Elucidating the role of exogenous melatonin in mitigating alkaline stress in soybeans across different growth stages: a transcriptomic and metabolomic approach.

BACKGROUND: Soybean (Glycine max), a vital grain and oilseed crop, serves as a primary source of plant protein and oil. Soil salinization poses a significant threat to soybean planting, highlighting the urgency to improve soybean resilience and adaptability to saline stress. Melatonin, recently identified as a key plant growth regulator, plays crucial roles in plant growth, development, and responses to environmental stress. However, the potential of melatonin to mitigate alkali stress in soybeans and the underlying mechanisms remain unclear. RESULTS: This study investigated the effects of exogenous melatonin on the soybean cultivar Zhonghuang 13 under alkaline stress. We employed physiological, biochemical, transcriptomic, and metabolomic analyses throughout both vegetative and pod-filling growth stages. Our findings demonstrate that melatonin significantly counteracts the detrimental effects of alkaline stress on soybean plants, promoting plant growth, photosynthesis, and antioxidant capacity. Transcriptomic analysis during both growth stages under alkaline stress, with and without melatonin treatment, identified 2,834 and 549 differentially expressed genes, respectively. These genes may play a vital role in regulating plant adaptation to abiotic stress. Notably, analysis of phytohormone biosynthesis pathways revealed altered expression of key genes, particularly in the ARF (auxin response factor), AUX/IAA (auxin/indole-3-acetic acid), and GH3 (Gretchen Hagen 3) families, during the early stress response. Metabolomic analysis during the pod-filling stage identified highly expressed metabolites responding to melatonin application, such as uteolin-7-O-(2''-O-rhamnosyl)rutinoside and Hederagenin-3-O-glucuronide-28-O-glucosyl(1,2)glucoside, which helped alleviate the damage caused by alkali stress. Furthermore, we identified 183 differentially expressed transcription factors, potentially playing a critical role in regulating plant adaptation to abiotic stress. Among these, the gene SoyZH13_04G073701 is particularly noteworthy as it regulates the key differentially expressed metabolite, the terpene metabolite Hederagenin-3-O-glucuronide-28-O-glucosyl(1,2)glucoside. WGCNA analysis identified this gene (SoyZH13_04G073701) as a hub gene, positively regulating the crucial differentially expressed metabolite of terpenoids, Hederagenin-3-O-glucuronide-28-O-glucosyl(1,2)glucoside. Our findings provide novel insights into how exogenous melatonin alleviates alkali stress in soybeans at different reproductive stages. CONCLUSIONS: Integrating transcriptomic and metabolomic approaches, our study elucidates the mechanisms by which exogenous melatonin ameliorates the inhibitory effects of alkaline stress on soybean growth and development. This occurs through modulation of biosynthesis pathways for key compounds, including terpenes, flavonoids, and phenolics. Our findings provide initial mechanistic insights into how melatonin mitigates alkaline stress in soybeans, offering a foundation for molecular breeding strategies to enhance salt-alkali tolerance in this crop.

A new experimental setup for combined fast differential scanning calorimetry and X-ray photon correlation spectroscopy.

Synchrotron-radiation-based techniques are a powerful tool for the investigation of materials. In particular, the availability of highly brilliant sources has opened the possibility to develop techniques sensitive to dynamics at the atomic scale such as X-ray photon correlation spectroscopy (XPCS). XPCS is particularly relevant in the study of glasses, which have been often investigated at the macroscopic scale by, for example, differential scanning calorimetry. Here, we show how to adapt a Flash calorimeter to combine XPCS and calorimetric scans. This setup paves the way to novel experiments requiring dynamical and thermodynamic information, ranging from the study of the crystallization kinetics to the study of the glass transition in systems that can be vitrified thanks to the high cooling rates reachable with an ultrafast calorimeter.

Genome Survey and Chromosome-Level Draft Genome Assembly of Glycine max var. Dongfudou 3: Insights into Genome Characteristics and Protein Deficiencies.

