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Molecular cloning and functional expression of a stress-induced multifunctional O-methyltransferase with pinosylvin methyltransferase activity from Scots pine (Pinus sylvestris L.)

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Abstract

Formation of pinosylvin (PS) and pinosylvin 3-O-monomethyl ether (PSM), as well as the activities of stilbene synthase (STS) and S-adenosyl-l-methionine (SAM):pinosylvin O-methyltransferase (PMT), were induced strongly in needles of Scots pine seedlings upon ozone treatment, as well as in cell suspension cultures of Scots pine upon fungal elicitation. A SAM-dependent PMT protein was purified and partially characterised. A cDNA encoding PMT was isolated from an ozone-induced Scots pine cDNA library. Southern blot analysis of the genomic DNA suggested the presence of a gene family. The deduced protein sequence showed the typical highly conserved regions of O-methyltransferases (OMTs), and average identities of 20–56% to known OMTs. PMT expressed in Escherichia coli corresponded to that of purified PMT (40 kDa) from pine cell cultures. The recombinant enzyme catalysed the methylation of PS, caffeic acid, caffeoyl-CoA and quercetin. Several other substances, such as astringenin, resveratrol, 5-OH-ferulic acid, catechol and luteolin, were also methylated. Recombinant PMT thus had a relatively broad substrate specificity. Treatment of 7-year old Scots pine trees with ozone markedly increased the PMT mRNA level. Our results show that PMT represents a new SAM-dependent OMT for the methylation of stress-induced pinosylvin in Scots pine needles.

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Authors and Affiliations

  1. Institute of Biochemical Plant Pathology, GSF-National Research Center for Environment and Health, 85764, Neuherberg, Germany

    Hélène Chiron, Monika Trost, Werner Heller, Dieter Ernst & Heinrich Sandermann Jr

  2. Biology Department, University of Orléans, BP 6749, 45067, Orléans Cedex 2, France

    Hélène Chiron, Alain Drouet & Anne-Catherine Claudot

  3. MPI für Biochemie, 82512 , Martinsried, Germany

    Christoph Eckerskorn

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  1. Hélène Chiron
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  2. Alain Drouet
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  3. Anne-Catherine Claudot
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  4. Christoph Eckerskorn
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Chiron, H., Drouet, A., Claudot, AC. et al. Molecular cloning and functional expression of a stress-induced multifunctional O-methyltransferase with pinosylvin methyltransferase activity from Scots pine (Pinus sylvestris L.). Plant Mol Biol 44, 733–745 (2000). https://doi.org/10.1023/A:1026507707186

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