Travels in a world of small science*

As a boy, I read Sinclair Lewis's Arrowsmithand dreamed of doing research of potential benefit to society. I describe the paths of my scientific career that followed. Several distinguished scientists served as my mentors and I present their profiles. Much of my career was in a small department at a small institution where independent researchers collaborated informally. I describe the unique method of carrying on research there. My curiosity about glycolate metabolism led to unraveling the enzymatic mechanism of the glycolate oxidase reaction and showing the importance of H₂O₂ as a byproduct. I discovered enzymes catalyzing the reduction of glyoxylate and hydroxypyruvate. I found α-hydroxysulfonates were useful competitive inhibitors of glycolate oxidase. In a moment of revelation, I realized that glycolate metabolism was an essential part of photorespiration, a process that lowers net photosynthesis in C₃ plants. I added inhibitors of glycolate oxidase to leaves and showed: (1) glycolate was synthesized only in light as an early product of photosynthetic CO₂ assimilation, (2) the rate of glycolate oxidation consumed a sizable fraction of net photosynthesis in C₃ but not in C₄ plants, and (3) that glycolate metabolism increased greatly at higher temperatures. For a while I studied the control of stomatal opening in leaves, and this led to the finding that potassium ions are a key solute in guard cells. I describe experiments that show that when photorespiration rates are high, as occurs at higher temperatures, genetically increasing leaf catalase activity reduces photorespiration and increases net photosythetic CO₂ assimilation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Enhanced Chlorella vulgaris Buitenzorg growth by photon flux density alteration in serial photobioreactors

Microalgae perform oxygenic photosynthesis and are capable of taking up a large amount of CO₂, using an inducible CO₂ concentrating mechanism (CCM), and fixing CO₂ into higher compounds. These characteristics make the microalgae potentially useful for removal and utilization of CO₂ emitted from industrial plants and, generally, the usage of photosynthetic microorganisms has increased and significantly improved as a solution for CO₂ emissions. In this light and based on previous research using Anabaena cylindrica IAM M1 and Spirulina platensis IAM M 135, enhancement was sought for CO₂ fixation and biomass production by Chlorella vulgaris Buitenzorg by increasing the photon flux density concurrent with increases in culture biomass during the cellular growth phase and was compared to cultures of Chlorella grown at optimal constant illumination, with all cultures grown using Bennick basal medium, 29°C, and a flow of 1.0 atm. 10% CO₂ enriched air delivered to three in serial photobioreactors of 0.200 dm³ capacity each. The results showed that increasing illumination during culture increased biomass production of Chlorella by ∼60% as well as increased CO₂ fixation ability by ∼7.0%. It was also demonstrated that the non-competitive inhibition of [HCO₃ ⁻] as a carbon source significantly affected the cultivation in both the increasing and constant photon flux density regimes.     

PHOTOSYNTHETIC UTILIZATION OF INORGANIC CARBON IN THE ECONOMIC BROWN ALGA,HIZIKIA FUSIFORME (SARGASSACEAE) FROM THE SOUTH CHINA SEA1

The mechanism of inorganic carbon (C_i) acquisition by the economic brown macroalga, Hizikia fusiforme (Harv.) Okamura (Sargassaceae), was investigated to characterize its photosynthetic physiology. Both intracellular and extracellular carbonic anhydrase (CA) were detected, with the external CA activity accounting for about 5% of the total. Hizikia fusiforme showed higher rates of photosynthetic oxygen evolution at alkaline pH than those theoretically derived from the rates of uncatalyzed CO₂ production from bicarbonate and exhibited a high pH compensation point (pH 9.66). The external CA inhibitor, acetazolamide, significantly depressed the photosynthetic oxygen evolution, whereas the anion‐exchanger inhibitor 4,4′‐diisothiocyano‐stilbene‐2,2′‐disulfonate had no inhibitory effect on it, implying the alga was capable of using HCO₃^? as a source of C_i for its photosynthesis via the mediation of the external CA. CO₂ concentrations in the culture media affected its photosynthetic properties. A high level of CO₂ (10,000 ppmv) resulted in a decrease in the external CA activity; however, a low CO₂ level (20 ppmv) led to no changes in the external CA activity but raised the intracellular CA activity. Parallel to the reduction in the external CA activity at the high CO₂ was a reduction in the photosynthetic CO₂ affinity. Decreased activity of the external CA in the high CO₂ grown samples led to reduced sensitiveness of photosynthesis to the addition of acetazolamide at alkaline pH. It was clearly indicated that H. fusiforme, which showed CO₂‐limited photosynthesis with the half‐saturating concentration of C_i exceeding that of seawater, did not operate active HCO₃^? uptake but used it via the extracellular CA for its photosynthetic carbon fixation.     

