High light triggers flavonoid and polysaccharide synthesis through DoHY5-dependent signaling in Dendrobium officinale.

Dendrobium officinale is edible and has medicinal and ornamental functions. Polysaccharides and flavonoids, including anthocyanins, are important components of D. officinale that largely determine the nutritional quality and consumer appeal. There is a need to study the molecular mechanisms regulating anthocyanin and polysaccharide biosynthesis to enhance D. officinale quality and its market value. Here, we report that high light (HL) induced the accumulation of polysaccharides, particularly mannose, as well as anthocyanin accumulation, resulting in red stems. Metabolome and transcriptome analyses revealed that most of the flavonoids showed large changes in abundance, and flavonoid and polysaccharide biosynthesis was significantly activated under HL treatment. Interestingly, DoHY5 expression was also highly induced. Biochemical analyses demonstrated that DoHY5 directly binds to the promoters of DoF3H1 (involved in anthocyanin biosynthesis), DoGMPP2, and DoPMT28 (involved in polysaccharide biosynthesis) to activate their expression, thereby promoting anthocyanin and polysaccharide accumulation in D. officinale stems. DoHY5 silencing decreased flavonoid- and polysaccharide-related gene expression and reduced anthocyanin and polysaccharide accumulation, whereas DoHY5 overexpression had the opposite effects. Notably, naturally occurring red-stemmed D. officinale plants similarly have high levels of anthocyanin and polysaccharide accumulation and biosynthesis gene expression. Our results reveal a previously undiscovered role of DoHY5 in co-regulating anthocyanin and polysaccharide biosynthesis under HL conditions, improving our understanding of the mechanisms regulating stem color and determining nutritional quality in D. officinale. Collectively, our results propose a robust and simple strategy for significantly increasing anthocyanin and polysaccharide levels and subsequently improving the nutritional quality of D. officinale.

DhuFAP: a platform for gene functional analysis in Dendrobium huoshanense.

BACKGROUND: Dendrobium huoshanense, a traditional medicinal and food plant, has a rich history of use. Recently, its genome was decoded, offering valuable insights into gene function. However, there is no comprehensive gene functional analysis platform for D. huoshanense. RESULT: To address this, we created a platform for gene function analysis and comparison in D. huoshanense (DhuFAP). Using 69 RNA-seq samples, we constructed a gene co-expression network and annotated D. huoshanense genes by aligning sequences with public protein databases. Our platform contained tools like Blast, gene set enrichment analysis, heatmap analysis, sequence extraction, and JBrowse. Analysis revealed co-expression of transcription factors (C2H2, GRAS, NAC) with genes encoding key enzymes in alkaloid biosynthesis. We also showcased the reliability and applicability of our platform using Chalcone synthases (CHS). CONCLUSION: DhuFAP ( www.gzybioinformatics.cn/DhuFAP ) and its suite of tools represent an accessible and invaluable resource for researchers, enabling the exploration of functional information pertaining to D. huoshanense genes. This platform stands poised to facilitate significant biological discoveries in this domain.

MIR396-GRF/GIF enhances in planta shoot regeneration of Dendrobium catenatum.

Recent studies on co-transformation of the growth regulator, TaGRF4-GIF1 chimera (Growth Regulating Factor 4-GRF Interacting Factor 1), in cultivated wheat varieties (Triticum aestivum), showed improved regeneration efficiency, marking a significant breakthrough. Here, a simple and reproducible protocol using the GRF4-GIF1 chimera was established and tested in the medicinal orchid Dendrobium catenatum, a monocot orchid species. TaGRF4-GIF1 from T. aestivum and DcGRF4-GIF1 from D. catenatum were reconstructed, with the chimeras significantly enhancing the regeneration efficiency of D. catenatum through in planta transformation. Further, mutating the microRNA396 (miR396) target sites in TaGRF4 and DcGRF4 improved regeneration efficiency. The target mimicry version of miR396 (MIM396) not only boosted shoot regeneration but also enhanced plant growth. Our methods revealed a powerful tool for the enhanced regeneration and genetic transformation of D. catenatum.

Physiological and transcriptomic comparisons shed light on the cold stress response mechanisms of Dendrobium spp.

