Kiriakos Kotzabasis

Polyamines in the photosynthetic apparatus : Photosystem II highly resolved subcomplexes are enriched in spermine.

The three main polyamines putrescine (Put), spermidine (Spd) and spermine (Spm) were characterized by HPLC in intact spinach leaf cells, intact chloroplasts, thylakoid membranes, Photosystem II membranes, the light-harvesting complex and the PS II complex. All contain the three polyamines in various ratios; the HPLC polyamine profiles of highly resolved PS II species (a Photosystem II core and the rection center) suggest an enrichment in the polyamine Spm.

Ozone impact on the photosynthetic apparatus and the protective role of polyamines

One of the primary plant mechanisms protecting leaf cells against enhanced atmospheric ozone is the accumulation of polyamines, generally observed as an increase in putrescine level, and in particular its bound form to thylakoid membranes. Ozone-sensitive plants of tobacco (cultivar Bel W3) in contrast to ozone-tolerant Bel B, are not able to increase their endogenous thylakoid membrane-bound putrescine when they are exposed to an atmosphere with enhanced ozone concentration, resulting in reduction of their photosynthetic rates and consequently reduction in plant biomass formation. In comparison to the tolerant cultivar Bel B, a prolongation of ozone exposure thus can lead to typical visible symptoms (necrotic spots) in leaves of the sensitive plant. Exogenously manipulated increase of the cellular putrescine levels of the ozone-sensitive Bel W3 is sufficient to revert these effects, whereas a reduction in endogenous putrescine levels of the tolerant cultivar Bel B renders them sensitive to ozone treatment. The results of this work reveal a regulator role for polyamines in adaptation of the photosynthetic apparatus and consequently to its protection in an environment polluted by ozone.

Methanol as alternative carbon source for quicker efficient production of the microalgae Chlorella minutissima: Role of the concentration and frequence of administration

Autotrophic cultures of the marine microalgae Chlorella minutissima were performed at 13 000 lux continuous illumination in 1 l chambers fertilised with 0.25 g l−1 F2 medium and different doses of methanol. This was administered in two ways during two parallel experimental series of 10 days: 0.05, 0.1, 0.5, 1.0 and 5.0% methanol (v/v) in one unique dose at the beginning of the culture and 1/10 of these (i.e. 0.005, 0.01, 0.05, 0.1 and 0.5% methanol (v/v)) in daily doses for the 10-day culture period. Low concentrations of methanol induced a faster increase of cell density and dry weight than control, while high concentrations induced symptoms of toxicity. The higher cell densities and quicker growth were observed in the experiments with daily administration of 0.005 and 0.1% (v/v) methanol, while those with one dose presented an initial boosted growth but a final cell density lower than control. The role of methanol as alternative carbon source for microalgae, as well as its possible impact on the quality of biomass production and on the environment, are discussed.

Remodeling of tobacco thylakoids by over-expression of maize plastidial transglutaminase

Transglutaminases (TGases, EC 2.3.2.13) are intra- and extra-cellular enzymes that catalyze post-translational modification of proteins by establishing epsilon-(gamma-glutamyl) links and covalent conjugation of polyamines. In chloroplast it is well established that TGases specifically polyaminylate the light-harvesting antenna of Photosystem (PS) II (LHCII, CP29, CP26, CP24) and therefore a role in photosynthesis has been hypothesised (Della Mea et al. [23] and refs therein). However, the role of TGases in chloroplast is not yet fully understood. Here we report the effect of the over-expression of maize (Zea mays) chloroplast TGase in tobacco (Nicotiana tabacum var. Petit Havana) chloroplasts. The transglutaminase activity in over-expressers was increased 4 times in comparison to the wild-type tobacco plants, which in turn increased the thylakoid associated polyamines about 90%. Functional comparison between Wt tobacco and tgz over-expressers is shown in terms of fast fluorescence induction kinetics, non-photochemical quenching of the singlet excited state of chlorophyll a and antenna heterogeneity of PSII. Both in vivo probing and electron microscopy studies verified thylakoid remodeling. PSII antenna heterogeneity in vivo changes in the over-expressers to a great extent, with an increase of the centers located in grana-appressed regions (PSIIalpha) at the expense of centers located mainly in stroma thylakoids (PSIIbeta). A major increase in the granum size (i.e. increase of the number of stacked layers) with a concomitant decrease of stroma thylakoids is reported for the TGase over-expressers.

