Publications of NIPGR Scientists

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Author: search.filters.author.Senthil-Kumar, MuthappaSubject: search.filters.subject.Arabidopsis thaliana

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Now showing 1 - 7 of 7
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    A method for isolation and transfection of arabidopsis protoplast for sucrose feeding assay
    (Springer Nature Publishing AG, 2026) Anjali, Anjali; Senthil-Kumar, Muthappa
    Protoplasts serve as a powerful system to study various plant physiological processes and to understand crucial signaling pathways within the plant system. The isolation of Arabidopsis protoplasts is a well-established technique and being utilized for wide range of assays. The method described herein includes the precise cutting of Arabidopsis leaves using extraction buffer containing cellulase and macerozyme. Further we present a polyethylene glycol [PEG]-mediated protoplast transformation method. Here, we have elucidated the methodology for isolation of protoplast for the AtSWEET-mediated sucrose uptake assay in control, and Pseudomonas syringe pv tomato DC3000 (Pst DC3000) treated leaves by utilizing GC/MS analysis. Our approach includes certain modifications to the previously published protoplast isolation technique, streamlining the process and providing a more accessible alternative to the highly specialized Xenopus oocyte uptake assay.
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    Drought attenuates plant defence against bacterial pathogens by suppressing the expression of CBP60g/SARD1 during combined stress
    (John Wiley & Sons, 2022) Choudhary, Aanchal; Senthil-Kumar, Muthappa
    In nature, plants are frequently exposed to drought and bacterial pathogens simultaneously. However, information on how the drought and defence pathways interact and orchestrate global transcriptional regulation is limited. Here, we show that moderate drought stress enhances the susceptibility of Arabidopsis thaliana to Pseudomonas syringae pv. tomato DC3000. Using transcriptome meta-analysis, we found that drought and bacterial stress antagonistically modulate a large set of genes predominantly involved in salicylic acid (SA) and abscisic acid (ABA) signalling networks. We identified that the levels of SA and ABA are dynamically regulated during the course of stress. Importantly, under combined stress, drought through the ABA pathway downregulates the induction of CALMODULIN-BINDING PROTEIN 60g (CBP60g) and SYSTEMIC ACQUIRED RESISTANCE DEFICIENT 1 (SARD1), two transcription factors crucial for SA production upon bacterial infection. We also identified an important role of NPR1-LIKE PROTEIN 3 and 4 (NPR3/4) transcriptional repressors in the drought-mediated negative regulation of CBP60g/SARD1 expression. Using a genetic approach, we show that CBP60g/SARD1 expression is the key determinant of plant defence against bacterial pathogens under combined stress. Thus, these transcription factors act as critical nodes for the crosstalk between drought and bacterial stress signalling under combined stress in plants.
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    Investigation of the novel transcriptional changes under combined drought and bacterial stress underpins the role of AtMYB96 in imparting tolerance
    (Springer Nature Publishing AG, 2021) Choudhary, Aanchal; Senthil-Kumar, Muthappa
    The physiological and molecular responses instigated to combat drought and bacterial pathogens often work antagonistically and, in most cases, the impact of combined stress is more detrimental to plant growth. Interestingly, plants exposed to this stress combination show a novel transcriptome fingerprint with a significant set of genes that are uniquely altered under combined stress. Despite this being reported in several transcriptomic datasets, our molecular understanding of these combined stress-specific genes and pathways is still in its nascent stages. These unique genes and the dedicated regulatory pathways are important for understanding the molecular aspects of signaling responses under combined stress. In this study, a previously available microarray dataset was extensively reanalysed to identify the novel genes and pathways specifically altered under combined stress. Using a combination of bioinformatic and data-guided approaches, we identified major biological pathways, transcription factor (TF) network and novel motifs potentially involved in the combined stress-specific responses. The candidate gene analysis using gene expression and mutant analysis identified AtMYB96 as an important TF involved in combined stress response. Taken together, our analysis pinpoints many novel genes that can be used for understanding the molecular mechanism of how plants deal with the combination of drought and bacterial pathogen.
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    An efficient, high-throughput method for the simultaneous exposure of drought stress and bacterial infection in plants
    (John Wiley & Sons, 2020) Choudhary, Aanchal; Senthil-Kumar, Muthappa
    PREMISE: We developed a systematic protocol for the easy, high-throughput, qualitative, and quantitative assessment of the patho-morphological, physiological, and molecular responses of Arabidopsis thaliana plants simultaneously subjected to drought and bacterial infection. This approach will assist studies elucidating plant adaptation strategies to combat combined stresses. METHODS AND RESULTS: Plants were grown in small screw-capped containers, individual pots, or pot strips. Watering was withheld from 30-day-old plants, which were subsequently infected with Pseudomonas syringae pv. tomato DC3000 using a dip inoculation. The natural development of both drought and bacterial infection was successfully and rapidly replicated in large numbers of plants, which is difficult to achieve with existing protocols. CONCLUSIONS: Our protocol offers a simple, low-cost, high-throughput strategy for the rapid and easy bacterial infection of large numbers of plants. It can be used in large-scale mutant and ecotype screenings under combined stresses and for other foliar pathogens in different plant species.