BACKGROUND: Dendrobium spp. comprise a group of tropical orchids with ornamental and medicinal value. Dendrobium spp. are sensitive to low temperature, and the underlying cold response regulatory mechanisms in this group are unclear. To understand how these plants respond to cold stress, we compared the transcriptomic responses of the cold-tolerant cultivar 'Hongxing' (HX) and the cold-sensitive cultivar 'Sonia Hiasakul' (SH) to cold stress. RESULTS: Chemometric results showed that the physiological response of SH in the later stages of cold stress is similar to that of HX throughout the cold treatment. Orthogonal partial least squares discriminant analysis (OPLS-DA) revealed that soluble protein content and peroxidase activity are key physiological parameters for assessing the cold tolerance of these two Dendrobium spp. cultivars. Additionally, weighted gene co-expression network analysis (WGCNA) results showed that many cold response genes and metabolic pathways significantly associated with the physiological indices were enriched in the 12 detected modules. The Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene ontology (GO) enrichment analyses of the 105 hub genes showed that Dendrobium spp. adapt to cold stress by regulating signal transduction, phytohormones, transcription factors, protein translation and modification, functional proteins, biosynthesis and metabolism, cell structure, light, and the circadian clock. Hub genes of the cold stress response network included the remorin gene pp34, the abscisic acid signaling pathway-related genes PROTEIN PHOSPATASE 2 C (PP2C), SNF1-RELATED PROTEIN KINASE 2 (SnRK2), ABRE-BINDING FACTOR 1 (ABF1) and SKI-INTERACTING PROTEIN 17 (SKIP17), the Ca2+ signaling-related GTP diphosphokinase gene CRSH1, the carbohydrate-related gene STARCH SYNTHASE 2 (SS2), the cell wall biosynthesis gene CINNAMYL ALCOHOL DEHYDROGENASE (CAD7), and the endocytosis-related gene VACUOLAR PROTEIN SORTING-ASSOCIATED PROTEIN 52 A (VPS52A). CONCLUSIONS: The cold-responsive genes and metabolic pathways of Dendrobium spp. revealed in this study provide important insight to enable the genetic enhancement of cold tolerance in Dendrobium spp., and to facilitate cold tolerance breeding in related plants.

Genomic identification and expression analysis of acid invertase (AINV) gene family in Dendrobium officinale Kimura et Migo.

BACKGROUND: Dendrobium officinale Kimura et Migo, a renowned traditional Chinese orchid herb esteemed for its significant horticultural and medicinal value, thrives in adverse habitats and contends with various abiotic or biotic stresses. Acid invertases (AINV) are widely considered enzymes involved in regulating sucrose metabolism and have been revealed to participate in plant responses to environmental stress. Although members of AINV gene family have been identified and characterized in multiple plant genomes, detailed information regarding this gene family and its expression patterns remains unknown in D. officinale, despite their significance in polysaccharide biosynthesis. RESULTS: This study systematically analyzed the D. officinale genome and identified four DoAINV genes, which were classified into two subfamilies based on subcellular prediction and phylogenetic analysis. Comparison of gene structures and conserved motifs in DoAINV genes indicated a high-level conservation during their evolution history. The conserved amino acids and domains of DoAINV proteins were identified as pivotal for their functional roles. Additionally, cis-elements associated with responses to abiotic and biotic stress were found to be the most prevalent motif in all DoAINV genes, indicating their responsiveness to stress. Furthermore, bioinformatics analysis of transcriptome data, validated by quantitative real-time reverse transcription PCR (qRT-PCR), revealed distinct organ-specific expression patterns of DoAINV genes across various tissues and in response to abiotic stress. Examination of soluble sugar content and interaction networks provided insights into stress release and sucrose metabolism. CONCLUSIONS: DoAINV genes are implicated in various activities including growth and development, stress response, and polysaccharide biosynthesis. These findings provide valuable insights into the AINV gene amily of D. officinale and will aid in further elucidating the functions of DoAINV genes.

Symbiosis between Dendrobium catenatum protocorms and Serendipita indica involves the plant hypoxia response pathway.

