EuSHT Acts as a Hub Gene Involved in the Biosynthesis of 6-Hydroxyluteolin and Quercetin Induced by Salt Stress in Eucommia ulmoides

Salt stress inhibits plant growth and affects the biosynthesis of its secondary metabolites.Flavonoids are natural compounds that possess many important biological activities,playing a significant role in the medicinal activity of Eucommia ulmoides(E.ulmoides).To investigate the mechanism by which salt stress affects the biosynthesis of flavonoids in E.ulmoides,a comprehensive analysis of metabolomics and transcriptomics was conducted.The results indicated that salt stress led to the wilting and darkening of E.ulmoides leaves,accompanied by a decrease in chlorophyll levels,and significantly induced malondialdehyde(MDA)and relative electrical conductivity.During salt stress,most metabolites in the flavonoid biosynthesis pathway of E.ulmoides were upregulated,indicating that flavonoid biosynthesis is likely the main induced pathway under salt stress.Among them,secondary metabolites such as 6-Hydroxyluteolin and Quercetin are likely key metabolites induced by salt stress.The correlation analysis of transcriptomics and metabolomics revealed that EuSHT is a hub gene induced by salt stress,promoting the production of flavonoids such as 6-Hydroxyluteolin.The co-expression network showed a strong positive correlation between EuSHT and the biosynthesis of 6-Hydroxyluteolin and Quercetin,while it exhibited a negative correlation with Catechin biosynthesis.The branches leading to Luteolin and Dihydroquercetin are likely the main pathways for flavonoid compound biosynthesis in the plant stress response during salt stress.The results of this study provided a preliminary mechanism of secondary metabolites such as flavonoids in the medicinal plant E.ulmoides induced by salt stress and provided new theoretical support for discussing the mechanism of plant stress response.It also provided useful information for subsequent exploration of resistance genes in E.ulmoides.

Comprehensive Assessment of the Safety of Eucommia ulmoides Leaf Extract for Consumption as a Traditional Chinese Health Food

To ensure the export quality of Eucommia ulmoides leaf extract(ELE)and facilitate E.ulmoides leaf inclusion in the directory of traditional Chinese health foods,an overall safety assessment of ELE was performed,including genotoxicity and long-term toxicity,according to the national food safety standards of China.No variations in the reverse mutation number of the nominal bacterial strains were observed under ELE treatment in comparison with the solvent control.Additionally,the micronucleus rates of in vivo mammalian erythrocytes and in vitro mammalian cells under ELE treatment were equivalent to or significantly lower than those of the solvent control.The fold change in the trifluorothymidine resistance mutation frequency of the thymidine kinase gene under ELE treatment was less than three times in comparison with the solvent control,suggesting that ELE did not cause genotoxicity.Moreover,animal experiments showed that the growth performance of rats under ELE treatment was enhanced because the body weights of rats increased.No oxidative injury or inflammatory responses were induced and no histopathological lesions of tissues were detected under ELE treatment.In addition,plasma triglycerides and low-density lipoprotein cholesterol levels significantly decreased,and plasma high-density lipoprotein cholesterol levels significantly increased with ELE treatment,suggesting that ELE was health-promoting.Furthermore,moderate to excellent antimicrobial activities,a favorable anticancer capacity,and superior antioxidant abilities of ELE were found,implying ELE possesses good bioactivities.Therefore,we affirmed ELE is safe to consume as a traditional Chinese health food.

Cloning and Function Identification of a Phytoene Desaturase Gene from Eucommia ulmoides

The phytoene desaturase(PDS)encodes a crucial enzyme in the carotenoid biosynthesis pathway.Silencing or inhibiting PDS expression leads to the appearance of mottled,chlorosis,or albino leaves.In this study,the CDS sequence of EuPDS(Eucommia ulmoides Phytoene Desaturase)was first cloned and then PDS was silenced in Nicotiana benthamiana.Result showed the expression level of EuPDS in leaves was higher than that in the roots and stems.In N.benthamiana leaves,which were treated by Agrobacterium for 24 h,photo-bleaching was shown on the fresh leaves one week after injection and the transcript level of PDS was down-regulated during the period of emersion.This suggested that EuPDS could silence PDS of N.benthamiana,so as to cause the phenotype of leaf whitening.PDS is the main reporter gene involved in virus-induced gene silencing(VIGS).This study offered molecular evidence for identifying PDS gene involved in Carotenoid’s biosynthesis pathway and the regulation networks in E.ulmides.It also laid a useful foundation for study on leaf discoloration mechanism of other woody plants.

