FUNCTIONAL AND STRUCTURAL IMPORTANCE OF LIPIDS IN PHOTOSYNTHETIC MEMBRANES

The lipid composition of photosynthetic membranes is essential in the construction of the photosynthetic machinery and in developing resistance to various stress conditions. We use a combination of physiological, biophysical and molecular biological approaches to investigate the structural and functional importance of phosphatidylglycerol in the functions and the formation of the photosynthetic apparatus, the role of lipid-protein, lipid-carotenoid and carotenoid-protein interaction in stress responses.

The role of phosphatidylglycerol in photosynthetic electron transport processes

Among glycoglycerolipids, dominant lipids of photosynthetic membranes, there is only one phospholipid species, phosphatidylglycerol (PG). PG is an indispensable lipid in cyanobacteria and higher plants, which are able to carry out oxygenic photosynthesis. Crystallographic studies of protein complexes involved in photosynthesis have revealed the presence of PG molecules in the reaction centre (RC) of PSI and PSII complexes.

We generated several PG deficient mutants of various cyanobacterial strains. These PG mutants needed an exogenous supply of PG for their growth. The growth rate of the mutant cells in the presence of PG was hardly distinguishable from that of the wild-type; however, that of the mutant cells was gradually suppressed during PG depletion. Re-addition of PG molecules restored cell division (Fig. 1).

Short-term depletion of PG hardly influenced the activity of PSI; however, that of PSII was severely affected. PG depletion suppressed the electron transport in PSII around primary and secondary quinones, inhibited the assembly of the CP43 subunit to PSII RC and disturbed the formation of the functionally active dimers of PSII. Long-term PG depletion suppressed the oligomerization of PSI RCs (Fig. 2) and decreased the PSI activity of the PG-deficient cells.

We concluded that PG depletion affects cyanobacterial cell division and PG is necessary for the oligomerization and functioning of photosynthetic reaction centres.

We investigated the remodeling of PG molecules in PG-deficient mutant cells of Synechocystis PCC6803 without manipulation of the enzymes involved in this remodeling process. The exogenously added artificial dioleoyl-PG was transformed into photosynthetically more essential natural PG derivatives. Our experiments demonstrated remodeling of lipids in a prokaryotic photosynthetic bacterium.

Carotenoids, indispensable components for assembly and functions of photosystems

A gradual elimination of PG from the photosynthetic membranes of the mutant cells resulted in increased sensitivity to light, which resulted in photobleaching of photosynthetic pigments and ultimately to cell death. The induction of photobleaching can be explained by the formation of a triplet state of chlorophyll, which initiates the formation of reactive oxygen species that in turn induce the degradation of photosynthetic pigments in the PG-depleted cells. We demonstrated that in PG-deficient mutant Synechocystis PCC6803 cells, PG depletion induces an accumulation of myxoxanthophyll and echinenone in both thylakoid and cytoplasmic membranes. An increase in myxoxanthophyll and echinenone content upon PG depletion suggests that PG depletion regulates the biosynthetic pathway of specific carotenoids.

In order to get a better understanding of carotenoid involvement in photosynthetic functions, we constructed the first oxygenic photosynthetic prokaryotic mutant completely deficient in carotenoid (Car) synthesis.

Complete elimination of Cars (Fig. 3) did not significantly affect the assembly of PSI and cytochrome b₆f complexes. The carotenoid deficient mutant strain of Synechocystis PCC6803 did not have any oxygen-evolving activity. The cells became light sensitive. The lack of Cars blocked the construction of PSII RCs and the functions related to PSII protein complexes. Elimination of Cars does not affect severely the transcription of the studied photosynthetic RC genes, although the translation of protein subunits of PSII was remarkably suppressed.

We concluded that Cars are indispensable constituents of the photosynthetic apparatus and essential for protection of chlorophylls against photooxidation. Furthermore, they are also required for the synthesis and assembly of PSII subunits.

The role of membrane lipids in low temperature stress processes in wheat

Cell membranes are dynamic structures, which are constantly formed and degraded. Changes in the lipid composition of membranes play an essential role in the adaptation of living organisms to various environmental effects. Freezing temperature is one of the major abiotic stresses living organisms, especially plants, have to face. For example, the growing season and the geographical diversity of plant species are highly influenced by frost damage. The major food grain crop consumed by humans is bread wheat. Appropriate frost tolerance of winter wheat is essential for its safe production in countries under temperate climate. The primary sites of freezing injury in plants are cell membranes, especially the plasma membranes. Because of the importance of the plasma membrane in plant adaptation against the deleterious effects of low temperature, detailed analysis of the alterations of their lipids will substantially help our understanding of the role of the structural modifications of the membrane and the signals generated in the membrane.

Thus the major objective of this project is comprehensive lipid species profiling using special wheat genetic stocks such as substitution and recombinant lines. We hope to localize and map genes involved in the metabolism of lipids. This may enable us to provide useful molecular markers linked to the mapped genes, thus helping MAS.