Physiological response of the Cyanobacterium synechocystis sp. PCC 6803 to fluctuating light

Abstract

Photosynthetic microbes are a promising feedstock for renewable biofuels, but the yields of industrial cultivation systems will need significant improvements if they are to be economically viable and succeed. One particular challenge faced by photosynthetic microbes in commercial production systems is the highly dynamic light environment created by vertical mixing within dense cultures. Rapid changes in light intensity make it difficult for these microbes to acclimate and utilize the available light efficiently. Attempts to identify targets for genetically improving photosynthetic microbes to flourish in these environments are hampered by a poor understanding of the physiological response to fluctuating light. My thesis is focused on developing our fundamental understanding of the photophysiology and acclimation responses associated with light environments in industrial conditions. The aim is to eventually increase areal productivity in industrial cultivation systems by applying insights from physiological characterization into future strain engineering approaches. The first chapter introduces the issues associated with industrial cultivation of photosynthetic microbes. I present some background on the need for biofuels, why there has been a focus on using photosynthetic microbes as a feedstock, and how photosynthetic microbes are cultivated industrially. I then present some of the physiological challenges faced by photosynthetic organisms in industrial cultivation, with particular focus on the challenge of a dynamic light environments. I give a brief background on photosynthesis to explain some of the acclimation responses that can be altered in a fluctuating light environment. Chapter 2 presents a proteomic and physiological comparison of the cyanobacterium Synechocystis sp. PCC6803 cultivated in a fluctuating light environment (30s light on/off) to a continuous light environment which had the same average photon flux density. We found that cultures in fluctuating light grew at half the exponential growth rate of continuous light cultures. Reduced growth did not appear to be due to photo-oxidative stress, as we detected reduced levels of reactive oxygen species and oxidative-stress responsive proteins in fluctuating light. We show evidence that reduced growth could be due to a partial shift to a respiratory state in fluctuating light. Reduced growth could also be due to increased dissipation of electrons, as suggested by the higher capacity for photosynthesis at light levels from 106-174 μmol photon m⁻² s⁻¹. We found other surprising changes such as increases in some carbon concentration mechanism components and decreases in some others. These components were thought to be regulated by a similar mechanism due to their co-expression in high CO₂ to ambient CO₂ shift experiments. This suggesting some unusual signaling is occurring due to the fluctuating light. We also found a number of hypothetical and poorly characterized proteins were significantly different in fluctuating and continuous light. In the appendix, I present the preliminary characterization of slr1719, a poorly characterized protein identified in the proteomic analysis of fluctuating and continuous light. This study generated a knock out of slr1719. We found that the Δslr1719 strain compared to a control strain grew much slower in low CO₂ conditions in saturating and sub saturating light conditions. However in replete CO₂ conditions, there was no difference in growth rate, regardless of light intensity tested in Δslr1719 versus the control. We argue that this finding, paired with evidence from the literature, suggests slr1719 may participate in cyclic electron flow. Other roles suggested by the literature for this protein could have important roles in the acclimation to fluctuating light, and warrant further study.