Mycorrhizae are ubiquitous symbioses established between fungi and plant roots. Orchids, in particular, require compatible mycorrhizal fungi for seed germination and protocorm development. Unlike arbuscular mycorrhizal fungi, which have wide host ranges, orchid mycorrhizal fungi are often highly specific to their host orchids. However, the molecular mechanism of orchid mycorrhizal symbiosis is largely unknown compared to that of arbuscular mycorrhizal and rhizobial symbiosis. Here, we report that an endophytic Sebacinales fungus, Serendipita indica, promotes seed germination and the development of protocorms into plantlets in several epiphytic Epidendroideae orchid species (6 species in 2 genera), including Dendrobium catenatum, a critically endangered orchid with high medicinal value. Although plant-pathogen interaction and high meristematic activity can induce the hypoxic response in plants, it has been unclear whether interactions with beneficial fungi, especially mycorrhizal ones, also involve the hypoxic response. By studying the symbiotic relationship between D. catenatum and S. indica, we determined that hypoxia-responsive genes, such as those encoding alcohol dehydrogenase (ADH), are highly induced in symbiotic D. catenatum protocorms. In situ hybridization assay indicated that the ADH gene is predominantly expressed in the basal mycorrhizal region of symbiotic protocorms. Additionally, the ADH inhibitors puerarin and 4-methylpyrazole both decreased S. indica colonization in D. catenatum protocorms. Thus, our study reveals that S. indica is widely compatible with orchids and that ADH and its related hypoxia-responsive pathway are involved in establishing successful symbiotic relationships in germinating orchids.

DcERF109 regulates shoot branching by participating in strigolactone signal transduction in Dendrobium catenatum.

Shoot branching fundamentally influences plant architecture and agricultural yield. However, research on shoot branching in Dendrobium catenatum, an endangered medicinal plant in China, remains limited. In this study, we identified a transcription factor DcERF109 as a key player in shoot branching by regulating the expression of strigolactone (SL) receptors DWARF 14 (D14)/ DECREASED APICAL DOMINANCE 2 (DAD2). The treatment of D. catenatum seedlings with GR24rac/TIS108 revealed that SL can significantly repress the shoot branching in D. catenatum. The expression of DcERF109 in multi-branched seedlings is significantly higher than that of single-branched seedlings. Ectopic expression in Arabidopsis thaliana demonstrated that overexpression of DcERF109 resulted in significant shoot branches increasing and dwarfing. Molecular and biochemical assays demonstrated that DcERF109 can directly bind to the promoters of AtD14 and DcDAD2.2 to inhibit their expression, thereby positively regulating shoot branching. Inhibition of DcERF109 by virus-induced gene silencing (VIGS) resulted in decreased shoot branching and improved DcDAD2.2 expression. Moreover, overexpression of DpERF109 in A. thaliana, the homologous gene of DcERF109 in Dendrobium primulinum, showed similar phenotypes to DcERF109 in shoot branch and plant height. Collectively, these findings shed new insights into the regulation of plant shoot branching and provide a theoretical basis for improving the yield of D. catenatum.

Drought Stress Induced Different Response Mechanisms in Three Dendrobium Species under Different Photosynthetic Pathways.

Many Dendrobium species, which hold a high status and value in traditional Chinese medicine, grow on barks and rocks in the wild, often encountering harsh environments and facing droughts. However, the molecular mechanisms underlying the shift in the photosynthetic pathway induced by drought remain unclear. To address this issue, three Dendrobium species with different photosynthetic pathways were selected for sequencing and transcriptome data analysis after drought treatment. The findings included 134.43 GB of sequencing data, with numerous Differentially Expressed Genes (DEGs) exhibiting different response mechanisms under drought stress. Gene Ontology (GO)-KEGG-based enrichment analysis of DEGs revealed that metabolic pathways contributed to drought tolerance and alterations in photosynthetic pathways. Phosphoenolpyruvate Carboxylase (PEPC) was subjected to phylogenetic tree construction, sequence alignment, and domain analysis. Under drought stress, variations were observed in the PEPC gene structure and expression among different Dendrobium species; the upregulation of Dc_gene2609 expression may be caused by dof-miR-384, which resulted in the shift from C3 photosynthesis to CAM, thereby improving drought tolerance in Dendrobium. This study revealed the expression patterns and roles of PEPC genes in enhancing plant drought tolerance and will provide an important basis for in-depth research on Dendrobium's adaptation mechanisms in arid environments.

The Evolution of the WUSCHEL-Related Homeobox Gene Family in Dendrobium Species and Its Role in Sex Organ Development in D. chrysotoxum.

