Schulz Margot, Filary Barbara, Kühn Sabine, Colby Thomas, Harzen Anne, Schmidt Jürgen, Sicker Dieter, Hennig Lothar, Hofmann Diana, Disko Ulrich, Anders Nico
a IMBIO Institut für Biotechnologie der Pflanzen, Universität Bonn , Karlrobert Kreiten Str. 13, 53115 Bonn , Germany.
b Max-Planck-Institut für Pflanzenzüchtungsforschung, Carl von Linné Weg 10 , 50829 Köln , Germany.
Plant Signal Behav. 2016;11(1):e1119962. doi: 10.1080/15592324.2015.1119962.
The major detoxification product in maize roots after 24 h benzoxazolin-2(3H)-one (BOA) exposure was identified as glucoside carbamate resulting from rearrangement of BOA-N-glucoside, but the pathway of N-glucosylation, enzymes involved and the site of synthesis were previously unknown. Assaying whole cell proteins revealed the necessity of H2O2 and Fe(2+) ions for glucoside carbamate production. Peroxidase produced BOA radicals are apparently formed within the extraplastic space of the young maize root. Radicals seem to be the preferred substrate for N-glucosylation, either by direct reaction with glucose or, more likely, the N-glucoside is released by glucanase/glucosidase catalyzed hydrolysis from cell wall components harboring fixed BOA. The processes are accompanied by alterations of cell wall polymers. Glucoside carbamate accumulation could be suppressed by the oxireductase inhibitor 2-bromo-4´-nitroacetophenone and by peroxidase inhibitor 2,3-butanedione. Alternatively, activated BOA molecules with an open heterocycle may be produced by microorganisms (e.g., endophyte Fusarium verticillioides) and channeled for enzymatic N-glucosylation. Experiments with transgenic Arabidopsis lines indicate a role of maize glucosyltransferase BX9 in BOA-N-glycosylation. Western blots with BX9 antibody demonstrate the presence of BX9 in the extraplastic space. Proteomic analyses verified a high BOA responsiveness of multiple peroxidases in the apoplast/cell wall. BOA incubations led to shifting, altered abundances and identities of the apoplast and cell wall located peroxidases, glucanases, glucosidases and glutathione transferases (GSTs). GSTs could function as glucoside carbamate transporters. The highly complex, compartment spanning and redox-regulated glucoside carbamate pathway seems to be mainly realized in Poaceae. In maize, carbamate production is independent from benzoxazinone synthesis.
在暴露于苯并恶唑啉 - 2(3H)-酮（BOA）24小时后，玉米根中的主要解毒产物被鉴定为BOA - N - 葡萄糖苷重排产生的氨基甲酸酯葡萄糖苷，但此前N - 糖基化途径、相关酶及合成位点均未知。对全细胞蛋白质的分析表明，H₂O₂和Fe(2+)离子对于氨基甲酸酯葡萄糖苷的产生是必需的。过氧化物酶产生的BOA自由基显然是在玉米幼根的质外体空间内形成的。自由基似乎是N - 糖基化的首选底物，要么通过与葡萄糖直接反应，要么更有可能是通过葡聚糖酶/葡萄糖苷酶催化水解从含有固定BOA的细胞壁成分中释放出N - 葡萄糖苷。这些过程伴随着细胞壁聚合物的改变。氨基甲酸酯葡萄糖苷的积累可被氧化还原酶抑制剂2 - 溴 - 4´ - 硝基苯乙酮和过氧化物酶抑制剂2,3 - 丁二酮抑制。另外，具有开放杂环的活化BOA分子可能由微生物（如内生真菌串珠镰刀菌）产生，并被引导进行酶促N - 糖基化。对转基因拟南芥品系的实验表明玉米糖基转移酶BX9在BOA - N - 糖基化中起作用。用BX9抗体进行的蛋白质免疫印迹显示BX9存在于质外体空间。蛋白质组学分析证实了质外体/细胞壁中多种过氧化物酶对BOA具有高反应性。BOA孵育导致质外体和细胞壁中过氧化物酶、葡聚糖酶、葡萄糖苷酶和谷胱甘肽转移酶（GST）的迁移、丰度改变和种类变化。GST可能作为氨基甲酸酯葡萄糖苷的转运体。高度复杂、跨越不同区室且受氧化还原调节的氨基甲酸酯葡萄糖苷途径似乎主要在禾本科植物中实现。在玉米中，氨基甲酸酯的产生独立于苯并恶嗪酮的合成。
