7th joint conference "Joint Microbiology & Infection Conference 2024" of the DGHM and VAAM

Prof. Dr.

Chair

PhotoSynthetics: Elucidating energy balancing mechanisms in cyanobacteria via metabolic engineering

Invited Speaker

Invited Talk

Abstract: Phycobilisomes (PBS) are the elaborated light-harvesting antennas in cyanobacteria. To balance the harvesting of light energy against the risks of photodamage, many cyanobacteria have evolved a photoprotective mechanism that relies on the interaction between a photoreceptor, the Orange Carotenoid Protein (OCP), and the PBS. Here we present four cryo-electron microscopy structures, with and without OCP, of the 6.2 MDa PBS from the model organism Synechocystis PCC 6803 at overall resolution 2.1-3.5 Å. The structures revealed the existence of three different conformational states of the antenna, two previously unknown, for the unquenched PBS. We found that two of the rods can switch conformation within the complex, suggestive of a potentially new type of regulation. We also discovered a novel linker protein, named ApcG, that binds to the membrane facing side of the PBS. In addition, the structure of the PBS-OCP complex shows four 34 kDa OCPs organized as two dimers quench the PBS. The complex also reveals for the first time, the structure of the active form of the OCP, revealing an ~60 Å displacement of its regulatory C-terminal domain. Finally, we elucidate energy transfer pathways based on structural and spectroscopic properties. These results provide detailed insights into the cyanobacterial light-harvesting and place a foundation for future bioengineering applications.

Revealing the structure of the light-harvesting antenna and its photoprotected state

Abstract: Because of their photosynthesis-dependent lifestyle, cyanobacteria evolved sophisticated regulatory mechanisms to adapt to oscillating day-night metabolic changes. How they coordinate the metabolic switch between autotrophic and glycogen-catabolic metabolism in light and darkness is poorly understood. Recently, we showed that the second messenger c-di-AMP is implicated in diurnal regulation1. To unravel the signaling functions of c-di-AMP, we identified the cyanobacterial c-di-AMP receptor proteins. The carbon-sensor protein SbtB was identified as a major c-di-AMP receptor1. We found that c-di-AMP–bound SbtB interacts with the glycogen-branching enzyme GlgB. Accordingly, both c-di-AMP-free (ΔdacA) and ΔsbtB–deficient mutants displayed impaired glycogen synthesis and nighttime survival1. To gain better understanding of cellular processes regulated by SbtB or c-di-AMP, we compared the metabolomic, transcriptomics, and proteomic landscapes of both mutants2,3. While our results indicate that the cellular roles of SbtB is restricted to carbon/glycogen metabolism1-5, the ΔdacA lethality seems a result of dysregulation of multiple metabolic processes2,3. These processes include photosynthesis and redox regulation, which lead to elevated levels of intracellular ROS and glutathione2. Further, we showed an impact of c-di-AMP on transcription/transitional regulations, ion homeostasis, and especially on central carbon metabolism. Adding to its cellular functions catalog, for first time, we show physiologically that c-di-AMP signaling influences on nitrogen metabolism, implying new cellular roles for c-di-AMP in controlling nitrogen homeostasis2. Additionally, we found that the cyanobacterial pilus biogenesis and natural competence are regulated by c-di-AMP and show that the ComFB signaling protein is a novel c-di-AMP-receptor protein6, widespread in bacterial phyla, and required for DNA uptake. 
 
1.	Selim KA, Haffner M, Burkhardt M, Mantovani O, Neumann N, Albrecht R, Seifert R, Krüger L, Stülke J, Hartmann MD, Hagemann M, Forchhammer K. Diurnal metabolic control in cyanobacteria requires perception of second messenger signaling molecule c-di-AMP by the carbon control protein SbtB. Sci Adv. 2021; 7(50):eabk0568.
2.	Haffner M, Mantovani O, Spät P, Maček B, Hagemann M, Forchhammer K, Selim KA. Diurnal rhythm causes metabolic crises in the cyanobacterial mutants of c-di-AMP signalling cascade. BioRxiv 2023; doi: https://doi.org/10.1101/2023.11.14.567006.
3.	Mantovani O, Reimann V, Haffner M, Herrmann FP, Selim KA, Forchhammer K, Hess WR, Hagemann M. The impact of the cyanobacterial carbon-regulator protein SbtB and of the second messengers cAMP and c-di-AMP on CO2 -dependent gene expression. New Phytol. 2022; 234(5):1801-1816.
4.	Selim KA, Haffner M, Mantovani O, Albrecht R, Zhu H, Hagemann M, Forchhammer K, Hartmann MD. Carbon signaling protein SbtB possesses atypical redox-regulated apyrase activity to facilitate regulation of bicarbonate transporter SbtA. PNAS. 2023; 120(8):e2205882120.
5.	Selim KA, Haase F, Hartmann MD, Hagemann M, Forchhammer K. PII-like signaling protein SbtB links cAMP sensing with cyanobacterial inorganic carbon response. PNAS. 2018; 115(21):E4861-E4869.
6.	Samir S, Doello S, Zimmer E, Haffner M, Enkerlin AM, Müller T, Dengler L, Lambidis SP, Sivabalasarma S, Albers S-V, Selim KA. The second messenger c-di-AMP controls natural competence via ComFB signaling protein. BioRxiv 2023; doi: https://doi.org/10.1101/2023.11.27.568819 

