Integrative analysis of the metabolome and transcriptome reveals the mechanism of polyphenol biosynthesis in Taraxacum mongolicum.

Introduction: Dandelion is widely used in clinical practice due to its beneficial effects. Polyphenolic compounds are considered the main anti-inflammatory active ingredient of dandelion, but the gene expression patterns of polyphenolic compounds in different dandelion tissues are still unclear. Methods: In this study, we combined a nontargeted metabolome, PacBio Iso-seq transcriptome, and Illumina RNA-seq transcriptome to investigate the relationship between polyphenols and gene expression in roots, flowers, and leaves of flowering dandelion plants. Results: Eighty-eight flavonoids and twenty-five phenolic acids were identified, and 64 candidate genes involved in flavonoid biosynthesis and 63 candidate genes involved in chicoric acid biosynthesis were identified. Most flavonoid and chicoric acid-related genes demonstrated the highest content in flowers. RNA-seq analysis revealed that genes involved in polyphenol biosynthesis pathways, such as CHS, CHI, F3H, F3'H, FLS, HQT, and CAS, which are crucial for the accumulation of flavonoids and chicoric acid, were upregulated in flowers. Discussion: The combination of transcriptomic and metabolomic data can help us better understand the biosynthetic pathways of polyphenols in dandelion. These results provide abundant genetic resources for further studying the regulatory mechanism of dandelion polyphenol biosynthesis.

SlTrxh functions downstream of SlMYB86 and positively regulates nitrate stress tolerance via S-nitrosation in tomato seedling.

Nitric oxide (NO) is a redox-dependent signaling molecule that plays a crucial role in regulating a wide range of biological processes in plants. It functions by post-translationally modifying proteins, primarily through S-nitrosation. Thioredoxin (Trx), a small and ubiquitous protein with multifunctional properties, plays a pivotal role in the antioxidant defense system. However, the regulatory mechanism governing the response of tomato Trxh (SlTrxh) to excessive nitrate stress remains unknown. In this study, overexpression or silencing of SlTrxh in tomato led to increased or decreased nitrate stress tolerance, respectively. The overexpression of SlTrxh resulted in a reduction in levels of reactive oxygen species (ROS) and an increase in S-nitrosothiol (SNO) contents; conversely, silencing SlTrxh exhibited the opposite trend. The level of S-nitrosated SlTrxh was increased and decreased in SlTrxh overexpression and RNAi plants after nitrate treatment, respectively. SlTrxh was found to be susceptible to S-nitrosation both in vivo and in vitro, with Cysteine 54 potentially being the key site for S-nitrosation. Protein interaction assays revealed that SlTrxh physically interacts with SlGrx9, and this interaction is strengthened by S-nitrosation. Moreover, a combination of yeast one-hybrid (Y1H), electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR), and transient expression assays confirmed the direct binding of SlMYB86 to the SlTrxh promoter, thereby enhancing its expression. SlMYB86 is located in the nucleus and SlMYB86 overexpressed and knockout tomato lines showed enhanced and decreased nitrate stress tolerance, respectively. Our findings indicate that SlTrxh functions downstream of SlMYB86 and highlight the potential significance of S-nitrosation of SlTrxh in modulating its function under nitrate stress.

Identification of autophagy-related signatures in doxorubicin-induced cardiotoxicity.

