Function and Benefits of Natural Competence in Cyanobacteria: From Ecology to Targeted Manipulation

Schirmacher, Alexandra M.; Hanamghar, Sayali S; Zedler, Julie A Z

doi:10.3390/life10110249

Cited by 15 publications

(17 citation statements)

References 118 publications

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“…Albeit the mechanisms of transformation based on natural competence are not experimentally verified for Synechocystis , the anticipated process likely results in the linearization of supplied plasmids, which are then taken up as single-stranded DNA. , Consequently, the linearized, single-stranded DNA needs to be repaired to maintain a replicative plasmid within the cell. This, however, might interfere with the overall transformation procedure and/or raise the possibility of recombination events, e.g., into the endogenous plasmids pCA2.4, pCB2.4, or pCC5.2.…”

Section: Discussionmentioning

confidence: 99%

“…Some cyanobacteria, including Synechocystis, are naturally competent, enabling the uptake of exogenous DNA and to integrate it into their genome via homologous recombination. 15 Although neutral chromosomal sites for the integration of heterologous genes devoid of pleiotropic effects are available, 16 the method is impaired by chromosome polyploidy, 17 which requires additional alternatives to introduce genes. In this regard, replicative plasmids could be used.…”

Section: ■ Introductionmentioning

confidence: 99%

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Generation of Synthetic Shuttle Vectors Enabling Modular Genetic Engineering of Cyanobacteria

et al. 2022

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Cyanobacteria have raised great interest in biotechnology due to their potential for a sustainable, photosynthesis-driven production of fuels and value-added chemicals. This has led to a concomitant development of molecular tools to engineer the metabolism of those organisms. In this regard, however, even cyanobacterial model strains lag behind compared to their heterotrophic counterparts. For instance, replicative shuttle vectors that allow gene transfer independent of recombination into host DNA are still scarce. Here, we introduce the pSOMA shuttle vector series comprising 10 synthetic plasmids for comprehensive genetic engineering of Synechocystis sp. PCC 6803. The series is based on the small endogenous plasmids pCA2.4 and pCB2.4, each combined with a replicon from Escherichia coli, different selection markers as well as features facilitating molecular cloning and the insulated introduction of gene expression cassettes. We made use of genes encoding green fluorescent protein (GFP) and a Baeyer−Villiger monooxygenase (BVMO) to demonstrate functional gene expression from the pSOMA plasmids in vivo. Moreover, we demonstrate the expression of distinct heterologous genes from individual plasmids maintained in the same strain and thereby confirmed compatibility between the two pSOMA subseries as well as with derivatives of the broad-host-range plasmid RSF1010. We also show that gene transfer into the filamentous model strain Anabaena sp. PCC 7120 is generally possible, which is encouraging to further explore the range of cyanobacterial host species that could be engineered via pSOMA plasmids. Altogether, the pSOMA shuttle vector series displays an attractive alternative to existing plasmid series and thus meets the current demand for the introduction of complex genetic setups and to perform extensive metabolic engineering of cyanobacteria.

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Section: Discussionmentioning

confidence: 99%

Section: ■ Introductionmentioning

confidence: 99%

Generation of Synthetic Shuttle Vectors Enabling Modular Genetic Engineering of Cyanobacteria

et al. 2022

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“…T4P are also known to be important for natural competence (Piepenbrink, 2019; Schirmacher et al., 2020; Yoshihara et al., 2001). Therefore, we investigated the ability of the minor pilin mutants described above to be transformed by exogenous DNA.…”

Section: Resultsmentioning

confidence: 99%

Minor pilins are involved in motility and natural competence in the cyanobacterium Synechocystis sp. PCC 6803

Oeser

Wallner

Schuergers

et al. 2021

Molecular Microbiology

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Cyanobacteria synthesize type IV pili, which are known to be essential for motility, adhesion and natural competence. They consist of long flexible fibers that are primarily composed of the major pilin PilA1 in Synechocystis sp. PCC 6803. In addition, Synechocystis encodes less abundant pilin‐like proteins, which are known as minor pilins. In this study, we show that the minor pilin PilA5 is essential for natural transformation but is dispensable for motility and flocculation. In contrast, a set of minor pilins encoded by the pilA9‐slr2019 transcriptional unit are necessary for motility but are dispensable for natural transformation. Neither pilA5‐pilA6 nor pilA9‐slr2019 are essential for pilus assembly as mutant strains showed type IV pili on the cell surface. Three further gene products with similarity to PilX‐like minor pilins have a function in flocculation of Synechocystis. The results of our study indicate that different minor pilins facilitate distinct pilus functions. Further, our microarray analysis demonstrated that the transcription levels of the minor pilin genes change in response to surface contact. A total of 122 genes were determined to have altered transcription between planktonic and surface growth, including several plasmid genes which are involved exopolysaccharide synthesis and the formation of bloom‐like aggregates.

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“…The molecular mechanism of natural transformation has been recently described in Synechocystis PCC 6803 [ 248 ]. First, DNA uptake from the environment is mediated by binding to the (type IV) pili appendages, which are also involved in cell adhesion and biofilm formation as well as twitching motility.…”

Section: Genetic Characteristics Of the Model Cyanobacteria Synechocystis Pcc 6803 Synechococcus Pcmentioning

confidence: 99%

Genetic, Genomics, and Responses to Stresses in Cyanobacteria: Biotechnological Implications

Cassier‐Chauvat

Blanc-Garin

Chauvat

2021

Genes

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Cyanobacteria are widely-diverse, environmentally crucial photosynthetic prokaryotes of great interests for basic and applied science. Work to date has focused mostly on the three non-nitrogen fixing unicellular species Synechocystis PCC 6803, Synechococcus PCC 7942, and Synechococcus PCC 7002, which have been selected for their genetic and physiological interests summarized in this review. Extensive “omics” data sets have been generated, and genome-scale models (GSM) have been developed for the rational engineering of these cyanobacteria for biotechnological purposes. We presently discuss what should be done to improve our understanding of the genotype-phenotype relationships of these models and generate robust and predictive models of their metabolism. Furthermore, we also emphasize that because Synechocystis PCC 6803, Synechococcus PCC 7942, and Synechococcus PCC 7002 represent only a limited part of the wide biodiversity of cyanobacteria, other species distantly related to these three models, should be studied. Finally, we highlight the need to strengthen the communication between academic researchers, who know well cyanobacteria and can engineer them for biotechnological purposes, but have a limited access to large photobioreactors, and industrial partners who attempt to use natural or engineered cyanobacteria to produce interesting chemicals at reasonable costs, but may lack knowledge on cyanobacterial physiology and metabolism.

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Function and Benefits of Natural Competence in Cyanobacteria: From Ecology to Targeted Manipulation

Cited by 15 publications

References 118 publications

Generation of Synthetic Shuttle Vectors Enabling Modular Genetic Engineering of Cyanobacteria

Generation of Synthetic Shuttle Vectors Enabling Modular Genetic Engineering of Cyanobacteria

Minor pilins are involved in motility and natural competence in the cyanobacterium Synechocystis sp. PCC 6803

Genetic, Genomics, and Responses to Stresses in Cyanobacteria: Biotechnological Implications

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