Enhanced stable production of ethylene in photosynthetic cyanobacterium Synechococcus elongatus PCC 7942

Carbonell, Verónica; Vuorio, E; Aro, Eva‐Mari; Kallio, Pauli T.

doi:10.1007/s11274-019-2652-7

Cited by 25 publications

(22 citation statements)

References 41 publications

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“…In agreement with an earlier study [38], the addition of arginine did not increase the ethylene productivity, thus this amino acid is not a limiting substrate in the engineered strains under our standard treatment. However, when in the same study 2-oxoglutarate and arginine were both added to the cells, ethylene productivity was increased [38]. Thus, the combination of both substrates enhances the ethylene production.…”

Section: Discussionsupporting

confidence: 93%

“…In order to discover which are the bottlenecks for ethylene production, we decided to investigate the effect of increasing the concentrations of the two substrates of Efe, 2-oxoglutarate and arginine. It has been observed that addition of 2-oxoglutarate did not increase ethylene production in Synechocystis but it did in Synechococcus [38]. In addition, it is known that when cultivating the cells without a nitrogen source, the 2-oxoglutarate levels increase when a switch in metabolism occurs [21,39,40].…”

Section: Discussionmentioning

confidence: 99%

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Increased ethylene production by overexpressing phosphoenolpyruvate carboxylase in the cyanobacterium Synechocystis PCC 6803

Durall

Lindberg

et al. 2020

Biotechnol Biofuels

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Background: Cyanobacteria can be metabolically engineered to convert CO 2 to fuels and chemicals such as ethylene. A major challenge in such efforts is to optimize carbon fixation and partition towards target molecules. Results: The efe gene encoding an ethylene-forming enzyme was introduced into a strain of the cyanobacterium Synechocystis PCC 6803 with increased phosphoenolpyruvate carboxylase (PEPc) levels. The resulting engineered strain (CD-P) showed significantly increased ethylene production (10.5 ± 3.1 µg mL −1 OD −1 day −1) compared to the control strain (6.4 ± 1.4 µg mL −1 OD −1 day −1). Interestingly, extra copies of the native pepc or the heterologous expression of PEPc from the cyanobacterium Synechococcus PCC 7002 (Synechococcus) in the CD-P, increased ethylene production (19.2 ± 1.3 and 18.3 ± 3.3 µg mL −1 OD −1 day −1 , respectively) when the cells were treated with the acetyl-CoA carboxylase inhibitor, cycloxydim. A heterologous expression of phosphoenolpyruvate synthase (PPSA) from Synechococcus in the CD-P also increased ethylene production (16.77 ± 4.48 µg mL −1 OD −1 day −1) showing differences in the regulation of the native and the PPSA from Synechococcus in Synechocystis. Conclusions: This work demonstrates that genetic rewiring of cyanobacterial central carbon metabolism can enhance carbon supply to the TCA cycle and thereby further increase ethylene production.

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Section: Discussionsupporting

confidence: 93%

Section: Discussionmentioning

confidence: 99%

Increased ethylene production by overexpressing phosphoenolpyruvate carboxylase in the cyanobacterium Synechocystis PCC 6803

Durall

Lindberg

et al. 2020

Biotechnol Biofuels

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“…Although S. elongatus is of great scientific interest in the context of direct bioconversion of carbon dioxide into fuels and plastic precursors (Atsumi et al ., 2009; Carbonell et al ., 2019), metabolic engineering to enhance production of fatty acids (Santos‐Merino et al ., 2018), circadian rhythm biology (Fleming and O'Shea, 2018) and many additional applications, its interaction with mammalian immunological processes has not previously been detailed. This study comprehensively assessed the safety of intravenous administration of S. elongatus cyanobacteria in rats in vivo to inform future exploration of this organism's utility in symbiotic therapeutics.…”

Section: Discussionmentioning

confidence: 99%

Safety of photosynthetic Synechococcus elongatus for in vivo cyanobacteria–mammalian symbiotic therapeutics

