title: Production of superoxide from photosystem II-light harvesting complex II supercomplex in STN8 kinase knock-out rice mutants under photoinhibitory illumination.
abstract: When phosphorylation of Photosystem (PS) II core proteins is blocked in STN8 knock-out mutants of rice (Oryza sativa) under photoinhibitory illumination, the mobilization of PSII supercomplex is prevented. We have previously proposed that more superoxide (O2(-)) is produced from PSII in the mutant (Nath et al., 2013, Plant J. 76, 675-686). Here, we clarify the type and site for the generation of reactive oxygen species (ROS). Using both histochemical and fluorescence probes, we observed that, compared with wild-type (WT) leaves, levels of ROS, including O2(-) and hydrogen peroxide (H2O2), were increased when leaves from mutant plants were illuminated with excess light. However, singlet oxygen production was not enhanced under such conditions. When superoxide dismutase was inhibited, O2(-) production was increased, indicating that it is the initial event prior to H2O2 production. In thylakoids isolated from WT leaves, kinase was active in the presence of ATP, and spectrophotometric analysis of nitrobluetetrazolium absorbance for O2(-) confirmed that PSII-driven superoxide production was greater in the mutant thylakoids than in the WT. This contrast in levels of PSII-driven superoxide production between the mutants and the WT plants was confirmed by conducting protein oxidation assays of PSII particles from osstn8 leaves under strong illumination. Those assays also demonstrated that PSII-LHCII supercomplex proteins were oxidized more in the mutant, thereby implying that PSII particles incur greater damage even though D1 degradation during PSII-supercomplex mobilization is partially blocked in the mutant. These results suggest that O2(-) is the major form of ROS produced in the mutant, and that the damaged PSII in the supercomplex is the primary source of O2(-).
authors.affiliation: Department of Integrated Biological Science, Department of Molecular Biology, Pusan National University, Busan 609-735, Republic of Korea.
authors.forename: Roshan Sharma
authors.initials: RS
authors.lastname: Poudyal
authors.affiliation: Department of Integrated Biological Science, Department of Molecular Biology, Pusan National University, Busan 609-735, Republic of Korea; Department of Biological Sciences, Western Michigan University, Kalamazoo, MI, USA.
authors.forename: Krishna
authors.initials: K
authors.lastname: Nath
authors.affiliation: Department of Integrated Biological Science, Department of Molecular Biology, Pusan National University, Busan 609-735, Republic of Korea; Department of Biology, North-Eastern Federal University, 58 Belinsky Str., Yakutsk 677-027, Republic of Sakha (Yakutia), Russian Federation; Institute of Molecular Biology and Biotechnology, Azerbaijan National Academy of Sciences, PatamdarShosse 40, Baku AZ 1073, Azerbaijan.
authors.forename: Ismayil S
authors.initials: IS
authors.lastname: Zulfugarov
authors.affiliation: Department of Integrated Biological Science, Department of Molecular Biology, Pusan National University, Busan 609-735, Republic of Korea. Electronic address: chlee@pusan.ac.kr.
authors.forename: Choon-Hwan
authors.initials: CH
authors.lastname: Lee
mesh_terms: D055786:Gene Knockout Techniques; D006861:Hydrogen Peroxide / Q000378:metabolism; D008027:Light; D045342:Light-Harvesting Protein Complexes / Q000378:metabolism*; D009154:Mutation; D012275:Oryza / Q000166:cytology / Q000201:enzymology / Q000235:genetics* / Q000528:radiation effects; D045332:Photosystem II Protein Complex / Q000378:metabolism*; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D011494:Protein Kinases / Q000172:deficiency / Q000235:genetics*; D013481:Superoxides / Q000378:metabolism*; D020524:Thylakoids / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects
chemical_list: D045342:Light-Harvesting Protein Complexes; D045332:Photosystem II Protein Complex; D013481:Superoxides; D006861:Hydrogen Peroxide; D011494:Protein Kinases
keywords: High light; Photooxidation; Photosystem II; Rice; STN8 kinase; Superoxide