Circadian clocks regulate biological activities, providing organisms a fitness advantage under diurnal changing conditions by allowing them to anticipate and adapt to recurring external changes. In recent years attention was drawn to the entrainment by intracellular cycles. Photosynthetic Cyanobacteria coordinate their gene expression, metabolism, and other activities in a circadian fashion. Solely, three proteins, KaiA, KaiB, and KaiC, constitute the well-studied circadian clock of the cyanobacterial model,Synechococcus elongatusPCC 7942. It remained inconclusive for a long time whetherSynechocystissp. PCC 6803, an important organism for biotechnological applications, can also maintain circadian rhythms under continuous illumination. Using an approach, which does not require genetic modification, we investigated the growth behavior ofSynechocystisvia non-invasive online backscattering measurement and verified all three criteria for true circadian oscillators: temperature compensation, entrainment by external stimuli, and a self-sustained freerunning period of about 24 hours. Since manipulation of the circadian clock (SynechocystisΔkaiA1B1C1) led to a significant reduction in glycogen content, disruption of glycogen synthesis (SynechocystisΔglgC) entirely inhibited glycogen formation and both mutants lost oscillations, we hypothesize that the oscillations reflect glycogen metabolism.
Monitoring circadian rhythms in cyanobacteria usually requires genetically modified reporter strains or intensive sampling for downstream analysis. Even for the main cyanobacterial modelSynechocystissp. PCC 6803 it was debated for years to which extent undamped circadian oscillations are really present until a suitable reporter strain was developed. We applied online backscatter measurements as an alternative readout to monitor circadian oscillations in cyanobacteria. InSynechocystisthe temperature-compensatedkaiA1B1C1-driven 24 h metabolic oscillations did not require light-dark entrainment, highlighting the relevance of the clock for the carbon metabolism even under continuous light, an aspect which should be considered for industrial set-ups. Our method opens the possibility to extend circadian analysis to non-GMO and monitor metabolic rhythmicity during high-density cultivation.
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