Energy storage by cyclic electron flow through photosystem I (PSI) was measured in vivo using the photoacoustic technique. A wide variety of photosynthetic organisms were considered and all showed measurable energy storage by PSI-cyclic electron flow except for higher plants using the C-3 carbon fixation pathway. The capacity for energy storage by PSI-cyclic electron flow alone was found to be small in comparison to that of linear and cyclic electron flows combined but may be significant, nonetheless, under conditions when photosystem 11 is damaged, particularly in cyanobacteria. Light-induced dynamics of energy storage by PSI-cyclic electron flow were evident, demonstrating regulation under changing environmental conditions.The oxygen-evolving photosynthetic apparatus contains two reaction center complexes, designated PSI electron flow through PSI in C3 plants has been proposed to generate ATP over and above that produced by linear electron flow, adjusting the ratio of ATP to NADPH generated by the light reactions of photosynthesis in accordance with the needs of the plant (1). As an example, cyclic electron flow through PSI has been proposed as a source of ATP for repair of PSII units damaged by environmental stress, since PSI is typically much less susceptible to stress than PSII (8, 9). Much of the ambiguity concerning the function and significance of cyclic electron flow through PSI is due to the difficulty of measuring it in whole cells and tissues. Most previous studies ofPSI-cyclic electron flow and accompanying phosphorylation have necessarily used either in vitro measurements of thylakoid fragments or somewhat ambiguous light-induced absorbance changes in whole cells or tissues (12,17). The photoacoustic method for measurement of photosynthetic energy storage is well suited for study of PSI-cyclic electron flow, however, because it is capable of simple, direct, and quantitative measurement of energy storage by cyclic electron flow in intact leaves and algae as well as in thylakoid preparations (9,10,16,18). Presumably, the bulk of such energy storage by PSI-cyclic electron flow represents photophosphorylation of ADP.In the present report, the occurrence, capacity, and regulation of energy storage by PSI-cyclic electron flow in whole tissues of a variety of photosynthetic organisms are characterized using the photoacoustic method. MATERIALS AND METHODS Plant Material

NifS-like proteins provide the sulfur (S) for the formation of iron-sulfur (Fe-S) clusters, an ancient and essential type of cofactor found in all three domains of life. Plants are known to contain two distinct NifS-like proteins, localized in the mitochondria (MtNifS) and the chloroplast (CpNifS). In the chloroplast, five different Fe-S cluster types are required in various proteins. These plastid Fe-S proteins are involved in a variety of biochemical pathways including photosynthetic electron transport and nitrogen and sulfur assimilation. In vitro, the chloroplastic cysteine desulfurase CpNifS can release elemental sulfur from cysteine for Fe-S cluster biogenesis in ferredoxin. However, because of the lack of a suitable mutant allele, the role of CpNifS has not been studied thus far in planta. To study the role of CpNifS in Fe-S cluster biogenesis in vivo, the gene was silenced by using an inducible RNAi (interference) approach. Plants with reduced CpNifS expression exhibited chlorosis, a disorganized chloroplast structure, and stunted growth and eventually became necrotic and died before seed set. Photosynthetic electron transport and carbon dioxide assimilation were severely impaired in the silenced plant lines. The silencing of CpNifS decreased the abundance of all chloroplastic Fe-S proteins tested, representing all five Fe-S cluster types. Mitochondrial Fe-S proteins and respiration were not affected, suggesting that mitochondrial and chloroplastic Fe-S assembly operate independently. These findings indicate that CpNifS is necessary for the maturation of all plastidic Fe-S proteins and, thus, essential for plant growth.Fe-S proteins ͉ inducible RNAi ͉ photosynthesis ͉ Arabidopsis thaliana

