiotic stress tolerance. Plants protect themselves from numerous stresses by inducing HSP, besides some stress-responsive proteins, suggesting analogous response mechanisms. A close association between the HSP and ROS also co-exists, indicating that plants have evolved to gain a higher degree of regulation over ROS toxicity and can use ROS as elicitor to induce HSP for better adaptations through activating an array of molecules. Therefore, unraveling the mechanisms of plant response against various stress and the role of HSP in acquired stress tolerance is utmost important to delineate their specific function as a part of stress-responsive module. The HSP have been well characterized in different crop species, albeit the knowledge about their correlation with genome sequence information as well as their functional plasticity is limited.
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    RNA-seq analysis identifies key genes involved in chickpea (Cicer arietinum L.) shoot development
    (Botanical Society of Bengal, 2016) Biswas, S; Aggarwal, PR; Tayal, R; Sarkar, MP; Chakraborty, Niranjan; Chakraborty, Subhra
    Understanding the mechanism of plant development is particularly important for studying the regulatory genes and their specific functions. Here, we used paired end RNA sequencing to understand the complexity of legume shoot development in a non-model crop chickpea (Cicer arietinum L.). A total of 149.5 million paired end reads were obtained from two developmental stages of chickpea shoot. Differential gene expression analysis revealed the enrichment of transcripts associated with shoot development. Validation of few genes showed the reproducibility of our RNA-seq data. Collectively, the present study will not only help to generate a comprehensive organ specific transcriptome map but will also serve as a resource for gene discovery and future crop improvement program.
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    Plant fungus interaction proteomics: An update
    (Springer, 2016) Narula, Kanika; Aggarwal, Pooja R.; Chakraborty, Niranjan; Chakraborty, Subhra
    Diversity of angiosperm is renowned and mechanism of perception and interaction with different environmental conditions is also variable. Patho-stress response in different plant families varies during the invasion of same or different fungal species. A major puzzle is how interaction and communication could increase fitness in plant at molecular level. Global proteome analysis of plant-pathosystem provides an invaluable resource for the identification of host as well as pathogen proteins involved in disease progression or immunity development. At protein level plant-fungal interaction upsurge the need to understand protein homeostasis and molecular adaptation of building blocks of cell to manifest natural selection for the host. Here, we examine the multilayered facets of interaction between organisms of two diverse kingdoms, namely plant and fungi at protein level based on more than 3000 identified host proteins till date.