Browsing by Author "Elagamey, Eman"

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Chitosan-triggered immunity to Fusarium in chickpea is associated with changes in the plant extracellular matrix architecture, stomatal closure and remodelling of the plant metabolome and proteome
(John Wiley & Sons, 2020) Narula, Kanika; Elagamey, Eman; Abdellatef, Magdi A.E.; Sinha, Arunima; Ghosh, Sudip; Chakraborty, Niranjan; Chakraborty, Subhra
Pathogen/microbe associated molecular patterns (PAMPs/MAMPs) initiate complex defense responses by reorganizing the biomolecular dynamics of the host cellular machinery. The extracellular matrix (ECM) acts as a physical scaffold that prevents recognition and entry of phyto‐pathogens, while guard cells perceive and integrate signals metabolically. Although chitosan is known MAMP implicated in plant defense, the precise mechanism of chitosan‐triggered immunity (CTI) remains unknown. Here, we show how chitosan imparts immunity against fungal disease. Morpho‐histological examination revealed stomatal closure accompanied by reductions in stomatal conductance and transpiration rate as early responses in chitosan‐treated seedlings upon vascular fusariosis. Electron microscopy and Raman spectroscopy showed ECM fortification leading to oligosaccharide signaling, as documented by increased galactose, pectin and associated secondary metabolites. Multiomics approach using quantitative ECM proteomics and metabolomics identified 325 chitosan‐triggered immune‐responsive proteins (CTIRPs) notably novel ECM structural proteins, LYM2 and receptor‐like kinases, and 65 chitosan‐triggered immune‐responsive metabolites (CTIRMs), including sugars, sugar alcohols, fatty alcohols, organic and amino acids. Identified proteins and metabolites are linked to ROS production, stomatal movement, root nodule development and root architecture coupled with oligosaccharide signaling that leads to Fusarium resistance. The cumulative data demonstrate that ROS, NO and eATP govern CTI, in addition to induction of PR proteins, CAZymes and PAL activities, besides accumulation of phenolic compounds downstream of CTI. The immune‐related correlation network identified functional hubs in the CTI pathway. Altogether, these shifts led to the discovery of chitosan‐responsive networks that cause significant ECM and guard cell remodeling and translate ECM cues into cell fate decisions during fusariosis.
Combining extracellular matrix proteome and phosphoproteome of chickpea and meta-analysis reveal novel proteoforms and evolutionary significance of clade-specific wall-associated events in plant
(John Wiley & Sons, 2024) Narula, Kanika; Sinha, Arunima; Choudhary, Pooja; Ghosh, Sudip; Elagamey, Eman; Sharma, Archana; Sengupta, Atreyee; Chakraborty, Niranjan; Chakraborty, Subhra
Extracellular matrix (ECM) plays central roles in cell architecture, innate defense andcell wall integrity (CWI) signaling. During transition to multicellularity, modulardomain structures of ECM proteins and proteoforms have evolved due to continuousadaptation across taxonomic clades under different ecological niche. Although thisincredible diversity has to some extent been investigated at protein level, extracellu-lar phosphorylation events and molecular evolution of ECM proteoform familiesremains unexplored. We developed matrisome proteoform atlas in a grain legume,chickpea and performed meta-analyses of 74 plant matrisomes. MS/MS analysisidentified 1,424 proteins and 315 phosphoproteins involved in diverse functions.Cross-species ECM protein network identified proteoforms associated with CWImaintenance system. Phylogenetic characterization of eighteen matrix protein fami-lies highlighted the role of taxon-specific paralogs and orthologs. Novel informationwas acquired on gene expansion and loss, co-divergence, sub functionalization andneofunctionalization during evolution. Modular networks of matrix protein familiesand hub proteins showed higher diversity across taxonomic clades than amongorgans. Furthermore, protein families differ in nonsynonymous to synonymous sub-stitution rates. Our study pointed towards the matrix proteoform functionality,sequence divergence variation, interactions between wall remodelers and molecularevolution using a phylogenetic framework. This is the first report on comprehensivematrisome proteoform network illustrating presence of CWI signaling proteins inland plants.