PURPOSE: Doxorubicin is an antibiotic drug used clinically to treat infectious diseases and tumors. Unfortunately, it is cardiotoxic. Autophagy is a cellular self-decomposition process that is essential for maintaining homeostasis in the internal environment. Accordingly, the present study was proposed to characterize the autophagy-related signatures of doxorubicin-induced cardiotoxicity. METHODS: Datasets related to doxorubicin-induced cardiotoxicity were retrieved by searching the GEO database and differentially expressed genes (DEGs) were identified. DEGs were taken to intersect with autophagy-related genes to obtain autophagy-related signatures, and Gene Ontology (GO)/Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis and protein-protein interaction (PPI) network were performed on them. Further, construction of miRNA-hub gene networks and identification of target drugs to reveal potential molecular mechanisms and therapeutic strategies. Animal models of doxorubicin-induced cardiotoxicity were constructed to validate differences in gene expression for autophagy-related signatures. RESULTS: PBMC and heart samples from the GSE37260 dataset were selected for analysis. There were 995 and 2357 DEGs in PBMC and heart samples, respectively, and they had 23 intersecting genes with autophagy-related genes. RT-qPCR confirmed the differential expression of 23 intersecting genes in doxorubicin-induced cardiotoxicity animal models in general agreement with the bioinformatics results. An autophagy-related signatures consisting of 23 intersecting genes is involved in mediating processes and pathways such as autophagy, oxidative stress, apoptosis, protein ubiquitination and phosphorylation. Moreover, Akt1, Hif1a and Mapk3 are hub genes in autophagy-associated signatures and their upstream miRNAs are mainly rno-miR-1188-5p, rno-miR-150-3p and rno-miR-326-3p, and their drugs are mainly CHEMBL55802, Carboxyamidotriazole and 3-methyladenine. CONCLUSION: This study identifies for the first-time autophagy-related signatures in doxorubicin's cardiotoxicity, which could provide potential molecular mechanisms and therapeutic strategies for doxorubicin-induced cardiotoxicity.

Identification of a novel marker and its associated laccase gene for regulating ear length in tropical and subtropical maize lines.

KEY MESSAGE: This study revealed the identification of a novel gene, Zm00001d042906, that regulates maize ear length by modulating lignin synthesis and reported a molecular marker for selecting maize lines with elongated ears. Maize ear length has garnered considerable attention due to its high correlation with yield. In this study, six maize inbred lines of significant importance in maize breeding were used as parents. The temperate maize inbred line Ye107, characterized by a short ear, was crossed with five tropical or subtropical inbred lines featuring longer ears, creating a multi-parent population displaying significant variations in ear length. Through genome-wide association studies and mutation analysis, the A/G variation at SNP_183573532 on chromosome 3 was identified as an effective site for discriminating long-ear maize. Furthermore, the associated gene Zm00001d042906 was found to correlate with maize ear length. Zm00001d042906 was functionally annotated as a laccase (Lac4), which showed activity and influenced lignin synthesis in the midsection cells of the cob, thereby regulating maize ear length. This study further reports a novel molecular marker and a new gene that can assist maize breeding programs in selecting varieties with elongated ears.

Rho2 involved in development, stress response and pathogenicity of Fusarium oxysporum.

Rho GTPases regulate the activity of cell wall biosynthesis, actin assembly and polar cell secretion. However, the function of Rho GTPase in filamentous fungi is poorly understood. To understand the role of Rho2 GTPase in Fusarium oxysporum, which is one of root rot pathogens of Panax notoginseng, △rho2 mutant was constructed. Phenotypes of △rho2, including conidiation, germination of spores, stresses (osmotic-, cell membrane-, cell wall disturbing-, metal-, and high temperature-) tolerance and pathogenicity were analyzed. The results showed that the growth of △rho2 was destroyed under cell wall disturbing stress and high temperature stress, suggesting that Rho2 regulated the response of F. oxysporum to cell wall synthesis inhibitors and high temperature stress. Germination of spores and pathogenicity to P. notoginseng were reduced in △rho2 mutant. Western blot results showed that rho2 deletion increased the phosphorylation level of Mpk1. To identify genes regulated by Rho2, transcriptome sequencing was carried out. 2477 genes were identified as upregulated genes and 2177 genes were identified as downregulated genes after rho2 was deleted. These genes provide clues for further study of rho2 function.

14-3-3 proteins regulate the HCHO stress response by interacting with AtMDH1 and AtGS1 in tobacco and Arabidopsis.

