Enzymatic regulation of shikonin biosynthesis in Lithospermum erythrorhizon cell cultures

Heide, Lutz; Nishioka, Naoya; Fukui, Hiroshi; Tabata, Minoru

doi:10.1016/s0031-9422(00)97877-4

Cited by 86 publications

(39 citation statements)

References 16 publications

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“…41) Furthermore, Gaisser and Heide 44) reported that white light irradiation increased the activity of PHB-O-glucosyltransferase in L. erythrorhizon cells, while blue light did not affect it. No shikonin in cells cultured under white or blue light was produced.…”

Section: Discussionmentioning

confidence: 99%

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Regulation of Lithospermic Acid B and Shikonin Production in Lithospermum erythrorhizon Cell Suspension Cultures.

Yamamoto

Zhao

Yazaki

et al. 2002

CHEMICAL & PHARMACEUTICAL BULLETIN

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Caffeic acid oligomers such as rosmarinic acid and lithospermic acid B (LAB) are distributed in plants of Labiatae and Boraginaceae. Among these oligomers, LAB, the caffeic acid tetramer, has a variety of attractive pharmacological activities such as antioxidant and radical scavenging, 1,2) improvement of renal disease, [3][4][5][6] hypotensive, [7][8][9] improvement of myocardial infarction, 1,10) amplification of beating of myocardial muscles, 11) anti-hepatitis, 12) and aldose reductase inhibitory activity. 13) To utilize LAB as a medicinal resource, more than 30 plant sources were surveyed and only two medicinal plants, Salvia officinalis and Lithospermum erythrorhizon SIEB. et ZUCC, were found to contain this substance in an appreciable amount.14) Many attempts to produce this compound in high quantities using cell or hairy root cultures have been unsuccessful. [15][16][17][18][19][20][21][22] Recently, we found that L. erythrorhizon cell-suspension cultures, which were established for the industrial production of shikonin, 23) produced four caffeic acid oligomers, rosmarinic acid, LAB, a monoglucoside of LAB, and (ϩ)-rabdosiin. 24) In particular, the production of LAB in the cultured cells was highly stimulated in shikonin production medium M-9, 25) reaching almost the same amount as that of shikonin (LAB: ca. 6-10% of dry wt, and total shikonin derivatives: ca. 6-12% of dry wt.). 24,26) Since both LAB and shikonin are formed through a common phenylpropanoid pathway (Chart 1), the counterbalance between two metabolic routes either to LAB or to shikonin would be regulated in L. erythrorhizon cells, suggesting that the selection of suitable regulatory factors would result in higher production of LAB. However, the metabolic linkage and the regulatory mechanism for the biosyntheses of phenylpropanoid-derived compounds of diverse types in L. erythrorhizon cell cultures remain to be clarified. In this study, we investigated the critical regulatory factors to control these two biosynthetic pathways separately and to produce LAB efficiently in the culture system of L. erythrorhizon. ExperimentalCell Cultures The culture strain M18TOM 24) of Lithospermum erythrorhizon, which had been maintained in Linsmaier-Skoog (LS) medium 27) containing IAA 1 mM and kinetin 10 mM, was used in the present study. Cellsuspension cultures in 100-ml flasks each containing 30 ml of medium were agitated on a rotary shaker (80 rpm) at 23°C in the dark and subcultured at intervals of 2 weeks. The cells cultured in LS medium for 2 weeks were inoculated into M-9 medium, 25) modified M-9 or LS media containing 3% sucrose, and the same growth regulators as above (inoculum size: 1 g of fresh cells/30 ml of medium in a 100-ml flask) and cultured for 3 weeks under the above-mentioned culture conditions. For the investigation of the effect of light color on the production of secondary metabolites, each flask was covered with a commercially available colored cellophane sheet, 28) and irradiated with white light (10000 lx) from fluorescent lamps d...

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Section: Discussionmentioning

confidence: 99%

“…White or blue light strongly inhibits shikonin production by suppressing the gene expression of PHB geranyltransferase, a key enzyme in shikonin biosynthesis, 41,42) as well as 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, an enzyme responsible for the formation of the geranyl side chain of shikonin. 43) As shown in Fig.…”

Section: ϫmentioning

confidence: 99%

Regulation of Lithospermic Acid B and Shikonin Production in Lithospermum erythrorhizon Cell Suspension Cultures.