Comparative analyses of extracellular matrix proteome: an under-explored area in plant research
(intechopen.com, 2012) Narula, Kanika; Elagamey, Eman; Datta, Asis; Chakraborty, Niranjan; Chakraborty, Subhra
Within their social milieu, cells are petite and deformable, enclosed in a flimsy plasma membrane which swerves from their default spherical shape to more polar shapes due to the local deposition, complex interactions and the remodelling of the extracellular matrix (ECM). Consequently, multicellularity has evolved, albeit independently in plants and animals. Although animals are truly multicellular, plants are supracellular organisms because their immobile cells divide via phragmoplast-based incomplete cytokinesis, which results in the formation of cytoplasmic cell-to-cell channels known as plasmodesmata (Baluska et al., 2003). The ECM in plants, often referred as the cell wall, is integrated into the apoplast—a structurally coherent superstructure extending throughout the plant body. In lieu, plant cells are not fully separated and both the plasma membrane and endoplasmic reticulum traverse cellular borders through plasmodesmata (Baluska et al., 2003; Fincher, G. 2009.). The ECM is a fundamental component of the microenvironment of both animal and plant cells that has been substantially expanded during evolution. Throughout the plant kingdom, the formation and regulation of the ECM architecture has been shown to have the potential to influence many conduits of development, position-dependent differentiation, patterning and totipotent cell niches, besides environmental stress response and pathobiology (Brownlee & Berger, 1995; Degenhardt & Gimmer, 2000; Wilson, 2010). Furthermore, it has been reported that the ECM plays an important morphoregulatory role during somatic embryogenesis and organogenesis in plants, besides its pivotal role in cellular osmo- and volume-regulation (Šamaj et al., 1999; Rose et al., 2004). The plant ECM has biomechanical and morphogenetic functions with the immense ability to turn cells into hydraulic machines which establish a crucial functional difference between cell walls and other cellular surface structures. It encloses the cell hermetically and constrains the hydrostatic pressure evoked by osmotic gradients between the cell and its environment which controls cellular osmo- and volume-regulation (Peters et al., 2000; Cosgrove, D. J. 2005). Plasticity in the ECM allows the cellular uptake of massive amounts of water into a central vacuole while rigidity in the ECM determines the conductance of enormous amounts of water and dissolved solutes through vascular bundles. The secretion of an ECM by one cell can also influence the neighbouring cells, conceivably the best exemplified paracrine interaction known in the plant kingdom (for a review, see Brownlee, 2002). Beyond their paramount importance in the generation of form, cell walls are frequently considered ‘growth-controlling’ (Wolf et al., 2009). Cells devoid of the ECM inevitably lose their polar shape and the loss of cellular polarisation prevents cell-to-cell interactions and communication. The ECM/cell wall is evolutionary and inherently bestowed with information that can be both stored and relayed to cell interior via templating processes. It serves as the first line mediator in cell signalling for perceiving and transmitting extra- and intercellular signals in many cellular pathways. Communication between the cytoplasm and the cell wall is necessary and evident because of events such as cell expansion (Cosgrove, 1997, Schröder, F et al 2009), mechanical stress (Kumar et al., 2006; Telewski, 2006), environmental perturbation (Gail McLean et al., 1997; Thelen, J. and Peck, S. 2007) and pathogen infection (Hammond- Kosack & Jones, 1996) which lead to altered biosynthesis and the modification of wall components and downstream cytoplasmic events. In addition, it can act as a substrate for migration and has also been recognised as a surrogate for providing inputs into cell behaviour (Hall et al., 2002), although the available data is rather scarce for higher plants and critical linker molecules between the cytoskeleton and the ECM are still missing. Thus, the ECM/cell wall primarily serves a dual function, as a cell support system and for signalling during development and stress. The ECM/cell wall must therefore be dynamic as cells divide and elongate, modulating its composition and architecture during its synthesis and after it has been deposited. The wall function is a multi-step, complex process and the underlying mechanisms governing these steps are not fully understood.