Evidence That Putrescine Modulates the Higher Plant Photosynthetic Proton Circuit

The light reactions of photosynthesis store energy in the form of an electrochemical gradient of protons, or proton motive force (pmf), comprised of electrical (Δψ) and osmotic (ΔpH) components. Both components can drive the synthesis of ATP at the chloroplast ATP synthase, but the ΔpH component also plays a key role in regulating photosynthesis, down-regulating the efficiency of light capture by photosynthetic antennae via the qE mechanism, and governing electron transfer at the cytochrome b6f complex. Differential partitioning of pmf into ΔpH and Δψ has been observed under environmental stresses and proposed as a mechanism for fine-tuning photosynthetic regulation, but the mechanism of this tuning is unknown. We show here that putrescine can alter the partitioning of pmf both in vivo (in Arabidopsis mutant lines and in Nicotiana wild type) and in vitro, suggesting that the endogenous titer of weak bases such as putrescine represents an unrecognized mechanism for regulating photosynthetic responses to the environment.

Influence of polyamine inhibitors on light-independent and light- dependent chlorophyll biosynthesis and on the photosynthetic rate

Abstract The mutant C-2A′ of the unicellular green alga Scenedesmus obliquus behaves at 32°C like higher plants in that it accumulates protochlorophyllide in darkness and converts it to chlorophyll when exposed to light. By lowering the growth temperature from 32 to 22°C, protochlorophyllide can be reduced in darkness. Thus this organism allowed us to study the effects of the inhibition of several enzymes of polyamine biosynthesis on light-dependent and light-independent chlorophyll biosynthesis. Reduction of the intracellular level of putrescine by the use of 1,4-diamino-2-butanone inhibitor blocked the light-independent chlorophyll biosynthesis (dramatically increasing the photochlorophyllide levels with parallel reduction of chlorophyll). The influence of the inhibitor in question is even more dramatic in light-dependent chlorophyll biosynthesis, where small concentrations of 1,4-diamino-2-butanone resulted in considerable reduction of chlorophyll during illumination. The above reduction of chlorophyll was accompanied by an increase in photosynthetic activity and respiration rate, which suggests the formation of a photosynthetic apparatus that behaves similarly to one adapted to high light intensities. Decreases in the intracellular levels of spermidine and spermine by the inhibitors cyclohexylamine and 1,3-diaminopropane respectively, although not showing changes in light-independent chlorophyll biosynthesis, do exhibit a regular reduction of light-dependent chlorophyll biosynthesis and a decrease in the photosynthetic activity as well as in the respiration rate. These characteristics are typical for a photosynthetic apparatus adapted to low light intensities with a higher antenna composition per reaction centre. The possible effects of the reduction of putrescine, spermidine and spermine levels on the stabilization-destabilization of photosynthetic subcomplexes are discussed.

The regulatory role of polyamines in structure and functioning of the photosynthetic apparatus during photoadaptation

In this contribution we describe the changes in structure and functioning of the photosynthetic apparatus of the unicellular green alga Scenedesmus obliquus induced by inhibition or induction of polyamine biosynthesis. The synthesis or inhibition is controlled by white light of low and high intensities, as well as by blue and red irradiation. We observe that a decrease of the intracellular putrescine level and, vice versa, an increase of spermine, indicated by a raised ratio of spermine to putrescine, simulate a low-light photoadapted photosynthetic apparatus. Action spectra of the spermine/putrescine ratio compared with action spectra on characteristics of photoadaptation, e.g., chlorophyll biosynthesis, or the rate of primary photochemistry and electron transport, show that the photoreceptors for both the adaptation of the photosynthetic apparatus to low-light conditions and for the regulation of the spermine/putrescine ratio are, at least, very similar. We show that the photoreceptor is primarily a blue-light receptor with a superimposed red-light receptor that absorbs around 680 nm.

