Functional Characterization of PmDXR, a Critical Rate-Limiting Enzyme, for Turpentine Biosynthesis in Masson Pine (Pinus massoniana Lamb.).

As one of the largest and most diverse classes of specialized metabolites in plants, terpenoids (oprenoid compounds, a type of bio-based material) are widely used in the fields of medicine and light chemical products. They are the most important secondary metabolites in coniferous species and play an important role in the defense system of conifers. Terpene synthesis can be promoted by regulating the expressions of terpene synthase genes, and the terpene biosynthesis pathway has basically been clarified in Pinus massoniana, in which there are multiple rate-limiting enzymes and the rate-limiting steps are difficult to determine, so the terpene synthase gene regulation mechanism has become a hot spot in research. Herein, we amplified a PmDXR gene (GenBank accession no. MK969119.1) of the MEP pathway (methyl-erythritol 4-phosphate) from Pinus massoniana. The DXR enzyme activity and chlorophyll a, chlorophyll b and carotenoid contents of overexpressed Arabidopsis showed positive regulation. The PmDXR gene promoter was a tissue-specific promoter and can respond to ABA, MeJA and GA stresses to drive the expression of the GUS reporter gene in N. benthamiana. The DXR enzyme was identified as a key rate-limiting enzyme in the MEP pathway and an effective target for terpene synthesis regulation in coniferous species, which can further lay the theoretical foundation for the molecularly assisted selection of high-yielding lipid germplasm of P. massoniana, as well as provide help in the pathogenesis of pine wood nematode disease.

Genome-Wide Identification and Expression Analysis of YTH Gene Family for Abiotic Stress Regulation in Camellia chekiangoleosa.

N6-methyladenosine (m6A) is essential for RNA metabolism in cells. The YTH domain, conserved in the kingdom of Eukaryotes, acts as an m6A reader that binds m6A-containing RNA. In plants, the YTH domain is involved in plant hormone signaling, stress response regulation, RNA stability, translation, and differentiation. However, little is known about the YTH genes in tea-oil tree, which can produce edible oil with high nutritional value. This study aims to identify and characterize the YTH domains within the tea-oil tree (Camellia chekiangoleosa Hu) genome to predict their potential role in development and stress regulation. In this study, 10 members of the YTH family containing the YTH domain named CchYTH1-10 were identified from C. chekiangoleosa. Through analysis of their physical and chemical properties and prediction of subcellular localization, it is known that most family members are located in the nucleus and may have liquid-liquid phase separation. Analysis of cis-acting elements in the CchYTH promoter region revealed that these genes could be closely related to abiotic stress and hormones. The results of expression profiling show that the CchYTH genes were differentially expressed in different tissues, and their expression levels change under drought stress. Overall, these findings could provide a foundation for future research regarding CchYTHs in C. chekiangoleosa and enrich the world in terms of epigenetic mark m6A in forest trees.

Analysis on metabolic functions of rhizosphere microbial communities of Pinus massoniana provenances with different carbon storage by Biolog Eco microplates.

Introduction: Rhizosphere microorganisms are influenced by vegetation. Meanwhile, they respond to vegetation through their own changes, developing an interactive feedback system between microorganisms and vegetation. However, it is still unclear whether the functional diversity of rhizosphere soil microorganisms varies with different carbon storage levels and what factors affect the functional diversity of rhizosphere soil microorganisms. Methods: In this study, the Biolog-Eco microplate technique was used to analyze the metabolic diversity of carbon source of rhizosphere soil microorganisms from 6 Pinus massoniana provenances with three levels of high, medium and low carbon storage. Results: The results showed that the average well color development(AWCD) value of rhizosphere microorganisms was significantly positive correlated with carbon storage level of Pinus massoniana (p < 0.05). The AWCD value, Simpson and Shannon diversity of high carbon sequestrance provenances were 1.40 (144h incubation) 0.96 and 3.24, respectively, which were significantly higher (p < 0.05) than those of other P. massoniana provenances. The rhizosphere microbial AWCD, Shannon and Simpson diversity of the 6 provenances showed the same variation trend (SM>AY>QJ>SX>HF>SW). Similarly, microbial biomass carbon (MBC) content was positively correlated with carbon storage level, and there were significant differences among high, medium and low carbon storage provenances. The PCA results showed that the differences in the carbon source metabolism of rhizosphere microorganisms were mainly reflected in the utilization of amino acids, carboxylic acids and carbohydrates. Pearson correlation analysis showed that soil organic carbon (SOC), total nitrogen (TN) and pH were significantly correlated with rhizosphere AWCD (p < 0.05). Conclusion: Soil properties are important factors affecting rhizosphere microbial carbon source metabolism. The study confirmed that the microorganisms of high carbon storage provenances had relatively high carbon metabolic activity. Among them, the carbon metabolic activity of rhizosphere microorganisms of SM provenance was the highest, which was the preferred provenances in effective ecological service function.

