Monoterpenoid Biosynthesis Research Articles

Comparative Transcriptome Analysis of Flower Senescence of Camellia lutchuensis.

BackgroundFlower senescence is the last stage of flower development and affects the ornamental and economic value of flower plants. There is still less known on flower senescence of the ornamental plant Camellia lutchuensis, a precious species of Camellia with significant commercial application value.MethodsTranscriptome sequencing was used to investigate the flower senescence in five developmental stages of C. lutchuensis.ResultsBy Illumina HiSeq sequencing, we generated approximately 101.16 Gb clean data and 46649 differentially expressed unigenes. Based on the different expression pattern, differentially expressed unigenes were classified into 10 Sub Class. And Sub Class 9 including 8252 unigenes, was highly expressed in the flower senescent stage, suggesting it had a potential regulatory relationship of flower senescence. First, we found that ethylene biosynthesis genes ACSs, ACOs, receptor ETR genes and signaling genes EINs, ERFs all upregulated during flower senescence, suggesting ethylene might play a key role in the flower senescence of C. lutchuensis. Furthermore, reactive oxygen species (ROS) production related genes peroxidase (POD), lipase (LIP), polyphenoloxidase (PPO), and ROS scavenging related genes glutathione S-transferase (GST), glutathione reductase (GR) and superoxide dismutase (SOD) were induced in senescent stage, suggesting ROS might be involved in the flower senescence. Besides, the expression of monoterpenoid and isoflavonoid biosynthesis genes, transcription factors (WRKY, NAC, MYB and C2H2), senescence-associated gene SAG20 also were increased during flower senescence.ConclusionIn C. lutchuensis, ethylene pathway might be the key to regulate flower senescence, and ROS signal might play a role in the flower senescence.

Comparative transcriptome profiling reveals the defense pathways and mechanisms in the leaves and roots of blueberry to drought stress

Blueberry is an important horticultural plant that is very susceptible to drought. However, the molecular regulation mechanisms of blueberry tolerance to drought remain elusive. In this study, the transcriptome profile of blueberry by RNA-seq under different degree of drought treatment were conducted and drought-related genes and pathways were screened by using weighted gene co-expression network analysis (WGCNA) and Venn analysis. Interestingly, the leaves and roots of blueberry exhibited different expression patterns under drought treatment and differentially expressed genes (DEGs), GO terms and KEGG pathways showed more enriched in leaves than in roots. The majority of DEGs were associated with signal transduction, transcriptional regulation, and metabolism. 8 key pathways and 8 DEGs were shared both in leaves and roots. Notably, the pathway with the most of core genes in leaves is plant hormone signal transduction, and in roots are MAPK signaling pathway, reactive oxygen species metabolism and the key genes such as <italic>VcXTH3</italic>, <italic>VcPP2C51</italic>, and <italic>VcPNC1</italic> were identified. For transcription factors, <italic>VcABR1</italic>, <italic>VcABF2</italic>, <italic>VcMYB108</italic> and <italic>VcMYB93</italic> are likely involved in drought response. In metabolism category, <italic>VcCYP75A1</italic> was likely involved in anthocyanin biosynthesis, and <italic>VcPNC1</italic> in monoterpenoid biosynthesis pathway. The eight DEGs markedly induced under drought exhibited differential expression in leaves and roots. Correspondingly, physiological assays showed that POD activity, ABA content, and anthocyanin biosynthesis in leaves and roots were significantly increased. Collectively, our study revealed the synergistic and distinct defense pathways and mechanisms in leaves and roots of blueberry and explored the potential regulatory network in blueberry response to drought stress.

Transcriptomic profiling of the floral fragrance biosynthesis pathway of Liriodendron and functional characterization of the LtuDXR gene

