Leaf Transcriptome Research Articles

Identification and characteristics of differentially expressed genes under UV-B stress in Gossypium hirsutum

ObjectiveThis study aimed to screen the differentially expressed genes (DEGs) of Gossypium hirsutum under UV-B stress and identify the significant pathways based on gene enrichment analysis results.MethodsIn this study, the allotetraploid crop G. hirsutum was used to examine changes in various physiological indexes under UV-B stress, and screened out all DEGs under UV-B stress (16 kJ m-2 d-1) based on six leaf transcriptomes. The main enrichment pathways of DEGs were analyzed according to gene annotation. Finally, we predicted the regulatory genes of phenylpropanoid pathway under UV-B stress by co-expression network analysis, and selected GhMYB4 for verification.ResultsGene annotation analysis revealed that DEGs were predominantly enriched in pathways related to photosynthesis and secondary metabolism. Further analysis revealed that UV-B stress impaired photosynthesis mainly by damaging photosystem II (PSII) and inhibiting electron transport, whereas G. hirsutum responded to UV-B stress by synthesizing secondary metabolites such as anthocyanins and lignin. We selected the regulatory genes GhMYB4 for verification. It was found to be an anthocyanin negative regulator in response to UV-B stress.ConclusionsUV-B stress impaired photosynthesis mainly by damaging photosystem II (PSII) and inhibiting electron transport, whereas G. hirsutum responded to UV-B stress by synthesizing secondary metabolites such as anthocyanins and lignin.

Systems analysis of long-term heat stress responses in the C4 grass Setaria viridis.

Many C4 plants are used as food and fodder crops and often display improved resource use efficiency compared to C3 plants. However, the response of C4 plants to future extreme conditions such as heatwaves is less understood. Here, Setaria viridis, an emerging C4 model grass, was grown under long-term high temperature stress for two weeks (42°C, compared to 28°C). This resulted in stunted growth, but surprisingly had little impact on leaf thickness, leaf area-based photosynthetic rates and bundle sheath leakiness. Dark respiration rates increased and there were major alterations in carbon and nitrogen metabolism in the heat-stressed plants. Abscisic acid and indole-acetic acid-amino acid conjugates accumulated in the heat-stressed plants, consistent with transcriptional changes. Leaf transcriptomics, proteomics and metabolomics analyses were carried out and mapped onto the metabolic pathways of photosynthesis, respiration, carbon/nitrogen metabolism and phytohormone biosynthesis and signaling. An in-depth analysis of correlations between transcripts and their corresponding proteins revealed strong differences between groups in the strengths and signs of correlations. Overall, many stress signaling pathways were upregulated, consistent with multiple signals leading to reduced plant growth. A systems-based model of the plant response to long-term heat stress is presented based on the oxidative stress, phytohormone and sugar signaling pathways.

Transcriptome-guided breeding for Paspalum notatum: producing apomictic hybrids with enhanced omega-3 content

Key messageTranscriptomics- and FAME-GC-MS-assisted apomixis breeding generated Paspalum notatum hybrids with clonal reproduction and increased α-linolenic acid content, offering the potential to enhance livestock product's nutritional quality and reduce methane emissions A low omega-6:omega-3 fatty acid ratio is considered an indicator of the nutritional impact of milk fat on human health. In ruminants, major long-chain fatty acids, such as linoleic acid (18:2, omega-6) and α-linolenic acid (18:3, omega-3), originate from dietary sources and reach the milk via the bloodstream. Since forages are the primary source of long-chain fatty acids for such animals, they are potential targets for improving milk lipid composition. Moreover, a high 18:3 content in their diet is associated with reduced methane emissions during grazing. This work aimed to develop genotypes of the forage grass Paspalum notatum with high leaf 18:3 content and the ability for clonal reproduction via seeds (apomixis). We assembled diploid and polyploid Paspalum notatum leaf transcriptomes and recovered sequences of two metabolism genes associated with the establishment of lipid profiles, namely SUGAR-DEPENDENT 1 (SDP1) and PEROXISOMAL ABC TRANSPORTER 1 (PXA1). Primers were designed to amplify all expressed paralogs in leaves. qPCR was used to analyse SDP1 and PXA1 expression in seven divergent genotypes. Reduced levels of SDP1 and PXA1 were found in the polyploid sexual genotype Q4188. Fatty acid methyl esters/gas chromatography/mass spectrometry (FAME/GC/MS) assays confirmed an increased percentage of 18:3 in this genotype. Crosses between Q4188 and the obligate apomictic pollen donor Q4117 resulted in two apomictic F1 hybrids (JS9 and JS71) with reduced SDP1 and PXA1 levels, increased 18:3 content, and clonal maternal reproduction. These materials could enhance milk and meat quality while reducing greenhouse gas emissions during grazing.

