Imaging of photo-oxidative stress responses in leaves.
High resolution digital imaging was used to identify sites of photo-oxidative stress responses in Arabidopsis leaves non-invasively, and to demonstrate the potential of using a suite of imaging techniques for the study of oxidative metabolism in planta. Tissue-specific photoinhibition of photosynthesis in individual chloroplasts in leaves was imaged by chlorophyll fluorescence microscopy. Singlet oxygen production was assessed by imaging the quenching of the fluorescence of dansyl-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrole (DanePy) that results from its reaction with singlet oxygen. Superoxide and hydrogen peroxide accumulation were visualized by the reduction of nitroblue tetrazolium (NBT) to formazan deposits and by polymerization with 3,3'-diaminobenzidine (DAB), respectively. Stress-induced expression of a gene involved with antioxidant metabolism was imaged from the bioluminescence from leaves of an Arabidopsis APX2-LUC transformant, which co-expresses an ascorbate peroxidase (APX2) with firefly luciferase. Singlet oxygen and superoxide production were found to be primarily located in mesophyll tissues whereas hydrogen peroxide accumulation and APX2 gene expression were primarily localized in the vascular tissues.
Fryer
Michael J
MJ
Department of Biological Sciences, University of Essex, Colchester, Essex CO4 3SQ, UK.
Oxborough
Kevin
K
Mullineaux
Phillip M
PM
Baker
Neil R
NR
D029681:Arabidopsis Proteins; D060387:Ascorbate Peroxidases; D002734:Chlorophyll / Q000528:radiation effects; D018506:Gene Expression Regulation, Plant / Q000528:radiation effects; D006861:Hydrogen Peroxide / Q000378:metabolism; D008027:Light; D018384:Oxidative Stress / Q000502:physiology*; D010100:Oxygen / Q000378:metabolism; D010544:Peroxidases / Q000235:genetics / Q000378:metabolism; D010788:Photosynthesis / Q000528:radiation effects; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism
D029681:Arabidopsis Proteins; D017382:Reactive Oxygen Species; D013481:Superoxides; D002734:Chlorophyll; D006861:Hydrogen Peroxide; D010544:Peroxidases; C555293:APX2 protein, Arabidopsis; D060387:Ascorbate Peroxidases; D010100:Oxygen