Fast and reversible response of thylakoid-associated polyamines during and after UV-B stress: a comparative study of the wild type and a mutant lacking chlorophyll b of unicellular green alga Scenedesmus obliquus.

The functional and biochemical aspects of the photosynthetic apparatus in response to UV-B radiation were examined in unicellular oxygenic algae Scenedesmus obliquus. The wild type (Wt) and a chlorophyll b-less mutant (Wt-lhc) were used as a specific tool for the understanding of antenna role. Photosynthesis was monitored during and after UV-B stress by time resolved fluorescence spectroscopy and polarography. Carotenoids, such as neoxanthin, loroxanthin, lutein, violaxanthin, antheraxanthin, zeaxanthin, α- and β-carotene, cellular and thylakoid-associated putrescine, spermidine, spermine and subcomplexes of light-harvesting complex (LHCII) of photosystem II (PSII) were investigated to assess their possible involvement in response to UV-B. Oxygen evolution depression by UV-B was higher in the Wt-lhc mutant than in the Wt. Photosynthesis recovery occurred in the Wt, but not in the mutant. The dissipation of excess excitation energy during UV-B stress was accompanied by changes in the thylakoid-associated polyamines which were much higher than changes in xanthophylls. We conclude that, at least in the unicellular green alga S. obliquus, mutants lacking chlorophyll b have significant lower capacity for recovery after UV-B stress. In addition, the comparison of xanthophylls and thylakoid-associated polyamines reveals that the latter are more responsive to UV-B stress and in a reversible manner.

Changes in the biosynthesis and catabolism of polyamines in isolated plastids during chloroplast photodevelopment

Polyamines (PAs), diamine oxidase (DAd) activity and putrescine formation via ornithine (ornithine decarboxylase, ODC) and arginine ~arginine decarboxylase and arginasc pathway, ADC*) were found in isolated etioplasts, chloroplasts and the intermediate plastids of chloroplast photodevelopmcnt. The isolated etioplasts contain large amounts of PAs (putrcscinc (Put), spermidine (Spd). sperminc (Spin) ), and high activitics of the main PA biosynthetic enzymes ODC and ADC*, combincd with very low activity of the |nain PA catabolic enzyme DAd, were found, During the photodcvelopment of ctioplasts to chloroplasts, a two-step decrease of intraplastidic PAs, ODC and ADC* takes place: the tirst very fast and the second stronger after a very hmg exposure to light. The correlation tffthese two steps with the protochlorophyllide (PChlide) to chlorophyllide (Chlide) photoconvcrsion and the developmc|at of the photosynthetic apparatus is discussed. Our experiments indicate that the almost zero DAd activity in etioplasts increases rapidly and shows a maximum after exposure to light for 9 h, whereas the intracellular DAd activity remains almost unchanged. All the plastidic activities (ODC, ADC*, DAd) and the level of PAs ( Put, Spd, Spin) during the photodevclopment of chltm~plasts arc correlated with the corresponding intracellular values and the differences are discussed.

Effects of ammonia from livestock farming on lichen photosynthesis

This study investigated if atmospheric ammonia (NH3) pollution around a sheep farm influences the photosynthetic performance of the lichens Evernia prunastri and Pseudevernia furfuracea. Thalli of both species were transplanted for up to 30 days in a semi-arid region (Crete, Greece), at sites with concentrations of atmospheric ammonia of ca. 60 microg/m3 (at a sheep farm), ca. 15 microg/m3 (60 m from the sheep farm) and ca. 2 microg/m3 (a remote area 5 km away). Lichen photosynthesis was analysed by the chlorophyll a fluorescence emission to identify targets of ammonia pollution. The results indicated that the photosystem II of the two lichens exposed to NH(3) is susceptible to this pollutant in the gas-phase. The parameter PI(ABS), a global index of photosynthetic performance that combines in a single expression the three functional steps of the photosynthetic activity (light absorption, excitation energy trapping, and conversion of excitation energy to electron transport) was much more sensitive to NH3 than the FV/FM ratio, one of the most commonly used stress indicators.