The enzyme superoxide dismutase is ubiquitous in aerobic organisms where it plays a major role in alleviating oxygen-radical toxicity. An insertion mutation introduced into the iron superoxide dismutase locus (designated sodE) of the cyanobacterium Synechococcus sp. PCC 7942 created a mutant strain devoid of detectable iron superoxide dismutase activity. Both wild-type and mutant strains exhibited similar photosynthetic activity and viability when grown with 17 jmolm-2-s'1 illumination in liquid culture supplemented with 3% carbon dioxide. In contrast, the sodB mutant exhibited significantly greater damage to its photosynthetic system than the wild-type strain when grown under increased oxygen tension or with methyl viologen. Although damage occurs at both photosystems I and II, it is primarily localized at photosystem I in the sodB mutant. Growth in 100% molecular oxygen for 24 hr decreased photoacoustically measured energy storage in 3-(3,4-dichlorophenyl)-1,1-dimethylurea and abolished the fluorescence state 2 to state 1 transition in the sodB mutant, indicating interruption of cyclic electron flow around photosystem I. Analysis of the flash-induced absorption transient at 705 nm indicated that the interruption of cyclic electron flow occurred in the return part of the cycle, between the two [4 Fe-4 SI centers of photosystem I, FA and FB, and cytochrome f. Even though the sodB mutant was more sensitive to damage by active oxygen than wild-type cells, both strains were equally sensitive to the photoinhibition of photosystem II caused by exposure to strong light.

Light-mediated chloroplast movements are common in plants. When leaves of Alocasia brisbanensis (F.M. Bailey) Domin are exposed to dim light, mesophyll chloroplasts spread along the periclinal walls normal to the light, maximizing absorbance. Under high light, the chloroplasts move to anticlinal walls. It has been proposed that movement to the high-light position shortens the diffusion path for CO 2 from the intercellular air spaces to the chloroplasts, thus reducing CO 2 limitation of photosynthesis. To test this hypothesis, we used pulsed photoacoustics to measure oxygen diffusion times as a proxy for CO 2 diffusion in leaf cells. We found no evidence that chloroplast movement to the high-light position enhanced gas diffusion. Times for oxygen diffusion were not shorter in leaves pretreated with white light, which induced chloroplast movement to the high-light position, compared with leaves pretreated with 500 to 700 nm light, which did not induce movement. From the oxygen diffusion time and the diffusion distance from chloroplasts to the intercellular gas space, we calculated an oxygen permeability of 2.25 ϫ 10 Ϫ6 cm 2 s Ϫ1 for leaf cells at 20°C. When leaf temperature was varied from 5°C to 40°C, the permeability for oxygen increased between 5°C and 20°C but changed little between 20°C and 40°C, indicating changes in viscosity or other physical parameters of leaf cells above 20°C. Resistance for CO 2 estimated from oxygen permeability was in good agreement with published values, validating photoacoustics as another way of assessing internal resistances to CO 2 diffusion.Light-mediated chloroplast movements in the leaves of some plants are so striking that they create patterns visible to the naked eye. They have attracted the attention of plant physiologists for more than a century. Chloroplast movements of all types have been the subject of numerous reviews (Britz, 1979; Haupt and Scheuerlein, 1990; Wada et al., 1993; Yatsuhashi, 1996; Haupt, 1999; Wada and Kagawa, 2001; Kagawa and Wada, 2002). In leaves, chloroplasts spread along the periclinal walls of mesophyll cells (the face position) in low light, whereas in high light, they move toward the anticlinal walls (the profile position), effectively forming cylinders of chloroplasts in palisade tissue. In darkness, the chloroplasts are generally in an intermediate position, although this varies among species (Inoue and Shibata, 1974) and depends on the growth environment (Trojan and Gabrys, 1996). Because of the optical sieve effect (Britz and Briggs, 1987), these marked chloroplast movements alter light absorption (Zurzycki, 1961), which, in many leaves, gives rise to the visible color changes that have attracted attention for so long.Chloroplast movements are widespread in algae, mosses, ferns, and seed plants. Among seed plants, they are common in both monocots and dicots, and they occur in plants with widely differing leaf anatomy, from submerged aquatic plants (Zurzycki and Lelatko, 1969) to sclerophyllous evergreens (Del Hierro et al., 2000). Chl...