Floral fragrance, which has the function of attracting pollinators, is a class of volatile secondary metabolites mainly released by the secretory tissue of petals. Terpenoids are key components of floral volatile substances. Previous studies have shown that there are significant differences in the concentration and composition of volatile floral fragrances, especially terpenoids, between Liriodendron chinense and L. tulipifera. At present, the mechanism by which the synthesis of floral fragrance is regulated in Liriodendron remains unexplored. In this study, we analyzed the differentially expressed genes (DEGs) of L. chinense and L. tulipifera, and identified 130 DEGs related to terpenoid synthesis. A KEGG enrichment analysis of DEGs related to terpenoid biosynthesis revealed that the monoterpenoid biosynthesis pathway was the most significant. We cloned the LtuDXR gene from L. tulipifera using RACE technology. RT-qPCR results showed that the expression of the LtuDXR gene was the highest in the early florescence petals, indicating that the LtuDXR gene may play a role in the synthesis of volatile terpenoids. Subcellular localization showed that the LtuDXR protein is mainly localized in the chloroplast. Overexpression of LtuDXR in Arabidopsis thaliana significantly increased the plant height, DXR enzyme activity, and carotenoid content. In this study, we identified and functionally characterized LtuDXR, which is involved in terpenoid synthesis in Liriodendron. Our work lays the foundation for further exploration of the molecular mechanism by which terpenoid biosynthesis is regulated in Liriodendron.

Selectivity of enzymes involved in the formation of opposite enantiomeric series of p-menthane monoterpenoids in peppermint and Japanese catnip

Peppermint (Mentha x piperita L.) and Japanese catnip (Schizonepeta tenuifolia (Benth.) Briq.) accumulate p-menthane monoterpenoids with identical functionalization patterns but opposite stereochemistry. In the present study, we investigate the enantioselectivity of multiple enzymes involved in monoterpenoid biosynthesis in these species. Based on kinetic assays, mint limonene synthase, limonene 3-hydroxylase, isopiperitenol dehydrogenase, isopiperitenone reductase, and menthone reductase exhibited significant enantioselectivity toward intermediates of the pathway that proceeds through (-)-4S-limonene. Limonene synthase, isopiperitenol dehydrogenase and isopiperitenone reductase of Japanese catnip preferred intermediates of the pathway that involves (+)-4R-limonene, whereas limonene 3-hydroxylase was not enantioselective, and the activities of pulegone reductase and menthone reductase were too low to acquire meaningful kinetic data. Molecular modeling studies with docked ligands generally supported the experimental data obtained with peppermint enzymes, indicating that the preferred enantiomer was aligned well with the requisite cofactor and amino acid residues implicated in catalysis. A striking example for enantioselectivity was peppermint (-)-menthone reductase, which binds (-)-menthone with exquisite affinity but was predicted to bind (+)-menthone in a non-productive orientation that positions its carbonyl functional group at considerable distance to the NADPH cofactor. The work presented here lays the groundwork for structure-function studies aimed at unraveling how enantioselectivity evolved in closely related species of the Lamiaceae and beyond.

Monoterpenoid biosynthesis by engineered microbes.

Monoterpenoids are C10 isoprenoids and constitute a large family of natural products. They have been used as ingredients in food, cosmetics, and therapeutic products. Many monoterpenoids such as linalool, geraniol, limonene, and pinene are volatile and can be found in plant essential oils. Conventionally, these bioactive compounds are obtained from plant extracts by using organic solvents or by distillation method, which are costly and laborious if high-purity product is desired. In recent years, microbial biosynthesis has emerged as alternative source of monoterpenoids with great promise for meeting the increasing global demand for these compounds. However, current methods of production are not yet at levels required for commercialization. Production efficiency of monoterpenoids in microbial hosts is often restricted by high volatility of the monoterpenoids, a lack of enzymatic activity and selectivity, and/or product cytotoxicity to the microbial hosts. In this review, we summarize advances in microbial production of monoterpenoids over the past 3 years with particular focus on the key metabolic engineering strategies for different monoterpenoid products. We also provide our perspective on the promise of future endeavors to improve monoterpenoid productivity.

New high‐quality peach (Prunus persica L. Batsch) genome assembly to analyze the molecular evolutionary mechanism of volatile compounds in peach fruits

