|Title||Light harvesting, light adaptation and photoprotection in aquatic photosynthesis studies by time-resolved fluorescence spectroscopy|
|Source||Wageningen University. Promotor(en): Herbert van Amerongen. - Wageningen : Wageningen University - ISBN 9789462572454 - 190|
|Publication type||Dissertation, internally prepared|
|Keyword(s)||licht - adaptatie - bacillariophyta - cyanobacteriën - verdedigingsmechanismen - fotochemie - spectrofluorimetrie - light harvesting complexen - light - adaptation - bacillariophyta - cyanobacteria - defence mechanisms - photochemistry - spectrofluorimetry - light harvesting complexes|
|Categories||Algae / Analytical Chemistry|
Aquatic photosynthetic organisms unavoidably experience light fluctuations that vary in amplitude, duration and origin, compromising their photosynthetic efficiency. Weather conditions and underwater flow cause continuous changes in irradiance to which the organisms have to adapt. Many light-adaptation strategies of photosynthetic organisms, such as light acclimation, photoprotection and state transitions are still not well understood. In this thesis, time-resolved fluorescence spectroscopy is used to obtain insight into the response of diatoms and cyanobacteria, both aquatic photosynthetic organisms, to changing light conditions.
In chapter 2, photoacclimation (long-term acclimation to irradiance conditions) of the diatom Cyclotella meneghiniana is discussed. It is shown that the diatom cells fine-tune the amount of absorbed light energy by modifying their antenna size: cells grown in high light intensity have smaller antennas than those grown in low light. At the same time, the increase of growth light intensity leads to a decrease of the relative amount of photosystem I (PSI) as compared to PSII. Such a strategy might be beneficial for diatoms, since they are known to have an electron transfer cycle around PS II to release excess electrons produced in high light intensities. Besides discussing photoacclimation, we give a detailed description the fluorescence kinetics in C. meneghiniana. It is concluded that the diatom antenna, represented by light-harvesting fucoxanthin chlorophyll proteins (FCPs), transfer their excitation energy predominantly to PSII. FCPs associated with PSII are slightly richer in red-absorbing fucoxanthin than the FCPs associated with PSI, suggesting that PSII antennas (partly) constitute the antenna form FCPb (i.e. oligomeric antenna complexes).
In chapter 3 the process of non-photochemical quenching (NPQ, thermal dissipation of excess absorbed light energy) of chlorophyll a fluorescence was studied in the same diatom species. Diatoms can rapidly switch on/off NPQ to respond to fast light-intensity changes in moving waters. They are capable to induce higher NPQ values than plants or other photosynthetic organisms. The reason for such high NPQ values, however, is not clear. We performed picosecond fluorescence measurements at 77K on cells locked in three different states: Besides using conventional unquenched and quenched states of the cells (in the absence and presence of the total NPQ component, respectively), we also performed measurements on the dark-adapted state directly following NPQ. In this state, diatoxantin (Dtx, a carotenoid related to NPQ), accumulated during the NPQ period and Dtx-related NPQ persists, while ΔpH-related NPQ has relaxed. In this way we revealed the following sequence of events during full development of NPQ. First, the pH gradient across the thylakoid membrane induces quenching of FCP trimers (FCPa complexes), while they are still part of PSII. This is followed by (partial) detachment of FCPa from PSII after which quenching persists. The pH gradient also causes the formation of Dtx, which leads to further quenching of isolated PSII cores and some aggregated FCPa. To summarize, quenching of PSII -both cores and complexes- and FCPa substantially contribute to NPQ in diatoms. The FCPb antenna form on the other hand does not contribute to the NPQ process.
Certain aquatic photosynthetic organisms, such as cyanobacteria and green algae, can also cope with changing light conditions by dynamically varying the relative antenna size of PSI and of PSII. Consequently, a redistribution of light energy between the PSs is achieved. This phenomenon is called “state transitions”. It is known to be driven via a change in the redox status of electron carriers between PSII and PSI. In cyanobacteria, this redox change can be achieved via dark-light transitions. However, the cascade of microscopic events that lead to subsequent energy redistribution in cyanobacteria is still not completely clear. In chapter 4, a study on dark-light transitions using the cyanobacterium Synechocystis sp. PCC 6803 as a model organism is described. It is demonstrated that during dark to light transitions, there is mainly detachment of phycobilisomes (PBSs) (cyanobacterial antennas) from PSI, generally not followed by their attachment to PSII: only 15 % of the PBSs that detach from PSI actually move to PSII, while the major part remains detached from both PSs. We conclude that PSI-PSII-PBS megacomplexes, which were recently isolated using chemical cross-linking, are not involved in dark/light state transitions, suggesting that, if present, they are only transiently formed in cyanobacteria. To summarize, the findings presented in chapter 4 suggest that in cyanobacteria, unlike in green algae or higher plants, the main role of state transitions is to change the absorption cross-section of PSI, rather than that of PSII.
In chapter 5, a study of the role of flv4-2 operon-encoded proteins in Synechocystis is described. Three genes are found in the operon: Flv4, Sll0218, and Flv2. Only recently flv4-2 operon-encoded proteins were found to constitute an additional photoprotective mechanism in a number of cyanobacteria by safeguarding PSII activity via an alternative electron chain. Its contribution becomes vital for the cells in high light and in air-level CO2, when the photosynthetic electron transport chain is over-reduced. It is demonstrated that deletion of the operon induces 20% PBS detachment. The reduced PSII dimer to monomer ratio, as a result of the absence of the small Sll0218 protein, favors a relative decrease of the PSII dimer content of about 20%, showing a direct correlation between PSII dimer destabilization and PBS detachment from reaction centers. On the other hand, the suggested binding of the Flv2/Flv4 heterodimer closely to the quinone B (QB) pocket in PSII increases the QB redox potential, thereby promoting forward electron transfer and increasing the charge separation rates in PSII. This activity of the Flv2/Flv4 heterodimer in combination with its earlier reported role as an electron acceptor in alternative electron chain provides more oxidized state of the PQ pool in high light and in air-level CO2.