Overexpression ofβ-1,4-Glucanase Gene EuEG1 Improves Micrografting of Eucommia ulmoides

Adventitious root formation poses a major constraint on the tissue culture and genetic transformation of Eucommia ulmoides.Micrografting offers a new method for the transplantation of genetic transformation,and its success depends on the formation of graft unions.This study used transgenic rootless test-tube seedlings as scions and seedlings from seed as rootstocks during micrografting to avoid the rooting issues that occur during tissue culture and to investigate the role of the EuEG1 gene in the graft healing process.We found that the EuEG1 gene is a vital regulator of graft,and its overexpression contributes to the survival of Eucommia ulmoides micrografting.The EuEG1 gene transgenic plants(TP)used as scions for micrografting presented a significantly higher survival rate than the wild type(WT)and empty vector(EV)regenerated scions.During the grafting healing process,the expression of the EuEG1 gene was higher during the period of callus proliferation,suggesting that the EuEG1 gene was involved in the graft healing process.Histological observation revealed that more calluses tissue appeared at the junction of transgenic scions,and the connection with the rootstock was stronger,which benefits wound healing.These results provide new insights into Eucommia ulmoides micrografting and indicate that the EuEG1 gene can promote wound healing and improve the micrografting survival rate.

Role and Effects of Eucommia ulmoides Leaves in the Growth Process of Livestock and Poultry

Substitutes of feed antibiotics have been a key research topic in the new stage of animal husbandry.Chinese veterinary medicine refers to decoction pieces and their preparations processed from natural plants,animals,and minerals and used for animal disease prevention and improvement of animal production performance under the guidance of Chinese veterinary pharmacy theory.Eucommia ulmoides leaf extract has many active functional components such as chlorogenic acid,E.ulmoides polysaccharides and flavonoids,which have many biological properties such as antibacterial,antioxidant,immune-regulation,sugar ester-and bone metabolism-regulation effects.This paper explored active ingredients and biological properties of E.ulmoides leaf extract,as well as its role and effects in livestock and poultry breeding,providing a scientific basis for the use of E.ulmoides leaves in livestock and poultry breeding.

南方红壤区杜仲(Eucommia ulmoides)树干液流动态

采用根据热平衡原理设计的热扩散探针(Thermal dissipation sap flow velocity probe,TDP),于2004年7月到10月对南方红壤区的杜仲人工林的树干液流进行连续监测,结合所测定的相关因子,分析杜仲液流的变化规律及其与各因子的关系。结果表明,在不同月份,杜仲液流速率的日变化规律基本一致,呈单峰曲线,但是树干液流在启动时间、峰值出现时间上存在差异;从7月到10月份,杜仲树干液流速率逐渐增大,10月份液流速率达到1.818 g/(cm2.h);杜仲液流速率与太阳辐射、空气相对湿度、温度、风速等气象因子相关性显著,但是太阳辐射和空气相对湿度是影响杜仲液流的重要因子,其中7月到9月份,太阳辐射是主要影响因子,而10月份,空气相对湿度占主导地位;树干液流与胸径、冠层厚度以及胸径平方与树高之积呈现出显著的相关性。

Dynamic Changes in Distribution of Lignin and Hemicelluloses in Cell Walls During Differentiation of Secondary Xylem in Eucommia ulmoides

The dynamic changes in the distribution of lignin and hemicelluloses (xylans and xyloglucans) in cell walls during the differentiation of secondary xylem in Eucommia ulmoides Oliv. were studied by means of ultraviolet light microscopy and transmission electron microscopy combined with immunogold labelling. In the cambial zone and cell expansion zone, xyloglucans were localized both in the tangential and radial walls, but no xylans or lignin were found in these regions. With the formation of secondary wall S-1 layer, lignin occurred in the cell corners and middle lamella, while xylans appeared in S-1 layer, and xyloglucans were localized in the primary walls and middle lamella. In pace with the formation of secondary wall S-2 and S-3 layer, lignification extended to S-1, S-2 and S-3 layer in sequence, showing a patchy style of lignin deposition. Concurrently, xylans distributed in the whole secondary walls and xyloglucans, on the other hand, still localized in the primary walls and middle lamella. The results indicated that along with the formation and lignification of the secondary wall, great changes had taken place in the cell walls. Different parts of cell walls, such as cell corners, middle lamella, primary walls and various layers of secondary walls, had different kinds of hemicelluloses, which formed various cell wall architecture combined with lignin and other cell wall components.

