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Tartary buckwheat is rich in flavonoids, which not only play an important role in the plant-environment interaction, but are also beneficial to human health. Rutin is a therapeutic flavonol which is massively accumulated in Tartary buckwheat. It has been demonstrated that transcription factors control rutin biosynthesis. However, the transcriptional regulatory network of rutin is not fully clear. In this study, through transcriptome and target metabolomics, we validated the role of FtMYB102 and FtbHLH4 TFs at the different developmental stages of Tartary buckwheat. The elevated accumulation of rutin in the sprout appears to be closely associated with the expression of FtMYB102 and FtbHLH4. Yeast two-hybrid, transient luciferase activity and co-immunoprecipitation demonstrated that FtMYB102 and FtbHLH4 can interact and form a transcriptional complex. Moreover, yeast one-hybrid showed that both FtMYB102 and FtbHLH4 directly bind to the promoter of chalcone isomerase (CHI), and they can coordinately induce CHI expression as shown by transient luciferase activity assay. Finally, we transferred FtMYB102 and FtbHLH4 into the hairy roots of Tartary buckwheat and found that they both can promote the accumulation of rutin. Our results indicate that FtMYB102 and FtbHLH4 can form a transcriptional complex by inducing CHI expression to coordinately promote the accumulation of rutin.

Tanshinone and phenolic acids are the most important active substances of Salvia miltiorrhiza, and the insight into their transcriptional regulatory mechanisms is an essential process to increase their content in vivo. SmMYB36 has been found to have important regulatory functions in the synthesis of tanshinone and phenolic acid; paradoxically, its mechanism of action in S. miltiorrhiza is not clear. Here, we demonstrated that SmMYB36 functions as a promoter of tanshinones accumulation and a suppressor of phenolic acids through the generation of SmMYB36 overexpressed and chimeric SmMYB36-SRDX (EAR repressive domain) repressor hairy roots in combination with transcriptomic-metabolomic analysis. SmMYB36 directly down-regulate the key enzyme gene of primary metabolism, SmGAPC, up-regulate the tanshinones biosynthesis branch genes SmDXS2, SmGGPPS1, SmCPS1 and down-regulate the phenolic acids biosynthesis branch enzyme gene, SmRAS. Meanwhile, SmERF6, a positive regulator of tanshinone synthesis activating SmCPS1, was up-regulated and SmERF115, a positive regulator of phenolic acid biosynthesis activating SmRAS, was down-regulated. Furthermore, the seven acidic amino acids at the C-terminus of SmMYB36 are required for both self-activating domain and activation of target gene expression. As a consequence, this study contributes to reveal the potential relevance of transcription factors synergistically regulating the biosynthesis of tanshinone and phenolic acid.

Tartary buckwheat is rich in flavonoids, which not only play an important role in plant-environment interaction, but are also beneficial to human health. Rutin is a therapeutic flavonol which is massively accumulated in Tartary buckwheat. It has been demonstrated that transcription factors control rutin biosynthesis. However, the transcriptional regulatory network of rutin is not fully clear. In this study, through transcriptome and target metabolomics, we validated the role of FtMYB102 and FtbHLH4 TFs at the different developmental stages of Tartary buckwheat. The elevated accumulation of rutin in the sprout appears to be closely associated with the expression of FtMYB102 and FtHLH4. Yeast two-hybrid, transient luciferase activity and co-immunoprecipitation demonstrated that FtMYB102 and FtbHLH4 can interact and form a transcriptional complex. Moreover, yeast one-hybrid showed that both FtMYB102 and FtbHLH4 directly bind to the promoter of chalcone isomerase (CHI), and they can coordinately induce CHI expression as shown by transient luciferase activity assay. Finally, we transferred the FtMYB102 and FtbHLH4 into the hairy roots of Tartary buckwheat and found that they both can promote the accumulation of rutin. Our results indicate that FtMYB102 and FtbHLH4 can form a transcriptional complex by inducing CHI expression to coordinately promote the accumulation of rutin.