The air‐lift photobioreactors with flow patterning for high‐density cultures of microalgae and carbon dioxide removal

A photobioreactor containing microalgae is a highly efficient system for converting carbon dioxide (CO₂) into biomass. Using a microalgal photobioreactor as a CO₂ mitigation system is a practical approach to the problem of CO₂ emission from waste gas. In this study, a marine microalga, Chlorella sp. NCTU‐2, was applied to assess biomass production and CO₂ removal. Three types of photobioreactors were designed and used: (i) without inner column (i.e. a bubble column), (ii) with a centric‐tube column and (iii) with a porous centric‐tube column. The specific growth rates (μ) of the batch cultures in the bubble column, the centric‐tube and the porous centric‐tube photobioreactor were 0.180, 0.226 and 0.252 day^?1, respectively. The porous centric‐tube photobioreactor, operated in semicontinuous culture mode with 10% CO₂ aeration, was evaluated. The results show that the maximum biomass productivity was 0.61 g/L when one fourth of the culture broth was recovered every 2 days. The CO₂ removal efficiency was also determined by measuring the influent and effluent loads at different aeration rates and cell densities of Chlorella sp. NCTU‐2. The results show that the CO₂ removal efficiency was related to biomass concentration and aeration rate. The maximum CO₂ removal efficiency of the Chlorella sp. NCTU‐2 culture was 63% when the biomass was maintained at 5.15 g/L concentration and 0.125 vvm aeration (volume gas per volume broth per min; 10% CO₂ in the aeration gas) in the porous centric‐tube photobioreactor.     

Enzyme activities of the carbon reduction cycle in some photosynthetic organisms

Profile analyses of the enzymes comprising the photosynthetic carbon reduction cycle have been performed in extracts of dark grown and greening Euglena gracilis var. bacillaris. Chlorella pyrenoidosa grown photoautotrophically, in the light with glucose or in the dark with glucose, Tolypothrix tenuis, Chromatium and leaves of spinach. Amounts of activity are compared with the level of photosynthetic CO₂ fixation. Only in Chromatium were all enzyme activities sufficient to support the in vivo rate of CO₂ fixation. In organisms other than Chromatium, some enzymes and particularly fructose 1,6-phosphatase and ribulose 1.5-diphosphate carboxylase appeared to be present in insufficient amounts to support the photosynthetic rate of the intact cell. Developmental studies with Euglena and growth studies with Chlorella led to the conclusion that these enzymes were associated with the cycle. Suppression of CO₂ fixation in heterotrophically grown Chlorella was accompanied by a striking decrease in the same enzymes whose activities increased in greening Euglena.     

Evidence of de novo synthesis of maltose excreted by the endosymbiotic Chlorella from Paramecium bursaria

The endosymbiotic Chlorella sp. from Paramecium bursaria excretes maltose both in the light and in the dark. Experiments on photosynthetic ¹⁴CO₂ fixation and ¹⁴CO₂ pulse-chase experiments show that maltose is synthesized in the light directly from compounds of the Calvin cycle, whereas in the dark it results from starch degradation.     

Inhibition of photosynthesis by intracellular carbonic anhydrase in microalgae under excess concentrations of CO2

When cells of Chlorococcum littorale that had been grown in air (air-grown cells) were transferred to extremely high CO₂ concentrations (>20%), active photosynthesis resumed after a lag period which lasted for 1–4 days. In contrast, C. littorale cells which had been grown in 5% CO₂ (5% CO₂-grown cells) could grow in 40% CO₂ without any lag period. When air-grown cells were transferred to 40% CO₂, the quantum efficiency of PS II (Φ_II) decreased greatly, while no decrease in Φ_II was apparent when the 5% CO₂-grown cells were transferred to 40% CO₂. In contrast to air-grown cells, 5% CO₂-grown cells showed neither extracellular nor intracellular carbonic anhydrase (CA) activity. Upon the acclimation of 5% CO₂-grown cells to air, photosynthetic susceptibility to 40% CO₂ was induced. This change was associated with the induction of CA. In addition, neither suppression of photosynthesis nor arrest of growth was apparent when ethoxyzolamide (EZA), a membrane-permeable inhibitor of CA, had been added before transferring air-grown cells of C. littorale to 40% CO₂. The intracellular pH value (pH_i) decreased from 7.0 to 6.4 when air-grown C. littorale cells were exposed to 40% CO₂ for 1–2 h, but no such decrease in pH_i was apparent in the presence of EZA. Both air- and 5% CO₂-grown cells of Chlorella sp. UK001, which was also resistant to extremely high CO₂ concentrations, grew in 40% CO₂ without any lag period. The activity of CA was much lower in air-grown cells of this alga than those in air-grown C. littorale cells. These results prompt us to conclude that intracellular CA caused intracellular acidification and hence inhibition of photosynthetic carbon fixation when air-grown C. littorale cells were exposed to excess concentrations of CO₂. No such harmful effect of intracellular CA was observed in Chlorella sp. UK001 cells. This revised version was published online in June 2006 with corrections to the Cover Date.     

