Dendrobium Multi-Omics Reveal Lipid Remodeling in Response to Freezing

Freezing damage is a common phenomenon responsible for reduced yields of economic crops. Regulation of lipid metabolism plays an important role in plant growth and adaptation during freezing. We previously carried out transcriptome and untargeted metabolome analyses to determine the regulation of flavonol and anthocyanin biosynthesis during freezing treatment (FT) and post-freezing recovery (FR) in Dendrobium catenatum. However, changes in lipid levels are hard to confirm by untargeted metabolomics analysis alone. Regulation of lipid metabolism in response to freezing is largely unknown in Dendrobium. In this study, a multi-omics strategy was used to offer a better means of studying metabolic flow during FT and FR. To this end, 6976 proteins were identified by the 4D_label-free proteome, including 5343 quantified proteins. For each of the two conditions, we enriched differentially accumulated proteins (DAPs) into 15 gene ontology (GO) terms, including primary metabolism, lipid metabolism, and photosynthesis processes. We also identified 7 lipid categories and 3672 lipid species using lipidome assays. We found significant remodeling occurring in the phospholipid category during FT and FR. We also found that most sphingolipids were significantly upregulated. An integrated multi-omics analysis revealed significant changes in the expression levels of 141 mRNAs and encoding proteins under both FT and FR conditions. During FT, phospholipase A (PLA) and phospholipase D (PLD) were associated with phospholipid editing and galactolipid remodeling. These results provide valuable new insights into how the freezing tolerance of D. catenatum might be improved by genetic engineering

<i>Dendrobium</i> Multi-Omics Reveal Lipid Remodeling in Response to Freezing

Freezing damage is a common phenomenon responsible for reduced yields of economic crops. Regulation of lipid metabolism plays an important role in plant growth and adaptation during freezing. We previously carried out transcriptome and untargeted metabolome analyses to determine the regulation of flavonol and anthocyanin biosynthesis during freezing treatment (FT) and post-freezing recovery (FR) in Dendrobium catenatum. However, changes in lipid levels are hard to confirm by untargeted metabolomics analysis alone. Regulation of lipid metabolism in response to freezing is largely unknown in Dendrobium. In this study, a multi-omics strategy was used to offer a better means of studying metabolic flow during FT and FR. To this end, 6976 proteins were identified by the 4D_label-free proteome, including 5343 quantified proteins. For each of the two conditions, we enriched differentially accumulated proteins (DAPs) into 15 gene ontology (GO) terms, including primary metabolism, lipid metabolism, and photosynthesis processes. We also identified 7 lipid categories and 3672 lipid species using lipidome assays. We found significant remodeling occurring in the phospholipid category during FT and FR. We also found that most sphingolipids were significantly upregulated. An integrated multi-omics analysis revealed significant changes in the expression levels of 141 mRNAs and encoding proteins under both FT and FR conditions. During FT, phospholipase A (PLA) and phospholipase D (PLD) were associated with phospholipid editing and galactolipid remodeling. These results provide valuable new insights into how the freezing tolerance of D. catenatum might be improved by genetic engineering

Metabolic Profiling of Terpene Diversity and the Response of Prenylsynthase-Terpene Synthase Genes during Biotic and Abiotic Stresses in <i>Dendrobium catenatum</i>

Dendrobium catenatum is a widely cultivated Chinese orchid herb rich in abundant secondary metabolites, such as terpenes. However, terpene distribution and characterization of terpene biosynthesis-related genes remain unknown in D. catenatum. In this study, metabolic profiling was performed to analyze terpene distribution in the root, stem, leaf, and flower of D. catenatum. A total of 74 terpene compounds were identified and classified. Clustering analysis revealed that terpene compounds exhibited a tissue-specific accumulation, including monoterpenes in the flowers, sesquiterpenes in the stems, and triterpenes in the roots. Transcriptome analysis revealed that the ‘terpenoid backbone biosynthesis’ pathway was only significantly enriched in root vs. flower. The expression of terpene biosynthesis-related genes was spatiotemporal in the flowers. Prenylsynthase-terpene synthases (PS-TPSs) are the largest and core enzymes for generating terpene diversity. By systematic sequence analysis of six species, 318 PS-TPSs were classified into 10 groups and 51 DcaPS-TPSs were found in eight of them. Eighteen DcaPS-TPSs were regulated by circadian rhythm under drought stress. Most of the DcaPS-TPSs were influenced by cold stress and fungi infection. The cis-element of the majority of the DcaPS-TPS promoters was related to abiotic stress and plant development. Methyl jasmonate levels were significantly associated with DcaTPSs expression and terpene biosynthesis. These results provide insight into further functional investigation of DcaPS-TPSs and the regulation of terpene biosynthesis in Dendrobium

