Browsing by Author "Mishra, Shruti"

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The Arabidopsis eATP receptor DORN1 and CNGC19 calcium channel act in tandem to regulate plant defense upon Spodoptera litura herbivory
(Oxford University Press, 2025) Kundu, Pritha; Kumari, Misha; Meena, Mukesh Kumar; Mishra, Shruti; Vadassery, Jyothilakshmi
Plants deploy cellular Ca2+ elevation as a signal for environmental stress signaling. Extracellular ATP (eATP) is released into the extracellular matrix when cells are wounded. DOES NOT RESPOND TO NUCLEOTIDES 1 (DORN1), a key legume-type lectin receptor, senses and binds eATP and activates Ca2+ signaling. No evidence directly links calcium-mediated eATP signaling to resistance against insect herbivores. Here, we report upregulation of DORN1 transcripts upon wounding and Spodoptera litura feeding in Arabidopsis. Loss-of-function of DORN1 resulted in increased S. litura feeding compared to that on wildtype. Plant immunity is compromised in dorn1 mutants as they show reduced S. litura oral secretion mediated Ca2+ elevation, jasmonic acid accumulation, and expression of jasmonate responsive genes. The herbivory-induced calcium channel, CYCLIC NUCLEOTIDE GATED CHANNEL 19 (CNGC19), co-expresses with DORN1. We found that eATP-induced Ca2+ elevation requires functional CNGC19. Loss-of-function of DORN1 and CNGC19 highly increased the susceptibility to S. litura, mediated by reduced accumulation of jasmonates. We also demonstrate a plausible interaction of CNGC19 with DORN1. The data implicate the role of damage-released eATP and its receptor DORN1 in herbivory-induced defense signaling. DORN1 together with the Ca2+ channel CNGC19 generate the eATP-induced Ca2+ elevation, leading to the activation of immune signaling.
Deep metabolomics revealed trajectories of jasmonate signaling-mediated primary metabolism in Arabidopsis upon Spodoptera litura herbivory
(John Wiley & Sons, 2025) Kundu, Anish; Bera, Paramita; Mishra, Shruti; Vadassery, Jyothilakshmi
Plants defend against chewing herbivores by up-regulating jasmonic acid (JA) signaling, which activates downstream signaling cascades and produces numerous secondary metabolites that act as defense molecules against the herbivores. Although secondary metabolism always remains a focus of research, primary metabolism is also reported to be realigned upon herbivory. However, JA signaling-mediated modulation of primary metabolites and their metabolic pathways in plants are mostly unexplored. Here, we applied gas chromatography–mass spectrometry-based untargeted metabolomics aided with computational statistical frameworks on wild type Arabidopsis, mutants of active JA receptor (i.e., CORONATINE-INSENSITIVE 1, COI1-1) and downstream transcription factor (i.e., MYC2) to navigate the JA signaling-mediated primary metabolism alterations during herbivory. Pathway and metabolite's chemical class enrichment analysis revealed JA signaling is crucial for constitutive as well as herbivore-induced primary metabolism and topology of their interaction networks. JA signaling majorly modulated alterations of sugars, amino acids and related metabolites. Herbivory-mediated sugar depletion and induction of methionine for aliphatic glucosinolates are also dependent on JA signaling. Taken together, our results demonstrate trails of JA signaling-mediated primary metabolic alterations associated with herbivory.
External jasmonic acid isoleucine mediates amplification of plant elicitor peptide receptor (PEPR) and jasmonate-based immune signalling
(John Wiley & Sons, 2024) Mittal, Deepika; Gautam, Janesh Kumar; Varma, Mahendra; Laie, Amrutha; Mishra, Shruti; Behera, Smrutisanjita; Vadassery, Jyothilakshmi
Jasmonic acid-isoleucine (JA-Ile) is a plant defence hormone whose cellular levels are elevated upon herbivory and regulate defence signalling. Despite their pivotal role, our understanding of the rapid cellular perception of bioactive JA-Ile is limited. This study identifies cell type-specific JA-Ile-induced Ca2+ signal and its role in self-amplification and plant elicitor peptide receptor (PEPR)-mediated signalling. Using the Ca2+ reporter, R-GECO1 in Arabidopsis, we have characterized a monophasic and sustained JA-Ile-dependent Ca2+ signature in leaf epidermal cells. The rapid Ca2+ signal is independent of positive feedback by the JA-Ile receptor, COI1 and the transporter, JAT1. Microarray analysis identified up-regulation of receptors, PEPR1 and PEPR2 upon JA-Ile treatment. The pepr1 pepr2 double mutant in R-GECO1 background exhibits impaired external JA-Ile induced Ca2+cyt elevation and impacts the canonical JA-Ile responsive genes. JA responsive transcription factor, MYC2 binds to the G-Box motif of PEPR1 and PEPR2 promoter and activates their expression upon JA-Ile treatment and in myc2 mutant, this is reduced. External JA-Ile amplifies AtPep-PEPR pathway by increasing the AtPep precursor, PROPEP expression. Our work shows a previously unknown non-canonical PEPR-JA-Ile-Ca2+-MYC2 signalling module through which plants sense JA-Ile rapidly to amplify both AtPep-PEPR and jasmonate signalling in undamaged cells.