Comparative nuclear proteomics analysis provides insight into the mechanism of signaling and immune response to blast disease caused by Magnoporthe oryzae in rice
(John Wiley & Sons, 2019) Narula, Kanika; Choudhary, Pooja; Ghosh, Sudip; Elagamey, Eman; Chakraborty, Niranjan; Chakraborty, Subhra
Modulation of plant immune system by extrinsic/intrinsic factors and host-specific determinants fine-tunes cellular components involving multiple organelles, particularly nucleus to mount resistance against pathogen attack. Rice blast, caused by hemibiotrophic fungus Magnaporthe oryzae, is one of the most devastating diseases that adversely affect rice productivity. However, the role of nuclear proteins and their regulation in response to M. oryzae remains unknown. Here, we elucidate the nucleus-associated immune pathways in blast resistant rice genotype. Temporal analysis of nuclear proteome was carried out using 2-DE coupled MS/MS analysis. A total of 140 immune responsive proteins (IRPs) were identified associated with nuclear reorganization, cell division, energy production/deprivation, signaling and gene regulation. We interrogated the proteome data using correlation network analysis that identified significant functional modules pointing towards immune related coinciding processes through a common mechanism of remodelling and homeostasis. Novel clues regarding blast resistance include nucleus associated redox homeostasis and glycolytic enzyme mediated chromatin organization which manipulates cell division and immunity. Taken together, our study provides evidence that coordination of nuclear function and reprogramming of host translational machinery regulate resistance mechanism against blast disease.
Extracellular matrix proteome and phosphoproteome of potato reveals functionally distinct and diverse canonical and non-canonical proteoforms
(MDPI AG, 2016) Elagamey, Eman; Narula, Kanika; Sinha, Arunima; Aggarwal, Pooja Rani; Ghosh, Sudip; Chakraborty, Niranjan; Chakraborty, Subhra
The extracellular matrix (ECM) has a molecular machinery composed of diverse proteins and proteoforms that combine properties of tensile strength with extensibility exhibiting growth-regulatory functions and self- and non-self-recognition. The identification of ECM proteoforms is the prerequisite towards a comprehensive understanding of biological functions accomplished by the outermost layer of the cell. Regulatory mechanisms of protein functions rely on post-translational modifications, phosphorylation in particular, affecting enzymatic activity, interaction, localization and stability. To investigate the ECM proteoforms, we have isolated the cell wall proteome and phosphoproteome of a tuberous crop, potato (Solanum tuberosum). LC-MS/MS analysis led to the identification of 38 proteins and 35 phosphoproteins of known and unknown functions. The findings may provide a better understanding of biochemical machinery and the integrated protein and phosphoprotein network of ECM for future functional studies of different developmental pathways and guidance cues in mechanosensing and integrity signaling.
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.
Molecular dissection of extracellular matrix proteome reveals discrete mechanism regulating Verticillium dahliae triggered vascular wilt disease in potato
(John Wiley & Sons, 2017) Elagamey, Eman; Sinha, Arunima; Narula, Kanika; Abdellatef, Magdi A.E.; Chakraborty, Niranjan; Chakraborty, Subhra
Plants exposed to patho-stress mostly succumb due to disease by disruption of cellular integrity and changes in the composition of the extracellular matrix (ECM). Vascular wilt, caused by the soil borne hemibiotrophic filamentous fungus Verticillium dahliae, is one of the most significant diseases that adversely affects plant growth and productivity. The virulence of the pathogen associated with the ECM-related susceptibility of the host plant is far from being understood. To better understand ECM-associated disease responses that allow the pathogen to supress plant immunity, a temporal analysis of ECM proteome was carried out in vascular wilt susceptible potato cultivar upon V. dahliae infection. The proteome profiling led to the identification of 75 patho-stress responsive proteins (PSRPs), predominantly involved in wall hydration, architecture, and redox homeostasis. Two novel clues regarding wilt disease of potato were gained from this study. First, wall crosslinking and salicylic acid signaling significantly altered during patho-stress. Second, generation of reactive oxygen species and scavenging proteins increased in abundance leading to cell death and necrosis of the host. We provide evidence for the first time that how fungal invasion affects the integrity of ECM components and host reprogramming for susceptibility may function at the cell surface by protein plasticity. This article is protected by copyright. All rights reserved.
