Publications of NIPGR Scientists
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Item Role of Nod factor receptors and its allies involved in nitrogen fixation(Springer Nature Publishing AG, 2023) Singh, Jawahar; Verma, Praveen K.The availability of water and nutrients in the soil is a major limiting factor affecting crop productivity. Plants of the Leguminosae family form a symbiotic association with nitrogen-fixing Gram-negative soil bacteria, rhizobia for nitrogen fixation. This symbiotic relationship between legumes and rhizobia depends on the signal exchange between them. Plant receptor-like kinases (RLKs) containing lysin motif (LysM) and/or leucine-rich repeat (LRR) play an important role in the perception of chemical signals from rhizobia for initiation and establishment of root nodule symbiosis (RNS) that results in nitrogen fixation. This review highlights the diverse aspects of LysM-RLK and LRR receptors including their specificity, functions, interacting partners, regulation, and associated signaling in RNS. The activation of LysM-RLKs and LRR-RLKs is important for ensuring the successful interaction between legume roots and rhizobia. The intracellular regions of the receptors enable additional layers of signaling that help in the transduction of signals intracellularly. Additionally, symbiosis receptor-like kinase (SYMRK) containing the LRR motif acts as a co-receptor with Nod factors receptors (LysM-RLK). Cleavage of the malectin-like domain from the SYMRK ectodomain is a mechanism for controlling SYMRK stability. Overall, this review has discussed different aspects of legume receptors that are critical to the perception of signals from rhizobia and their subsequent role in creating the mutualistic relationship necessary for nitrogen fixation. Additionally, it has been discussed how crucial it is to extrapolate the knowledge gained from model legumes to crop legumes such as chickpea and common bean to better understand the mechanism underlying nodule formation in crop legumes. Future directions have also been proposed in this regard.Item The R2R3-MYB-SG7 transcription factor CaMYB39 orchestrates surface phenylpropanoid metabolism and pathogen resistance in chickpea(John Wiley & Sons, 2023) Saxena, Samiksha; Pal, Lalita; Naik, Jogindra; Singh, Yeshveer; Verma, Praveen K.; Chattopadhyay, Debasis; Pandey, AshutoshFlavonoids are important plant pigments and defense compounds; understanding the transcriptional regulation of flavonoid biosynthesis may enable engineering crops with improved nutrition and stress tolerance. Here, we characterize R2R3-MYB domain subgroup 7 transcription factor CaMYB39, which regulates flavonol biosynthesis primarily in chickpea trichomes. CaMYB39 overexpression in chickpea was accompanied by a change in flux availability for the phenylpropanoid pathway, particularly flavonol biosynthesis. Lines overexpressing CaMYB39 showed higher isoflavonoid levels, suggesting its role in regulating isoflavonoid pathway. CaMYB39 transactivates the transcription of early flavonoid biosynthetic genes (EBG). FLAVONOL SYNTHASE2, an EBG, encodes an enzyme with higher substrate specificity for dihydrokaempferol than other dihydroflavonols explaining the preferential accumulation of kaempferol derivatives as prominent flavonols in chickpea. Interestingly, CaMYB39 overexpression increased trichome density and enhanced accumulation of diverse flavonol derivatives in trichome-rich tissues. Moreover, CaMYB39 overexpression reduced ROS levels and induced defense gene expression which aids in partially blocking the penetration efficiency of the fungal pathogen, Ascochyta rabiei, resulting in lesser symptoms, thus establishing its role against deadly Ascochyta blight(AB) disease. Overall, our study reports an instance where R2R3-MYB-SG7 member, CaMYB39, besides regulating flavonol biosynthesis, modulates diverse pathways like general phenylpropanoid, isoflavonoid, trichome density and defense against necrotrophic fungal infection in chickpea.Item BAR domain is essential for early endosomal trafficking and dynamics in Ascochyta rabiei(Springer Nature Publishing AG, 2023) Shree, Ankita; Sinha, Manisha; Verma, Praveen K.Ascochyta blight disease is a devastating disease caused by the fungal pathogen Ascochyta rabiei that threatens chickpea production around the globe. Endocytic mechanism has a significant role in fungal growth and virulence. The underlying biology of biogenesis of central component of endocytosis viz Rab5 vesicles, is not completely understood. The involvement of F-BAR domain containing protein (ArF-BAR) in various cellular processes that collectively make ArF-BAR as an important virulence determinant. Here, we report that ArF-BAR is involved in biogenesis and motility of early endosome. In the absence of ArF-BAR gene (Δarf-bar), fungal mutants exhibited reduced number of EGFP coated ArRab5 vesicles, along with the considerable reduction in their dynamics. Here, we show that ArF-BAR interacts with clathrin light chain (ArCLC), specifically with its F-BAR domain. These findings suggests the novel role of ArF-BAR in biogenesis and dynamics of early endosome. Additionally, ArF-BAR is involved in clathrin-mediated mechanism of endocytosis which is required for host infection and disease development. Identification of this pathway offers new impending targets for disease intervention in plants.Item The nuclear effector ArPEC25 from the necrotrophic fungus Ascochyta rabiei targets