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Irradiance is the main driver of photosynthesis. In natural conditions, irradiance incident on a leaf often fluctuates, due to the movement of leaves, clouds and the sun. These fluctuations force photosynthesis to respond dynamically, however with delays that are subject to rate constants of underlying processes, such as regulation of electron transport, activation states of enzymes in the Calvin cycle, and stomatal conductance (gs). For example, in leaves adapted to low irradiance that are suddenly exposed to high irradiance, photosynthesis increases slowly (within tens of minutes); this process is called photosynthetic induction. Photosynthesis in fluctuating irradiance (dynamic photosynthesis) is limited by several physiological processes, and is further modulated by environmental factors other than irradiance, such as CO2 concentration, air humidity and temperature. Studying dynamic photosynthesis and its environmental and physiological control can help to identify targets for improvements of crop growth, improve the accuracy of mathematical models of photosynthesis, and explore new, dynamic lighting strategies in greenhouses. In this thesis, the limitations acting on dynamic photosynthesis are explored by reviewing the literature, by experimenting with a suite of environmental factors (CO2 concentration, temperature, air humidity, irradiance intensity and spectrum), genetic diversity in the form of mutants, genetic transformants and ecotypes, and by mathematical modelling. Several genotypes of tomato (Solanum lycopersicum) and the model plant Arabidopsis thaliana, all grown in climate chambers, were used in the experiments. The main findings of the thesis are that a) CO2 concentration and air humidity strongly affect the rate of change of dynamic photosynthesis through a combination of diffusional and biochemical limitations; b) Rubisco activation kinetics are pivotal in controlling rates of photosynthesis increase after a stepwise increase in irradiance, and are further affected by CO2 concentration; c) gs limits photosynthetic induction kinetics in A. thaliana but not in tomato in ambient conditions, and becomes a stronger limitation in low CO2 concentration or air humidity; and d) mesophyll conductance, non-photochemical quenching (NPQ) and sucrose synthesis do not limit dynamic photosynthesis under the conditions used. In Chapter 1, the rationale for the research conducted is described, by introducing the concept of fluctuating irradiance and its effects on photosynthesis rates. The chapter discusses how dynamic photosynthesis is measured and described, and provides a range of possible applications of the insights gained by the research conducted in this dissertation. In Chapter 2, the current literature is reviewed and a mechanistic framework is built to explore the effects that the environmental factors CO2 concentration, temperature and air humidity have on rates of dynamic photosynthesis. Across data from literature, higher CO2 concentration and temperature speed up photosynthetic induction and slow down its loss, thereby facilitating higher rates of dynamic photosynthesis. Major knowledge gaps exist regarding the loss of photosynthetic induction in low irradiance, dynamic changes in mesophyll conductance, and the extent of limitations imposed by gs across species and environmental conditions. Chapter 3 is an experimental exploration of the effects of CO2 concentration, leaf temperature, air humidity and percentage of blue irradiance on rates of photosynthetic induction in dark-adapted tomato leaves. Rubisco activation, changes in stomatal and mesophyll conductance, diffusional and biochemical limitations, efficiency of electron transport through photosystem II, NPQ and transient water use efficiency, were examined to give a comprehensive overview of the environmental modulation of dynamic photosynthesis. Unlike the percentage of blue irradiance, increases in CO2 concentration, leaf temperature and air humidity all positively affected the rates of photosynthetic induction, and these effects were explained by changes in diffusional and biochemical limitations. Maximising the rates of Rubisco activation would increase CO2 assimilation by 6-10%, while maximising the rates of stomatal opening would increase assimilation by at most 1-2%, at the same time negatively affecting intrinsic water use efficiency. In Chapter 4 it is explored whether the effects of CO2 concentration on dynamic photosynthesis are similar across various irradiance environments. Gain and loss of photosynthetic induction in several low irradiance treatments, as well as sinusoidal changes in irradiance, were studied using tomato leaves. Elevated CO2 concentration (800 ppm) enhanced the rate of photosynthetic induction by 4-12% (compared to 400 ppm) and decreased the loss of photosynthetic induction by 21-25%. Elevated CO2 concentration enhanced rates of dynamic photosynthesis regardless of initial photosynthetic induction state to a similar extent. Therefore, rising global CO2 concentration will benefit integrated assimilation throughout whole canopies, where different leaf layers experience widely differing irradiance regimes. In Chapter 5 it is tested whether stomatal limitation exists during photosynthetic induction in tomato leaves. The abscisic acid-deficient flacca mutant and its wildtype were exposed to various CO2 concentrations to change the diffusion gradient. Despite gs being much larger in flacca, photosynthetic induction proceeded with the same speed in both genotypes in ambient CO2 concentration. This suggested that stomata did not limit photosynthetic induction in the wildtype. Using these findings, several indices of stomatal limitations were compared. Diffusional limitation, a new index, was found to be the most useful. In Chapter 6, an exploration of some physiological limitations underlying dynamic photosynthesis is undertaken. Several mutants, transformants and ecotypes of A. thaliana, affecting rates of Rubisco activation, gs, NPQ and sucrose metabolism, were used. Next to a characterisation of their steady-state responses to CO2 concentrations and irradiance, leaves were exposed to stepwise increases and decreases in irradiance (using several intensities) and to lightflecks of several amplitudes and frequencies. Rubisco activase isoform and concentration, as well as various levels of gs, strongly affected rates of dynamic photosynthesis, while this was not the case with low NPQ or sucrose phosphate synthase concentration. This suggests Rubisco activase and gs as targets for improvement of photosynthesis in fluctuating irradiance. Chapter 7 is a modelling exercise of dynamic photosynthesis, based on data obtained from measurements on mutants of A. thaliana (Chapter 6). This includes a goal-seeking model that allows reproducing the regulation of Rubisco by irradiance and CO2 concentration. The model also includes a full description of leaf-level NPQ, incorporates mesophyll conductance and accounts for the fundamental physics of delays introduced by open gas exchange systems on CO2 measurements. Different data sets for model calibration and validation were used. It was found that the model accurately predicted the effects of the mutants, suggesting that the assumptions of the model were sound and represented the underlying mechanisms correctly. In Chapter 8, the findings in this thesis are synthesized. The insights gained throughout this dissertation are related to existing literature to give a comprehensive overview of the state of knowledge about the limitations of dynamic photosynthesis. The methodology of assessing transient stomatal limitations, and some aspects of using chlorophyll fluorescence measurements during photosynthetic induction, are discussed. Finally, possible applications and ideas for future research on photosynthesis in fluctuating irradiance are discussed. |
