17 O of the O 2 evolved were essentially identical to those of the substrate water. The fractionation slopes for the oxygenase reaction of Rubisco and respiration were identical (0.518 6 0.001) and that of glycolate oxidation was 0.503 6 0.002. There was a considerable difference in the slopes of O 2 photoreduction (the Mehler reaction) in the cyanobacterium Synechocystis sp. strain PCC 6803 (0.497 6 0.004) and that of pea (Pisum sativum) thylakoids (0.526 6 0.001). These values provided clear and independent evidence that the mechanism of O 2 photoreduction differs between higher plants and cyanobacteria. We used our method to assess the magnitude of O 2 photoreduction in cyanobacterial cells maintained under conditions where photorespiration was negligible. It was found that electron flow to O 2 can be as high as 40% that leaving photosystem II, whereas respiratory activity in the light is only 6%. The implications of our findings to the evaluation of specific O 2 -producing or -consuming reactions, in vivo, are discussed. O/ 16 O ratios in air bubbles in polar ice cores or dissolved O 2 in water bodies have been used to assess the photosynthetic rates on broad spatial and temporal scales (Luz et al., 1999;Luz and Barkan, 2000;Blunier et al., 2002). To accurately assess these rates, it is essential to know, with high precision, the isotope fractionation effects due to biological producing and consuming mechanisms. To date, precise measurements of these fractionations are available only for oxygen uptake in dark respiration by the cytochrome oxidase and the alternative oxidase pathways (Angert et al., 2003;Luz and Barkan, 2005 (Helman et al., 2003). In higher plants, electron transfer from PSI to oxygen, either from the Fx center of PSI or ferredoxin (Foyer and Noctor, 2000), results in the formation of superoxide. The latter is disproportionated by superoxide dismutase, and the H 2 O 2 produced is reduced to water by ascorbate peroxidase (Asada, 1999(Asada, , 2000. In cyanobacteria, NADPH donates electrons to A-type flavoproteins that reduce the oxygen directly to water without the formation of reactive oxygen intermediates (Vicente et al., 2002;Helman et al., 2003). Hence, the isotopic fractionation during photoreduction might differ between plants and cyanobacteria.In this study, we measured the triple isotopic O 2 fractionation by oxygen-producing and -consuming reactions in order to assess the overall photosynthetic oxygen production and the extent of these reactions in vivo. We show that the Mehler reaction of cyanobacteria has a different triple isotopic signature of oxygen compared to other oxygen-consuming processes. Thus, besides serving as an important tool for measurements of photosynthetic productivity, the relationship between the three oxygen isotopes can be used to assess the magnitude of the Mehler reaction under natural conditions. Determination of Triple Isotope Fractionations Fractionations during O 2 Uptake in the Absence of PhotosynthesisIn cases where O 2 -consuming reaction occurs withou...

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Some of the oxygen produced during oxygenic photosynthesis is consumed but little is known about the extent of the processes involved. We measured the (17)O/(16)O and (18)O/(16)O ratios in O(2) produced by certain marine and freshwater phytoplankton representing important groups of primary producers. When the cells were performing photosynthesis under very low dissolved oxygen concentrations (< 3 μM), we observed significant enrichment in both (18)O and (17)O with respect to the substrate water. The difference in δ(18)O between O(2) and water was about 4.5, 3, 5.5, and 7‱ in the diatom Phaeodactylum tricornutum, Nannochloropsis sp. (Eustigmatophyceae), the coccolithophore Emiliania huxleyi and the green alga Chlamydomonas reinhardtii, respectively. The difference in δ(17)O was about 0.52 that of δ(18)O. As explained, the observed enrichments most probably stem from considerable oxygen consumption during photosynthesis even when major O(2)-consuming reactions such as photorespiration were minimized. These enrichments increased linearly with rising O(2) levels but with different δ(17)O/δ(18)O slopes for the various organisms, suggesting engagements of different O(2)-consuming reactions with rising O(2) levels. Consumption of O(2) may be important for energy dissipation during photosynthesis. The isotope enrichment observed here, not accounted for in earlier assessments, closes an important gap in our understanding of the difference between the isotopic compositions of atmospheric oxygen and that of seawater, i.e., the Dole effect.

Diatoms are an important group of primary producers in the aquatic environment. They are able to acclimate to fast changes in the light intensity by various mechanisms including a rise in non-photochemical fluorescence quenching (NPQ). The latter has been attributed to the xanthophyll cycle (XC) following activation of diadinoxanthin de-epoxidase by the acidification of the thylakoid lumen. To examine whether fluorescence quenching in the diatom Phaeodactylum tricornutum depends on the DeltapH generated by the photosynthetic electron transport, we arrested the latter by 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU). This treatment hardly affected the NPQ or XC, even when methylviologen was present. Dissipation of the DeltapH by 2,4-dinitrophenol inhibited the XC but did not alter NPQ. Similar results, i.e. inhibition of the XC but normal fluorescence quenching, were observed when the experiments were performed at 3 degrees C. Measurements of thermoluminescence showed that excess light treatment caused a marked decline in the signals obtained as a result of recombination of Q(B) (-) with the S(3) state of the Mn cluster; this was also observed in cells treated with DCMU (recombination of Q(A) (-) with S(2)). Light treatment also diminished the Q(A) (-) re-oxidation signals. The data suggest that changes in PSII core centre itself due to exposure to excess light conditions play an important part in the acclimation of P. tricornutum to the changing light conditions.

Regulated programmed cell death (PCD) processes have been documented in several phytoplankton species and are hypothesized to play a role in population dynamics. However, the mechanisms leading to the coordinated collapse of phytoplankton blooms are poorly understood. We showed that the collapse of the annual bloom of Peridinium gatunense, an abundant dinoflagellate in Lake Kinneret, Israel, is initiated by CO2 limitation followed by oxidative stress that triggers a PCD-like cascade. We provide evidences that a protease excreted by senescing P. gatunense cells sensitizes younger cells to oxidative stress and may consequently trigger synchronized cell death of the population. Ageing of the P. gatunense cultures was characterized by a remarkable rise in DNA fragmentation and enhanced sensitivity to H2O2. Exposure of logarithmic phase (young) cultures to conditioning media from stationary phase (old) cells sensitized them to H2O2 and led to premature massive cell death. We detected the induction of specific extracellular protease activity, leupeptin-sensitive, in ageing cultures and in lake waters during the succession of the P. gatunense bloom. Partial purification of the conditioned media revealed that this protease activity is responsible for the higher susceptibility of young cells to oxidative stress. Inhibition of the protease activity lowered the sensitivity to oxidative stress, whereas application of papain to logarithmic phase P. gatunense cultures mimicked the effect of the spent media and enhanced cell death. We propose a novel mechanistic framework by which a population of unicellular phytoplankton orchestrates a coordinated response to stress, thereby determine the fate of its individuals.