The WUSCHEL-related homeobox (WOX) transcription factor plays a vital role in stem cell maintenance and organ morphogenesis, which are essential processes for plant growth and development. Dendrobium chrysotoxum, D. huoshanense, and D. nobile are valued for their ornamental and medicinal properties. However, the specific functions of the WOX gene family in Dendrobium species are not well understood. In our study, a total of 30 WOX genes were present in the genomes of the three Dendrobium species (nine DchWOXs, 11 DhuWOXs, and ten DnoWOXs). These 30 WOXs were clustered into ancient clades, intermediate clades, and WUS/modern clades. All 30 WOXs contained a conserved homeodomain, and the conserved motifs and gene structures were similar among WOXs belonging to the same branch. D. chrysotoxum and D. huoshanense had one pair of fragment duplication genes and one pair of tandem duplication genes, respectively; D. nobile had two pairs of fragment duplication genes. The cis-acting regulatory elements (CREs) in the WOX promoter region were mainly enriched in the light response, stress response, and plant growth and development regulation. The expression pattern and RT-qPCR analysis revealed that the WOXs were involved in regulating the floral organ development of D. chrysotoxum. Among them, the high expression of DchWOX3 suggests that it might be involved in controlling lip development, whereas DchWOX5 might be involved in controlling ovary development. In conclusion, this work lays the groundwork for an in-depth investigation into the functions of WOX genes and their regulatory role in Dendrobium species' floral organ development.

Metabolic and Transcriptomic Profile Revealing the Differential Accumulating Mechanism in Different Parts of Dendrobium nobile.

Dendrobium nobile is an important orchid plant that has been used as a traditional herb for many years. For the further pharmaceutical development of this resource, a combined transcriptome and metabolome analysis was performed in different parts of D. nobile. First, saccharides, organic acids, amino acids and their derivatives, and alkaloids were the main substances identified in D. nobile. Amino acids and their derivatives and flavonoids accumulated strongly in flowers; saccharides and phenols accumulated strongly in flowers and fruits; alkaloids accumulated strongly in leaves and flowers; and a nucleotide and its derivatives and organic acids accumulated strongly in leaves, flowers, and fruits. Simultaneously, genes for lipid metabolism, terpenoid biosynthesis, and alkaloid biosynthesis were highly expressed in the flowers; genes for phenylpropanoids biosynthesis and flavonoid biosynthesis were highly expressed in the roots; and genes for other metabolisms were highly expressed in the leaves. Furthermore, different members of metabolic enzyme families like cytochrome P450 and 4-coumarate-coA ligase showed differential effects on tissue-specific metabolic accumulation. Members of transcription factor families like AP2-EREBP, bHLH, NAC, MADS, and MYB participated widely in differential accumulation. ATP-binding cassette transporters and some other transporters also showed positive effects on tissue-specific metabolic accumulation. These results systematically elucidated the molecular mechanism of differential accumulation in different parts of D. nobile and enriched the library of specialized metabolic products and promising candidate genes.

[Cloning and functional analysis of flavanone synthase â ¡ from Dendrobium huoshanense].

Flavonoid C-glycosides are a class of natural products that are widely involved in plant defense responses and have diverse pharmacological activities. They are also important active ingredients of Dendrobium huoshanense. Flavanone synthase Ⅱ has been proven to be a key enzyme in the synthesis pathway of flavonoid C-glycosides in plants, and their catalytic product 2-hydroxyflavanone is the precursor compound for the synthesis of various reported flavonoid C-glycosides. In this study, based on the reported amino acid sequence of flavanone synthase Ⅱ, a flavanone synthase Ⅱ gene(DhuFNSⅡ) was screened and verified from the constructed D. huoshanense genome localization database. Functional validation of the enzyme showed that it could in vitro catalyze naringenin and pinocembrin to produce apigenin and chrysin, respectively. The open reading frame(ORF) of DhuFNSⅡ was 1 644 bp in length, encoding 547 amino acids. Subcellular localization showed that the protein was localized on the endoplasmic reticulum. RT-qPCR results showed that DhuFNSⅡ had the highest expression in stems, followed by leaves and roots. The expression levels of DhuFNSⅡ and other target genes in various tissues of D. huoshanense were significantly up-regulated after four kinds of abiotic stresses commonly encountered in the growth process, but the extent of up-regulation varied among treatment groups, with drought and cold stress having more significant effects on gene expression levels. Through the identification and functional analysis of DhuFNSⅡ, this study is expected to contribute to the elucidation of the molecular mechanism of the formation of quality metabolites of D. huoshanense, flavonoid C-glycosides, and provide a reference for its quality formation and scientific cultivation.

