Glutathione oxidation in response to intracellular H2O2: Key but overlapping roles for dehydroascorbate reductases

Glutathione is a pivotal molecule in oxidative stress, during which it is potentially oxidized by several pathways linked to H2O2 detoxification. We have investigated the response and functional importance of 3 potential routes for glutathione oxidation pathways mediated by glutathione S-transferases (GST), glutaredoxin-dependent peroxiredoxins (PRXII), and dehydroascorbate reductases (DHAR) in Arabidopsis during oxidative stress. Loss-of-function gstU8, gstU24, gstF8, prxIIE and prxIIF mutants as well as double gstU8 gstU24, gstU8 gstF8, gstU24 gstF8, prxIIE prxIIF mutants were obtained. No mutant lines showed marked changes in their phenotype and glutathione profiles in comparison to the wild-type plants in either optimal conditions or oxidative stress triggered by catalase inhibition. By contrast, multiple loss of DHAR functions markedly decreased glutathione oxidation triggered by catalase deficiency. To assess whether this effect was mediated directly by loss of DHAR enzyme activity, or more indirectly by upregulation of other enzymes involved in glutathione and ascorbate recycling, we measured expression of glutathione reductase (GR) and expression and activity of monodehydroascorbate reductases (MDHAR). No evidence was obtained that either GRs or MDHARs were upregulated in plants lacking DHAR function. Hence, interplay between different DHARs appears to be necessary to couple ascorbate and glutathione pools and to allow glutathione-related signaling during enhanced H2O2 metabolism.谷胱甘肽(Glutathione)是氧化应激过程中的关键分子，在此过程中它可通过多条与过氧化氢(H₂O₂)解毒相关的通路发生氧化。本研究探究了拟南芥在氧化应激状态下，由谷胱甘肽S-转移酶(GST)、谷氧还蛋白依赖型过氧化物酶(PRXII)以及脱氢抗坏血酸还原酶(DHAR)介导的3条潜在谷胱甘肽氧化通路的响应及其功能重要性。我们成功获得了gstU8、gstU24、gstF8、prxIIE和prxIIF单功能缺失突变体，以及gstU8 gstU24、gstU8 gstF8、gstU24 gstF8、prxIIE prxIIF双功能缺失突变体。在正常生长条件或过氧化氢酶抑制诱导的氧化应激条件下，所有突变株系的表型及谷胱甘肽谱均与野生型植株无显著差异。与之相反，同时缺失多种DHAR功能会显著降低由过氧化氢酶缺乏诱导的谷胱甘肽氧化水平。为探究该效应是直接由DHAR酶活性丧失介导，还是通过上调谷胱甘肽与抗坏血酸循环相关的其他酶类间接产生，我们检测了谷胱甘肽还原酶(GR)的表达水平，以及单脱氢抗坏血酸还原酶(MDHAR)的表达与酶活性。结果未发现DHAR功能缺失植株中GR或MDHAR表达上调的证据。因此，不同DHAR之间的协同作用似乎是连接抗坏血酸与谷胱甘肽库、并在过氧化氢代谢增强时实现谷胱甘肽相关信号传导的必要条件。