Yamamoto

Zhao

Yazaki

et al. 2002

CHEMICAL & PHARMACEUTICAL BULLETIN

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“…This enzyme plays a critical role in the regulation of shikonin biosynthesis in cell cultures of L. erythrorhizon, i.e. up-and down-regulation of this enzyme activity directly affects the production of shikonin (21,22). The enzyme activity is strongly suppressed by light irradiation, ammonium ion, and the synthetic auxin 2,4-dichlorophenoxyacetic acid (2,4-D) (23), whereas this activity is enhanced with the addition of oligogalacturonide (OG) (24) and methyl jasmonate (MJ) (25) to the medium.…”

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confidence: 99%

Geranyl Diphosphate:4-Hydroxybenzoate Geranyltransferase fromLithospermum erythrorhizon

Yazaki

Kunihisa

Fujisaki

et al. 2002

Journal of Biological Chemistry

141

117

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Two cDNAs encoding geranyl diphosphate:4-hydroxybenzoate 3-geranyltransferase were isolated from Lithospermum erythrorhizon by nested PCR using the conserved amino acid sequences among polyprenyltransferases for ubiquinone biosynthesis. They were functionally expressed in yeast COQ2 disruptant and showed a strict substrate specificity for geranyl diphosphate as the prenyl donor, in contrast to ubiquinone biosynthetic enzymes, suggesting that they are involved in the biosynthesis of shikonin, a naphthoquinone secondary metabolite. Regulation of their expression by various culture conditions coincided with that of geranyltransferase activity and the secondary metabolites biosynthesized via this enzyme. This is the first established plant prenyltransferase that transfers the prenyl chain to an aromatic substrate.

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“…For instance, it has been demonstrated that PAL is activated in the presence of the cell wall fragments of yeast fungal elicitor (33). PAL converts phenylalanine to cinnamic acid, an important precursor in the shikalkin pathway ( Figure 1) (10). In this research, the highest amount of pigment production was observed in the presence of R. solani (40 mg.L -1 ) and the best result was belonged to the collar-originated callus ( Figure 5).…”

Section: Discussionmentioning

confidence: 51%

“…p-Hydroxybenzoic acid (PHB), derived from the phenylpropanoid pathway, and geranyl-pyrophosphate (GPP), from the mevalonic acid pathway, join together under the effect of PHBgeranyl transferase to initiate shikalkin biosynthesis ( Figure 1). Phenylalanine lyase (PAL) and 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGR) play key roles in the shikalkin pathway (10). Light, ammonium and 2,4-dichlorophenoxyacetic acid (2,4-D) inhibit the shikalkin formation while 3-indoleacetic acid (IAA) and copper ion (Cu 2+ ) induce its biosynthesis (11).…”

Section: Introductionmentioning

confidence: 99%

Enhancement of Shikalkin Production in Arnebia euchroma Callus by a Fungal Elicitor, Rhizoctonia solani

2015

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Background: There is a growing demand for mass production of shikalkin (a natural pigment consisted of shikonin and alkannin) due to its increasing applications in cosmetics, pharmaceutical and nutrition industries. The root of Iranian Arnebia euchroma produces shikalkin. The promising capability of this plant for shikalkin production has already been demonstrated in cell culture studies. Objectives: Elicitation effect of Rhizoctonia solani (R. solani) in comparison with the effects of Cu 2+ , methyl jasmonate (MJ), and salicylic acid (SA) on the shikalkin production was investigated in A. euchroma callus. Materials and Methods:The calli from different origins (leaf, collar and root) were proliferated on a modified LinsmaierSkoog (mLS) medium and were subsequently transferred onto the pigment production medium containing various amounts of the desirable elicitor. Observations were quantified and the pigment production was precisely measured spectrophotometrically. Results: Pigment biosynthesis was induced on White medium containing IAA (1 μM) and kinetin (10 μM) in dark at 25°C. Use of R. solani increased the pigment production by 7 fold greater than normal White medium. Cu 2+ only doubled the shikalkin production. MJ and SA showed enhancing effects comparable to that of Cu 2+ . Discussions: It is assumed that upon binding of the polysaccharides of the fungal cells to the plant cell surface, a cascade of signaling is initiated that led to expression of genes involving in the biosynthesis of shikalkin.

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Enzymatic regulation of shikonin biosynthesis in Lithospermum erythrorhizon cell cultures

Cited by 86 publications

References 16 publications

Regulation of Lithospermic Acid B and Shikonin Production in Lithospermum erythrorhizon Cell Suspension Cultures.

Regulation of Lithospermic Acid B and Shikonin Production in Lithospermum erythrorhizon Cell Suspension Cultures.

Geranyl Diphosphate:4-Hydroxybenzoate Geranyltransferase fromLithospermum erythrorhizon

Enhancement of Shikalkin Production in Arnebia euchroma Callus by a Fungal Elicitor, Rhizoctonia solani

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