Tartary buckwheat ( Fagopyrum tataricum) is rich in flavonoids, which not only play an important role in plant-environment interaction, but are also beneficial to human health. Rutin is a therapeutic flavonol which is massively accumulated in Tartary buckwheat. It has been demonstrated that transcription factors (TFs) control rutin biosynthesis. However, the transcriptional regulatory network of rutin is not fully clear. In this study, through transcriptome and target metabolomics profiling analysis together with homolog prediction, we identified and validated the role of FtMYB102 and FtbHLH4 TFs at the different developmental stages of Tartary buckwheat. The elevated accumulation of rutin in the sprout appears to be closely associated with the expression of FtMYB102 and FtHLH4. Yeast two-hybrid, transient luciferase activity and co-immunoprecipitation analysis demonstrated that FtMYB102 and FtbHLH4 can interact and form a transcriptional complex. Moreover, yeast one-hybrid showed that both FtMYB102 and FtbHLH4 directly bind to the promoter of chalcone isomerase ( CHI), and they can coordinately induce CHI expression as shown by transient luciferase activity assay. Finally, we transferred the FtMYB102 and FtbHLH4 into the hairy roots of Tartary buckwheat and found that they both can promote the accumulation of rutin. Our results indicate that FtMYB102 and FtbHLH4 can form a transcriptional complex by inducing CHI expression to coordinately promote the accumulation of rutin.

Jasmonate (JA) signaling regulates plant growth and development, biotic and abiotic stress tolerance, and primary and secondary metabolism biosynthesis. It is extensively modulated by JA-ZIM-domain (

Edible fungi resources have good application prospects in the research and development of food, medicine, and health products. Polyporus umbellatus (Pers.) Fries, as a precious edible and medicinal fungus, has long been used by Chinese medicine to treat urinary systems and related kidney diseases.In recent years, researchers have discovered and isolated a variety of active compounds from P. umbellatus. Modern phytochemical and pharmacological experiments showed that the crude extract of P. umbellatus had many biological functions and could be widely used in the fields of food, pharmaceutical and cosmetics. This paper summarizes the active components of P. umbellatus, through elaborating its mechanism of action, further clarify the action substances, in order to improve the utilization rate of P. umbellatus, promote the development and application of P. umbellatus in food, pharmaceutical and cosmetics industry.In this paper, the literatures related to P. umbellatus were summarized and classified by "China National Knowledge Instructure (CNKI)", "Google Scholar" and "Web of Science". Compared with other articles, this work systematically sorted out all the active substances with clear structures in P. umbellatus. On this basis, combined with the chemical composition of P. umbellatus, its functional efficacy was expounded, and the effects of different types of active substances in P. umbellatus were further presented.The main chemical constituents of P. umbellatus include polysaccharide and sterol, and the secondary compounds include fatty acids, phenols and other small molecules. These active substances endowed P. umbellatus anti-cancer, antibacterial, diuretic, antioxidant, enhance immune system, promote hair growth and other pharmacological activities, which has been verified many times in vivo and in vitro experiments.Modern in vitro or in vivo pharmacological experiments and clinical practice for the efficacy of P. umbellatus provides a strong support, and the separation of compounds in P. umbellatus has also deepened people's understanding of this traditional Chinese medicine, greatly promoted the development and application of P. umbellatus. However, the complex active substances of poring also hinder the research of P. umbellatus to some extent, and the mechanism of action and potential synergistic or antagonistic effect of the mixture of various active ingredients have not been clearly analyzed. How to use the bioactivity-guided separation strategy to identify more bioactive components and analyze the molecular mechanism of the main active components have become the main problems of P. umbellatus research, but also provides a direction for the further study of it.

Tartary buckwheat sprouts have a high nutritional value and are gluten-free, and polyphenols are their main active constituents. However, information regarding the active constituents' difference of Tartary buckwheat sprouts grown from seeds with different morphology, at different developmental stages and environments is limited. Here, we developed a LC-MS-based targeted metabolomics approach to analyze polyphenols (46 flavonoids and 6 anthraquinones) in 40 Tartary buckwheat sprouts varieties. Both flavonoids and anthraquinones contributed to significant differences in sprouts grown from seed with different color or shape. Twenty-seven differential compounds were all at a higher level in 3-day-old sprouts, and the fold change from 3-day-old to 8-day-old sprouts was 1.42-6.64. A total of 25 differential compounds were all significantly upregulated upon UV-B radiation, especially for epicatechin. This study is valuable not only for better breeding cultivars of Tartary buckwheat sprouts, but also assessing their metabolic quality.