Peach (Prunus persica L. Batsch) is an economically important fruit crop worldwide. Although a high-quality peach genome has previously been published, Sanger sequencing was used for its assembly, which generated short contigs. Here, we report a chromosome-level genome assembly and sequence analysis of Chinese Cling, an important founder cultivar for peach breeding programs worldwide. The assembled genome contained 247.33Mb with a contig N50 of 4.13Mb and a scaffold N50 of 29.68Mb, representing 99.8% of the estimated genome. Comparisons between this genome and the recently published one (Lovell peach) uncovered 685407 single nucleotide polymorphisms, 162655 insertions and deletions, and 16248 structural variants. Gene family analysis highlighted the contraction of the gene families involved in flavone, flavonol, flavonoid, and monoterpenoid biosynthesis. Subsequently, the volatile compounds of 256 peach varieties were quantitated in mature fruits in 2015 and 2016 to perform a genome-wide association analysis. A comparison with the identified domestication genomic regions allowed us to identify 25 quantitative trait loci, associated with seven volatile compounds, in the domestication region, which is consistent with the differences in volatile compounds between wild and cultivated peaches. Finally, a gene encoding terpene synthase, located within a previously reported quantitative trait loci region, was identified to be associated with linalool synthesis. Such findings highlight the importance of this new assembly for the analysis of evolutionary mechanisms and gene identification in peach species. Furthermore, this high-quality peach genome provides valuable information for future fruit improvement.

Integrating GC-MS and ssRNA-Seq analysis to identify long non-coding RNAs related to terpenoid biosynthesis in Cinnamomum camphora

Terpenoids are the main constituents in the leaf essential oil (LEO) of Cinnamomum camphora, which has long been considered an important raw material for food, cosmetic and pharmaceutical industries. However, most previous studies have concentrated more on the functions of structural genes and microRNAs in terpenoid biosynthesis pathways. Long non-coding RNAs (lncRNAs) related to terpenoid biosynthesis are rarely reported. Here, the components of LEOs in cineole-, linalool- and nerolidol-type C. camphora were determined by gas chromatography-mass spectrometry (GC–MS). Strand-specific RNA sequencing (ssRNA-seq) was used to identify lncRNAs involved in terpenoid biosynthesis in three chemotypes of C. camphora. A total of 8826 lncRNAs were first classified in C. camphora: 4331 long intergenic noncoding RNAs (lincRNAs), 1410 antisense lncRNAs, and 3085 sense overlapping lncRNAs. Among these lncRNAs, 243, 343, and 252 were classified as cineole-, linalool- and nerolidol-specific lncRNAs, respectively. Many target genes of differentially expressed lncRNAs (DELs) were enriched in terpenoid biosynthesis pathways, and 17 DELs were related to monoterpenoid and sesquiterpenoid biosynthesis. Moreover, we also constructed lncRNA–protein-coding gene (PCgenes) regulatory networks for terpenoid biosynthesis. This study supplies precious genetic resources to further research on lncRNA functions and their role in the regulatory mechanism of terpenoid biosynthesis.

Integrating RNA-seq with functional expression to analyze the regulation and characterization of genes involved in monoterpenoid biosynthesis in Nepeta tenuifolia Briq.

Nepeta tenuifolia Briq. (Lamiaceae) is a medicinal plant historically used in the East Asia region to treat cold and fever, and it is currently used as a clinically effective treatment for respiratory diseases. We previously found that monoterpenoids are the dominant volatile secondary metabolites in N. tenuifolia and their biosynthesis occurs in peltate glandular trichomes. To gain an insight into the molecular mechanisms underlying monoterpenoid biosynthesis in N. tenuifolia, we conducted transcriptome sequencing and examined the expression differences in monoterpene molecular pathway-related genes in different tissues and growth stages by qRT-RCR. In total, six p-menthane monoterpene biosynthetic genes in the (+)-menthone pathway were identified and cloned successfully based on transcriptome data. Moreover, the major constituents, including (+)-limonene, (−)-pulegone and (+)-menthone showed greater accumulation in the spikes than in other organs, such as the expression levels of related key enzyme genes. Additionally, the relative expression of pulegone reductase was the highest at 84 days, showing an inverse trend from (−)-pulegone relative content and leading to (+)-menthone accumulation in peltate glandular trichomes. Finished cloning of the gene for limonene 3-hydroxylase in N. tenuifolia (NtL3OH), heterologous expression in yeast, and in vitro assays were performed for functional characterization. Our study provides an important resource for further research of secondary metabolism of monoterpenes in peltate glandular trichomes of N. tenuifolia and other homologous species.

Metabolism and transcriptome profiling provides insight into the genes and transcription factors involved in monoterpene biosynthesis of borneol chemotype of Cinnamomum camphora induced by mechanical damage.