Genome-wide identification, characterization and transcriptional profile of the SWEET gene family in Dendrobium officinale.

BACKGROUND: Dendrobium officinale Kimura et Migo (D. officinale) is a well-known traditional Chinese medicine with high content polysaccharides in stems. The SWEET (Sugars Will Eventually be Exported Transporters) family is a novel class of sugar transporters mediating sugar translocation among adjacent cells of plants. The expression patterns of SWEETs and whether they are associated with stress response in D. officinale remains uncovered. RESULTS: Here, 25 SWEET genes were screened out from D. officinale genome, most of which typically contained seven transmembrane domains (TMs) and harbored two conserved MtN3/saliva domains. Using multi-omics data and bioinformatic approaches, the evolutionary relationship, conserved motifs, chromosomal location, expression patterns, correlationship and interaction network were further analyzed. DoSWEETs were intensively located in nine chromosomes. Phylogenetic analysis revealed that DoSWEETs were divided into four clades, and conserved motif 3 specifically existed in DoSWEETs from clade II. Different tissue-specific expression patterns of DoSWEETs suggested the division of their roles in sugar transport. In particular, DoSWEET5b, 5c, and 7d displayed relatively high expression levels in stems. DoSWEET2b and 16 were significantly regulated under cold, drought, and MeJA treatment, which were further verified using RT-qPCR. Correlation analysis and interaction network prediction discovered the internal relationship of DoSWEET family. CONCLUSIONS: Taken together, the identification and analysis of the 25 DoSWEETs in this study provide basic information for further functional verification in D. officinale.

Genome-wide identification of Aux/IAA and ARF gene families reveal their potential roles in flower opening of Dendrobium officinale.

BACKGROUND: The auxin indole-3-acetic acid (IAA) is a vital phytohormone that influences plant growth and development. Our previous work showed that IAA content decreased during flower development in the medicinally important orchid Dendrobium officinale, while Aux/IAA genes were downregulated. However, little information about auxin-responsive genes and their roles in D. officinale flower development exists. RESULTS: This study validated 14 DoIAA and 26 DoARF early auxin-responsive genes in the D. officinale genome. A phylogenetic analysis classified the DoIAA genes into two subgroups. An analysis of cis-regulatory elements indicated that they were related by phytohormones and abiotic stresses. Gene expression profiles were tissue-specific. Most DoIAA genes (except for DoIAA7) were sensitive to IAA (10 µmol/L) and were downregulated during flower development. Four DoIAA proteins (DoIAA1, DoIAA6, DoIAA10 and DoIAA13) were mainly localized in the nucleus. A yeast two-hybrid assay showed that these four DoIAA proteins interacted with three DoARF proteins (DoARF2, DoARF17, DoARF23). CONCLUSIONS: The structure and molecular functions of early auxin-responsive genes in D. officinale were investigated. The DoIAA-DoARF interaction may play an important role in flower development via the auxin signaling pathway.

The climate changes promoted the chloroplast genomic evolution of Dendrobium orchids among multiple photosynthetic pathways.

Dendrobium orchids have multiple photosynthetic pathways, which can be used as a model system for studying the evolution of crassulacean acid metabolism (CAM). In this study, based on the results of the net photosynthetic rates (Pn), we classified Dendrobium species into three photosynthetic pathways, then employed and compared their chloroplast genomes. The Dendrobium chloroplast genomes have typical quartile structures, ranging from 150,841-153,038 bp. The apparent differences in GC content, sequence variability, and IR junctions of SSC/IRB junctions (JSBs) were measured within chloroplast genomes among different photosynthetic pathways. The phylogenetic analysis has revealed multiple independent CAM origins among the selected Dendrobium species. After counting insertions and deletions (InDels), we found that the occurrence rates and distribution densities among different photosynthetic pathways were inconsistent. Moreover, the evolution patterns of chloroplast genes in Dendrobium among three photosynthetic pathways were also diversified. Considering the diversified genome structure variations and the evolution patterns of protein-coding genes among Dendrobium species, we proposed that the evolution of the chloroplast genomes was disproportional among different photosynthetic pathways. Furthermore, climatic correlation revealed that temperature and precipitation have influenced the distribution among different photosynthetic pathways and promoted the foundation of CAM pathway in Dendrobium orchids. Based on our study, we provided not only new insights into the CAM evolution of Dendrobium but also provided beneficial genetic data resources for the further systematical study of Dendrobium.

