Changes in carbohydrate metabolism and endogenous hormone regulation during bulblet initiation and development in Lycoris radiata.

BACKGROUND: Lycoris species have great ornamental and medicinal values; however, their low regeneration efficiency seriously restricts their commercial production. Understanding the mechanism of bulblet propagation in this genus, which has remained underexplored to date, could provide a theoretical basis for improving the reproductive efficiency. Therefore, we studied the bulblet initiation and developmental processes in Lycoris radiata. RESULTS: We found that bulblets are formed on the junctions of the innermost layers of scales and the basal plate, and initially present as an axillary bud and gradually develop into a bulblet. We also determined the changes in carbohydrate and endogenous hormone contents during bulblet initiation and development, as well as the expression patterns of genes involved in carbohydrate metabolism and hormone biosynthesis and signaling through transcriptome analysis. Soluble sugars derived from starch degradation in the outer scales are transported to and promote bulblet initiation and development through starch synthesis in the inner scales. This process is mediated by several genes involved in carbohydrate metabolism, especially genes encoding ADP glucose pyrophosphorylase, a crucial starch synthesis enzyme. As for hormones, endogenous IAA, GA, and ABA content showed an increase and decrease during bulblet initiation and development, respectively, which were consistent with the expression patterns of genes involved in IAA, GA, and ABA synthesis and signal transduction. In addition, a decrease in ZR content may be down- and up-regulated by CK biosynthesis and degradation related genes, respectively, with increasing auxin content. Furthermore, expression levels of genes related to BR, JA, and SA biosynthesis were increased, while that of ethylene biosynthesis genes was decreased, which was also consistent with the expression patterns of their signal transduction genes. CONCLUSIONS: The present study provides insights into the effect of carbohydrate metabolism and endogenous hormone regulation on control of L. radiata bulblet initiation and development. Based on the results, we propose several suggestions to improve L. radiata propagation efficiency in production, which will provide directions for future research.

The interaction between nitrogen availability and auxin, cytokinin, and strigolactone in the control of shoot branching in rice (Oryza sativa L.).

KEY MESSAGE: Nitrogen availability and cytokinin could promote shoot branching in rice, whereas auxin and strigolactone inhibited it. The interaction between nitrogen availability and the three hormones is discussed. Rice shoot branching is strongly affected by nitrogen availability and the plant hormones auxin, cytokinin, and strigolactone; however, the interaction of them in the regulation of rice shoot branching remains a subject of debate. In the present study, nitrogen and the three hormones were used to regulate rice tiller bud growth in the indica rice variety Yangdao 6. Both nitrogen and CK promoted shoot branching in rice, whereas auxin and SL inhibited it. We used HPLC to determine the amounts of endogenous IAA and CK, and we used quantitative real-time PCR analysis to quantify the expression levels of several genes. Nitrogen enhanced the amount of CK by promoting the expression levels of OsIPTs in nodes. In addition, both nitrogen and CK downregulated the expression of genes related to SL synthesis in root and nodes, implying that the inhibition of SL synthesis by nitrogen may occur at least partially through the CK pathway. SL did not significantly reduce the amount of CK or the expression levels of OsIPT genes, but it did significantly reduce the amount of auxin and the auxin transport capacity in nodes. Auxin itself inhibited CK synthesis and promoted SL synthesis in nodes rather than in roots. Furthermore, we found that CK and SL quickly reduced and increased the expression of FC1 in buds, respectively, implying that FC1 might be a common target for the CK and SL pathways. Nitrogen and auxin delayed expression change patterns of FC1, potentially by changing the downstream signals for CK and SL.

Differential regulation of proteins in rice (Oryza sativa L.) under iron deficiency.

KEY MESSAGE: Sixty-three proteins were identified to be differentially accumulated due to iron deficiency in shoot and root. The importance of these proteins alterations on shoot physiology is discussed. Iron (Fe) is an essential micronutrient for plant growth and its accumulation affects the quality of edible plant organs. To investigate the adaptive mechanism of a Chinese rice variety grown under iron deficiency, proteins differentially accumulated in leaves and roots of Yangdao 6, an indica cultivar, under Fe deficiency growth condition, were profiled using a two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/MS). The accumulations of seventy-three proteins were detected to be increased or decreased upon iron deficiency, and sixty-three of them were successfully identified. Among the sixty-three proteins, a total of forty proteins were identified in rice leaves, and twenty-three proteins were in roots. Most of these proteins are involved in photosynthesis, C metabolism, oxidative stress, Adenosine triphosphate synthesis, cell growth or signal transduction. The results provide a comprehensive way to understand, at the level of proteins, the adaptive mechanism used by rice shoots and roots under iron deficiency.

