Search

Engineering anthocyanin biosynthesis in herbs could provide health‐promoting foods for improving human health. Rehmannia glutinosa is a popular medicinal herb in Asia, and was a health food for the emperors of the Han Dynasty (59 B.C.). In this study, we revealed the differences in anthocyanin composition and content between three Rehmannia species. On the 250, 235 and 206 identified MYBs in the respective species, six could regulate anthocyanin biosynthesis by activating the ANTHOCYANIDIN SYNTHASE (ANS) gene expression. Permanent overexpression of the Rehmannia MYB genes in tobacco strongly promoted anthocyanin content and expression levels of NtANS and other genes. A red appearance of leaves and tuberous/roots was observed, and the total anthocyanin content and the cyanidin‐3‐O‐glucoside content were significantly higher in the lines overexpressing RgMYB41, RgMYB42, and RgMYB43 from R. glutinosa, as well as RcMYB1 and RcMYB3 in R. chingii and RhMYB1 from R. henryi plants. Knocking out of RcMYB3 by CRISPR/Cas9 gene editing resulted in the discoloration of the R. chingii corolla lobes, and decreased the content of anthocyanin. R. glutinosa overexpressing RcMYB3 displayed a distinct purple color in the whole plants, and the antioxidant activity of the transgenic plants was significantly enhanced compared to WT. These results indicate that Rehmannia MYBs can be used to engineer anthocyanin biosynthesis in herbs to improve their additional value, such as increased antioxidant contents. [ABSTRACT FROM AUTHOR]

Tetrastigma hemsleyanum Diels et Gilg is a valuable medicinal herb. Chilling sensitivity is the major limiting factor of the artificial cultivation of the plant. Some key genes, such as ZEP and NCED genes of ABA biosynthesis, GA2ox, GA3ox, and GA20ox genes of GA biosynthesis, ACO genes of ET biosynthesis pathway were screened to be crucial in cold response. The response of ABA and ABA/GA1+3 to cold stress was prior to that of GA1+3, ZR, ABA/IAA, and ABA/ZR. The increasing changes in ABA/GA1+3 turned to a steep decline with the extension of stress time, which might be one factor contributing to cold sensitivity of T. hemsleyanum. The cold tolerance of T. hemsleyanum would be repressed by GA3 but enhanced by ABA when ABA orGA3 was used alone. Both the ABA-mediated promotion and GA-mediated repression of cold tolerance could be attenuated using a combination of the two phytohormones within 6 h. When the biosynthesis of ABA and GA were inhibited by FLU and PAC, respectively, the effects of GA and ABA treatment were reversed partially. In summary, we presented the first study of global hormone-regulated transcripts expression patterns in T. hemsleyanum. This study suggested that GA and ABA could work antagonistically to balance the cold stress responses. PAC, a GA biogenesis inhibitor, as well as exogenous ABA, might be potential plant growth regulators that can promote cold tolerance of T. hemsleyanum. The study also provided valuable hints in revealing the potential candidate genes that regulated cold tolerance of T. hemsleyanum. [ABSTRACT FROM AUTHOR]

Summary of main observation and conclusion: Collismycins (COLs) are antibiotics characterized by a 2,2′‐bipyridine (2,2′‐BP) core composed of a trisubstituted ring A and an unmodified ring B. The 2,2′‐BP core, which possesses metal‐chelating ability and plays key roles in various biological activities of COLs, is biosynthesized by a nonribosomal peptide synthetase (NRPS)‐polyketide synthase (PKS) hybrid machinery. The starter module of the NRPS‐PKS hybrid machinery consists of a type II peptidyl carrier protein (PCP) ColA1a and an adenylation protein ColA1b. We here report the functional characterization of ColA1a and ColA1b in vitro, confirming their functions in selection and loading of picolinic acid (PA), instead of normal amino acid substrates, as the origin of ring B in COLs. The 2.1 Å crystal structure of ColA1a was solved, revealing structural features including the additional helices α1a, α1b and missing helix α3, which may reflect unique interactions of ColA1a with other NRPS‐PKS proteins/domains or substrate. Primary and tertiary structural comparison of ColA1a with other PCPs revealed the structural basis for their typical α‐helical bundle, providing a better understanding of the structural flexibility of PCPs. These results facilitate the starter module engineering for the generation of COL derivatives with ring B modifications in the future. [ABSTRACT FROM AUTHOR]