BackgroundThe borneol chemotype of Cinnamomum camphora (BCC), a monoterpene-rich woody plant species, is the sole source prescribed by the Chinese Pharmacopoeia for the production of natural D-borneol, a major monoterpene in BCC used for millennia as a topical analgesic in China. Nevertheless, the possible gene-regulatory roles of transcription factors (TFs) in BCC’s monoterpenoid biosynthesis remained unknown. Here, a joint analysis of the transcriptome and terpenoid metabolome of BCC induced by mechanical damage (MD) was used to comprehensively explore the interaction between TFs and terpene synthase (TPS) unigenes that might participate in monoterpene biosynthesis in BCC.ResultsGas chromatography–mass spectrometry analysis detected 14 monoterpenes and seven sesquiterpenes. All but two monoterpenes underwent a significantly increased accumulation after the MD treatment. RNA sequencing data revealed that 10 TPS, 82 MYB, 70 AP2/ERF, 38 BHLH, 31 WRKY, and 29 bZIP unigenes responded to the MD treatment. A correlation analysis revealed that three monoterpene synthase genes (CcTPS1, CcTPS3, CcTPS4) highly correlated with multiple monoterpenes, namely D-borneol, camphor, and bornyl acetate, which could be responsible for monoterpenoid biosynthesis in BCC. Furthermore, five WRKY, 15 MYB, 10 ERF/AP2, five bZIP, and two BHLH genes had strong, positive correlations with CcTPS1 or CcTPS4, judging by their high coefficient values (R2 > 0.8). The bioinformatics results were verified by quantitative real-time PCR.ConclusionThis study provides insight into the genes involved in the biosynthesis and regulation of monoterpene in BCC and thus provides a pool of candidate genes for future mechanistic analyses of how monoterpenes accumulate in BCC.

Population transcriptomic sequencing reveals allopatric divergence and local adaptation in Pseudotaxus chienii (Taxaceae)

BackgroundElucidating the effects of geography and selection on genetic variation is critical for understanding the relative importance of adaptation in driving differentiation and identifying the environmental factors underlying its occurrence. Adaptive genetic variation is common in tree species, especially widely distributed long-lived species. Pseudotaxus chienii can occupy diverse habitats with environmental heterogeneity and thus provides an ideal material for investigating the process of population adaptive evolution. Here, we characterize genetic and expression variation patterns and investigate adaptive genetic variation in P. chienii populations.ResultsWe generated population transcriptome data and identified 13,545 single nucleotide polymorphisms (SNPs) in 5037 unigenes across 108 individuals from 10 populations. We observed lower nucleotide diversity (π = 0.000701) among the 10 populations than observed in other gymnosperms. Significant negative correlations between expression diversity and nucleotide diversity in eight populations suggest that when the species adapts to the surrounding environment, gene expression and nucleotide diversity have a reciprocal relationship. Genetic structure analyses indicated that each distribution region contains a distinct genetic group, with high genetic differentiation among them due to geographical isolation and local adaptation. We used FST outlier, redundancy analysis, and latent factor mixed model methods to detect molecular signatures of local adaptation. We identified 244 associations between 164 outlier SNPs and 17 environmental variables. The mean temperature of the coldest quarter, soil Fe and Cu contents, precipitation of the driest month, and altitude were identified as the most important determinants of adaptive genetic variation. Most candidate unigenes with outlier signatures were related to abiotic and biotic stress responses, and the monoterpenoid biosynthesis and ubiquitin-mediated proteolysis KEGG pathways were significantly enriched in certain populations and deserve further attention in other long-lived trees.ConclusionsDespite the strong population structure in P. chienii, genomic data revealed signatures of divergent selection associated with environmental variables. Our research provides SNPs, candidate unigenes, and biological pathways related to environmental variables to facilitate elucidation of the genetic variation in P. chienii in relation to environmental adaptation. Our study provides a promising tool for population genomic analyses and insights into the molecular basis of local adaptation.

Genome-Wide Analysis of Potassium Channel Genes in Rice: Expression of the OsAKT and OsKAT Genes under Salt Stress.