Omics based approaches to decipher the leaf ionome and transcriptome changes in Solanum lycopersicum L. upon Tomato Brown Rugose Fruit Virus (ToBRFV) infection.

The Tomato Brown Rugose Fruit Virus (ToBRFV) is a pathogen that mostly affects plants from the Solanaceae family. This virus severely affects the yield of tomato (Solanum lycopersicum L.) plants, thus creating an urgent need to research the basis of resistance to manage the disease. To understand the molecular basis of resistance, we employed omics-based approaches involving leaf ionomics and transcriptomics to help us decipher the interaction between elemental and nutritional composition and investigate its effect on the gene expression profile upon the ToBRFV infection in tomatoes. Ionomics was used to investigate the accumulation of trace elements in leaves, showcasing that the plants resistant to the virus had significantly higher concentrations of iron (p-value = 0.039) and nickel (p-value = 0.042) than the susceptible ones. By correlating these findings with transcriptomics, we identified some key genes involved in iron homeostasis and abscisic acid pathways, potentially responsible for conferring resistance against the pathogen. From the obtained list of differentially expressed genes, a panel of mutation profile was discovered with three important genes-Solyc02g068590.3.1 (K+ transporter), Solyc01g111890.3.1 (LRR), and Solyc02g061770.4.1 (Chitinase) showing persistent missense mutations. We ascertain the role of these genes and establish their association with plant resistance using genotyping assays in various tomato lines. The targeted selection of these genetic determinants can further enhance plant breeding and crop yield management strategies.

The Impact of Nutrient Solution Electrical Conductivity on Leaf Transcriptome Contributing to the Fruit Quality of Cucumber Grown in Coir Cultivation.

Soilless cultivation is increasingly utilized in supplying essential nutrients for greenhouse crops. However, the impact of coir cultivation under varying electrical conductivity (EC) conditions on cucumber growth and fruit quality, particularly through the regulation of gene expression during the vegetative stage, remains uncertain. In this study, we performed metabolic measurements on cucumber in both vegetative and reproductive stages under three different EC conditions and found metabolic products such as some primary metabolites (cellulose, many uncharged amino acids) and some secondary metabolites (rutin, cucurbitacin B) accumulated the most under EC of 5 dS·m-1. Next, we conducted transcriptome profiling in cucumber leaves, revealing that the function of genes significantly regulated by EC was associated with photosynthesis, many anabolic processes, and membrane transport. Finally, a set of genes contributed to metabolites related to the fruit quality of cucumber were identified by the Orthogonal Partial Least Squares (O2PLS) analysis, including genes involved in the biosynthesis of amino acids, polysaccharides, and many secondary metabolites. Taken together, these findings suggest that coir cultivation in greenhouses with moderate EC can induce a transcriptome-wide change in gene expression, thereby contributing to enhancing the abundance of metabolites associated with cucumber fruit quality.

Evolution of Duplicated Glutathione Metabolic Pathway in Gossypium hirsutum and Its Response to UV-B Stress.

