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Agitation down-regulates immunoglobulin binding protein EibG expression in Shiga toxin-producing Escherichia coli (STEC).

Kuczius T, Zhang W, Merkel V, Mellmann A, Tarr PI, Karch H - PLoS ONE (2015)

Bottom Line: EibG proteins as well as corresponding mRNA were highly expressed under static growth conditions while shearing stress created by agitation during growth repressed protein synthesis.High and low EibG expression was reversible indicating a process with up- and down-regulation of the protein expression.Our findings indicate that shear stress represses EibG expression and might reduce bacterial attachments to cells and surfaces.

View Article: PubMed Central - PubMed

Affiliation: Institute for Hygiene, Westfälische Wilhelms-University and University Hospital Münster, Robert Koch-Strasse 41, 48149, Münster, Germany.

ABSTRACT
Shiga toxin (Stx)-producing Escherichia coli (STEC) carrying eibG synthesize Escherichia coli immunoglobulin binding protein (EibG). EibG nonspecifically binds to immunoglobulins and tends to aggregate in multimers but is poorly expressed in wild-type strains. To study synthesis of the proteins and their regulation in the pathogens, we identified natural growth conditions that increased EibG synthesis. EibG proteins as well as corresponding mRNA were highly expressed under static growth conditions while shearing stress created by agitation during growth repressed protein synthesis. Further regulation effects were driven by reduced oxygen tension, and pH up-regulated EibG expression, but to a lesser extent than growth conditions while decreased temperature down-regulated EibG. Bacteria with increased EibG expression during static growth conditions showed a distinct phenotype with chain formation and biofilm generation, which disappeared with motion. High and low EibG expression was reversible indicating a process with up- and down-regulation of the protein expression. Our findings indicate that shear stress represses EibG expression and might reduce bacterial attachments to cells and surfaces.

No MeSH data available.


Related in: MedlinePlus

EibG expression is down-regulated by shear stress.STEC strain 2875/96 was grown in LB medium at varying revolutions per minute (rpm), as indicated. After overnight inoculation at 37°C, bacteria were harvested by centrifugation. For comparison, identical protein quantities, ranging from 3.0 to 0.1 μg per lane were separated by SDS-PAGE and immunoblotted. EibG signals were visualized using mab human Fc fragment and chemiluminescence (A). Marker sizes are indicated (M). EibG signals were quantified densitometrically and intensities are provided (B; n.d. = not determined). To analyse differences in signal intensities among protein amounts loaded on the gel and among gel runs, EibG signals of three independent immunoblots were quantified (C). The highest signal intensity was defined as 1.0 and the ratios of lower signals were estimated and reported as means (± standard deviations). The ratios of EibG signals are provided for 3 and 1 μg protein (black bars and grey bars, respectively).
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pone.0119583.g002: EibG expression is down-regulated by shear stress.STEC strain 2875/96 was grown in LB medium at varying revolutions per minute (rpm), as indicated. After overnight inoculation at 37°C, bacteria were harvested by centrifugation. For comparison, identical protein quantities, ranging from 3.0 to 0.1 μg per lane were separated by SDS-PAGE and immunoblotted. EibG signals were visualized using mab human Fc fragment and chemiluminescence (A). Marker sizes are indicated (M). EibG signals were quantified densitometrically and intensities are provided (B; n.d. = not determined). To analyse differences in signal intensities among protein amounts loaded on the gel and among gel runs, EibG signals of three independent immunoblots were quantified (C). The highest signal intensity was defined as 1.0 and the ratios of lower signals were estimated and reported as means (± standard deviations). The ratios of EibG signals are provided for 3 and 1 μg protein (black bars and grey bars, respectively).

Mentions: Incubation of the strains in identical volume flasks (100 ml), the EibG expression varied inversely with the agitation frequency (Fig. 2). At 180 rpm, we identified modest synthesis of EibG from strain 2875/96 (Fig. 2A) with faint but visible bands diminishing in signal intensities with decreasing protein amounts (Fig. 2B), and more prominent signals with reduced agitation rates (110 to 40 rpm). The strongest signals were observed for bacteria grown under static conditions, and immunoblots showed a slight decrease in EibG signals at low rates of agitation (40 rpm) (Fig. 2C). High frequency shaking (110–180 rpm) elicited the weakest signal intensities.