Potassium (K+), as a vital element, is involved in regulating important cellular processes such as enzyme activity, cell turgor, and nutrient movement in plant cells, which affects plant growth and production. Potassium channels are involved in the transport and release of potassium in plant cells. In the current study, three OsKAT genes and two OsAKT genes, along with 11 nonredundant putative potassium channel genes in the rice genome, were characterized based on their physiochemical properties, protein structure, evolution, duplication, in silico gene expression, and protein–protein interactions. In addition, the expression patterns of OsAKTs and OsKATs were studied in root and shoot tissues under salt stress using real-time PCR in three rice cultivars. K+ channel genes were found to have diverse functions and structures, and OsKATs showed high genetic divergence from other K+ channel genes. Furthermore, the Ka/Ks ratios of duplicated gene pairs from the K+ channel gene family in rice suggested that these genes underwent purifying selection. Among the studied K+ channel proteins, OsKAT1 and OsAKT1 were identified as proteins with high potential N-glycosylation and phosphorylation sites, and LEU, VAL, SER, PRO, HIS, GLY, LYS, TYR, CYC, and ARG amino acids were predicted as the binding residues in the ligand-binding sites of K+ channel proteins. Regarding the coexpression network and KEGG ontology results, several metabolic pathways, including sugar metabolism, purine metabolism, carbon metabolism, glycerophospholipid metabolism, monoterpenoid biosynthesis, and folate biosynthesis, were recognized in the coexpression network of K+ channel proteins. Based on the available RNA-seq data, the K+ channel genes showed differential expression levels in rice tissues in response to biotic and abiotic stresses. In addition, the real-time PCR results revealed that OsAKTs and OsKATs are induced by salt stress in root and shoot tissues of rice cultivars, and OsKAT1 was identified as a key gene involved in the rice response to salt stress. In the present study, we found that the repression of OsAKTs, OsKAT2, and OsKAT2 in roots was related to salinity tolerance in rice. Our findings provide valuable insights for further structural and functional assays of K+ channel genes in rice.

The Volatilome: A Vital Piece of the Complete Soil Metabolome

Soils harbor complex biological processes intertwined with metabolic inputs from microbes and plants. Measuring the soil metabolome can reveal active metabolic pathways, providing insight into the presence of specific organisms and ecological interactions. A subset of the metabolome is volatile; however, current soil studies rarely consider volatile organic compounds (VOCs), contributing to biases in sample processing and metabolomic analytical techniques. Therefore, we hypothesize that overall, the volatility of detected compounds measured using current metabolomic analytical techniques will be lower than undetected compounds, a reflection of missed VOCs. To illustrate this, we examined a peatland metabolomic dataset collected using three common metabolomic analytical techniques: nuclear magnetic resonance (NMR), gas chromatography-mass spectroscopy (GC-MS), and fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR-MS). We mapped the compounds to three metabolic pathways (monoterpenoid biosynthesis, diterpenoid biosynthesis, and polycyclic aromatic hydrocarbon degradation), chosen for their activity in peatland ecosystems and involvement of VOCs. We estimated the volatility of the compounds by calculating relative volatility indices (RVIs), and as hypothesized, the average RVI of undetected compounds within each of our focal pathways was higher than detected compounds (p&amp;lt; 0.001). Moreover, higher RVI compounds were absent even in sub-pathways where lower RVI compounds were observed. Our findings suggest that typical soil metabolomic analytical techniques may overlook VOCs and leave missing links in metabolic pathways. To more completely represent the volatile fraction of the soil metabolome, we suggest that environmental scientists take into consideration these biases when designing and interpreting their data and/or add direct online measurement methods that capture the integral role of VOCs in soil systems.

Genome-wide identification of MIKC-type genes related to stamen and gynoecium development in Liriodendron