Identification and Expression Patterns of WOX Transcription Factors under Abiotic Stresses in Pinus massoniana.

WUSCHEL-related homeobox (WOX) transcription factors (TFs) play a crucial role in regulating plant development and responding to various abiotic stresses. However, the members and functions of WOX proteins in Pinus massoniana remain unclear. In this study, a total of 11 WOX genes were identified, and bioinformatics methods were used for preliminary identification and analysis. The phylogenetic tree revealed that most PmWOXs were distributed in ancient and WUS clades, with only one member found in the intermediate clade. We selected four highly conserved WOX genes within plants for further expression analysis. These genes exhibited expressions across almost all tissues, while PmWOX2, PmWOX3, and PmWOX4 showed high expression levels in the callus, suggesting their potential involvement in specific functions during callus development. Expression patterns under different abiotic stresses indicated that PmWOXs could participate in resisting multiple stresses in P. massoniana. The identification and preliminary analysis of PmWOXs lay the foundation for further research on analyzing the resistance molecular mechanism of P. massoniana to abiotic stresses.

Microbial communities and functions changed in rhizosphere soil of Pinus massoniana provenances with different carbon storage.

Introduction: The average carbon storage of Pinus massoniana is much higher than the average carbon storage of Chinese forests, an important carbon sink tree species in subtropical regions of China. However, there are few studies on the differences in rhizosphere microorganisms of P. massoniana with different carbon storages. Methods: To clarify the relationships between plant carbon storage level, environmental parameters and microbial community structure, we identified three carbon storage levels from different P. massoniana provenances and collected rhizosphere soil samples. We determined chemical properties of soil, extracellular enzyme activity, and microbial community structures at different carbon storage levels and examined how soil factors affect rhizosphere microorganisms under different carbon storage levels. Results: The results revealed that soil organic carbon (SOC), nitrate nitrogen (NO3--N), ammonium nitrogen (NH4+-N) contents all increased with increasing carbon storage levels, while pH decreased accordingly. In contrast, the available phosphorus (AP) content did not change significantly. The soil AP content was within the range of 0.91 ~ 1.04 mg/kg. The microbial community structure of P. massoniana changed with different carbon storage, with Acidobacteria (44.27%), Proteobacteria (32.57%), and Actinobacteria (13.43%) being the dominant bacterial phyla and Basidiomycota (73.36%) and Ascomycota (24.64%) being the dominant fungal phyla across the three carbon storage levels. Soil fungi were more responsive to carbon storage than bacteria in P. massoniana. C/N, NH4+-N, NO3--N, and SOC were the main drivers (p < 0.05) of changes in rhizosphere microbial communities. Discussion: The results revealed that in the rhizosphere there were significant differences in soil carbon cycle and microorganism nutrient preferences at different carbon storages of P. massoniana provenance, which were significantly related to the changes in rhizosphere microbial community structure. Jiangxi Anyuan (AY) provenance is more suitable for the construction of high carbon storage plantation.

Transcriptome-Wide Identification of TCP Transcription Factor Family Members in Pinus massoniana and Their Expression in Regulation of Development and in Response to Stress.