the chickpea transcription factor CaβLIM1a and negatively modulates lignin biosynthesis, increasing host susceptibility(Oxford University Press, 2023) Singh, Shreenivas Kumar; Shree, Ankita; Verma, Sandhya; Singh, Kunal; Kumar, Kamal; Srivastava, Vikas; Singh, Ritu; Saxena, Samiksha; Singh, Agam Prasad; Pandey, Ashutosh; Verma, Praveen K.Fungal pathogens deploy a barrage of secreted effectors to subvert host immunity, often by evading, disrupting, or altering key components of transcription, defense signaling, and metabolic pathways. However, the underlying mechanisms of effectors and their host targets are largely unexplored in necrotrophic fungal pathogens. Here, we describe the effector protein Ascochyta rabiei PEXEL-like Effector Candidate 25 (ArPEC25), which is secreted by the necrotroph A. rabiei, the causal agent of Ascochyta blight disease in chickpea (Cicer arietinum), and is indispensable for virulence. After entering host cells, ArPEC25 localizes to the nucleus and targets the host LIM transcription factor CaβLIM1a. CaβLIM1a is a transcriptional regulator of CaPAL1, which encodes phenylalanine ammonia lyase, the regulatory, gatekeeping enzyme of the phenylpropanoid pathway. ArPEC25 inhibits the transactivation of CaβLIM1a by interfering with its DNA binding ability, resulting in negative regulation of the phenylpropanoid pathway and decreased levels of intermediates of lignin biosynthesis, thereby suppressing lignin production. Our findings illustrate the role of fungal effectors in enhancing virulence by targeting a key defense pathway that leads to the biosynthesis of various secondary metabolites and antifungal compounds. This study provides a template for the study of less explored necrotrophic effectors and their host target functions.Item Rapid and precise detection of cryptic tea pathogen Exobasidium vexans: RealAmp validation of LAMP approach(Springer Nature Publishing AG, 2023) Chaliha, Chayanika; Srivastava, Richa; Kalita, Eeshan; Sahoo, Lingaraj; Verma, Praveen K.This work embodies the development of a real time loop mediated isothermal amplification (RealAmp) assay for the rapid detection of the cryptic tea phytopathogen, Exobasidium vexans, the causal organism of blister blight disease. Due to the widespread popularity of tea as a beverage and the associated agro-economy, the rapid detection and management of the fast-spreading blister blight disease have been a longstanding necessity. Loop-mediated isothermal amplification (LAMP) primers were designed targeting the E. vexans ITS rDNA region and the reaction temperature was optimized at 62 °C with a 60 min reaction time. Amplification of the E. vexans isolates in the initial LAMP reactions was confirmed by both agarose gel electrophoresis and SYBR Green I dye based colour change visualization. The specificity of the LAMP primers for E. vexans was validated by negative testing of seven different phytopathogenic test fungi using LAMP and RealAmp assay. The positive findings in RealAmp assay for E. vexans strain were corroborated via detecting fluorescence signals in real-time. Further, the LAMP assays performed with gDNA isolated from infected tea leaves revealed positive amplification for the presence of E. vexans. The results demonstrate that this rapid and precise RealAmp assay has the potential to be applied for field-based detection of E. vexans in real-time.Item Invasion and colonization of pathogenic Fusarium oxysporum R1 in Crocus sativus L. during corm rot disease progression(MDPI AG, 2022) Bhagat, Nancy; Magotra, Shanu; Gupta, Rikita; Sharma, Shikha; Verma, Sandhya; Verma, Praveen K.; Ali, Tahir; Shree, Ankita; Vakhlu, JyotiThe corm rot of saffron caused by Fusarium oxysporum (Fox) has been reported to be the most destructive fungal disease of the herb globally. The pathogen, Fusarium oxysporum R1 (Fox R1) isolated by our group from Kashmir, India, was found to be different from Fusarium oxysporum f.sp. gladioli commonly reported corm rot agent of saffron. In the present study, Fox R1 was further characterized using housekeeping genes and pathogenicity tests, as Fusarium oxysporum R1 f.sp. iridacearum race 4. Though Fox R1 invaded the saffron plant through both corm and roots, the corm was found to be the preferred site of infection. In addition, the route of pathogen movement wastracked by monitoring visual symptoms, semi-quantitative PCR, quantitative-PCR (q-PCR), real-time imaging of egfp-tagged Fusarium oxysporum R1, and Fox R1 load quantification. This study is the first study of its kind on the bidirectional pathogenesis from corm to roots and vice-versa, as the literature only reports unidirectional upward movement from roots to other parts of the plant. In addition, the colonization pattern of Fox R1 in saffron corms and roots was studied. The present study involved a systematic elucidation of the mode and mechanism of pathogenesis in the saffron Fusarium oxysporum strain R1 pathosystem.Item Comparative genomic analysis of GARP transcription factor family in legumes and identification of stress-responsive candidate genes(Springer Nature Publishing AG, 2023) Singh, Ritu; Pandey, Ashutosh; Verma, Praveen K.The GARP transcription factors have been identified for multiple biological functions throughout the life cycle of a plant. Despite of its involvement in crucial functions, systematic study of GARPs remains obscure in plants. In this study, we explored the genomic, molecular, and evolutionary perspectives of the GARP