Uncovering the involvement of DoDELLA1-interacting proteins in development by characterizing the DoDELLA gene family in Dendrobium officinale.

BACKGROUND: Gibberellins (GAs) are widely involved in plant growth and development. DELLA proteins are key regulators of plant development and a negative regulatory factor of GA. Dendrobium officinale is a valuable traditional Chinese medicine, but little is known about D. officinale DELLA proteins. Assessing the function of D. officinale DELLA proteins would provide an understanding of their roles in this orchid's development. RESULTS: In this study, the D. officinale DELLA gene family was identified. The function of DoDELLA1 was analyzed in detail. qRT-PCR analysis showed that the expression levels of all DoDELLA genes were significantly up-regulated in multiple shoots and GA3-treated leaves. DoDELLA1 and DoDELLA3 were significantly up-regulated in response to salt stress but were significantly down-regulated under drought stress. DoDELLA1 was localized in the nucleus. A strong interaction was observed between DoDELLA1 and DoMYB39 or DoMYB308, but a weak interaction with DoWAT1. CONCLUSIONS: In D. officinale, a developmental regulatory network involves a close link between DELLA and other key proteins in this orchid's life cycle. DELLA plays a crucial role in D. officinale development.

Mitochondrial genome comparison and phylogenetic analysis of Dendrobium (Orchidaceae) based on whole mitogenomes.

BACKGROUND: Mitochondrial genomes are essential for deciphering the unique evolutionary history of seed plants. However, the rules of their extreme variation in genomic size, multi-chromosomal structure, and foreign sequences remain unresolved in most plant lineages, which further hindered the application of mitogenomes in phylogenetic analyses. RESULTS: Here, we took Dendrobium (Orchidaceae) which shows the great divergence of morphology and difficulty in species taxonomy as the study focus. We first de novo assembled two complete mitogenomes of Dendrobium wilsonii and Dendrobium henanense that were 763,005 bp and 807,551 bp long with multichromosomal structures. To understand the evolution of Dendrobium mitogenomes, we compared them with those of four other orchid species. The results showed great variations of repetitive and chloroplast-derived sequences in Dendrobium mitogenomes. Moreover, the intergenic content of Dendrobium mitogenomes has undergone expansion during evolution. We also newly sequenced mitogenomes of 26 Dendrobium species and reconstructed phylogenetic relationships of Dendrobium based on genomic mitochondrial and plastid data. The results indicated that the existence of chloroplast-derived sequences made the mitochondrial phylogeny display partial characteristics of the plastid phylogeny. Additionally, the mitochondrial phylogeny provided new insights into the phylogenetic relationships of Dendrobium species. CONCLUSIONS: Our study revealed the evolution of Dendrobium mitogenomes and the potential of mitogenomes in deciphering phylogenetic relationships at low taxonomic levels.

Complete genome of Sphingomonas paucimobilis ZJSH1, an endophytic bacterium from Dendrobium officinale with stress resistance and growth promotion potential.

Sphingomonas paucimobilis ZJSH1 is an endophytic bacterium isolated from the roots of Dendrobium officinale with the ability to promote plant growth. It was found that the genome of strain ZJSH1 had gene fragment rearrangement compared with the genomes of the other four strains of S. paucimobilis, and the genome was integrated with phage genes. Functional analysis showed that the strain contained colonization-related genes, chemotaxis and invasion. A variety of genes encoding active materials, such as hormones (IAA, SA, ABA and zeaxanthin), phosphate cycle, antioxidant enzymes, and polysaccharides were identified which provide the strain with growth promotion and stress-resistant characteristics. Experiments proved that S. paucimobilis ZJSH1 grew well in media containing 80 g/L sodium chloride, 240 g/L polyethylene glycol and 800 µmol/L Cd2+, indicating its potential for resistance to stresses of salt, drought and cadmium, respectively. S. paucimobilis ZJSH1 is the only endophytic bacterium of this species that has been reported to promote plant growth. The analysis of its genome is conducive to understanding its growth-promoting mechanism and laying a foundation for the development and utilization of this species in the field of agriculture.

Transcriptome and metabolome profiling unveil the accumulation of flavonoids in Dendrobium officinale.