Pinus massoniana is an important coniferous tree species for barren mountain afforestation with enormous ecological and economic significance. It has strong adaptability to the environment. TEOSINTE BRANCHED 1/CYCLOIDEA/PCF (TCP) transcription factors (TFs) play crucial roles in plant stress response, hormone signal transduction, and development processes. At present, TCP TFs have been widely studied in multiple plant species, but research in P. massoniana has not been carried out. In this study, 13 PmTCP TFs were identified from the transcriptomes of P. massoniana. The phylogenetic results revealed that these PmTCP members were divided into two categories: Class I and Class II. Each PmTCP TF contained a conserved TCP domain, and the conserved motif types and numbers were similar in the same subgroup. According to the transcriptional profiling analysis under drought stress conditions, it was found that seven PmTCP genes responded to drought treatment to varying degrees. The qRT-PCR results showed that the majority of PmTCP genes were significantly expressed in the needles and may play a role in the developmental stage. Meanwhile, the PmTCPs could respond to several stresses and hormone treatments at different levels, which may be important for stress resistance. In addition, PmTCP7 and PmTCP12 were nuclear localization proteins, and PmTCP7 was a transcriptional suppressor. These results will help to explore the regulatory factors related to the growth and development of P. massoniana, enhance its stress resistance, and lay the foundation for further exploration of the physiological effects on PmTCPs.

Genome-Wide Identification and Characterization of the YTH Domain-Containing RNA-Binding Protein Family in Liriodendron chinense.

N6-methyladenosine (m6A) is becoming one of the most important RNA modifications in plant growth and development, including defense, cell differentiation, and secondary metabolism. YT521-B homology (YTH) domain-containing RNA-binding proteins, identified as m6A readers in epitranscriptomics, could affect the fate of m6A-containing RNA by recognizing and binding the m6A site. Therefore, the identification and study of the YTH gene family in Liriodendron chinense (L. chinense) can provide a molecular basis for the study of the role of m6A in L. chinense, but studies on the YTH gene in L. chinense have not been reported. We identified nine putative YTH gene models in the L. chinense genome, which can be divided into DF subgroups and DC subgroups. Domain sequence analysis showed that the LcYTH protein had high sequence conservation. A LcYTH aromatic cage bag is composed of tryptophan and tryptophan (WWW). PrLDs were found in the protein results of YTH, suggesting that these genes may be involved in the process of liquid-liquid phase separation. LcYTH genes have different tissue expression patterns, but the expression of LcYTHDF2 is absolutely dominant in all tissues. In addition, the expression of the LcYTH genes is changed in response to ABA and MeJA. In this study, We identified and analyzed the expression pattern of LcYTH genes. Our results laid a foundation for further study of the function of the LcYTH gene and further genetic and functional analyses of m6A RNA modification in forest trees.

Cloning of PmMYB6 in Pinus massoniana and an Analysis of Its Function.

Phenylpropanoids are crucial for the growth and development of plants and their interaction with the environment. As key transcriptional regulators of plant growth and development, MYB-like transcription factors play a vital role in the biosynthesis of phenylpropanoid metabolites. In this study, we functionally characterized PmMYB6, a Pinus massoniana gene that encodes an R2R3-MYB transcription factor. It was confirmed by qPCR that PmMYB6 was highly expressed in the flowers, xylem, and phloem of P. massoniana. By overexpressing PmMYB6 in tobacco and poplar, we found that transgenic plants had enlarged xylem, increased content of lignin and flavonoids, and up-regulated expression of several enzyme genes of the phenylpropane metabolism pathway to different degrees. The above research results indicate that PmMYB6 is involved in the metabolic flux distribution of different branches of the phenylpropane metabolic pathway, and the results may provide clues for the regulation of metabolic fluxes between flavonoids and the lignin biosynthesis pathways of P. massoniana, as well as provide a basis for the molecular breeding of P. massoniana.

The Transcriptomic Analysis of the Response of Pinus massoniana to Drought Stress and a Functional Study on the ERF1 Transcription Factor.

