Microbiome-metabolomics-based insight into the protective effects of dietary fiber from sweetpotato residues on the high-fat diet-induced intestinal integrity damage.

Dietary fibers have attracted much attention due to their multiple benefits on gut health. In this work, the protective mechanism of dietary fiber from sweetpotato residues (SRDF) on the high-fat diet (HFD)-induced intestinal barrier injury was investigated using microbiome-metabolomics-based approach. The physicochemical property analysis demonstrated a thermal stability below 200 °C and porous pectin-polysaccharide structure of SRDF with high in vitro functional activities. The biochemical analysis indicated that SRDF significantly ameliorated intestinal barrier function by improving intestinal morphology and permeability and inhibiting inflammatory response. Microbiome analysis demonstrated that SRDF significantly reversed the HFD-induced dysbacteriosis, decreased the ratio of Firmicutes/Bacteroides and enhanced the relative abundance of probiotics, such as Muribaculaceae and Bifidobacteriaceae. Metabolomics analysis showed that SRDF also significantly altered the metabolic profile in the colon, wherein the differential metabolites were mainly involved in amino acid metabolism (especially tryptophan). Pearson correlation coefficient identified the beneficial relationship between intestinal microbiome and metabolome induced by SRDF. The limitation of this study was that the mouse model may not fully replicate the human intestinal responses due to the difference between the standard environmental conditions and natural world. Generally, our results implied the great potential of SRDF as a functional food ingredient.

Effects of water-soluble and water-insoluble α-glucans produced in situ by Leuconostoc citreum SH12 on physicochemical properties of fermented soymilk and their structural analysis.

This study investigated the effect of in situ produced water-soluble α-glucan (LcWSG) and water-insoluble α-glucan (LcWIG) from Leuconostoc citreum SH12 on the physicochemical properties of fermented soymilk. α-Glucans produced by Leuc. citreum SH12 improved water-holding capacity, viscosity, viscoelasticity and texture of fermented soymilk. Gtf1365 and Gtf836 of the five putative glucansucrases were responsible for synthesizing LcWSG and LcWIG during soymilk fermentation, respectively. Co-fermentation of soymilk with Gtf1365 and Gtf836 and non-exopolysaccharide-producing Lactiplantibacillus plantarum D1031 indicated that LcWSG effectively hindered the whey separation of fermented soymilk by increasing viscosity, while LcWIG improved hardness, springiness and accelerated protein coagulation. Fermented soymilk gel formation was mainly based on hydrogen bonding and hydrophobic interactions, which were promoted by both LcWSG and LcWIG. LcWIG has a greater effect on α-helix to ß-sheet translation in fermented soymilk, causing more rapid protein aggregation and thicker cross-linked gel network. Structure-based exploration of LcWSG and LcWIG from Leuc. citreum SH12 revealed their distinct roles in the physicochemical properties of fermented soymilk due to their different ratio of α-1,6 and α-1,3 glucosidic linkages and various side chain length. This study may guide the application of the water-soluble and water-insoluble α-glucans in fermented plant protein foods for their quality improvement.

Constructing Protein-Scaffolded Multienzyme Assembly Enhances the Coupling Efficiency of the P450 System for Efficient Daidzein Biosynthesis from (2S)-Naringenin.

Daidzein is a major isoflavone compound with an immense pharmaceutical value. This study applied a novel P450 CYP82D26 which can biosynthesize daidzein from (2S)-naringenin. However, the recombinant P450 systems often suffer from low coupling efficiency, leading to an electron transfer efficiency decrease and harmful reactive oxygen species release, thereby compromising their stability and catalytic efficiency. To address these challenges, the SH3-GBD-PDZ (SGP) protein scaffold was applied to assemble a multienzyme system comprising CYP82D26, P450 reductase, and NADP+-dependent aldehyde reductase in desired stoichiometric ratios. Results showed that the coupling efficiency of the P450 system was significantly increased, primarily attributed to the channeling effect of NADPH resulting from the proximity of tethered enzymes and the electrostatic interactions between NADPH and SGP. Assembling this SGP-scaffolded assembly system in Escherichia coli yielded a titer of 240.5 mg/L daidzein with an 86% (2S)-naringenin conversion rate, which showed a 9-fold increase over the free enzymes of the P450 system. These results underscore the potential application of the SGP-scaffolded multienzyme system in enhancing the coupling and catalytic efficiency of the P450 system.