Proteomic signatures uncover phenotypic plasticity of susceptible and resistant genotypes by wall remodelers in rice blast
(John Wiley & Sons, 2024) Sinha, Arunima; Narula, Kanika; Bhola, Latika; Sengupta, Atreyee; Choudhary, Pooja; Nalwa, Pragya; Kumar, Mohit; Elagamey, Eman; Chakraborty, Niranjan; Chakraborty, Subhra
Molecular communication between macromolecules dictates extracellular matrix (ECM) dynamics during pathogen recognition and disease development. Extensive research has shed light on how plant immune components are activated, regulated and function in response to pathogen attack. However, two key questions remain largely unresolved: (i) how does ECM dynamics govern susceptibility and disease resistance, (ii) what are the components that underpin these phenomena? Rice blast, caused by Magnaporthe oryzae adversely affects rice productivity. To understand ECM regulated genotype-phenotype plasticity in blast disease, we temporally profiled two contrasting rice genotypes in disease and immune state. Morpho-histological, biochemical and electron microscopy analyses revealed that increased necrotic lesions accompanied by electrolyte leakage governs disease state. Wall carbohydrate quantification showed changes in pectin level was more significant in blast susceptible compared to blast resistant cultivar. Temporally resolved quantitative disease- and immune-responsive ECM proteomes identified 308 and 334 proteins, respectively involved in wall remodelling and integrity, signalling and disease/immune response. Pairwise comparisons between time and treatment, messenger ribonucleic acid expression, diseasome and immunome networks revealed novel blast-related functional modules. Data demonstrated accumulation of α-galactosidase and phosphatase were associated with disease state, while reactive oxygen species, induction of Lysin motif proteins, CAZymes and extracellular Ca-receptor protein govern immune state.
Quantitative extracellular matrix proteomics suggests cell wall reprogramming in host-specific immunity during vascular wilt caused by Fusarium oxysporum in chickpea
(John Wiley & Sons, 2017) Elagamey, Eman; Narula, Kanika; Sinha, Arunima; Ghosh, Sudip; Abdellatef, Magdi A.E.; Chakraborty, Niranjan; Chakraborty, Subhra
Extracellular matrix (ECM) is the unique organelle that perceives stress signals and reprograms molecular events of host cell during patho-stress. However, our understanding of how ECM dictates plant immunity is largely unknown. Vascular wilt caused by the soil borne filamentous fungus Fusarium oxysporum is a major impediment for global crop productivity. To elucidate the role of ECM proteins and molecular mechanism associated with cell wall mediated immunity, the temporal changes of ECM proteome was studied in vascular wilt resistant chickpea cultivar upon F. oxysporum infection. The 2-DE protein profiling coupled with mass spectrometric analysis identified 166 immune responsive proteins (IRPs) involved in variety of functions. Our data suggest that wall remodeling; protein translocation, stabilization and chitin triggered immunity; extracellular ATP signaling are major players in early, middle and later phases of ECM signaling during fungal attack. Furthermore, we interrogated the proteome data using network analysis that identified modules enriched in known and novel immunity-related prognostic proteins centered around nascent aminopolypeptide complex (NAC), amine oxidase, thioredoxin and chaperonin. This study for the first time provides an insight into the complex network operating in the ECM and impinges on the surveillance mechanism of innate immunity during patho-stress in crop plant.