Academic Journal
CRISPR/Cas9-mediated genome editing reveals the involvement of a polyphenol oxidase in the shikonin-specific biosynthesis in Lithospermum erythrorhizon.
| Τίτλος: | CRISPR/Cas9-mediated genome editing reveals the involvement of a polyphenol oxidase in the shikonin-specific biosynthesis in Lithospermum erythrorhizon. |
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| Συγγραφείς: | Nakanishi K; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan.; Institute of Natural Medicine, Toyama University, 2630 Sugitani, Toyama 930-0194, Japan., Takano Y; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Yamamoto K; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Matsuda Y; Graduate School of Science and Technology, Shinshu University, 3-1-1 Asahi, Matsumoto, Nagano 390-8621, Japan., Yano M; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Mito K; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Ichino T; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan.; Laboratory of Medicinal Cell Biology, Kobe Pharmaceutical University, 4-19-1 Motoyamakita-machi, Higashinada, Hyogo, Kobe 658-8558, Japan., Tatsumi K; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Li H; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Ohara K; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Munakata R; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Suzuki H; Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan., Sakurai N; Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan., Shibata D; Kazusa DNA Research Institute, Kisarazu, Chiba 292-0818, Japan., Osakabe K; Graduate School of Technology, Industrial and Social Science, Tokushima University, 3-18-15 Kuramoto, Tokushima 770-8503, Japan., Watanabe B; Chemistry Laboratory, The Jikei University School of Medicine, 8-3-1 Kokuryo, Chofu, Tokyo 182-8570, Japan., Okada T; Division of Molecular Cell Biology, Department of Biomolecular Sciences, Faculty of Medicine, Saga University 5-1-1 Nabeshima, Saga, Saga 840-8502, Japan., Shimomura K; Graduate School of Life Sciences, Toyo University, 1-1-1 Izumino, Itakura, Ora, Gunma 374-0193, Japan., Takanashi K; Graduate School of Science and Technology, Shinshu University, 3-1-1 Asahi, Matsumoto, Nagano 390-8621, Japan., Sugiyama A; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan., Yazaki K; Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan. |
| Πηγή: | Plant & cell physiology [Plant Cell Physiol] 2026 Sep 29; Vol. 67 (9), pp. 1557-1572. |
| Τύπος έκδοσης: | Journal Article |
| Γλώσσα: | English |
| Στοιχεία περιοδικού: | Publisher: Oxford University Press Country of Publication: Japan NLM ID: 9430925 Publication Model: Print Cited Medium: Internet ISSN: 1471-9053 (Electronic) Linking ISSN: 00320781 NLM ISO Abbreviation: Plant Cell Physiol Subsets: MEDLINE |
| Imprint Name(s): | Publication: Tokyo : Oxford University Press Original Publication: Kyoto, Japan : Japanese Society of Plant Physiologists |
| Ιατρικοί όροι (MeSH): | Lithospermum*/genetics , Lithospermum*/metabolism , Lithospermum*/enzymology , Naphthoquinones*/metabolism , CRISPR-Cas Systems*/genetics , Catechol Oxidase*/metabolism , Catechol Oxidase*/genetics, Plant Roots/metabolism ; Plant Roots/genetics ; Plant Proteins/metabolism ; Plant Proteins/genetics ; Gene Expression Regulation, Plant |
| Περίληψη: | Shikonin, a 1,4-naphthoquinone derivative produced by some limited Boraginaceae species, exhibits unique pharmacological properties and is also used as a natural dye. The regulatory factors of shikonin production have been demonstrated using a cell culture system of Lithospermum erythrorhizon. Among these factors, copper is known to be the strongest enhancer of shikonin production, and the copper-requiring biosynthetic step is shikonin-specific, unlike the byproduct benzoquinones. Despite the long history of shikonin biosynthesis studies for over 40 years, the copper-involved reaction has been still unknown. Here, we explored candidate genes associated with shikonin production using a PCR-select subtraction experiment. Genes encoding PPO, a dicopper-dependent oxidoreductase, was highlighted by the strong synchronous expression with shikonin production. Transcriptome analysis of hairy roots and cultured cells of this plant revealed that, of the five PPO genes expressed in L. erythrorhizon (LePPOs), only LePPO1 showed a close correlation with shikonin production. Then, we generated genome-edited hairy roots of LePPO1 using CRISPR/Cas9-mediated mutagenesis to analyze its impact on shikonin derivative and other specialized metabolite production. The results showed that shikonin content was markedly reduced in all LePPO1-ge lines, while the content of deoxyshikonofuran, a hydroquinone-type shunt product that branches after 3''-hydroxygeranylhydroquinone in the shikonin biosynthetic pathway, remained unaffected in the LePPO1-ge lines. These findings address the question of why a copper ion is crucial for shikonin biosynthesis and suggest that LePPO1 participates in naphthalene ring formation. Interestingly, LePPO1 is localized in plastids, whereas shikonin accumulates in the apoplast. (© The Author(s) 2026. Published by Oxford University Press on behalf of the Japanese Society of Plant Physiologists. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.) |
| Grant Information: | 19H05638 Japan Society for the Promotion of Science KAKENHI; 22K05438 Japan Society for the Promotion of Science KAKENHI; 25K08838 Japan Society for the Promotion of Science KAKENHI; JPMJSP2110 Japan Science and Technology Agency (JST) SPRING; NEDO 16100890-0 New Energy and Industrial Technology Development Organization; RIKEN Cluster for Science, Technology and Innovation Hub; Institute for the Future of Human Society of Kyoto University for 2023; Jikei University Research Fund; Humanosphere Science Research on Sustainable Humanosphere Science; Kyoto University |
| Contributed Indexing: | Keywords: Lithospermum erythrorhizon; 1,4-naphthoquinone derivatives; copper ion; polyphenol oxidase; red gromwell; shikonin biosynthesis |
| Substance Nomenclature: | 0 (Naphthoquinones) 3IK6592UBW (shikonin) EC 1.10.3.1 (Catechol Oxidase) 0 (Plant Proteins) |
| Entry Date(s): | Date Created: 20260529 Date Completed: 20260929 Latest Revision: 20260930 |
| Update Code: | 20260930 |
| DOI: | 10.1093/pcp/pcag074 |
| PMID: | 42213070 |
| Βάση Δεδομένων: | MEDLINE |
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