Polarographische Messung der Nitritreduktion von Chlorella im monochromatischen Licht

Summary In green plant cells nitrite is reduced by two systems, one dependent on photosynthesis and the other upon respiration. Using a polarographic method for continuous measurement of nitrite uptake, the relationship between light driven and respiration linked nitrite reduction of Chlorella cells was studied.Photosynthetic nitrite reduction is characterized by a pronounced increase in the velocity of nitrite uptake upon illumination. After the light is turned off the velocity immediately returns to the preillumination value. Photosynthetic nitrite reduction of Chlorella is separated from respiration linked nitrite reduction by illumination with red light under anaerobic conditions; it is stimulated by CO₂ and is inhibited by DCMU, findings which confirm earlier observations.In white light a special blue light stimulation of nitrite uptake is overlapped by photosynthetic nitrite reduction. In contrast to photosynthetic nitrite reduction this type of light stimulation is characterized by a lag period of about I min from the onset of illumination; it continues about 10 min when the light is turned off. It is separated from photosynthetic nitrite reduction by irradiation of the algae with low intensities of short wavelength light (<500 nm). Blue light stimulation of nitrite uptake of Chlorella is strongly dependent on the developmental stage of the cells. It is observed with young cells (autospores) of synchronized algae only.There is no evidence for any connection between blue light stimulation of nitrite uptake and photosynthesis. From the sensitivity of this process towards anaerobic conditions and antimycin A it is concluded to be a stimulation of respiration linked nitrite reduction.Under conditions of low exogenous nitrite concentration a temporary inhibition of steady state dark nitrite reduction appears immediately after the light is turned off. From several observations it is concluded that the inhibition already exists during the preceding illumination and decreases the rate of total nitrite uptake in the light. This process is suppressed by inhibition of respiration as well as by the inhibitor of photosynthesis, DCMU.If nitrate is the source of nitrogen an excretion of nitrite is found following illumination. The kinetics of this process agree with those observed for the light induced inhibition of steady state dark nitrite reduction immediately after illumination.     

Comparison of photosynthetic acclimation to elevated CO2 and limited nitrogen supply in soybean

Plants grown at elevated CO₂ often acclimate such that their photosynthetic capacities are reduced relative to ambient CO₂-grown plants. Reductions in synthesis of photosynthetic enzymes could result either from reduced photosynthetic gene expression or from reduced availability of nitrogen-containing substrates for enzyme synthesis. Increased carbohydrate concentrations resulting from increased photosynthetic carbon fixation at elevated CO₂ concentrations have been suggested to reduce the expression of photosynthetic genes. However, recent studies have also suggested that nitrogen uptake may be depressed by elevated CO₂, or at least that it is not increased enough to keep pace with increased carbohydrate production. This response could induce a nitrogen limitation in elevated-CO₂ plants that might account for the reduction in photosynthetic enzyme synthesis. If CO₂ acclimation were a response to limited nitrogen uptake, the effects of elevated CO₂ and limiting nitrogen supply on photosynthesis and nitrogen allocation should be similar. To test this hypothesis we grew non-nodulating soybeans at two levels each of nitrogen and CO₂ concentration and measured leaf nitrogen contents, photosynthetic capacities and Rubisco contents. Both low nitrogen and elevated CO₂ reduced nitrogen as a percentage of total leaf dry mass but only low nitrogen supply produced significant decreases in nitrogen as a percentage of leaf structural dry mass. The primary effect of elevated CO₂ was to increase non-structural carbohydrate storage rather than to decrease nitrogen content. Both low nitrogen supply and elevated CO₂ also decreased carboxylation capacity (V_cmax) and Rubisco content per unit leaf area. However, when V_cmax and Rubisco content were expressed per unit nitrogen, low nitrogen supply generally caused them to increase whereas elevated CO₂ generally caused them to decrease. Finally, elevated CO₂ significantly increased the ratio of RuBP regeneration capacity to V_cmax whereas neither nitrogen supply nor plant age had a significant effect on this parameter. We conclude that reductions in photosynthetic enzyme synthesis in elevated CO₂ appear not to result from limited nitrogen supply but instead may result from feedback inhibition by increased carbohydrate contents.     

