Unknown,Transcriptomics,Genomics,Proteomics

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Transcriptomic response to long time ethanol production in the cyanobacterium Synechocystis sp. PCC6803

ABSTRACT: The production of biofuels in photosynthetic microalgae and cyanobacteria is considered a promising alternative to the generation of fuels from fossil resources. However, to be economically competitive, producer strains need to be established that synthesize the targeted product at high yield and over a long time. Engineering cyanobacteria to forced fuel producers should considerably interfere with overall cell homeostasis, which in turn might counteract productivity and sustainability of the process. Therefore, in-depth characterization of the cellular response upon long-term production is of high interest for the targeted improvement of a desired strain. Here we report the results of a monitoring experiment, in which the transcriptome-wide response to continuous ethanol production in the unicellular model cyanobacterium Synechocystis sp. PCC6803 was examined using high resolution microarrays. In two independent experiments, ethanol production rates of 0.0338% (v/v) EtOH d-1 M-BM-1 0.002 and 0.0303% (v/v) d-1 M-BM-1 0.002 were obtained over 18 consecutive days, measuring biological triplicates in fully automated photobioreactors. Ethanol production caused a significant (~40%) delay in biomass accumulation in the producer strains and the development of a bleaching phenotype. Absorption spectroscopy indicated in particular a down-regulation of light harvesting capacity. Microarray analyses performed at day 4, 7, 11 and 18 of the experiment revealed only three mRNAs with a strongly modified accumulation level throughout the course of the experiment. In addition to the overexpressed adhA (slr1192) gene, this was an about 4 fold reduction in cpcB (sll1577) and an about 3 to 6 fold increase in rps8 (sll1809) mRNA levels. Much weaker modifications of expression level or modifications restricted to day 18 of the experiment were observed for genes involved in carbon assimilation (Ribulose bisphosphate carboxylase and Glutamate decarboxylase). Molecular analysis of the reduced cpcB levels revealed a post-transcriptional operon discoordination in the cpcBA operon leaving a truncated mRNA cpcA* likely not competent for translation. Moreover, Western blots and zinc-enhanced bilin fluorescence blots confirmed a severe reduction in the amounts of both phycocyanin subunits, explaining the cause of the bleaching phenotype. We conclude that the changes in gene expression upon induction of long term ethanol production in Synechocystis sp. PCC6803 are highly specific. in particular we did not observe a comprehensive stress response contributing to a complex phenotype as might have been expected. Gene expression of Synechocystis sp. PCC 6803 WT (#621) and the isogenic ethanol producing strain #309 was monitored at 4, 7, 11 and 18 days after induction of ethanol production by copper depletion. Each condition was sampled in biological duplicates

ORGANISM(S): Synechocystis sp. PCC 6803

SUBMITTER: Jens Georg

PROVIDER: E-GEOD-49552 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Montagud2010 - Genome-scale metabolic network of Synechocystis sp. PCC6803 (iSyn669) This model is described in the article: Reconstruction and analysis of genome-scale metabolic model of a photosynthetic bacterium. Montagud A, Navarro E, Fernández de Córdoba P, Urchueguía JF, Patil KR. BMC Syst Biol 2010; 4: 156 Abstract: BACKGROUND: Synechocystis sp. PCC6803 is a cyanobacterium considered as a candidate photo-biological production platform--an attractive cell factory capable of using CO2 and light as carbon and energy source, respectively. In order to enable efficient use of metabolic potential of Synechocystis sp. PCC6803, it is of importance to develop tools for uncovering stoichiometric and regulatory principles in the Synechocystis metabolic network. RESULTS: We report the most comprehensive metabolic model of Synechocystis sp. PCC6803 available, iSyn669, which includes 882 reactions, associated with 669 genes, and 790 metabolites. The model includes a detailed biomass equation which encompasses elementary building blocks that are needed for cell growth, as well as a detailed stoichiometric representation of photosynthesis. We demonstrate applicability of iSyn669 for stoichiometric analysis by simulating three physiologically relevant growth conditions of Synechocystis sp. PCC6803, and through in silico metabolic engineering simulations that allowed identification of a set of gene knock-out candidates towards enhanced succinate production. Gene essentiality and hydrogen production potential have also been assessed. Furthermore, iSyn669 was used as a transcriptomic data integration scaffold and thereby we found metabolic hot-spots around which gene regulation is dominant during light-shifting growth regimes. CONCLUSIONS: iSyn669 provides a platform for facilitating the development of cyanobacteria as microbial cell factories. This model is hosted on BioModels Database and identified by: MODEL1507180008. 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.

Project description:The production of biofuels in photosynthetic microalgae and cyanobacteria is considered a promising alternative to the generation of fuels from fossil resources. However, to be economically competitive, producer strains need to be established that synthesize the targeted product at high yield and over a long time. Engineering cyanobacteria to forced fuel producers should considerably interfere with overall cell homeostasis, which in turn might counteract productivity and sustainability of the process. Therefore, in-depth characterization of the cellular response upon long-term production is of high interest for the targeted improvement of a desired strain. Here we report the results of a monitoring experiment, in which the transcriptome-wide response to continuous ethanol production in the unicellular model cyanobacterium Synechocystis sp. PCC6803 was examined using high resolution microarrays. In two independent experiments, ethanol production rates of 0.0338% (v/v) EtOH d-1 ± 0.002 and 0.0303% (v/v) d-1 ± 0.002 were obtained over 18 consecutive days, measuring biological triplicates in fully automated photobioreactors. Ethanol production caused a significant (~40%) delay in biomass accumulation in the producer strains and the development of a bleaching phenotype. Absorption spectroscopy indicated in particular a down-regulation of light harvesting capacity. Microarray analyses performed at day 4, 7, 11 and 18 of the experiment revealed only three mRNAs with a strongly modified accumulation level throughout the course of the experiment. In addition to the overexpressed adhA (slr1192) gene, this was an about 4 fold reduction in cpcB (sll1577) and an about 3 to 6 fold increase in rps8 (sll1809) mRNA levels. Much weaker modifications of expression level or modifications restricted to day 18 of the experiment were observed for genes involved in carbon assimilation (Ribulose bisphosphate carboxylase and Glutamate decarboxylase). Molecular analysis of the reduced cpcB levels revealed a post-transcriptional operon discoordination in the cpcBA operon leaving a truncated mRNA cpcA* likely not competent for translation. Moreover, Western blots and zinc-enhanced bilin fluorescence blots confirmed a severe reduction in the amounts of both phycocyanin subunits, explaining the cause of the bleaching phenotype. We conclude that the changes in gene expression upon induction of long term ethanol production in Synechocystis sp. PCC6803 are highly specific. in particular we did not observe a comprehensive stress response contributing to a complex phenotype as might have been expected.

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Transcriptomic response to long time ethanol production in the cyanobacterium Synechocystis sp. PCC6803

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