SummaryPhosphate (Pi) deficiency in soil system is a limiting factor for rice growth and yield. Majority of the soil phosphorus (P) is organic in nature, not readily available for root uptake. Low Pi‐inducible purple acid phosphatases (PAPs) are hypothesized to enhance the availability of Pi in soil and cellular system. However, information on molecular and physiological roles of rice PAPs is very limited. Here, we demonstrate the role of a novel rice PAP, OsPAP21b in improving plant utilization of organic‐P. OsPAP21b was found to be under the transcriptional control of OsPHR2 and strictly regulated by plant Pi status at both transcript and protein levels. Biochemically, OsPAP21b showed hydrolysis of several organophosphates at acidic pH and possessed sufficient thermostability befitting for high‐temperature rice ecosystems with acidic soils. Interestingly, OsPAP21b was revealed to be a secretory PAP and encodes a distinguishable major APase (acid phosphatase) isoform under low Pi in roots. Further, OsPAP21b‐overexpressing transgenics showed increased biomass, APase activity and P content in both hydroponics supplemented with organic‐P sources and soil containing organic manure as sole P source. Additionally, overexpression lines depicted increased root length, biomass and lateral roots under low Pi while RNAi lines showed reduced root length and biomass as compared to WT. In the light of these evidences, present study strongly proposes OsPAP21b as a useful candidate for improving Pi acquisition and utilization in rice.

Phosphate (Pi) deficiency severely affects crop yield. Modern high yielding rice genotypes are sensitive to Pi deficiency whereas traditional rice genotypes are naturally compatible with low Pi ecosystems. However, the underlying molecular mechanisms for low Pi tolerance in traditional genotypes remain largely elusive. To delineate the molecular mechanisms for low Pi tolerance, two contrasting rice genotypes, Dular (low Pi tolerant), and PB1 (low Pi sensitive), have been selected. Comparative morphophysiological, global transcriptome and lipidome analyses of root and shoot tissues of both genotypes grown under Pi deficient and sufficient conditions revealed potential low Pi tolerance mechanisms of the traditional genotype. Most of the genes associated with enhanced internal Pi utilization (phospholipid remobilization) and modulation of root system architecture (RSA) were highly induced in the traditional rice genotype, Dular. Higher reserves of phospholipids and greater accumulation of galactolipids under low Pi in Dular indicated it has more efficient Pi utilization. Furthermore, Dular also maintained greater root growth than PB1 under low Pi, resulting in larger root surface area due to increased lateral root density and root hair length. Genes involved in enhanced low Pi tolerance of the traditional genotype can be exploited to improve the low Pi tolerance of modern high yielding rice cultivars.

Phosphorus (P) deficiency limits plant growth and yield. Since plants can absorb only the inorganic form of P (Pi), a large portion of soil P (organic and inorganic P complexes) remains unused. Here, we identified and characterized a PHR2-regulated, novel low-Pi-responsive haloacid dehalogenase (HAD)-like hydrolase, While OsHAD1 is a functional HAD protein having both acid phosphatase and phytase activities, it showed little homology with other known low-Pi-responsive HAD superfamily members. Recombinant OsHAD1 is active at acidic pH and dephosphorylates a broad range of organic and inorganic P-containing substrates, including phosphorylated serine and sodium phytate. Exogenous application of recombinant OsHAD1 protein in growth medium supplemented with phytate led to marked increases in growth and total P content of Pi-deficient wild-type rice () seedlings. Furthermore, overexpression of in rice resulted in enhanced phosphatase activity, biomass, and total and soluble P contents in Pi-deficient transgenic seedlings treated with phytate as a restricted Pi source. Gene expression and metabolite profiling revealed enhanced Pi starvation responses, such as up-regulation 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 with the wild type. To elucidate the underlying regulatory mechanisms of OsHAD1, we performed in vitro pull-down assays, which revealed the association of OsHAD1 with protein kinases such as OsNDPKs. We conclude that, besides dephosphorylation of cellular organic P, OsHAD1 in coordination with kinases may regulate the phosphorylation status of downstream targets to accomplish Pi homeostasis under limited Pi supply.

