Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwake-cho, Sakyo-ku, Kyoto, 606-8502, Japan.
Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8572, Japan.
Plant Mol Biol. 2020 Mar;102(4-5):403-416. doi: 10.1007/s11103-019-00954-3. Epub 2020 Jan 3.
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. 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.
CYP81A P450s 使稗草对多种和多种除草剂化学物质具有抗药性。CYP81A 底物的偏好可以作为稗草和其他相关物种中交叉抗性预测和管理的基础。基于代谢的除草剂抗性是农业的主要威胁，因为它是不可预测的，并且可能会扩展到不同的化学基团和作用模式，包括现有的、新的和有待发现的除草剂。关于参与除草剂代谢的酶的信息有限，这阻碍了杂草中交叉抗性的预测。先前在具有多种除草剂抗性 (MHR) 的稗草中鉴定出多个 CYP81A 亚家族成员，这些成员赋予了对六种不相关除草剂类别的交叉抗性。这表明 CYP81As 在赋予稗草不可预测的交叉抗性方面起着关键作用，因此对其九个假定的功能性 CYP81A 基因对来自 24 个化学组的 33 种除草剂进行了功能表征。在拟南芥中异位表达鉴定出了能够赋予多种和多种除草剂抗性的 CYP81A。进一步在大肠杆菌中对 CYP81A 进行了酶功能表征。通过修饰 N 端、与 HemA 基因共表达和在最佳温度下培养优化了大肠杆菌中 CYP81A 的表达。CYP81A 将其除草剂底物代谢为羟基化、N-/O-去甲基化或两者的产物。CYP81A 赋予的交叉抗性模式针对乙酰乳酸合酶抑制剂的所有化学基团，并扩展到抑制光合作用系统 II、脱叶烯酶、原卟啉原氧化酶、4-羟苯基丙酮酸双加氧酶和 1-脱氧-D-木酮糖 5-磷酸合酶的除草剂。在 MHR 稗草中预测和证实了对除草剂嘧啶肟、丙酯磺隆和噻酮磺隆的交叉抗性。本研究表明，对除草剂代谢关键酶的功能表征可以揭示交叉抗性模式，并确定适当的化学选择来管理稗草中现有的和意外的交叉抗性。