Formaldehyde (HCHO) is one of the most essential common carcinogenic environmental pollutants. While 14-3-3 proteins are known to regulate the response of plants to HCHO stress, the regulatory mechanisms responsible for a tolerant phenotype remain unclear. We first performed qPCR analysis of HCHO-treated Arabidopsis and tobacco and determined that the expression of At14-3-3PSI and Nt14-3-3C genes was rapidly upregulated after HCHO stress. Furthermore, overexpression of 14-3-3, AtMDH1 or AtGS1 genes enhanced plant HCHO absorption capacity and resistance, and knockdown or knockout of 14-3-3, AtMDH1 or AtGS1 genes reduced plant HCHO absorption capacity and resistance. However, overexpression of the AtGS1 and AtMDH1 genes in the At14-3-3 psi mutant restored HCHO uptake and resistance in Arabidopsis. Moreover, 14-3-3 bound to the N-terminus of AtMDH1 and the C-terminus of AtGS1, respectively, and repressed and enhanced their expression. The 13C NMR results of HCHO stress mutants Atgs1 and Atmdh1 showed that the metabolites Glu and Asp rapidly increased, indicating that AtGS1 and AtMDH1 were indeed indispensable for Arabidopsis to metabolize HCHO. In conclusion, we uncovered a HCHO stress response mechanism mediated by 14-3-3, which enhances the plant's ability to absorb HCHO, deepening our understanding of how plants respond to HCHO stress.

Whole genome sequencing and analysis of Armillaria gallica Jzi34 symbiotic with Gastrodia elata.

BACKGROUND: Armillaria species are plant pathogens, but a few Armillaria species can establish a symbiotic relationship with Gastrodia elata, a rootless and leafless orchid, that is used as a Chinese herbal medicine. Armillaria is a nutrient source for the growth of G. elata. However, there are few reports on the molecular mechanism of symbiosis between Armillaria species and G. elata. The genome sequencing and analysis of Armillaria symbiotic with G. elata would provide genomic information for further studying the molecular mechanism of symbiosis. RESULTS: The de novo genome assembly was performed with the PacBio Sequel platform and Illumina NovaSeq PE150 for the A. gallica Jzi34 strain, which was symbiotic with G. elata. Its genome assembly contained ~ 79.9 Mbp and consisted of 60 contigs with an N50 of 2,535,910 bp. There were only 4.1% repetitive sequences in the genome assembly. Functional annotation analysis revealed a total of 16,280 protein coding genes. Compared with the other five genomes of Armillaria, the carbohydrate enzyme gene family of the genome was significantly contracted, while it had the largest set of glycosyl transferase (GT) genes. It also had an expansion of auxiliary activity enzymes AA3-2 gene subfamily and cytochrome P450 genes. The synteny analysis result of P450 genes reveals that the evolutionary relationship of P450 proteins between A. gallica Jzi34 and other four Armillaria was complex. CONCLUSIONS: These characteristics may be beneficial for establishing a symbiotic relationship with G. elata. These results explore the characteristics of A. gallica Jzi34 from a genomic perspective and provide an important genomic resource for further detailed study of Armillaria. This will help to further study the symbiotic mechanism between A. gallica and G. elata.

Resected lymph nodes and survival of patients with esophageal squamous cell carcinoma: an observational study.

BACKGROUND: The incidence and mortality of esophageal cancer are high. Therefore, the authors aimed to investigate how the number of dissected lymph nodes (LNs) during esophagectomy for esophageal squamous cell carcinoma impacts overall survival (OS), particularly that of patients with positive LNs. MATERIALS AND METHODS: Data from 2010 to 2017 were obtained from the Sichuan Cancer Hospital and Institute Esophageal Cancer Case Management Database. Participants were divided into two groups: patients with negative lymph nodes (N0) and patients with positive lymph nodes (N+). The median number of resected LNs during surgery was 24; therefore, patients with 15-23 and those with 24 or more resected LNs were assigned to subgroups A and B, respectively. RESULTS: After a median follow-up of 60.33 months, 1624 patients who underwent esophagectomy were evaluated; 60.53 and 39.47% had a pathological diagnosis of N+ or N0, respectively. The median OS was 33.9 months for the N+ group; however, the N0 group did not achieve the median OS. The mean OS was 84.9 months. In the N+ group, the median OS times of subgroups A and B were 31.2 and 37.1 months, respectively. The OS rates at 1, 3, and 5 years were 82, 43, and 34%, respectively, for subgroup A of the N+ group; they were 86, 51, and 38%, respectively, for subgroup B of the N+ group. Subgroups A and B of the N0 group exhibited no statistically significant differences. CONCLUSION: Increasing the number of LNs harvested during surgery to 24 or more could improve the OS of patients with positive LNs but not that of patients with negative LNs.