Transcriptomic and Metabolomic Analyses Reveals That Exogenous Methyl Jasmonate Regulates Galanthamine Biosynthesis in Lycoris longituba Seedlings.

The Amaryllidaceae alkaloid galanthamine (Gal) in Lycoris longituba is a secondary metabolite that has been used to treat Alzheimer's disease. Plant secondary metabolism is affected by methyl jasmonate (MeJA) exposure, although the regulatory mechanisms of MeJA on L. longituba seedlings remains largely unknown. In the present study, 75, 150, and 300 µM MeJA were used as treatments on L. longituba seedlings for 7, 14, 21, and 28 days, while 0 µM MeJA was used as the control (MJ-0). The effect of exogenous MeJA on Gal synthesis in L. longituba was then investigated using transcriptomic sequencing and metabolite profiling via GC-MS and LC-MS analysis. Galanthamine (Gal), lycorine (Lyc), and lycoramine (Lycm) abundances were 2. 71-, 2. 01-, and 2.85-fold higher in 75 µM MeJA (MJ-75) treatment plants compared to MJ-0 treatment plants after 7 days of cultivation. Transcriptomic analysis further showed that MJ-75 treatment significantly induced the expression of norbelladine synthase (NBS) and norbelladine 4'-O-methyltransferase (OMT), which are involved in the Gal biosynthesis pathway. In addition, increased expression was observed in MJ-75 treatment plants for genes in the JA synthesis and JA signaling pathways including those of allene oxide cyclase (AOC), 12-oxo-phytodienoic acid reductase (OPR), jasmonic acid amino acid synthase (JAR), and transcription factor MYC. The L. longituba tyrosine decarboxylase (LlTYDC) enzyme was identified and proposed to be involved in the Gal biosynthetic pathway. Metabolomics results demonstrated that the accumulation of Amaryllidaceae alkaloids, and especially alkaloids in the Gal biosynthesis pathway, could be induced by MJ-75 treatment. Interestingly, metabolites in the JA synthesis pathway were also affected by MeJA treatment. Overall, this multi-omics study suggests that both the JA synthesis/JA signaling and Gal biosynthesis pathways were affected by exogenous MeJA treatment. This comprehensive study of gene expression and metabolite contents can help us better understand the molecular mechanisms underlying MeJA-mediated Gal biosynthesis in L. longituba.

Transcriptome Analysis of Different Tissues Reveals Key Genes Associated With Galanthamine Biosynthesis in Lycoris longituba.

L ycoris longituba is a traditional medicinal plant containing the bioactive compound galanthamine (Gal), a type of Amaryllidaceae alkaloid and can be used to treat Alzheimer's disease. However, research on its genome or transcriptome and associated genes in the biosynthetic pathway is incomplete. In this study, we estimated the nuclear genome size of this species to be 29.33 Gb by flow cytometry. Then, RNA sequencing of the leaves, roots, and bulbs of L. longituba was carried out. After de novo assembly, 474,589 all-transcripts and 333,440 all-unigenes were finally generated. In addition, the differentially expressed genes (DEGs) were identified, and genes involved in the galanthamine metabolic pathway encoding tyrosine decarboxylase (TYDC), phenylalanine ammonia-lyase (PAL), cinnamate 4-hydroxylase (C4H), p-coumarate 3-hydroxylase (C3H), norbelladine synthase (NBS), norbelladine 4'-O-methyltransferase (OMT), and noroxomaritidine synthase (CYP96T1) were detected and validated by real-time quantitative PCR analysis. One candidate gene, Lycoris longituba O-Methyltransferase (LlOMT), was identified in the proposed galanthamine biosynthetic pathway. Sequence analysis showed that LlOMT is a class I OMT. LlOMT is localized in the cytoplasm, and biochemical analysis indicated that the recombinant LlOMT catalyzes norbelladine to generate 4'-O-methylnorbelladine. The protoplast transformation result showed that the overexpression of LlOMT could increase the Gal content. Our results indicate that LlOMT may play a role in galanthamine biosynthesis in L. longituba. This work provides a useful resource for the metabolic engineering of Amaryllidaceae alkaloids.

