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FliZ is a global regulatory protein affecting the expression of flagellar and virulence genes in individual Xenorhabdus nematophila bacterial cells.

Jubelin G, Lanois A, Severac D, Rialle S, Longin C, Gaudriault S, Givaudan A - PLoS Genet. (2013)

Bottom Line: Heterogeneity in the expression of various bacterial genes has been shown to result in the presence of individuals with different phenotypes within clonal bacterial populations.Moreover, studies of a bacterial population exposed to a graded series of FliZ concentrations showed that FliZ functioned as a rheostat, controlling the rate of transition between the "OFF" and "ON" states in individuals.FliZ thus plays a key role in cell fate decisions, by transiently creating individuals with different potentials for motility and host interactions.

View Article: PubMed Central - PubMed

Affiliation: INRA, UMR 1333 Laboratoire DGIMI, Montpellier, France ; Université Montpellier 2, UMR 1333 Laboratoire DGIMI, Montpellier, France.

ABSTRACT
Heterogeneity in the expression of various bacterial genes has been shown to result in the presence of individuals with different phenotypes within clonal bacterial populations. The genes specifying motility and flagellar functions are coordinately regulated and form a complex regulon, the flagellar regulon. Complex interplay has recently been demonstrated in the regulation of flagellar and virulence gene expression in many bacterial pathogens. We show here that FliZ, a DNA-binding protein, plays a key role in the insect pathogen, Xenorhabdus nematophila, affecting not only hemolysin production and virulence in insects, but efficient swimming motility. RNA-Seq analysis identified FliZ as a global regulatory protein controlling the expression of 278 Xenorhabdus genes either directly or indirectly. FliZ is required for the efficient expression of all flagellar genes, probably through its positive feedback loop, which controls expression of the flhDC operon, the master regulator of the flagellar circuit. FliZ also up- or downregulates the expression of numerous genes encoding non-flagellar proteins potentially involved in key steps of the Xenorhabdus lifecycle. Single-cell analysis revealed the bimodal expression of six identified markers of the FliZ regulon during exponential growth of the bacterial population. In addition, a combination of fluorescence-activated cell sorting and RT-qPCR quantification showed that this bimodality generated a mixed population of cells either expressing ("ON state") or not expressing ("OFF state") FliZ-dependent genes. Moreover, studies of a bacterial population exposed to a graded series of FliZ concentrations showed that FliZ functioned as a rheostat, controlling the rate of transition between the "OFF" and "ON" states in individuals. FliZ thus plays a key role in cell fate decisions, by transiently creating individuals with different potentials for motility and host interactions.

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Bimodal expression of flagellin and hemolysin genes in Xenorhabdus.The strains indicated were grown in LB medium to mid-exponential growth phase (strains with PfliC-gfp[AAV], pPROBE-gfp[AAV] constructs or PD31-gfp[AAV]constructs) or mid-stationary growth phase (strains with PxaxAB-gfp[AAV] or PxhlBA-gfp[AAV] constructs) and GFP fluorescence signals were quantified in individual bacterial cells by flow cytometry. The pPROBE-gfp[AAV] vector carrying a promoter-less gfp gene and the PD31-gfp[AAV] construct carrying the gfp gene under the control of a constitutive promoter were used as negative and positive controls, respectively. Data are shown on two-dimensional dot plots, with the GFP signal on the x-axis and the forward scatter parameter (FSC) on the y-axis. Gates corresponding to GFP-negative and GFP-positive populations are shown and the corresponding percentages are indicated at the bottom left and bottom right of each image, respectively.
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pgen-1003915-g002: Bimodal expression of flagellin and hemolysin genes in Xenorhabdus.The strains indicated were grown in LB medium to mid-exponential growth phase (strains with PfliC-gfp[AAV], pPROBE-gfp[AAV] constructs or PD31-gfp[AAV]constructs) or mid-stationary growth phase (strains with PxaxAB-gfp[AAV] or PxhlBA-gfp[AAV] constructs) and GFP fluorescence signals were quantified in individual bacterial cells by flow cytometry. The pPROBE-gfp[AAV] vector carrying a promoter-less gfp gene and the PD31-gfp[AAV] construct carrying the gfp gene under the control of a constitutive promoter were used as negative and positive controls, respectively. Data are shown on two-dimensional dot plots, with the GFP signal on the x-axis and the forward scatter parameter (FSC) on the y-axis. Gates corresponding to GFP-negative and GFP-positive populations are shown and the corresponding percentages are indicated at the bottom left and bottom right of each image, respectively.

Mentions: Microscopic observations performed during insect infection have revealed that the expression of the fliC and xaxAB genes in Xenorhabdus is heterogeneous differing between individual cells [23]. We investigated the heterogeneity of fliC, xaxAB and xhlBA gene expression during bacterial growth in vitro, by measuring the expression of these genes in the wild type strain carrying the PfliC-gfp[AAV], PxaxAB-gfp[AAV] or PxhlBA-gfp[AAV] fusion by flow cytometry. Two distinct populations (OFF [GFP-negative cells] and ON [GFP-positive cells]) were visualized, providing evidence of a bimodal distribution of cells, in terms of the expression of flagellin and hemolysin genes, in Xenorhabdus (Figure 2 and S3).


