Norio Murata

We provide here an overview of the remarkable life and outstanding research of David (Dave) Charles Fork (March 4, 1929–December 13, 2021) in oxygenic photosynthesis. In the words of the late Jack Edgar Myers, he was a top ‘photosynthetiker’. His research dealt with novel findings on light absorption, excitation energy distribution, and redistribution among the two photosystems, electron transfer, and their relation to dynamic membrane change as affected by environmental changes, especially temperature. David was an attentive listener and a creative designer of experiments and instruments, and he was also great fun to work with. He is remembered here by his family, coworkers, and friends from around the world including Australia, France, Germany, Japan, Sweden, Israel, and USA.

Various compatible solutes enable plants to tolerate abiotic stress, and glycinebetaine (GB) is one of the most‐studied among such solutes. Early research on GB focused on the maintenance of cellular osmotic potential in plant cells. Subsequent genetically engineered synthesis of GB‐biosynthetic enzymes and studies of transgenic plants demonstrated that accumulation of GB increases tolerance of plants to various abiotic stresses at all stages of their life cycle. Such GB‐accumulating plants exhibit various advantageous traits, such as enlarged fruits and flowers and/or increased seed number under non‐stress conditions. However, levels of GB in transgenic GB‐accumulating plants are relatively low being, generally, in the millimolar range. Nonetheless, these low levels of GB confer considerable tolerance to various stresses, without necessarily contributing significantly to cellular osmotic potential. Moreover, low levels of GB, applied exogenously or generated by transgenes for GB biosynthesis, can induce the expression of certain stress‐responsive genes, including those for enzymes that scavenge reactive oxygen species. Thus, transgenic approaches that increase tolerance to abiotic stress have enhanced our understanding of mechanisms that protect plants against such stress.

In the late 1960s, I identified an aspect of the kinetics of chlorophyll fluorescence in algal cells that I was unable to explain in terms of photochemical quenching. I proposed a novel regulatory mechanism for the distribution of light energy to photosystems I and II, which is now known by the term of “state transitions.” I also examined the “cation-dependent redistribution of light energy” to photosystems I and II and the “energy-dependent quenching” of chlorophyll fluorescence. At that time, financial constraints prevented me from measuring the emission and action spectra of chlorophyll fluorescence at liquid-nitrogen temperature and the light quality-dependent changes in the yield of chlorophyll fluorescence at room temperature. The financial problems were solved by constructing several pieces of electronic equipment using skills obtained by repairing radios when I was a high-school and college student.

Recent studies of responses of cyanobacterial cells to salt stress have revealed that the NaCl-induced decline in the photosynthetic activities of photosystems II and I involves rapid and slow changes. The rapid decreases in the activities of both photosystems, which occur within a few minutes, are reversible and are associated with osmotic effects, which induce the efflux of water from the cytosol through water channels and rapidly increase intracellular concentrations of salts. Slower decreases in activity, which occur within hours, are irreversible and are associated with ionic effects that are due to the influx of Na⁺ and Cl⁻ ions through K⁺(Na⁺) channels and, probably, Cl⁻ channels, with resultant dissociation of extrinsic proteins from photosystems. In combination with light stress, salt stress significantly stimulates photoinhibition by inhibiting repair of photodamaged photosystem II. Tolerance of photosystems to salt stress can be enhanced by genetically engineered increases in the unsaturation of fatty acids in membrane lipids and by intracellular synthesis of compatible solutes, such as glucosylglycerol and glycinebetaine. In this review, we summarize recent progress in research on the effects of salt stress on photosynthesis in cyanobacteria.

Since the first identification of phosphatidylglycerol in Scenedesmus by Benson and Maruo in 1958, researchers have studied many biological functions of this phospholipid. Genetic, biochemical, and structural studies of photosynthetic organisms have revealed that phosphatidylglycerol is crucial to the photosynthetic transport of electrons, the development of chloroplasts, and tolerance to chilling. In this review, we summarize our present understanding of the biochemical and physiological functions of phosphatidylglycerol in cyanobacteria and higher plants.

Recent investigations of photoinhibition have revealed that photodamage to photosystem II (PSII) involves two temporally separated steps: the first is the inactivation of the oxygen-evolving complex by light that has been absorbed by the manganese cluster and the second is the impairment of the photochemical reaction center by light that has been absorbed by chlorophyll. Our studies of photoinhibition in Synechocystis sp. PCC 6803 at various temperatures demonstrated that the first step in photodamage is not completed at low temperatures, such as 10°C. Further investigations suggested that an intermediate state, which is stabilized at low temperatures, might exist at the first stage of photodamage. The repair of PSII involves many steps, including degradation and removal of the D1 protein, synthesis de novo of the precursor to the D1 protein, assembly of the PSII complex, and processing of the precursor to the D1 protein. Detailed analysis of photodamage and repair at various temperatures has demonstrated that, among these steps, only the synthesis of the precursor to D1 appears to proceed at low temperatures. Investigations of photoinhibition at low temperatures have also indicated that prolonged exposure of cyanobacterial cells or plant leaves to strong light diminishes their ability to repair PSII. Such non-repairable photoinhibition is caused by inhibition of the processing of the precursor to the D1 protein after prolonged illumination with strong light at low temperatures.

Summary: The tolerance of various species of plant to abiotic stress has been enhanced by genetic engineering with certain genes. However, the use of such transgenes is often associated with negative effects on growth and productivity under non‐stress conditions. The codA gene from Arthrobacter globiformis is of particular interest with respect to the engineering of desirable productive traits in crop plants. The expression of this gene in tomato plants resulted in significantly enlarged flowers and fruits under non‐stress conditions. The enlargement of flowers and fruits was associated with high levels of glycinebetaine that accumulated in reproductive organs, such as flower buds and fruits. The enlargement of flowers was related to an increase in the size and number of cells, and reflected the pleiotropic effect of the codA transgene on the expression of genes involved in the regulation of cell division.

Summary: Oxidative stress caused by reactive oxygen species and, in particular, to hydrogen peroxide (H₂O₂) has a major impact on all biological systems, including plants and microorganisms. We investigated the H₂O₂‐inducible expression of genes in the cyanobacterium Synechocystis sp. PCC 6803 using genome‐wide DNA microarrays. Our systematic screening of a library of mutant lines with defects in histidine kinases (Hiks) by RNA slot‐blot hybridization and DNA‐microarray analysis suggested that four Hiks, namely, Hik33, Hik34, Hik16 and Hik41, are involved in the perception and transduction of H₂O₂ signals that regulate the gene expression of 26 of the 77 H₂O₂‐inducible genes with induction factors higher than 4.0. Among the four Hiks, Hik33 was the main contributor and was responsible for 22 of the 26 H₂O₂‐inducible genes under the control of the Hiks. By contrast to Hik33, PerR encoding putative peroxide‐sensing protein is involved in the regulation of only nine H₂O₂‐inducible genes.