Dongfudou 3 is a highly sought-after soybean variety due to its lack of beany flavor. To support molecular breeding efforts, we conducted a genomic survey using next-generation sequencing. We determined the genome size, complexity, and characteristics of Dongfudou 3. Furthermore, we constructed a chromosome-level draft genome and speculated on the molecular basis of protein deficiency in GmLOX1, GmLOX2, and GmLOX3. These findings set the stage for high-quality genome analysis using third-generation sequencing. The estimated genome size is approximately 1.07 Gb, with repetitive sequences accounting for 72.50%. The genome is homozygous and devoid of microbial contamination. The draft genome consists of 916.00 Mb anchored onto 20 chromosomes, with annotations of 46,446 genes and 77,391 transcripts, achieving Benchmarking Single-Copy Orthologue (BUSCO) completeness of 99.5% for genome completeness and 99.1% for annotation. Deletions and substitutions were identified in the three GmLox genes, and they also lack corresponding active proteins. Our proposed approach, involving k-mer analysis after filtering out organellar DNA sequences, is applicable to genome surveys of all plant species, allowing for accurate assessments of size and complexity. Moreover, the process of constructing chromosome-level draft genomes using closely related reference genomes offers cost-effective access to valuable information, maximizing data utilization.

Multitarget therapy with a corticosteroid, cyclosporine and mycophenolate mofetil for idiopathic membranous nephropathy: a prospective randomized controlled trial.

BACKGROUND: The effectiveness of multitarget combination therapy with a corticosteroid, cyclosporine and mycophenolate mofetil for idiopathic membranous nephropathy (IMN) is unclear. In the present study, we aimed to compare the efficacy and safety of multitarget therapy with a cyclical corticosteroid-cyclophosphamide regimen in patients with IMN. METHODS: This was a single-centre, prospective, randomized, controlled trial. We randomly assigned patients with IMN to receive multitarget therapy (a combination of prednisone, cyclosporine and mycophenolate mofetil) or 6-month cyclical treatment with a corticosteroid and cyclophosphamide. The study patients were followed up for 12 months. The primary outcome was a composite of complete or partial remissions at 12 months. Adverse events were also assessed. RESULTS: The study cohort comprised 78 patients, 39 of whom received multitarget therapy and the other 39 cyclical alternating treatment with a corticosteroid and cyclophosphamide. At 12 months, 31 of 39 patients (79%) in the multitarget therapy group and 34 of 39 (87%) in the corticosteroid-cyclophosphamide group had achieved complete or partial remissions (relative risk 0.93; 95% confidence interval 0.72-1.21; P = .85; log-rank test). The prevalence of adverse events was significantly lower in the multitarget therapy group than in the corticosteroid-cyclophosphamide group [46% (18 of 39) vs 74% (29 of 39); P < .05]. CONCLUSIONS: Multitarget therapy for IMN patients is noninferior to cyclical alternating treatment with corticosteroid and cyclophosphamide in inducing proteinuria remission and has a better safety profile than the corticosteroid-cyclophosphamide combination.

Altered Cingulum Functioning in Major Depressive Disorder Patient With Suicide Attempts: A Resting-State Functional Magnetic Resonance Imaging Study.

Background: Major depressive disorder (MDD) with suicide attempts (SA) poses a significant public health issue. This study aims to identify neurobiological markers for MDD with SA on resting-state brain functional magnetic resonance imaging (rs-fMRI). Methods: Fifty-one unmedicated adult MDD participants, 27 with SA on the Beck Scale for Suicidal Ideation and 24 without SA, underwent rs-fMRI scanning. A group of 30 healthy controls (HC) matched for age, gender, and education-level with MDD were chosen. A whole brain analysis of regional homogeneity (ReHo) was performed on subjects to identify regions where brain activity was associated with SA. Multiple comparison analysis was performed for ReHo. Pearson's correlation analysis was performed between HAMD-SA scores and ReHo. The statistical significance level was set at p < 0.05. Results: We examined whether there were significant differences among the three groups in whole brain ReHo during resting state. Subjects with SA showed significant increase of ReHo in the right Cingulum Post in comparison with those without SA. Subjects with SA showed significant decrease of ReHo in the right Cingulate Gyrus/Precuneus in comparison with HC. The mean ReHo from the significant brain region was associated with HAMD-SA (item 3 of the HAMD) scores (r = 0.349, P = 0.012) but was not associated with HAMD-24 scores. Conclusion: These results indicate that SA is associated with altered resting-state brain activity. The pattern of elevated activity in the cingulum functioning may be related to SA. Identifying cingulum activity associated with SA may help to elucidate its pathogenesis and etiology.

