Quality control of photosystem II: direct imaging of the changes in the thylakoid structure and distribution of FtsH proteases in spinach chloroplasts under light stress.

Under light stress, the reaction center-binding protein D1 of PSII is photo-oxidatively damaged and removed from PSII complexes by proteases located in the chloroplast. A protease considered to be responsible for degradation of the damaged D1 protein is the metalloprotease FtsH. We showed previously that the active hexameric FtsH protease is abundant at the grana margin and the grana end membranes, and this homo-complex removes the photodamaged D1 protein in the grana. Here, we showed a change in the distribution of FtsH in spinach thylakoids during excessive illumination by transmission electron microscopy (TEM) and immunogold labeling of FtsH. The change in distribution of the protease was accompanied by structural changes to the thylakoids, which we detected using spinach leaves by TEM after chemical fixation of the samples. Quantitative analyses showed several characteristic changes in the structure of the thylakoids, including shrinkage of the grana, outward bending of the marginal portions of the thylakoids and an increase in the height of the grana stacks under excessive illumination. The increase in the height of the grana stacks may include swelling of the thylakoids and an increase in the partition gaps between the thylakoids. These data strongly suggest that excessive illumination induces partial unstacking of the thylakoids, which enables FtsH to access easily the photodamaged D1 protein. Finally three-dimensional tomography of the grana was recorded to observe the effect of light stress on the overall structure of the thylakoids.

Quality control of PSII: behavior of PSII in the highly crowded grana thylakoids under excessive light.

The grana thylakoids of higher plant chloroplasts are crowded with PSII and the associated light-harvesting complexes (LHCIIs). They constitute supercomplexes, and often form semi-crystalline arrays in the grana. The crowded condition of the grana may be necessary for efficient trapping of excitation energy by LHCII under weak light, but it might hinder proper movement of LHCII necessary for reversible aggregation of LHCII in the energy-dependent quenching of Chl fluorescence under moderate high light. When the thylakoids are illuminated with extreme high light, the reaction center-binding D1 protein of PSII is photodamaged, and the damaged protein migrates to the grana margins for degradation and subsequent repair. In both moderate and extreme high-light conditions, fluidity of the thylakoid membrane is crucial. In this review, we first provide an overview of photoprotective processes, then discuss changes in membrane fluidity and mobility of the protein complexes in the grana under excessive light, which are closely associated with photoprotection of PSII. We hypothesize that reversible aggregation of LHCII, which is necessary to avoid light stress under moderate high light, and swift turnover of the photodamaged D1 protein under extreme high light are threatened by irreversible protein aggregation induced by reactive oxygen species in photochemical reactions.

Quality control of Photosystem II: reversible and irreversible protein aggregation decides the fate of Photosystem II under excessive illumination.

In response to excessive light, the thylakoid membranes of higher plant chloroplasts show dynamic changes including the degradation and reassembly of proteins, a change in the distribution of proteins, and large-scale structural changes such as unstacking of the grana. Here, we examined the aggregation of light-harvesting chlorophyll-protein complexes and Photosystem II core subunits of spinach thylakoid membranes under light stress with 77K chlorophyll fluorescence; aggregation of these proteins was found to proceed with increasing light intensity. Measurement of changes in the fluidity of thylakoid membranes with fluorescence polarization of diphenylhexatriene showed that membrane fluidity increased at a light intensity of 500-1,000 µmol photons m(-) (2) s(-) (1), and decreased at very high light intensity (1,500 µmol photons m(-) (2) s(-) (1)). The aggregation of light-harvesting complexes at moderately high light intensity is known to be reversible, while that of Photosystem II core subunits at extremely high light intensity is irreversible. It is likely that the reversibility of protein aggregation is closely related to membrane fluidity: increases in fluidity should stimulate reversible protein aggregation, whereas irreversible protein aggregation might decrease membrane fluidity. When spinach leaves were pre-illuminated with moderately high light intensity, the qE component of non-photochemical quenching and the optimum quantum yield of Photosystem II increased, indicating that Photosystem II/light-harvesting complexes rearranged in the thylakoid membranes to optimize Photosystem II activity. Transmission electron microscopy revealed that the thylakoids underwent partial unstacking under these light stress conditions. Thus, protein aggregation is involved in thylakoid dynamics and regulates photochemical reactions, thereby deciding the fate of Photosystem II.