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Functional characterization of cytochrome P450 CYP81A subfamily to disclose the pattern of cross-resistance in Echinochloa phyllopogon

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CYP81A P450s armor Echinochloa phyllopogon against diverse and several herbicide chemistries. CYP81A substrate preferences can be a basis for cross-resistance prediction and management in E. phyllopogon and other related species.

Abstract

Metabolism-based herbicide resistance is a major threat to agriculture, as it is unpredictable and could extend resistance to different chemical groups and modes of action, encompassing existing, novel and to-be-discovered herbicides. Limited information on the enzymes involved in herbicide metabolism has hindered the prediction of cross-resistance in weeds. Members of CYP81A subfamily in multiple herbicide resistant (MHR) Echinochloa phyllopogon were previously identified for conferring cross-resistance to six unrelated herbicide classes. This suggests a critical role of CYP81As in endowing unpredictable cross-resistances in E. phyllopogon, thus the functions of all its nine putative functional CYP81A genes to 33 herbicides from 24 chemical groups were characterized. Ectopic expression in Arabidopsis thaliana identified the CYP81As that can confer resistance to multiple and diverse herbicides. The CYP81As were further characterized for their enzymatic functions in Escherichia coli. CYP81A expression in E. coli was optimized via modification of the N-terminus, co-expression with HemA gene and culture at optimal temperature. CYP81As metabolized its herbicide substrates into hydroxylated, N-/O-demethylated or both products. The cross-resistance pattern conferred by CYP81As is geared towards all chemical groups of acetolactate synthase inhibitors and is expanded to herbicides inhibiting photosystem II, phytoene desaturase, protoporphyrinogen oxidase, 4-hydroxyphenylpyruvate dioxygenase, and 1-deoxy-d-xylulose 5-phosphate synthase. Cross-resistance to herbicides pyrimisulfan, propyrisulfuron, and mesotrione was predicted and confirmed in MHR E. phyllopogon. This study demonstrated that the functional characterization of the key enzymes for herbicide metabolism could disclose the cross-resistance pattern and identify appropriate chemical options to manage the existing and unexpected cross-resistances in E. phyllopogon.

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Acknowledgements

All the authors would like to thank Kumiai Chemical Industry Co., Ltd. for providing the herbicides fenoxasulfone and thiobencarb. This research was partly supported by JSPS KAKENHI Grant Number 17KI5234 to SI and by WSSJ Research Initiative Grants Program No. 1901 to TY. The authors wish to thank the anonymous reviewers and the editor for their constructive comments on the manuscript.

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Author notes

  1. Niña Gracel Dimaano and Takuya Yamaguchi contributed equally to this work.

Authors and Affiliations

  1. Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwake-cho, Sakyo-ku, Kyoto, 606-8502, Japan

    Niña Gracel Dimaano, Kanade Fukunishi, Tohru Tominaga & Satoshi Iwakami

  2. Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8572, Japan

    Takuya Yamaguchi

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  1. Niña Gracel Dimaano
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  2. Takuya Yamaguchi
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  3. Kanade Fukunishi
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  4. Tohru Tominaga
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  5. Satoshi Iwakami
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Contributions

SI and TY conceived the research and experiment plans. ND, TY, KF and SI conducted the experiments. ND, TY and SI analyzed and interpreted the data. ND, TY and SI wrote the manuscript with valuable contributions of TT.

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Correspondence to Satoshi Iwakami.

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Dimaano, N.G., Yamaguchi, T., Fukunishi, K. et al. Functional characterization of cytochrome P450 CYP81A subfamily to disclose the pattern of cross-resistance in Echinochloa phyllopogon. Plant Mol Biol 102, 403–416 (2020). https://doi.org/10.1007/s11103-019-00954-3

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