Publications of NIPGR Scientists
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Item 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, MuthappaIn 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.Item 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, MuthappaThe 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.Item 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, MuthappaPREMISE: 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.Item ath-miR164c influences plant responses to the combined stress of drought and bacterial infection by regulating proline metabolism(Elsevier B.V., 2020) Gupta, Aarti; Patil, Mahesh; Qamar, Aarzoo; Senthil-Kumar, MuthappaPlants under combined stresses exhibit a prominent shift in molecular responses compared with plants exposed to the same stresses independently. Profiling responses to individual and combined stressors at the gene expression level have identified several genes with intersecting responses to these stressors. However, the upstream regulators at the intersection of plant responses to individual and combined stresses are not known. Here, using the transcriptome of Arabidopsis thaliana under individual and combined drought and Pseudomonas syringae infection, we identified several genes whose expression overlaps between individual and combined stresses. To study the key regulator of such an overlapping gene, we predicted that the expression of 1-Pyrroline-5-carboxylate synthase 1 (AtP5CS1) is regulated by ath-miR164c at post-transcriptional level. Our results from the stem-loop RT-PCR based expression analysis revealed significant downregulation of ath-miR164c in response to P. syringae infection under both well-irrigated (pathogen only) and drought stress (combined stress) conditions. Furthermore, an Arabidopsis loss-of-function mutant of the miRNA ath-miR164c exhibited resistance to pathogen infection under combined stress, unlike the wild-type plants, implicating the role of ath-miR164c in regulating plant immunity. AtP5CS1 gene expression and proline accumulation were enhanced in the ath-miR164c mutant plants relative to the wild-type plants, demonstrating that ath-miR164c regulates AtP5CS1 of the proline biosynthesis pathway, which was also validated by 5’RLM-RACE results. This miRNA-mediated modulation of AtP5CS1 gene expression under combined stress fills crucial gaps in identifying the key convergent players in the current understanding of plant stress responses.Item AtGBF3 confers tolerance to Arabidopsis thaliana against combined drought and Pseudomonas syringae stress(Elsevier B.V., 2019) Dixit, Sandeep Kumar; Gupta, Aarti; Fatima, Urooj; Senthil-Kumar, MuthappaIn field conditions, plants are often exposed to a combination of abiotic and biotic stresses, for instance, drought and pathogen infection. Transcriptome studies on Arabidopsis thaliana and other plants under individual and combined drought and pathogen stresses have unveiled the activation of shared molecular defense mechanisms. These shared plant responses are characterized by commonly regulated genes under both individual as well as combined stresses. Therefore, the identification of commonly regulated genes during individual and combined stress conditions can reveal plant responses towards combined stress. Available transcriptome studies on combined-stressed plants have hinted at G-Box Binding Factor 3 (GBF3) as one of the regulatory components of the shared response. However, the mechanistic understanding of the role of AtGBF3 under combined drought and pathogen stress is not yet decoded. In the current study, we used genetic approaches to identify the role of AtGBF3 in conferring tolerance to individual and combined drought and pathogen stress. Atgbf3 mutant plants showed increased susceptibility, while AtGBF3-overexpressing plants were tolerant under individual and combined drought and Pseudomonas syringae pv. tomato infection stresses as compared to wild-type plants. We further analyzed the global transcriptome of Atgbf3 mutant plants under combined stress to identify its downstream targets. We also established a high-throughput method to apply combined polyethylene glycol and pathogen stress on Murashige and Skoog medium-grown plants to further validate the role of AtGBF3 in combined stress.Item Transcriptome changes in Arabidopsis thaliana infected with Pseudomonas syringae during drought recovery(Nature Publishing Group, 2017) Gupta, Aarti; Senthil-Kumar, MuthappaField-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.Item Global transcriptional analysis reveals unique and shared responses in Arabidopsis thaliana exposed to combined drought and pathogen stress(Frontiers Media S.A., 2016) Gupta, Aarti; Sarkar, Ananda K.; Senthil-Kumar, MuthappaWith frequent fluctuations in global climate, plants are exposed to co-occurring drought and pathogen infection and this combination adversely affects plant survival. In the past, some studies indicated that morpho-physiological responses of plants to the combined stress are different from the individual stressed plants. However, interaction of drought stressed plants with pathogen has not been widely studied at molecular level. Such studies are important to understand the defense pathways that operate as part of combined stress tolerance mechanism. In this study, Arabidopsis thaliana was exposed to individual drought stress, Pseudomonas syringae pv tomato DC3000 (Pst DC3000) infection and their combination. Using Affymetrix WT gene 1.0 ST array, global transcriptome profiling of leaves under individual drought stress and pathogen infection was compared with their combination. The results obtained from pathway mapping (KAAS and MAPMAN) demonstrated the modulation in defense pathways in A. thaliana under drought and host pathogen Pst DC3000 infection. Further, our study revealed ‘tailored’ responses under combined stress and the time of occurrence of each stress during their concurrence has showed differences in transcriptome profile. Our results from microarray and RT-qPCR revealed unique regulation of 20 novel genes exclusively during the stress interaction. This study indicates that plants exposed to concurrent drought and pathogen stress experience a new state of stress. Thus, under frequently changing climatic conditions each combination of stressor and their timing defines the plant responses and should thus be studied explicitly.