FliZ is a global regulatory protein affecting the expression of flagellar and virulence genes in individual Xenorhabdus nematophila bacterial cells.

Jubelin G, Lanois A, Severac D, Rialle S, Longin C, Gaudriault S, Givaudan A - PLoS Genet. (2013)

Bimodal expression of flagellin and hemolysin genes in Xenorhabdus.The strains indicated were grown in LB medium to mid-exponential growth phase (strains with PfliC-gfp[AAV], pPROBE-gfp[AAV] constructs or PD31-gfp[AAV]constructs) or mid-stationary growth phase (strains with PxaxAB-gfp[AAV] or PxhlBA-gfp[AAV] constructs) and GFP fluorescence signals were quantified in individual bacterial cells by flow cytometry. The pPROBE-gfp[AAV] vector carrying a promoter-less gfp gene and the PD31-gfp[AAV] construct carrying the gfp gene under the control of a constitutive promoter were used as negative and positive controls, respectively. Data are shown on two-dimensional dot plots, with the GFP signal on the x-axis and the forward scatter parameter (FSC) on the y-axis. Gates corresponding to GFP-negative and GFP-positive populations are shown and the corresponding percentages are indicated at the bottom left and bottom right of each image, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814329&req=5

pgen-1003915-g002: Bimodal expression of flagellin and hemolysin genes in Xenorhabdus.The strains indicated were grown in LB medium to mid-exponential growth phase (strains with PfliC-gfp[AAV], pPROBE-gfp[AAV] constructs or PD31-gfp[AAV]constructs) or mid-stationary growth phase (strains with PxaxAB-gfp[AAV] or PxhlBA-gfp[AAV] constructs) and GFP fluorescence signals were quantified in individual bacterial cells by flow cytometry. The pPROBE-gfp[AAV] vector carrying a promoter-less gfp gene and the PD31-gfp[AAV] construct carrying the gfp gene under the control of a constitutive promoter were used as negative and positive controls, respectively. Data are shown on two-dimensional dot plots, with the GFP signal on the x-axis and the forward scatter parameter (FSC) on the y-axis. Gates corresponding to GFP-negative and GFP-positive populations are shown and the corresponding percentages are indicated at the bottom left and bottom right of each image, respectively.
Mentions: Microscopic observations performed during insect infection have revealed that the expression of the fliC and xaxAB genes in Xenorhabdus is heterogeneous differing between individual cells [23]. We investigated the heterogeneity of fliC, xaxAB and xhlBA gene expression during bacterial growth in vitro, by measuring the expression of these genes in the wild type strain carrying the PfliC-gfp[AAV], PxaxAB-gfp[AAV] or PxhlBA-gfp[AAV] fusion by flow cytometry. Two distinct populations (OFF [GFP-negative cells] and ON [GFP-positive cells]) were visualized, providing evidence of a bimodal distribution of cells, in terms of the expression of flagellin and hemolysin genes, in Xenorhabdus (Figure 2 and S3).

Bottom Line: Heterogeneity in the expression of various bacterial genes has been shown to result in the presence of individuals with different phenotypes within clonal bacterial populations.Moreover, studies of a bacterial population exposed to a graded series of FliZ concentrations showed that FliZ functioned as a rheostat, controlling the rate of transition between the "OFF" and "ON" states in individuals.FliZ thus plays a key role in cell fate decisions, by transiently creating individuals with different potentials for motility and host interactions.

View Article: PubMed Central - PubMed

Affiliation: INRA, UMR 1333 Laboratoire DGIMI, Montpellier, France ; Université Montpellier 2, UMR 1333 Laboratoire DGIMI, Montpellier, France.

ABSTRACT
Heterogeneity in the expression of various bacterial genes has been shown to result in the presence of individuals with different phenotypes within clonal bacterial populations. The genes specifying motility and flagellar functions are coordinately regulated and form a complex regulon, the flagellar regulon. Complex interplay has recently been demonstrated in the regulation of flagellar and virulence gene expression in many bacterial pathogens. We show here that FliZ, a DNA-binding protein, plays a key role in the insect pathogen, Xenorhabdus nematophila, affecting not only hemolysin production and virulence in insects, but efficient swimming motility. RNA-Seq analysis identified FliZ as a global regulatory protein controlling the expression of 278 Xenorhabdus genes either directly or indirectly. FliZ is required for the efficient expression of all flagellar genes, probably through its positive feedback loop, which controls expression of the flhDC operon, the master regulator of the flagellar circuit. FliZ also up- or downregulates the expression of numerous genes encoding non-flagellar proteins potentially involved in key steps of the Xenorhabdus lifecycle. Single-cell analysis revealed the bimodal expression of six identified markers of the FliZ regulon during exponential growth of the bacterial population. In addition, a combination of fluorescence-activated cell sorting and RT-qPCR quantification showed that this bimodality generated a mixed population of cells either expressing ("ON state") or not expressing ("OFF state") FliZ-dependent genes. Moreover, studies of a bacterial population exposed to a graded series of FliZ concentrations showed that FliZ functioned as a rheostat, controlling the rate of transition between the "OFF" and "ON" states in individuals. FliZ thus plays a key role in cell fate decisions, by transiently creating individuals with different potentials for motility and host interactions.

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