A three-colour stress biosensor reveals multimodal response in single cells and spatiotemporal dynamics of biofilms.
Ahmed E ZoheirMorgan S SobolLaura MeischDiana Ordoñez-RuedaAnne-Kristin KasterChristof M NiemeyerKersten S RabePublished in: NPJ biofilms and microbiomes (2023)
The plethora of stress factors that can damage microbial cells has evolved sophisticated stress response mechanisms. While existing bioreporters can monitor individual responses, sensors for detecting multimodal stress responses in living microorganisms are still lacking. Orthogonally detectable red, green, and blue fluorescent proteins combined in a single plasmid, dubbed RGB-S reporter, enable simultaneous, independent, and real-time analysis of the transcriptional response of Escherichia coli using three promoters which report physiological stress (PosmY for RpoS), genotoxicity (PsulA for SOS), and cytotoxicity (PgrpE for RpoH). The bioreporter is compatible with standard analysis and Fluorescent Activated Cell Sorting (FACS) combined with subsequent transcriptome analysis. Various stressors, including the biotechnologically relevant 2-propanol, activate one, two, or all three stress responses, which can significantly impact non-stress-related metabolic pathways. Implemented in microfluidic cultivation with confocal fluorescence microscopy imaging, the RGB-S reporter enabled spatiotemporal analysis of live biofilms revealing stratified subpopulations of bacteria with heterogeneous stress responses.
Keyphrases
- escherichia coli
- induced apoptosis
- label free
- quantum dots
- crispr cas
- cell cycle arrest
- high resolution
- single cell
- stress induced
- high throughput
- single molecule
- oxidative stress
- candida albicans
- gene expression
- cell death
- optical coherence tomography
- living cells
- microbial community
- endoplasmic reticulum stress
- signaling pathway
- pseudomonas aeruginosa
- cell proliferation
- heat stress
- heat shock
- klebsiella pneumoniae
- biofilm formation
- heat shock protein