Ultracytochemical Localization of ATPase During the Secondary Xylem Differentiation and Dedifferentiation in Eucommia ulmoides Trunk

The ultracytochemical localization of ATPase in the secondary xylem cells during their differentiation and dedifferentiation in the girdled Eucommia ulmoides Oliv. was carried out using a lead phosphate precipitation technique. Throughout the differentiation, which is a typical programmed cell death (PCD) process, ATPase deposits increased in the nucleus but decreased and progressively disappeared in the cell organelles. At the same time, the distribution of ATPase increased in the inner face of the cell wall and pits with cytoplasmic degeneration. The results demonstrated that the PCD was an energy dependent active process and was controlled by nuclear genes. On the other hand, the distribution of ATPase in the intercellular spaces increased with the formation of the new cambium resulted from the dedifferentiation of the secondary xylem cells after girdling. However, ATPase was not found in the nucleus of the dividing cells, suggesting that nutrients were transported through protoplast during differentiation, and through both protoplast and apoplast during dedifferentiation. Thus, the energy required in cell division was provided mainly by intercellular spaces. These findings indicate that the dynamic distribution of ATPase reflected which cell component was actively taking part in the cell metabolism at various stages of the plant development, and its distribution was associated with the physiological state of the cell. Based on the characteristic distributions of ATPase, the critical stage of cell differentiation and the relationship between the critical stage and dedifferentiation were discussed.

Effect of Regeneration After Girdling on Tree Growth in Eucommia ulmoides

Annual trunk diameters and tree heights were measured in July after initial_girdling and regirdling in Eucommia ulmoides Oliv. And the percentages of fallen leaves in all leaves and burst buds in all buds were estimated respectively on every autumn or spring. The results indicated that although the tree growth slowed down in the first year after girdling, but in the second year after girdling, the growth of the girdled trees tended to become normalized and up to the third year, it was similar to that of the normal trees. According to its developmental trait after girdling, the authors suggested that optimal girdling of the trees is in every 4 or 5 year intervals. With this method it was able to improve bark production and to protect the resource from extinction as well.

Programmed Cell Death During Secondary Xylem Differentiation in Eucommia ulmoides

Programmed cell death (PCD) during secondary xylem differentiation in Eucommia ulmoides Oliv. was examined using electron microscopy and by investigation of DNA fragmentation and degradation of caspase-like proteases (CLPs). DNA ladders were detected in developing secondary xylem by gel electrophoresis. DNA fragmentation was further confirmed by using the TdT-mediated dUTP nick-end labeling (TUNEL) method. Western blotting analysis showed that CLPs (caspase-8- and caspase-3-like proteases) and PARP (poly (ADP-ribose) polymerase) were degraded during secondary xylem differentiation. The results thus indicated that secondary xylem differentiation in E ulmoides was a typical process of PCD and the degradation of CLPs might be a constitutive PCD event during secondary xylem differentiation.

Ultrastructural Observation of Changes of Polysaccharide Grains in Dormant Shoots of Eucommia ulmoides

Cytochemical and ultrastructural studies indicated that compound spherical bodies observed near the sieve plate in the sieve tube members during the period of cambial dormancy in the shoots of Eucommia ulmoides Oliv. were polysaccharide bodies different from the polysaccharide grains in other parenchyma cells in shoots, and were similar to cell walls in their staining properties. The compound spherical bodies occurred in the sieve elements during the period of cambial rest and disappeared in the sieve elements during the period of cambial quiescence and activity.

Mechanism of Dormancy in the Buds and Cambium of Eucommia ulmoides

This paper describes changes in the structure of buds and vascular cambium as well as in protein content of one-year-old dormant branches of Eucommia ulmoides Oliv. in water-culture conditions. Results confirm the existence of 2 quiescences (Q1, Q2) and 1 rest (R) phases in this tree during the dormancy period. In the R time, the E. ulmoides cambium was unable to reactivate even though the tree was subjected to exogenous IAA, suitable temperature or required luminosity. Furthermore, pistillate trees entered the dormant phase earlier than staminate ones. The protein content in the bark during Q1 increased significantly ( P < 0.01), but drastically decreased in the R period, before rising up again at the onset of Q2 ( P < 0.01). Even though the change pattern of protein content was similar, its occurrence is much earlier in staminate than in pistillate plants. SDS-PAGE revealed a 'special' protein band of 11.8 kD in the transitional Q1-R-Q2 stages. This 'special' protein bands would be related to the cambial dormancy and to the regulation of the Q1-R-Q2 transition.