Effect of Exogenous Gibberellin, Paclobutrazol, Abscisic Acid, and Ethrel Application on Bulblet Development in Lycoris radiata.

Lycoris species have great ornamental and medicinal values; however, their low regeneration efficiency significantly restricts their commercial production. Exogenous hormone application is an effective way to promote bulblet development, but their effect on Lycoris radiata has not been verified to date. In the present study, we examined the effect of different exogenous hormones on bulblet development in L. radiata, and found that gibberellic acid (GA) significantly inhibited, whereas paclobutrazol (PBZ), abscisic acid (ABA), and ethrel promoted bulblet development, especially PBZ, a GA biosynthesis inhibitor. Furthermore, GA reduced endogenous cytokinin (CK) content, as well as the activities of carbohydrate metabolism enzymes, including sucrose synthase (SUS) and glucose-1-phosphate adenylyltransferase (AGPase), by downregulating the expression levels of LrSUS1, LrSUS2, and genes encoding AGPase large and small subunits. This resulted in the decrease in carbohydrate accumulation in the bulblets, thus hindering their development. PBZ had the opposite effect to GA on carbohydrate metabolism; it decreased endogenous GA15 and GA24, thereby promoting bulblet development. ABA promoted endogenous auxin content and the activities of starch synthesis enzymes, especially soluble starch synthase (SSS) and granule-bound SS (GBSS), through the up-regulation of the expression levels of LrSS1, LrSS2, and LrGBSS1 genes, which could also result in the accumulation of carbohydrates in the bulblets and promote their development. In addition, ethrel application partly promoted bulblet development by promoting endogenous CK content. Although the accumulation of carbohydrates and the activity of starch enzymes were increased by ethrel treatment, we hypothesized that the effect of ethrel on regulating carbohydrate metabolism may be indirect. Our results could provide a basis for improving the propagation efficiency of L. radiata for production, as well as propose some directions for future research.

[Prognostic value of microRNA-122 combined with APACHE II score in patient with acute respiratory distress syndrome].

OBJECTIVE: To investigate the prognostic value of microRNA-122 (miR-122) combined with acute physiology and chronic health evaluation II (APACHE II) score in patient with acute respiratory distress syndrome (ARDS), and to provide evidence for the diagnosis and treatment of ARDS. METHODS: ARDS patients admitted to the Third People's Hospital of Haikou City from January 2016 to December 2018 were enrolled. The general data, serum miR-122 expression level and APACHE II score within 24 hours were collected. The patients were divided into survival group and death group according to the survival status of ARDS patients. ARDS patients were divided into low-risk group (< 10 scores), medium-risk group (10-20 scores) and high-risk group (> 20 scores) according to APACHE II score. Predictive values of miR-122 and APACHE II scores on prognosis in ARDS patients were evaluated by the receiver operating characteristic (ROC) curve. The correlation between the serum miR-122 expression and APACHE II score in patients with ARDS was calculated by Pearson correlation analysis. RESULTS: A total of 142 ARDS patients were selected, 94 male and 48 female; with age (56.80±11.30) years old; 55 deaths and 87 survivors; 67 of high-risk, 48 of medium-risk and 27 of low-risk. The expression of serum miR-122 and APACHE II score in the death group were significantly higher than those in the survival group [miR-122 (2-ΔΔCt): 0.26±0.12 vs. 0.07±0.03, APACHE II: 31.84±4.25 vs. 15.30±2.60, both P < 0.01]. With the severity increase of the disease, the serum miR-122 expression level, APACHE II score, and mortality rate of ARDS patients gradually elevated, and the difference between the two groups was significant in the low-risk group, medium-risk group, and high-risk group [miR-122 (2-ΔΔCt): 0.05±0.02, 0.14±0.06, 0.23±0.09; APACHE II: 12.30±2.15, 20.62±3.40, 28.90±3.60; mortality rate: 11.1%, 31.2%, 55.2%, respectively, all P < 0.05]. ROC curve analysis showed that miR-122 and APACHE II score could predict the death of ARDS patients, and the area under the ROC curve (AUC) was 0.835 [95% confidence interval (95%CI) = 0.776-0.893] and 0.790 (95%CI = 0.732-0.854); the predicted value of the miR-122 combined with APACHE II score (AUC = 0.918, 95%CI = 0.857-0.972) was higher than the single miR-122 and APACHE II score (both P < 0.05), with sensitivity and specificity were 91.3% and 86.4% respectively. The correlation analysis showed that the expression of serum miR-122 was positively correlated with APACHE II score in death patient with ARDS (r = 0.825, P < 0.01). CONCLUSIONS: Elevated serum miR-122 expression level is associated with disease severity and prognosis of ARDS patients; miR-122 combination with APACHE II score has a high evaluation value on prognosis of ARDS patients.

