Genome-Wide Identification of OsZIPs in Rice and Gene Expression Analysis under Manganese and Selenium Stress.

Zinc (Zn)- and iron (Fe)-regulating transport-like proteins (ZIPs) are a class of proteins crucial for metal uptake and transport in plants, particularly for Zn and Fe absorption and distribution. These proteins ensure the balance of trace elements essential for plant growth, development, and metabolic activities. However, the role of the rice (Oryza sativa) OsZIP gene family in manganese (Mn) and selenium (Se) transport remains underexplored. This research conducted an all-sided analysis of the rice OsZIPs and identified 16 OsZIP sequences. Phylogenetic analysis categorized the OsZIPs predominantly within the three subfamilies. The expression levels of OsZIPs in rice root and leaf subjected to Mn and Se toxicity stress were examined through quantitative real-time PCR (qRT-PCR). The findings revealed significant differential expression of many OsZIPs under these conditions, indicating a potential regulating effect in the response of rice to Mn and Se toxicity. This work lays a foundation for further functional studies of OsZIPs, enhancing our understanding of the response mechanisms of rice to Mn and Se toxicity and their roles in growth, development, and environmental adaptation.

High Concentrations of Se Inhibited the Growth of Rice Seedlings.

Selenium (Se) is crucial for both plants and humans, with plants acting as the main source for human Se intake. In plants, moderate Se enhances growth and increases stress resistance, whereas excessive Se leads to toxicity. The physiological mechanisms by which Se influences rice seedlings' growth are poorly understood and require additional research. In order to study the effects of selenium stress on rice seedlings, plant phenotype analysis, root scanning, metal ion content determination, physiological response index determination, hormone level determination, quantitative PCR (qPCR), and other methods were used. Our findings indicated that sodium selenite had dual effects on rice seedling growth under hydroponic conditions. At low concentrations, Se treatment promotes rice seedling growth by enhancing biomass, root length, and antioxidant capacity. Conversely, high concentrations of sodium selenite impair and damage rice, as evidenced by leaf yellowing, reduced chlorophyll content, decreased biomass, and stunted growth. Elevated Se levels also significantly affect antioxidase activities and the levels of proline, malondialdehyde, metal ions, and various phytohormones and selenium metabolism, ion transport, and antioxidant genes in rice. The adverse effects of high Se concentrations may directly disrupt protein synthesis or indirectly induce oxidative stress by altering the absorption and synthesis of other compounds. This study aims to elucidate the physiological responses of rice to Se toxicity stress and lay the groundwork for the development of Se-enriched rice varieties.

The Stylo Cysteine-Rich Peptide SgSnakin1 Is Involved in Aluminum Tolerance through Enhancing Reactive Oxygen Species Scavenging.

Stylo (Stylosanthes spp.) is an important pasture legume with strong aluminum (Al) resistance. However, the molecular mechanisms underlying its Al tolerance remain fragmentary. Due to the incomplete genome sequence information of stylo, we first conducted full-length transcriptome sequencing for stylo root tips treated with and without Al and identified three Snakin/GASA genes, namely, SgSnakin1, SgSnakin2, and SgSnakin3. Through quantitative RT-PCR, we found that only SgSnakin1 was significantly upregulated by Al treatments in stylo root tips. Histochemical localization assays further verified the Al-enhanced expression of SgSnakin1 in stylo root tips. Subcellular localization in both tobacco and onion epidermis cells showed that SgSnakin1 localized to the cell wall. Overexpression of SgSnakin1 conferred Al tolerance in transgenic Arabidopsis, as reflected by higher relative root growth and cell vitality, as well as lower Al concentration in the roots of transgenic plants. Additionally, overexpression of SgSnakin1 increased the activities of SOD and POD and decreased the levels of O2·- and H2O2 in transgenic Arabidopsis in response to Al stress. These findings indicate that SgSnakin1 may function in Al resistance by enhancing the scavenging of reactive oxygen species through the regulation of antioxidant enzyme activities.

The Physiological Response Mechanism of Peanut Leaves under Al Stress.

