Browsing by Author "Bhadouria, Jyoti"

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A citrate efflux transporter important for manganese distribution and phosphorus uptake in rice
(John Wiley & Sons, 2023) Panchal, Poonam; Bhatia, Chitra; Chen, Yi; Sharma, Meenakshi; Bhadouria, Jyoti; Verma, Lokesh; Maurya, Kanika; Miller, Anthony J.; Giri, Jitender
The plant citrate transporters, functional in mineral nutrient uptake and homeostasis, usually belong to the multidrug and toxic compound extrusion transporter family. We identified and functionally characterized a rice (Oryza sativa) citrate transporter, OsCT1, which differs from known plant citrate transporters and is structurally close to rice silicon transporters. Domain analysis depicted that OsCT1 carries a bacterial citrate-metal transporter domain, CitMHS. OsCT1 showed citrate efflux activity when expressed in Xenopus laevis oocytes and is localized to the cell plasma membrane. It is highly expressed in the shoot and reproductive tissues of rice, and its promoter activity was visible in cells surrounding the vasculature. The OsCT1 knockout (KO) lines showed a reduced citrate content in the shoots and the root exudates, whereas overexpression (OE) line showed higher citrate exudation from their roots. Further, the KO and OE lines showed variations in the manganese (Mn) distribution leading to changes in their agronomical traits. Under deficient conditions (Mn-sufficient conditions followed by 8 days of 0 μm MnCl2 · 4H2 O treatment), the supply of manganese towards the newer leaf was found to be obstructed in the KO line. There were no significant differences in phosphorus (P) distribution; however, P uptake was reduced in the KO and increased in OE lines at the vegetative stage. Further, experiments in Xenopus oocytes revealed that OsCT1 could efflux citrate with Mn. In this way, we provide insights into a mechanism of citrate-metal transport in plants and its role in mineral homeostasis, which remains conserved with their bacterial counterparts.
Identification of purple acid phosphatases in chickpea and potential roles of CaPAP7 in seed phytate accumulation
(Nature Publishing Group, 2017) Bhadouria, Jyoti; Singh, Ajit Pal; Mehra, Poonam; Verma, Lokesh; Srivastawa, Rishi; Parida, Swarup K.; Giri, Jitender
Purple acid phosphatases (PAPs) play important roles in phosphate (Pi) acquisition and utilization. These PAPs hydrolyze organic Phosphorus (P) containing compounds in rhizosphere as well as inside the plant cell. However, roles of PAPs in one of the most widely cultivated legumes, chickpea (Cicer arietnum L.), have not been unraveled so far. In the present study, we identified 25 putative PAPs in chickpea (CaPAPs) which possess functional PAP motifs and domains. Differential regulation of CaPAPs under different nutrient deficiencies revealed their roles under multiple nutrient stresses including Pi deficiency. Interestingly, most of the CaPAPs were prominently expressed in flowers and young pods indicating their roles in flower and seed development. Association mapping of SNPs underlying CaPAPs with seed traits revealed significant association of low Pi inducible CaPAP7 with seed weight and phytate content. Biochemical characterization of recombinant CaPAP7 established it to be a functional acid phosphatase with highest activity on most abundant organic-P substrate, phytate. Exogenous application of recombinant CaPAP7 enhanced biomass and Pi content of Arabidopsis seedlings supplemented with phytate as sole P source. Taken together, our results uncover the PAPs in chickpea and potential roles of CaPAP7 in seed phytate accumulation.
Monogalactosyl Diacylglycerol Synthase 3 affects phosphate utilization and acquisition in rice
(Oxford University Press, 2022) Verma, Lokesh; Bhadouria, Jyoti; Rupam, Bhunia Kumar; Singh, Shweta; Panchal, Poonam; Bhatia, Chitra; Eastmond, Peter J; Giri, Jitender
Galactolipids are essential for compensating for the loss of phospholipids by “membrane lipid remodeling” in plants under phosphorus (P) deficiency. Monogalactosyl diacylglycerol (MGDG) synthases catalyze the synthesis of MGDG which is further converted into digalactosyl diacylglycerol (DGDG), later replacing phospholipids in the extraplastidial membranes. However, the roles of these enzymes are not well explored in rice. In this study, the rice MGDG synthase gene, OsMGD3 was identified and functionally characterized. We showed that plant phosphate (Pi) status and transcription factor OsPHR2 are involved in the transcriptional regulation of OsMGD3. CRISPR/Cas9 knockout (KO) and overexpression (OE) lines of OsMGD3 were generated to explore its potential role in rice adaptation to Pi deficiency. Compared to WT, OsMGD3 KO lines displayed a reduction while OE lines showed an enhanced, Pi acquisition and utilization. Further, OsMGD3 showed a predominant role in roots, altering lateral root growth. Our comprehensive lipidomic analysis revealed the role of OsMGD3 in membrane lipid remodeling in addition to a role in regulating diacylglycerol and phosphatidic acid levels that affected the expression of Pi transporters. Our study highlights the role of OsMGD3 in affecting both internal P utilization and P acquisition in rice.