Mapping of Lymph Node Metastasis and Efficacy Index in Thoracic Esophageal Squamous Cell Carcinoma: A Large-Scale Retrospective Analysis.

BACKGROUND: Esophageal squamous cell carcinoma has a high mortality rate in China. The metastatic pattern in the lymph nodes and the value of their dissection on the overall survival of these patients remain controversial. The primary aim of this study was to provide a basis for accurate staging of esophageal cancer and to identify the relationship between esophageal cancer surgery, lymph node dissection, and overall survival rates. METHODS: We utilized our hospital database to retrospectively review the data of 1727 patients with esophageal cancer who underwent R0 esophagectomy from January 2010 to December 2017. The lymph nodes were defined according to Japanese Classification of Esophageal Cancer, 11th Edition. The Efficacy Index (EI) was calculated by multiplying the frequency (%) of metastases to a zone and the 5-year survival rate (%) of patients with metastases to that zone, and then dividing by 100. RESULTS: The EI was high in the supraclavicular and mediastinal zones in patients with upper esophageal tumors, and the EI of 101R was 17.39, which was the highest among the lymph node stations. In patients with middle esophageal tumors, the EI was highest in the mediastinal zone, followed by the celiac and supraclavicular zones. Furthermore, the EI was highest in the celiac zone, followed by the mediastinal zones in patients with lower esophageal tumors. CONCLUSIONS: The EI of resected lymph nodes was found to vary between stations and was related to the primary location of the tumor.

Impact of two­field or three­field lymphadenectomy on overall survival in middle and lower thoracic esophageal squamous cell carcinoma: A single­center retrospective analysis.

Squamous cell carcinoma is the main subtype of esophageal cancer in East Asia. The effect of the number of lymph nodes (LNs) removed to treat middle and lower thoracic esophageal squamous cell carcinoma (ESCC) in China remains controversial. Therefore, the present study aimed to investigate the impact of the number of LNs removed during lymphadenectomy on the survival of patients with middle and lower thoracic ESCC. Data were obtained from the Sichuan Cancer Hospital and Institute Esophageal Cancer Case Management Database from January 2010 to April 2020. Either three-field systematic lymphadenectomy (3F group) or two-field systematic lymphadenectomy (2F group) was performed for ESCC cases with or without suspicious tumor-positive cervical LNs, respectively. Subgroups were designed for further analysis based on the quartile number of resected LNs. After 50.7 months of median follow-up, 1,659 patients who underwent esophagectomy were enrolled. The median overall survival (OS) of the 2F and 3F groups was 50.0 months and 58.5 months, respectively. The OS rates at 1, 3 and 5 years were 86, 57 and 47%, respectively, in the 2F group, and 83, 52 and 47%, respectively, in the 3F group (P=0.732). The average OS of the 3F B and D groups was 57.7 months and 30.2 months, respectively (P=0.006). In the 2F group, the OS between subgroups was not significantly different. In conclusion, resection of >15 LNs during two-field dissection in patients with ESCC undergoing esophagectomy did not affect their survival outcomes. In three-field lymphadenectomy, the extent of LNs removed could lead to different survival outcomes.

Overexpression of tomato glutathione reductase (SlGR) in transgenic tobacco enhances salt tolerance involving the S-nitrosylation of GR.

