uan Y

Water Deficit Affected Flavonoid Accumulation by Regulating Hormone Metabolism in Scutellaria baicalensis Georgi Roots

The content of flavonoids especially baicalin and baicalein determined the medical quality of Scutellaria baicalensis which is a Chinese traditional medicinal plant. Here, we investigated the mechanism responsible for the content and composition of flavonoids in S. baicalensis under water deficit condition. The transcription levels of several genes which are involved in flavonoid biosynthesis were stimulated by water deficit. Under water deficit condition, fifteen up-regulated proteins, three down-regulated proteins and other six proteins were detected by proteomic analysis. The identified proteins include three gibberellin (GA)- or indoleacetic acid (IAA)-related proteins. Decreased endogenous GAs level and increased IAA level were observed in leaves of S. baicalensis which was treated with water deficit. Exogenous application of GA or α-naphthalene acelic acid (NAA) to plants grown under water deficit conditions led to the increase of endogenous GAs and the decrease of IAA and flavonoids, respectively. When the synthesis pathway of GA or IAA in plants was inhibited by application with the inhibitors, flavonoid levels were recovered. These results indicate that water deficit affected flavonoid accumulation might through regulating hormone metabolism in S. baicalensis Georgi.

A Scutellaria baicalensis R2R3-MYB gene, SbMYB8, regulates flavonoid biosynthesis and improves drought stress tolerance in transgenic tobacco

R2R3-MYB proteins are involved in the primary and secondary metabolism, developmental processes and the responses to biotic and abiotic stresses. Little is known about the functions of R2R3-MYB proteins in Scutellaria baicalensis Georgi which is a traditional Chinese medicinal plants. In this study, the function of a S. baicalensis R2R3-MYB protein, SbMYB8, was investigated. SbMYB8 had similar expression pattern with SbC4H and SbCHS in ABA-treated S. baicalensis, indicating that SbMYB8 might be involved in the flavonoid metabolism. SbMYB8 protein could bind to the GmMYB92 BS3 sequence of SbCHS promoter region, regulating the expression of SbCHS. The SbMYB8 protein was localized to the nucleus where it activated transcription. The transgenic tobacco plants over-expressing SbMYB8 had higher caffeoylquinic acid contents, compared to that in wild type plants. Overexpression of SbMYB8 also changed the expression level of some flavonoid biosynthesis-related genes. It was found that overexpression of SbMYB8 can improve stress tolerance of transgenic plants, and can alter the activity and expression levels of some antioxidant enzymes. These results indicate that SbMYB8 plays important roles in flavonoid biosynthesis and stress tolerance of plant.

The Scutellaria baicalensis R2R3-MYB Transcription Factors Modulates Flavonoid Biosynthesis by Regulating GA Metabolism in Transgenic Tobacco Plants

R2R3-MYB proteins play role in plant development, response to biotic and abiotic stress, and regulation of primary and secondary metabolism. Little is known about the R2R3-MYB proteins in Scutellaria baicalensis which is an important Chinese medical plant. In this paper, nineteen putative SbMYB genes were identified from a S. baicalensis cDNA library, and eleven R2R3-MYBs were clustered into 5 subgroups according to phylogenetic reconstruction. In the S. baicalensis leaves which were sprayed with GA3, SbMYB2 and SbMYB7 had similar expression pattern with SbPALs, indicating that SbMYB2 and SbMYB7 might be involved in the flavonoid metabolism. Transactivation assay results showed that SbMYB2 and SbMYB7 can function as transcriptional activator. The expression of several flavonoid biosynthesis-related genes were induced or suppressed by overexpression of SbMYB2 or SbMYB7 in transgenic tobacco plants. Consistent with the change of the expression of NtDH29 and NtCHI, the contents of dicaffeoylspermidine and quercetin-3,7-O-diglucoside in SbMYB2-overexpressing or SbMYB7-overexpressing transgenic tobacco plants were decreased. The transcriptional level of NtUFGT in transgenic tobacco overexpressing SbMYB7 and the transcriptional level of NtHCT in SbMYB2-overexpressing tobacco plants were increased; however the application of GA3 inhibited the transcriptional level of these two genes. These results suggest that SbMYB2 and SbMYB7 might regulate the flavonoid biosynthesis through GA metabolism.

Genetic stability, active constituent, and pharmacoactivity of Salvia miltiorrhiza hairy roots and wild plant.