gene family in the three major leguminous plants, namely chickpea (Cicer arietinum), soybean (Glycine max), and barrel clover (Medicago truncatula). Here, we identified 53, 56, and 107 GARP genes in Cicer, Medicago, and Glycine, respectively. They were classified into four clades and two sub-clades as per phylogenetic analysis, and the result was supported by consensus motifs, domain organization, and exon–intron structures. Detailed comparative analysis indicates conservation of the GARP gene family in plants. Identification of paralogous and orthologous gene pairs revealed that the expansion of this family occurs mainly through genome duplication in legumes. Additionally, the three-dimensional structure and functional enrichment analysis indicated their major role in signaling, growth, development, and stress processes. The chickpea GARP genes were also characterized for their transcript modulation in diverse plant organs and during pathogenic stress. Differential regulation of 24 CaGARP genes was observed during Ascochyta Blight (AB) stress. Characterization of AB-responsive genes reveals an over-representation of stress and hormone-binding elements on the promoter of CaGARPs. Additionally, interactome analysis also confirms the role of GARPs in plant stress and development. Our findings not only provide a handful of stress-responsive genes but also lay the foundation for prospective functional studies of GARPs in legumes.Item Global transcriptome and co-expression analysis reveals robust host defence pathway reprogramming and identifies key regulators of early phases of Cicer-Ascochyta interactions(American Phytopathological Society, 2022) Singh, Ritu; Dwivedi, Aditi; Singh, Yeshveer; Kumar, Kamal; Ranjan, Aashish; Verma, Praveen K.Ascochyta blight (AB) caused by a filamentous fungus Ascochyta rabiei is a major threat to global chickpea production. The mechanisms underlying chickpea response to A. rabiei remain elusive. Here, we investigated the comparative transcriptional dynamics of AB-resistant and susceptible chickpea genotypes upon A. rabiei infection to understand the early host defence response. Our findings revealed that AB-resistant plants underwent rapid and extensive transcriptional reprogramming compared to susceptible host. At early stage (24-hpi), mainly cell wall remodeling and secondary metabolite pathways were highly activated, while DEGs related with signaling components viz. protein kinases, transcription factors, and hormonal pathways show remarkable upsurge at 72-hpi, especially in resistant genotype. Notably, our data suggests imperative role of JA, ET, and ABA signaling in providing immunity against A. rabiei. Furthermore, gene co-expression networks and modules corroborated the importance of cell wall remodeling, signal transduction, and phytohormone pathways. The hub genes such as MYB14, PRE6, and MADS-SOC1 discovered in these modules might be the master regulators governing chickpea immunity. Overall, we not only provide novel insights for comprehensive understanding of immune signaling components mediating AB resistance/susceptibility at early Cicer-Ascochyta interactions, but also offer a valuable resource for developing AB-resistant chickpea.Item Genome-wide identification, expression, and characterization of CaLysM-RLKs in chickpea root nodule symbiosis(Elsevier B.V., 2022) Singh, Jawahar; Verma, Praveen K.Legumes establish a nitrogen-fixing symbiosis with rhizobia which depends on the exchange of chemical signals between the two partners. Successful interaction between legumes and rhizobia depends on the perception of Nod factors (NFs) by the receptor-like kinases. A few cell surface receptors have been identified that perceive the rhizobial NFs and activate the downstream signaling pathway for nodulation in model legumes like Lotus japonicus and Medicago truncatula. However, crop legume such as chickpea lacks detailed analysis of LysM-RLKs genes for NFs perception. Here, we identified and characterized CaLysM-RLK genes and analyzed their gene structure, motif composition, chromosomal localization, phylogenetic relationship with Arabidopsis, Oryza sativa, and Medicago truncatula. To understand the spatiotemporal expression patterns, we performed transcript profiling of CaLysM-RLKs genes at a different time interval of the chickpea-rhizobia symbiotic interaction that revealed CaNFP and CaLYK3 can act as candidate genes for NF perception. Down-regulation of CaNFP and CaLYK3 in chickpea by agrobacterium rhizogenes mediated hairy root sysytem leads to signifificant reduction in nodule numbers. This study will be useful for improving root nodule symbiosis in chickpea.Item Ascochyta rabiei: A threat to global chickpea production(John Wiley & Sons, 2022) Singh, Ritu; Kumar, Kamal; Purayannur, Savithri; Chen, Weidong; Verma, Praveen K.The necrotrophic fungus Ascochyta rabiei causes Ascochyta blight (AB) disease in chickpea. A. rabiei infects all aerial parts of the plant, which results in severe yield loss. At present, AB disease occurs in most chickpea-growing countries. Globally increased incidences of A. rabiei infection and the emergence of new aggressive isolates directed the interest of researchers toward understanding the evolution of pathogenic determinants in this fungus. In this review, we summarize the molecular and genetic studies of the pathogen along with approaches that are helping in combating the disease. Possible areas of future research are also suggested.