Variants in multiple genes polymorphism association analysis of COPD in the Chinese Li population.

BACKGROUND: It is known that the contribution of risk alleles to chronic obstructive pulmonary disease (COPD) may vary between populations. Further, previous studies involving various ethnic groups have revealed associations between COPD and genetic polymorphisms in families with sequence similarity 13, member A (FAM13A), micro-RNA 2054 (MIR2054), SET domain containing protein 7 (SETD7), ring finger protein 150 (RNF150), hedgehog interacting protein (HHIP), and vascular endothelial growth factor A (VEGFA). Our objective was to explore the association between these gene polymorphism and COPD in members of Chinese Li minority population. MATERIALS AND METHODS: The Chinese Li population case-control study was conducted to assess genetic associations with COPD risk. Seven single nucleotide polymorphisms (SNPs) located on chromosome 4, including FAM13A, MIR2054, SETD7, RNF150, and HHIP, and nine SNPs in the VEGFA gene were genotyped among 234 cases and 240 controls using Sequenom Mass-ARRAY(®) platform. Linkage disequilibrium (LD) analysis was performed using Haploview software and the associations of the SNP frequencies with COPD were analyzed using chi-square (χ(2)) tests, genetic models analysis, and haplotype analysis. RESULTS: By χ(2) we found the minor allele "G" of rs17050782 was with increased COPD risk in allele model. In genetic models, we found the minor allele of rs7671167 (P=0.028 by dominant model) and rs17050782 (P=0.008 by recessive model) was associated with the increased risk of COPD disease. Likewise, an increased risk of developing COPD was associated with the "GGCGC" haplotype of VEGFA (odds ratio =1.48, 95% confidence interval =1.02-2.12, P=0.037). CONCLUSION: Our results were the first time to reveal that SNPs from FAM13A (rs7671167), SETD7 (rs17050782), and a haplotype of VEGFA ("GGCGC") are potential susceptibility loci associated with increased COPD risk in Chinese Li minority population.

The analyses of risk factors for COPD in the Li ethnic group in Hainan, People's Republic of China.

OBJECTIVE: To study the risk factors for chronic obstructive pulmonary disease (COPD) in Li population in Hainan province, People's Republic of China. METHODS: Li people above 40 years of age from Hainan were chosen by stratified random cluster sampling between 2012 and 2014. All participants were interviewed with a home-visiting questionnaire, and spirometry was performed on all eligible participants. Patients with airflow limitation (forced expiratory volume in 1 second [FEV1]/forced vital capacity [FVC] <0.70) were further examined by postbronchodilator spirometry, and those with a postbronchodilator FEV1/FVC <0.70 was diagnosed with COPD. The information of physical condition and history, smoking intensity, smoking duration, second-hand smoking, education, job category, monthly household income, working years, residential environment, primary fuel for cooking and heating (biomass fuel including wood, crop residues, dung, and charcoal, or modern fuel such as natural gas, liquefied petroleum gas, electricity, and solar energy), ventilated kitchen, heating methods, air pollution, recurrent respiratory infections, family history of respiratory diseases, cough incentives, and allergies of COPD and non-COPD subjects was analyzed by univariate and multivariate logistic regression models to identify correlated risk factors for COPD. RESULTS: Out of the 5,463 Li participants, a total of 277 COPD cases were identified by spirometry, and 307 healthy subjects were randomly selected as controls. Univariate logistic regression analyses showed that older people (65 years and above), low body mass index (BMI), biomass smoke, 11-20 and >20 cigarettes/day, smoking for 40 years or more, second-hand smoking, recurrent respiratory infections, and induced cough were risk factors for COPD, whereas high BMI, high education level, and presence of ventilated kitchen were protective factors. Subsequent multivariate logistic regression model further demonstrated that aging, low BMI, biomass smoke, >20 cigarettes/day, and recurrent respiratory tract infections were high-risk factors for COPD in the Li population. CONCLUSION: The incidence of COPD has a strong correlation with age, BMI, biomass smoke, >20 cigarettes/day, and recurrent respiratory infections, suggesting they were high-risk factors for COPD in Li population.