S-nitrosylation, a post-translational modification (PTM) dependent on nitric oxide, is essential for plant development and environmental responsiveness. However, the function of S-nitrosylation of glutathione reductase (GR) in tomato (SlGR) under NaCl stress is yet uncertain. In this study, sodium nitroprusside (SNP), an exogenous NO donor, alleviated the growth inhibition of tomato under NaCl treatment, particularly at 100 µM. Following NaCl treatment, the transcripts, enzyme activity, and S-nitrosylated level of GR were increased. In vitro, the SlGR protein was able to be S-nitrosylated by S-nitrosoglutathione (GSNO), significantly increasing the activity of GR. SlGR overexpression transgenic tobacco plants exhibited enhanced germination rate, fresh weight, and increased root length in comparison to wild-type (WT) seedlings. The accumulation of reactive oxygen species (ROS) was lower, whereas the expression and activities of GR, ascorbate peroxidase (APX), superoxide dismutase (SOD), and catalase (CAT); the ratio of ascorbic acid/dehydroascorbic acid (AsA/DHA), reduced glutathione/oxidized glutathione (GSH/GSSG), total soluble sugar and proline contents; and the expression of stress-related genes were higher in SlGR overexpression transgenic plants in comparison to the WT plants following NaCl treatment. The accumulation of NO and S-nitrosylated levels of GR in transgenic plants was higher in comparison to WT plants following NaCl treatment. These results indicated that S-nitrosylation of GR played a significant role in salt tolerance by regulating the oxidative state.

Optimal discharge planning for esophagectomy patients with enhanced recovery after surgery: Recommendations.

Background: Studies have suggested that the postoperative length of stay (PLOS) of esophagectomy patients under the enhanced recovery after surgery (ERAS) pathway should be >10 days as against the previously recommended 7 days. We investigated the distribution and influencing factors of PLOS in the ERAS pathway in order to recommend an optimal planned discharge time. Methods: This was a single-center retrospective study of 449 patients with thoracic esophageal carcinoma who underwent esophagectomy and perioperative ERAS between January 2013 and April 2021. We established a database to prospectively document the causes of delayed discharge. Results: The mean and median PLOS were 10.2 days and 8.0 days (range: 5-97), respectively. Patients were divided into four groups: group A (PLOS ≤ 7 days), 179 patients (39.9%); group B (8 ≤ PLOS ≤ 10 days), 152 (33.9%); group C (11 ≤ PLOS ≤ 14 days), 68 (15.1%); group D (PLOS > 14 days), 50 patients (11.1%). The main cause of prolonged PLOS in group B was minor complications (prolonged chest drainage, pulmonary infection, recurrent laryngeal nerve injury). Severely prolonged PLOS in groups C and D were due to major complications and comorbidities. On multivariable logistic regression analysis, open surgery, surgical duration >240 min, age >64 years, surgical complication grade >2, and critical comorbidities were identified as risk factors for delayed discharge. Conclusions: The optimal planned discharge time for patients undergoing esophagectomy with ERAS should be 7-10 days with a 4-day discharge observation window. Patients at risk of delayed discharge should be managed adopting PLOS prediction.

Enhancement production of lipid and unsaturation of fatty acids in Cryptococcus humicola via addition of calcium ion.

Lipids synthesized by oleaginous yeasts are considered to be the best candidates for biodiesel production. Cryptococcus humicola as an oleaginous yeast accumulated lipid in cells. In order to optimize the conditions for lipid production, different carbon and nitrogen sources were used and metals were added into the medium. Ca2+ addition increased the lipid production greatly. Xylose and peptone were optimal carbon source and nitrogen source, respectively for lipid accumulation. Response surface experiment results revealed that the accumulation of lipid could be maximized when the xylose, peptone and Ca2+ concentration was 61 g/L, 4.31 g/L, 0.67 mmol/L. C16 and C18 fatty acid account for about 91% of the total fatty acids. The most abundant fatty acid was oleic acid (42.68%), followed by palmitic acid (29.7%) and stearic acid (13.87%). The addition of Ca2+ increased the content of unsaturated fatty acids (such as C16:1 and C18:1) and improved the unsaturation of fatty acids. Quantitative real time PCR analysis revealed that expression of genes related to lipid biosynthesis showed up-regulated by Ca2+ treatment. This study provided a strategy for increase in lipid production and content of unsaturated fatty acids.

Transcriptome analysis reveals the regulatory mode by which NAA promotes the growth of Armillaria gallica.