Rice and chickpea GDPD s are transcriptionally influenced by mineral deficiencies; especially, by phosphate starvation and CaGDP1 encodes an active glycerophosphodiester phosphodiesterase enzyme. Glycerophosphodiester phosphodiesterases (GDPDs) are enzymes involved in the degradation of glycerophosphodiesters into sn-glycerol-3-phosphate and corresponding alcohols. These phospholipid remodeling genes have been suggested to play important roles in phosphate homeostasis. However, comprehensive information about the role of GDPDs under low phosphate (P) and other nutrient deficiencies (N, K, Fe, Zn) in rice and chickpea is missing. Here, we identified 13 OsGDPDs and 6 CaGDPDs in rice and chickpea, respectively, and partly characterized their roles in multiple nutrient stresses. Expression profiling after 7 and 15 days of deficiency treatments revealed unique and overlapping differential expression patterns of OsGDPDs and CaGDPDs under different nutrient stresses. Principal component analysis on the expression patterns of OsGDPDs and CaGDPDs revealed their preferential role in P starvation. Some of the GDPDs were also induced by N, K, Fe and Zn deficiency in temporal manner in both crops suggesting their roles in multiple nutrient stresses. Biochemical characterization of highly responsive chickpea GDPD, CaGDPD1, confirmed its in vitro GDPD activity and revealed its optimal temperature, pH and cofactor requirements. Further, CaGDPD1 showed its accumulation in ER and endomembranes. We hereby propose CaGDPD1 and various OsGDPDs as low P responsive marker genes in chickpea and rice, respectively. Our data uphold role of GDPDs in multinutrient responses and suggest them as candidates for rice and chickpea improvement for tolerance to various nutrient deficiencies.

Soil Phosphorus (P) deficiency is one of the major challenges to rice crop world-wide. Modern rice genotypes are highly P-responsive and rely on high input of P fertilizers. However, low P tolerant traditional cultivars and landraces have genetic potential to sustain well under low P. Identification of high resolution DNA polymorphisms (SNPs and InDels) in such contrasting genotypes is largely missing for low P response at gene levels. Here, we report high quality DNA polymorphisms in low P sensitive genotype, PB1 and tolerant traditional genotype, Dular. We performed whole genome resequencing using Illumina NGS platform and identified a total of 5,157,939 sequence variants in PB1 and Dular with reference to Nipponbare genome. We have identified approximately 2.3 million and 2.9 million high quality polymorphisms in PB1 and Dular, respectively, with an average read depth of ≥24X. We further mapped several DNA polymorphisms (non-synonymous and regulatory variants) having potential functional significance to key Phosphate Starvation Responsive (PSR) and root architecture genes in Dular and Kasalath using a compiled list of low P responsive genes. These identified variants can serve as a useful source of genetic variability for improving low P tolerance and root architecture of high yielding modern genotypes.

Soil phosphate (Pi) deficiency is major constraint for rice cultivation worldwide. Cellular membranes account for one third of cellular organic phosphorus (P) in the form of phospholipids. Therefore, remobilization of Pi from membrane phospholipids under Pi deficiency can be an important strategy to improve phosphorus use efficiency. Glycerophosphodiester phosphodiesterases (GDPDs) hydrolyse intermediate product of phospholipid catabolism, glycerophosphodiesters to glycerol‐3‐phosphate, a precursor for P and non P‐lipid biosynthesis. Here, we show that OsGDPD2 is a Pi deficiency responsive gene, which is transcriptionally regulated by OsPHR2. In silico analysis of active site residues and enzymatic assays confirmed phosphodiesterase activity of OsGDPD2. All overexpression lines showed higher GDPD activity, Pi content, root growth, and biomass accumulation as compared with wild type. Conversely, silencing of OsGDPD2 led to decreased GDPD activity and Pi content. Notably, most of the P‐containing metabolites and fatty acids were elevated in transgenic lines. Further, quantitative analysis of polar lipids revealed higher accumulation of several classes of phospholipids and galactolipids in overexpression lines indicating a potential role of OsGDPD2 in de novo glycerolipid biosynthesis. Thus, present study provides insights into novel physiological roles of OsGDPD2 in low Pi acclimation in rice.

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.

Intensive farming has depleted the soil zinc (Zn) availability resulting in decreased crop productivity. Here, we attempt to understand the Zn deficiency response in rice through temporal transcriptome analysis. For this, rice seedlings were raised under Zn-deficient conditions up to 4 weeks followed by Zn re-supply for 3 days. Zn-deficient plants developed characteristic deficiency symptoms such as leaf bronzing, decrease in biomass, total chlorophyll, PSII efficiency, decreased carbonic anhydrase activity and increased ROS production. Interestingly, severe alterations in root system architecture were also observed. Comprehensive transcriptome analyses of rice seedlings were carried out after 2 (DEF2W) and 4 weeks (DEF4W) of Zn deficiency with respect to transcriptome profiles of corresponding Zn sufficient conditions (SUF2W, SUF4W). Additionally, to detect the potential Zn-responsive genes, transcriptome profile of Zn-recovered seedlings was compared with DEF4W. All differentially expressed Zn-responsive genes were categorized into early and late Zn deficiency response, and a set of 77 genes, induced and repressed on Zn deficiency and re-supply, respectively, was identified. These genes could be used as low Zn-responsive marker genes. Further, genes involved in membrane transport, phytosiderophore activity and organic acid biosynthesis showed high differential expression. Additionally, the present study unravelled several genes putatively associated with alterations in root system architecture under Zn deficiency and provides novel insights into the interpretation of morpho-physiological, biochemical and molecular regulation of zinc deficiency responses in rice.