OsHAD1, a haloacid dehalogenase-like APase enhances phosphate accumulation
(American Society of Plant Biologists, 2017) Pandey, Bipin Kumar; Mehra, Poonam; Verma, Lokesh; Bhadouria, Jyoti; Giri, Jitender
Phosphorus (P) deficiency limits plant growth and crop yield. Since, plants can absorb only inorganic form of P (Pi), a large portion of soil P (organic and inorganic P complexes) remains largely unused. Here, we identified and characterized a PHR2 regulated; novel low Pi responsive haloacid dehalogenase (HAD)-like hydrolase, OsHAD1. While, OsHAD1 is a functional HAD protein having both acid phosphatase and phytase activity; it showed little homology with other known low Pi responsive HAD superfamily members. Recombinant OsHAD1 is highly active at acidic pH and dephosphorylates broad range of organic and inorganic P containing substrates including protein phosphates and Na-phytate. Exogenous application of recombinant OsHAD1 protein in growth media supplemented with phytate, led to marked increase in growth and total P content of Pi deficient WT rice seedlings. Further, overexpression of OsHAD1 in rice resulted in enhanced phosphatase activity, biomass, total and soluble P content in Pi deficient transgenic seedlings treated with phytate as restricted Pi source. Gene expression and metabolite profiling revealed enhanced Pi starvation responses such as upregulation of multiple genes involved in Pi uptake and solubilization, accumulation of organic acids, enhanced secretory phosphatase activity and depletion of ATP in overexpression lines as compared to WT. To elucidate the underlying regulatory mechanisms of OsHAD1, we performed in-vitro pull down assays which revealed association of OsHAD1 with protein kinases. We conclude that besides dephosphorylation of cellular organic-P, OsHAD1 in coordination with kinases may regulate phosphorylation status of downstream targets to accomplish Pi homeostasis under limited Pi supply.
Purple acid phosphatases: roles in phosphate utilization and new emerging functions
(Springer Nature Publishing AG, 2022) Bhadouria, Jyoti; Giri, Jitender
Plants strive for phosphorus (P), which is an essential mineral for their life. Since P availability is limiting in most of the world’s soils, plants have evolved with a complex network of genes and their regulatory mechanisms to cope with soil P defciency. Among them, purple acid phosphatases (PAPs) are predominantly associated with P remobilization within the plant and acquisition from the soil by hydrolyzing organic P compounds. P in such compounds remains otherwise unavailable to plants for assimilation. PAPs are ubiquitous in plants, and similar enzymes exist in bacteria, fungi, mammals, and unicellular eukaryotes, but having some diferences in their catalytic center. In the recent past, PAPs’ roles have been extended to multiple plant processes like fowering, seed development, senescence, carbon metabolism, response to biotic and abiotic stresses, signaling, and root development. While new functions have been assigned to PAPs, the underlying mechanisms remained understood poorly. Here, we review the known functions of PAPs, the regulatory mechanisms, and their relevance in crop improvement for P-use-efciency. We then discuss the mechanisms behind their functions and propose areas worthy of future research. Finally, we argue that PAPs could be a potential target for improving P utilization in crops. In turn, this is essential for sustainable agriculture.
Root-expressed rice PAP3b enhances secreted APase activity and helps utilize organic phosphate
(Oxford University Press, 2023) Bhadouria, Jyoti; Mehra, Poonam; Verma, Lokesh; Pazhamala, Lekha T; Rumi, Rumi; Panchal, Poonam; Sinha, Alok Krishna; Giri, Jitender
Phosphate (Pi) deficiency leads to the induction of purple acid phosphatases (PAPs) in plants, which dephosphorylates organic phosphorus complexes in the rhizosphere and intracellular compartments to release Pi. In this study, we demonstrate that OsPAP3b belongs to group III low molecular weight PAP, and is low Pi responsive, preferentially in roots. The expression of OsPAP3b is negatively regulated with Pi re-supply. Interestingly, OsPAP3b was found to be dual localized to the nucleus and secretome. Furthermore, OsPAP3b is transcriptionally regulated by OsPHR2 as substantiated by DNA-protein binding assay. Through in-vitro biochemical assays, we further demonstrate that OsPAP3b is a functional acid phosphatase with broad substrate specificity. Overexpression of OsPAP3b in rice led to increased secreted APase activity and improved mineralization of organic P sources, reflected in better growth of transgenics compared to wild type when grown on organic P as exogenous P substrate. Under Pi deprivation, OsPAP3b knockdown and knockout lines showed no significant changes in total P content and dry biomass. However, the expression of other phosphate starvation-induced (PSI) genes and the levels of metabolites were found to be altered in the overexpression and knockdown lines. In addition, in-vitro pull-down assay revealed multiple putative interacting proteins of OsPAP3b. Our data collectively suggest that OsPAP3b can aid in organic P utilization in rice. The APase isoforms behavior and nuclear localization indicate its additional role, possibly in stress signaling. Considering its important roles, OsPAP3b could be a potential target for improving low Pi adaptation in rice.