The organogenesis and development of reproductive organs, i.e., stamen and gynoecium, are important floral characteristics that are closely related to pollinators and reproductive fitness. As a genus from Magnoliaceae, Liriodendron has only two relict species: L. chinense and L. tulipifera. Despite the similar flower shapes of these species, their natural seed-setting rates differ significantly, implying interspecies difference in floral organogenesis and development. MADS-box genes, which participate in floral organogenesis and development, remain unexplored in Liriodendron. Here, to explore the interspecies difference in floral organogenesis and development and identify MADS-box genes in Liriodendron, we examined the stamen and gynoecium primordia of the two Liriodendron species by scanning electron microscopy combined with paraffin sectioning, and then collected two types of primordia for RNA-seq. A total of 12 libraries were constructed and 42,268 genes were identified, including 35,269 reference genes and 6,999 new genes. Monoterpenoid biosynthesis was enriched in L. tulipifera. Genome-wide analysis of 32 MADS-box genes was conducted, including phylogenetic trees, exon/intron structures, and conserved motif distributions. Twenty-six genes were anchored on 17 scaffolds, and six new genes had no location information. The expression profiles of MIKC-type genes via RT-qPCR acrossing six stamen and gynoecium developmental stages indicates that the PI-like, AG/STK-like, SEP-like, and SVP-like genes may contribute to the species-specific differentiation of the organogenesis and development of reproductive organs in Liriodendron. Our findings laid the groundwork for the future exploration of the mechanism underlying on the interspecific differences in reproductive organ development and fitness in Liriodendron.

Metabolomics characterizes the metabolic changes of Lonicerae Japonicae Flos under different salt stresses.

Salt stress affects the metabolic homeostasis of medicinal plants. However, medicinal plants are sessile organisms that cannot escape from salt stress. They acclimatize themselves to the stress by reprogramming their metabolic pathways. Lonicerae Japonicae Flos (LJF) with strong antioxidant activity is commonly used in traditional Chinese medicine, tea, and beverage. Nevertheless, the variation of integrated metabolites in LJF under different salt stresses remains unclear. In this study, High Performance Liquid Chromatography tandem triple time-of-flight mass spectrometry (HPLC- triple TOF-MS/MS) coupled with multivariate statistical analysis was applied to comparatively investigate the metabolites changes in LJF under different salt stress (0, 100, 200, 300 mM NaCl). Total 47 differential metabolites were screened from 79 metabolites identified in LJF under different salt stress. Low salt-treated group (100 mM NaCl) appeared to be the best group in terms of relative contents (peak areas) of the wide variety in bioactive components. Additionally, the phenylpropanoid pathway, monoterpenoid biosynthesis, glycolysis, TCA cycle, and alkaloid biosynthesis were disturbed in all salt-stress LJF. The results showed that LJF metabolisms were dramatically induced under salt stress and the quality of LJF was better under low salt stress. The study provides novel insights into the quality assessment of LJF under salt stress and a beneficial framework of knowledge applied to improvement the medicinal value of LJF.

Transcriptional Analysis of Metabolic Pathways and Regulatory Mechanisms of Essential Oil Biosynthesis in the Leaves of Cinnamomum camphora (L.) Presl.

The roots, bark, and leaves of Cinnamomum camphora are rich in essential oils, which mainly comprised monoterpenes and sesquiterpenes. Although the essential oils obtained from C. camphora have been widely used in pharmaceutical, medicinal, perfume, and food industries, the molecular mechanisms underlying terpenoid biosynthesis are poorly understood. To address this lack of knowledge, we performed transcriptome analysis to investigate the key regulatory genes involved in terpenoid biosynthesis in C. camphora. High-oil-yield trees of linalool type and low-oil-yield trees were used to assemble a de novo transcriptome of C. camphora. A total of 121,285 unigenes were assembled, and the total length, average length, N50, and GC content of unigenes were 87,869,987, 724, 1,063, and 41.1%, respectively. Comparison of the transcriptome profiles of linalool-type C. camphora with trees of low oil yield resulted in a total of 3,689 differentially expressed unigenes, among them 31 candidate genes had annotations associated with metabolism of terpenoids and polyketides, including four in the monoterpenoid biosynthesis pathway and three in the terpenoid backbone biosynthesis pathway. Collectively, this genome-wide transcriptome provides a valuable tool for future identification of genes related to essential oil biosynthesis. Additionally, the identification of a cohort of genes in the biosynthetic pathways of terpenoids provides a theoretical basis for metabolic engineering of essential oils in C. camphora.

Physiological and Transcriptome Analysis of Sugar Beet Reveals Different Mechanisms of Response to Neutral Salt and Alkaline Salt Stresses.

