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Lgt processing is an essential step in Streptococcus suis lipoprotein mediated innate immune activation.

Wichgers Schreur PJ, Rebel JM, Smits MA, van Putten JP, Smith HE - PLoS ONE (2011)

Bottom Line: Genetic inactivation of a putative prolipoprotein diacylglyceryl transferase (Lgt) gene resulted in deficient lipoprotein synthesis as evidenced by palmitate labeling.The Lgt mutant showed strongly reduced activation of porcine PBMCs, indicating that lipoproteins are dominant porcine PBMC activating molecules of S. suis.In addition, we provide evidence that Lgt processing of lipoproteins is required for lipoprotein mediated innate immune activation.

View Article: PubMed Central - PubMed

Affiliation: Central Veterinary Institute, Wageningen UR, Lelystad, The Netherlands. paul.wichgersschreur@wur.nl

ABSTRACT

Background: Streptococcus suis causes invasive infections in pigs and occasionally in humans. The host innate immune system plays a major role in counteracting S. suis infections. The main components of S. suis able to activate the innate immune system likely include cell wall constituents that may be released during growth or after cell wall integrity loss, however characterization of these components is still limited.

Methodology/principal findings: [corrected] A concentrated very potent innate immunity activating supernatant of penicillin-treated S. suis was SDS-PAGE fractionated and tested for porcine peripheral blood mononucleated cell (PBMC) stimulating activity using cytokine gene transcript analysis. More than half of the 24 tested fractions increased IL-1β and IL-8 cytokine gene transcript levels in porcine PBMCs. Mass spectrometry of the active fractions indicated 24 proteins including 9 lipoproteins. Genetic inactivation of a putative prolipoprotein diacylglyceryl transferase (Lgt) gene resulted in deficient lipoprotein synthesis as evidenced by palmitate labeling. The Lgt mutant showed strongly reduced activation of porcine PBMCs, indicating that lipoproteins are dominant porcine PBMC activating molecules of S. suis.

Conclusion/significance: This study for the first time identifies and characterizes lipoproteins of S. suis as major activators of the innate immune system of the pig. In addition, we provide evidence that Lgt processing of lipoproteins is required for lipoprotein mediated innate immune activation.

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Related in: MedlinePlus

Lipidation of wild type and Δlgt mutant bacteria.Lipidation was assessed by incubating the wild type and mutant bacteria with [3H]palmitic acid, followed by penicillin treatment and SDS-PAGE. Lipidation was visualized using autoradiography. As a control, total protein release of wild type and mutant bacteria was visualized with Silver staining.
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pone-0022299-g004: Lipidation of wild type and Δlgt mutant bacteria.Lipidation was assessed by incubating the wild type and mutant bacteria with [3H]palmitic acid, followed by penicillin treatment and SDS-PAGE. Lipidation was visualized using autoradiography. As a control, total protein release of wild type and mutant bacteria was visualized with Silver staining.

Mentions: To verify that lipoprotein processing had been abolished in the Δlgt mutant, lipidation of lipoproteins in the wild type and mutant was analyzed. Bacteria were grown in the presence of [3H]palmitic acid and subsequently treated with penicillin. Similar amounts of protein were released from wild type and (complemented) Δlgt mutant bacteria (Fig. 4). Several radiolabeled proteins were detected in the supernatant of the wild type and the Δlgt::pGA14-lgt mutant (Fig. 4), whereas no radiolabeled (lipo)proteins were detected in the supernatant of the Δlgt mutant and the Δlgt::pGA14-cm mutant. These data confirm that Lgt is responsible for lipid modification of prelipoproteins in S. suis.


Lgt processing is an essential step in Streptococcus suis lipoprotein mediated innate immune activation.

Wichgers Schreur PJ, Rebel JM, Smits MA, van Putten JP, Smith HE - PLoS ONE (2011)

Lipidation of wild type and Δlgt mutant bacteria.Lipidation was assessed by incubating the wild type and mutant bacteria with [3H]palmitic acid, followed by penicillin treatment and SDS-PAGE. Lipidation was visualized using autoradiography. As a control, total protein release of wild type and mutant bacteria was visualized with Silver staining.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3139625&req=5

pone-0022299-g004: Lipidation of wild type and Δlgt mutant bacteria.Lipidation was assessed by incubating the wild type and mutant bacteria with [3H]palmitic acid, followed by penicillin treatment and SDS-PAGE. Lipidation was visualized using autoradiography. As a control, total protein release of wild type and mutant bacteria was visualized with Silver staining.
Mentions: To verify that lipoprotein processing had been abolished in the Δlgt mutant, lipidation of lipoproteins in the wild type and mutant was analyzed. Bacteria were grown in the presence of [3H]palmitic acid and subsequently treated with penicillin. Similar amounts of protein were released from wild type and (complemented) Δlgt mutant bacteria (Fig. 4). Several radiolabeled proteins were detected in the supernatant of the wild type and the Δlgt::pGA14-lgt mutant (Fig. 4), whereas no radiolabeled (lipo)proteins were detected in the supernatant of the Δlgt mutant and the Δlgt::pGA14-cm mutant. These data confirm that Lgt is responsible for lipid modification of prelipoproteins in S. suis.

Bottom Line: Genetic inactivation of a putative prolipoprotein diacylglyceryl transferase (Lgt) gene resulted in deficient lipoprotein synthesis as evidenced by palmitate labeling.The Lgt mutant showed strongly reduced activation of porcine PBMCs, indicating that lipoproteins are dominant porcine PBMC activating molecules of S. suis.In addition, we provide evidence that Lgt processing of lipoproteins is required for lipoprotein mediated innate immune activation.

View Article: PubMed Central - PubMed

Affiliation: Central Veterinary Institute, Wageningen UR, Lelystad, The Netherlands. paul.wichgersschreur@wur.nl

ABSTRACT

Background: Streptococcus suis causes invasive infections in pigs and occasionally in humans. The host innate immune system plays a major role in counteracting S. suis infections. The main components of S. suis able to activate the innate immune system likely include cell wall constituents that may be released during growth or after cell wall integrity loss, however characterization of these components is still limited.

Methodology/principal findings: [corrected] A concentrated very potent innate immunity activating supernatant of penicillin-treated S. suis was SDS-PAGE fractionated and tested for porcine peripheral blood mononucleated cell (PBMC) stimulating activity using cytokine gene transcript analysis. More than half of the 24 tested fractions increased IL-1β and IL-8 cytokine gene transcript levels in porcine PBMCs. Mass spectrometry of the active fractions indicated 24 proteins including 9 lipoproteins. Genetic inactivation of a putative prolipoprotein diacylglyceryl transferase (Lgt) gene resulted in deficient lipoprotein synthesis as evidenced by palmitate labeling. The Lgt mutant showed strongly reduced activation of porcine PBMCs, indicating that lipoproteins are dominant porcine PBMC activating molecules of S. suis.

Conclusion/significance: This study for the first time identifies and characterizes lipoproteins of S. suis as major activators of the innate immune system of the pig. In addition, we provide evidence that Lgt processing of lipoproteins is required for lipoprotein mediated innate immune activation.

Show MeSH
Related in: MedlinePlus