Increasing levels of UV-B radiation caused by the greenhouse effect has become an emerging threat to crop health and yield. The glutathione (GSH) metabolic pathway is generally involved in plant stress responses through scavenging accumulated reactive oxygen species, and is therefore believed to play an essential role in enhancing plant tolerance to UV-B stress. However, the complex evolutionary details of this pathway in polyploid plants, especially under UV-B stress, remain largely unknown. Here, using the important allotetraploid crop, Gossypium hirsutum, as an example, we comprehensively investigated the composition and phylogenetic relationships of genes encoding 12 key structural enzymes in this pathway, and compared the expression changes of all the relevant genes under UV-B stress (16 kJ m-2 d-1) based on six leaf transcriptomes. Consequently, we identified 205 structural genes by genome-wide searching and predicted 98 potential regulatory genes under multiple stress conditions by co-expression network analysis. Furthermore, we revealed that 19 structural genes including 5 homoeologous pairs and 96 regulatory genes possessing 25 homoeologous pairs were reticulately correlated without homoeologous selection preference under UV-B stress. This result suggests a complex rewiring and reassignment between structural genes and their regulatory networks in the duplicated metabolic pathways of polyploid cotton. This study extends our understanding of the molecular dynamics of the GSH metabolic pathway in response to UV-B stress in G. hirsutum and, more broadly, in polyploid plants.

Transcriptome dynamics in developing leaves from C3 and C4 Flaveria species.

C4 species have evolved more than 60 times independently from C3 ancestors. This multiple and parallel evolution of the complex C4 trait suggests common underlying evolutionary mechanisms, which could be identified by comparative analysis of closely related C3 and C4 species. Efficient C4 function depends on a distinctive leaf anatomy that is characterised by enlarged, chloroplast-rich bundle sheath cells and narrow vein spacing. To elucidate the molecular mechanisms that generate the Kranz anatomy, we analysed a developmental series of leaves from the C4 plant Flaveria bidentis and the closely related C3 species Flaveria robusta by comparing anatomies and transcriptomes. Vascular density measurements of all nine leaf developmental stages identified three leaf anatomical zones whose proportions vary with respect to the developmental stage. We then deconvoluted the transcriptome datasets using non-negative matrix factorisation, which identified four distinct transcriptome patterns in the growing leaves of both species. By integrating the leaf anatomy and transcriptome data, we were able to correlate the different transcriptional profiles with different developmental zones in the leaves. These comparisons revealed an important role for auxin metabolism, in particular auxin homeostasis (conjugation and deconjugation), in establishing the high vein density typical of C4 species.

Functional characterization of seven terpene synthases from Ophiopogon japonicus via engineered Escherichia coli

Abstract Background Ophiopogon japonicus (L.f) Ker-Gawl. growing in Zhejiang is recognized as the Dao-di medicinal herb for the production of Ophiopogonis Radix. Borneol-7-O-[β-D-apiofuranosyl-(1→6)]-β-D-glucopyranoside, a prominent pharmacologically active compound, serves as a marker distinguishing O. japonicus in Zhejiang from those in other geographical areas. It is synthesized from borneol through glycosylation, with terpene synthase (TPS) being the critical enzyme catalyzing the conversion of terpene precursors into borneol. Objective The aim of the study was to define key genes involved in biosynthesis of borneol in O. japonicus. Methods The candidate terpene synthase genes were identified from the root and leaf transcriptome data of O. japonicus in Zhejiang and the functions of these enzymes were identified using engineered Escherichia coli. Results This study developed a rapid expression system for monoterpene and sesquiterpene synthases based on engineered E. coli. Seven terpene synthase genes (OjTPS1 to OjTPS7) were identified in different terpene synthase subfamilies, including 2 from TPS-a, 4 from TPS-b, and 1 from TPS-g. Biochemical analysis using an engineered system E. coli demonstrated that all the 7 terpene synthases produced monoterpenes, and OjTPS3, OjTPS5, and OjTPS6 also yielded sesquiterpenes. Conclusions These 7 terpene synthases produced 13 monoterpenes and 8 sesquiterpenes. Notably, OjTPS1 produced borneol establishing the groundwork for elucidating the biosynthetic pathways of borneol-7-O-[β-D-apiofuranosyl-(1→6)]-β-D-glucopyranoside and other volatile oil components.

