Real-time RNA profiling within a single bacterium.
Characterizing the dynamics of specific RNA levels requires real-time RNA profiling in a single cell. We show that the combination of a synthetic modular genetic system with fluorescence correlation spectroscopy allows us to directly measure in real time the activity of any specific promoter in prokaryotes. Using a simple inducible gene expression system, we found that induced RNA levels within a single bacterium of Escherichia coli exhibited a pulsating profile in response to a steady input of inducer. The genetic deletion of an efflux pump system, a key determinant of antibiotic resistance, altered the pulsating transcriptional dynamics and caused overexpression of induced RNA. In contrast with population measurements, real-time RNA profiling permits identifying relationships between genotypes and transcriptional dynamics that are accessible only at the level of the single cell.
Le
Thuc T
TT
Institute for Biophysical Dynamics and The James Franck Institute and Department of Biochemistry and Molecular Biology, University of Chicago, 5640 South Ellis Avenue, Chicago, IL 60637, USA.
Harlepp
Sébastien
S
Guet
Calin C
CC
Dittmar
Kimberly
K
Emonet
Thierry
T
Pan
Tao
T
Cluzel
Philippe
P
D001426:Bacterial Proteins / Q000737:chemistry; D002453:Cell Cycle; D049109:Cell Proliferation; D004305:Dose-Response Relationship, Drug; D018432:Drug Resistance, Multiple; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D005838:Genotype; D049452:Green Fluorescent Proteins / Q000378:metabolism; D015233:Models, Statistical; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012313:RNA / Q000737:chemistry* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D017381:Spliceosomes / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D012313:RNA