| Reference | [1]. Plant Physiol. 2015 Jun;168(2):428-42. doi: 10.1104/pp.15.00074. Epub 2015 Apr 10.<br />
Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins.<br />
Sircar D(1), Gaid MM(1), Chizzali C(1), Reckwell D(1), Kaufholdt D(1), Beuerle T(1), Broggini GA(1), Flachowsky H(1), Liu B(1), Hänsch R(1), Beerhues L(2).<br />
Author information: (1)Institute of Pharmaceutical Biology (D.S., M.M.G., D.R., T.B., B.L., L.B.) and Institute of Plant Biology (D.K., R.H.), Technische Universität Braunschweig, 38106 Braunschweig, Germany;Plant Pathology, Institute of Integrative Biology, Eidgenössische Technische Hochschule Zürich, 8092 Zurich, Switzerland (C.C., G.A.L.B.); andJulius Kühn-Institute, Federal Research Centre for Cultivated Plants, Institute for Breeding Research on Horticultural and Fruit Crops, 01326 Dresden, Germany (H.F.). (2)Institute of Pharmaceutical Biology (D.S., M.M.G., D.R., T.B., B.L., L.B.) and Institute of Plant Biology (D.K., R.H.), Technische Universität Braunschweig, 38106 Braunschweig, Germany;Plant Pathology, Institute of Integrative Biology, Eidgenössische Technische Hochschule Zürich, 8092 Zurich, Switzerland (C.C., G.A.L.B.); andJulius Kühn-Institute, Federal Research Centre for Cultivated Plants, Institute for Breeding Research on Horticultural and Fruit Crops, 01326 Dresden, Germany (H.F.) [email protected].<br />
Upon pathogen attack, fruit trees such as apple (Malus spp.) and pear (Pyrus spp.) accumulate biphenyl and dibenzofuran phytoalexins, with aucuparin as a major biphenyl compound. 4-Hydroxylation of the biphenyl scaffold, formed by biphenyl synthase (BIS), is catalyzed by a cytochrome P450 (CYP). The biphenyl 4-hydroxylase (B4H) coding sequence of rowan (Sorbus aucuparia) was isolated and functionally expressed in yeast (Saccharomyces cerevisiae). SaB4H was named CYP736A107. No catalytic function of CYP736 was known previously. SaB4H exhibited absolute specificity for 3-hydroxy-5-methoxybiphenyl. In rowan cell cultures treated with elicitor from the scab fungus, transient increases in the SaB4H, SaBIS, and phenylalanine ammonia lyase transcript levels preceded phytoalexin accumulation. Transient expression of a carboxyl-terminal reporter gene construct directed SaB4H to the endoplasmic reticulum. A construct lacking the amino-terminal leader and transmembrane domain caused cytoplasmic localization. Functional B4H coding sequences were also isolated from two apple (Malus × domestica) cultivars. The MdB4Hs were named CYP736A163. When stems of cv Golden Delicious were infected with the fire blight bacterium, highest MdB4H transcript levels were observed in the transition zone. In a phylogenetic tree, the three B4Hs were closest to coniferaldehyde 5-hydroxylases involved in lignin biosynthesis, suggesting a common ancestor. Coniferaldehyde and related compounds were not converted by SaB4H.<br />
DOI: 10.1104/pp.15.00074 PMCID: PMC4453778 PMID: 25862456 [Indexed for MEDLINE]<br />
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[2]. J Med Food. 2021 Feb;24(2):151-160. doi: 10.1089/jmf.2020.4861. Epub 2021 Jan 28.<br />
Aucuparin Suppresses Bleomycin-Induced Pulmonary Fibrosis Via Anti-Inflammatory Activity.<br />
Lee SY(1), Park SY(1), Lee GE(1), Kim H(1), Kwon JH(1), Kim MJ(1), Yoon HG(1).<br />
Author information: (1)Department of Biochemistry and Molecular Biology, Severance Medical Research Institute, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul, Korea.<br />