A symbiotic relationship is observed between Armillaria and the Chinese herbal medicine Gastrodia elata (G. elata). Armillaria is a nutrient source for the growth of G. elata, and its nutrient metabolism efficiency affects the growth and development of G. elata. Auxin has been reported to stimulate Armillaria species, but the molecular mechanism remains unknown. We found that naphthalene acetic acid (NAA) can also promote the growth of A. gallica. Moreover, we identified a total of 2071 differentially expressed genes (DEGs) by analyzing the transcriptome sequencing data of A. gallica at 5 and 10 hour of NAA treatment. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that these unigenes were significantly enriched in the metabolism pathways of arginine, proline, propanoate, phenylalanine and tryptophan. The expression levels of the general amino acid permease (Gap), ammonium transporter (AMT), glutamate dehydrogenase (GDH), glutamine synthetase (GS), Zn(II) 2Cys6 and C2H2 transcription factor genes were upregulated. Our transcriptome analysis showed that the amino acid and nitrogen metabolism pathways in Armillaria were rapidly induced within hours after NAA treatment. These results provide valuable insights into the molecular mechanisms by which NAA promotes the growth of Armillaria species.

Pseudomonas fluorescens BsEB-1: an endophytic bacterium isolated from the root of Bletilla striata that can promote its growth.

An endophytic Pseudomonas fluorescens (BsEB-1) was obtained from the roots of Bletilla striata. We investigated its growth-promoting properties and observed the impact of its inoculation on both the growth and polysaccharide content of Bletilla striata tubers. It was found that BsEB-1 possessed three growth-promoting activities: phosphate-solubilizing, produced indoleacetic acid (IAA) and siderophores, but had no nitrogen-fixing activity. BsEB-1 could rapidly attach to the root hairs of Bletilla striata tissue culture seedlings and endophytically colonize the region of maturation in the roots. It also significantly promoted the rooting and transplant survival rate of the seedlings, as well as the growth and expansion of the tubers, but did not increase their polysaccharide content. Pseudomonas fluorescens BsEB-1 exhibits potential for applications in the artificial planting of Bletilla striata.

Overexpression of tomato thioredoxin h (SlTrxh) enhances excess nitrate stress tolerance in transgenic tobacco interacting with SlPrx protein.

The thioredoxin (Trx) system plays a vital function in cellular antioxidative defense. However, little is known about Trx in tomato under excess nitrate. In this study, we isolated the tomato gene encoding h-type Trx gene (SlTrxh). The mRNA transcript of SlTrxh in roots and leaves of tomato was induced incrementally under excess nitrate for 24 h. Subcellular localization showed that SlTrxh might localize in the cytoplasm, nucleus and plasma membrane. Enzymatic activity characterization revealed that SlTrxh protein possesses the disulfide reductase function and Cysteine (Cys) 54 is important for its activity. Overexpressing SlTrxh in tobacco resulted in increasing seed germination rate, root length and decreasing H2O2 and O2- accumulation, compared with the wild type (WT) tobacco under nitrate stress. While overexpressing SlTrxhC54S (Cysteine 54 mutated to Serine) in tobacco showed decreased germination rate and root length compared with the WT after nitrate treatment. After nitrate stress treatment, SlTrxh overexpressing transgenic tobacco plants have lower malonaldehyde (MDA), H2O2 contents and Reactive Oxygen Species (ROS) accumulation, and higher mRNA transcript level of NtP5CS, NtDREB2, higher ratio of ASA/DHA and GSH/GSSG, higher activities of ascorbate peroxidase and NADP thioredoxin reductase. Besides, SlTrxh overexpressing plants showed higher tolerance to Methyl Viologen (MV) in the seed germination and seedling stage. The yeast two-hybrid, pull-down, Co-immunoprecipitation and Bimolecular luciferase complementation assay confirmed that SlTrxh physically interacted with tomato peroxiredoxin (SlPrx). These results suggest that SlTrxh contributes to maintaining ROS homeostasis under excess nitrate stress interacting with SlPrx and Cys54 is important for its enzyme activity.

The interaction of MYB, bHLH and WD40 transcription factors in red pear (Pyrus pyrifolia) peel.