Soil salinity significantly affects both Salvia miltiorrhiza growth and development as well as seed germination throughout field cultivation and production. The basic helix-loop-helix (bHLH) transcription factor (TF) MYC2 contributes significantly to plant stress resistance as a key regulator of the jasmonic acid signaling pathway. In transgenic S. miltiorrhiza hairy roots, SmMYC2 has been shown to promote the accumulation of tanshinone and salvianolic acid, but its role in S. miltiorrhiza of resistance to abiotic stress is unclear. Herein, we found methyl jasmonate (MeJA), NaCl, and PEG treatment all significantly increased SmMYC2 expression. In response to salt stress, SmMYC2 overexpression in yeast increased its rate of growth. Additionally, overexpression of SmMYC2 transgenic Arabidopsis thaliana and S. miltiorrhiza hairy root showed that it might improve salt resistance in transgenic plant. In particular, compared to WT, overexpression of SmMYC2 transgenic Arabidopsis had higher levels of three antioxidant enzymes (superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT)), proline (Pro) content, and ABA-dependent and ABA-independent genes expression. They also had lower levels of malondialdehyde (MDA) and reactive oxygen species (ROS) accumulation. What's more, overexpression of SmMYC2 increases the expression of flavonoid synthesis genes and the accumulation of related components in Arabidopsis. These findings imply that SmMYC2 functions as a positive regulator that regulates plant tolerance to salt through ABA-dependent and independent signaling pathways.

Casein kinase CK2 is a highly conserved serine/threonine protein kinase and exists in all eukaryotes. It has been demonstrated to be widely involved in the biological processes of plants. The CK2 holoenzyme is a heterotetramer consisting of two catalytic subunits (α and/or α') and two regulatory subunits (β). CK2 in plants is generally encoded by multiple genes, with monomeric and oligomeric forms present in the tissue. Various subunit genes of CK2 have been cloned and characterized from Arabidopsis thaliana, tobacco, maize, wheat, tomato, and other plants. This paper reviews the structural features of CK2, provides a clear classification of its physiological functions and mechanisms of action, and elaborates on the regulation of CK2 activity to provide a knowledge base for subsequent studies of CK2 in plants.

Salvia miltiorrhiza Bunge has been widely used in the treatment of cardiovascular and cerebrovascular diseases, due to the pharmacological action of its active components such as the tanshinones. Plasma membrane (PM) H+-ATPase plays key roles in numerous physiological processes in plants. However, little is known about the PM H+-ATPase gene family in S. miltiorrhiza (Sm). Here, nine PM H+-ATPase isoforms were identified and named SmPHA1–SmPHA9. Phylogenetic tree analysis showed that the genetic distance of SmPHAs was relatively far in the S. miltiorrhiza PM H+-ATPase family. Moreover, the transmembrane structures were rich in SmPHA protein. In addition, SmPHA4 was found to be highly expressed in roots and flowers. HPLC revealed that accumulation of dihydrotanshinone (DT), cryptotanshinone (CT), and tanshinone I (TI) was significantly reduced in the SmPHA4-OE lines but was increased in the SmPHA4-RNAi lines, ranging from 2.54 to 3.52, 3.77 to 6.33, and 0.35 to 0.74 mg/g, respectively, suggesting that SmPHA4 is a candidate regulator of tanshinone metabolites. Moreover, qRT-PCR confirmed that the expression of tanshinone biosynthetic-related key enzymes was also upregulated in the SmPHA4-RNAi lines. In summary, this study highlighted PM H+-ATPase function and provided new insights into regulatory candidate genes for modulating secondary metabolism biosynthesis in S. miltiorrhiza.

Phenolic acids are the major bioactive metabolites produced in Salvia miltiorrhiza, a traditional Chinese medicine called Danshen. Many phytohormone elicitor treatments induce phenolic acid biosynthesis, even though the underlying mechanism remains obscure. Expression pattern analysis showed that SmMAPK3 was highly expressed in leaves, and SmMAPK3 was significantly induced by salicylic acid (SA) and methyl jasmonate (JA). Bioinformatics analysis revealed that SmMAPK3 belongs to group A and contains a TEY motif in the activation loop together with three conserved regions (P-loop, C-loop and CD-domain). A previous study speculated that SmMAPK3 is likely a positive regulator in the biosynthesis of phenolic acids in S. miltiorrhiza. In this study, overexpression of SmMAPK3 increased phenolic acid biosynthetic gene expression and enhanced the accumulation of phenolic acids in S. miltiorrhiza plantlets. Yeast two-hybrid (Y2H) analysis and firefly luciferase complementation imaging (LCI) assays revealed that SmMAPKK2/4/5/7-SmMAPK3-SmJAZs form a cascade that regulates the accumulation of phenolic acids. In summary, this work deepens our understanding of the posttranscriptional regulatory mechanisms of phenolic acid biosynthesis and sheds new light on metabolic engineering in S. miltiorrhiza.