Physiological and Transcriptomic Characterization of Rice Genotypes under Drought Stress

Drought is a primary abiotic stress that inhibits rice (Oryza sativa L.) growth and development, and during the reproductive stage it has a negative impact on the rice seed-setting rate. This research study examined two rice lines, La-96 (drought sensitive) and La-163 (drought resistant), for drought stress treatment (with soil moisture at 20% for 7 days) and control (normal irrigation and kept soil moisture ≥40%). To elucidate the photosynthesis and molecular mechanisms underlying drought tolerance in rice, leaf photosynthetic traits and transcriptome sequencing were used to compare differences between two contrasting recombinant inbred lines (RIL) during drought and subsequent recovery at the booting stage. The rice line La-96 showed a significant decrease in seed-setting rate after being treated for seven days’ drought stress (from 86.64% to 22.75%), while La-163 was slightly affected (from 89.04% to 79.33%). The photosynthetic activities of both lines significantly decreased under the drought treatment, and these traits of La-163 recovered to a comparable level with the control after three days of rewatering. The transcriptome of both lines in three treatments (the control, drought stress, and subsequent recovery) were tested, and a total of 16,051 genes were identified, among which 10,566 genes were differentially expressed in various treatments and rice lines. Comprehensive gene expression profiles revealed that the specifically identified DEGs were involved in the ribosome synthesis and the metabolic pathway of photosynthesis, starch, and sucrose metabolism. The DEGs that are activated and respond quickly, as seen during recovery in the tolerant rice line, may play essential roles in regulating subsequent growth and development. This study uncovered the molecular genetic pathways of drought tolerance and extended our understanding of the drought tolerance mechanisms and subsequent recovery regulation in rice.

Combined transcriptome and metabolome analysis identifies triterpenoid-induced defense responses in Myzus persicae Sülzer-infested peach.

Piercing/sucking insects such as green peach aphid (GPA) (Myzus persicae) cause direct damage by obtaining phloem nutrients and indirect damage by spreading plant viruses. To investigate the response of peach trees (Prunus persica) to aphids, the leaf transcriptome and metabolome of two genotypes with different sensitivities to GPA were studied. The gene expression of aphid-susceptible plants infested with aphids was similar to that of control plants, whereas the gene expression of aphid-resistant plants infested with aphids showed strong induced changes in gene expression compared with control plants. Furthermore, gene transcripts in defense-related pathways, including plant-pathogen interaction, MAPK signaling, and several metabolic pathways, were strongly enriched upon aphid infestation. Untargeted secondary metabolite profiling confirmed that aphid infestation induced larger changes in aphid-resistant than in aphid-susceptible peaches. Consistent with transcriptomic alterations, nine triterpenoids showed highly significant GPA-induced accumulation in aphid-resistant peaches, whereas triterpenoid abundance remained predominantly unchanged or undetected in aphid-susceptible peaches. Furthermore, some types of transcription factors (including WRKYs, ERFs, and NACs) were strongly induced upon GPA infestation in aphid-resistant, but not in aphid-susceptible peaches. These results suggested that the accumulation of specialized triterpenoids and the corresponding pathway transcripts may play a key role in peach GPA resistance.

