Kenneth E. Wilson

Fourier transform infrared (FTIR) spectromicroscopy was used to study individual living cells of three closely-related species of the green algae Chlamydomonas . This study differentiated these three species based on differences in lipid and protein profiles, as well explored sources of variation in our measurements. Significant spectral variation was observed between individual cells within a single culture, as well as between control cultures of a species obtained on different days. Despite this, we were able to differentiate between the three close-related species. Differences in the spectra were confirmed using principal component analysis. Understanding the interplay of underlying variation and the degree of induced spectral differences is essential for the deployment of FTIR measurements in both bulk cultures and for individual living cells.

Industrial activity associated with oil‐sands extraction in Canada's Athabasca region produces a variety of contaminants of concern, including naphthenic acid fraction components (NAFCs). NAFCs are a complex mixture of organic compounds that are poorly understood both in terms of their chemical composition and effects on the environment. NAFC toxicity in the unicellular green algae Chlamydomonas reinhardtii P.A.Dangeard was correlated with the presence of the algal cell wall. It was suggested that the toxicity of NAFCs in C. reinhardtii was due to surfactant effects. Surfactant‐cell wall interactions are specific and governed by the compound class and structure, and by the nature of the biological material. Here, we investigate the effects of wildtype (WT) C. reinhardtii and two cell‐wall mutants on specific classes of NAFCs when growing cultures were treated with a 100 mg · L⁻¹ solution of NAFCs. Changes in the NAFC composition in the media were examined using high resolution mass spectrometry over a period of 4 d. Algal mediated changes in the NAFCs were limited to specific classes of NAFCs. In particular, the removal of large, classical naphthenic acids, with a double bond equivalent of 8, was observed in WT C. reinhardtii cultures. The observed algal mediated changes in NAFC composition would have been masked by low resolution mass spectrometry and highlight the importance of this tool in examining bioremediation of complex mixtures of NAFCs.

Photosystem I (PSI) utilizes light energy to excite electrons for the reduction of NADP⁺, and like photosystem II, it is sensitive to excess light. When PSI is excited and unable to be reduced by the electron transport chain, the special pair of chlorophyll molecules, P700⁺, will take electrons from neighboring sources leading to cellular damage. A Chlamydomonas reinhardtii mutant, which is defective in the production of the PsaF subunit of PSI, provides an ideal platform for studying the processes involved in protecting PSI from excess light. This strain dies following the exposure to high light (HL) because of photo‐oxidative damage. We used a second‐site suppressor screen to identify genes involved in protecting PsaF‐deficient PSI from excess light. In doing so, we demonstrated that the absence of the STT7 protein, which is required for LHCII phosphorylation and the process of state transitions suppresses the psaF HL‐lethal phenotype. On the basis of chlorophyll fluorescence measurements, the psaF mutant has a more reduced plastoquinone pool at a given photosynthetic photon flux density than the wild‐type cells. Under these conditions the process of state transitions will become active, resulting in the transfer of phosphorylated LHCII proteins to PSI, further increasing the excitation of PSI. However, in the psaF stt7 double mutant, the LHCII proteins will not be transferred to PSI, and thus the level of PSI excitation will remain lower. This study provides clear genetic evidence that the HL‐lethal phenotype of the psaF mutant is because of PSI overexciation.

We demonstrate the capability of synchrotron-based Fourier-Transform Infrared spectromicroscopy to detect metabolite formation in single, living cells of the unicellular algae Chlamydomonas reinhardtii. We show that the high brightness of the source provides a sufficient signal-to-noise ratio to detect small molecular species accumulating in a spot about 15 µm in size. Time resolved measurements are carried out on cells grown heterotrophically under low-light conditions to study the evolution of products of anaerobic metabolism. The formation of small molecular species, including ethanol and at least one carbonyl containing compound, can be detected with a time resolution of the order of one minute.