Pinus massoniana is a major fast-growing timber tree species planted in arid areas of south China, which has a certain drought-resistant ability. However, severe drought and long-term water shortage limit its normal growth and development. Therefore, in this study, physiological indices, and the transcriptome sequencing and cloning of AP2/ERF transcription factor of P. massonsiana were determined to clarify its molecular mechanism of drought stress. The results showed that stomatal conductance (Gs) content was significantly decreased, and superoxide dismutase (SOD) activity, and malondialdehyde (MDA) and abscisic acid (ABA) content were significantly increased under drought stress. Transcriptomic analysis revealed that compared to the control, 9, 3550, and 4142 unigenes with differential expression were identified by comparing plants subjected to light, moderate or severe drought. AP2/ERF with high expression was screened out for cloning. To investigate the biological functions of ERF1, it was over-expressed in wild-type Populus davdianaand × P. bolleana via the leaf disc method. Under drought stress, compared to wild-type plants, ERF1 over-expressing poplar lines (OE) maintained a higher photosynthetic rate and growth, while the transpiration rate and stomatal conductance significantly decreased and water use efficiency was improved, indicating that drought tolerance was enhanced. This study provides an insight into the molecular mechanism of drought stress adaptation in P. massoniana.

Transcriptome-Wide Identification of the GRAS Transcription Factor Family in Pinus massoniana and Its Role in Regulating Development and Stress Response.

Pinus massoniana is a species used in afforestation and has high economic, ecological, and therapeutic significance. P. massoniana experiences a variety of biotic and abiotic stresses, and thus presents a suitable model for studying how woody plants respond to such stress. Numerous families of transcription factors are involved in the research of stress resistance, with the GRAS family playing a significant role in plant development and stress response. Though GRASs have been well explored in various plant species, much research remains to be undertaken on the GRAS family in P. massoniana. In this study, 21 PmGRASs were identified in the P. massoniana transcriptome. P. massoniana and Arabidopsis thaliana phylogenetic analyses revealed that the PmGRAS family can be separated into nine subfamilies. The results of qRT-PCR and transcriptome analyses under various stress and hormone treatments reveal that PmGRASs, particularly PmGRAS9, PmGRAS10 and PmGRAS17, may be crucial for stress resistance. The majority of PmGRASs were significantly expressed in needles and may function at multiple locales and developmental stages, according to tissue-specific expression analyses. Furthermore, the DELLA subfamily members PmGRAS9 and PmGRAS17 were nuclear localization proteins, while PmGRAS9 demonstrated transcriptional activation activity in yeast. The results of this study will help explore the relevant factors regulating the development of P. massoniana, improve stress resistance and lay the foundation for further identification of the biological functions of PmGRASs.

Identification, Classification and Characterization of LBD Transcription Factor Family Genes in Pinus massoniana.

Transcription factors (TFs) are a class of proteins that play an important regulatory role in controlling the expression of plant target genes by interacting with downstream regulatory genes. The lateral organ boundary (LOB) structural domain (LBD) genes are a family of genes encoding plant-specific transcription factors that play important roles in regulating plant growth and development, nutrient metabolism, and environmental stresses. However, the LBD gene family has not been systematically identified in Pinus massoniana, one of the most important conifers in southern China. Therefore, in this study, we combined cell biology and bioinformatics approaches to identify the LBD gene family of P. massoniana by systematic gene structure and functional evolutionary analysis. We obtained 47 LBD gene family members, and all PmLBD members can be divided into two subfamilies, (Class I and Class II). By treating the plants with abiotic stress and growth hormone, etc., under qPCR-based analysis, we found that the expression of PmLBD genes was regulated by growth hormone and abiotic stress treatments, and thus this gene family in growth and development may be actively involved in plant growth and development and responses to adversity stress, etc. By subcellular localization analysis, PmLBD is a nuclear protein, and two of the genes, PmLBD44 and PmLBD45, were selected for functional characterization; secondly, yeast self-activation analysis showed that PmLBD44, PmLBD45, PmLBD46 and PmLBD47 had no self-activating activity. This study lays the foundation for an in-depth study of the role of the LBD gene family in other physiological activities of P. massoniana.

Transcriptome-Wide Identification of CCCH-Type Zinc Finger Proteins Family in Pinus massoniana and RR-TZF Proteins in Stress Response.

CCCH-type zinc finger proteins play an important role in multiple biotic and abiotic stresses. More and more reports about CCCH functions in plant development and stress responses have appeared over the past few years, focusing especially on tandem CCCH zinc finger proteins (TZFs). However, this has not been reported in Pinaceae. In this study, we identified 46 CCCH proteins, including 6 plant TZF members in Pinus massoniana, and performed bioinformatic analysis. According to RT-PCR analysis, we revealed the expression patterns of five RR-TZF genes under different abiotic stresses and hormone treatments. Meanwhile, tissue-specific expression analysis suggested that all genes were mainly expressed in needles. Additionally, RR-TZF genes showed transcriptional activation activity in yeast. The results in this study will be beneficial in improving the stress resistance of P. massoniana and facilitating further studies on the biological and molecular functions of CCCH zinc finger proteins.

