Journal Article

Co-expression of Cytochrome <i>b</i> <sub>561</sub> and Ascorbate Oxidase in Leaves of Wild Watermelon under Drought and High Light Conditions

Yoshihiko Nanasato, Kinya Akashi and Akiho Yokota

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 46, issue 9, pages 1515-1524
Published in print September 2005 | ISSN: 0032-0781
Published online September 2005 | e-ISSN: 1471-9053 | DOI: https://dx.doi.org/10.1093/pcp/pci164
Co-expression of Cytochrome b 
561 and Ascorbate Oxidase in Leaves of Wild Watermelon under Drought and High Light Conditions

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Despite carrying out C3 photosynthesis, wild watermelon (Citrullus lanatus sp.) exhibits exceedingly good tolerance to severe drought at high light intensities. However, the mechanism(s) by which this plant protects itself from photodamage has yet to be elucidated. In this study, we characterized wild watermelon cytochrome b 561 (cyt b 561), which potentially mediates regeneration of apoplastic ascorbate by transferring electrons from cytosolic ascorbate across the plasma membrane. Two cDNA species for wild watermelon cyt b 561, designated CLb561A and CLb561B, were isolated. Levels of both CLb561A mRNA and protein were significantly elevated in the leaves during drought at a light intensity of 700 µmol photons m–2 s–1. The transcript of CLb561B was detected to a much lesser extent, but no CLb561B protein was produced under any condition used in this study. A transient expression assay with the CLb561A::green fluorescent protein fusion construct showed clear fluorescence on the plasma membrane of onion epidermal cells. The CLb561A protein was enriched in the plasma membrane fraction in leaves of transgenic tobacco expressing CLb561A. Moreover, the high activity of apoplastic ascorbate oxidase (AO), which was able to dispose of cyt b 561-transferred reducing equivalents, increased in leaves of wild watermelon grown at high light intensity, but not lower light intensities. Taken together, these observations suggest the occurrence of a novel pathway for excess light energy dissipation in wild watermelon leaves, where excessive energy absorbed by chloroplasts can be transported to and dissipated safely in the apoplasts through the cooperative action of cyt b 561 and AO.

Keywords: Ascorbate oxidase; Cytochrome b561; Drought tolerance; Excess energy dissipation; Oxidative stress; Wild watermelon (Citrullus lanatus sp.); AO, ascorbate oxidase; Asc, ascorbate; CaMV, cauliflower mosaic virus; DHA, dehydroascorbate; G6PDH, glucose-6-phosphate dehydrogenase; GFP, green fluorescent protein; GR, glutathione reductase; HL, high light (700 µmol photons m–2 s–1); IWF, intercellular washing fluid; LL, low light (100 µmol photons m–2 s–1); MDA, monodehydroascorbate; PEG, polyethylene glycol; PMSF, phenylmethylsulfonyl fluoride; PVPP, polyvinylpolypyrrolidone; ROS, reactive oxygen species

Journal Article.  7941 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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