Dataset Information

Absence of Flv3 results in metabolic and bioenergetic rearrangements towards mixotrophy in engineered sucrose-excreting Synechocystis sp. PCC6803

ABSTRACT: In the cyanobacterium Synechocystis sp. PCC 6803, the flavodiiron protein Flv3 functions as an alternative electron acceptor from PSI and represents an interesting engineering target for reorganizing the electron and carbon fluxes in the cell. We have shown that the inactivation of Flv3 in engineered sucrose-excreting Synechocystis (S02:Δflv3) triggers sucrose reuptake from the growth medium, and leads to a transition from photoautotrophy to mixotrophy. Such an acclimation maintains optimal growth of the S02:Δflv3 strain through enhanced respiratory metabolism evidenced by extensive but gradual proteomic and metabolomic changes over the course of the nine-day experiment. In the absence of Flv3, a downregulation of photosynthetic light reactions and CO2 assimilation with concurrent upregulation of glycolytic pathways occurred before any differences in sucrose production between S02 and S02:Δflv3 strains were observed. Subsequently, the components of the respiratory pathways showed distinct upregulation in S02:Δflv3, including the plastoquinone reductase complexes NDH-11 and NDH-2, which function at the interphase between glycolytic metabolism and the electron transport pathways, and the terminal respiratory oxidases Cyd and Cox, which direct electrons to O2. Increased oxygen consumption together with lower rates of photosynthetic light reactions generating oxygen led to activation fermentative pathways in S02:Δflv3 as an alternative source of energy for the cells. Our results show that although Synechocystis cannot directly compensate for the lack of Flv3 by other alternative electron acceptors under continuous high light, high CO2 and high osmotic pressure, the cells respond to the increased electron pressure by extensive reorganization of the electron transfer pathways. While glycolytic metabolism is significantly up-regulated to provide energy for the cell, the accumulation of storage compounds and increased respiration serve as indirect electron sinks for photosynthetic electrons that would normally be consumed by the Flv1/Flv3 heterodimer.

ORGANISM(S): Synechocystis Sp. Pcc 6803

SUBMITTER: Dorota Muth-Pawlak

PROVIDER: PXD050257 | panorama | Fri Oct 31 00:00:00 GMT 2025

REPOSITORIES: PanoramaPublic

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Interplay between photosynthetic electron flux and organic carbon sinks in sucrose-excreting Synechocystis sp. PCC 6803 revealed by omics approaches.

Muth-Pawlak Dorota D Kakko Lauri L Kallio Pauli P Aro Eva-Mari EM

Microbial cell factories 20240701 1

<h4>Background</h4>Advancing the engineering of photosynthesis-based prokaryotic cell factories is important for sustainable chemical production and requires a deep understanding of the interplay between bioenergetic and metabolic pathways. Rearrangements in photosynthetic electron flow to increase the efficient use of the light energy for carbon fixation must be balanced with a strong carbon sink to avoid photoinhibition. In the cyanobacterium Synechocystis sp. PCC 6803, the flavodiiron protein ...[more]

PMID: 38951789

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Project description:When the yeast Saccharomyces cerevisiae is subjected to increasing glycolytic fluxes under aerobic conditions, there is a threshold value of the glucose uptake rate at which the metabolism shifts from being purely respiratory to mixed respiratory and fermentative. This shift is characterized by ethanol production, a phenomenon known as the Crabtree effect due to its analogy with lactate overflow in cancer cells. It is well known that at high glycolytic fluxes there is glucose repression of respiratory pathways resulting in a decrease in the respiratory capacity. Despite many years of detailed studies on this subject, it is not known whether the onset of the Crabtree effect (or overflow metabolism) is due to a limited respiratory capacity or caused by glucose-mediated repression of respiration. We addressed this issue by increasing respiration in S. cerevisiae by introducing a heterologous alternative oxidase, and observed reduced aerobic ethanol formation. In contrast, increasing non-respiratory NADH oxidation by overexpression of a water-forming NADH oxidase reduced aerobic glycerol formation. The metabolic response to elevated alternative oxidase occurred predominantly in the mitochondria, while NADH oxidase affected genes that catalyze cytosolic reactions. Moreover, NADH oxidase restored the deficiency of cytosolic NADH dehydrogenases in S. cerevisiae. These results indicate that NADH oxidase localizes in the cytosol, while alternative oxidase is directed to the mitochondria. The onset of aerobic ethanol formation is demonstrated to be a consequence of an imbalance in mitochondrial redox balancing. In addition to answering fundamental physiological questions, our findings are relevant for all biomass derived applications of S. cerevisiae. Experiment Overall Design: Heterologous gene expression in chemostats using Affymetrix Yeast Genome 2.0 arrays. Total RNA extraction and sample preparation, hybridization was done according to the manufacturer's protocol.

Project description:Nogales2012 - Genome-scale metabolic network of Synechocystis sp. (iJN678) This model is described in the article: Detailing the optimality of photosynthesis in cyanobacteria through systems biology analysis. Nogales J, Gudmundsson S, Knight EM, Palsson BO, Thiele I. Proc. Natl. Acad. Sci. U.S.A. 2012 Feb; 109(7): 2678-2683 Abstract: Photosynthesis has recently gained considerable attention for its potential role in the development of renewable energy sources. Optimizing photosynthetic organisms for biomass or biofuel production will therefore require a systems understanding of photosynthetic processes. We reconstructed a high-quality genome-scale metabolic network for Synechocystis sp. PCC6803 that describes key photosynthetic processes in mechanistic detail. We performed an exhaustive in silico analysis of the reconstructed photosynthetic process under different light and inorganic carbon (Ci) conditions as well as under genetic perturbations. Our key results include the following. (i) We identified two main states of the photosynthetic apparatus: a Ci-limited state and a light-limited state. (ii) We discovered nine alternative electron flow pathways that assist the photosynthetic linear electron flow in optimizing the photosynthesis performance. (iii) A high degree of cooperativity between alternative pathways was found to be critical for optimal autotrophic metabolism. Although pathways with high photosynthetic yield exist for optimizing growth under suboptimal light conditions, pathways with low photosynthetic yield guarantee optimal growth under excessive light or Ci limitation. (iv) Photorespiration was found to be essential for the optimal photosynthetic process, clarifying its role in high-light acclimation. Finally, (v) an extremely high photosynthetic robustness drives the optimal autotrophic metabolism at the expense of metabolic versatility and robustness. The results and modeling approach presented here may promote a better understanding of the photosynthetic process. They can also guide bioengineering projects toward optimal biofuel production in photosynthetic organisms. This model is hosted on BioModels Database and identified by: MODEL1507180046. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

Dataset Information

Absence of Flv3 results in metabolic and bioenergetic rearrangements towards mixotrophy in engineered sucrose-excreting Synechocystis sp. PCC6803

Publications

Interplay between photosynthetic electron flux and organic carbon sinks in sucrose-excreting Synechocystis sp. PCC 6803 revealed by omics approaches.

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