Expression and promoter analysis of MEP pathway enzyme-encoding genes in Pinus massoniana Lamb.

The methylerythritol phosphate (MEP) pathway provides the universal basic blocks for the biosynthesis of terpenoids and plays a critical role in the growth and development of higher plants. Pinus massoniana is the most valuable oleoresin producer tree with an extensive terrestrial range. It has the potential to produce more oleoresin with commercial value, while being resistant to pine wood nematode (PWN) disease. For this study, eleven MEP pathway associated enzyme-encoding genes and ten promoters were isolated from P. massoniana. Three PmDXS and two PmHDR existed as multi-copy genes, whereas the other six genes existed as single copies. All eleven of these MEP enzymes exhibited chloroplast localization with transient expression. Most of the MEP genes showed higher expression in the needles, while PmDXS2, PmDXS3, and PmHDR1 had high expression in the roots. The expressions of a few MEP genes could be induced under exogenous elicitor conditions. The functional complementation in a dxs-mutant Escherichia coli strain showed the DXS enzymatic activities of the three PmDXSs. High throughput TAIL PCR was employed to obtain the upstream sequences of the genes encoding for enzymes in the MEP pathway, whereby abundant light responsive cis-elements and transcription factor (TF) binding sites were identified within the ten promoters. This study provides a theoretical basis for research on the functionality and transcriptional regulation of MEP enzymes, as well as a potential strategy for high-resin generation and improved genetic resistance in P. massoniana.

Characterization and Interaction Analysis of the Secondary Cell Wall Synthesis-Related Transcription Factor PmMYB7 in Pinus massoniana Lamb.

In vascular plants, the importance of R2R3-myeloblastosis (R2R3-MYB) transcription factors (TFs) in the formation of secondary cell walls (SCWs) has long been a controversial topic due to the lack of empirical evidence of an association between TFs and downstream target genes. Here, we found that the transcription factor PmMYB7, which belongs to the R2R3-MYB subfamily, is involved in lignin biosynthesis in Pinus massoniana. PmMYB7 was highly expressed in lignified tissues and upon abiotic stress. As a bait carrier, the PmMYB7 protein had no toxicity or autoactivation in the nucleus. Forty-seven proteins were screened from the P. massoniana yeast library. These proteins were predicted to be mainly involved in resistance, abiotic stress, cell wall biosynthesis, and cell development. We found that the PmMYB7 protein interacted with caffeoyl CoA 3-O-methyltransferase-2 (PmCCoAOMT2)-which is involved in lignin biosynthesis-but not with beta-1, 2-xylosyltransferase (PmXYXT1) yeast two-hybrid (Y2H) studies. Our in vivo coimmunoprecipitation (Co-IP) assay further showed that the PmMYB7 and PmCCoAOMT2 proteins could interact. Therefore, we concluded that PmMYB7 is an upstream TF that can interact with PmCCoAOMT2 in plant cells. These findings lay a foundation for further research on the function of PmMYB7, lignin biosynthesis and molecular breeding in P. massoniana.

Identification, classification, and characterization of AP2/ERF superfamily genes in Masson pine (Pinus massoniana Lamb.).

Transcription factors (TFs) play crucial regulatory roles in controlling the expression of the target genes in plants. APETALA2/Ethylene-responsive factors (AP2/ERF) are part of a large superfamily of plant-specific TFs whose members are involved in the control of plant metabolism, development and responses to various biotic and abiotic stresses. However, the AP2/ERF superfamily has not been identified systematically in Masson pine (Pinus massoniana), which is one of the most important conifer in southern China. Therefore, we performed systematic identification of the AP2/ERF superfamily using transcriptome sequencing data from Masson pine. In the current study, we obtained 88 members of the AP2/ERF superfamily. All PmAP2/ERF members could be classified into 3 main families, AP2 (7 members), RAV (7 members), ERF (73 members) families, and a soloist protein. Subcellular localization assays suggested that two members of PmAP2/ERF were nuclear proteins. Based on pine wood nematode (PWN) inoculated transcriptome and qPCR analysis, we found that many members of PmAP2/ERF could respond to PWN inoculation and PWN related treatment conditions in vitro. In general, members of the AP2/ERF superfamily play an important role in the response of Masson pine responds to PWN. Furthermore, the roles of the AP2/ERF superfamily in other physiological activities of Masson pine remain to be further studied.