The salinization and alkalization of soil are widespread environmental problems. Sugar beet (B. vulgaris L.) is a moderately salt tolerant glycophyte, but little is known about the different mechanisms of sugar beet response to salt and alkaline stresses. The aim of this study was to investigate the influence of neutral salt (NaCl:Na2SO4, 1:1) and alkaline salt (Na2CO3) treatment on physiological and transcriptome changes in sugar beet. We found that a low level of neutral salt (NaCl:Na2SO4; 1:1, Na+ 25 mM) or alkaline salt (Na2CO3, Na+ 25 mM) significantly enhanced total biomass, leaf area and photosynthesis indictors in sugar beet. Under a high concentration of alkaline salt (Na2CO3, Na+ 100 mM), the growth of plants was not significantly affected compared with the control. But a high level of neutral salt (NaCl: Na2SO4; 1:1, Na+ 100 mM) significantly inhibited plant growth and photosynthesis. Furthermore, sugar beet tends to synthesize higher levels of soluble sugar and reducing sugar to cope with high neutral salt stress, and more drastic changes in indole acetic acid (IAA) and abscisic acid (ABA) contents were detected. We used next-generation RNA-Seq technique to analyze transcriptional changes under neutral salt and alkaline salt treatment in sugar beet. Overall, 4,773 and 2,251 differentially expressed genes (DEGs) were identified in leaves and roots, respectively. Kyoto encyclopedia of genes and genomes (KEGG) analysis showed that genes involving cutin, suberine and wax biosynthesis, sesquiterpenoid and triterpenoid biosynthesis and flavonoid biosynthesis had simultaneously changed expression under low neutral salt or alkaline salt, so these genes may be related to stimulating sugar beet growth in both low salt treatments. Genes enriched in monoterpenoid biosynthesis, amino acids metabolism and starch and sucrose metabolism were specifically regulated to respond to the high alkaline salt. Meanwhile, compared with high alkaline salt, high neutral salt induced the expression change of genes involved in DNA replication, and decreased the expression of genes participating in cutin, suberine and wax biosynthesis, and linoleic acid metabolism. These results indicate the presence of different mechanisms responsible for sugar beet responses to neutral salt and alkaline salt stresses.

Tandem mass tag-based (TMT) quantitative proteomics analysis reveals the response of fine roots to drought stress in cotton (Gossypium hirsutum L.)

BackgroundCotton (Gossypium hirsutum L.) is one of the most important cash crops worldwide. Fine roots are the central part of the root system that contributes to plant water and nutrient uptake. However, the mechanisms underlying the response of cotton fine roots to soil drought remains unclear. To elucidate the proteomic changes in fine roots of cotton plants under drought stress, 70–75% and 40–45% soil relative water content treatments were imposed on control (CK) and drought stress (DS) groups, respectively. Then, tandem mass tags (TMT) technology was used to determine the proteome profiles of fine root tissue samples.ResultsDrought significantly decreased the value of average root diameter of cotton seedlings, whereas the total root length and the activities of antioxidases were increased. To study the molecular mechanisms underlying drought response further, the proteome differences between tissues under CK and DS treatments were compared pairwise at 0, 30, and 45 DAD (days after drought stress). In total, 118 differentially expressed proteins (DEPs) were up-regulated and 105 were down-regulated in the ‘DS30 versus CK30’ comparison; 662 DEPs were up-regulated, and 611 were down-regulated in the ‘DS45 versus CK45’ comparison. The functions of these DEPs were classified according to their pathways. Under early stage drought (30 DAD), some DEPs involved in the ‘Cutin, suberin, and wax synthesis’ pathway were up-regulated, while the down-regulated DEPs were mainly enriched within the ‘Monoterpenoid biosynthesis’ pathway. Forty-five days of soil drought had a greater impact on DEPs involved in metabolism. Many proteins involving ‘Carbohydrate metabolism,’ ‘Energy metabolism,’ ‘Fatty acid metabolism,’ ‘Amino acid metabolism,’ and ‘Secondary metabolite biosynthesis’ were identified as DEPs. Additionally, proteins related to ion transport, stress/defense, and phytohormones were also shown to play roles in determining the fine root growth of cotton plants under drought stress.ConclusionsOur study identified potential biological pathways and drought-responsive proteins related to stress/defense responses and plant hormone metabolism under drought stress. Collectively, our results provide new insights for further improving drought tolerance in cotton and other crops.

Effect of Thidiazuron on Terpene Volatile Constituents and Terpenoid Biosynthesis Pathway Gene Expression of Shine Muscat (Vitis labrusca × V. vinifera) Grape Berries.