Rehmannia glutinosa production is affected by replanting disease, in which autotoxic harm to plants is mediated by endogenous phenolic acids as allelopathic compounds found in root exudates. These phenolic acids are mostly phenylpropanoid products of plants' secondary metabolisms. The molecular mechanism of their biosynthesis and release has not been explored in R. glutinosa. P-coumarate-3-hydroxylase (C3H) is the second hydroxylase gene involved in the phenolic acid/phenylpropanoid biosynthesis pathways. C3Hs have been functionally characterized in several plants. However, limited information is available on the C3H gene in R. glutinosa. Here, we identified a putative RgC3H gene and predicted its potential function by in silico analysis and subcellular localization. Overexpression or repression of RgC3H in the transgenic R. glutinosa roots indicated that the gene was involved in allelopathic phenolic biosynthesis. Moreover, we found that these phenolic acid release amount of the transgenic R. glutinosa roots were altered, implying that RgC3H positively promotes their release via the molecular networks of the activated phenolic acid/phenylpropanoid pathways. This study revealed that RgC3H plays roles in the biosynthesis and release of allelopathic phenolic acids in R. glutinosa roots, laying a basis for further clarifying the molecular mechanism of the replanting disease development. [ABSTRACT FROM AUTHOR]

Rehmannia glutinosa L., a perennial plant of Scrophulariaceae, is one of the most commonly used herbs in traditional Chinese medicine (TCM) that have been widely cultivated in China. However, to date, the biosynthetic pathway of its two quality-control components, catalpol and acteoside, are only partially elucidated and the mechanism for their tissue-specific accumulation remains unknown. To facilitate the basic understanding of the key genes and transcriptional regulators involved in the biosynthesis of catalpol and acteoside, transcriptome sequencing of radial striation (RS) and non-radial striation (nRS) from four R. glutinosa cultivars was performed. A total of 715,158,202 (~107.27 Gb) high quality reads obtained using paired-end Illumina sequencing were de novo assembled into 150,405 transcripts. Functional annotation with multiple public databases identified 155 and 223 unigenes involved in catalpol and acteoside biosynthesis, together with 325 UGTs, and important transcription factor (TF) families. Comparative analysis of the transcriptomes identified 362 unigenes, found to be differentially expressed in all RS vs. nRS comparisons, with 143 upregulated unigenes, including those encoding enzymes of the catalpol and acteoside biosynthetic pathway, such as geranyl diphosphate synthase (RgGPPS), geraniol 8-hydroxylase (RgG10H), and phenylalanine ammonia-lyase (RgPAL). Other differentially expressed unigenes predicted to be related to catalpol and acteoside biosynthesis fall into UDP-dependent glycosyltransferases (UGTs), as well as transcription factors. In addition, 16 differentially expressed genes were selectively confirmed by real-time PCR. In conclusion, a large unigene dataset of R. glutinosa generated in the current study will serve as a resource for the identification of potential candidate genes for investigation of the tuberous root development and biosynthesis of active components. [ABSTRACT FROM AUTHOR]

Mirabilis himalaica (Edgew.) Heimerl is one of the most important genuine medicinal plants in Tibet, in which the special plateau habitat has been associated with its excellent medicinal quality and efficacy. However, the mechanisms by which environmental factors affect biosynthesis of secondary metabolic components remain unclear in this species. In this study, RNA sequencing and iTRAQ (isobaric Tags for Relative and Absolute Quantification) techniques were used to investigate the critical molecular "events" of rotenoid biosynthesis responding to UV-B radiation, a typical plateau ecological factor presented in native environment-grown M. himalaica plants. A total of 3641 differentially expressed genes (DEGs) and 106 differentially expressed proteins (DEPs) were identified in M. himalaica between UV-B treatment and control check (CK). Comprehensive analysis of protein and transcript data sets resulted in 14 and 7 DEGs from the plant hormone signal transduction and phosphatidylinositol signaling system pathways, respectively, being significantly enriched. The result showed that the plant hormone signal transduction and phosphatidylinositol signaling system might be the key metabolic strategy of UV-B radiation to improve the biosynthesis of rotenoid in M. himalaica. At same time, most of the DEGs were associated with auxin and calcium signaling, inferring that they might drive the downstream transmission of these signal transduction pathways. Regarding those pathways, two chalcone synthase enzymes, which play key roles in the biosynthesis of rotenoid that were thought as the representative medicinal component of M. himalaica, were significantly upregulated in UV-B radiation. This study provides a theoretical basis for further exploration of the adaptation mechanism of M. himalaica to UV-B radiation, and references for cultivation standardization. [ABSTRACT FROM AUTHOR]