Characterization and Function of the 1-Deoxy-D-xylose-5-Phosphate Synthase (DXS) Gene Related to Terpenoid Synthesis in Pinus massoniana.

In the methyl-D-erythritol-4-phosphate (MEP) pathway, 1-deoxy-D-xylose-5-phosphate synthase (DXS) is considered the key enzyme for the biosynthesis of terpenoids. In this study, PmDXS (MK970590) was isolated from Pinus massoniana. Bioinformatics analysis revealed homology of MK970590 with DXS proteins from other species. Relative expression analysis suggested that PmDXS expression was higher in roots than in other plant parts, and the treatment of P. massoniana seedlings with mechanical injury via 15% polyethylene glycol 6000, 10 mM H2O2, 50 µM ethephon (ETH), 10 mM methyl jasmonate (MeJA), and 1 mM salicylic acid (SA) resulted in an increased expression of PmDXS. pET28a-PmDXS was expressed in Escherichia coli TransB (DE3) cells, and stress analysis showed that the recombinant protein was involved in resistance to NaCl and drought stresses. The subcellular localization of PmDXS was in the chloroplast. We also cloned a full-length 1024 bp PmDXS promoter. GUS expression was observed in Nicotiana benthamiana roots, stems, and leaves. PmDXS overexpression significantly increased carotenoid, chlorophyll a, and chlorophyll b contents and DXS enzyme activity, suggesting that DXS is important in isoprenoid biosynthesis. This study provides a theoretical basis for molecular breeding for terpene synthesis regulation and resistance.

Transcriptome-Wide Identification of WRKY Transcription Factors and Their Expression Profiles under Different Types of Biological and Abiotic Stress in Pinus massoniana Lamb.

Pinus massoniana Lamb, an economically important conifer tree, is widely distributed in China. WRKY transcription factors (TFs) play important roles in plant growth and development, biological and abiotic stress. Nevertheless, there is little information about the WRKY genes in P. massoniana. By searching for conserved WRKY motifs in transcriptomic RNA sequencing data for P. massoniana, 31 sequences were identified as WRKY TFs. Then, phylogenetic and conserved motif analyses of the WRKY family in P. massoniana, Pinus taeda and Arabidopsis thaliana were used to classify WRKY genes. The expression patterns of six PmWRKY genes from different groups were determined using real-time quantitative PCR for 2-year-old P. massoniana seedings grown in their natural environment and challenged by phytohormones (salicylic acid, methyl jasmonate, or ethephon), abiotic stress (H2O2) and mechanical damage stress. As a result, the 31 PmWRKY genes identified were divided into three major groups and several subgroups based on structural and phylogenetic features. PmWRKY genes are regulated in response to abiotic stress and phytohormone treatment and may participate in signaling to improve plant stress resistance. Some PmWRKY genes behaved as predicted based on their homology with A. thaliana WRKY genes, but others showed divergent behavior. This systematic analysis lays the foundation for further identification of WRKY gene functions to aid further exploration of the functions and regulatory mechanisms of PmWRKY genes in biological and abiotic stress in P. massoniana.

Genome-wide investigation of WRKY transcription factors in sweet osmanthus and their potential regulation of aroma synthesis.