The Efficacy of an Initial Prednisone Monotherapy Regimen on PLA2R-associated Idiopathic Membranous Nephropathy.

The aim of this study was to evaluate the efficacy and safety of prednisone monotherapy for phospholipase A2 receptor (PLA2R)-associated idiopathic membranous nephropathy (IMN). This was a retrospective cohort study involving 32 patients enrolled between January 2017 and June 2019 at Beijing Friendship Hospital, Capital Medical University, Beijing, China. Seventeen patients received prednisone monotherapy, and 15 received the Ponticelli regimen. The primary outcome was nephrotic syndrome remission at 12th month. The secondary outcomes were the incidence of adverse events and recurrence over the 12-month follow-up. At 12th month, 16/17 patients (94.1%) in the prednisone monotherapy group and 14/15 patients (93.3%) in the Ponticelli regimen group achieved remission (p = 0.19). The adverse events occurred in 9/17 and 9/15 of the patients receiving prednisone monotherapy and the Ponticelli regimen, respectively (p = 0.74). Four and three patients relapsed in the prednisone monotherapy and Ponticelli regimen groups, respectively (p = 0.99). Initial prednisone monotherapy had similar efficacy and safety compared with the Ponticelli regimen for PLA2R-associated IMN. Key Words: PLA2R-associated idiopathic membranous nephropathy, Ponticelli regimen, Prednisone monotherapy.

Leaf transcriptomic response mediated by cold stress in two maize inbred lines with contrasting tolerance levels.

Maize (Zea mays L.) is a thermophilic plant and a minor drop in temperature can prolong the maturity period. Plants respond to cold stress through structural and functional modification in cell membranes as well as changes in the photosynthesis and energy metabolism. In order to understand the molecular mechanisms underlying cold tolerance and adaptation, we employed leaf transcriptome sequencing together with leaf microstructure and relative electrical conductivity measurements in two maize inbred lines, having different cold stress tolerance potentials. The leaf physiological and transcriptomic responses of maize seedlings were studied after growing both inbred lines at 5 °C for 0, 12 and 24 h. Differentially expressed genes were enriched in photosynthesis antenna proteins, MAPK signaling pathway, plant hormone signal transduction, circadian rhythm, secondary metabolites related pathways, ribosome, and proteasome. The seedlings of both genotypes employed common stress responsive pathways to respond to cold stress. However, the cold tolerant line B144 protected its photosystem II from photooxidation by upregulating D1 proteins. The sensitive line Q319 was unable to close its stomata. Collectively, B144 exhibited a cold tolerance owing to its ability to mediate changes in stomata opening as well as protecting photosystem. These results increase our understanding on the cold stress tolerance in maize seedlings and propose multiple key regulators of stress responses such as modifications in photosystem II, stomata guard cell opening and closing, changes in secondary metabolite biosynthesis, and circadian rhythm. This study also presents the signal transduction related changes in MAPK and phytohormone signaling pathways in response to cold stress during seedling stage of maize.

Crystal structure of hydroxyquinol 1,2-dioxygenase PnpC from Pseudomonas putida DLL-E4 and its role of N-terminal domain for catalysis.

