title: Hygromycin B-induced cell death is partly mediated by reactive oxygen species in rice (Oryza sativa L.).
abstract: The aminoglycoside antibiotic hygromycin B (Hyg) inhibits prokaryotic, chloroplast and mitochondrial protein synthesis. Because of the toxic effect of Hyg on plant cells, the HPT gene, encoding hygromycin phosphotransferase, has become one of the most widely used selectable markers in plant transformation. Yet the mechanism behind Hyg-induced cell lethality in plants is not clearly understood. In this study, we aimed to decipher this mechanism. With Hyg treatment, rice calli exhibited cell death, and rice seedlings showed severe growth defects, leaf chlorosis and leaf shrinkage. Rice seedlings also exhibited severe lipid peroxidation and protein carbonylation, for oxidative stress damage at the cellular level. The production of reactive oxygen species such as O2(·-), H2O2 and OH(·) was greatly induced in rice seedlings under Hyg stress, and pre-treatment with ascorbate increased resistance to Hyg-induced toxicity indicating the existence of oxidative stress. Overexpression of mitochondrial Alternative oxidase1a gene without HPT selection marker in rice enhanced tolerance to Hyg and attenuated the degradation of protein content, whereas the rice plastidial glutathione reductase 3 mutant showed increased sensitivity to Hyg. These results demonstrate that Hyg-induced cell lethality in rice is not only due to the inhibition of protein synthesis but also mediated by oxidative stress.
authors.affiliation: Department of Agricultural Chemistry, College of Bioresources and Agriculture, National Taiwan University, Taipei, 10617, Taiwan.
authors.forename: Hui-Min
authors.initials: HM
authors.lastname: Oung
authors.affiliation: Department of Agricultural Chemistry, College of Bioresources and Agriculture, National Taiwan University, Taipei, 10617, Taiwan.
authors.forename: Ke-Chun
authors.initials: KC
authors.lastname: Lin
authors.affiliation: Department of Agricultural Chemistry, College of Bioresources and Agriculture, National Taiwan University, Taipei, 10617, Taiwan.|Department of Aquaculture, National Pingtung University of Science and Technology, Pingtung, 91201, Taiwan.
authors.forename: Tsung-Meng
authors.initials: TM
authors.lastname: Wu
authors.affiliation: Department of Agricultural Chemistry, College of Bioresources and Agriculture, National Taiwan University, Taipei, 10617, Taiwan.
authors.forename: Nulu Naga Prafulla
authors.initials: NN
authors.lastname: Chandrika
authors.affiliation: Department of Agricultural Chemistry, College of Bioresources and Agriculture, National Taiwan University, Taipei, 10617, Taiwan. cyhong@ntu.edu.tw.
authors.forename: Chwan-Yang
authors.initials: CY
authors.lastname: Hong
mesh_terms: D000900:Anti-Bacterial Agents / Q000633:toxicity; D016923:Cell Death / Q000187:drug effects; D017343:Genes, Plant; D005980:Glutathione Reductase / Q000235:genetics; D006921:Hygromycin B / Q000633:toxicity*; D024101:Mitochondrial Proteins / Q000235:genetics; D009154:Mutation; D012275:Oryza / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D010088:Oxidoreductases / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics; D030821:Plants, Genetically Modified; D017382:Reactive Oxygen Species / Q000378:metabolism; D036226:Seedlings / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D000099188:Alternative Oxidase
chemical_list: D000900:Anti-Bacterial Agents; D005980:Glutathione Reductase; D006921:Hygromycin B; D024101:Mitochondrial Proteins; D010088:Oxidoreductases; D017853:Phosphotransferases (Alcohol Group Acceptor); D010940:Plant Proteins; D017382:Reactive Oxygen Species; D000099188:Alternative Oxidase; C063242:hygromycin-B kinase
keywords: Alternative oxidase1a; Cell death; Glutathione reductase 3; Hygromycin B; Reactive oxygen species; Rice