Salvia miltiorrhiza is an annual plant growing in China, Mongolia, Korea and some other Asian countries. The extract from S. miltiorrhiza roots has been used for supporting healthy cardiovascular and circulatory systems during the last decade. The active constituents of S. miltiorrhiza from different areas vary significantly, and the wild resources are overexploited. To adapt the demand for active constituents of S. miltiorrhiza against cardiovascular-related diseases, alternative materials need to be developed. The aim of the present work was to investigate the possibility of S. miltiorrhiza hairy roots as the alternative materials. The results showed that S. miltiorrhiza hairy roots are genetically stable. The contents of salvianolic acid B and tanshinone IIA, two main active constituents in hairy roots, determined by the assessment of combining fl ow cytometry and phytochemical analysis, are comparable to or significantly lower than in wild plant roots. The extract from S. miltiorrhiza hairy roots also had similar protection activity for hypoxia and reoxygenation injury in rat cardiac myocytes like that from wild plant roots. S. miltiorrhiza hairy roots may be alternative materials to obtain the drug or healthy food for cardiovascular-related diseases.

Overexpression of two R2R3-MYB genes from Scutellaria baicalensis induces phenylpropanoid accumulation and enhances oxidative stress resistance in transgenic tobacco

MYB proteins are involved in many significant physiological and biochemical processes, including regulation of primary and secondary metabolism, flavonoid biosynthesis, response to various biotic and abiotic stresses, hormone synthesis and signal transduction. The functions of R2R3-MYB proteins in Scutellaria baicalensis Georgi under abiotic stress, however, has not been elucidated. To study the molecular mechanism by which MYB2 and MYB7 respond to abiotic stress in S.xa0baicalensis, we analyzed the phenylpropanoid content, growth phenotype, antioxidant enzyme activity and flavonoid synthesis-associated gene expression in SbMYB2 or SbMYB7-overexpressing transgenic tobacco plants after treatment with NaCl, mannitol and abscisic acid (ABA). The transgenic tobacco showed a higher fresh weight than did the wild type (WT) tobacco. In contrast, antioxidant enzyme activity and flavonoid synthesis-related gene expression were markedly higher in WT tobacco after treatment with NaCl, mannitol and ABA, as compared to transgenic plants, This is likely because increased phenylpropanoid accumulation in transgenic tobacco plants played a central role in abiotic stress resistance. These results indicate that overexpression of SbMYB2 or SbMYB7 increased phenylpropanoid accumulation and enhanced NaCl, mannitol and ABA stresses tolerance in transgenic tobacco.

ITS2 Secondary Structure Improves Discrimination between Medicinal "Mu Tong" Species when Using DNA Barcoding.

DNA barcoding is a promising species identification method, but it has proved difficult to find a standardized DNA marker in plant. Although the ITS/ITS2 RNA transcript has been proposed as the core barcode for seed plants, it has been criticized for being too conserved in some species to provide enough information or too variable in some species to align it within the different taxa ranks. We selected 30 individuals, representing 16 species and four families, to explore whether ITS2 can successfully resolve species in terms of secondary structure. Secondary structure was predicted using Mfold software and sequence-structure was aligned by MARNA. RNAstat software transformed the secondary structures into 28 symbol code data for maximum parsimony (MP) analysis. The results showed that the ITS2 structures in our samples had a common four-helix folding type with some shared motifs. This conserved structure facilitated the alignment of ambiguous sequences from divergent families. The structure alignment yielded a MP tree, in which most topological relationships were congruent with the tree constructed using nucleotide sequence data. When the data was combined, we obtained a well-resolved and highly supported phylogeny, in which individuals of a same species were clustered together into a monophyletic group. As a result, the different species that are often referred to as the herb “Mu tong” were successfully identified using short fragments of 250 bp ITS2 sequences, together with their secondary structure. Thus our analysis strengthens the potential of ITS2 as a promising DNA barcode because it incorporates valuable secondary structure information that will help improve discrimination between species.

Flavonoids and antioxidative enzymes in temperature-challenged roots of Scutellaria baicalensis Georgi.

The active compounds in the roots of Scutellaria baicalensis Georgi, a traditional Chinese medicinal plant, are mainly fl avonoids which have anti-inflammatory, antitumour, and anti- HIV activity, respectively. The increasing annual average temperature has rendered the S. baicalensis plants grown in some ancient producing regions no longer suitable for their medicinal usage. Hydrogen peroxide plays an important role in root responses to abnormal temperature in S. baicalensis. Baicalin and baicalein and antioxidative enzymes were anticipated to detoxify H2O2 in S. baicalensis. Here, we show that abnormal temperatures (10 and 40 °C) decreased the content of flavonoids as compared with the normal temperature (30 °C), and the transcripts of UDP-glucuronate:baicalein 7-O-glucuronosyltransferase and β-glucuronidase involved in the interconversion between baicalin and baicalein were affected by the 40-°C treatment. High temperature also increased the activities of catalase and peroxidase. Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that the transcript levels of peroxidase 2, peroxidase 3, monodehydroascorbate reductase 2, and dehydroascorbate reductase were significantly increased under high-temperature conditions. The respective genes would be candidates for improvement of the adaptation of S. baicalensis plants to abnormal temperatures and for regulation of the contents of the active compounds.