Energetic responses to transient high temperature stress in cyanobacteria. A dynamic system examined in vivo

Abstract

This thesis investigated the energetic response of a moderately thermophillic cyanobacterium, Thermosynechococcus elongatus BP.1 (BP.1), to high-temperature damage. Previous published work has investigated the physiological impacts of high-temperature stress on BP.1, and has focused primarily on understanding the absolute functional stability of discrete cellular components, exposed to high temperatures. The majority of this work was conducted in vitro, on isolated components, or through the addition of artificial electron donors and acceptors to cells. The results have clearly shown that photosystem two (PSII) is the thermally weakest cellular component of BP.1. Furthermore, the temperatures that inhibit PSII are very close to the optimal temperature for growth in this species. Strong evidence suggests that thylakoid components downstream of PSII, specifically photosystem one (PSI), and primary components of the electron transport chain, exhibit significantly greater tolerance to high temperatures than PSII. There has been no published work investigating the possible physiological significance of this. Furthermore, no data has been published on the tolerance to high temperatures of crucial cellular processes, carbon fixation and respiration. A key aim of the research was to identify whether any cellular functions, such as respiration, still operated in cells after high temperatures had inhibited PSII. To undertake this work, an in vivo methodology was developed to measure functionality of key cellular components, following high-temperature exposure. The method involved the integration of membrane inlet mass spectrometry (MIMS), including simultaneous gas flux measurements of CO2 and stable isotope differentiation of concurrent oxygen evolution and consumption fluxes, with pulse/probe spectroscopy of P700, the reaction-centre chlorophyll of PSI. Gas fluxes and P700 redox dynamics were measured from samples under steady state conditions of photosynthesis, following ten minutes of dark incubation across a range of damaging temperatures. The results showed clearly that respiratory systems, and probably CO2 fixation, were more stable than PSII. Furthermore, as PSII function declined to < 95 % of maximal rates, respiration was no longer inhibited by illumination. viii The P700 data indicated that PSI activity was maintained at a level that could not be supported solely by the remaining function of PSII, following high-temperature incubations. This suggested that published in vitro data, demonstrating greater thermal tolerance for PSI than PSII, may be functionally significant. Evidence that reductant sourced from respiration was driving PSI photochemistry, after PSII inhibition, was obtained. A hypothesis is proposed that the cyanobacterium utilises stored reductant to poise the thylakoid membrane for cyclic electron flux (CEF), once PSII is inhibited. The mechanism proposed in this thesis enables cells to maximise the utilisation of their stored energy, by enabling PSI to continue harnessing light energy if PSII is inhibited. A thylakoid proton gradient is maintained through PSI function, instead of standard chlororespiratory pathways that utilise a terminal oxidase. A new term, “delayed cyclic electron flux”, (CEFd) is proposed to describe this mechanism. It differentiates CEF around PSI, driven by PSII, from that supported by alternative sources of reductant, such as the respiratory complex succinate dehydrogenase (SDH). Delayed implies that the reductant has been stored in the cell.