Agitation down-regulates immunoglobulin binding protein EibG expression in Shiga toxin-producing Escherichia coli (STEC).

Kuczius T, Zhang W, Merkel V, Mellmann A, Tarr PI, Karch H - PLoS ONE (2015)

EibG expression is down-regulated by shear stress.STEC strain 2875/96 was grown in LB medium at varying revolutions per minute (rpm), as indicated. After overnight inoculation at 37°C, bacteria were harvested by centrifugation. For comparison, identical protein quantities, ranging from 3.0 to 0.1 μg per lane were separated by SDS-PAGE and immunoblotted. EibG signals were visualized using mab human Fc fragment and chemiluminescence (A). Marker sizes are indicated (M). EibG signals were quantified densitometrically and intensities are provided (B; n.d. = not determined). To analyse differences in signal intensities among protein amounts loaded on the gel and among gel runs, EibG signals of three independent immunoblots were quantified (C). The highest signal intensity was defined as 1.0 and the ratios of lower signals were estimated and reported as means (± standard deviations). The ratios of EibG signals are provided for 3 and 1 μg protein (black bars and grey bars, respectively).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4352079&req=5

pone.0119583.g002: EibG expression is down-regulated by shear stress.STEC strain 2875/96 was grown in LB medium at varying revolutions per minute (rpm), as indicated. After overnight inoculation at 37°C, bacteria were harvested by centrifugation. For comparison, identical protein quantities, ranging from 3.0 to 0.1 μg per lane were separated by SDS-PAGE and immunoblotted. EibG signals were visualized using mab human Fc fragment and chemiluminescence (A). Marker sizes are indicated (M). EibG signals were quantified densitometrically and intensities are provided (B; n.d. = not determined). To analyse differences in signal intensities among protein amounts loaded on the gel and among gel runs, EibG signals of three independent immunoblots were quantified (C). The highest signal intensity was defined as 1.0 and the ratios of lower signals were estimated and reported as means (± standard deviations). The ratios of EibG signals are provided for 3 and 1 μg protein (black bars and grey bars, respectively).
Mentions: Incubation of the strains in identical volume flasks (100 ml), the EibG expression varied inversely with the agitation frequency (Fig. 2). At 180 rpm, we identified modest synthesis of EibG from strain 2875/96 (Fig. 2A) with faint but visible bands diminishing in signal intensities with decreasing protein amounts (Fig. 2B), and more prominent signals with reduced agitation rates (110 to 40 rpm). The strongest signals were observed for bacteria grown under static conditions, and immunoblots showed a slight decrease in EibG signals at low rates of agitation (40 rpm) (Fig. 2C). High frequency shaking (110–180 rpm) elicited the weakest signal intensities.

Bottom Line: EibG proteins as well as corresponding mRNA were highly expressed under static growth conditions while shearing stress created by agitation during growth repressed protein synthesis.High and low EibG expression was reversible indicating a process with up- and down-regulation of the protein expression.Our findings indicate that shear stress represses EibG expression and might reduce bacterial attachments to cells and surfaces.

View Article: PubMed Central - PubMed

Affiliation: Institute for Hygiene, Westfälische Wilhelms-University and University Hospital Münster, Robert Koch-Strasse 41, 48149, Münster, Germany.

ABSTRACT
Shiga toxin (Stx)-producing Escherichia coli (STEC) carrying eibG synthesize Escherichia coli immunoglobulin binding protein (EibG). EibG nonspecifically binds to immunoglobulins and tends to aggregate in multimers but is poorly expressed in wild-type strains. To study synthesis of the proteins and their regulation in the pathogens, we identified natural growth conditions that increased EibG synthesis. EibG proteins as well as corresponding mRNA were highly expressed under static growth conditions while shearing stress created by agitation during growth repressed protein synthesis. Further regulation effects were driven by reduced oxygen tension, and pH up-regulated EibG expression, but to a lesser extent than growth conditions while decreased temperature down-regulated EibG. Bacteria with increased EibG expression during static growth conditions showed a distinct phenotype with chain formation and biofilm generation, which disappeared with motion. High and low EibG expression was reversible indicating a process with up- and down-regulation of the protein expression. Our findings indicate that shear stress represses EibG expression and might reduce bacterial attachments to cells and surfaces.

No MeSH data available.


Related in: MedlinePlus