Contrasting Effects of Carbon Dioxide and Irradiance on the Acclimation of Photosynthesis in Developing Soybean Leaves

Leaves developed at high irradiance (I) often have higher photosynthetic capacity than those developed at low I, while leaves developed at elevated CO₂ concentration [CO₂] often have reduced photosynthetic capacity compared with leaves developed at lower [CO₂]. Because both high I and elevated [CO₂] stimulate photosynthesis of developing leaves, their contrasting effects on photosynthetic capacity at maturity suggest that the extra photosynthate may be utilized differently depending on whether I or [CO₂] stimulates photosynthesis. These experiments were designed to test whether relationships between photosynthetic income and the net accumulation of soluble protein in developing leaves, or relationships between soluble protein and photosynthetic capacity at full expansion differed depending on whether I or [CO₂] was varied during leaf development. Soybean plants were grown initially with a photosynthetic photon flux density (PPFD) of 950 µmol m^–2 s^–1 and 350 µmol [CO₂] mol^–1, then exposed to [CO₂] ranging from 135 to 1400 µmol mol^–1 for the last 3 d of expansion of third trifoliolate leaves. These results were compared with experiments in which I was varied at a constant [CO₂] of 350 µmol mol^–1 over the same developmental period. Increases in area and dry mass over the 3 d were determined along with daily photosynthesis and respiration. Photosynthetic CO₂ exchange characteristics and soluble protein content of leaves were determined at the end of the treatment periods. The increase in leaflet mass was about 28 % of the dry mass income from photosynthesis minus respiration, regardless of whether [CO₂] or I was varied, except that very low I or [CO₂] increased this percentage. Leaflet soluble protein per unit of area at full expansion had the same positive linear relationship to photosynthetic income whether [CO₂] or I was varied. For variation in I, photosynthetic capacity varied directly with soluble protein per unit area. This was not the case for variation in [CO₂]. Increasing [CO₂] reduced photosynthetic capacity per unit of soluble protein by up to a factor of 2.5, and photosynthetic capacity exhibited an optimum with respect to growth [CO₂]. Thus CO₂ did not alter the relationship between photosynthetic income and the utilization of photosynthate in the net accumulation of soluble protein, but did alter the relationship between soluble protein content and photosynthetic characteristics in this species.     