Biogeographic, climatic, morphological, cytological and molecular data reveal a new diploid species from China in the genus Xanthocyparis (Cupressaceae)

AbstractThe genus Xanthocyparis was described in 2002 for a new conifer species X. vietnamensis from northern Vietnam, which became well‐known for its rarity. Recently, natural diploid populations were found in a small area of northern Guangxi, China, whereas material from Vietnam plus newly discovered populations from southern Guangxi were determined to be tetraploids. We integrated evidence from multiple data sources to perform a taxonomic evaluation of the two ploidy levels present in Xanthocyparis. Morphometric analyses detected statistically significant differences in cone and leaf characters, whereas microsatellite and transcriptome analyses revealed clear genetic divergence between diploid and tetraploid material. Furthermore, analysis of bioclimatic variables confirmed divergence in ecological niches. Gene trees from homologous sequences indicate that tetraploid material might have arisen via allopolyploidy from the extant diploid material and a third, possibly extinct lineage. Therefore, we recognize and describe a new diploid species from northern Guangxi China: Xanthocyparis guangxiensis.

Leaf transcriptomes from C3, C3-C4 intermediate, and C4Neurachne species give insights into C4 photosynthesis evolution.

The C4 photosynthetic pathway is hypothesized to have evolved from the ancestral C3 pathway through progressive changes in leaf anatomy and biochemistry with extant C3-C4 photosynthetic intermediate species representing phenotypes between species demonstrating full C3 and full C4 states. The Australian endemic genus Neurachne is the only known grass group that contains distinct, closely related species that carry out C3, C3-C4 intermediate, or C4 photosynthesis. To explore and understand the molecular mechanisms underlying C4 photosynthesis evolution in this genus, leaf transcriptomes were generated from two C3, three photosynthetic intermediate (proto-Kranz, C2-like, and C2), and two C4Neurachne species. The data were used to reconstruct phylogenetic relationships in Neurachne, which confirmed two independent C4 origins in the genus. Relative transcript abundances substantiated the photosynthetic phenotypes of individual species and highlighted transcriptional investment differences between species, including between the two C4 species. The data also revealed proteins potentially involved in C4 cycle intermediate transport and identified molecular mechanisms responsible for the evolution of C4-associated proteins in the genus.

Characterization of bZIP Transcription Factors in Transcriptome of Chrysanthemum mongolicum and Roles of CmbZIP9 in Drought Stress Resistance.

bZIP transcription factors play important roles in regulating plant development and stress responses. Although bZIPs have been identified in many plant species, there is little information on the bZIPs in Chrysanthemum. In this study, bZIP TFs were identified from the leaf transcriptome of C. mongolicum, a plant naturally tolerant to drought. A total of 28 full-length bZIP family members were identified from the leaf transcriptome of C. mongolicum and were divided into five subfamilies based on their phylogenetic relationships with the bZIPs from Arabidopsis. Ten conserved motifs were detected among the bZIP proteins of C. mongolicum. Subcellular localization assays revealed that most of the CmbZIPs were predicted to be localized in the nucleus. A novel bZIP gene, designated as CmbZIP9, was cloned based on a sequence of the data of the C. mongolicum transcriptome and was overexpressed in tobacco. The results indicated that the overexpression of CmbZIP9 reduced the malondialdehyde (MDA) content and increased the peroxidase (POD) and superoxide dismutase (SOD) activities as well as the expression levels of stress-related genes under drought stress, thus enhancing the drought tolerance of transgenic tobacco lines. These results provide a theoretical basis for further exploring the functions of the bZIP family genes and lay a foundation for stress resistance improvement in chrysanthemums in the future.

De novo transcriptome analysis identifies RpMYB1 as an activator of anthocyanin biosynthesis in Rehmannia piasezkii