Aluminum (Al) toxicity in acidic soils can significantly reduce peanut yield. The physiological response of peanut leaves to Al poisoning stress still has not been fully explored. This research examined the influences of Al toxicity on peanut leaves by observing the leaf phenotype, scanning the leaf area and perimeter, and by measuring photosynthetic pigment content, physiological response indices, leaf hormone levels, and mineral element accumulation. Fluorescence quantitative RT-PCR (qPCR) was utilized to determine the relative transcript level of specific genes. The results indicated that Al toxicity hindered peanut leaf development, reducing their biomass, surface area, and perimeter, although the decrease in photosynthetic pigment content was minimal. Al toxicity notably affected the activity of antioxidative enzymes, proline content, and MDA (malondialdehyde) levels in the leaves. Additionally, Al poisoning resulted in the increased accumulation of iron (Fe), potassium (K), and Al in peanut leaves but reduced the levels of calcium (Ca), manganese (Mn), copper (Cu), zinc (Zn), and magnesium (Mg). There were significant changes in the content of hormones and the expression level of genes connected with hormones in peanut leaves. High Al concentrations may activate cellular defense mechanisms, enhancing antioxidative activity to mitigate excess reactive oxygen species (ROS) and affecting hormone-related gene expression, which may impede leaf biomass and development. This research aimed to elucidate the physiological response mechanisms of peanut leaves to Al poisoning stress, providing insights for breeding new varieties resistant to Al poisoning.

Regulatory Mechanism through Which Old Soybean Leaves Respond to Mn Toxicity Stress.

Manganese (Mn) is a heavy metal that can cause excessive Mn poisoning in plants, disrupting microstructural homeostasis and impairing growth and development. However, the specific response mechanisms of leaves to Mn poisoning have not been fully elucidated. This study revealed that Mn poisoning of soybean plants resulted in yellowing of old leaves. Physiological assessments of these old leaves revealed significant increases in the antioxidant enzymes activities (peroxidase (POD), superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT)) and elevated levels of malondialdehyde (MDA), proline, indoleacetic acid (IAA), and salicylic acid (SA), under 100 µM Mn toxicity. Conversely, the levels of abscisic acid (ABA), gibberellin 3 (GA3), and jasmonic acid (JA) significantly decreased. The Mn content in the affected leaves significantly increased, while the levels of Ca, Na, K, and Cu decreased. Transcriptome analysis revealed 2258 differentially expressed genes in the Mn-stressed leaves, 744 of which were upregulated and 1514 were downregulated; these genes included genes associated with ion transporters, hormone synthesis, and various enzymes. Quantitative RT-PCR (qRT-PCR) verification of fifteen genes confirmed altered gene expression in the Mn-stressed leaves. These findings suggest a complex gene regulatory mechanism under Mn toxicity and stress, providing a foundation for further exploration of Mn tolerance-related gene regulatory mechanisms in soybean leaves. Using the methods described above, this study will investigate the molecular mechanism of old soybean leaves' response to Mn poisoning, identify key genes that play regulatory roles in Mn toxicity stress, and lay the groundwork for cultivating high-quality soybean varieties with Mn toxicity tolerance traits.

Transcriptomic and Lipidomic Analysis Reveals Complex Regulation Mechanisms Underlying Rice Roots' Response to Salt Stress.

Rice (Oryza sativa L.), a crucial food crop that sustains over half the world's population, is often hindered by salt stress during various growth stages, ultimately causing a decrease in yield. However, the specific mechanism of rice roots' response to salt stress remains largely unknown. In this study, transcriptomics and lipidomics were used to analyze the changes in the lipid metabolism and gene expression profiles of rice roots in response to salt stress. The results showed that salt stress significantly inhibited rice roots' growth and increased the roots' MDA content. Furthermore, 1286 differentially expressed genes including 526 upregulated and 760 downregulated, were identified as responding to salt stress in rice roots. The lipidomic analysis revealed that the composition and unsaturation of membrane lipids were significantly altered. In total, 249 lipid molecules were differentially accumulated in rice roots as a response to salt stress. And most of the major phospholipids, such as phosphatidic acid (PA), phosphatidylcholine (PC), and phosphatidylserine (PS), as well as major sphingolipids including ceramide (Cer), phytoceramide (CerP), monohexose ceramide (Hex1Cer), and sphingosine (SPH), were significantly increased, while the triglyceride (TG) molecules decreased. These results suggested that rice roots mitigate salt stress by altering the fluidity and integrity of cell membranes. This study enhances our comprehension of salt stress, offering valuable insights into changes in the lipids and adaptive lipid remodeling in rice's response to salt stress.

