Mechanismus und Funktion von nicht-standard circadianen Uhrsystemen in Cyanobakterien
Biochemie und Biophysik der Pflanzen
Zusammenfassung der Projektergebnisse
Through the rising and setting of the sun, most organisms on Earth are subjected to predictable environmental changes, such as light and darkness or temperature variations during a day and night cycle. To adapt to these ever-changing conditions, eukaryotic and prokaryotic organisms evolved endogenous biological clock systems. Currently, cyanobacteria are the only prokaryotes exhibiting a well-understood and robust circadian rhythm. Cyanobacteria are organisms which use, just like plants, the energy of the sun, water and carbon dioxide to proliferate in nearly every environment and ecological niche on our Earth. Using the power of photosynthesis, they strongly contribute to carbon dioxide fixation and oxygen production in the ocean and offer a large potential for biotechnological applications to produce clean and sustainable energy and other valuable products. The cyanobacterial circadian clock is fundamentally different from eukaryotic biological clock systems. The central oscillator which defines the timer is encoded by only three genes, named kaiA, kaiB and kaiC. The KaiC protein forms a homohexamer and harbors three enzymatic activities: autokinase, autophosphatase and ATPase activity. Through binding to KaiC, KaiA and KaiB modulate the autokinase and ATPase activities of KaiC and thus its phosphorylation state. The central oscillator then controls rhythms of biological activity, such as gene expression. In this project, we combined bioinformatics expertise with experimental science in order to reveal the function of divergent clock proteins. Our bioinformatic analyses defined the minimal core set of a prokaryotic clock and gave insights into shared and distinct features of timing systems in different prokaryotes, including also heterotrophic bacteria and Archaea. In the following we focused on the model cyanobacterial strain Synechocystis, which harbors more than one set of clock components. Several cyanobacteria and other prokaryotes harbor homologs of such non-standard clock proteins but their function is largely unknown. We demonstrated that the highly conserved output clock component, which is a transcription factor called RpaA, connects the standard oscillator (KaiA-KaiB1-KaiC1) to regulation of gene expression, thereby controlling mainly dark metabolism and carbon supply. Deletion of the second (kaiB2C2) cluster was not possible in Synechocystis, implying an essential but nonclock related function. Most importantly, we demonstrated that the non-standard KaiB3-KaiC3 system interacts with the standard oscillator and modulates its activity. Respective mutants show deficiencies when grown in the dark with glucose as energy and carbon source. Autophosphorylation and ATPase activity of KaiC3 was independent of KaiA suggesting that this system might work as an hourglass rather than as a bona fide oscillator. But surprisingly, using bioinformatics we have identified a new putative component of the KaiB3-KaiC3 system which might fulfill a KaiA-like function and which we showed to occur also with other nonstandard clock systems. The function of this new component awaits further analyses. Our analyses of the multi-timer setup in Synechocystis revealed that this complex regulatory system might represent the base for phenotypic plasticity for an organism which can switch between photoautotrophic and heterotrophic metabolic modes. Therefore, manipulation of the clock can be used to redirect biochemical pathways for metabolic optimization of biotechnologically important production strains.
Projektbezogene Publikationen (Auswahl)
- (2017) Minimal tool set for a prokaryotic circadian clock. BMC Evolutionary Biology. 1, 169
Schmelling NM, Lehmann R, Chaudhury P, Beck C, Albers SV, Axmann IM, Wiegard A
(Siehe online unter https://doi.org/10.1186/s12862-017-0999-7) - (2017) Structures of the cyanobacterial circadian oscillator frozen in a fully assembled state. Science. 6330, 1181-1184
Snijder J, Schuller JM, Wiegard A, Lössl P, Schmelling N, Axmann IM, Plitzko JM, Förster F, Heck AJ
(Siehe online unter https://doi.org/10.1126/science.aag3218) - (2018) Computational modelling unravels the precise clockwork of cyanobacteria. Interface Focus. 571, 20180038
Schmelling NM and Axmann IM
(Siehe online unter https://doi.org/10.1098/rsfs.2018.0038) - (2018) The role of the Synechocystis sp. PCC 6803 homolog of the circadian clock output regulator RpaA in day-night transitions. Mol. Microbiol. 110, 847-861
Köbler C, Schultz SJ, Kopp D, Voigt K, Wilde A
(Siehe online unter https://doi.org/10.1111/mmi.14129) - (2019) The Kai-protein clock - Keeping track of cyanobacteria’s daily life. In: Harris J, Marles-Wright J (eds) Macromolecular protein complexes II: Structure and function. Subcellular Biochemistry, Springer. 93, pp 359-391
Snijder J, Axmann IM
(Siehe online unter https://doi.org/10.1007/978-3-030-28151-9_12) - (2020) Synechocystis KaiC3 displays temperature and KaiB dependent ATPase activity and is important for growth in darkness. J. Bacteriol. 202, e00478-19
Wiegard A, Köbler C, Oyama K, Dörrich AK, Azai C, Terauchi K, Wilde A, Axmann IM
(Siehe online unter https://doi.org/10.1128/JB.00478-19)