Rehmannia piasezkii is a kind of medicinal plants, of the Orobanchaceae family, and well known for its large pink or purple corolla. However, no research on the molecular mechanism of flower color formation in R. piasezkii has been conducted so far. In this study, we investigated the transcriptome of root, stem, leaf and corollas of R. piasezkii using transcriptome sequencing technology and assembled 144,582 unigenes. A total of 58 anthocyanin biosynthetic genes were identified in the R. piasezkii transcriptome, fourteen of which were highly correlated with anthocyanin content, especially RpF3H2, RpDFR2, RpANS1, RpANS2 and RpUFGT. Totally, 35 MYB genes with FPKM values greater than 5 were identified in the R. piasezkii transcriptome, including an R2R3 MYB transcriptional factor RpMYB1, which belongs to subgroup 6 of the R2R3 MYB family. Agrobacterium-mediated transient expression of Nicotiana benthamiana revealed that overexpression of RpMYB1 could activate the expression of structural genes in anthocyanin synthesis pathway and promote the accumulation of anthocyanins in N. benthamiana leaves, indicating that RpMYB1 is a positive regulator of anthocyanin synthesis. Furthermore, combined transient overexpression of RpMYB1 with RpANS1, RpMYB1+RpANS1 with other structural genes all could further enhance the accumulation of anthocyanins in N. benthamiana leaves. Permanent overexpression of RpMYB1 in R. glutinosa promoted anthocyanin accumulation and expression levels of RgCHS, RgF3H, RgDFR and RgANS. Further evidence from dual-luciferase assay suggested that RpMYB1 could bind to the promoter of RpDFR2 and hence activating its expression. These findings provide insight into the molecular regulation in anthocyanin biosynthesis in R. piasezkii and provide valuable genetic resources for the genetic improvement of flower color.

Identification of full-length genes involved in the biosynthesis of β-caryophyllene and lupeol from the leaf transcriptome of Ayapana triplinervis.

β-Caryophyllene possesses potential anticancer properties against various cancers, including breast, colon, and lung cancer. Therefore, the essential oil of Ayapana triplinervis, which is rich in β-caryophyllene, can be a potential herbal remedy for treating cancer. However, molecular and genomic studies on A. triplinervis are still sparse. In this study, we obtained 14.7 Gb of RNA-Seq data from A. triplinervis leaf RNA and assembled 137 554 transcripts with an N50 value of 1437bp. We annotated 72 436 (52.7%) transcripts and mapped 10 640 transcripts to 156 biochemical pathways. Among them, 218 were related to terpenoid backbone biosynthesis, while 27 were linked to sesquiterpenoid and triterpenoid pathways. Ninety-four transcripts were annotated in the β-caryophyllene and lupeol pathways. From these transcripts, for the first time, we identified 25 full-length genes encoding all the 17 enzymes involved in β-caryophyllene biosynthesis and an additional five genes involved in lupeol biosynthesis. These genes will be useful for the metabolic engineering of β-caryophyllene and lupeol biosynthesis, not just in A. triplinervis but also in other species.

Effects of high-temperature stress on gene expression related to photosynthesis in two jujube (Ziziphus jujuba Mill.) varieties

ABSTRACT Elevated temperatures critically impact crop growth, development, and yield, with photosynthesis being the most temperature-sensitive physiological process in plants. This study focused on assessing the photosynthetic response and genetic adaptation of two different heat-resistant jujube varieties ‘Junzao’ (J) and ‘Fucuimi’ (F), to high-temperature stress (42°C Day/30°C Night). Comparative analyses of leaf photosynthetic indices, microstructural changes, and transcriptome sequencing were conducted. Results indicated superior high-temperature adaptability in F, evidenced by alterations in leaf stomatal behavior – particularly in J, where defense cells exhibited significant water loss, shrinkage, and reduced stomatal opening, alongside a marked increase in stomatal density. Through transcriptome sequencing 13,884 differentially expressed genes (DEGs) were identified, significantly enriched in pathways related to plant-pathogen interactions, amino acid biosynthesis, starch and sucrose metabolism, and carbohydrate metabolism. Key findings include the identification of photosynthetic pathway related DEGs and HSFA1s as central regulators of thermal morphogenesis and heat stress response. Revealing their upregulation in F and downregulation in J. The results indicate that these genes play a crucial role in improving heat tolerance in F. This study unveils critical photosynthetic genes involved in heat stress, providing a theoretical foundation for comprehending the molecular mechanisms underlying jujube heat tolerance.

