Leaf senescence in a stay-green mutant of Arabidopsis thaliana, ore10, was investigated during dark-incubation of its detached leaves. During this dark-induced senescence (DIS), Chl loss was delayed in ore10 mutants, as compared with wild type, but the rate of decline in the photochemical efficiency of PSII was not delayed in mutant leaves. After 2 d of DIS, native green gel electrophoresis of ore 10 leaf proteins resulted in a significant amount of pigment remaining as aggregates on top of the stacking gel. In addition, the accumulation of aggregates coincided with the emergence of a new band near 700 nm (F 699 ) in the 77 K fluorescence emission spectrum of the aggregates. At 4 d, F 699 became a major band, both in the isolated aggregates and in intact leaves. Prolonged treatment with detergents revealed that light-harvesting complex II (LHCII) remaining after 2 d was highly stable, and the accumulation of aggregates coincided with the appearance of truncated LHCII in senescing ore10 leaves. These results suggest that increased LHCII stability is due to the formation of aggregates of trimmed LHCII. Thus, the LHCII protein degradation step that follows proteolysis of its terminal peptides is a possible lesion site of the ore10 mutant.

We attempted to obtain a transcriptomic profile of ionizing radiation-responsive genes in Arabidopsis plants using Affymetrix ATH1 whole-genome microarrays. The Arabidopsis plants were irradiated with 200 Gy gamma rays at the early reproduction stage, 33 days after sowing. Rosette leaves were harvested during the postirradiation period from 36 to 49 days after sowing and used for the microarray analysis. The most remarkable changes in the genome-wide expression were observed at 42 days after sowing (9 days after the irradiation). We identified 2165 genes as gamma-ray inducible and 1735 genes as gamma-ray repressible. These numbers of affected genes were almost two to seven times higher than those at other times. In a comparison of the control and irradiated groups, we also identified 354 differentially expressed genes as significant by applying Welch's t test and fold change analysis. The gene ontology analysis showed that radiation up-regulated defense/ stress responses but down-regulated rhythm/growth responses. Specific expression patterns of 10 genes for antioxidant enzymes, photosynthesis or chlorophyll synthesis after irradiation were also obtained using real-time quantitative PCR analysis. We discuss physiological and genetic alterations in the antioxidative defense system, photosynthesis and chlorophyll metabolism after irradiation at the reproductive stage.

Photosystem I (PSI) is severely damaged by chilling at 4 degrees C in low light, especially in the chilling sensitive plant cucumber. To investigate the early events in PSI photoinhibition, we examined structural changes in the level of pigment-protein complexes in cucumber leaves in comparison with pea leaves. The complexes were separated on a native green gel and an increase in the intensity of a band was observed only in light-chilled cucumber leaves. The 77 K fluorescence emission spectrum of this green band indicated that the band was mainly composed of PSI with light-harvesting complex I. Each lane was cut from the green gel and separated on a fully denaturing SDS-PAGE in the second dimension. The new green gel band observed after light-chilling in cucumber leaves lacked 19, 18, and 16.5 kDa polypeptides. These results suggest that light-chilling facilitates the release of three peripheral polypeptides as an early event of chilling stress in vivo, which results in the inactivation of PSI in intact cucumber leaves.

During dark-induced leaf senescence (DIS), the non-functional stay-green mutant orelO showed delayed chlorophyll (Chl) degradation and increased stability in its light-harvesting complex II (LHCII). These phenomena were closely related to the formation of aggregates that mainly consisted of terminal-truncated LHCII (Oh et al., 2003). The orelO mutant apparently lacks the protease needed to degrade the truncated LHCII. In wild-type 0NT) plants, protease was found in the thylakoid fraction, but not the soluble fraction. A similar experiment using dansylated LHCII revealed that the protease degraded both WT and ore10 LHCII, indicating that its stability in orelO perhaps did not result from a defect in the LHCII polypeptides themselves. Although protease activity was not present in non.senesced WT leaves, it was induced during DIS. It also was possible to diminish the high level of protease present in the thylakoids through high-salt washing, suggesting that this enzyme is extrinsically bound to the outer surface of the stromaexposed thylakoid regions.