WRKY transcription factors, one of the largest transcription factor families, play important roles in regulating the synthesis of secondary metabolites. In sweet osmanthus (Osmanthus fragrans), the monoterpenes have been demonstrated as the most important volatile compounds, and the W-box, which is the cognate binding site of WRKY transcription factors, could be identified in most of the terpene-synthesis-related genes' promoters. However, the role of the WRKY family in terpene synthesis in sweet osmanthus has rarely been examined. In this study, 154 WRKY genes with conserved WRKY domain were identified and classified into three groups. The group II was further divided into five subgroups, and almost all members of IId contained a plant zinc cluster domain. Eight OfWRKYs (OfWRKY7/19/36/38/42/84/95/139) were screened from 20 OfWRKYs for their flower-specific expression patterns in different tissues. Simultaneously, the expression patterns of OfWRKYs and emission patterns of volatile compounds during the flowering process were determined and gas chromatography-mass spectrometry results showed that monoterpenes, such as linalool and ocimene, accounted for the highest proportion, contributing to the floral scent of sweet osmanthus in two cultivars. In addition, correlation analysis revealed the expression patterns of OfWRKYs (OfWRKY7/19/36/139) were each correlated with distinct monoterpenes (linalool, linalool derivatives, ocimene and ocimene derivatives). Subcellular localization analysis showed that p35S::GFP-OfWRKY7/38/95/139 were localized in the nucleus and OfWRKY139 had very strong transactivation activity. Collectively, the results indicated potential roles of OfWRKY139 and OfWRKYs with plant zinc cluster domain in regulating synthesis of aromatic compounds in sweet osmanthus, laying the foundation for use of OfWRKYs to improve the aroma of ornamental plants.

Transcriptional Analysis of Masson Pine (Pinus massoniana) under High CO2 Stress.

To explore the molecular mechanism of the response of Masson pine (Pinus massoniana), the main coniferous tree in southern China, to high CO2 stress, transcriptome sequencing was carried out to analyze the genome-wide responses of annual seedlings under different durations (0 h, 6 h, 12 h and 24 h) of high CO2 stress. The results showed that a total of 3080/1908, 3110/2115 and 2684/1483 genes were up-/down-regulated after 6 h, 12 h and 24 h of treatment, respectively, compared with control check group (CK, 0 h). Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that most of these differentially expressed genes (DEGs) were enriched in energy metabolism, carbohydrate synthesis, cell wall precursor synthesis and hormone regulation pathways. For energy metabolism, the expression of most genes involved in photosynthesis (including the light reaction and Calvin cycle) was generally inhibited, while the expression of genes related glycolysis, the tricarboxylic acid (TCA) cycle and PPP pathway was up-regulated. In addition, the increase in the CO2 concentration induced the up-regulation of gene expression in the sucrose synthesis pathway. Among all starch synthesis genes, GBSS (granule-bound starch synthase) had the highest expression level. On the other hand, during the synthesis of hemicellulose and pectin (cell wall precursor substances), the expression levels of GMD (GDP-mannose 4,6-dehydratase), MGP (Mannose-1-phosphate guanylyl transferase) and RHM (Rhamnose biosynthetic enzyme) were the highest, suggesting that the synthesis of the raw materials hemicellulose and pectin in Masson pine under stress were mainly supplied by GDP-Man, GDP-Fuc and UDP-Rha. Finally, stress inhibited gene expression in the ABA (Abscisic Acid) synthesis pathway and induced gene expression in the GA (Gibberellin), SA (Salicylic acid), BR(Brassinolide) and MeJA (Methyl Jasmonate) pathways. Stomatal switches were regulated by hormonal interactions. This experiment elaborated on the response and molecular mechanism of Masson pine to CO2 stress and aided in screening carbon sequestration genes for the corresponding molecular research of Masson pine in the future.

Characterization of Cellulose Synthesis in Plant Cells.

Cellulose is the most significant structural component of plant cell wall. Cellulose, polysaccharide containing repeated unbranched ß (1-4) D-glucose units, is synthesized at the plasma membrane by the cellulose synthase complex (CSC) from bacteria to plants. The CSC is involved in biosynthesis of cellulose microfibrils containing 18 cellulose synthase (CesA) proteins. Macrofibrils can be formed with side by side arrangement of microfibrils. In addition, beside CesA, various proteins like the KORRIGAN, sucrose synthase, cytoskeletal components, and COBRA-like proteins have been involved in cellulose biosynthesis. Understanding the mechanisms of cellulose biosynthesis is of great importance not only for improving wood production in economically important forest trees to mankind but also for plant development. This review article covers the current knowledge about the cellulose biosynthesis-related gene family.