Identification of Two GDSL-Type Esterase/Lipase Genes Related to Tissue-Specific Lipolysis in <i>Dendrobium catenatum</i> by Multi-Omics Analysis

Dendrobium catenatum is an important herb and widely cultivated in China. GDSL-Type Esterase/Lipase proteins (GELPs) are widely distributed in plants and play crucial roles in stress responses, plant growth, and development. However, no identification or functional analysis of GELPs was reported in D. catenatum. This study identifies 52 GELPs in D. catenatum genome, which is classified into four groups by phylogenetic analysis. Four conservative blocks (Ser-Gly-Asn-His) are found in most GELP domains. Transcriptome analysis reveals the expression profiles of GELPs in different organs and flowering phases. Co-expression analysis of the transcriptome and lipidome identifies a GELP gene, Dca016600, that positively correlates with 23 lipids. The purified Dca016600 protein shows the optimum pH is active from 8.0 to 8.5, and the optimum temperature is active from 30 °C to 40 °C. The kinetic study provides Vmax (233.43 μmol·min−1·mg−1) and Km (1.49 mM) for substrate p-nitrophenyl palmitate (p-NPP). Integrated analysis of the transcriptome and proteome identifies a GELP gene, Dca005399, which is specially induced by freezing. Interestingly, Dca005399 shows high expression in symbiotic germination seeds and sepals. This study provides new insights into the function of D. catenatum GELPs in plant development and stress tolerance

Identification and Expression Profiling of Nonphosphorus Glycerolipid Synthase Genes in Response to Abiotic Stresses in Dendrobium catenatum

Dendrobium&nbsp;catenatum, a valuable Chinese herb, frequently experiences abiotic stresses, such as cold and drought, under natural conditions. Nonphosphorus glycerolipid synthase (NGLS) genes are closely linked to the homeostasis of membrane lipids under abiotic stress in plants. However, there is limited information on NGLS genes in D. catenatum. In this study, a total of eight DcaNGLS genes were identified from the D. catenatum genome; these included three monogalactosyldiacylglycerol synthase (DcaMGD1, 2, 3) genes, two digalactosyldiacylglycerol synthase (DcaDGD1, 2) genes, and three sulfoquinovosyldiacylglycerol synthase (DcaSQD1, 2.1, 2.2) genes. The gene structures and conserved motifs in the DcaNGLSs showed a high conservation during their evolution. Gene expression profiling showed that the DcaNGLSs were highly expressed in specific tissues and during rapid growth stages. Furthermore, most DcaNGLSs were strongly induced by freezing and post-freezing recovery. DcaMGD1 and DcaSQDs were greatly induced by salt stress in leaves, while DcaDGDs were primarily induced by salt stress in roots. Under drought stress, most DcaNGLSs were regulated by circadian rhythms, and DcaSQD2 was closely associated with drought recovery. Transcriptome analysis also revealed that MYB might be regulated by circadian rhythm and co-expressed with DcaNGLSs under drought stress. These results provide insight for the further functional investigation of NGLS and the regulation of nonphosphorus glycerolipid biosynthesis in Dendrobium

The Sulfoquinovosyltransferaselike Enzyme SQD2.2 is Involved in Flavonoid Glycosylation, Regulating Sugar Metabolism and Seed Setting in Rice