Idiopathic pulmonary fibrosis (IPF) is a lung disease that results in scarring of the lungs for an unknown reason. Although many studies have been conducted on IPF, precise mechanisms and treatments have not yet been identified. In this study, we found that aucuparin, a natural product isolated from Sorbus aucuparia, inhibited pulmonary fibrosis in a bleomycin (BLM)-induced lung fibrosis mouse model. In the lung samples of mice treated with aucuparin, the gene expression of inflammation and macrophage activation markers was reduced compared to those treated with BLM alone. Moreover, aucuparin decreased the expression of profibrotic marker genes and increased the expression of antifibrotic marker genes. Finally, we observed that aucuparin significantly suppressed transforming growth factor-β-induced activation of inflammatory cytokine production and collagen synthesis from macrophages and fibroblasts, respectively. Taken together, these data demonstrate that aucuparin inhibits lung fibrosis via its anti-inflammatory action and support its potential to be a therapeutic drug for IPF treatment.<br />
DOI: 10.1089/jmf.2020.4861 PMID: 33512266 [Indexed for MEDLINE]<br />
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[3]. Phytochemistry. 2013 Dec;96:101-9. doi: 10.1016/j.phytochem.2013.09.003. Epub 2013 Sep 24.<br />
Biosynthesis of the biphenyl phytoalexin aucuparin in Sorbus aucuparia cell cultures treated with Venturia inaequalis.<br />
Khalil MN(1), Beuerle T, Müller A, Ernst L, Bhavanam VB, Liu B, Beerhues L.<br />
Author information: (1)Institute of Pharmaceutical Biology, Technische Universität Braunschweig, Mendelssohnstr. 1, 38106 Braunschweig, Germany.<br />
Aucuparin is the most widely distributed biphenyl phytoalexin in the rosaceous subtribe Pyrinae, which includes the economically important fruit trees apple and pear. The biphenyl scaffold is formed by biphenyl synthase, which catalyzes biosynthesis of 3,5-dihydroxybiphenyl. Conversion of this precursor to aucuparin (3,5-dimethoxy-4-hydroxybiphenyl) was studied in cell cultures of Sorbus aucuparia after treatment with an elicitor preparation from the scab-causing fungus Venturia inaequalis. The sequence of the biosynthetic steps detected was O-methylation – 4-hydroxylation – O-methylation. The two alkylation reactions were catalyzed by distinct methyltransferases, which differed in pH and temperature optima as well as stability. Biphenyl 4-hydroxylase was a microsomal cytochrome P450 monooxygenase, whose activity was appreciably decreased by the addition of established P450 inhibitors. When fed to V. inaequalis-treated S. aucuparia cell cultures, radioactively labeled 3,5-dihydroxybiphenyl was not only incorporated into aucuparin but also into the dibenzofuran eriobofuran, the accumulation of which paralleled that of aucuparin. However, biphenyl 2'-hydroxylase activity proposed to be involved in dibenzofuran formation was detected in neither microsomes nor cell-free extracts in the presence of NADPH and 2-oxoglutarate, respectively. Nevertheless, a basis for studying biphenyl biosynthesis at the gene level is provided.<br />
DOI: 10.1016/j.phytochem.2013.09.003 PMID: 24074553 [Indexed for MEDLINE]<br />
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[4]. Planta. 2019 Mar;249(3):677-691. doi: 10.1007/s00425-018-3031-6. Epub 2018 Oct 24.<br />
Molecular cloning and functional analysis of a biphenyl phytoalexin-specific O-methyltransferase from apple cell suspension cultures.<br />
Sarkate A(1), Saini SS(1), Gaid M(2), Teotia D(1), Mir JI(3), Agrawal PK(4), Beerhues L(2), Sircar D(5).<br />
Author information: (1)Plant Molecular Biology Group, Biotechnology Department, Indian Institute of Technology Roorkee, Roorkee, 247667, India. (2)Institute of Pharmaceutical Biology, Technische Universität Braunschweig, Mendelssohnstrasse 1, 38106, Braunschweig, Germany. (3)Plant Biotechnology Department, Central Institute of Temperate Horticulture (ICAR-CITH), Srinagar, 190005, Jammu and Kashmir, India. (4)Krishi Anusandhan Bhawan-I, ICAR, New Delhi, India. (5)Plant Molecular Biology Group, Biotechnology Department, Indian Institute of Technology Roorkee, Roorkee, 247667, India. [email protected].<br />