Unregulated GmAGL82 due to Phosphorus Deficiency Positively Regulates Root Nodule Growth in Soybean.

Nitrogen fixation, occurring through the symbiotic relationship between legumes and rhizobia in root nodules, is crucial in sustainable agriculture. Nodulation and soybean production are influenced by low levels of phosphorus stress. In this study, we discovered a MADS transcription factor, GmAGL82, which is preferentially expressed in nodules and displays significantly increased expression under conditions of phosphate (Pi) deficiency. The overexpression of GmAGL82 in composite transgenic plants resulted in an increased number of nodules, higher fresh weight, and enhanced soluble Pi concentration, which subsequently increased the nitrogen content, phosphorus content, and overall growth of soybean plants. Additionally, transcriptome analysis revealed that the overexpression of GmAGL82 significantly upregulated the expression of genes associated with nodule growth, such as GmENOD100, GmHSP17.1, GmHSP17.9, GmSPX5, and GmPIN9d. Based on these findings, we concluded that GmAGL82 likely participates in the phosphorus signaling pathway and positively regulates nodulation in soybeans. The findings of this research may lay the theoretical groundwork for further studies and candidate gene resources for the genetic improvement of nutrient-efficient soybean varieties in acidic soils.

Physiological Mechanism through Which Al Toxicity Inhibits Peanut Root Growth.

Al (Aluminum) poisoning is a significant limitation to crop yield in acid soil. However, the physiological process involved in the peanut root response to Al poisoning has not been clarified yet and requires further research. In order to investigate the influence of Al toxicity stress on peanut roots, this study employed various methods, including root phenotype analysis, scanning of the root, measuring the physical response indices of the root, measurement of the hormone level in the root, and quantitative PCR (qPCR). This research aimed to explore the physiological mechanism underlying the reaction of peanut roots to Al toxicity. The findings revealed that Al poisoning inhibits the development of peanut roots, resulting in reduced biomass, length, surface area, and volume. Al also significantly affects antioxidant oxidase activity and proline and malondialdehyde contents in peanut roots. Furthermore, Al toxicity led to increased accumulations of Al and Fe in peanut roots, while the contents of zinc (Zn), cuprum (Cu), manganese (Mn), kalium (K), magnesium (Mg), and calcium (Ca) decreased. The hormone content and related gene expression in peanut roots also exhibited significant changes. High concentrations of Al trigger cellular defense mechanisms, resulting in differentially expressed antioxidase genes and enhanced activity of antioxidases to eliminate excessive ROS (reactive oxygen species). Additionally, the differential expression of hormone-related genes in a high-Al environment affects plant hormones, ultimately leading to various negative effects, for example, decreased biomass of roots and hindered root development. The purpose of this study was to explore the physiological response mechanism of peanut roots subjected to aluminum toxicity stress, and the findings of this research will provide a basis for cultivating Al-resistant peanut varieties.

Impaired glycosylation of GmPAP15a, a root-associated purple acid phosphatase, inhibits extracellular phytate-P utilization in soybean.

