A portable epigenetic switch for bistable gene expression in bacteria

Olivenza, David R.; Nicoloff, Hervé; Sánchez-Romero, María Antonia; Cota, Ignacio; Andersson, Dan I.; Casadesús, Josep

doi:10.1038/s41598-019-47650-2

Cited by 20 publications

(16 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The resulting PCR product ( llp::gfp ) was integrated at the lac locus of DR3. A synthetic ribosome‐binding site named BI‐RBS used in a previous study (Olivenza et al ., 2019) was inserted upstream of the llp gene to optimize Llp production.…”

Section: Methodsmentioning

confidence: 99%

“…Escherichia coli K12 MG1655 was provided by the E. coli Genetic Stock Center. DR3, an MG1655 derivative with the lactose operon deleted (Olivenza et al, 2019), was used as an intermediate strain for the construction of DR29 and DR30. For construction of DR29, the opvAB::gfp fusion was amplified from SV6727 using the oligonucleotides MG-opvA and MG-opvB (Table 2).…”

Section: Bacterial Strainsmentioning

confidence: 99%

See 1 more Smart Citation

Epigenetic biosensors for bacteriophage detection and phage receptor discrimination

Olivenza

Casadesús

Ansaldi

2020

Environmental Microbiology

Self Cite

View full text Add to dashboard Cite

Summary Environmental monitoring of bacteria using phage‐based biosensors has been widely developed for many different species. However, there are only a few available methods to detect specific bacteriophages in raw environmental samples. In this work, we developed a simple and efficient assay to rapidly monitor the phage content of a given sample. The assay is based on the bistable expression of the Salmonella enterica opvAB operon. Under regular growth conditions, opvAB is only expressed by a small fraction of the bacterial subpopulation. In the OpvABON subpopulation, synthesis of the OpvA and OpvB products shortens the O‐antigen and confers resistance to phages that use LPS as a receptor. As a consequence, the OpvABON subpopulation is selected in the presence of such phages. Using an opvAB::gfp fusion, we could monitor LPS‐binding phages in various media, including raw water samples. To enlarge our phage‐biosensor panoply, we also developed biosensors able to detect LPS, as well as protein‐binding coliphages. Moreover, the combination of these tools allowed to identify the bacterial receptor triggering phage infection. The epigenetic opvAB::gfp biosensor thus comes in different flavours to detect a wide range of bacteriophages and identify the type of receptor they recognize.

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Bacterial Strainsmentioning

confidence: 99%

Epigenetic biosensors for bacteriophage detection and phage receptor discrimination

Olivenza

Casadesús

Ansaldi

2020

Environmental Microbiology

Self Cite

View full text Add to dashboard Cite

show abstract

“…E. coli K12 MG1655 was provided by the E. coli Genetic Stock Center. DR3, an MG1655 derivative with the lactose operon deleted (40), was used as an intermediate strain for the construction of DR29 and DR30. For construction of DR29, the opvAB :: gfp fusion was amplified from SV6727 using the oligonucleotides MG-opvA and MG-opvB.…”

Section: Methodsmentioning

confidence: 99%

“…The resulting PCR product ( llp::gfp ) was integrated at the lac locus of DR3. A synthetic ribosome binding site named BI-RBS used in a previous study (40) was inserted upstream of the llp gene to optimize Llp production.…”

Section: Methodsmentioning

confidence: 99%

Phase variation-based biosensors for bacteriophage detection and phage receptor discrimination