This manuscript describes the cloning and functional characterization of a biphenyl phytoalexin biosynthetic gene, 3,5-dihydroxybiphenyl O-methyltransferase from elicitor-treated cell cultures of scab resistant apple cultivar 'Florina'. Apples belong to the subtribe Malinae of the Rosaceae family. Biphenyls and dibenzofurans are the specialized phytoalexins of Malinae, of which aucuparin is the most widely distributed biphenyl. The precursor of aucuparin, 3,5-dihydroxybiphenyl, is a benzoate-derived polyketide, which is formed by the sequential condensation of three molecules of malonyl-CoA and one molecule of benzoyl-CoA in a reaction catalyzed by biphenyl synthase (BIS). This 3,5-dihydroxybiphenyl then undergoes sequential 5-O-methylation, 4-hydroxylation, and finally 3-O-methylation to form aucuparin. A cDNA encoding O-methyltransferase (OMT) was isolated and functionally characterized from the cell cultures of scab-resistant apple cultivar 'Florina' (Malus domestica cultivar 'Florina'; MdOMT) after treatment with elicitor prepared from the apple scab causing fungus Venturia inaequalis. MdOMT catalyzed the regiospecific O-methylation of 3,5-dihydroxybiphenyl at the 5-position to form 3-hydroxy-5-methoxybiphenyl. The enzyme showed absolute substrate preference for 3,5-dihydroxybiphenyl. The elicitor-treated apple cell cultures showed transient increases in the MdOMT (GenBank ID MF740747) and MdBIS3 (GenBank ID JQ390523) transcript levels followed by the accumulation of biphenyls (aucuparin and noraucuparin) and dibenzofuran (eriobofuran) phytoalexins. MdOMT fused with N- and C-terminal yellow fluorescent protein showed cytoplasmic localization in the epidermis of Nicotiana benthamiana leaves. In scab inoculated greenhouse-grown 'Florina' plants, the expression of MdOMT was transiently induced in the stem followed by the accumulation of biphenyl phytoalexins.<br />
DOI: 10.1007/s00425-018-3031-6 PMID: 30357505 [Indexed for MEDLINE]<br />
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[5]. Planta. 2012 Jan;235(1):217-23. doi: 10.1007/s00425-011-1545-2. Epub 2011 Nov 16.<br />
Endogenous hydrogen peroxide is a key factor in the yeast extract-induced activation of biphenyl biosynthesis in cell cultures of Sorbus aucuparia.<br />
Qiu X(1), Lei C, Huang L, Li X, Hao H, Du Z, Wang H, Ye H, Beerhues L, Liu B.<br />
Author information: (1)Key Laboratory of Photosynthesis and Environmental Molecular Physiology, Institute of Botany, The Chinese Academy of Sciences, Nanxincun 20, Haidian District, Beijing, 100093, China.<br />
Biphenyls are unique phytoalexins produced by plants belonging to Pyrinae, a subtribe of the economically important Rosaceae family. The formation of aucuparin, a well-known biphenyl, is induced by yeast extract (YE) in cell cultures of Sorbus aucuparia. However, the molecular mechanism underlying YE-induced activation of biphenyl biosynthesis remains unknown. Here we demonstrate that the addition of YE to the cell cultures results in a burst of reactive oxygen species (ROS; H(2)O(2) and O(2) (-)), followed by transcriptional activation of the biphenyl synthase 1 gene (BIS1) encoding the key enzyme of the biphenyl biosynthetic pathway and aucuparin accumulation. Pretreatment of the cell cultures with ROS scavenger dihydrolipoic acid and NADPH oxidase-specific inhibitor diphenylene iodonium abolished all of the above YE-induced biological events. However, when the cell cultures was pretreated with superoxide dismutase specific inhibitor N,N-diethyldithiocarbamic acid, although O(2) (-) continued to be generated, the H(2)O(2) accumulation, BIS1 expression and aucuparin production were blocked. Interestingly, exogenous supply of H(2)O(2) in the range of 0.05-10 mM failed to induce aucuparin accumulation. These results indicate that endogenous generation of H(2)O(2) rather than that of O(2) (-) is a key factor in YE-induced accumulation of biphenyl phytoalexins in cell cultures of S. aucuparia.<br />
DOI: 10.1007/s00425-011-1545-2 PMID: 22086110 [Indexed for MEDLINE]
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