Bridging fungal resistance and plant growth through constitutive overexpression of Thchit42 gene in Pelargonium graveolens.

Thchit42 constitutive expression for fungal resistance showed synchronisation with leaf augmentation and transcriptome analysis revealed the Longifolia and Zinc finger RICESLEEPER gene is responsible for plant growth and development. Pelargonium graveolens essential oil possesses significant attributes, known for perfumery and aromatherapy. However, optimal yield and propagation are predominantly hindered by biotic stress. All biotechnological approaches have yet to prove effective in addressing fungal resistance. The current study developed transgenic geranium bridging molecular mechanism of fungal resistance and plant growth by introducing cassette 35S::Thchit42. Furthermore, 120 independently putative transformed explants were regenerated on kanamycin fortified medium. Primarily transgenic lines were demonstrated peak pathogenicity and antifungal activity against formidable Colletotrichum gloeosporioides and Fusarium oxysporum. Additionally, phenotypic analysis revealed ~ 2fold increase in leaf size and ~ 2.1fold enhanced oil content. To elucidate the molecular mechanisms for genotypic cause, de novo transcriptional profiles were analyzed to indicate that the auxin-regulated longifolia gene is accountable for augmentation in leaf size, and zinc finger (ZF) RICESLEEPER attributes growth upregulation. Collectively, data provides valuable insights into unravelling the mechanism of Thchit42-mediated crosstalk between morphological and chemical alteration in transgenic plants. This knowledge might create novel opportunities to cultivate fungal-resistant geranium throughout all seasons to fulfil demand.

Identification and analysis of MAPK cascade gene families of Camellia oleifera and their roles in response to cold stress.

Low-temperature severely limits the growth and development of Camellia oleifera (C. oleifera). The mitogen-activated protein kinase (MAPK) cascade plays a key role in the response to cold stress. Our study aims to identify MAPK cascade genes in C. oleifera and reveal their roles in response to cold stress. In our study, we systematically identified and analyzed the MAPK cascade gene families of C. oleifera, including their physical and chemical properties, conserved motifs, and multiple sequence alignments. In addition, we characterized the interacting networks of MAPKK kinase (MAPKKK)-MAPK kinase (MAPKK)-MAPK in C. oleifera. The molecular mechanism of cold stress resistance of MAPK cascade genes in wild C. oleifera was analyzed by differential gene expression and real-time quantitative reverse transcription-PCR (qRT-PCR). In this study, 21 MAPKs, 4 MAPKKs and 55 MAPKKKs genes were identified in the leaf transcriptome of C. oleifera. According to the phylogenetic results, MAPKs were divided into 4 groups (A, B, C and D), MAPKKs were divided into 3 groups (A, B and D), and MAPKKKs were divided into 2 groups (MEKK and Raf). Motif analysis showed that the motifs in each subfamily were conserved, and most of the motifs in the same subfamily were basically the same. The protein interaction network based on Arabidopsis thaliana (A. thaliana) homologs revealed that MAPK, MAPKK, and MAPKKK genes were widely involved in C. oleifera growth and development and in responses to biotic and abiotic stresses. Gene expression analysis revealed that the CoMAPKKK5/CoMAPKKK43/CoMAPKKK49-CoMAPKK4-CoMAPK8 module may play a key role in the cold stress resistance of wild C. oleifera at a high-elevation site in Lu Mountain (LSG). This study can facilitate the mining and utilization of genetic resources of C. oleifera with low-temperature tolerance.

Comparative transcriptome and metabolite profiling reveal diverse pattern of CYP-TS gene expression during corosolic acid biosynthesis in Lagerstroemia speciosa (L.) Pers.

