Assessing the redox state of the plastoquinone pool in algae and cyanobacteria via OJIP fluorescence: perspectives and limitations
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Background
The redox state of the plastoquinone pool (PQ-redox) acts as a central element in a variety of intracellular signal pathways. Several methods for determining PQ-redox have been established. Although some methods are fully quantitative, such as those based on liquid chromatography, they are typically sensitive to sample preparation. Here, we critically evaluate the use of fast chlorophyll a fluorescence induction kinetics (the so-called OJIP transient) for semi-quantitative PQ-redox assessment in algae ( Chlorella vulgaris ) and cyanobacteria ( Synechocystis sp. PCC 6803). The method, based on the evaluation of relative fluorescence yield at the J-step of the OJIP transient (V J ), has already been reported; however, thus far, it has been used mostly for studying dark-acclimated leaves, which limits its range of use.
Results
In this work, we show that the OJIP transient can be used for semi-quantitative estimation of PQ-redox in algal and cyanobacterial cell cultures, in addition to plants. We further show that it can reflect PQ-redox in both dark-acclimated and light-acclimated samples. To estimate PQ-redox in light-acclimated cells, we introduce a new parameter V J ’. This parameter is derived from F O ’, F J ’ and F M ’ values – the fluorescence yields at the O and J steps of the OJIP transient, and the maximum fluorescence yield, respectively, measured in light-acclimated samples. Our systematic comparison of Multi-Color PAM, AquaPen, and FL 6000 fluorometers demonstrates that accurate measurement of V J and V J ’ in algal and cyanobacterial cultures requires low culture density and a high-intensity saturation pulse. We further show that with increasing light intensity to which the cells are exposed, the state of photosystem II (PSII) changes due to light-induced reduction of quinone A (Q − ) and conformational changes, which in turn influence both the sensitivity and dynamic range of the V J ’ parameter towards PQ-redox assessment. A comparison of fluorescence curves in algae and cyanobacteria revealed high homeostatic control over PQ-redox in Synechocystis sp. PCC 6803, maintained by terminal oxidases present at the thylakoid membrane.
Conclusions
While certain limitations exist, our critical assessment suggests that the OJIP method has great potential to become a routine tool for semi-quantitative PQ-redox assessment under a wide range of experimental conditions.