Seed setting is an important trait that contributes to seed yield and relies greatly on starchaccumulation. In this study, a sulfoquinovosyl transferase-like protein, designated as SQD2.2involved in seed setting and flavonoid accumulation, was identified and characterized in rice.Rice SQD2.2 is localized to the cytoplasm, and the SQD2.2 transcript was highest in leaves. RiceSQD2.2-overexpressing (OE) plants exhibited a decreased seed setting rate and diminished tillernumber simultaneously with an increased glycosidic flavonoid level compared with wild-type(WT) plants. SQD2.2 catalyzes the glycosylation of apigenin to produce apigenin 7-O-glucosideusing uridine diphosphate-glucose (UDPG) as a sugar donor, but it failed to compensate forsulfoquinovosyldiacylglycerol (SQDG) synthesis in the Arabidopsis sqd2 mutant. Furthermore,apigenin 7-O-glucoside inhibited starch synthase (SS) activity in a concentration-dependent manner,and SQD2.2-OE plants exhibited reduced SS activity accompanied by a significant reduction in starchlevels and an elevation in soluble sugar levels relative to WT plants. Both adenosine diphosphateglucose (ADPG) and UDPG levels in SQD2.2-OE plants were notably lower than those in WT plants.Taken together, rice SQD2.2 exhibits a novel role in flavonoid synthesis and plays an important role inmediating sugar allocation between primary and secondary metabolism in rice

Evaluation of Virus-Free Chrysanthemum ‘Hangju’ Productivity and Response to Virus Reinfection in the Field: Molecular Insights into Virus–Host Interactions

The shoot apical meristem culture has been used widely to produce virus-free plantlets which have the advantages of strong disease resistance, high yield, and prosperous growth potential. However, this virus-free plant will be naturally reinfected in the field. The physiological and metabolic responses in the reinfected plant are still unknown. The flower of chrysanthemum ‘Hangju’ is a traditional medicine which is unique to China. In this study, we found that the virus-free ‘Hangju’ (VFH) was reinfected with chrysanthemum virus B/R in the field. However, the reinfected VFH (RVFH) exhibited an increased yield and medicinal components compared with virus-infected ‘Hangju’ (VIH). Comparative analysis of transcriptomes was performed to explore the molecular response mechanisms of the RVFH to CVB infection. A total of 6223 differentially expressed genes (DEGs) were identified in the RVFH vs. the VIH. KEGG enrichment and physiological analyses indicated that treatment with the virus-free technology significantly mitigated the plants’ lipid and galactose metabolic stress responses in the RVFH. Furthermore, GO enrichment showed that plant viral diseases affected salicylic acid (SA)-related processes in the RVFH. Specifically, we found that phenylalanine ammonia-lyase (PAL) genes played a major role in defense-related SA biosynthesis in ‘Hangju’. These findings provided new insights into the molecular mechanisms underlying plant virus–host interactions and have implications for developing strategies to improve plant resistance against viruses

Dual Activities of Plant cGMP-Dependent Protein Kinase and its Roles in Gibberellin Signaling and Salt Stress.

Cyclic GMP (cGMP) is an important regulator in eukaryotes, and cGMP-dependent protein kinase (PKG) plays a key role in perceiving cellular cGMP in diverse physiological processes in animals. However, the molecular identity, property, and function of PKG in plants remain elusive. In this study, we have identified PKG from plants and characterized its role in mediating the gibberellin (GA) response in rice (Oryza sativa). PKGs from plants are structurally unique with an additional type 2C protein phosphatase domain. Rice PKG possesses both protein kinase and phosphatase activities, and cGMP stimulates its kinase activity but inhibits its phosphatase activity. One of PKG’s targets is GAMYB, a transcription factor in GA signaling, and the dual activities of PKG catalyze the reversible phosphorylation of GAMYB at Ser6 and modulate the nucleocytoplasmic distribution of GAMYB in response to GA. Loss of PKG impeded the nuclear localization of GAMYB and abolished GAMYB function in the GA response, leading to defects in GA-induced seed germination, internode elongation, and pollen viability. In addition to GAMYB, PKG has multiple potential targets and thus has broad effects, particularly in the salt stress response