Dendrobium officinale is a Chinese herbal medicine with a long history of use in China. Flavonoids are known to be an important secondary metabolite in Dendrobium officinale, but very little is known about their molecular regulation mechanism in D. officinale. In this study, we collected one to four years old D. officinale stems for the purpose of RNA-sequencing and mass spectrometry data collection. The results showed that metabolome analysis detected 124 different flavonoid metabolites of which flavonol metabolites were significantly increased in biennial samples. In the transcriptome analysis, 30 different genes involved in the synthesis of flavonoid were identified. The key genes FLS (LOC110101392, LOC110107557, LOC110114894) that regulate the synthesis of flavonols are highly expressed in biennial samples. The present study contributes a new insight into the molecular mechanism of flavonoid accumulation in D. officinale.

Metabolic Pathway Engineering Improves Dendrobine Production in Dendrobium catenatum.

The sesquiterpene alkaloid dendrobine, widely recognized as the main active compound and a quality control standard of medicinal orchids in the Chinese Pharmacopoeia, demonstrates diverse biological functions. In this study, we engineered Dendrobium catenatum as a chassis plant for the production of dendrobine through the screening and pyramiding of key biosynthesis genes. Initially, previously predicted upstream key genes in the methyl-D-erythritol 4-phosphate (MEP) pathway for dendrobine synthesis, including 4-(Cytidine 5'-Diphospho)-2-C-Methyl-d-Erythritol Kinase (CMK), 1-Deoxy-d-Xylulose 5-Phosphate Reductoisomerase (DXR), 2-C-Methyl-d-Erythritol 4-Phosphate Cytidylyltransferase (MCT), and Strictosidine Synthase 1 (STR1), and a few downstream post-modification genes, including Cytochrome P450 94C1 (CYP94C1), Branched-Chain-Amino-Acid Aminotransferase 2 (BCAT2), and Methyltransferase-like Protein 23 (METTL23), were chosen due to their deduced roles in enhancing dendrobine production. The seven genes (SG) were then stacked and transiently expressed in the leaves of D. catenatum, resulting in a dendrobine yield that was two-fold higher compared to that of the empty vector control (EV). Further, RNA-seq analysis identified Copper Methylamine Oxidase (CMEAO) as a strong candidate with predicted functions in the post-modification processes of alkaloid biosynthesis. Overexpression of CMEAO increased dendrobine content by two-fold. Additionally, co-expression analysis of the differentially expressed genes (DEGs) by weighted gene co-expression network analysis (WGCNA) retrieved one regulatory transcription factor gene MYB61. Overexpression of MYB61 increased dendrobine levels by more than two-fold in D. catenatum. In short, this work provides an efficient strategy and prospective candidates for the genetic engineering of D. catenatum to produce dendrobine, thereby improving its medicinal value.

Transcriptional and Physiological Analysis Reveal New Insights into the Regulation of Fertilization (N, P, K) on the Growth and Synthesis of Medicinal Components of Dendrobium denneanum.

Dendrobium denneanum is an important medicinal and ornamental plant. Its ornamental and medicinal values are affected by its vegetative growth conditions and chemical composition accumulation. This study adopted an orthogonal experimental design to treat D. denneanum with nine different levels of nitrogen (N), potassium (K), and phosphorus (P). The morphological indicators of the plant were positively correlated with the nitrogen concentration. The polysaccharide content was the highest at 1500 mg·L-1 nitrogen and 3000 mg·L-1 phosphorous and was 26.84% greater than the control. The flavonoid content increased by 36.2% at 500 mg·L-1 nitrogen, 2000 mg·L-1 phosphorous, and 300 mg·L-1 potassium. Principal component score analysis showed that nitrogen had the most significant impact on the various indicators of D. denneanum, followed by phosphorus and potassium. The comprehensive score showed that the T9 treatment (N: 1500 mg·L-1, P: 3000 mg·L-1, K: 500 mg·L-1) had the strongest effect on D. denneanum. Transcriptional analysis showed that compared with the control, the T9 treatment led to 2277 differentially expressed genes (1230 upregulated and 1047 downregulated). This includes fifteen genes enriched in the MAPK signaling pathway, five genes in phenylpropanoid biosynthesis, and two genes in flavonoid biosynthesis. These genes may be involved in regulating plant growth and the biosynthesis of polysaccharides and flavonoids. This study provides guidance for the optimal use of N, P, and K in the cultivation of D. denneanum.