Hydroxyquinol 1,2-dioxygenase is a key enzyme in the hydroxyquinol pathway of p-nitrophenol (PNP) degradation, and catalyzes the ring cleavage of benzenetriol to maleylacetate. Here, we report the first structure of a hydroxyquinol 1,2-dioxygenase from the Gram-negative bacterium Pseudomonas putida DLL-E4 (PnpC) at the resolution of 2.1 Å. The tertiary structure of PnpC resembles that of the homologous intradiol dioxygenases. The catalytic Fe(III) is pentacoordinated by the conserved Tyr160, Tyr194, His218 and His220, the citrate anion and one water molecule. Among the residues expected to interact with the substrate, structural comparison with the (chloro)catechol dioxygenases suggested that Asp80, Thr81 and Val248 are responsible for the substrate specificity. Moreover, truncation of the N-terminal α-helix of PnpC suggested the N-terminal domain is required for its soluble expression and enzyme catalysis. Our results might provide insights in the substrate recognition and rational design of this enzyme class to be used in bioremediation.

Crystal structure of p-nitrophenol 4-monooxygenase PnpA from Pseudomonas putida DLL-E4: The key enzyme involved in p-nitrophenol degradation.

p-Nitrophenol 4-monooxygenase PnpA, the key enzyme in the hydroquinone pathway of p-nitrophenol (PNP) degradation, catalyzes the monooxygenase reaction of PNP to p-benzoquinone in the presence of FAD and NADH. Here, we determined the first crystal structure of PnpA from Pseudomonas putida DLL-E4 in its apo and FAD-complex forms to a resolution of 2.04 Šand 2.48 Å, respectively. The PnpA structure shares a common fold with hydroxybenzoate hydroxylases, despite a low amino sequence identity of 14-18%, confirming it to be a member of the Class A flavoprotein monooxygenases. However, substrate docking studies of PnpA indicated that the residues stabilizing the substrate in an orientation suitable for catalysis are not observed in other homologous hydroxybenzoate hydroxylases, suggesting PnpA employs a unique catalytic mechanism. This work expands our understanding on the reaction mode for this enzyme class.

PnpB involvement in the regulation of temperature-sensitive para-nitrophenol degradation in Pseudomonas putida MT54 via PnpA.

Pseudomonas putida DLL-E4 can efficiently degrade para-nitrophenol and its intermediate metabolite hydroquinone at 37 °C and 30 °C. However, mutant strain Pseudomonas putida MT54, obtained by transposon mutagenesis from P. putida DLL-E4, could not degrade para-nitrophenol at 37 °C. The mutant genes including DW66_0143, DW66_0153 and pnpB were discovered in strain MT54 by whole genome resequencing. Gene knockout and complementation confirmed the necessity of PnpB in PNP degradation by temperature-sensitive strain MT54. PnpA catalyzes the first step in complete degradation of PNP, and we found its activity was significantly enhanced by PnpB. The measurement of bacterial two-hybrid system indicated that the effect was not mediated by the direct interaction between PnpA and PnpB, but caused by the elimination of product inhibition of PnpA. Furthermore, PnpA was characterized as a psychrophilic enzyme with optimum temperature of 20 °C. We concluded that the lowered activity of PnpA resulted from inactivation of PnpB at the restrictive temperature induced the temperature-sensitive characteristic of P. putida MT54.

Characterization of a novel GH36 α-galactosidase from Bacillus megaterium and its application in degradation of raffinose family oligosaccharides.

A novel α-galactosidase gene (agaB) from Bacillus megaterium 3-7 was cloned and expressed in Escherichia coli. The gene coded for a protein with 741 amino acids and a calculated molecular mass of 85.4kDa. The native structure of the recombined AgaB was determined to be a homotrimer. AgaB showed the highest identity of 57% with the characterized glycosyl hydrolase family 36 α-galactosidase from Clostridium stercorarium F-9. The enzyme exhibited a specific activity of 362.6U/mg at 37°C and pH 6.8. The enzyme showed strong resistance to proteases and great tolerance to galactose (Ki=12.5mM). AgaB displayed wide substrate specificity toward pNPGal, melibiose, raffinose and stachyose, with a Km of 0.42, 12.1, 17.0 and 25.4mM, respectively. Furthermore, AgaB completely hydrolyzed raffinose and stachyose present in soybean milk at 37°C within 4h when combined with trypsin. These favorable properties make AgaB a potential candidate for applications in the food and feed industries.