Phosphorus (P) is an essential nutrient, but easily fixed in soils. Therefore, most of soil P exists in the form of inaccessible organic phosphorus (Po), particularly phytate-P. Root-associated purple acid phosphatases (PAPs) are considered to play a crucial role in phosphate (Pi) scavenging in soils. However, evidence for regulating root-associated PAPs in utilization of extracellular phytate-P remain largely unknown in plants at both transcriptional and posttranslational levels. In this study, a Pi-starvation responsive GmPAP15a was identified in soybean (Glycine max). Overexpressing GmPAP15a led to significant increases in root-associated phytase activities, as well as total P content when phytate-P was supplied as the sole P resource in soybean hairy roots. Meanwhile, mass spectrometry (MS) analysis showed GmPAP15a was glycosylated at Asn144 and Asn502 , and its glycan structures of N-linked oligosaccharide chains exhibited microheterogeneity. Moreover, two homologues of AtPHR1, GmPHR9 and GmPHR32 were found to activate GmPAP15a transcription through luciferase activity analysis. Taken together, it is strongly suggested that GmPAP15a plays a vital role in phytate-P utilization in soybean, which might be regulated at both transcriptional and glycosylation modification levels. Our results highlight the GmPHR9/GmPHR32-GmPAP15a signalling pathway might present, and control phytate-P utilization in soybean.

Genome-Wide Identification of AhMDHs and Analysis of Gene Expression under Manganese Toxicity Stress in Arachis hypogaea.

Malate dehydrogenase (MDH) is one kind of oxidation-reduction enzyme that catalyzes the reversible conversion of oxaloacetic acid to malic acid. It has vital functions in plant development, photosynthesis, abiotic stress responses, and so on. However, there are no reports on the genome-wide identification and gene expression of the MDH gene family in Arachis hypogaea. In this study, the MDH gene family of A. hypogaea was comprehensively analyzed for the first time, and 15 AhMDH sequences were identified. According to the phylogenetic tree analysis, AhMDHs are mainly separated into three subfamilies with similar gene structures. Based on previously reported transcriptome sequencing results, the AhMDH expression quantity of roots and leaves exposed to manganese (Mn) toxicity were explored in A. hypogaea. Results revealed that many AhMDHs were upregulated when exposed to Mn toxicity, suggesting that those AhMDHs might play an important regulatory role in A. hypogaea's response to Mn toxicity stress. This study lays foundations for the functional study of AhMDHs and further reveals the mechanism of the A. hypogaea signaling pathway responding to high Mn stress.

Transcriptome Sequencing Analysis of Root in Soybean Responding to Mn Poisoning.

Manganese (Mn) is among one of the essential trace elements for normal plant development; however, excessive Mn can cause plant growth and development to be hindered. Nevertheless, the regulatory mechanisms of plant root response to Mn poisoning remain unclear. In the present study, results revealed that the root growth was inhibited when exposed to Mn poisoning. Physiological results showed that the antioxidase enzyme activities (peroxidase, superoxide dismutase, ascorbate peroxidase, and catalase) and the proline, malondialdehyde, and soluble sugar contents increased significantly under Mn toxicity stress (100 µM Mn), whereas the soluble protein and four hormones' (indolebutyric acid, abscisic acid, indoleacetic acid, and gibberellic acid 3) contents decreased significantly. In addition, the Mn, Fe, Na, Al, and Se contents in the roots increased significantly, whereas those of Mg, Zn, and K decreased significantly. Furthermore, RNA sequencing (RNA-seq) analysis was used to test the differentially expressed genes (DEGs) of soybean root under Mn poisoning. The results found 45,274 genes in soybean root and 1430 DEGs under Mn concentrations of 5 (normal) and 100 (toxicity) µM. Among these DEGs, 572 were upregulated and 858 were downregulated, indicating that soybean roots may initiate complex molecular regulatory mechanisms on Mn poisoning stress. The results of quantitative RT-PCR indicated that many DEGs were upregulated or downregulated markedly in the roots, suggesting that the regulation of DEGs may be complex. Therefore, the regulatory mechanism of soybean root on Mn toxicity stress is complicated. Present results lay the foundation for further study on the molecular regulation mechanism of function genes involved in regulating Mn tolerance traits in soybean roots.

Effect of plant growth regulators DA-6 and COS on drought tolerance of pineapple through bromelain and oxidative stress.