Deciphering the Crucial Roles of the Quorum-Sensing Transcription Factor SdiA in NADPH Metabolism and (S)-Equol Production in Escherichia coli Nissle 1917.

The active metabolite (S)-equol, derived from daidzein by gut microbiota, exhibits superior antioxidative activity compared with its precursor and plays a vital role in human health. As only 25% to 50% of individuals can naturally produce equol when supplied with isoflavone, we engineered probiotic E. coli Nissle 1917 (EcN) to convert dietary isoflavones into (S)-equol, thus offering a strategy to mimic the gut phenotype of natural (S)-equol producers. However, co-fermentation of EcN-eq with fecal bacteria revealed that gut microbial metabolites decreased NADPH levels, hindering (S)-equol production. Transcriptome analysis showed that the quorum-sensing (QS) transcription factor SdiA negatively regulates NADPH levels and (S)-equol biosynthesis in EcN-eq. Screening AHLs showed that SdiA binding to C10-HSL negatively regulates the pentose phosphate pathway, reducing intracellular NADPH levels in EcN-eq. Molecular docking and dynamics simulations investigated the structural disparities in complexes formed by C10-HSL with SdiA from EcN or E. coli K12. Substituting sdiA_EcN in EcN-eq with sdiA_K12 increased the intracellular NADPH/NADP+ ratio, enhancing (S)-equol production by 47%. These findings elucidate the impact of AHL-QS in the gut microbiota on EcN NADPH metabolism, offering insights for developing (S)-equol-producing EcN probiotics tailored to the gut environment.

Engineering Escherichia coli for cost-effective production of medium-chain fatty acids from soy whey using an optimized galactose-based autoinduction system.

Medium-chain fatty acids (MCFAs) are essential chemical feedstocks. Microbial production of MCFAs offers an attractive alternative to conventional methods, but the costly media and external inducers limit its practical application. To address this issue and make MCFA production more cost-effective, an E.coli platform was developed using soy whey as a medium and galactose as an autoinducer. We first designed an efficient, stringent, homogeneous, and robust galactose-based autoinduction system for the expression of pathway enzymes by rationally engineering the promoter of the galactose-proton symporter (GalP). Subsequently, the intracellular acetyl-CoA availability and NADH regeneration were enhanced to improve the reversal of the ß-oxidation cycle. The resulting strain yielded 8.20 g/L and 16.42 g/L MCFA in pH-controlled batch fermentation and fed-batch fermentation with glucose added using soy whey as medium, respectively. This study provided a cost-effective and promising platform for MCFA production, as well as future strain development for other value-added chemicals production.

Biosynthesis of (S)-Equol from Soy Whey by Metabolically Engineered Escherichia coli.

(S)-Equol is one of the most bioactive metabolites of the isoflavones with immense nutritional and pharmaceutical value. Soy whey is the major liquid byproduct of the soy product processing industries that is rich in nutrients and (S)-equol biosynthetic precursor daidzin. However, it is usually disposed into the sewage, causing high environmental contamination. Herein, we constructed a recombinant Escherichia coli for the biosynthesis of (S)-equol from soy whey. First, we evaluated daidzin-specific transporters and optimized the anaerobically induced Pnar in the (S)-equol biosynthesis cassette to produce (S)-equol from daidzin. Then, sucrase and α-galactosidase were co-expressed to confer sucrose, stachyose, and raffinose utilization capacity on E. coli. Meanwhile, EIIBCAglc was inactivated to eliminate the daidzin transport inhibition induced by glucose. Finally, combining these strategies and optimizing the fermentation conditions, the optimal strain produced 91.5 mg/L of (S)-equol with a yield of 0.96 mol/mol substrates in concentrated soy whey. Overall, this new strategy is an attractive route to broaden the applications of soy whey and achieve the eco-friendly production of (S)-equol.

Tofu by-product soy whey substitutes urea: Reduced ammonia volatilization, enhanced soil fertility and improved fruit quality in cherry tomato production.