The Effect of Oxygen Concentration on Photosynthesis in Higher Plants

The influence of oxygen concentration in the range 0–21% on photosynthesis in intact leaves of a number of higher plants has been investigated. Photosynthetic Co₂ fixation of higher plants is markedly inhibited by oxygen in concentrations down to less than 2%. The inhibition increases with oxygen concentration and is about 30% in an atmosphere of 21% O₂ and 0.03% Co.₂. Undoubtedly, therefore, oxygen in normal air exerts a strong inhibitory effect on photosynthetic Co₂ fixation of land plants under natural conditions. The inhibitory effect of oxygen is rapidly produced and fully reversible. The degree of inhibition is independent of light intensity. The quantum yield for Co₂ fixation, i.e. the slope of the linear part of the curve for Co₂ uptake versus absorbed quanta, is inhibited to the same degree as the light saturated rate at all oxygen concentrations studied. Diverse species of higher plants, varying greatly in photosynthetic response to light intensity and Co₂ concentration, and with light saturated roles of Co₂ fixation differing by a factor of more than 10 times, show a remarkable similarity in their response to oxygen concentration. By contrast, when studied under the same conditions as the higher plants, the green algae Chlorella and Ulva did not show-any measurable inhibition of photosynthetic Co₂ fixation. Similarity, the increase in fluorescence intensity with increasing oxygen concentrations found in higher plants also was not seen in Chlorella. The present results, together with previous data on the photosynthetic response of algae to oxygen concentration, indicate that the photosynthetic apparatus of higher plants differs considerably from that of algae in its sensitivity to oxygen. The inhibitory effect of oxygen on photosynthetic Co₂ fixation in higher plants is somewhat higher at wavelengths which excite preferentially photosystem I. Also, the Emerson enhancement of Co₂ fixation measured when a far red beam of low intensity is imposed on a background of red light is greater under low oxygen concontrution than under air. Measurements of reversible light-induced absorbance changes reveal that the change at 591 nm, probably caused by pla.stocyanin, is affected by oxygen concentration only if photosystem II is excited. the reducing effect on plastocyanin, caused by excitation of this system, decreases with increasing oxygen concentration. From these results it is suggested that a possible site of the inhibition by oxygen is in the electron carrier chain between the two photosystems. Oxygen might act as an electron acceptor at this site, causing reducing power to react back with molecular oxygen. However, this hypothesis does not account for equal inhibitions of the quantum yield and the light saturated rate of photosynthetic CO₂ uptake. Through the photosynthetic process plants take up carbon dioxide and evolve oxygen. The present high concentration of molecular oxygen in the atmosphere is generally considered to have arisen from the activity of photo-synthetic organisms. The effect of oxygen concentration would seem, therefore, to he a problem of great interest, not only in the field of the biophysics and biochemistry of photosynthesis, but in ecology and other branches of biology as well. It was discovered by Warburg (1920) that high concentrations of oxygen inhibit the rate of photosynthetic oxygen evolution in the unicellular alga Chlorella. Since then, it has been confirmed by various authors that oxygen cconcentrations in the range 21–100 per cent have a marked inhibitory effect on photosynthesis, particularly at saturating light intensities. There is some evidence that under conditions when carbon dioxide concentration limits photosynthesis, the inhibition may become obvious even in 21 per cent oxygen. The inhibition has not been considered to operate at low light intensities. A review on the subject has been given by Turner and Brittain (1962). Various hypotheses have been put forward to explain the inhibitory effect of oxygen, commonly referred to as the Warhurg effect. Some authors favor the idea of enzyme inhibition; Turner et al. (1958) that one or more enzymes of the carbon reduction cycle are inactivated by oxygen: lirianlals (1962) that enzymes of the oxygen-evolving complex are inhihited. Other hypotheses concern back-reactions in which molecular oxygen is taken up, thus reversing the photosynthetic process. These reactions include photo-oxidation, photorespiration, and the Mehler reaction (Tamiya et al., 1957). At present, there is no generally accepted hypothesis explaining the effect. The often conflicting results on which these hypotheses were based have been obtained mostly on algae. The first observation of an inhibitory effect on photosynthesis in a higher plant was made hy McAlister and Myers (1940) in wheat leaves. They found that the photosyntlietic CO₂ uptake was markedly lower in air than in an atmosphere of about 0.5 per cent oxygen. At the CO₂ concentration used (0.03%) the inhibition was present both at high and moderate light intensities. No data were obtained at low light intensities. Although the study of the effect of oxygen concentration on photosynthesis in higher plants would seem to be of great interest, particularily since the natural environment of most land plants is an atmosphere with an oxygen content of 21 per cent, it has attracted very little attention. To the author's knowledge no thorough investigation on the subject has been published. The present investigalion is directed toward elucidatirng the photosynthetic response of higher plants to oxygen concentrations up to that of normal air. Data are presented showing that the photosynthetic CO₂ fixation in intact leaves of higher plants, regardless of light intensity, is strongly inhibited by oxygen in normal air, and that the pholosynthetic response to oxygen differs considerably from that of green algae. The present investigalion is directed toward elucidatirng the photosynthetic response of higher plants to oxygen concentrations up to that of normal air. Data are presented showing that the photosynthetic CO₂ fixation in intact leaves of higher plants, regardless of light intensity, is strongly inhibited by oxygen in normal air, and that the pholosynthetic response to oxygen differs considerably from that of green algae.     

Transfer of Photosynthetically Produced Carbohydrate from Endosymbiotic Chlorellae to Paramecium bursaria*

Paramecium bursaria (Ehrenberg) and an endozoic zoochlorella Chlorella conductrix (Brandt) live in a symbiotic relationship. Uptake of NaH¹⁴CO₃ was studied to determine if carbohydrate products of photosynthesis are transferred to the host paramecium. Paramecium bursaria containing the algal symbionts took up NaH¹⁴CO₃ but those without the algal symbionts did not. Radioactive maltose, glucose, fructose and malate were identified from the ethanolic extract of paramecia. Transfer of materials from Paramecium to Chlorella and the transfer of other materials from Chlorella to Paramecium, led to the conclusion that this is a mutualistic relationship, both organisms benefiting from the relationship.     