BACKGROUND: Due to global warming, drought climates frequently occur on land, and despite being drought resistant, pineapples are still subjected to varying degrees of drought stress. Plant growth regulators can regulate the stress tolerance of plants through hormonal effects. This experiment aims to investigate the regulatory effects of different plant growth regulators on Tainong- 16 and MD-2 Pineapple when subjected to drought stress. RESULTS: In this experiment, we examined the regulatory effects of two different plant growth regulators, sprayed on two pineapple varieties: MD-2 Pineapple and Tainong-16. The main component of T1 was diethyl aminoethyl hexanoate (DA-6) and that of T2 is chitosan oligosaccharide (COS). An environment similar to a natural drought was simulated in the drought stress treatments. Then, pineapples at different periods were sampled and a series of indicators were measured. The experimental results showed that the drought treatments treated with T1 and T2 plant growth regulators had a decrease in malondialdehyde, an increase in bromelain and antioxidant enzyme indicators, and an increase in phenotypic and yield indicators. CONCLUSION: This experiment demonstrated that DA-6 and COS can enhance the drought resistance of pineapple plants to a certain extent through bromelain and oxidative stress. Therefore, DA-6 and COS have potential applications and this experiment lays the foundation for further research.

Comparative Transcriptome Analysis Reveals Complex Physiological Response and Gene Regulation in Peanut Roots and Leaves under Manganese Toxicity Stress.

Excess Manganese (Mn) is toxic to plants and reduces crop production. Although physiological and molecular pathways may drive plant responses to Mn toxicity, few studies have evaluated Mn tolerance capacity in roots and leaves. As a result, the processes behind Mn tolerance in various plant tissue or organ are unclear. The reactivity of peanut (Arachis hypogaea) to Mn toxicity stress was examined in this study. Mn oxidation spots developed on peanut leaves, and the root growth was inhibited under Mn toxicity stress. The physiological results revealed that under Mn toxicity stress, the activities of antioxidases and the content of proline in roots and leaves were greatly elevated, whereas the content of soluble protein decreased. In addition, manganese and iron ion content in roots and leaves increased significantly, but magnesium ion content decreased drastically. The differentially expressed genes (DEGs) in peanut roots and leaves in response to Mn toxicity were subsequently identified using genome-wide transcriptome analysis. Transcriptomic profiling results showed that 731 and 4589 DEGs were discovered individually in roots and leaves, respectively. Furthermore, only 310 DEGs were frequently adjusted and controlled in peanut roots and leaves, indicating peanut roots and leaves exhibited various toxicity responses to Mn. The results of qRT-PCR suggested that the gene expression of many DEGs in roots and leaves was inconsistent, indicating a more complex regulation of DEGs. Therefore, different regulatory mechanisms are present in peanut roots and leaves in response to Mn toxicity stress. The findings of this study can serve as a starting point for further research into the molecular mechanism of important functional genes in peanut roots and leaves that regulate peanut tolerance to Mn poisoning.

Characterization of phosphate transporter genes and the function of SgPT1 involved in phosphate uptake in Stylosanthes guianensis.

Phosphorus (P) is one of the principal macronutrients for plant growth and productivity. Although the phosphate (Pi) transporter (PT) of the PHT1 family has been functionally characterized as participating in Pi uptake and transport in plants, information about PT genes in stylo (Stylosanthes guianensis), an important tropical forage legume that exhibits good adaptability to low-P acid soils, is limited. In this study, stylo root growth was found to be stimulated under P deficiency. The responses of PT genes to nutrient deficiencies and their roles in Pi uptake were further investigated in stylo. Four novel PT genes were identified in stylo and designated SgPT2 to SgPT5. Like SgPT1, which had been previously identified, all five SgPT proteins harboured the major facilitator superfamily (MFS) domain. Variations in tissue-specific expression were observed among the SgPT genes, which displayed diverse responses to deficiencies in nitrogen (N), P and potassium (K) in stylo roots. Four of the five SgPTs exhibited high levels of transcriptional responsiveness to P deficiency in roots. Furthermore, SgPT1, a Pi-starvation-induced gene closely related to legume PT homologues that participate in Pi transport, was selected for functional analysis. SgPT1 was localized to the plasma membrane. Analysis of transgenic Arabidopsis showed that overexpression of SgPT1 led to increased Pi accumulation and promoted root growth in Arabidopsis plants. Taken together, the results of this study suggest the involvement of SgPTs in the stylo response to nutrient deprivation. SgPT1 might mediate Pi uptake in stylo, which is beneficial for root growth during P deficiency.