Williams

Wang

Paulsen

et al. 2020

Microbial Biotechnology

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The cyanobacterium Synechococcus elongatus (SE) has been shown to rescue ischaemic heart muscle after myocardial infarction by photosynthetic oxygen production. Here, we investigated SE toxicity and hypothesized that systemic SE exposure does not elicit a significant immune response in rats. Wistar rats intravenously received SE (n = 12), sterile saline (n = 12) or E. coli lipopolysaccharide (LPS, n = 4), and a subset (8 SE, 8 saline) received a repeat injection 4 weeks later. At baseline, 4 h, 24 h, 48 h, 8 days and 4 weeks after injection, clinical assessments, blood cultures, blood counts, lymphocyte phenotypes, liver function tests, proinflammatory cytokines and immunoglobulins were assessed. Across all metrics, SE rats responded comparably to saline controls, displaying no clinically significant immune response. As expected, LPS rats exhibited severe immunological responses. Systemic SE administration does not induce sepsis or toxicity in rats, thereby supporting the safety of cyanobacteria-mammalian symbiotic therapeutics using this organism.

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“…In integrative systems, the challenge is to find a redundant genomic locus ( neutral site ), that can be disrupted without compromising native functions that would affect maintenance and cell growth [ 3 ], or a gene of known function that is not required under the specific cultivation conditions used. Applicable target sites are therefore strain-specific, and linked with the experimental setup, that may influence production efficiency and system stability [ 4 ]. In addition, in polyploid organisms such as the model cyanobacterium Synechocystis sp.…”

Section: Introductionmentioning

confidence: 99%

Comparison of alternative integration sites in the chromosome and the native plasmids of the cyanobacterium Synechocystis sp. PCC 6803 in respect to expression efficiency and copy number

et al. 2021

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Background Synechocystis sp. PCC 6803 provides a well-established reference point to cyanobacterial metabolic engineering as part of basic photosynthesis research, as well as in the development of next-generation biotechnological production systems. This study focused on expanding the current knowledge on genomic integration of expression constructs in Synechocystis, targeting a range of novel sites in the chromosome and in the native plasmids, together with established loci used in literature. The key objective was to obtain quantitative information on site-specific expression in reference to replicon copy numbers, which has been speculated but never compared side by side in this host. Results An optimized sYFP2 expression cassette was successfully integrated in two novel sites in Synechocystis chromosome (slr0944; sll0058) and in all four endogenous megaplasmids (pSYSM/slr5037-slr5038; pSYSX/slr6037; pSYSA/slr7023; pSYSG/slr8030) that have not been previously evaluated for the purpose. Fluorescent analysis of the segregated strains revealed that the expression levels between the megaplasmids and chromosomal constructs were very similar, and reinforced the view that highest expression in Synechocystis can be obtained using RSF1010-derived replicative vectors or the native small plasmid pCA2.4 evaluated in comparison. Parallel replicon copy number analysis by RT-qPCR showed that the expression from the alternative loci is largely determined by the gene dosage in Synechocystis, thereby confirming the dependence formerly proposed based on literature. Conclusions This study brings together nine different integrative loci in the genome of Synechocystis to demonstrate quantitative differences between target sites in the chromosome, the native plasmids, and a RSF1010-based replicative expression vector. To date, this is the most comprehensive comparison of alternative integrative sites in Synechocystis, and provides the first direct reference between expression efficiency and replicon gene dosage in the context. In the light of existing literature, the findings support the view that the small native plasmids can be notably more difficult to target than the chromosome or the megaplasmids, and that the RSF1010-derived vectors may be surprisingly well maintained under non-selective culture conditions in this cyanobacterial host. Altogether, the work broadens our views on genomic integration and the rational use of different integrative loci versus replicative plasmids, when aiming at expressing heterologous genes in Synechocystis.

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Enhanced stable production of ethylene in photosynthetic cyanobacterium Synechococcus elongatus PCC 7942

Cited by 25 publications

References 41 publications

Increased ethylene production by overexpressing phosphoenolpyruvate carboxylase in the cyanobacterium Synechocystis PCC 6803

Increased ethylene production by overexpressing phosphoenolpyruvate carboxylase in the cyanobacterium Synechocystis PCC 6803

Safety of photosynthetic Synechococcus elongatus for in vivo cyanobacteria–mammalian symbiotic therapeutics

Comparison of alternative integration sites in the chromosome and the native plasmids of the cyanobacterium Synechocystis sp. PCC 6803 in respect to expression efficiency and copy number

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