Additional file 2 :Figure S1. Conserved motifs in the products of genes belonging to the SmMAPK family, as predicted by MEME software. Figure S2. A proposed interaction network involving the products of genes belonging to the SmMAPK family. Figure S3. Pearson correlation coefficient of R value between the Ct value of the qRT-PCR results and the log2 RPKM values from the RNA-seq analysis. Pearson correlation coefficient of R values were visualized by TBtools.

Background: Flammulina velutipes (F. velutipes) is one of the most important mushrooms in Japan and China because of its potential medicinal and nutritional value. Ergosterol is an important precursor of vitamin D2, progesterone, hydrocortisone, brassinolide, and novel anticancer and anti-HIV drugs. One of the main methods for obtaining ergosterol is extraction of its natural products from fungi. However, because of the low production of ergosterol due to its inefficient biosynthesis, its approach cannot meet human needs. Therefore, increasing the ergosterol level in fungi is an important goal to be pursued. Although genes encoding key enzymes in the biosynthesis pathway of ergosterol have been identified in Saccharomyces cerevisiae, the mechanism and regulation of ergosterol biosynthesis in F. velutipes remain unclear, and are the focus of this study. Results: In this work, nine cDNA libraries produced from the three stages of F. velutipes were sequenced by using the Illumina HiSeqTM 4000 platform, resulting in at least 6.63 Gb of clean reads from each library. De novo sequence assemblers were used to generate 220,523 transcripts and 28,330 unigenes, which were annotated according to seven protein databases. Here, we combined transcriptomics and metabolomics approaches to investigate the biosynthesis of bioactive ergosterol from F. velutipes. The contents of 16 intermediates or metabolites from different stages (young fruiting body stage and mycelium stage) were found to be significantly different. Then, transcriptional profiling of the genes involved in ergosterol biosynthesis was carried out, and the transcripts of key genes involved in the metabolism were analysed. A total of 51 key unigenes (12 upregulated unigenes and 39 downregulated unigenes) were identified as differentially expressed. Furthermore, four genes (i.e., ERG10s, ERG1s, ERG11s and ERG26s) were identified as the most important genes that played roles in the regulation of the pathway flux towards ergosterol synthesis in F. velutipes. Combining transcriptomics and metabolomics, we explored the regulatory relationship between ergosterol biosynthesis genes and metabolites in F. velutipes. Conclusions: The data obtained in this work provide useful information for understanding the biosynthesis, metabolism and regulation of sterols in F. velutipes and for channelling the metabolic flux towards ergosterol at different growth stages.

Salicylic acid (SA) is an elicitor widely used to promote the synthesis of secondary metabolites. In order to determine the relevance between SA-induced pH changes and secondary metabolite synthesis, Salvia miltiorrhiza suspension cells were exposed to either SA, fusicoccin (FC), sodium orthovanadate (OVA), methylamine (ME) and their combination (FC+SA and ME+SA) for investigating the effect of cytosolic pH change on phenolic acids production, such as caffeic acid (CA), rosmarinic acid (RA), and salvianolic acid B (Sal B). The variations of cytosolic pH were sensed by using the fluorescent probe BCECF-AM, and combination with laser scanning confocal microscopic technique. Meanwhile, the genes expression of phenylalanine ammonia-lyase (PAL), tyrosine aminotransferase (TAT), and rosmarinic acid synthase (RAS) were determined using real time PCR. The contents of CA, RA, and Sal B were measured by high performance liquid chromatography. The results showed that SA decreased the cytosolic pH by inhibiting the activity of plasma membrane H+-ATPase. OVA induced cytosolic acidification too, while both FC and ME inhibited the acidification induced by SA. SA also up-regulated the genes expression of TAT, PAL, and RAS, and as a result enhanced the accumulation of those phenolic acids.