Soy whey is an abundant, nutrient-rich and safe wastewater produced in tofu processing, so it is necessary to valorize it instead of discarding it as sewage. Whether soy whey can be used as a fertilizer substitute for agricultural production is unclear. In this study, the effects of soy whey serving as a nitrogen source to substitute urea on soil NH3 volatilization, dissolved organic matter (DOM) components and cherry tomato qualities were investigated by soil column experiment. Results showed that the soil NH4+-N concentrations and pH values of the 50% soy whey fertilizer combined with 50% urea (50%-SW) and 100% soy whey fertilizer (100%-SW) treatments were lower than those of 100% urea treatment (CKU). Compared with CKU, 50%-SW and 100%-SW treatments increased the abundance of ammonia oxidizing bacteria (AOB) by 6.52-100.89%, protease activity by 66.22-83.78%, the contents of total organic carbon (TOC) by 16.97-35.64%, humification index (HIX) of soil DOM by 13.57-17.99%, and average weight per fruit of cherry tomato by 13.46-18.56%, respectively. Moreover, soy whey as liquid organic fertilizer reduced the soil NH3 volatilization by 18.65-25.27% and the fertilization cost by 25.94-51.87% compared with CKU. This study provides a promising option with economic and environmental benefits for soy whey utilization and cherry tomato production, which contributes to the win-win effectiveness of sustainable production for both the soy products industry and agriculture.

The Next Frontier of 3D Bioprinting: Bioactive Materials Functionalized by Bacteria.

3D bioprinting has become a flexible technical means used in many fields. Currently, research on 3D bioprinting is mainly focused on the use of mammalian cells to print organ and tissue models, which has greatly promoted progress in the fields of tissue engineering, regenerative medicine, and pharmaceuticals. In recent years, bacterial bioprinting has gradually become a rapidly developing research fields, with a wide range of potential applications in basic research, biomedicine, bioremediation, and other field. Here, this works reviews new research on bacterial bioprinting, and discuss its future research direction.

Sustainable production of genistin from glycerol by constructing and optimizing Escherichia coli.

Genistin is one of the bioactive isoflavone glucosides found in legumes, which have great nutraceutical and pharmaceutical significance. The market available isoflavones are currently produced by direct plant extraction. However, its low abundance in plant and structural complexity hinders access to this phytopharmaceutical via plant extraction or chemical synthesis. Here, the E. coli cell factory for sustainable production of genistin from glycerol was constructed. First, we rebuilt the precursor genistein biosynthesis pathway in E. coli, and its titer was then increased by 668% by identifying rate-limiting steps and applying an artificial protein scaffold system. Then de novo production of genistin from glycerol was achieved by functional screening and introduction of glycosyl-transferases, UDP-glucose pathway and specific genistin efflux pumps, and 48.1 mg/L of genistin was obtained. A further engineered E. coli strain equipped with an improved malonyl-CoA pathway, alternative glycerol-utilization pathways, acetyl-CoA carboxylase (ACC), and CRISPR interference (CRISPRi) mediated regulation produced up to 137.8 mg/L of genistin in shake flask cultures. Finally, 202.7 mg/L genistin was achieved through fed-batch fermentation in a 3-L bioreactor. This study represents the de novo genistin production from glycerol for the first time and will lay the foundation for low-cost microbial production of glucoside isoflavones. In addition, the multiphase workflow may provide a reference for engineering the biosynthetic pathways in other microbial hosts as well, for green manufacturing of complex natural products.

Soy Isoflavones Protect Neuronal PC12 Cells against Hypoxic Damage through Nrf2 Activation and Suppression of p38 MAPK and AKT-mTOR Pathways.

Isoflavones are a class of major phenolic compounds, derived from soybeans, that possess unique therapeutic and biological properties. The possible mechanisms of isoflavone-mediated protection of neuronal PC12 cells against hypoxic damage was investigated in this study. Isoflavones showed potential neuroprotective effects by increasing cell viability, decreasing the level of reactive oxygen species (ROS), and inhibiting apoptosis and cell cycle arrest in cobalt chloride (CoCl2)-induced hypoxic damage. A Western blot analysis indicated that isoflavones decreased apoptosis by up-regulating the Bcl-xL protein and down-regulating the Bax protein. They further reduced the S-phase fraction of the cell cycle by down-regulating the p21 protein and up-regulating the cyclin A protein levels. Additionally, isoflavones activated Nrf2 protein translocation and inhibited the p38 MAPK and AKT-mTOR pathways. A molecular docking analysis further revealed that isoflavones displayed a potential competitive interaction with the Nrf2 protein for Keap1. Our findings suggest that isoflavones could be a potent neuroprotective phytochemical in soybeans and their products.