Einfluß kurzer Dunkelperioden auf CO2-Aufnahme und Phosphoenolpyruvat-Carboxylierung während der Photosynthese-Induktion bei Chlorella vulgaris

Zusammenfassung Nach einer Dunkelperiode von 40 min und 40 sec wurden die CO₂-Aufnahme und die ¹⁴C-markierten Produkte während der Photosynthese-Induktion bei Chlorella vulgaris (211-11f) bestimmt. Die mit Preßluft (0,03 Vol.-% CO₂) begasten Algen sind bei +27°C kultiviert und bei +10° oder +25°C gemessen worden. Ein Induktionseffekt der photosynthetischen CO₂-Aufnahme konnte nur nach einer längeren Dunkelperiode (>3 min) beobachtet werden. Unter diesen Bedingungen wurde ¹⁴CO₂ am Anfang der Belichtung in Malat, Aspartat und 3-Phosphoglycerat eingebaut. Nach einer kurzen Dunkelperiode (40 sec) waren zu Beginn der Belichtung vor allem die Produkte des Calvin-Cyclus markiert. Die Wirkung von Intermediaten auf die Ausbildung der Induktionseffekte wird diskutiert.

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Effect of short dark periods on CO₂ uptake and carboxylation of phosphoenolpyruvate during the photosynthetic induction period in Chlorella vulgaris

Summary CO₂ exchange, ¹⁴CO₂ fixation and ¹⁴C labelled products of Chlorella vulgaris (strain 211-11f) were studied during the photosynthetic induction period at +10° and +25°C after a dark period of 40 min and 40 sec. The algae were grown under normal aerated conditions (0.03 vol.-% CO₂) at +27°C. Transient changes in CO₂ uptake, measured with an infrared gas analyzer, could be observed only after a dark period of >3 min; no such changes occurred after a dark period of 40 sec. The autoradiographic studies of the kinetics of the appearance of labelled products at +10° and +25°C showed that after a long dark period (40 min) at the beginning of illumination ¹⁴CO₂ was incorporated into malate, aspartate and 3-phosphoglycerate. Under these conditions, the intermediates of the Calvin cycle were labelled after 30 sec (+25°C) or 2 min (+10°C) of photosynthesis. After a dark period of 40 sec (at +10° and +25°C), however, ¹⁴C incorporation into malate and aspartate was rather low at the beginning of illumination; moreover, the intermediates of the Calvin cycle appeared earlier and were more strongly labelled after this short dark period. The results are discussed with reference to the influence of intermediates on the formation of the transient changes of CO₂ uptake in Chlorella.

     

Gene expression analysis of a Louisiana native Chlorella vulgaris (Chlorophyta)/Leptolyngbya sp. (Cyanobacteria) co‐culture using suppression subtractive hybridization

A locally isolated co‐culture of two photosynthetic species [Chlorella vulgaris (Chlorophyta) and Leptolyngbya sp. (Cyanobacteria)] displayed enhanced growth when compared to a Chlorella monoculture; however, the biological mechanisms driving such improvement are currently not well understood. To investigate these mechanisms, this study examined the differential gene expression in the Chlorella between the co‐culture and the monoculture. Suppression subtractive hybridization was performed between mRNA from Chlorella in the co‐culture and in a monoculture, and 105 genes were identified as being putatively differentially expressed. Nine of these genes, corresponding to the key functional categories of energy, metabolism, and protein synthesis, were further examined using quantitative real‐time PCR and showed differential regulation of photosystem I and photosystem II and upregulation of stress‐response genes and a gene encoding an oil‐globule‐associated gene in the co‐culture Chlorella. This differential gene expression study of a Chlorella/cyanobacteria co‐culture will aid in the development of culture strategies capable of taking advantage of these differences for the production of biomass and bioproducts of interest. Knowledge of the underlying genetic causes of the changes in growth and productivity of the species in co‐culture provides insights on possible target genes for optimization of the culture.     