Tanshinones, the major bioactive components in Salvia miltiorrhiza Bunge (Danshen), are synthesized via the mevalonic acid (MVA) pathway or the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway and the downstream biosynthesis pathway. In this study, the bacterial component lipopolysaccharide (LPS) was utilized as a novel elicitor to induce the wild type hairy roots of S. miltiorrhiza. HPLC analysis revealed that LPS treatment resulted in a significant accumulation of cryptotanshinone (CT) and dihydrotanshinone I (DTI). qRT-PCR analysis confirmed that biosynthesis genes such as SmAACT and SmHMGS from the MVA pathway, SmDXS and SmHDR from the MEP pathway, and SmCPS, SmKSL and SmCYP76AH1 from the downstream pathway were markedly upregulated by LPS in a time-dependent manner. Furthermore, transcription factors SmWRKY1 and SmWRKY2, which can activate the expression of SmDXR, SmDXS and SmCPS, were also increased by LPS. Since Ca2+ signaling is essential for the LPS-triggered immune response, Ca2+ channel blocker LaCl3 and CaM antagonist W-7 were used to investigate the role of Ca2+ signaling in tanshinone biosynthesis. HPLC analysis demonstrated that both LaCl3 and W-7 diminished LPS-induced tanshinone accumulation. The downstream biosynthesis genes including SmCPS and SmCYP76AH1 were especially regulated by Ca2+ signaling. To summarize, LPS enhances tanshinone biosynthesis through SmWRKY1- and SmWRKY2-regulated pathways relying on Ca2+ signaling. Ca2+ signal transduction plays a key role in regulating tanshinone biosynthesis in S. miltiorrhiza.

Phenylpropanoid-derived compounds represent a diverse family of secondary metabolites that originate from phenylalanine. These compounds have roles in plant growth and development, and in defense against biotic and abiotic stress. Many of these compounds are also beneficial to human health and welfare. V-myb myeloblastosis viral oncogene homolog (MYB) proteins belong to a large family of transcription factors and are key regulators of the synthesis of phenylpropanoid-derived compounds. This review summarizes the current understanding of MYB proteins and their roles in the regulation of phenylpropanoid metabolism in plants.

Salvia miltiorrhiza (Chinese name: dānshēn) is used in traditional Chinese medicine for the treatment of cardiovascular and cerebrovascular diseases. The tanshinones represent the most important biological active class of compounds present in dānshēn extracts. They are synthesized via either the cytoplasmic mevalonate or the plastidial 2-C-methyl-D-erythritol-4-phosphate pathway. Here, we summarize recent discoveries regarding the mechanisms underlying tanshinone biosynthesis and how the process is regulated. Tanshinone accumulation in planta is affected by a range of elicitors and by the composition of the culture medium. Its production in hairy root cultures can be enhanced by the over-expression of genes encoding 1-deoxy-D-xylulose 5-phosphate synthase, 3-hydroxy-3-methylglutaryl-CoA reductase, geranylgeranyl diphosphate synthase and allene oxide cyclase. The pathway leading to the biosynthesis of the tanshinone precursors miltiradiene and ferruginol, has been engineered in yeast.

Our study found that except Novosphingobium resinovorum (B5) Salvia miltiorrhiza root endophytic bacteria Pseudomonas brassicacearum sub sp. neoaurantiaca (B1), Rhizobium radiobacter (B2), Pseudomonas thivervalensis (B3), Pseudomonas frederiksbergensis (B4) significantly improved the activity of key enzymes 3-hydroxy-3-methyglutary1-CoA reductase and 1-deoxy-d-xylulose-5-phosphate synthase in the biosynthetic pathway of tanshinones. Specifically, HMGR activity with B1 treatment increased 2.1-fold that of control, 1-deoxy-d-xylulose-5-phosphate synthase activity with B2 treatment increased 5.0-fold that of control, which caused a significant increase in tanshinone content in the hairy roots. The dihydrotanshinone I and cryptotanshinone content under B1 treatment increased 19.2-fold and 11.3-fold, respectively, and total tanshinone content increased 3.7-fold that of control. The five endophytic bacteria B1, B2, B3, B4 and B5 all significantly decreased phenylalanine ammonia-lyase and tyrosine aminotransferase activity in hairy roots, of which, B3 treatment decreased phenylalanine ammonia-lyase activity by 46.2 %, and B2 treatment decreased tyrosine aminotransferase activity by 44.7 % compared with the control. Each of the five endophytic bacteria decomposed rosmarinic acid and salvianolic acid B, which caused a significant decrease in rosmarinic acid and salvianolic acid B content in hairy roots, with B2 treatment decreasing rosmarinic acid and salvianolic acid B content by 94.5 and 89.0 %, respectively, compared with the control. The five endophytic bacteria also inhibited the growth of S. miltiorrhiza hairy roots, of which, B2 and B4 treatment decreased hairy root biomass by 55.2 and 51.3 %, respectively, compared with the control, while hairy roots promoted the growth of B4 and B5 and inhibited the growth of B1 and B3.