c-di-AMP signaling in cyanobacteria: a new paradigm in controlling cellular homeostasis

Jun.-Prof. Dr.

Abstract: Cyanobacteria use a circadian clock to predict environmental daily cycles and organize a multitude of cellular processes in matching daily rhythms. In the clock model organism Synechococcus elongatus PCC 7942 the clock is composed of the three proteins KaiA, KaiB and KaiC. Some cyanobacteria express multiple diverged Kai proteins, which implies that circadian regulation might be more complex in those organisms. In Synechocystis sp. PCC 6803 three KaiB and KaiC proteins are present. We chose it as a representative to study the function and interplay of diverged clock proteins in an otherwise well characterized model organism (1- 4). Based on homology searches, it was believed that all cyanobacteria contain only one copy of KaiA, but we discovered a second KaiA protein (KaiA3) in Synechocystis (5). Only the C-terminal domain of this new chimeric KaiA3 protein resembles a KaiA domain. The N-terminal domain is similar to a NarL-type response regulator receiver domain. Mixing KaiA3, KaiB3 and KaiC3, we could reconstitute low amplitude, temperature compensated ~24h oscillations of KaiC3 phosphorylation in vitro. This demonstrates that KaiA3B3C3 form a bona fide circadian oscillator. In the cell, KaiC3 displayed 24h phosphorylation rhythms that were phase-locked with KaiC1 phosphorylation cycles. Circadian rhythms can be simply read out via online monitoring of the backscatter properties of cells (6). Studies of kai gene deletion strains indicated that the KaiA1B1C1 oscillator and KaiA3B3C3 oscillator are both required to maintain stable backscatter oscillations, which implies that two interconnected oscillators are present in Synechocystis.  We are currently investigating the interplay of these two oscillators. 
 
(1) Wiegard A, Dörrich AK, Deinzer HT, Beck C, Wilde A, Holtzendorff J, Axmann IM. Microbiology (Reading). 2013 May;159(Pt 5):948-958. doi: 10.1099/mic.0.065425-0.  
(2) Beck C, Hertel S, Rediger A, Lehmann R, Wiegard A, Kölsch A, Heilmann B, Georg J, Hess WR, Axmann IM. Appl Environ Microbiol. 2014 Sep;80(17):5195-206. doi: 10.1128/AEM.01086-14.  
(3) Schmelling NM, Lehmann R, Chaudhury P, Beck C, Albers SV, Axmann IM, Wiegard A. BMC Evol Biol. 2017 Jul 21;17(1):169. doi: 10.1186/s12862-017-0999-7. P 
(4) Wiegard A, Köbler C, Oyama K, Dörrich AK, Azai C, Terauchi K, Wilde A, Axmann IM.  J Bacteriol. 2020 Jan 29;202(4):e00478-19. doi: 10.1128/JB.00478-19.  
(5) Köbler , Schmelling NM, Pawlowski A, Spät P, Scheurer NM, Sebastian K, Berwanger LC, Maček B, Wiegard A, Axmann IM, Wilde A. preprint,  bioRxiv 2021.07.20.453058; doi: https://doi.org/10.1101/2021.07.20.453058 
(6) Berwanger LC, Thumm N, Gholamipoor R, Wiegard A, Schlebusch J, Kollmann M, Axmann IM. preprint, bioRxiv 2023.09.26.559469; doi: https://doi.org/10.1101/2023.09.26.559469 

Symposium
Molecular Mechanisms of Cyanobacterial Photosynthesis and Metabolism

Termin

Chair

Programm

SymposiumMolecular Mechanisms of Cyanobacterial Photosynthesis and Metabolism

Termin

Chair

Programm

Symposium
Molecular Mechanisms of Cyanobacterial Photosynthesis and Metabolism