title: Epitope-Tagged Autotransporters as Single-Cell Reporters for Gene Expression by a Salmonella Typhimurium wbaP Mutant.
abstract: Phenotypic diversity is an important trait of bacterial populations and can enhance fitness of the existing genotype in a given environment. To characterize different subpopulations, several studies have analyzed differential gene expression using fluorescent reporters. These studies visualized either single or multiple genes within single cells using different fluorescent proteins. However, variable maturation and folding kinetics of different fluorophores complicate the study of dynamics of gene expression. Here, we present a proof-of-principle study for an alternative gene expression system in a wbaP mutant of Salmonella Typhimurium (S. Tm) lacking the O-sidechain of the lipopolysaccharide. We employed the hemagglutinin (HA)-tagged inverse autotransporter invasin (invAHA) as a transcriptional reporter for the expression of the type three secretion system 1 (T1) in S. Tm. Using a two-reporter approach with GFP and the InvAHA in single cells, we verify that this reporter system can be used for T1 gene expression analysis, at least in strains lacking the O-antigen (wbaP), which are permissive for detection of the surface-exposed HA-epitope. When we placed the two reporters gfp and invAHA under the control of either one or two different promoters of the T1 regulon, we were able to show correlative expression of both reporters. We conclude that the invAHA reporter system is a suitable tool to analyze T1gene expression in S. Tm and propose its applicability as molecular tool for gene expression studies within single cells.
authors.affiliation: Institute of Microbiology, Department of Biology, ETH Zürich, Zürich, Switzerland.
authors.forename: Ismeta
authors.initials: I
authors.lastname: Curkić
authors.affiliation: Institute of Microbiology and Hygiene, University of Tübingen, Tübingen, Germany.
authors.forename: Monika
authors.initials: M
authors.lastname: Schütz
authors.affiliation: Institute of Microbiology and Hygiene, University of Tübingen, Tübingen, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Oberhettinger
authors.affiliation: Institute of Microbiology, Department of Biology, ETH Zürich, Zürich, Switzerland.
authors.forename: Médéric
authors.initials: M
authors.lastname: Diard
authors.affiliation: Institute of Molecular Systems Biology, ETH Zürich, Zürich, Switzerland.
authors.forename: Manfred
authors.initials: M
authors.lastname: Claassen
authors.affiliation: Department of Biosciences, EVOGENE Section, University of Oslo, Oslo, Norway.
authors.forename: Dirk
authors.initials: D
authors.lastname: Linke
authors.affiliation: Institute of Microbiology, Department of Biology, ETH Zürich, Zürich, Switzerland.
authors.forename: Wolf-Dietrich
authors.initials: WD
authors.lastname: Hardt
mesh_terms: D002352:Carrier Proteins / Q000235:genetics*; D002876:Chromosomes, Bacterial; D000939:Epitopes / Q000235:genetics*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D009154:Mutation; D010957:Plasmids; D012486:Salmonella typhimurium / Q000235:genetics*
chemical_list: D002352:Carrier Proteins; D000939:Epitopes