In higher plants, calcium is a ubiquitous second messenger in eukaryotic signal transduction cascades. The plant-specific calcium-dependent protein kinases (CDPKs) play important roles regulating downstream components of calcium signaling. We conducted a genome-wide analysis of maize (Zea mays) CDPKs and identified 35 CDPK genes. Maize CDPKs were found to be similar to their counterparts in rice in gene structure, GC content and subgroup classification. Divergence time estimation suggested that maize-rice orthologs were largely consistent with the time when these two species diverged from the last common ancestor. Semiquantitative RT-PCR revealed that the 29 of total 35 maize CDPK genes were expressed in all tissues, including root, stem, leaf, tassel, ear, and kernel. Our genomic and bioinformatics analyses will provide an important foundation for further functional dissection of the maize CDPK gene family.

More than 20 post-transcriptional gene silencing (PTGS) suppressors have been found since HC-Pro, the first gene silencing suppressor, was found in 1998. The silencing suppressor strongly suggested that gene silencing functions as natural defense mechanisms against viruses. It also represented a valuable tool for the dissection of the gene silencing pathway. We have used P1/HC-Pro RNA silencing suppressor activity to increase green fluorescent protein (GFP) expression in tobacco using an Agrobacterium-mediated transient expression system. P1/HC-Pro stimulated GFP-gene expression but not dsGFP-gene expression was shown by RT-PCR, Northern and Western blot analysis. Expression of the gene silencing suppressor and the target gene provided a new strategy of heterogeneous gene expressing in plants. It may be of commercial significance to produce foreign proteins using plant bioreactors.

Bacterium strain PJ3, isolated from wastewater and identified as Arthrobacter sp. bacterium based on its 16S rDNA gene, could use carbazole as the sole carbon, nitrogen and energy source. The genomic library of strain PJ3 was constructed and a positive clone JM109 (pUCW402) was screened out for the expression of dioxygenase by the ability to form yellow ring-fission product. A 2,3-dihydroxybiphenyl dioxygenase (23DHBD) gene of 933 bp was found in the 3360 bp exogenous fragment of pUCW402 by GenSCAN software and BLAST analysis. The phylogenetic analysis showed that 23DHBD from strain PJ3 formed a deep branch separate from a cluster containing most known 23DHBD in GenBank. Southern hybridization confirmed for the first time that the 23DHBD gene was from the genomic DNA of Arthrobacter sp. PJ3. In order to test the gene function, recombinant bacterium BL21 (pETW-8) was constructed to express 23DHBD. The expression level in BL21 (pETW-8) was highest compared with the recombinant bacteria JM109 (pUCW402) and strain PJ3. We observed that 23DHBD was not absolute specific. The enzyme activity was higher with 2,3-dihydroxybiphenyl as a substrate than with catechol. The substrate specificity assay suggested that 23DHBD was essential for cleavage of bi-cyclic aromatic compounds during the course of aromatic compound biodegradation in Arthrobacter sp. strain PJ3.

A number of members of the SAP (“stress-associated protein”) gene family have been implicated in the plant stress response. Here, a SAP gene has been isolated using PCR RACE from the perennial grass Leymus chinensis, a species which has reputation for ecological adaptability. The 17.6 kDa LcSAP product comprised 161 residues, including both an A20 domain and an AN1 domain, a feature of type I SAPs. Using a semi-quantitative RT-PCR assay to profile its transcription, it was shown that LcSAP was more strongly transcribed in the leaf than in the root under control conditions. The level of LcSAP transcription began to rise 6 h after the plant’s exposure to 400 mM NaCl, and the abundance of transcript remained stable for at least 24 h. Exposing the plant to 100 mM Na2CO3 also induced LcSAP transcription, but the abundance of SAP transcript faded after 6 h. When LcSAP was introduced into yeast cells, the transgenic cells grew better than wild type ones when the medium contained 1.4 M NaCl. The ability of LcSAP to respond to salinity stress in yeast suggests that it also makes a contribution to the stress tolerance shown by L. chinensis.

Salvia miltiorrhiza (Chinese name: dānshēn) is used in traditional Chinese medicine for the treatment of cardiovascular and cerebrovascular diseases. The tanshinones represent the most important biological active class of compounds present in dānshēn extracts. They are synthesized via either the cytoplasmic mevalonate or the plastidial 2-C-methyl-D-erythritol-4-phosphate pathway. Here, we summarize recent discoveries regarding the mechanisms underlying tanshinone biosynthesis and how the process is regulated. Tanshinone accumulation in planta is affected by a range of elicitors and by the composition of the culture medium. Its production in hairy root cultures can be enhanced by the over-expression of genes encoding 1-deoxy-D-xylulose 5-phosphate synthase, 3-hydroxy-3-methylglutaryl-CoA reductase, geranylgeranyl diphosphate synthase and allene oxide cyclase. The pathway leading to the biosynthesis of the tanshinone precursors miltiradiene and ferruginol, has been engineered in yeast.