Soybean Whey Bio-Processed Using Weissella hellenica D1501 Protects Neuronal PC12 Cells Against Oxidative Damage.

Soybean whey, as a byproduct of soybean industry, has caused considerable concern recently because of its abundant nutrients. To further utilize soybean whey, it was fermented with Weissella hellenica D1501, and the neuroprotective potency of this beverage was studied in the present work. The phenolic profile and antioxidant capacity of fermented soybean whey (FSBW) were analyzed. The neuroprotective effects were evaluated based on the hydrogen peroxide-stimulated oxidative damage model in a neural-like cell (PC12). Results demonstrated that soybean whey's phenolic contents and antioxidant activities were markedly improved after fermentation. Glycoside isoflavones were efficiently converted into aglycones by W. hellenica D1501. FSBW extract apparently increased cell viability, decreased reactive oxide species levels, and protected antioxidant enzymes in oxidative damage. Furthermore, FSBW effectively reduced apoptosis rate by inhibiting Bax protein and improving Bcl-2 and Bcl-xL proteins. FSBW ameliorated the cell cycle through the decrease of p21 protein and an increase of cyclin A protein. The findings of this study thus suggested that W. hellenica D1501-fermented soybean whey could potentially protect nerve cells against oxidative damage.

Neuroprotective potency of a soy whey fermented by Cordyceps militaris SN-18 against hydrogen peroxide-induced oxidative injury in PC12 cells.

PURPOSE: Soy whey is a byproduct generated from the processing of several soybean products. Its valorization has continued to attract significant research interest in recent times due to the nutritional and bioactive potency of its chemical composition. Herein, the neuroprotective potency of a soy whey fermented by Cordyceps militaris SN-18 against hydrogen peroxide (H2O2)-induced oxidative injury in PC12 cells was investigated. METHODS: The phenolic compositions were analyzed by high-performance liquid chromatography. Antioxidant activities were assessed by ABTS•+ scavenging assay, DPPH radical scavenging assay, reducing power assay, and ferric reducing antioxidant power assay. The neuroprotective effects of fermented soy whey (FSW) were investigated based on the oxidative injury model in PC12 cells. RESULTS: FSW possessed higher total phenolic content and antioxidant activities compared with unfermented soy whey (UFSW) and that most of the isoflavone glycosides were hydrolyzed into their corresponding aglycones during fermentation. The extract from FSW exhibited a greater protective effect on PC12 cells against oxidative injury by promoting cell proliferation, restoring cell morphology, inhibiting lactic dehydrogenase leakage, reducing reactive oxygen species levels, and enhancing antioxidant enzyme activities compared with that from UFSW. Additionally, cell apoptosis was significantly inhibited by FSW through down-regulation of caspase-3, caspase-9, and Bax and up-regulation of Bcl-2 and Bcl-xL. S-phase cell arrest was attenuated by FSW through increasing cyclin A, CDK1 and CDK2, and decreasing p21 protein. CONCLUSION: Fermentation with C. militaris SN-18 could significantly improve the bioactivity of soy whey by enhancing the ability of nerve cells to resist oxidative damage.

Neuroprotective Potency of Tofu Bio-Processed Using Actinomucor elegans against Hypoxic Injury Induced by Cobalt Chloride in PC12 Cells.

Fermented soybean products have attracted great attention due to their health benefits. In the present study, the hypoxia-injured PC12 cells induced by cobalt chloride (CoCl2) were used to evaluate the neuroprotective potency of tofu fermented by Actinomucor elegans (FT). Results indicated that FT exhibited higher phenolic content and antioxidant activity than tofu. Moreover, most soybean isoflavone glycosides were hydrolyzed into their corresponding aglycones during fermentation. FT demonstrated a significant protective effect on PC12 cells against hypoxic injury by maintaining cell viability, reducing lactic dehydrogenase leakage, and inhibiting oxidative stress. The cell apoptosis was significantly attenuated by the FT through down-regulation of caspase-3, caspases-8, caspase-9, and Bax, and up-regulation of Bcl-2 and Bcl-xL. S-phase cell arrest was significantly inhibited by the FT through increasing cyclin A and decreasing the p21 protein level. Furthermore, treatment with the FT activated autophagy, indicating that autophagy possibly acted as a survival mechanism against CoCl2-induced injury. Overall, FT offered a potential protective effect on nerve cells in vitro against hypoxic damage.