Advances in Plant Lipid Metabolism Responses to Phosphate Scarcity.

Low phosphate (Pi) availability in soils severely limits crop growth and production. Plants have evolved to have numerous physiological and molecular adaptive mechanisms to cope with Pi starvation. The release of Pi from membrane phospholipids is considered to improve plant phosphorus (P) utilization efficiency in response to Pi starvation and accompanies membrane lipid remodeling. In this review, we summarize recent discoveries related to this topic and the molecular basis of membrane phospholipid alteration and triacylglycerol metabolism in response to Pi depletion in plants at different subcellular levels. These findings will help to further elucidate the molecular mechanisms underlying plant adaptation to Pi starvation and thus help to develop crop cultivars with high P utilization efficiency.

Dissection of Crop Metabolome Responses to Nitrogen, Phosphorus, Potassium, and Other Nutrient Deficiencies.

Crop growth and yield often face sophisticated environmental stresses, especially the low availability of mineral nutrients in soils, such as deficiencies of nitrogen, phosphorus, potassium, and others. Thus, it is of great importance to understand the mechanisms of crop response to mineral nutrient deficiencies, as a basis to contribute to genetic improvement and breeding of crop varieties with high nutrient efficiency for sustainable agriculture. With the advent of large-scale omics approaches, the metabolome based on mass spectrometry has been employed as a powerful and useful technique to dissect the biochemical, molecular, and genetic bases of metabolisms in many crops. Numerous metabolites have been demonstrated to play essential roles in plant growth and cellular stress response to nutrient limitations. Therefore, the purpose of this review was to summarize the recent advances in the dissection of crop metabolism responses to deficiencies of mineral nutrients, as well as the underlying adaptive mechanisms. This review is intended to provide insights into and perspectives on developing crop varieties with high nutrient efficiency through metabolite-based crop improvement.

Phosphate starvation responsive GmSPX5 mediates nodule growth through interaction with GmNF-YC4 in soybean (Glycine max).

Phosphorus (P) deficiency adversely affects nodule development as reflected by reduced nodule fresh weight in legume plants. Though mechanisms underlying nodule adaptation to P deficiency have been studied extensively, it remains largely unknown which regulator mediates nodule adaptation to P deficiency. In this study, GUS staining and quantitative reverse transcription-PCR analysis reveal that the SPX member GmSPX5 is preferentially expressed in soybean (Glycine max) nodules. Overexpression of GmSPX5 enhanced soybean nodule development particularly under phosphate (Pi) sufficient conditions. However, the Pi concentration was not affected in soybean tissues (i.e., leaves, roots, and nodules) of GmSPX5 overexpression or suppression lines, which distinguished it from other well-known SPX members functioning in control of Pi homeostasis in plants. Furthermore, GmSPX5 was observed to interact with the transcription factor GmNF-YC4 in vivo and in vitro. Overexpression of either GmSPX5 or GmNF-YC4 significantly upregulated the expression levels of five asparagine synthetase-related genes (i.e., GmASL2-6) in soybean nodules. Meanwhile, yeast one-hybrid and luciferase activity assays strongly suggested that interactions of GmSPX5 and GmNF-YC4 activate GmASL6 expression through enhancing GmNF-YC4 binding of the GmASL6 promoter. These results not only demonstrate the GmSPX5-GmNF-YC4-GmASL6 regulatory pathway mediating soybean nodule development, but also considerably improve our understanding of SPX functions in legume crops.

Complex gene regulation between young and old soybean leaves in responses to manganese toxicity.