A novel approach to virus-induced post-transcriptional gene silencing for studying the function of the ribulose bisphosphate carboxylase small subunit (rbcS) gene was established and optimized using potato virus X vector and Nicotiana benthamiana as experimental material. The analysis of silencing phenomena, transcriptional level, protein expression, and pigment measurement showed that the expression of the rbcS endogenous gene was inactivated by the expression of a 500-bp homologous cDNA fragment carried in the virus vector.

A Gram-positive, aerobic, rod-shaped, motile, endospore-forming bacterium, designated strain A12(T), was isolated from a saline and alkali soil samples in Baicheng City, western of Jilin Province, China. Growth occurred in 15-45 °C (optimum, 30 °C) and at pH 7.0-11.5 (optimum, pH 9.0) and in the presence of 0-10 % (w/v) NaCl [optimum, 1-3 % (w/v) NaCl]. Meso-DAP was present in the peptidoglycan. The predominant menaquinone was MK-7. The major polar lipid profile was phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidyl inositol-methyl and phosphotidylinositol dimannosid. The major fatty acid (10 % of total fatty acids) was anteiso-C15:0. DNA G + C content was 36.2 mol %. The level of 16S rRNA gene sequence similarity between strain A12(T) and other recognized species of the family was below 95.6 %. Phylogenetic analysis based on 16S rRNA gene sequence data indicated that the strain A12(T) fell with the family Bacillaceae and formed a distinct taxon. Based on physiological, chemotaxonomic and phylogenetic analyses, strain A12(T) was considered to represent a novel species of a new genus, for which the name Jilinibacillus soli gen. nov., sp. nov. was proposed. The type strain of Jilinibacillus soli was A12(T) (=GIMN1.014(T) = CCTCC M2011164(T) = KCTC 33417(T)).

Salvia miltiorrhiza Bunge (Lamiaceae) root, generally called Danshen, is an important herb in Chinese medicine widely used for treatment of various diseases. Phenolic acids in S. miltiorrhiza, as important effective compounds, have become a new research focus in plant secondary metabolism in recent years. This review summarizes the recent advances in the regulation of water-soluble phenolic acid biosynthesis in S. miltiorrhiza via regulators at molecular level, such as the phenylalanine ammonia-lyase gene (PAL), cinnamic acid 4-hydroxylase gene (C4H), 4-coumarate-CoA ligase gene (4CL), tyrosine aminotransferase gene (TAT), 4-hydroxyphenylpyruvate reductase gene (HPPR), 4-hydroxyphenylpyruvated dioxygenase gene (HPPD), hydroxycinnamoyl-CoA:hydroxyphenyllactate hydroxycinnamoyl transferase-like gene (RAS-like), and v-myb avian myeloblastosis viral oncogene homolog 4 gene (MYB4), and production of anthocyanin pigmentation 1 gene (AtPAP1), and via regulators at cell level, such as methyl jasmonate, salicylic acid, abscisic acid, polyamines, metal ions, hydrogen peroxide (H2O2), ultraviolet-B radiation, and yeast elicitor.

Tanshinones, a group of active ingredients in Salvia miltiorrhiza, are derived from at least two biosynthetic pathways, which are the mevalonate (MVA) pathway in the cytosol and the 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway in the plastids. Abscisic acid (ABA) and methyl jasmonate (MJ) are two well-known plant hormones induced by water stress. In this study, effects of polyethylene glycol (PEG), ABA and MJ on tanshinone production in S. miltiorrhiza hairy roots were investigated, and the role of MJ in PEG- and ABA-induced tanshinone production was further elucidated. The results showed that tanshinone production was significantly enhanced by treatments with PEG, ABA and MJ. The mRNA levels of 3-hydroxy-3-methylglutaryl co-enzyme A reductase (HMGR), 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR) and 1-deoxy-d-xylulose 5-phosphate synthase (DXS), as well as the enzyme activities of HMGR and DXS were stimulated by all three treatments. PEG and ABA triggered MJ accumulation. Effects of PEG and ABA on tanshinone production were completely abolished by the ABA biosynthesis inhibitor [tungstate (TUN)] and the MJ biosynthesis inhibitor [ibuprofen (IBU)], while effects of MJ were almost unaffected by TUN. In addition, MJ-induced tanshinone production was completely abolished by the MEP pathway inhibitor [fosmidomycin (FOS)], but was just partially arrested by the MVA pathway inhibitor [mevinolin (MEV)]. In conclusion, a signal transduction model was proposed that exogenous applications of PEG and ABA triggered endogenous MJ accumulation by activating ABA signaling pathway to stimulate tanshinone production, while exogenous MJ could directly induce tanshinone production mainly via the MEP pathway in S. miltiorrhiza hairy roots.