An analysis of photosynthetic response and adaptation to temperature in higher plants: temperature acclimation in the desert evergreen Nerium oleander L*

Abstract. Factors underlying the process of photosynthetic acclimation to temperature were investigated for the shrub Nerium oleander L. Ramets of a single clone were grown under day/night temperature regimes of 20°C/15°C or 45°C/32°C. Plants grown at the lower temperature regime possessed rates of photosynthesis twice that of the high-temperature grown plants when CO₂ fixation was measured at 20°C. In contrast, the plants grown at the high-temperature regime had twice the rate of CO₂ fixation of the 20°C/l 5°C-grown plants at a measurement temperature of 45° C. It was determined that the ability to acclimate to changes in temperature regime was present in fully mature leaves. A reciprocal transfer of plants between the two growth regimes resulted in the appearance of the CO₂ fixation characteristics appropriate to the new growth temperature after 12–14d. The response of CO₂ fixation to light, temperature, and CO₂ partial pressure and the temperature responses of soluble and membrane-bound photosynthetic enzyme systems were analysed to determine which components might be responsible for the superior photosynthetic performance of the 20°C/I5°C-grown plants at 20°C, and the enhanced high-temperature stability of the 45°C/32°C plants. The measured photosynthetic capacity of the 20°C/15°C plants could not be attributed to gross morphological, stomatal, or other physical changes, or to a general increase in the concentration of components of the photosynthetic process. Only a single enzyme, Fru-P₂ phosphatase, was affected to an extent similar to that of photosynthesis. The enhanced thermal stability of the 45°C/32°C plants may be attributed primarily to an enhanced stability of the chloroplast membrane-bound enzymatic activities and the stability of the photosynthetic carbon metabolism enzymes which require lighl for activation.     

Electron transport is the functional limitation of photosynthesis in field-grown Pima cotton plants at high temperature

Restrictions to photosynthesis can limit plant growth at high temperature in a variety of ways. In addition to increasing photorespiration, moderately high temperatures (35–42 °C) can cause direct injury to the photosynthetic apparatus. Both carbon metabolism and thylakoid reactions have been suggested as the primary site of injury at these temperatures. In the present study this issue was addressed by first characterizing leaf temperature dynamics in Pima cotton (Gossypium barbadense) grown under irrigation in the US desert south‐west. It was found that cotton leaves repeatedly reached temperatures above 40 °C and could fluctuate as much as 8 or 10 °C in a matter of seconds. Laboratory studies revealed a maximum photosynthetic rate at 30–33 °C that declined by 22% at 45 °C. The majority of the inhibition persisted upon return to 30 °C. The mechanism of this limitation was assessed by measuring the response of photosynthesis to CO₂ in the laboratory. The first time a cotton leaf (grown at 30 °C) was exposed to 45 °C, photosynthetic electron transport was stimulated (at high CO₂) because of an increased flux through the photorespiratory pathway. However, upon cooling back to 30 °C, photosynthetic electron transport was inhibited and fell substantially below the level measured before the heat treatment. In the field, the response of assimilation (A) to various internal levels of CO₂ (C_i) revealed that photosynthesis was limited by ribulose‐1,5‐bisphosphate (RuBP) regeneration at normal levels of CO₂ (presumably because of limitations in thylakoid reactions needed to support RuBP regeneration). There was no evidence of a ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco) limitation at air levels of CO₂ and at no point on any of 30 A–C_i curves measured on leaves at temperatures from 28 to 39 °C was RuBP regeneration capacity measured to be in substantial excess of the capacity of Rubisco to use RuBP. It is therefore concluded that photosynthesis in field‐grown Pima cotton leaves is functionally limited by photosynthetic electron transport and RuBP regeneration capacity, not Rubisco activity.     

A microbiologist's odyssey: Bacterial viruses to photosynthetic bacteria

Perspective can be defined as the relationships or relative importance of facts or matters from any special point of view. Thus, my Personal perspective reflects the threads I followed in a 50-year journey of research in the complex tapestry of bioenergetics and various aspects of microbial metabolism. An early interest in biochemical and microbial evolution led to the fertile hunting grounds of anoxygenic photosynthetic bacteria. Viewed as a physiological class, these organisms show remarkable metabolic versatility in that certain individual species are capable of using all the known major types of energy conversion (photosynthetic, respiratory, and fermentative) to support growth. Since such anoxyphototrophs are readily amenable to molecular genetic/biological manipulation, it can be expected that they will eventually provide important clues for unraveling the evolutionary relationships of the several kinds of energy conversion. I gradually came to believe that understanding the evolution of phototrophs would require detailed knowledge not only of how light is converted to chemical energy, but also of a) pathways of monomer production from extracellular sources of carbon and nitrogen and b) mechanisms cells use for integrating ATP regeneration with the energy-requiring biosyntheses of biological macromolecules. Serendipic observation of photoproduction of H₂ from organic compounds by Rhodospirillum rubrum in 1949 led to discovery of N₂ fixation by anoxyphototrophs, and this capacity was later exploited for the isolation of hitherto unknown species of photosynthetic prokaryotes, including the heliobacteria. Recent studies on the reaction centers of the heliobacteria suggest the possibility that these bacteria are descendents of early phototrophs that gave rise to oxygenic photosynthetic organisms.Abbreviations AMP adenosine monophosphate - ADP adenosine diphosphate - ATP adenosine triphosphate - ATPase adenosine triphosphatase - Bchl bacteriochlorophyll - DMSO dimethyl sulfoxide - NADH reduced nicotinamide adenine dinucleotide - nif ^– genes for dinitrogen fixation - Nif^– bacterial mutants incapable of dinitrogen fixation - O/R oxidation/reduction - P_i inorganic orthophosphate - R. capsulatus Rhodobacter capsulatus - R. sphaeroides Rhodobacter sphaeroides - Rps. Rhodopseudomonas - TMAO trimethyl amine-N-oxide Written at the invitation of Govindjee.     