Manganese (Mn) is an essential micronutrient for plant growth. However, excess manganese is toxic and inhibits crop production. Although it is widely known that physiological and molecular mechanisms underlie plant responses to Mn toxicity, few studies have been conducted to compare Mn tolerance capabilities between young and old leaves in plants; thus, the mechanisms underlying Mn tolerance in different plant tissues or organs are not fully understood. In this study, the dose responses of soybean to Mn availability were investigated. Genome-wide transcriptomic analysis was subsequently conducted to identify the differentially expressed genes (DEGs) in both young and old leaves of soybean in responses to Mn toxicity. Our results showed that excess Mn severely inhibited soybean growth and increased both Mn accumulation in and brown spots on soybean leaves, especially for the old leaves, strongly suggesting that more Mn was allocated to old leaves in soybean. Transcriptomic profiling revealed that totals of 4410 and 2258 DEGs were separately identified in young leaves and old leaves. Furthermore, only 944 DEGs were found to be commonly regulated in both young and old leaves of soybean, strongly suggesting distinct responses present in soybean young and old leaves in responses to Mn toxicity.

Characterization of Purple Acid Phosphatase Family and Functional Analysis of GmPAP7a/7b Involved in Extracellular ATP Utilization in Soybean.

Low phosphate (Pi) availability limits crop growth and yield in acid soils. Although root-associated acid phosphatases (APases) play an important role in extracellular organic phosphorus (P) utilization, they remain poorly studied in soybean (Glycine max), an important legume crop. In this study, dynamic changes in intracellular (leaf and root) and root-associated APase activities were investigated under both Pi-sufficient and Pi-deficient conditions. Moreover, genome-wide identification of members of the purple acid phosphatase (PAP) family and their expression patterns in response to Pi starvation were analyzed in soybean. The functions of both GmPAP7a and GmPAP7b, whose expression is up regulated by Pi starvation, were subsequently characterized. Phosphate starvation resulted in significant increases in intracellular APase activities in the leaves after 4 days, and in root intracellular and associated APase activities after 1 day, but constant increases were observed only for root intracellular and associated APase activities during day 5-16 of P deficiency in soybean. Moreover, a total of 38 GmPAP members were identified in the soybean genome. The transcripts of 19 GmPAP members in the leaves and 17 in the roots were upregulated at 16 days of P deficiency despite the lack of a response for any GmPAP members to Pi starvation at 2 days. Pi starvation upregulated GmPAP7a and GmPAP7b, and they were subsequently selected for further analysis. Both GmPAP7a and GmPAP7b exhibited relatively high activities against adenosine triphosphate (ATP) in vitro. Furthermore, overexpressing GmPAP7a and GmPAP7b in soybean hairy roots significantly increased root-associated APase activities and thus facilitated extracellular ATP utilization. Taken together, these results suggest that GmPAP7a and GmPAP7b might contribute to root-associated APase activities, thus having a function in extracellular ATP utilization in soybean.

Efficient mesophyll protoplast isolation and development of a transient expression system for castor-oil plant (Ricinus communis L.).

INTRODUCTION: We investigated the main factors affecting the efficacy of protoplast isolation, including leaf-obtaining period, cutting shapes of leaf material, enzyme concentration, enzymolysis time, and centrifugal speed. METHODS: Protoplast isolation was optimal on the condition of 20 days of leaf materials, 2-mm filament of leaves, 1.6% RS and 0.8% R-10, 80 min of enzymolysis, and 700 rpm of centrifugation, resulting in the best yield (1.19 X 106 protoplasts/g FW) and vitality (80.34%) of mesophyll protoplasts. The transient expression vector pGFPl with green fluorescent protein was transfected into the obtained protoplasts from castor by polyethylene glycol-mediated method with a transformation efficiency of 12.37%. RESULTS: Moreover, the applicability of the system for studying the subcellular localization of Re FATA (an acyl-ACP thioesterase) was validated via the protoplast isolation and transient expression protocol in this study. DISCUSSION: Collectively, the efficient mesophyll protoplast isolation and protoplast transient expression system facilitate to analyze the function of specific gene in castor (Ricinus communis L).