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Self-regulation of phytoalexin production: a non-biosynthetic enzyme controls alkaloid biosynthesis in cultured cells of Eschscholzia californica

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Abstract

Benzophenanthridine alkaloids are strong antimicrobials of Papaveraceae and attractive lead compounds for drug development. The cytotoxicity of these compounds requires the producing plant to limit the pathogen-triggered burst of biosynthesis. Cells of Eschscholzia californica excrete early benzophenanthridines to the cell wall, followed by re-uptake and reduction in the cytoplasm by the detoxifying enzyme sanguinarine reductase. We now discovered that this enzyme is a core component of self-control in alkaloid production. RNAi-based silencing of sanguinarine reductase gave rise to mutants that either show a complete stop of elicitor-triggered alkaloid production or a burst of biosynthesis that severalfold surpasses the wild type level. These unexpected phenotypes reflect impacts of substrate or product of sanguinarine reductase: the substrate, sanguinarine, inhibits phospholipase A2 at the plasma membrane, an initial component of the signal path towards expression of biosynthetic enzymes. The product, dihydrosanguinarine, inhibits enzymes of early biosynthesis, prior to reticuline formation. By tuning these steady states, sanguinarine reductase adjusts the capacity of alkaloid biosynthesis: a minimum activity is sufficient to prevent the blockade of the induction pathway by sanguinarine, while the full activity of the same enzyme causes a limitation of the biosynthetic flow via dihydrosanguinarine.

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Acknowledgments

The work was supported by the Deutsche Forschungsgemeinschaft (W.R., grant Nr. Ro 889/12-1) and the Martin-Luckner Foundation, Halle (M.H.). The help of Dr. Angelika Schierhorn, Max-Planck-Unit for Enzymology of Protein Folding, Halle, with the ESI–MS of some alkaloids is gratefully acknowledged.

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Authors and Affiliations

  1. Lab of Molecular Cell Biology, Department of Pharmaceutical Biology, Institute of Pharmacy, Martin-Luther-University Halle-Wittenberg, Kurt-Mothes-Str. 3, 06120, Halle (Saale), Germany

    Henriette Müller, Michael Heinze & Werner Roos

  2. Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, 06120, Halle (Saale), Germany

    Ramona Heinke & Jürgen Schmidt

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  1. Henriette Müller
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  2. Michael Heinze
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  3. Ramona Heinke
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  4. Jürgen Schmidt
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  5. Werner Roos
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Correspondence to Werner Roos.

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Müller, H., Heinze, M., Heinke, R. et al. Self-regulation of phytoalexin production: a non-biosynthetic enzyme controls alkaloid biosynthesis in cultured cells of Eschscholzia californica . Plant Cell Tiss Organ Cult 119, 661–676 (2014). https://doi.org/10.1007/s11240-014-0565-6

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