KEY MESSAGE: Sunlight enhanced peel color and significantly up-regulated the expression of PyMYB10 and PybHLH genes. MYB-bHLH-WD40 transcriptional complex forms in the light and is involved in regulating anthocyanin accumulation in the peel. Anthocyanin is the major pigment in the peel of Yunnan red pear (Pyrus pyrifolia (Burm.) Nak.). A transcriptional activation protein complex, involving members of the transcription factor classes of MYB, bHLH and WD40, regulates anthocyanin biosynthesis. This complex was examined in the peel of red pear. In order to clarify the interaction of PyMYB10, PybHLH and PyWD40, fruit were bagged then peel samples collected 0, 3, 5, and 7 days after bag removal. Samples were used for Western blotting and protein interaction analysis. The results showed that sunlight enhanced peel color and significantly up-regulated the expression of both PyMYB10 and PybHLH genes. Co-immunoprecipitation (Co-IP) analysis showed that PybHLH interacted with PyMYB10 or PyWD40, and PyMYB10 interacted with PyWD40. Using onion cells as a model system, bimolecular fluorescence complementation (BiFC) confirmed these interactions and showed that the interaction localized to the nuclei. GST Pull down and Far-Western blotting assays demonstrated that PybHLH interacted with PyMYB10 or PyWD40, respectively, and PyMYB10 interacted with PyWD40 in vitro. In addition, EMSA assay showed that PyMYB10 can directly bind to the promoter of the gene encoding the anthocyanin biosynthesis enzyme anthocyanidin synthase (PyANS). Taken together, these results showed that the ternary complex of PyMYB10, PybHLH and PyWD40 transcription factors forms to regulate anthocyanin biosynthesis and accumulation in Yunnan red pear.

Co-overexpression of AtSHMT1 and AtFDH induces sugar synthesis and enhances the role of original pathways during formaldehyde metabolism in tobacco.

Serine hydroxymethyltransferase 1 (SHMT1) is a key enzyme in the photorespiration pathway in higher plants. Our previous study showed that AtSHMT1 controls the assimilation of HCHO to sugars in Arabidopsis. The expression of SHMT1 was induced in Arabidopsis but was inhibited in tobacco under HCHO stress. To investigate whether the function of AtSHMT1 in the HCHO assimilation could be exerted in tobacco, AtSHMT1 was overexpressed alone (S5) or co-overexpressed (SF6) with Arabidopsis formate dehydrogenase (AtFDH) in leaves using a light-inducible promoter in this study. 13C NMR analyses showed that the 13C-metabolic flux from H13CHO was introduced to sugar synthesis in SF6 leaves but not in S5 leaves. The increase in the production of metabolites via the original pathways was particularly greater in SF6 leaves than in S5 leaves, suggesting that co-overexpression of AtSHMT1 and AtFDH is more effective than overexpression of AtSHMT1 alone in the enhancement of HCHO metabolism in tobacco leaves. Consequently, the increase in HCHO uptake and resistance was greater in SF6 leaves than in S5 leaves. The mechanism underlying the role of overexpressed AtSHMT1 and AtFDH was discussed based on changes in photosynthetic parameters, chlorophyll content, antioxidant enzyme activity and the oxidative level in leaves.

De novo RNA sequencing and analysis reveal the putative genes involved in diterpenoid biosynthesis in Aconitum vilmorinianum roots.

In this study, the putative genes involved in diterpenoid alkaloids biosynthesis in A. vilmorinianum roots were revealed by transcriptome sequencing. 59.39 GB of clean bases and 119,660 unigenes were assembled, of which 69,978 unigenes (58.48%) were annotated. We identified 27 classes of genes (139 candidate genes) involved in the synthesis of diterpenoid alkaloids, including the mevalonate (MVA) pathway, the methylerythritol 4-phosphate (MEP) pathway, the farnesyl diphosphate regulatory pathway, and the diterpenoid scaffold synthetic pathway. 12 CYP450 genes were identified. We found that hydroxymethylglutaryl-CoA reductase was the key enzyme in MVA metabolism, which was regulated by miR6300. Transcription factors, such as bHLH, AP2/EREBP, and MYB, used to synthesize the diterpenes were analyzed. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02646-6.