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    GBF3 transcription factor imparts drought tolerance in Arabidopsis thaliana
    (Nature Publishing Group, 2017) Ramegowda, Venkategowda; Gill, Upinder Singh; Sivalingam, Palaiyur Nanjappan; Gupta, Aarti; Gupta, Chirag; Govind, Geetha; Nataraja, Karaba N.; Pereira, Andy; Udayakumar, Makarla; Mysore, Kirankumar S.; Senthil-Kumar, Muthappa
    Drought transcriptome analysis of finger millet (Eleusine coracana) by cDNA subtraction identified drought responsive genes that have a potential role in drought tolerance. Through virus-induced gene silencing (VIGS) in a related crop species, maize (Zea mays), several genes, including a G-BOX BINDING FACTOR 3 (GBF3) were identified as candidate drought stress response genes and the role of GBF3 in drought tolerance was studied in Arabidopsis thaliana. Overexpression of both EcGBF3 and AtGBF3 in A. thaliana resulted in improved tolerance to osmotic stress, salinity and drought stress in addition to conferring insensitivity to ABA. Conversely, loss of function of this gene increased the sensitivity of A. thaliana plants to drought stress. EcGBF3 transgenic A. thaliana results also suggest that drought tolerance of sensitive plants can be improved by transferring genes from far related crops like finger millet. Our results demonstrate the role of GBF3 in imparting drought tolerance in A. thaliana and indicate the conserved role of this gene in drought and other abiotic stress tolerance in several plant species.
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    Transcriptome changes in Arabidopsis thaliana infected with Pseudomonas syringae during drought recovery
    (Nature Publishing Group, 2017) Gupta, Aarti; Senthil-Kumar, Muthappa
    Field-grown plants experience cycles of drought stress and recovery due to variation in soil moisture status. Physiological, biochemical and transcriptome responses instigated by recovery are expected to be different from drought stress and non-stressed state. Such responses can further aid or antagonize the plant’s interaction with the pathogen. However, at molecular level, not much is known about plantpathogen interaction during drought recovery. In the present study, we performed a microarray-based global transcriptome profiling and demonstrated the existence of unique transcriptional changes in Arabidopsis thaliana inoculated with Pseudomonas syringae pv. tomato DC3000 at the time of drought recovery (drought recovery pathogen, DRP) when compared to the individual drought (D) or pathogen (P) or drought recovery (DR). Furthermore, the comparison of DRP with D or DR and P transcriptome revealed the presence of a few common genes among three treatments. Notably, a gene encoding proline dehydrogenase (AtProDH1) was found to be commonly up-regulated under drought recovery (DR), DRP and P stresses. We also report an up-regulation of pyrroline-5-carboxylate biosynthesis pathway during recovery. We propose that AtProDH1 influences the defense pathways during DRP. Altogether, this study provides insight into the understanding of defense responses that operate in pathogen-infected plants during drought recovery.
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    Global profiling of phytohormone dynamics during combined drought and pathogen stress in Arabidopsis thaliana reveals ABA and JA as major regulators
    (Nature Publishing Group, 2017) Gupta, Aarti; Hisano, Hiroshi; Hojo, Yuko; Matsuura, Takakazu; Ikeda, Yoko; Mori, Izumi C.; Senthil-Kumar, Muthappa
    Global transcriptome studies demonstrated the existence of unique plant responses under combined stress which are otherwise not seen during individual stresses. In order to combat combined stress plants use signaling pathways and ‘cross talk’ mediated by hormones involved in stress and growth related processes. However, interactions among hormones’ pathways in combined stressed plants are not yet known. Here we studied dynamics of different hormones under individual and combined drought and pathogen infection in Arabidopsis thaliana by liquid chromatography-mass spectrometry (LC-MS) based profiling. Our results revealed abscisic acid (ABA) and salicylic acid (SA) as key regulators under individual drought and pathogen stress respectively. Under combined drought and host pathogen stress (DH) we observed non-induced levels of ABA with an upsurge in SA and jasmonic acid (JA) concentrations, underscoring their role in basal tolerance against host pathogen. Under a non-host pathogen interaction with drought (DNH) stressed plants, ABA, SA and JA profiles were similar to those under DH or non-host pathogen alone. We propose that plants use SA/JA dependent signaling during DH stress which antagonize ABA biosynthesis and signaling pathways during early stage of stress. The study provides insights into hormone modulation at different time points during combined stress.