Volatile compounds are considered to be essential for the flavor and aroma quality of grapes. Thidiazuron (TDZ) is a commonly used growth regulator in grape cultivation that stimulates larger berries and prevents fruit drop. This study was conducted to investigate the effect of TDZ on the production of aroma volatiles and to identify the key genes involved in the terpene biosynthesis pathways that are affected by this compound. Treatment with TDZ had a negative effect on the concentration of volatile compounds, especially on monoterpenes, which likely impacts the sensory characteristics of the fruit. The expression analysis of genes related to the monoterpenoid biosynthesis pathways confirmed that treatment with TDZ negatively regulated the key genes DXS1, DXS3, DXR, HDR, VvPNGer and VvPNlinNer1. Specifically, the expression levels of the aforementioned genes were down-regulated in almost all berry development stages in the TDZ-treated samples. The novel results from the present study can be used to aid in the development of food products which maintain the flavor quality and sensory characteristics of grape. Furthermore, these findings can provide the theoretical basis that can help to optimize the utilization of TDZ for the field production of grapes at a commercial scale.

Differential expression and functional analysis of two short-chain alcohol dehydrogenases/reductases in Hedychium coronarium

Abstract In this study, the full cDNA sequences of HcADH2 and HcADH3 were cloned from Hedychium coronarium. The amino acid sequences encoded by them contained three most conserved motifs of short-chain alcohol dehydrogenase (ADH), namely NAD+ binding domain, TGxxx[AG]xG and active site YxxxK. The highest similarity between two genes and ADH from other plants was 70%. Phylogenetic analysis showed that they belonged to a member of the short-chain dehydrogenases/reductases 110C subfamily, but they were distinctly clustered in different clades. Real-time polymerase chain reaction analyses showed that HcADH2 was specifically expressed in bract, and it was expressed higher in no-scented Hedychium forrestii than other Hedychium species, but was undetectable in Hedychium coccineum. HcADH3 was expressed higher in the lateral petal of the flower than in other vegetative organs, and it was expressed the most in H. coronarium that is the most scented among Hedychium species, and its expression levels peaked at the half opening stage. HcADH2 and HcADH3 had almost no significant expression in leaves, but HcADH2 was expressed in response to external stimuli. The mechanical injury and methyl jasmonate (MeJA) treatment could induce expression of HcADH2 in leaves, whereas HcADH3 could have an induced expression only by MeJA. The recombinant HcADH3 protein, but not HcADH2, expressed in Escherichia coli-catalysed conversion of geraniol into citral. It was speculated that HcADH3 had an induced expression in vegetative organ of H. coronarium and took part in monoterpenoid biosynthesis in H. coronarium flowers, but the role of HcADH2 is relevant only for defensive reactions.

Comparative transcriptomics characterized the distinct biosynthetic abilities of terpenoid and paeoniflorin biosynthesis in herbaceous peony strains.

The herbaceous peony (Paeonia lactiflora Pall.) is a perennial flowering plant of the Paeoniaceae species that is widely cultivated for medical and ornamental uses. The monoterpene glucoside paeoniflorin and its derivatives are the active compounds of the P. lactiflora roots. However, the gene regulation pathways associated with monoterpene and paeoniflorin biosynthesis in P. lactiflora are still unclear. Here, we selected three genotypes of P. lactiflora with distinct morphologic features and chemical compositions that were a result of long-term reproductive isolation. We performed an RNA-sequencing experiment to profile the transcriptome changes of the shoots and roots. Using de novo assembly analysis, we identified 36,264 unigenes, including 521 genes responsible for encoding transcription factors. We also identified 28,925 unigenes that were differentially expressed in different organs and/or genotypes. Pathway enrichment analysis showed that the P. lactiflora unigenes were significantly overrepresented in several secondary metabolite biosynthesis pathways. We identified and profiled 33 genes responsible for encoding the enzymescontrolling the major catalytic reactions in the terpenoid backbone and in monoterpenoid biosynthesis. Our study identified the candidate genes in the terpenoid biosynthesis pathways, providing useful information for metabolic engineering of P. lactiflora intended for pharmaceutical uses and facilitating the development of strategies to improve marker-assist P. lactiflora in the future.