Composition, antioxidant activity, and neuroprotective effects of anthocyanin-rich extract from purple highland barley bran and its promotion on autophagy.

Anthocyanin-rich purple highland barley has attracted great attention recently due to its health benefits in humans. The composition of the purified anthocyanin extract (PAE) from purple highland barley bran (PHBB) was characterized by liquid chromatography-mass spectrometry (LC-MS) with a high acylated anthocyanin profile. PAE exhibited high antioxidant activity and potential neuroprotective effects on cobalt chloride (CoCl2)-induced hypoxic damage in PC12 cells by maintaining cell viability, restoring cell morphology, inhibiting lactic dehydrogenase (LDH) leakage, reducing reactive oxygen species (ROS) levels, enhancing antioxidant enzyme activities, inhibiting cell apoptosis, and attenuating cell cycle arrest. Treatment cells (PC12 and U2OS) with PAE activated autophagy, indicating that autophagy possibly acted as a survival mechanism against CoCl2-induced injury. This study demonstrated that PAE from the PHBB was a high-quality natural functional food colorant and potentially could be used as a preventive agent for brain dysfunction caused by hypoxic damage.

Effects of lactic acid bacteria fermented yellow whey on the protein coagulation and isoflavones distribution in soymilk.

This study investigated the soymilk coagulation induced by fermented yellow whey (FYW), which is extensively used as a natural tofu coagulant in China. The aggregations involving proteins and isoflavone particles caused by FYW were analyzed using the proteomic technology and high-performance liquid chromatography, respectively. As indicated, the FYW-induced coagulation of soy proteins mainly occurred at pH 5.80-5.90. When the pH of soymilk decreased, the 7S ß, 11S A3 and some of 11S A1a subunits and SBP, Bd, lectin and TA aggregated the earliest, and later did the 11S A4, other 11S A1a, 11S A2 and 11S A1b subunits. The 7S α and α' subunits and TB showed an obvious delay in aggregation. Moreover, isoflavones in the form of aglycones were more likely to coprecipitate with proteins, compared with glycosides. These results could provide an important reference and assistance for future research on the development of traditional FYW-tofu.

Multistarter fermentation of glutinous rice with Fu brick tea: Effects on microbial, chemical, and volatile compositions.

A higher fermentation efficiency was achieved, using multistarter fermentation of glutinous rice supplemented with Fu brick tea (FGR-FBT), than when using traditional fermentation. The effects of multistarter fermentation on the microbial, chemical, and volatile compositions were determined. When FBT was incorporated during glutinous rice fermentation, increased population of yeasts and fungi, as well as enhanced α-amylase, proteinase and ß-glucosidase activities, were observed. Specific fungi were isolated and identified as Aspergillus spp., which are known to secrete extracellular enzymes that modify the chemical properties, including ethanol levels, pH, total acids, and total soluble solids. The aroma profile of fermented glutinous rice was studied in the absence and presence of FBT, using HS-SPME-GC-MS and the electronic-nose. This analysis indicated that 35 characteristic volatile compounds were only found in FGR-FBT. The results show that FBT can be added during the fermentation of food products to enhance microbial biotransformation and modify flavour metabolism.

Increased Phenolic Content and Enhanced Antioxidant Activity in Fermented Glutinous Rice Supplemented with Fu Brick Tea.

Glutinous rice-based foods have a long history are consumed worldwide. They are also in great demand for the pursuit of novel sensory and natural health benefits. In this study, we developed a novel fermented glutinous rice product with the supplementation of Fu brick tea. Using in vitro antioxidant evaluation and phenolic compounds analysis, fermentation with Fu brick tea increased the total phenolic content and enhanced the antioxidant activity of glutinous rice, including scavenging of 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) radical, 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical, and hydroxyl radical, ferric-reducing antioxidant power, and ferric ion reducing power and iron chelating capability. Besides, compared with traditional fermented glutinous rice, this novel functional food exhibited a stronger activity for protecting DNA against hydroxyl radical-induced oxidation damage. Quantitative analysis by HPLC identified 14 compounds covering catechins and phenolic acids, which were considered to be positively related to the enhanced antioxidant capability. Furthermore, we found that 80% ethanol was a suitable extract solvent compared with water, because of its higher extraction efficiency and stronger functional activities. Our results suggested that this novel fermented glutinous rice could serve as a nutraceutical food/ingredient with special sensory and functional activities.