Effect of jasmonic acid on the stomatal and nonstomatal limitation of leaf photosynthesis in barley leaves

The effect of long-term (7 days) and shortterm (up to 2 h) treatment of barley plants with jasmonic acid (JA) on the components contributing to stomatal and nonstomatal limitation of photosynthesis was studied. Net CO₂ assimilation rate (A) responses to intercellular CO₂ concentration (C _i ), i.e., A/C _i curves, were used to assess the photosynthetic ability. Long-term treatment of barley plants with JA led to a noticeable decrease in both the initial slope of the A/C _i curves and the maximum A at saturating C _i . The proportion of stomatal and nonstomatal factors in limitation of photosynthesis depended on the applied JA concentration. Short-term treatment with JA affected neither the stomatal conductivity for CO₂ nor the rate of photosynthetic CO₂ assimilation. We suggest that JA may affect photosynthesis indirectly, either as a stress-modulating substance, or through the alterations in gene expression.     

Similarity of growth patterns between plantlets and seedlings of Brassica campestris L. under different in vitro environmental conditions

Explants and seeds of Brassica campestris L. were cultured on Murashige & Skoog (1962) medium with and without sucrose in a vessel with different numbers of air changes per hour under different PPF (photosynthetic photon flux) conditions. The growth and development of plantlets in the vessel were similar to those of seedlings when cultured under the same in vitro environmental conditions. The growth and development of seedlings when cultured under the same in vitro environmental conditions. The growth and development of plantlets/seedlings were greater for treatments with a higher number of air changes per hour and a higher PPF regardless of the sucrose concentration in the culture medium.The CO₂ concentration in the vessel with a lower number of air changes per hour decreased to approximately 100 ppm during the photoperiod on day 21 due to the photosynthetic activities of the plantlets/seedlings. The low CO₂ concentration, in turn, reduced the net photosynthetic rate of plantlets/seedlings in the vessel, and thus affected their growth and development.Abbreviations C_in CO₂ concentration in the culture vessel - C_out CO₂ concentration in the culture room - MS mineral composition of Murashige & Skoog (1962) medium - PPF photosynthetic photon flux     

Limitations due to water stress on leaf net photosynthesis of Quercus coccifera in the Portuguese evergreen scrub

Summary Gas exchange characteristics in leaves of the sclerophyll shrub Quercus coccifera were studied in the natural habitat in Portugal during spring and during the summer dry period. Compared to other sclerophyll species growing at the same site, photosynthesis in leaves of Quercus coccifera was less affected by water stress. Moderate water stress after six weeks of drought led to large decreases in stomatal conductance but no change in mesophyll photosynthetic capacity as compared to late spring. Leaf internal CO₂ pressure remained near 220 bar during diurnal courses in the spring. On midsummer days, leaf internal CO₂ decreased from a late morning value of 200 bar to a late afternoon value of approximately 150 bar. In contrast to Quercus suber (Tenhunen et al. 1984), restriction of CO₂ supply due to stomatal closure reduced net CO₂ uptake at midday and in the afternoon during midsummer. A decrease in leaf carboxylation efficiency and an increase in CO₂ compensation point at midday also played an important role in determining the diurnal course of net photosynthesis. During the late stages of drought in September, severe water stress led to reduction in mesophyll photosynthetic capacity and further reduction in leaf conductance. The observed decrease in mesophyll photosynthetic capacity was correlated with decrease in the daily minimum leaf water potential to greater negative values than-30 bar. At this time, CO₂ saturated photosynthetic rates decreased as much as 50% over the course of a day when measured at constant saturating light, 32° C leaf temperature, and a water vapor mole fraction difference between leaf and air of 30 mbar bar^-1.