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Author: search.filters.author.Mishra, ShrutiSubject: search.filters.subject.Arabidopsis

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    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.
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    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/