Extensive leaf transcriptome profiling and differential gene expression analysis of field grown and elicited shoot cultures of L. speciosa suggest that differential synthesis of CRA is mediated primarily by CYP and TS genes, showing functional diversity. Lagerstroemia speciosa L. is a tree species with medicinal and horticultural attributes. The pentacyclic triterpene, Corosolic acid (CRA) obtained from this species is widely used for the management of diabetes mellitus in traditional medicine. The high mercantile value of the compound and limited availability of innate resources entail exploration of alternative sources for CRA production. Metabolic pathway engineering for enhanced bioproduction of plant secondary metabolites is an attractive proposition for which, candidate genes in the pathway need to be identified and characterized. Therefore, in the present investigation, we focused on the identification of cytochrome P450 (CYP450) and oxidosqualene cyclases (OSC) genes and their differential expression during biosynthesis of CRA. The pattern of differential expression of these genes in the shoot cultures of L. speciosa, elicited with different epigenetic modifiers (azacytidine (AzaC), sodium butyrate (NaBu) and anacardic acid (AA)), was studied in comparison with field grown plant. Further, in vitro cultures with varying (low to high) concentrations of CRA were systematically assessed for the expression of CYP-TS and associated genes involved in CRA biosynthesis by transcriptome sequencing. The sequenced samples were de novo assembled into 180,290 transcripts of which, 92,983 transcripts were further annotated by UniProt. The results are collectively given in co-occurrence heat maps to identify the differentially expressed genes. The combined transcript and metabolite profiles along with RT-qPCR analysis resulted in the identification of CYP-TS genes with high sequence variation. Further, instances of concordant/discordant relation between CRA biosynthesis and CYP-TS gene expression were observed, indicating functional diversity in genes.

Integrated Metabolome and Transcriptome Analyses Provide New Insights into the Leaf Color Changes in Osmanthus fragrans cv. ‘Wucaigui’

Osmanthus fragrans, belonging to the family Oleaceae, is listed as one of the most important traditional ornamental plant species in China. A new cultivar O. fragrans ‘Wucaigui’ has a very diversified form in terms of leaf colors, in which the leaf color changes from red to yellow-green and finally to dark green. To understand the mechanisms involved in leaf color changes, metabolome and transcriptome studies were performed on leaves at different developmental stages. A total of 79 metabolites, two chlorophyll, 26 carotenoids, and 51 anthocyanins, were detected in the 6 different developmental stages. An orthogonal partial least squares discriminant analysis identified key metabolites at different developmental stages, including lutein, pelargonidin-3-O-(6-O-p-coumaroyl)-glucoside, neoxanthin, and α-carotene. A total of 48,837 genes were obtained by transcriptome sequencing, including 3295 novel genes. Using a weighted gene co-expression network analysis to study the correlations between key metabolites and differentially expressed genes, we determined the characteristic modules having the highest correlations with key metabolites and selected associated candidate genes. Five genes (OfSHOU4L, OfATL1B, OfUGE5 OfEIF1AX, and OfUGE3) were finally identified as hub genes using real-time fluorescence quantitative PCR. In addition, we proposed a model based on the changes in key metabolite contents and the network regulatory map during the changes in O. fragrans ‘Wucaigui’ leaf color. The positive regulation of OfUGE3 led to an increase in the lutein content, which resulted in the leaves changing from grayish brown to moderate brown; during the change from moderate brown to dark greenish-yellow, the positive regulation of three genes (OfHOU4L, OfATL1B, and OfUGE5) increased the content of pelargonidin-3-O-(6-O-p-coumaroyl)-glucoside and the red color of the leaves gradually faded to dark greenish-yellow and then to strong yellow-green; the positive regulation of OfEIF1AX increased the content of neoxanthin; the stages in which the color changed from strong yellow-green to yellow-green and then to moderate olive-green were positively regulated by OfUGE3, which resulted in higher α-carotene content. These findings provided new insights into the mechanisms underlying the processes involved in O. fragrans ‘Wucaigui’ leaf color changes at the metabolic and transcriptional levels. This work seeks to contribute to the development of artificial regulate and control technology in the breeding and production of O. fragrans and other ornamental plants.