Improving medium chain fatty acid production in Escherichia coli by multiple transporter engineering.

Medium-chain fatty acids (C6-C10, MCFAs) represent valuable molecules for food and pharmaceutical industries. However, MCFAs in Escherichia coli induce stresses and toxicity, reducing final yields. These phenotypes could be alleviated if MCFA export from cells was accelerated, yet information on proteins responsible for exporting MCFAs remains limited. Here, a panel of 17 genes or operons was screened. Overexpressing resistance nodulation cell division family transporter acrE, mdtE and mdtC increased MCFA titer by 46.4%, 65.2% and 33.8%, whereas overexpressing transcriptional activators soxS, marA and multidrug efflux pumps cmr, fadL decreased MCFA titer by 42.5%, 27.8%, 37.6% and 29.5%, respectively. Furthermore, the effect of multiple transporter engineering was investigated. Overexpressing acrE, mdtE and mdtC together with deleting cmr increased MCFA production by more than two-fold. This study demonstrated the efficient efflux pump specifically for MCFAs and this multiple transporter engineering strategy can be employed to synthesize other bio-products toxic to microorganisms.

Improving metabolic efficiency of the reverse beta-oxidation cycle by balancing redox cofactor requirement.

Previous studies have made many exciting achievements on pushing the functional reversal of beta-oxidation cycle (r-BOX) to more widespread adoption for synthesis of a wide variety of fuels and chemicals. However, the redox cofactor requirement for the efficient operation of r-BOX remains unclear. In this work, the metabolic efficiency of r-BOX for medium-chain fatty acid (C6-C10, MCFA) production was optimized by redox cofactor engineering. Stoichiometric analysis of the r-BOX pathway and further experimental examination identified NADH as a crucial determinant of r-BOX process yield. Furthermore, the introduction of formate dehydrogenase from Candida boidinii using fermentative inhibitor byproduct formate as a redox NADH sink improved MCFA titer from initial 1.2g/L to 3.1g/L. Moreover, coupling of increasing the supply of acetyl-CoA with NADH to achieve fermentative redox balance enabled product synthesis at maximum titers. To this end, the acetate re-assimilation pathway was further optimized to increase acetyl-CoA availability associated with the new supply of NADH. It was found that the acetyl-CoA synthetase activity and intracellular ATP levels constrained the activity of acetate re-assimilation pathway, and 4.7g/L of MCFA titer was finally achieved after alleviating these two limiting factors. To the best of our knowledge, this represented the highest titer reported to date. These results demonstrated that the key constraint of r-BOX was redox imbalance and redox engineering could further unleash the lipogenic potential of this cycle. The redox engineering strategies could be applied to acetyl-CoA-derived products or other bio-products requiring multiple redox cofactors for biosynthesis.

Overexpression of the nitrate transporter, OsNRT2.3b, improves rice phosphorus uptake and translocation.

KEY MESSAGE: Overexpression of OsNRT2.3b in rice can increase Pi uptake and accumulation through advanced root system, enhanced OsPT and OsPHR genes expression, and the phloem pH homeostasis. Nitrogen (N) and phosphorus (P) are two essential macronutrients for plants. Overexpression of the rice nitrate transporter, OsNRT2.3b, can improve rice grain yield and nitrogen use efficiency (NUE). Here, OsNRT2.3b overexpression resulted in increased grain yield, straw yield, and grain:straw ratio, accompanied by increased P concentrations in the leaf blade, leaf sheath, culm, and unfilled rice hulls. Overexpression of OsNRT2.3b significantly increased 33Pi uptake compared with WT under 300-µM Pi but not 10-µM Pi condition in 24 h. Moreover, the OsNRT2.3b-overexpressing rice lines showed increased root and shoot biomass, root:shoot ratio, total root length root surface area and N, P accumulation under 300- and 10-µM Pi supply in hydroponic solution. The levels of OsPT2, OsPT8, and OsPHR2 expression in roots and of OsPT1 and OsPHR2 in shoots were upregulated in OsNRT2.3b-overexpressing rice. These results indicated that OsNRT2.3b overexpression can improve rice P uptake and accumulation, partially through the advanced root system, enhanced gene expression, and the phloem pH regulation function.