Olivenza

Casadesús

Ansaldi

2019

Preprint

Self Cite

View full text Add to dashboard Cite

13Environmental monitoring of bacteria using phage-based biosensors has been widely 14 developed for many different species. However, there are only a few available methods to 15 detect specific bacteriophages in raw environmental samples. In this work, we developed a 16 simple and efficient assay to rapidly monitor the phage content of a given sample. The assay is 17 based on the bistable expression of the Salmonella enterica opvAB operon. Under regular 18 growth conditions, opvAB is only expressed by a small fraction of the bacterial subpopulation. 19In the OpvAB ON subpopulation, synthesis of the OpvA and OpvB products shortens the O-20 antigen in the lipopolysaccharide and confers resistance to phages that use LPS as a receptor. 21As a consequence, the OpvAB ON subpopulation is selected in the presence of such phages. 22Using an opvAB::gfp fusion, we could monitor LPS-binding phages in various media, including 23 raw water samples. To enlarge our phage-biosensor panoply, we also developed several 24 coliphage biosensors that proved efficient to detect LPS-as well as protein-binding coliphages. 25Moreover, the combination of these tools allows to identify what is the bacterial receptor 26 triggering phage infection. The opvAB::gfp biosensor thus comes in different flavours to 27 efficiently detect a wide range of bacteriophages and identify the type of receptor they 28 recognize. 29 30 31 from environmental samples still constitutes a challenge for those interested in isolating and 35 characterizing bacteriophages for ecological or biotechnological purposes. This work provides 36 a simple and accurate method based on the bi-stable expression of genes that confer 37 resistance to certain classes of bacteriophages in different bacterial models. It paves the way 38 for future development of highly efficient phage biosensors that can discriminate among 39 several receptor-binding phages and that could be declined in many more versions. In a 40 context where phage ecology, research, and therapy are flourishing again, it becomes essential 41 to possess simple and efficient tools for phage detection.42 43 44 45 Keywords: bacteriophage, biosensor, phase variation, LPS, phage receptor 46 47 48 49Bacteriophages, the viruses that infect bacteria, are ubiquitous on Earth, very abundant and 50 highly diverse (1-4). They participate in the daily turnover of bacterial communities and are 51 hypothesized to be major drivers of carbon recycling (5). Their abundance and diversity have 52 long been acknowledged in the oceans and more recently associated to numerous 53 microbiomes as a major part of the viromes (2,(6)(7)(8). Bacteriophages are considered a vast 54 reservoir of genes and a major vector for horizontal gene transfer that allow the emergence of 55 new biological functions (9, 10). Furthermore, phages have been at the origin of many 56 discoveries and concepts in molecular biology (11)(12)(13). 57Among the applications resulting from a century of phage research, phage therapy stands out 58 by receiving more and more a...

show abstract

“…Artificial genetic and epigenetic signaling networks in cells have been used to detect, process and store information about external stimuli in bacteria and mammalian cells [4][5][6][7][8][9][10][11]. In addition, DNA methylation based bistable gene expression switches have been described in bacteria that were based on two model well-studied DNA methyltransferases, the Escherichia coli deoxyadenosine DNA methyltransferase (Dam) and the Caulobacter crescentus cell-cycle-regulated methyltransferase (CcrM) [9,12]. In a previous work, we have developed a DNA methylation based epigenetic memory system in E.coli [9].…”

Section: Introductionmentioning

confidence: 99%

Development of an epigenetic tetracycline sensor system based on DNA methylation

2020

View full text Add to dashboard Cite

Bacterial live cell sensors are potentially powerful tools for the detection of environmental toxins. In this work, we have established and validated a flow cytometry readout for an existing bacterial arabinose sensor system with DNA methylation based memory function (Maier et al., 2017, Nat. Comm., 8:15336). Flow cytometry readout is convenient and enables a multiparameter analysis providing information about single-cell variability, which is beneficial for further development of sensor systems of this type in the future. We then designed a tetracycline sensor system, because of the importance of antibiotics pollution in the light of multi-resistant pathogens. To this end, a tetracycline trigger plasmid was constructed by replacing the araC repressor gene and the ara operator of the arabinose trigger plasmid with the tetR gene coding for the tetracycline repressor and the tet operon. After combination with the memory plasmid, the tetracycline sensor system was shown to be functional in E. coli allowing to detect and memorize the presence of tetracycline. Due to a positive feedback between the trigger and memory systems, the combined whole-cell biosensor showed a very high sensitivity for tetracycline with a detection threshold at 0.1 ng/ml tetracycline, which may be a general property of sensors of this type. Moreover, acute presence of tetracycline and past exposure can be detected by this sensor using the dual readout of two reporter fluorophores.

show abstract

A portable epigenetic switch for bistable gene expression in bacteria

Cited by 20 publications

References 39 publications

Epigenetic biosensors for bacteriophage detection and phage receptor discrimination

Epigenetic biosensors for bacteriophage detection and phage receptor discrimination

Phase variation-based biosensors for bacteriophage detection and phage receptor discrimination

Development of an epigenetic tetracycline sensor system based on DNA methylation

Contact Info

Product

Resources

About