Comparative analysis of chemical components between barks and leaves of Eucommia ulmoides Oliver

The chemical components of the essential oils in the barks and leaves of Eucommia ulmoides Oliver were analyzed and compared by chromatograms and mass spectra technique, heuristic evolving latent projections (HELP), alternative moving window factor analysis (AMWFA) algorithms and normalization method based on the peak areas; the flavones in the barks and leaves of Eucommia ulmoides Oliver were separated on an ODS column by gradient elution carried out with the flow phase consisting of water, methanol and phosphoric acid (0.1%), and their contents were quantitatively determined by standard curve method and diode array detection (DAD) at 362 nm. The results show that 68 and 73 compounds respectively from essential oils of the barks and leaves of Eucommia ulmoides Oliver are identified, and there are 33 mutual compounds among 108 compounds determined. The total contents of these volatile components of the two samples possess 92.9% and 97.75% of the gross of the relevant essential oils, respectively; the contents of the rutin, quercetin and kaempferol in the barks and leaves of Eucommia ulmoides Oliver are 0.016 9, 0.003 6, 0.002 1 and 0.064 4, 0.030 2, 0.010 0 mg/g, respectively, and the determination recoveries are 95.2%-106.2%. The comparative analysis shows that for the barks and leaves of Eucommia ulmoides Oliver, there are significant differences in their components of the relevant essential oils and flavones.

Chemical constituents,biological functions and pharmacological effects for comprehensive utilization of Eucommia ulmoides Oliver

Eucommia ulmoides Oliver is a native plant and valuable tonic Chinese medicine in China with a long history,great economic value and comprehensive development potential.Traditionally,the comprehensive utilization rate of E.ulmoides Oliv.is still very low,only bark has been used as medicine and other parts of Eucommia ulmoides Oliv.cannot be fully utilized,even the leaves have been well utilized in food products in Japan in the past decades.In order to improve the comprehensive utilization efficiency of E.ulmoides Oliv.,in this review,we summarized the varieties and contents of main active compounds,biological functions and pharmacological effects in different parts of E.ulmoides Oliv.The findings suggest that other parts of E.ulmoides Oliv.could replace the bark of E.ulmoides Oliv.to some extent besides of their respective applications.The unique and extensive physiological functions between different parts of E.ulmoides Oliv.indicate that the comprehensive utilization of E.ulmoides Oliv.has a wide space to develop,which is also an effective way to protect E.ulmoides Oliv.resources and improve its the utilization rate.

Isolation of a novel antifungal peptide from the bark of Eucommia ulmoides Oliv

A novel Eucommia antifungal peptide, named EAFP3, was isolated from the bark of Eucommia ul- moides by NaC1 extract, gel filtration and reverse phase high performance liquid chromatography. The molecular mass of EAFP3 is 4157.3 Da, and its partial amino acid sequence is -. LYQQLIAGITLNK.-. EAFP3 exerts an inhibitory activity against Candida albicans in vitro and the drug concentration required for 50% growth inhibition （IC50） is 31.25μg/mL.

Determining the in vitro Anti-Aging Effect of the Characteristic Components from Eucommia ulmoides