Artemisia argyi is an herbaceous plant of the genus Artemisia. Its young and mature leaves are used as food and medicine, respectively. Glandular trichomes (GTs) are distributed on the leaf surface in A. argyi and are generally considered the location of flavonoid biosynthesis and accumulation. However, the mechanism of flavonoid biosynthesis and accumulation in A. argyi remains unclear. In this study, the coregulatory genes involved in flavonoid biosynthesis and trichome development in this species were screened and evaluated, and the biosynthetic pathways for key flavonoids in A. argyi were uncovered. AaMYB1 and AaYABBY1 were screened using weighted gene co-expression network analysis, and both genes were then genetically transformed into Nicotiana tabacum L. cv. K326 (tobacco). Simultaneously, AaYABBY1 was also genetically transformed into Arabidopsis thaliana. The total flavonoid and rutin contents were increased in tobacco plants overexpressing AaMYB1 and AaYABBY1 , and the expression levels of genes participating in the flavonoid synthesis pathway, such as PAL , FLS , and F3H , were significantly up-regulated in plants overexpressing these genes. These results indicated that AaMYB1 and AaYABBY1 promote flavonoid biosynthesis in tobacco. Furthermore, compared to that in the wild-type, the trichome density was significantly increased in tobacco and A. thaliana plants overexpressing AaYABBY1. These results confirm that AaYABBY1 might be involved in regulating trichome formation in A. argyi. This indicates the potential genes involved in and provides new insights into the development of trichome cellular factories based on the "development–metabolism" interaction network and the cultivation of high-quality A. argyi. [Display omitted] • AaMYB1 and AaYABBY1 were identified from Artemisia argyi. • Overexpression of AaMYB1 and AaYABBY1 promoted flavonoid biosynthesis in transgenic tobacco. • AaYABBY1 increased trichome density by enhancing the expression of TTG1 , TTG2 , MYB5 and MYB23 in transgenic Arabidopsis thaliana. [ABSTRACT FROM AUTHOR]

[Display omitted] • A set of Rehmannia enzyme-coding genes for salidroside biosynthesis was identified. • In vitro activity revealed these enzymes' function for salidroside biosynthesis. • Heterologous coexpression of these enzyme-encoding genes produced salidroside in vivo. Rehmannia glutinosa produces many phenylethanoid glycoside (PhG) compounds, including salidroside, which not only possesses various biological activities but also is a core precursor of some medicinal PhGs, so it is very important to elucidate the species' salidroside biosynthesis pathway to enhance the production of salidroside and its derivations. Although some plant copper-containing amine oxidases (CuAOs), phenylacetaldehyde reductases (PARs) and UDP-glucose glucosyltransferases (UGTs) are thought to be vital catalytic enzymes involved in the downstream salidroside biosynthesis pathways, to date, none of these proteins or the associated genes in R. glutinosa have been characterized. To verify a postulated R. glutinosa salidroside biosynthetic pathway starting from tyrosine, this study identified and characterized a set of R. glutinosa genes encoding RgCuAO, RgPAR and RgUGT enzymes for salidroside biosynthesis. The functional activities of these proteins were tested in vitro by heterologous expression of these genes in Escherichia coli , confirming these catalytic abilities in these corresponding reaction steps of the biosynthetic pathway. Importantly, four enzyme-encoding genes (including the previously reported RgTyDC2 encoding tyrosine decarboxylase and the RgCuAO1 , RgPAR1 and RgUGT2 genes) were cointegrated into Saccharomyces cerevisiae to reconstitute the R. glutinosa salidroside biosynthetic pathway, achieving an engineered strain that produced salidroside and validating these enzymes' catalytic functions. This study elucidates the complete R. glutinosa salidroside biosynthesis pathway from tyrosine metabolism in S. cerevisiae , establishing a basic platform for the efficient production of salidroside and its derivatives. [ABSTRACT FROM AUTHOR]

Summary of main observation and conclusion: Collismycins (COLs) are antibiotics characterized by a 2,2′‐bipyridine (2,2′‐BP) core composed of a trisubstituted ring A and an unmodified ring B. The 2,2′‐BP core, which possesses metal‐chelating ability and plays key roles in various biological activities of COLs, is biosynthesized by a nonribosomal peptide synthetase (NRPS)‐polyketide synthase (PKS) hybrid machinery. The starter module of the NRPS‐PKS hybrid machinery consists of a type II peptidyl carrier protein (PCP) ColA1a and an adenylation protein ColA1b. We here report the functional characterization of ColA1a and ColA1b in vitro, confirming their functions in selection and loading of picolinic acid (PA), instead of normal amino acid substrates, as the origin of ring B in COLs. The 2.1 Å crystal structure of ColA1a was solved, revealing structural features including the additional helices α1a, α1b and missing helix α3, which may reflect unique interactions of ColA1a with other NRPS‐PKS proteins/domains or substrate. Primary and tertiary structural comparison of ColA1a with other PCPs revealed the structural basis for their typical α‐helical bundle, providing a better understanding of the structural flexibility of PCPs. These results facilitate the starter module engineering for the generation of COL derivatives with ring B modifications in the future. [ABSTRACT FROM AUTHOR]