Forward genetic screen using transgenic calcium reporter aequorin to identify novel targets in calcium signaling
(Journal of Visualized Experiment, 2020) Mittal, Deepika; Mishra, Shruti; Prajapati, Ramgopal; Vadassery, Jyothilakshmi
Forward genetic screens have been important tools in the unbiased identification of genetic components involved in several biological pathways. The basis of the screen is to generate a mutant population that can be screened with a phenotype of interest. EMS (ethyl methane sulfonate) is a commonly used alkylating agent for inducing random mutation in a classical forward genetic screen to identify multiple genes involved in any given process. Cytosolic calcium (Ca2+) elevation is a key early signaling pathway that is activated upon stress perception. However the identity of receptors, channels, pumps and transporters of Ca2+ is still elusive in many study systems. Aequorin is a cellular calcium reporter protein isolated from Aequorea victoria and stably expressed in Arabidopsis. Exploiting this, we designed a forward genetic screen in which we EMS-mutagenized the aequorin transgenic. The seeds from the mutant plants were collected (M1) and screening for the phenotype of interest was carried out in the segregating (M2) population. Using a 96-well high-throughput Ca2+ measurement protocol, several novel mutants can be identified that have a varying calcium response and are measured in real time. The mutants with the phenotype of interest are rescued and propagated till a homozygous mutant plant population is obtained. This protocol provides a method for forward genetic screens in Ca2+ reporter background and identify novel Ca2+ regulated targets. The video component of this article can be found at https://www.jove.com/video/61259/
Piriformospora indica recruits host-derived putrescine for growth promotion in plants
(Oxford University Press, 2022) Kundu, Anish; Mishra, Shruti; Kundu, Pritha; Jogawat, Abhimanyu; Vadassery, Jyothilakshmi
Growth promotion induced by the endosymbiont Piriformospora indica has been observed in various plants; however, except growth phytohormones, specific functional metabolites involved in P. indica-mediated growth promotion are unknown. Here, we used a GC-MS based untargeted metabolite analysis to identify tomato (Solanum lycopersicum) metabolites whose levels were altered during P. indica-mediated growth promotion. Metabolomic multivariate analysis revealed several primary metabolites with altered levels, with putrescine induced most significantly in roots during the interaction. Further, our results indicated that P. indica modulates the arginine decarboxylase (ADC)-mediated putrescine biosynthesis pathway via induction of SlADC1 in tomato. P. indica did not promote growth in Sladc1-VIGS (virus-induced gene silencing of SlADC1) lines of tomato tomato and showed less colonization. Furthermore, using LC-MS/MS we showed that putrescine promoted growth by elevation of auxin (indole-3-acetic acid) and gibberellin (GA4, GA7) levels in tomato. In Arabidopsis (Arabidopsis thaliana) adc knock-out mutants, P. indica colonization also decreased and showed no plant growth promotion, and this response was rescued upon exogenous application of putrescine. Putrescine is also important for hyphal growth of P. indica, indicating that it is co-adapted by both host and microbe. Taken together, we conclude that putrescine is an essential metabolite and its biosynthesis in plants is crucial for P. indica-mediated plant growth promotion and fungal growth.
Regulatory role of phytohormones in the interaction of plants with insect herbivores
(Elsevier B.V., 2023) Kundu, Pritha; Bera, Paramita; Mishra, Shruti; Vadassery, Jyothilakshmi
Insect herbivores affect crop growth worldwide, causing significant yield losses annually. Phytohormones are prime signaling molecules that coordinate many physiological phenomena, besides triggering stress-responsive regulatory genes involved in plant defense. Here, we present a holistic picture of the central role of various phytohormones—jasmonic acid, salicylic acid, ethylene, abscisic acid, auxin, cytokinin, gibberellins, and brassinosteroids in regulating the diverse molecular interactions between plants and insect herbivores. With an initial overview of the early signaling response, the chapter majorly focuses on the role of different key phytohormones in defending plants against herbivory and their cross talk. We also describe the phytohormone-mediated production of secondary metabolites and other defense toxins to counteract harmful herbivores. With the well-established myriad of information in selected plant–herbivore model systems, it is now critical to translate this knowledge to less explored crop plants for sustainable development in agriculture.