To evaluate the potential anti-aging ability of Eucommia ulmoides,four characteristic components(chlorogenic acid,geniposidic acid,aucubin,quercetin)were selected to assess their effects on H_(2)O_(2)-induced oxidative damage model of human umbilical vein endothelial cell(HUVEC).Oxidative damage indexes,inflammatory factors,cell cycle,cell apoptosis,cell senescence,and their related proteins were analyzed by methyl thiazolyl tetrazolium(MTT)assay,enzyme-linked immunosorbent assay(ELISA),propidium iodide(PI)staining,annexin V-FITC/PI double staining,SAβ-galactosidase staining,and western blotting(WB).The results showed that H_(2)O_(2)-induced cell growth inhibition rate decreased as supplementation with characteristic components when compared to H_(2)O_(2) group.Meanwhile,the contents of antioxidant indexes(reactive oxygen species,lactate dehydrogenase,molondialdehyde,superoxide dismutase,glutathione),inflammatory factors(nuclear factor kappa-B,intercellular cell adhesion molecule-1,vascular cell adhesion protein 1),and functional factors(NO,Endothelin-1)in characteristic components treated groups improved if comparison with H_(2)O_(2) group,suggesting the characteristic components of E.ulmoides could alleviate H_(2)O_(2)-induced oxidative damage.Moreover,cell cycle,cell apoptosis,cell senescence,and their related proteins under characteristic components treatment exhibited a better effect than under H_(2)O_(2) treatment,implying the characteristic components could participate in anti-aging via multiple pathways.These results manifested that the characteristic components of E.ulmoides posses the capacity of anti-aging,which provided a basis for investigating the anti-aging ability of E.ulmoides itself.

The Extraction of Eucommia Ulmoides Oliv Polysaccharides and Its Protective Effect on Liver of Clophasphamidecy Injured Mice

Objective:To study the influenceof eucommia polysaccharide on the mice' liver damaged by clophasphamidecy (CY).Methods:Injecting CY build mice liver damage model,eucommia polysaccharide given different doses, measured blood serum ALT,AST and the liver's SOD,MDA. Results:After the injection CY,blood serum ALT,AST and the MDA of liver rise and the SOD of liver reduce comparedwith the blank group. The eucommia polysaccharide can improve these index.Conclusion:The Eucommia polysaccharide may protect the mice' liver damaged by CY.

Transcriptomic and Physiological Analyses of Pigment Accumulation in Eucommia ulmoides‘Hongye’

Eucommia ulmoides‘Hongye’is a new ornamental variety of E.ulmoides with excellent red or purple foliage.We found that E.ulmoides‘Hongye’exhibited a gradual change from green to red colour under light conditions.However,the colouring mechanism in the leaves of E.ulmoides‘Hongye’remains unclear.In this study,we compared the pigment content and leaf colour index of E.ulmoides‘Hongye’at five stages with those of E.ulmoides‘Xiaoye’,which was used as the control variety.The transcriptome sequencing data of the first-period(H1,green)and fifth-period(H5,red)leaves were also analysed and compared.The corresponding gene regulation in anthocyanin-related metabolic pathways was then analysed.Physiological results indicated that the contents of flavonoids and anthocyanins in red leaves(H5)were significantly higher than those in green leaves(H1),whereas the chlorophyll content in red leaves(H5)was lower than that in green leaves(H1).Moreover,the carotenoid content did not significantly differ between the two varieties.A transcriptome analysis identified 4240 differentially expressed genes(DEGs),and 20 of these genes were found to be involved in flavonoid and anthocyanin biosynthesis pathways.The results provide a reference for further study of the leaf colouration mechanism in E.ulmoides.

Cloning and Characterization of EuGID1 in Eucommia ulmoides Oliver

Gibberellic acid controlled the key developmental processes of the life cycle of landing plants,and regulated the growth and development of plants.In this study,a novel gibberellin receptor gene EuGID1 was obtained from Eucommia ulmoides Oliver.The cDNA of EuGID1 was 1556 bp,and the open reading frame was 1029 bp,which encoded 343 amino acids.EuGID1 had the homology sequence with the hormone-sensitive lipase family.Amino acid sequence alignment confirmed EuGID1 protein had the highest homology with the GID1 protein of Manihot esculenta.EuGID1 was located in the nucleus and cell membrane and had expression in four plant organs.Overexpression of EuGID1 in transgenic Arabidopsis plants promoted plant elongation and increased siliques yield.

杜仲Eucommia ulmoides的核型分析

以萌发种子的根尖为材料,对兰州地区引种的杜仲进行了体细胞染色体核型分析。结果显示:杜仲染色体较小,数目均为34条,与前人的研究结果一致;杜仲体细胞染色体核型有2类,第1类为2n=34=20m(2SAT)+10sm+4st,第2类为2n=34=20m(2SAT)+12sm+2st,在2类核型中均见到了随体染色体,随体均位于中部着丝点染色体的短臂上。2类核型中最长染色体与最短染色体的比值分别小于2,臂比值大于2∶1的染色体的比例在1%～50%之间,按照stebbins的核型分类标准应属2A核型,与前人的核型分析结果差异较大。