Plants of the genus Salvia produce various types of phenolic compounds and tanshinones which are effective for treatment of coronary heart disease. Salvia miltiorrhiza and S. castanea Diels f. tomentosa Stib are two important members of the genus. In this study, metabolic profiles and cDNA-AFLP analysis of four samples were employed to identify novel genes potentially involved in phenolic compounds and tanshinones biosynthesis, including the red roots from the two species and two tanshinone-free roots from S. miltiorrhiza. The results showed that the red roots of S. castanea Diels f. tomentosa Stib produced high contents of rosmarinic acid (21.77 mg/g) and tanshinone IIA (12.60 mg/g), but low content of salvianolic acid B (1.45 mg/g). The red roots of S. miltiorrhiza produced high content of salvianolic acid B (18.69 mg/g), while tanshinones accumulation in this sample was much less than that in S. castanea Diels f. tomentosa Stib. Tanshinones were not detected in the two tanshinone-free samples, which produced high contents of phenolic compounds. A cDNA-AFLP analysis with 128 primer pairs revealed that 2300 transcript derived fragments (TDFs) were differentially expressed among the four samples. About 323 TDFs were sequenced, of which 78 TDFs were annotated with known functions through BLASTX searching the Genbank database and 14 annotated TDFs were assigned into secondary metabolic pathways through searching the KEGGPATHWAY database. The quantitative real-time PCR analysis indicated that the expression of 9 TDFs was positively correlated with accumulation of phenolic compounds and tanshinones. These TDFs additionally showed coordinated transcriptional response with 6 previously-identified genes involved in biosynthesis of tanshinones and phenolic compounds in S. miltiorrhiza hairy roots treated with yeast extract. The sequence data in the present work not only provided us candidate genes involved in phenolic compounds and tanshinones biosynthesis but also gave us further insight into secondary metabolism in Salvia.

A novel plasma membrane intrinsic, LcPIP1, was isolated from Leymus chinensis using RACE method. The LcPIP1 has 288 amino acids with an estimated molecular mass of 30.6 kDa. Semi RT-PCR analysis indicated that the expression level of LcPIP1 was obviously higher in leaf than root. The LcPIP1 was also found to be induced by salt stress. In addition, transformed with the LcPIP1, Saccharomyces cerevisiae could increase tolerance to salt stress. These results indicate that the LcPIP1 gene seems to play a role in resistance against salt stress.

Spinach Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase) large (rbcL) and small (rbcS) subunits were separated by SDS-PAGE, and protein amount and purity were determined by Bradford assay. Polyclonal antibodies against rbcL and rbcS subunit were generated in female BALB/c mice and had no cross-reaction with each other. A total of 81 microg antigens were used and 0.3 ml anti-sera with titer of 1:5000 were yielded. The antibodies were also applicable to study rbcL and rbcS in tobacco plant Nicotiana benthamiana. Potato virus X vector pGR107 induced silencing of rbcS gene by Agrobacterium in Nicotiana benthamiana was performed. The expression level of rbcL and rbcS was lower in rbcS silenced plants than that in control plants as detected by the corresponding antibodies. This implied that the expression of rbcL was regulated by rbcS.

The aFGF (acidic fibroblast growth factor) plays an important role in morphogenesis, angiogenesis and wound healing and is therefore of potential medical interest. A DNA fragment encoding haFGF (human aFGF) has been cloned into the PVX (potato virus X)-based binary vector (pgR107) and transiently expressed in Nicotiana benthamiana (a wild Australian tobacco) by agroinfection. Approx. 1 week after agroinfection, the recombinant haFGF accumulated in the agroinfected plants reached up to 1% of the total soluble protein. haFGF was then purified on heparin-Sepharose CL-6B. The purified haFGF could stimulate the growth of NIH 3T3 cells, suggesting that the recombinant haFGF expressed via PVX viral vector in N. benthamiana was active biologically.